Background: Analysis of peripheral blood lymphocyte subsets has become an essential tool in the evaluation of outcome of diagnostic and research related questions in immunological and pathological conditions. Periodic evaluation and establishment of normal lymphocyte reference ranges are required in clinical and research settings of various immunodeficiency disorders for evaluation of the significance of observations. It is also important that age and gender specific lymphocyte subset reference ranges should be locally established for meaningful comparison and accurate result interpretation as age plays a significant role in the development of immune system.

Methods: We performed dual platform flow cytometry to determine reference ranges for lymphocyte subsets (CD3, CD4, CD8, CD19 [B cells] and CD16+CD56+ [Natural Killer - NK cells]) in 50 adolescents (age range: 12-18) and 100 adults (age range: 21-67) along with T cell maturation, activation and co-stimulatory molecules in healthy multiracial adult population of South Florida.

Results: The lymphocyte reference ranges percentages [absolute counts - Abs, cells/μl] unadjusted for gender differences for adolescents are: CD3: 49-83 [939-2959]; CD4: 27-53 [467-1563]; CD8: 16-40 [259-1262]; CD19+ B cells: 8-31 [169-1297] and CD16+CD56+ NK cells: 3-30 [59-1178] and for adults are: CD3: 65-88 [983-3572]; CD4: 26-62 [491-2000]; CD8: 14-44 [314-2,087]; CD19+ B cells: 2-27 [64-800] and CD16+CD56+ NK cells: 2-27 [27-693]. The ranges for CD4:CD8 ratio for adolescents and adults are 0.7-2.6 and 0.6-4.4, respectively. Gender based analysis of relative percentages of lymphocyte subsets showed no significant differences between adult and adolescent males and females. The mean CD4:CD8 ratio was significantly higher in adult females than males (P=0.04) and in adolescents this difference was not significant between genders. The mean CD3 and CD4 T cell percentages were higher and CD19 cell percentages were lower in adults compared to adolescents (P<0.0001). Absolute lymphocyte counts showed a positive correlation with the absolute counts of CD3+, CD4+, CD8+, CD19+, CD16+CD56+, CD45RO+ and CD45RA+ cells (all correlations with P<0.0001 except CD45RO [P=0.01] and CD45RA [P=0.03]).

Conclusion: The reference values of peripheral blood lymphocyte subsets were analyzed in healthy adolescent and adult population of South Florida. This study indicates the need for periodic evaluation and establishment of lymphocyte reference ranges for patient population served based on gender and age since these could influence immune status and treatment outcome.

Reference ranges of blood-circulating leukocyte populations by, e.g., age and sex, are required for monitoring immune-cell kinetics. Most previous reports in which flow cytometry has been used to define the reference ranges for leukocyte counts included a limited number of donors and/or cell populations and/or did not consider age and sex simultaneously. Moreover, other factors not previously considered in the definition of normal ranges, such as the presence of chronic-lymphocytic-leukemia (CLL)-like low-count monoclonal B-cell lymphocytosis (MBLlo), might also be associated with an altered distribution of leukocytes in blood in association with an immunodeficiency and increased risk of infection and cancer. Here, we established reference cell-count ranges for the major populations of leukocytes in blood of non-MBL and MBLlo adult Caucasians matched by age and sex using the EuroFlow Lymphocyte Screening Tube (LST). A total of 706 Caucasian adult donors—622 non-MBL and 84 MBLlo—were recruited from the general population. Among non-MBL donors, the total leukocyte, neutrophil, basophil dendritic cell and monocyte counts remained stable through adulthood, while the absolute numbers of T- and B-cell populations and plasma cells decreased with age. The number of eosinophils and NK-cell increased over time, with clear differences according to sex for certain age ranges. In MBLlo subjects, few differences in the absolute cell counts by age (vs. non-MBL) were observed, and MBLlo men and women showed similar trends to non-MBL subjects except for the B-cell count drop observed in >70 y-men, which was more pronounced in MBLlo vs. non-MBL controls. Building robust age- and sex-matched reference ranges for the most relevant immune-cell populations in the blood of non-MBL donors is essential to appropriately identify an altered immune status in different clinical settings and highlight the altered immune-cell profiles of MBLlo subjects.

Region-specific reference ranges for adult peripheral blood lymphocyte subsets have been established in few countries around the world, but most studies were restricted to younger adults for monitoring of HIV patients. The aim of this investigation was to establish age- and gender-specific reference ranges for healthy adults. Lymphocyte subsets were examined in 100 healthy volunteers (50 males, 50 females) aged 19-85 years by two-color flow cytometric analysis with a FACSCalibur. A statistically significant decline in the mean numbers of CD3+/CD8+ T cells and CD19+ B cells was observed beyond an age of 50 years, whereas the mean counts of NK cells and CD4+/CD8+ ratio significantly increased beyond the age of 50. Females < or = 50 years had significantly higher mean CD4+ T cell counts and lower NK cell counts than males < or = 50 years. Based on these results, we established reference values for three subgroups: males < or = 50 years, females < or = 50 years, and males/females > 50 years.

Existing normative flow cytometry data have several limitations including small sample sizes, incompletely described study populations, variable flow cytometry methodology, and limited depth for defining lymphocyte subpopulations. To overcome these issues, we defined high-dimensional flow cytometry reference ranges for the healthy human immune system using Human Immunology Project Consortium methodologies after carefully screening 127 subjects deemed healthy through clinical and laboratory testing. We enrolled subjects in the following age cohorts: 18-29 years, 30-39, 40-49, and 50-66 and enrolled cohorts to ensure an even gender distribution and at least 30% non-Caucasians. From peripheral blood mononuclear cells, flow cytometry reference ranges were defined for >50 immune subsets including T-cell (activation, maturation, T follicular helper and regulatory T cell), B-cell, and innate cells. We also developed a web tool for visualization of the dataset and download of raw data. This dataset provides the immunology community with a resource to compare and extract data from rigorously characterized healthy subjects across age groups, gender and race.

Blood was collected from the New York State Department of Health (NYSDOH) employees to assess variances in leukocyte numbers in January, May, and September throughout a year and over many years. Women and men of ages 20 to 80 volunteered to donate for this program. Most of the blood came from healthy individuals, and many remained healthy throughout the years of their blood donations. The major objective was to determine the extent that blood leukocyte numbers change so that transient vs more lingering changes may be helpful in assessing health status. Since some donors remained in the program for 14 years, age influences over time could be determined. Within a short period of 2-3 years, the flow cytometric immunophenotypic profile of blood lymphocyte is relatively stable with a CV% of < 20%. However, as humans age, the blood CD3 T cell, CD8 T cell, B cell, NKT cell, and CD4/CD8 double-negative T cell (DN-T cell) subsets declined in cell numbers/μL, but the double-positive CD4/CD8 T cells (DP-T cells) increased in numbers. The extent and chronology of a variance, e.g., a subset exceeding its 75th or 90th percentile, might be indicative of a transient or chronic physiological or psychosocial stress affecting health or a developing pathology; however, because of the wide ranges of cell numbers/μL for each subset among individuals reported as healthy, everyone's immunity and health must be carefully evaluated. A CD4 to CD8 ratio (4/8R) of < 1 has been used to define an immunodeficiency such as HIV-induced AIDS, but a high 4/8R is less well associated with health status. A high 4/8R or granulocyte to lymphocyte ratio (GLR) might be an indicator of a stress, infection, or immune-related pathology. Sporadic and longitudinal increases of GLRs are reported. The results suggest that there are some age and sex differences in leukocyte numbers; stress influences on the blood profile of leukocytes likely exist. However, some values exceeding 2 standard deviations from means do not necessarily predict a health concern, whereas a longitudinal increase or decline might be indicative of a need for further evaluations.

Some people remain healthier throughout life than others but the underlying reasons are poorly understood. Here we hypothesize this advantage is attributable in part to optimal immune resilience (IR), defined as the capacity to preserve and/or rapidly restore immune functions that promote disease resistance (immunocompetence) and control inflammation in infectious diseases as well as other causes of inflammatory stress. We gauge IR levels with two distinct peripheral blood metrics that quantify the balance between (i) CD8+ and CD4+ T-cell levels and (ii) gene expression signatures tracking longevity-associated immunocompetence and mortality-associated inflammation. Profiles of IR metrics in ~48,500 individuals collectively indicate that some persons resist degradation of IR both during aging and when challenged with varied inflammatory stressors. With this resistance, preservation of optimal IR tracked (i) a lower risk of HIV acquisition, AIDS development, symptomatic influenza infection, and recurrent skin cancer; (ii) survival during COVID-19 and sepsis; and (iii) longevity. IR degradation is potentially reversible by decreasing inflammatory stress. Overall, we show that optimal IR is a trait observed across the age spectrum, more common in females, and aligned with a specific immunocompetence-inflammation balance linked to favorable immunity-dependent health outcomes. IR metrics and mechanisms have utility both as biomarkers for measuring immune health and for improving health outcomes.

Immunosenescence is characterized by an age-related decline in immune system function. Major efforts have been made to identify changes in peripheral blood lymphocyte subsets accompanying immunosenescence in elderly adults. However, the change trends of some lymphocyte subsets with age are still controversial, and populations of advanced ages, such as people in their 80s or 90s, have not yet been thoroughly investigated. To provide further insight, we recruited 957 healthy donors without certain diseases with ages ranging from 20 to 95 years. Peripheral lymphocyte subsets, including T cells, CD4 T cells, CD8 T cells, B cells and NK cells, and the CD4/CD8 ratio were measured by flow cytometry. Additionally, regulatory CD4 T cells with inhibitory functions marked by CD3CD4CD25 and the expression of the costimulatory molecule CD28 on CD8 T cells were evaluated. Sex was considered at the same time. The data indicated that in elderly people, peripheral T (p < 0.001), CD4 T (p < 0.001) and B (p < 0.001) lymphocyte subsets decreased, but the NK cell population (p < 0.001) increased. More regulatory CD4 T cells may imply stronger inhibition in the elderly population. The decreased CD28 expression with age in females verified CD28 to be an immunosenescence marker and the sharply decreased CD28 expression after 75 years in males indicated a rapid immunosenescence at the late life span of the male populations. In addition, our study established reference values for peripheral lymphocyte subsets at different age stages in males and females, which are urgently needed for the clinical management and treatment of geriatric diseases.

Objective: To evaluate and quantitatively describe age-dependent homeostasis for a broad range of total T-cells and specific T-lymphocyte subpopulations in healthy human subjects. Methods: A systematic literature review was performed to identify and collect relevant quantitative information on T-lymphocyte counts in human blood and various organs. Both individual subject and grouped (aggregated) data on T-lymphocyte observations in absolute and relative values were digitized and curated; cell phenotypes, gating strategies for flow cytometry analyses, organs from which observations were obtained, subjects' number and age were also systematically inventoried. Age-dependent homeostasis of each T-lymphocyte subpopulation was evaluated via a weighted average calculation within pre-specified age intervals, using a piece-wise equal-effect meta-analysis methodology. Results: In total, 124 studies comprising 11722 unique observations from healthy subjects encompassing 20 different T-lymphocyte subpopulations - total CD45+ and CD3+ lymphocytes, as well as specific CD4+ and CD8+ naïve, recent thymic emigrants, activated, effector and various subpopulations of memory T-lymphocytes (total-memory, central-memory, effector-memory, resident-memory) - were systematically collected and included in the final database for a comprehensive analysis. Blood counts of most T-lymphocyte subpopulations demonstrate a decline with age, with a pronounced decrease within the first 10 years of life. Conversely, memory T-lymphocytes display a tendency to increase in older age groups, particularly after ~50 years of age. Notably, an increase in T-lymphocyte numbers is observed in neonates and infants (0 - 1 year of age) towards less differentiated T-lymphocyte subpopulations, while an increase into more differentiated subpopulations emerges later (1 - 5 years of age). Conclusion: A comprehensive systematic review and meta-analysis of T-lymphocyte age-dependent homeostasis in healthy humans was performed, to evaluate immune T-cell profiles as a function of age and to characterize generalized estimates of T-lymphocyte counts across age groups. Our study introduces a quantitative description of the fundamental parameters characterizing the maintenance and evolution of T-cell subsets with age, based on a comprehensive integration of available organ-specific and systems-level flow cytometry datasets. Overall, it provides the most up-to-date view of physiological T-cell dynamics and its variance and may be used as a consistent reference for gaining further mechanistic understanding of the human immune status in health and disease.

Background: Quantitatively assessing host immunity remains a challenge in clinical practice.

Results: Most parameters in lymphocyte number, function and phenotype were correlated with age. The reference ranges of these parameters were established in four age groups (children, adolescents, adults, and elders). The numbers of CD4 T cells, CD8 T cells, B cells, but not NK cells, were negatively correlated with age. However, the function of CD4 T cells, CD8 T cells and NK cells was positively correlated with age. The expression of CD28 on T cells gradually decreased with increasing age and was negatively correlated with their function. An opposite phenomenon was observed in the expressions of HLA-DR and CD45RO on T cells. An immune scoring model was established by using 8 parameters (CD4 T cell number × function, CD28CD4 T cell number, HLA-DRCD4 T cell number, CD45ROCD4 T cell number, CD8 T cell number × function, CD28CD8 T cell number, HLA-DRCD8 T cell number, NK cell number × function) from the results of lymphocyte number, function, and phenotype. This immune scoring model showed sensitivities of 70% and 71.4% in determining hyper-immune and hypo-immune status, respectively.

Conclusions: An immune scoring model based on combination of lymphocyte number, function, and phenotype shows potential value in quantitatively assessing host immunity.

Methods: 261 healthy individuals aged 1 to 82 years were recruited from Tongji Hospital. The number, function, and phenotype of CD4 T cells, CD8 T cells and NK cells were simultaneously determined.

1.

Of the inflammatory markers identified, C-reactive protein (CRP) has the analyte and assay characteristics that are the most conducive for use in practice.

2.

To obtain a CRP concentration in metabolically stable patients, 2 measurements, fasting or nonfasting, should be made (optimally 2 weeks apart) and the results averaged. If the CRP level is >10 mg/L, then the test should be repeated and the patient examined for sources of infection or inflammation.

3.

CRP results should be expressed only as milligrams per liter and expressed to 1 decimal point.

4.

Risk assessment should be modeled after the lipids approach via 3 risk categories: low risk, average risk, and high risk. On the basis of the CRP population distributions, the following tertiles are recommended for categorizing patients: low risk, <1.0 mg/L; average risk, 1.0 to 3.0 mg/L; and high risk, >3.0 mg/L. It should be recognized that other acute inflammatory conditions may result in mildly to moderately increased CRP levels, such as inflammatory bowel disease, rheumatoid arthritis, and long-term alcoholism.

5.

Performance goals for CRP measurement, similar to those developed for total cholesterol, HDL and LDL cholesterol, and triglycerides, need to be developed with a view toward better characterization of the total allowable error required to measure CRP reliably.

Background: Elevated systematic inflammation is a hallmark of aging, but the association of long-term inflammation trajectories with subsequent aging phenotypes has been little examined. We assessed inflammatory marker C-reactive protein (CRP) repeatedly over time and examined whether long-term changes predicted aging outcomes. Methods: A total of 2,437 men and women aged 47-87 years at baseline (1998-2001) who were participants in the English Longitudinal Study of Ageing had CRP measured on two or three occasions between 1998 and 2009. Inflammation trajectories were computed using latent-class growth mixture modeling and were related to aging outcomes measured in 2012/2013: physical functioning, cardiometabolic, respiratory, mental health, and a composite "healthy aging" outcome. Results: Four CRP trajectories were identified as follows: "stable-low" (71 per cent of the sample) with baseline mean 1.33 mg/L remaining <3 mg/L; "medium-to-high" (14 per cent) with baseline 2.7 mg/L rising to 5.3 mg/L; "high-to-medium" (10 per cent) with baseline 6.6 mg/L decreasing to 2.4 mg/L; and "stable-high" (5 per cent) with levels from 5.7 to 7.5 mg/L. Relative to the stable-low trajectory, individuals in the medium-to-high had a higher risk of limitations in basic activities of daily living (ADL, odds ratio; 95% confidence interval: 2.09; 1.51, 2.88), instrumental ADL (1.62; 1.15, 2.30), impaired balance (1.59; 1.20, 2.11) and walking speed (1.61; 1.15, 2.24), arthritis (1.55; 1.16, 2.06), hypertension (1.57; 1.21, 2.04), obesity (1.95; 1.36, 2.80), poor respiratory function (1.84; 1.36, 2.50), and depression (1.55; 1.13, 2.12). A lower odds of healthy aging was observed in people in the medium-to-high (0.57; 0.40, 0.79) and stable-high (0.50; 0.27, 0.91) trajectories. Conclusions: Older people who displayed an elevation in CRP levels over a decade experienced an increased risk of adverse aging outcomes.

Background: Inflammageing biomarkers have been implicated in frailty among middle-aged and older adults. However, there is a paucity of information regarding inflammageing biomarkers association with adverse clinical outcomes, particularly when it is elevated. Understanding this knowledge may allow clinicians to utilise these biomarkers to provide early preventive strategies. This review investigated the association between the elevated inflammageing biomarkers namely C-Reactive Proteins (CRP) including High-Sensitivity (Hs)-CRP, Interleukin-6 (IL-6) and TNF-α (TNF-α) in community-dwelling adults aged 40 years and above on the outcomes of the risk of all-cause mortality, risk of all-cause hospitalisation and risk of all-cause re-admission. Secondary outcomes included risk of depression and anxiety.

Method: Studies were searched in PubMed, Embase, Biosis Preview, Web of Science, Scopus and ProQuest Dissertations and Theses along with grey literature. A total of 22 cohort studies were included for analysis. Experimental, quasi-experimental, cross-sectional and case-control studies were excluded.

Results: Community-dwelling adults aged 40 years and above have a significantly higher risk of all-cause mortality by 1.54 times and by 1.47 times with elevated CRP/Hs-CRP and elevated IL-6 respectively (p=0.001; p=0.001). Those with elevated CRP/Hs-CRP has significantly higher odds of depression by 1.48 times (p<0.001). Limited evidence exists pertaining to the effect of the inflammageing biomarkers on the risk of hospitalisation, risk of re-admission and risk of anxiety.

Conclusion: Precision care interventions of biomarkers monitoring implemented into chronic disease follow-up on the CRP or Hs-CRP as well as IL-6 levels may reduce the risk of mortality and depression.

Context: Few studies have examined whether the inflammatory markers IL-6 and C-reactive protein (CRP) are associated with exceptional longevity.

Objective: Our objective was to determine the association of serum CRP and IL-6 with adult lifespan. DESIGN, SETTING,

And Participants: This was a prospective, population-based study of 610 men and 743 postmenopausal women, mean age 73 yr, who had serum IL-6 and CRP measurements at baseline (1984-1987) and who were followed for mortality for up to 23 yr (through 2008). Participants must have been old enough at baseline (57 yr) to have the potential to achieve age 80 during follow-up.

Main Outcome Measures: Relative survival time and age at death (lifespan) were assessed.

Results: During follow-up, overall mortality was 69%. After adjustment for cardiovascular disease risk factors, in men, each sd increase in IL-6 was associated with a 15% decrease in survival time and 0.94-yr shorter lifespan (P < 0.001); corresponding values for CRP were a 12% decrease in survival time and a 1.00-yr reduction in lifespan (P < 0.001). Among women not using estrogen therapy (n = 532), survival time decreased 7% per sd higher IL-6 and lifespan was 1.35 yr shorter (P < 0.001). IL-6 was not related to lifespan among women using estrogen. CRP levels were not significantly associated with survival time or lifespan in women regardless of estrogen status.

Conclusions: Higher levels of inflammatory markers predicted reduced survival time and shorter lifespan among older men, whereas only IL-6 was associated with longevity in postmenopausal women and only among those not using estrogen.

Background/purpose: Interleukin-6 (IL-6) and C-reactive protein (CRP) are inflammatory makers of potential interest in all-cause and cardiovascular death risk prediction, but their additive explanatory value to established risk factors is not well documented among nonwestern populations.

Methods: We investigated the additive value of IL-6 and CRP to the Framingham risk score and lifestyle factors in predicting all-cause and cardiovascular mortality among a population-representative sample of 1023 adults aged 54 years and above in Taiwan.

Results: A total of 351 deaths and 82 cardiovascular deaths were identified (median follow-up = 11.2 years). After adjustment for established risk factors, elevated IL-6 and CRP levels were associated with a higher risk of all-cause death: the hazard ratios for the highest risk quartile compared with the lowest quartile were 3.64 (95% confidence interval, 2.44-5.44) for IL-6 and 2.31 (95% confidence interval, 1.62-3.29) for CRP. IL-6 was also significantly associated with cardiovascular mortality. For both all-cause and cardiovascular mortality, IL-6 yielded a substantial and significant increase in the area under the receiver operator characteristic curve (change in the area under the receiver operator characteristic curve = 0.036 and 0.024, respectively), but CRP did not (change in the area under the receiver operator characteristic curve = 0.004 and 0.009, respectively).

Conclusion: Although both IL-6 and CRP were significantly associated with all-cause mortality, only IL-6 provided a substantial improvement in discrimination. Similarly, IL-6 demonstrated a notable prognostic value for predicting cardiovascular mortality, but not CRP. These findings provide further support for the role of inflammation in the deterioration of health at older ages among a nonwestern population.

Objective: to assess the levels and determinants of interleukin (IL)-1β, IL-6, tumour necrosis factor (TNF)-α and C-reactive protein (CRP) in a healthy Caucasian population.

Methods: population sample of 2884 men and 3201 women aged 35 to 75. IL-1β, IL-6 and TNF-α were assessed by a multiplexed particle-based flow cytometric assay and CRP by an immunometric assay.

Results: Spearman rank correlations between duplicate cytokine measurements (N = 80) ranged between 0.89 and 0.96; intra-class correlation coefficients ranged between 0.94 and 0.97, indicating good reproducibility. Among the 6085 participants, 2289 (37.6%), 451 (7.4%) and 43 (0.7%) had IL-1β, IL-6 and TNF-α levels below detection limits, respectively. Median (interquartile range) for participants with detectable values were 1.17 (0.48-3.90) pg/ml for IL-1β; 1.47 (0.71-3.53) pg/ml for IL-6; 2.89 (1.82-4.53) pg/ml for TNF-α and 1.3 (0.6-2.7) ng/ml for CRP. On multivariate analysis, greater age was the only factor inversely associated with IL-1β levels. Male sex, increased BMI and smoking were associated with greater IL-6 levels, while no relationship was found for age and leisure-time PA. Male sex, greater age, increased BMI and current smoking were associated with greater TNF-α levels, while no relationship was found with leisure-time PA. CRP levels were positively related to age, BMI and smoking, and inversely to male sex and physical activity.

Conclusion: Population-based levels of several cytokines were established. Increased age and BMI, and to a lesser degree sex and smoking, significantly and differentially impact cytokine levels, while leisure-time physical activity has little effect.

Unlabelled: To investigate whether interleukin-6 (IL-6), C-reactive protein (CRP) and tumor necrosis factor-alpha (TNF-α) protein levels predict all-cause mortality in older persons living in the community.

Design: Prospective cohort study.

Setting: Data were from the Aging and Longevity Study in the Sirente Geographic Area, a prospective cohort study.

Participants: Individuals aged 80 and older living in an Italian mountain community (N = 362).

Measurements: Participants were classified according to the median value of the three inflammation markers (IL-6, 2.08 pg/mL; TNF-α, 1.43 pg/mL; CRP, 3.08 mg/L). A composite summary score of inflammation was also created. The main outcome was risk of death after 4 years of follow-up.

Results: One hundred fifty deaths occurred during 4 years of follow-up. In the unadjusted model, high levels of each of the three markers were associated with greater mortality. After adjusting for potential confounders, high levels of IL-6 (hazard ratio (HR) = 2.18, 95% confidence interval (CI) = 1.29-3.69) and CRP (HR = 2.58, 95% CI = 1.52-4.40) were associated with a significantly greater risk of death, whereas the association between TNF-α protein levels and mortality was no longer significant (HR = 1.26, 95% CI = 0.74-2.15). The composite summary score of inflammation was strongly associated with mortality, with the highest risk estimated for individuals with all three inflammatory markers above the median.

Conclusion: Low levels of inflammatory markers are associated with better survival in older adults, independent of age and other clinical and functional variables.

Low-grade inflammatory indicators are widely used in the diagnosis and prognosis of different diseases, particularly, in cancers. The establishing of reference intervals (RIs) are necessary to provide baselines for the interpretation and application of the indicators in clinical. A retrospective review was organized to include ostensibly healthy subjects aged >18 years old to establish RIs based on gender and age for systemic immune-inflammation index (SII), neutrophil-to-lymphocyte ratio (NLR), platelet-to-lymphocyte ratio (PLR), mean platelet volume-to-platelet ratio (MPV/PLT), MPV and red blood cell distribution width - standard deviation (RDW-SD). Another 5012 healthy persons were included to validate the RIs. The differences among gender, age, and Body Mass Index (BMI) groups were compared to assess their influences on each indicator. A total of 26,242 males and 11,934 females were included. All indicators differed between males and females except NLR. Compared with males, the values of SII, PLR, MPV and RDW-SD were higher while MPV/PLT lower in females. The values of SII, NLR and PLR increased with age in females between 18 and 50 years old, declined in 51-60 years old group, then increased with age again. Values of MPV/PLT and RDW-SD increased with age. The values of SII increased with BMI while PLR, MPV, and MPV/PLT decreased. We effectively established RIs for the indicators. They will help with predicting disease progression in various clinical practices. Gender, age, and BMI were suggested to be taken into consideration when these parameters were applied in clinical.

Background: In recent years, increasing attention has been focused on the impact of red blood cell indices (RCIs) on disease prognosis. We aimed to investigate the association of mean corpuscular hemoglobin (MCH), mean corpuscular hemoglobin concentration (MCHC), and mean corpuscular volume (MCV) with mortality.

Methods: The study used cohort data from U.S. adults who participated in the 1999-2008 National Health and Nutrition Examination Survey. All-cause mortality was the primary outcome during follow-up, with secondary cardiovascular mortality outcomes. COX regression was applied to analyze the connection between RCIs and mortality. We adopted three models to minimize potential bias. Smooth-fit curves and threshold effect analyses were utilized to observe the dose-response relationship between RCIs and all-cause and cardiovascular mortality. In addition, we performed sensitivity analyses.

Results: 21,203 individuals were enrolled in our research. During an average 166.2 ± 54.4 months follow-up, 24.4% of the population died. Curve fitting indicated a U-shaped relationship between MCV and MCH with all-cause mortality, and the relationship of MCHC to all-cause mortality is L-shaped. We identified inflection points in the relationship between MCV, MCH, and MCHC and all-cause mortality as 88.56732 fl, 30.22054 pg, 34.34624 g/dl (MCV <88.56732 fl, adjusted HR 0.99, 95 CI% 0.97-1.00; MCV >88.56732 fl, adjusted HR 1.05, 95 CI% 1.04-1.06. MCH <30.22054 pg, adjusted HR 0.95, 95 CI% 0.92-0.98; MCH >30.22054 pg, adjusted HR 1.08, 95 CI% 1.04-1.12. MCHC <34.34624 g/dl, adjusted HR 0.88, 95 CI% 0.83-0.93). Besides, the MCV curve was U-shaped in cardiovascular mortality (MCV <88.56732 fl, adjusted HR 0.97, 95 CI% 0.94-1.00; MCV >88.56732 fl, adjusted HR 1.04, 95 CI% 1.01-1.06).

Conclusion: This cohort study demonstrated that RCIs (MCH, MCHC, and MCV) were correlated with mortality in the general population. Three RCIs were nonlinearly correlated with all-cause mortality. In addition, there were nonlinear relationships between MCH and MCV and cardiovascular mortality.

Red blood cells (RBCs) have emerged as biomarkers of the aging process as they undergo several changes in human aging and age-related diseases. The objectives of our study are to explore the effect of human aging on RBC indices, the strengths, therapeutic interventions, challenges, and future directions for using RBCs as a biomarker. Two online databases, PubMed and ScienceDirect, were used to search relevant studies using "RBCs as biomarkers of human aging," "red blood cells [MeSH Terms] AND biomarkers [MeSH Terms] AND human aging [MeSH Terms]," and "erythrocytes and human aging" as keywords. A total of 474 studies were identified, and after the removal of duplicates, excluding studies based on title, abstract, or full text, 74 studies and 48 additional studies found through cross-referencing were included in this systematic review. Based on the evidence, we concluded that RBC indices such as hemoglobin concentration, mean corpuscular volume, RBC distribution width, RBC membrane, oxidative stress, and metabolism change with human aging. Several studies have applied therapeutic interventions to RBCs, including dietary supplementation, phytochemicals, nanoparticles, and physical activity, to mitigate aging and related outcomes. Hence, the quality of life for older people and healthy aging can be improved by further investigating the RBC parameters, molecular mechanisms, and their implications for age-related health consequences.

Background: The normal ranges for clinical chemistry tests are usually defined by cut-offs given by the distribution in healthy individuals. This approach does however not indicate if individuals outside the normal range are more prone to disease.

Methods: We studied the associations and risk prediction of 11 plasma and serum biomarkers with all-cause mortality in two population-based cohorts: a Swedish cohort (X69) initiated in 1969, and the UK Biobank (UKB) initiated in 2006-2010, with up to 48- and 9-years follow-up, respectively.

Results: In X69 and in UKB, 18,529 and 425,264 individuals were investigated, respectively. During the follow-up time, 14,475 deaths occurred in X69 and 17,116 in UKB. All evaluated tests were associated with mortality in X69 (P<0.0001, except bilirubin P<0.005). For calcium, blood urea nitrogen, bilirubin, hematocrit, uric acid, and iron, U-shaped associations were seen (P<0.0001). For leukocyte count, gamma-glutamyl transferase, alkaline phosphatases and lactate dehydrogenase, linear positive associations were seen, while for albumin the association was negative. Similar associations were seen in UKB. Addition of all biomarkers to a model with classical risk factors improved mortality prediction (delta C-statistics: +0.009 in X69 and +0.023 in UKB, P<0.00001 in both cohorts).

Conclusions: Commonly used clinical chemistry tests were associated with all-cause mortality both in the medium- and long-term perspective, and improved mortality prediction beyond classical risk factors. Since both linear and U-shaped relationships were found, we propose to define the normal range of a clinical chemistry test based on its association with mortality, rather than from the distribution.

Vitamin D deficiency is one of the most common micronutrient deficiencies in the United States. An analysis of data from 71,685 participants in NHANES (2001 to 2018) showed that vitamin D levels were less than 25 nmol per liter, 25 to 50 nmol per liter, and 50 to 75 nmol per liter in 2.6%, 22.0%, and 40.9% of participants, respectively. A severe or moderate deficiency was more prevalent among women and non-Hispanic Black Americans; during winter, when ultraviolet B radiation is insufficient to synthesize the vitamin in skin at northern latitudes; and among persons 20 to 29 years of age. Behaviors that screened out sunlight and low milk intake also predicted a severe deficiency. The only good dietary sources of vitamin D are oily fish, eggs, and fortified foods, including dairy products and breakfast cereals.

There is still debate about the cutoff values for serum concentrations of the vitamin D biomarker (25-hydroxyvitamin D), but the National Academy of Medicine (known as the Institute of Medicine before 2015), National Osteoporosis Foundation, and American Geriatrics Society specify a level of less than 30 nmol per liter as deficiency. For most people, a level of 50 nmol per liter is adequate, although the Endocrine Society recommends a level of 75 nmol per liter to maximize the benefits for bone and muscle. The National Academy of Medicine recommends an intake of 600 IU of vitamin D per day for persons 1 to 70 years of age and 800 IU per day for those who are 71 years of age or older.

When health outcomes were compared for 25,000 people in the United States who took 2000 IU of vitamin D per day or placebo for 5 years, vitamin D supplementation did not help prevent cancer, heart disease, or depression or, surprisingly, preserve bone mineral density or prevent fractures. However, an updated meta-analysis suggests that vitamin D supplementation reduced total mortality from cancer by 25% in analyses that excluded the first 2 years of follow-up.

1. A true healthy normal ALT level in prospectively studied populations without identifiable risk factors for liver disease ranges from 29 to 33 IU/l for males and 19 to 25 IU/l for females, and levels above this should be assessed by physicians.
2. Elevated ALT or AST above the upper limit of normal (ULN) in a population without identifiable risk factors is associated with increased liver-related mortality.
3. There is a linear relationship between ALT level and body mass index (BMI) that should be assessed by physicians.
4. A normal ALT level may not exclude significant liver disease.
5. ALT levels are higher in males than females.
6. AST and ALT ULN ranges can vary between different labs.
7. Clinicians may rely on local lab ULN ranges for alkaline phosphatase and bilirubin.
Normal lab values are generally defined as the mean value of a healthy population±2 s.d.’s. This incorporates 95% of subjects. By definition, 2.5% of the population will be greater than the ULN of the reference population. For alkaline phosphatase and bilirubin levels, establishing normal liver enzyme levels that can be replicated across different reference labs has not been reported as problematic, which differs from the wide variations in ranges reported as normal for ALT levels. However, establishing normal ranges for ALT and AST levels have been problematic due to differences in the definition of healthy control populations that are used to establish the normal reference ranges. One report examined the local reference laboratory ranges for ALT used by the non-alcoholic steatohepatitis (NASH) Clinical Research Network and demonstrated significant differences in the defined ALT ULN (range 35–79 IU/l for men and 31–55 IU/l for women) (13). These wide ranges appeared to be due to the use of different reference populations utilized by the different laboratories with local populations used to establish the normal range of ALT, apparently without consideration of factors such as BMI.

Comparing biomarker profiles measured at similar ages, but earlier in life, among exceptionally long-lived individuals and their shorter-lived peers can improve our understanding of aging processes. This study aimed to (i) describe and compare biomarker profiles at similar ages between 64 and 99 among individuals eventually becoming centenarians and their shorter-lived peers, (ii) investigate the association between specific biomarker values and the chance of reaching age 100, and (iii) examine to what extent centenarians have homogenous biomarker profiles earlier in life. Participants in the population-based AMORIS cohort with information on blood-based biomarkers measured during 1985-1996 were followed in Swedish register data for up to 35 years. We examined biomarkers of metabolism, inflammation, liver, renal, anemia, and nutritional status using descriptive statistics, logistic regression, and cluster analysis. In total, 1224 participants (84.6% females) lived to their 100th birthday. Higher levels of total cholesterol and iron and lower levels of glucose, creatinine, uric acid, aspartate aminotransferase, gamma-glutamyl transferase, alkaline phosphatase, lactate dehydrogenase, and total iron-binding capacity were associated with reaching 100 years. Centenarians overall displayed rather homogenous biomarker profiles. Already from age 65 and onwards, centenarians displayed more favorable biomarker values in commonly available biomarkers than individuals dying before age 100. The differences in biomarker values between centenarians and non-centenarians more than one decade prior death suggest that genetic and/or possibly modifiable lifestyle factors reflected in these biomarker levels may play an important role for exceptional longevity.

The global increase in life expectancy has sparked growing interest in the factors that contribute to exceptional longevity. Between 1990 and 2015, the number of centenarians worldwide more than quadrupled. This study aimed to analyse the relationship between blood-based biomarkers and the likelihood of reaching 100 years of age in Catalonia (2015-2022), and to examine how biomarker variations during COVID-19 affected longevity. Using a retrospective cohort study based on primary care electronic health records from Catalonia, we compared centenarians with individuals aged 92 or older who died before reaching 100 years of age. We analysed anaemia, cholesterol, glycemia, kidney function, and liver function biomarkers. We employed multiple strategies to control for confounding including matching without replacement, adjusting for both observed confounders at both the individual and contextual level, and unobserved confounders, in particular spatial dependence. Our findings reveal that centenarians exhibit higher rates of chronic conditions, greater socioeconomic disadvantage, and increased neighbourhood inequality in urban areas. Biologically, longevity was linked to intermediate levels of ferritin and cholesterol, alongside lower glucose, creatinine, and uric acid levels. Glycaemic balance, indicated by HbA1c and fasting glucose, emerged as a key factor in survival to extreme old age. Additionally, biomarker improvements during the pandemic correlated with an increased likelihood of reaching centenarian age. These results emphasize the complex interplay between biological, behavioural, and contextual factors in determining longevity. While biomarkers provide valuable insights, they are insufficient indicators of healthy ageing. Future research should integrate multiple dimensions, among them, environmental, and social determinants for uncovering the mechanisms of longevity.

Clinical laboratories now routinely report estimated GFR (eGFR) based on filtration markers. The most common filtration marker used is creatinine, a 113 dalton byproduct of creatine metabolism and one for which laboratory assays have been standardized since 2003. The preferred estimating equation in the United States and much of the world is the CKD-EPI 2009 creatinine equation, which is more accurate than the earlier MDRD equation, particularly for eGFR values greater than 60 mL/min/1.73 m 2 ( https://www.kidney.org/professionals/kdoqi/gfr_calculator ). In situations requiring additional accuracy and precision, cystatin C can be used with creatinine in the CKD-EPI 2012 creatinine-cystatin C equation. Adding cystatin C may be particularly useful for individuals with altered creatinine production and/or metabolism (eg, extremely high or low body size or muscle mass, limb amputation, high-protein diet, use of creatinine supplements, or use of drugs affecting tubular secretion of creatinine).

Albuminuria should ideally be quantified by a urine ACR. Albuminuria staging is classified as A1 (urine ACR <30 mg/g), A2 (30-300 mg/g), and A3 (>300 mg/g). Guidelines recommend the use of urine ACR to stage CKD rather than urine protein-to-creatinine ratio because assays for the former are more likely to be standardized and have better precision at lower values of albuminuria. The most precise measurements come from a first morning sample or 24-hour collection, as there is high biological variability in urine albumin excretion over the course of the day. Random samples, however, are also acceptable in initial screening. Compared with urine protein-to-creatinine ratio, urine ACR is believed to be a more sensitive and specific marker of glomerular pathology since some urine proteins such as uromodulin are present (and may even be protective) in normal physiology. If tubular or overflow proteinuria is suspected, then urine protein electrophoresis or testing for the specific protein can be pursued (eg, immunoglobulin heavy and light chains, α 1 -microglobulin, and β 2 -microglobulin).

Kidney function, often assessed by estimated glomerular filtration rate (eGFR), declines naturally with age. However, there is a lack of eGFR reference values to describe normal and abnormal values for a specific age. The European Chronic Kidney Disease Burden Consortium is comprised of nine participating general population-based studies from seven European countries which provides European age- and sex-specific eGFR reference values in healthy adults using the European Kidney Function Consortium (EKFC) equation. Of 2,572,020 individuals, 1,535,253 (60%) were considered healthy, of which 45% were men. Ages ranged from 18 to 105 years old in men and 18 to 107 years old in women with a median age of 43 years in both sexes. At age 20 in men, the 5th, 50th and 95th eGFR percentiles were 78 ml/min per 1.73 m2, 99 ml/min per 1.73 m2, and 119 ml/min per 1.73 m2. In 20-year-old women this was 81 ml/min per 1.73 m2, 101 ml/min per 1.73 m2, and 121 ml/min per 1.73 m2. Consequently, in men aged 80 years old, the 5th, 50th and 95th eGFR percentiles were 49 ml/min per 1.73 m2, 66 ml/min per 1.73 m2, and 84 ml/min per 1.73 m2. In 80 year old women this was 46 ml/min per 1.73 m2, 63 ml/min per 1.73 m2, and 81 ml/min per 1.73 m2. Overall, our study shows that eGFR is not preserved with ageing in healthy individuals and these eGFR reference values can help determine abnormal and normal kidney function across the age range.

Chronic kidney disease (CKD) is a worldwide public health challenge and results in substantial morbidity, mortality, and health care costs. In adults, CKD is diagnosed based on the presence of abnormalities of kidney structure or function (ie, abnormal albuminuria or estimated glomerular filtration rate [eGFR]) for more than 3 months, with implications for health. The development of a uniform definition of CKD has been a useful paradigm that serves as a foundation for targeted screening and more precise diagnosis of CKD, appropriate patient care, and health care resource planning. Values of eGFR that are less than the threshold of 60 mL/min/1.73 m 2 indicate CKD because they reflect an average loss of at least 50% of the kidney function in a healthy young adult. Many large studies have found that, starting from this threshold, lower eGFR values are associated with a graded increase in the relative risks of adverse outcomes in adults, such as kidney failure, cardiovascular events, and mortality across all age categories.

Although there is consensus that persistent presence of abnormal albuminuria indicates CKD, the appropriateness of using a single eGFR threshold to define CKD, regardless of albuminuria, has been debated. Because eGFR declines with advancing age, a definition based on a fixed eGFR threshold may lead to underdiagnosis in young individuals and overdiagnosis in elderly individuals, whose eGFR physiologically declines with aging. A fixed-threshold definition may result in overestimation of the CKD burden in an aging population, largely attributable to people who may not have increased risks of adverse outcomes. An age-adapted CKD definition has been proposed, with eGFR thresholds of 75, 60, and 45 mL/min/1.73 m 2 for younger than 40, 40 to 64, and 65 years or older, respectively. The differences in size and outcomes of the CKD population identified using the 2 definitions are unknown. It is also unclear to what extent the outcomes of people who have CKD only according to the current fixed eGFR threshold definition may differ from those who do not have CKD.

Laboratories should ideally determine their own reference interval according to CLSI guidelines (CLSI Document C28A) even if the manufacturer has provided one. If a laboratory chooses to establish its own reference interval, test subjects without known diabetes should not have obesity and should have FPG <5.6 mmol/L (100 mg/dL) and, ideally, a 2-h glucose <11.1 mmol/L (200 mg/dL) during an OGTT. For many years, HbA 1c reference intervals were 4% to 6% (20 to 42 mmol/mol). This reflected mean ± 2 SD. Improvements in assay accuracy now allow a narrower range. For assay methods that are NGSP certified, reference intervals should not deviate substantially (e.g., >0.5%) from a mean of 5% (31 mmol/mol) i.e., 4.5% to 5.5% (26 to 37 mmol/mol). Many organizations and laboratories have lowered the upper limit of the reference interval to 5.6% (31 mmol/mol). Note that treatment target values recommended by the ADA and other clinical organizations, not the reference intervals, are used to evaluate metabolic control and diagnostic cutoffs.

Objectives: To determine reference values for laboratory tests in individuals aged 85 and older.

Design: Cross-sectional cohort study.

Setting: International.

Participants: Long Life Family Study (LLFS) participants (N~5,000, age: range 25-110, median 67, 45% male).

Measurements: Serum biomarkers were selected based on association with aging-related diseases and included complete blood count, lipids (triglycerides, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, total cholesterol), 25-hydroxyvitamin D2 and D3, vitamin D epi-isomer, diabetes mellitus-related biomarkers (adiponectin, insulin, insulin-like growth factor 1, glucose, glycosylated hemoglobin, soluble receptor for advanced glycation endproduct), kidney disease-related biomarkers (albumin, creatinine, cystatin), endocrine biomarkers (dehydroepiandrosterone, sex-hormone binding globulin, testosterone), markers of inflammation (interleukin 6, high-sensitivity C-reactive protein, N-terminal pro b-type natriuretic peptide), ferritin, and transferrin.

Results: Of 38 measured biomarkers, 34 were significantly correlated with age. Summary statistics were generated for all biomarkers according to sex and 5-year age increments from 50 and up after excluding participants with diseases and treatments that were associated with biomarkers. A biomarker data set was also generated that will be useful for other investigators seeking to compare biomarker levels between studies.

Conclusion: Levels of several biomarkers change with older age in healthy individuals. The descriptive statistics identified herein will be useful in future studies and, if replicated in additional studies, might also become useful in clinical practice. The availability of the reference data set will facilitate appropriate calibration of biomarkers measured in different laboratories.

Background: Reference intervals of thyroid-stimulating hormone (TSH) and free thyroxine (FT4) are statistically defined by the 2·5-97·5th percentiles, without accounting for potential risk of clinical outcomes. We aimed to define the optimal healthy ranges of TSH and FT4 based on the risk of cardiovascular disease and mortality.

Methods: This systematic review and individual participant data (IPD) meta-analysis identified eligible prospective cohorts through the Thyroid Studies Collaboration, supplemented with a systematic search via Embase, MEDLINE (Ovid), Web of science, the Cochrane Central Register of Controlled Trials, and Google Scholar from Jan 1, 2011, to Feb 12, 2017 with an updated search to Oct 13, 2022 (cohorts found in the second search were not included in the IPD). We included cohorts that collected TSH or FT4, and cardiovascular outcomes or mortality for adults (aged ≥18 years). We excluded cohorts that included solely pregnant women, individuals with overt thyroid diseases, and individuals with cardiovascular disease. We contacted the study investigators of eligible cohorts to provide IPD on demographics, TSH, FT4, thyroid peroxidase antibodies, history of cardiovascular disease and risk factors, medication use, cardiovascular disease events, cardiovascular disease mortality, and all-cause mortality. The primary outcome was a composite outcome including cardiovascular disease events (coronary heart disease, stroke, and heart failure) and all-cause mortality. Secondary outcomes were the separate assessment of cardiovascular disease events, all-cause mortality, and cardiovascular disease mortality. We performed one-step (cohort-stratified Cox models) and two-step (random-effects models) meta-analyses adjusting for age, sex, smoking, systolic blood pressure, diabetes, and total cholesterol. The study was registered with PROSPERO, CRD42017057576.

Findings: We identified 3935 studies, of which 53 cohorts fulfilled the inclusion criteria and 26 cohorts agreed to participate. We included IPD on 134 346 participants with a median age of 59 years (range 18-106) at baseline. There was a J-shaped association of FT4 with the composite outcome and secondary outcomes, with the 20th (median 13·5 pmol/L [IQR 11·2-13·9]) to 40th percentiles (median 14·8 pmol/L [12·3-15·0]) conveying the lowest risk. Compared with the 20-40th percentiles, the age-adjusted and sex-adjusted hazard ratio (HR) for FT4 in the 80-100th percentiles was 1·20 (95% CI 1·11-1·31) for the composite outcome, 1·34 (1·20-1·49) for all-cause mortality, 1·57 (1·31-1·89) for cardiovascular disease mortality, and 1·22 (1·11-1·33) for cardiovascular disease events. In individuals aged 70 years and older, the 10-year absolute risk of composite outcome increased over 5% for women with FT4 greater than the 85th percentile (median 17·6 pmol/L [IQR 15·0-18·3]), and men with FT4 greater than the 75th percentile (16·7 pmol/L [14·0-17·4]). Non-linear associations were identified for TSH, with the 60th (median 1·90 mIU/L [IQR 1·68-2·25]) to 80th percentiles (2·90 mIU/L [2·41-3·32]) associated with the lowest risk of cardiovascular disease and mortality. Compared with the 60-80th percentiles, the age-adjusted and sex-adjusted HR of TSH in the 0-20th percentiles was 1·07 (95% CI 1·02-1·12) for the composite outcome, 1·09 (1·05-1·14) for all-cause mortality, and 1·07 (0·99-1·16) for cardiovascular disease mortality.

Interpretation: There was a J-shaped association of FT4 with cardiovascular disease and mortality. Low concentrations of TSH were associated with a higher risk of all-cause mortality and cardiovascular disease mortality. The 20-40th percentiles of FT4 and the 60-80th percentiles of TSH could represent the optimal healthy ranges of thyroid function based on the risk of cardiovascular disease and mortality, with more than 5% increase of 10-year composite risk identified for FT4 greater than the 85th percentile in women and men older than 70 years. We propose a feasible approach to establish the optimal healthy ranges of thyroid function, allowing for better identification of individuals with a higher risk of thyroid-related outcomes.

Funding: None.

Background: Variations in thyroid function within reference ranges are associated with increased risk of diseases and death. However, the impact of thyroid function on life expectancy (LE) with and without noncommunicable diseases (NCDs) remains unknown. We therefore aimed to investigate the association of thyroid function with total LE and LE with and without NCD among euthyroid individuals.

Methods And Findings: The study was embedded in the Rotterdam Study, a prospective population-based study carried out in the Netherlands. In total, 7,644 participants without known thyroid disease and with thyroid-stimulating hormone (TSH) and free thyroxine (FT4) levels within reference ranges were eligible. NCDs were defined as presence of cardiovascular disease, diabetes mellitus type 2, or cancer. We used the demographic tool of multistate life tables to calculate LE estimates at the age of 50 years, using prevalence, incidence rates, and hazard ratios for three transitions (healthy to NCD, healthy to death, and NCD to death). The total LE and LE with and without NCD among TSH and FT4 tertiles were calculated separately in men and women. Analyses were adjusted for sociodemographic and cardiovascular risk factors. The mean (standard deviation) age of the participants was 64.5 (9.7) years, and 52.3% were women. Over a median follow-up of 8 years (interquartile range 2.7-9.9 years), 1,396 incident NCD events and 1,422 deaths occurred. Compared with those in the lowest TSH tertile, men and women in the highest TSH tertile were expected to live 1.5 years (95% confidence interval [CI] 0.8-2.3, p < 0.001) and 1.5 years (CI 0.8-2.2, p < 0.001) longer, respectively, of which 1.4 years (CI 0.5-2.3, p = 0.002) and 1.3 years (CI 0.3-2.1, p = 0.004) with NCD. Compared with those in the lowest FT4 tertile, the difference in LE for men and women in the highest FT4 tertile was -3.7 years (CI -5.1 to -2.2, p < 0.001) and -3.3 years (CI -4.7 to -1.9, p < 0.001), respectively, of which -1.8 years (CI -3.1 to -0.7, p = 0.003) and -2.0 years (CI -3.4 to -0.7, p = 0.003) without NCD. A limitation of the study is the observational design. Thus, the possibility of residual confounding cannot be entirely ruled out.

Conclusions: In this study, we found that people with low-normal thyroid function (i.e., highest tertile of TSH and lowest tertile of FT4 reference ranges) are expected to live more years with and without NCD than those with high-normal thyroid function (i.e., lowest tertile of TSH and highest tertile of FT4 reference ranges). These findings provide support for a re-evaluation of the current reference ranges of thyroid function.

Background: Current clinical practice uses a one-size-fits-all approach to define reference intervals for the results of diagnostic tests about thyroid function. This approach does not recognize subgroup differences according to age, sex, or race.

Objective: To identify age-, sex-, and race-specific reference intervals for the common diagnostic tests that measure thyroid function and to examine how these new reference intervals reclassify persons into disease categories when compared with current reference intervals.

Design: Cross-sectional analysis.

Setting: Data from the U.S. NHANES (National Health and Nutrition Examination Survey) supplemented with data from a multicenter Chinese study.

Participants: A nationally representative sample from NHANES aged 20 years or older (n = 8308) supplemented with a Chinese database of routine health checkups from 49 hospitals in 10 provinces aged 18 years or older (n = 314 302).

Measurements: The thyroid function reference interval was defined as the interval of diagnostic indicator levels from the 2.5th (lower limit) to the 97.5th (upper limit) percentile by age, sex, and race subgroups.

Results: In 8308 NHANES participants, the 97.5th percentile levels of thyroid-stimulating hormone (TSH) increased with age, whereas total triiodothyronine (TT3) levels declined with age and total thyroxine (TT4) levels were stable across different ages. Women had higher TT4 levels, and White participants had higher TSH levels. Using current reference intervals, the prevalence of subclinical hypothyroidism increased from 2.4% for ages 20 to 29 years to 5.9% for ages 70 years and older. In contrast, using age-, sex-, and race-specific reference intervals reclassified 48.5% of persons with subclinical hypothyroidism as normal, especially women and White participants, and reclassified 31.2% of persons with subclinical hyperthyroidism as normal, especially women, Black participants, and Hispanic participants. When compared with the findings from U.S. participants, many of the findings from 314 302 Chinese participants were similar.

Limitation: Cross-sectional data; sample size limitations for subgroup.

Conclusion: These findings should help establish more accurate reference intervals for thyroid diseases and facilitate development of a consensus about how to define and manage those diseases.

Primary Funding Source: National Key Research and Development Program of China and National Natural Science Foundation.

Background: Thyroid-stimulating hormone (TSH) and subsequent free thyroxine (FT4) concentrations outside the reference interval (RI) are used to diagnose thyroid diseases. Most laboratories do not provide age-specific RIs for TSH and FT4 beyond childhood, although TSH concentrations vary with age. Therefore, we aimed to establish TSH and FT4 age-specific RIs throughout life and aimed to determine whether using these RIs would result in reclassification of thyroid disease diagnoses in adults. Methods: This multicenter retrospective cross-sectional study used big data to determine indirect RIs for TSH and FT4. These RIs were determined by TMC and refineR-analysis, respectively, using four different immunoassay platforms (Roche, Abbott, Siemens, and Beckman Coulter). Retrospective data (2008-2022) from 13 Dutch laboratories for general practitioners and local hospitals were used. RIs were evaluated per manufacturer. Age groups were established from 2 to 20 years by 2-year categories and decade categories between 20 and 100 years. Results: We included totally 7.6 million TSH and 2.2 million FT4 requests. TSH upper reference limits (URLs) and FT4 lower reference limits were higher in early childhood and decreased toward adulthood. In adulthood, TSH URLs increased from 60 years in men, and from 50 years in women, while FT4 URLs increased from 70 years onward. Using adult age-specific RIs resulted in a decrease in diagnoses of subclinical and overt hypothyroidism in women above 50 and men above 60 years in our Roche dataset. Conclusion: This study stressed the known importance of using age-specific RIs for TSH and FT4 in children. This study also showed the clinical relevance of using age-specific RIs for TSH in adulthood to reduce diagnoses of subclinical hypothyroidism in older persons. Therefore, implementation of adult TSH age-specific RIs should be strongly considered. Data are less uniform regarding FT4 age-specific RIs and more research should be performed before implementing these in clinical practice.

Background: Many studies have reported that the thyroid-stimulating hormone (TSH) reference interval is susceptible to external factors, such as age, sex, race, region and iodine intake. However, no meta-analysis has comprehensively explored the effect of these factors on the TSH reference interval.

Methods: Articles published from January 1960 to January 2020 were searched in PubMed, Embase, Cochrane, Scopus, Medline English databases and CNKI, WanFang and CQVIP Chinese databases. In total, 19 studies were ultimately included. All data were analysed using Review Manager 5.3, STATA 16.0 software, GraphPad Prism 8.0 and Microsoft Excel 2010 to draw the TSH concentration curve.

Results: The TSH reference interval was significantly influenced by sex and age. The mean of TSH concentration in females was 0.27 mIU/L higher than that in males. Reference interval of TSH is divided into 20-59 years old and >60 years old age groups in males, and 20-39 years old and >40 years old age groups in females. Regardless of sex, TSH concentrations all increase with age. In iodine-deficient areas, TSH reference intervals were generally lower than those in iodine-sufficient or iodine-excessive areas. The TSH reference interval in Asia and North American countries was generally higher than that in most European countries. In the subgroup analyses of sample size, region and assay methods and manufacturers, the between-group differences were significant.

Conclusion: The TSH reference interval was significantly influenced by sex, age, iodine intake, sample size, region, and assay methods and manufacturers, but other factors should not be ignored. Therefore, it is necessary for each laboratory to validate an appropriate TSH reference interval based on local conditions.

Hypothyroidism is characterized by the relative or absolute reductions in serum levels of thyroxine (T4). Serum levels of total or free triiodothyronine (T3), however, are not reliable markers of hypothyroidism. Thyroid-stimulating hormone (TSH) is elevated in primary hypothyroidism. However, TSH is typically low or inappropriately normal in central hypothyroidism, which may be caused by pituitary disease (secondary hypothyroidism) or hypothalamic disorders (tertiary hypothyroidism). Patients with nonthyroidal chronic illness can have decreased serum levels of T3, T4, and TSH, and generally lack hypothyroid symptoms. Patients with generalized thyroid hormone resistance usually have high concentrations of T3 and T4, with inappropriately normal or elevated TSH and variable degrees of hypothyroid and hyperthyroid symptoms. Subclinical hypothyroidism is characterized by normal T3 and T4, with an elevated TSH (<10 mIU/L) and few, if any, symptoms attributable to hypothyroidism.
Complicating these diagnoses are data suggesting that euthyroid elderly patients have higher TSH values that continue to rise with increasing age and are especially notable as patients reach and continue through their 9 th decade ( 224 ). The significance of this is unclear, but there may be a genetic component, since children of nonagenarians with elevated TSH have higher TSH levels than age-matched controls ( 225 ).
Thyroid dysfunction has major effects on lipoprotein metabolism. Hypothyroidism is associated with reduced LDL receptors, reduced cholesterol 7 alpha-hydroxylase, low cholesterol ester transfer protein, and decreased lipoprotein lipase, which, together, may lead to an elevation in total cholesterol, TG, LDL-C, and apoB. High-density lipoprotein cholesterol has been reported to increase, remain unchanged, or decrease. One study found increased HDL-C in hypothyroid patients, but reduced cholesterol efflux ( 226 ).

Background: Whether circulating sex hormones modulate mortality and cardiovascular disease (CVD) risk in aging men is controversial.

Purpose: To clarify associations of sex hormones with these outcomes.

Data Sources: Systematic literature review to July 2019, with bridge searches to March 2024.

Study Selection: Prospective cohort studies of community-dwelling men with sex steroids measured using mass spectrometry and at least 5 years of follow-up.

Data Extraction: Independent variables were testosterone, sex hormone-binding globulin (SHBG), luteinizing hormone (LH), dihydrotestosterone (DHT), and estradiol concentrations. Primary outcomes were all-cause mortality, CVD death, and incident CVD events. Covariates included age, body mass index, marital status, alcohol consumption, smoking, physical activity, hypertension, diabetes, creatinine concentration, ratio of total to high-density lipoprotein cholesterol, and lipid medication use.

Data Synthesis: Nine studies provided individual participant data (IPD) (255 830 participant-years). Eleven studies provided summary estimates (n = 24 109). Two-stage random-effects IPD meta-analyses found that men with baseline testosterone concentrations below 7.4 nmol/L (<213 ng/dL), LH concentrations above 10 IU/L, or estradiol concentrations below 5.1 pmol/L had higher all-cause mortality, and those with testosterone concentrations below 5.3 nmol/L (<153 ng/dL) had higher CVD mortality risk. Lower SHBG concentration was associated with lower all-cause mortality (median for quintile 1 [Q1] vs. Q5, 20.6 vs. 68.3 nmol/L; adjusted hazard ratio [HR], 0.85 [95% CI, 0.77 to 0.95]) and lower CVD mortality (adjusted HR, 0.81 [CI, 0.65 to 1.00]). Men with lower baseline DHT concentrations had higher risk for all-cause mortality (median for Q1 vs. Q5, 0.69 vs. 2.45 nmol/L; adjusted HR, 1.19 [CI, 1.08 to 1.30]) and CVD mortality (adjusted HR, 1.29 [CI, 1.03 to 1.61]), and risk also increased with DHT concentrations above 2.45 nmol/L. Men with DHT concentrations below 0.59 nmol/L had increased risk for incident CVD events.

Limitations: Observational study design, heterogeneity among studies, and imputation of missing data.

Conclusion: Men with low testosterone, high LH, or very low estradiol concentrations had increased all-cause mortality. SHBG concentration was positively associated and DHT concentration was nonlinearly associated with all-cause and CVD mortality.

Primary Funding Source: Medical Research Future Fund, Government of Western Australia, and Lawley Pharmaceuticals. (

Prospero: CRD42019139668).

Background: While abundant research suggests a sex-specific role of endogenous sex steroid hormones in chronic diseases, research on mortality remains inconclusive. We quantified the sex-specific associations of endogenous sex steroid hormones including total testosterone (TT), free testosterone, bioavailable testosterone, estradiol (E2), dehydroepiandrosterone, and dehydroepiandrosterone-sulphate and sex hormone binding globulin (SHBG) with risk of all-cause and cause-specific mortality in the general population.

Methods: Embase, Medline, Web of Science, and Cochrane Central were searched and population-based cohort studies investigating the association of interested were included. The risk of bias was assessed using the ROBINS-E tool. The certainty of evidence was evaluated using GRADE framework. Pooled hazard ratios (HRs) and 95% confidence intervals (CI) were calculated using a random effects model for the top versus bottom tertile of sex hormones and risk of mortality.

Results: 53 publications with 359,047 participants were included in the systematic review. A significant association was observed between higher level of TT and risk of all-cause mortality (HR (95%CI): 0.89 (0.83 to 0.97), n=19 studies) in men, while no association was found in women. Dose-response analysis suggested a significant U-shaped association between TT and all-cause mortality in men and a J-shaped association in women. Higher level of SHBG was significantly associated with higher risk of all-cause mortality in women (1.25 (1.13 to 1.39), n=3) and no association was observed in men. Additionally, higher DHEAs levels were associated with lower risk of all-cause mortality in men (0.72 (0.57 to 0.91), n=6) and no association was observed in women.

Conclusions: This meta-analysis reveals a dose-response link between endogenous sex steroid hormones and mortality, highlighting the need for sex-specific studies on hormone modulation's impact on mortality and longevity.

Source Of Funding: None.

Prospero Registration Code: CRD42022329605.

Background And Aims: Availability of age- and sex-specific reference values for sex steroids and sex steroid-binding globulin (SHBG) levels allows for appropriate interpretation of research findings and their clinical applications. We report the sex-specific distribution and reference levels of sex steroids, including total estradiol, total testosterone and (calculated) free androgen index (cFAI), SHBG and other androgens dehydroepiandrosterone (DHEA), dehydroepiandrosterone sulphate (DHEAS) and androstenedione across age.

Methods: Using data from 3291 participants from the prospective population-based Rotterdam Study (2006-2008), we visualised the distribution of sex steroids and SHBG levels by calculating and depicting the 5th, 25th, 50th, 75th and 95th percentiles per year and per age-year across 5-year age bands to provide reference value ranges in men and women. Total estradiol and SHBG were measured using automated immunoassay and androgens using liquid chromatography-mass spectrometry (LC-MS/MS).

Result: Mean age was 56.8 (range 45.6-79.9) years in men and 56.9 (range 45.7-79.9) years in women. Amongst men, total estradiol and SHBG showed an increasing trend from 45 years onwards. In women, total estradiol and SHBG showed a decreasing trend from 45 years until the age of 60. From 60 years onwards, SHBG showed an increasing trend. For total testosterone, a clear declining trend was observed amongst men but not women. Other androgens showed a similar decreasing trend in both sexes from 45 years onwards.

Discussion And Conclusion: Our study underlines sex-specific trends in sex steroids and SHBG levels with ageing. This warrants taking into account sex- and age-specific reference values for sex steroids and SHBG when investigating their impact on health outcomes to prevent controversial results and allow for their appropriate clinical application.

LDL-C management should be individualized based on each patient's specific situation. Evidence supports an LDL-C target of <100 mg/dL for healthy adults in the general population. Individuals at risk for ASCVD, including those with very high baseline LDL-C levels suggestive of a genetic cause such as familial hypercholesterolemia (FH), individuals with multiple risk factors for ASCVD, or those already diagnosed with ASCVD, may require lower LDL-C levels. Higher plaque burden and higher plaque progression rate are situations where more intensive LDL-C lowering therapy should be considered.

The primary objective for individuals with TG levels ≥500 mg/dL is to achieve and maintain a TG concentration <500 mg/dL to reduce the risk for acute pancreatitis. This may be achieved through nutrition and other lifestyle interventions, as well as pharmacotherapy when appropriate. In such patients, a secondary objective is to manage levels of LDL-C and non-HDL-C, as well as other modifiable risk factors, to lower ASCVD risk. For those with mild-to-moderate hypertriglyceridemia, reducing ASCVD risk by attaining acceptable levels of LDL-C and non-HDL-C is the primary objective of intervention.
1
National Lipid Association Recommendations for Patient-Centered Management of Dyslipidemia: Part 1—Full Report
J Clin Lipidol.2015;
9:129-169
,
3
2018 AHA/ACC/AACVPR/AAPA/ABC/ACPM/ADA/AGS/APhA/ASPC/NLA/PCNA Guideline on the Management of Blood Cholesterol: A Report of the American College of Cardiology/American Heart Association Task Force on Clinical Practice Guidelines
J Am Coll Cardiol.2019;
73:e285-e350
,
5
,
7
2021 Canadian Cardiovascular Society guidelines for the management of dyslipidemia for the prevention of cardiovascular disease in adults
Can J Cardiol.2021;
37:1129-1150
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8
2021 ESC Guidelines on cardiovascular disease prevention in clinical practice
Eur Heart J.2021;
42:3227-3337
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32
Clinical review on triglycerides
Eur Heart J.2020;
41:99-109c
3

Over the past two decades, cardiovascular disease (CVD) prevention guidelines have progressively lowered LDL-C targets to <70 mg/dL for high-risk individuals based on the assumption of a linear relationship between LDL-C levels and CVD risk. However, the available evidence challenges this premise. Multiple studies demonstrate a weak or inconsistent association between LDL-C levels and atherosclerosis progression at the individual patient-level. Systematic reviews supporting the linearity assumption have notable limitations, including extrapolation beyond observed LDL-C ranges and potential ecological fallacy, as meta-regression analyses rely on study-level data, while patient-level data within the same trials often show no association between LDL-C reduction and CVD outcomes. Moreover, randomized controlled trials explicitly designed to assess LDL-C targets have yielded inconclusive and biased results. LDL-C itself is a heterogeneous marker, with particle size and composition influencing its atherogenicity. The cardiovascular benefits of lipid-lowering therapies may arise in part from pleiotropic effects unrelated to LDL-C lowering. Additionally, several studies indicate that higher LDL-C levels are paradoxically associated with longevity in elderly populations that is equal to or even greater than that of the general population. Collectively, this body of evidence raises questions about the validity of current LDL-C targets < 70 mg/dL in high-risk patients.

The optimal BP goal for individuals balances the benefits of BP reduction to prevent CVD events with the risks of adverse effects at that level of BP. Evidence supporting an SBP goal of less than 130 mm Hg for most adults comes from the SPRINT and STEP clinical trials, as well as multiple systematic reviews and meta-analyses. Although to our knowledge recent trials have not focused on DBP, a value of less than or equal to 80 mm Hg is a reasonable goal and recommended as a target for adults younger than 65 years. Thus, the new optimal BP goal for adults with hypertension is less than 130/80 mm Hg except in adults aged 65 years or older when the goal is SBP less than 130 mm Hg without regard to DBP.

For adults with diabetes and hypertension, clinical practice guidelines support an SBP goal of less than 130 mm Hg. For adults with chronic kidney disease who are not undergoing dialysis, with or without diabetes, the most recent Kidney Disease: Improving Global Outcomes guidelines recommended an SBP goal of less than 120 mm Hg when tolerated, but other guidelines recommend less than 130 mm Hg. The BP goal for patients with other comorbidities (eg, stroke, ischemic heart disease, peripheral artery disease, heart failure) is less than 130/80 mm Hg.

Suboptimal medication adherence and failure by clinicians (ie, physicians, nurse practitioners, and physician assistants) to appropriately increase medication doses are common causes of not achieving the BP goal and can prevent optimal reduction in CVD risk and death. Successful attainment of the ideal BP level requires continuous accurate BP monitoring (by patients and their clinicians), appropriate pharmacologic dose titration in response to current BP levels, and, in those who fail to respond to dose escalation, assessment of adherence to the antihypertensive regimen. Optimally, office-based BP monitoring should be combined with out-of-office measurements, such as home BP monitoring recordings obtained by a carefully instructed patient who uses the proper BP measurement technique and provides cumulative BP readings to the clinician’s office.

Quiz Ref IDVitamin D is a fat-soluble vitamin that performs an important role in calcium homeostasis and bone metabolism and also affects many other cellular regulatory functions outside the skeletal system.1-3Vitamin D requirements may vary by individual; thus, no one serum vitamin D level cutpoint defines deficiency, and no consensus exists regarding the precise serum levels of vitamin D that represent optimal health or sufficiency. According to the National Academy of Medicine, an estimated 97.5% of the population will have their vitamin D needs met at a serum level of 20 ng/mL (49.9 nmol/L) and risk for deficiency, relative to bone health, begins to occur at levels less than 12 to 20 ng/mL (29.9-49.9 nmol/L).1,4A report based on data from the 2014 National Health and Nutrition Examination Survey found that 5% of the population 1 year or older had very low 25-hydroxyvitamin D (25[OH]D) levels (<12 ng/mL) and 18% had levels between 12 and 19 ng/mL.5

Recent trends have shown a rise in screening rates for vitamin D status using serum 25(OH)D in the general population. Specifically, 25(OH)D testing frequency rose from 0.29 per 1000 person-years at risk in 2005 to 16.1 per 1000 person-years at risk by 2015, highlighting a growing interest by patients and physicians in assessing vitamin D status ( 263 , 265 ). Advocating for the routine screening of 25(OH)D levels in healthy adults is contingent upon demonstrating that such screenings can effectively identify individuals with low 25(OH)D who might not be detected through traditional risk factor assessments, and that vitamin D supplementation, following the identification of a low 25(OH)D level, leads to improvements in clinical outcomes (eg, prevention of osteoporosis, CVD, diabetes, respiratory infections, overall mortality).
Screening for low 25(OH)D in generally healthy adults (ie, those who are not at increased risk for vitamin D deficiency) would involve testing large numbers of people, with important implications for health care systems.
The US Preventive Services Task Force (USPSTF) recently concluded that there was insufficient evidence to inform a decision regarding the balance of benefits and harms of screening for vitamin D status with 25(OH)D in asymptomatic adults ( 266 ). A recommendation against population screening for vitamin D deficiency with 25(OH)D is included in the “Choosing Wisely” campaign, an initiative by the American Board of Internal Medicine to spark conversations between clinicians and patients about the value of common tests ( choosingwisely.org ).
• Question 12. Should screening with a 25(OH)D test (with vitamin D supplementation/treatment only if below a threshold) vs no screening with a 25(OH)D test be used for healthy adults?
Recommendation 12
In healthy adults, we suggest against routine screening for 25(OH)D levels. (2 | ⊕◯◯◯)
Technical remarks

Vitamin D3 has many important health benefits. Unfortunately, these benefits are not widely known among health care personnel and the general public. As a result, most of the world's population has serum 25-hydroxyvitamin D (25(OH)D) concentrations far below optimal values. This narrative review examines the evidence for the major causes of death including cardiovascular disease, hypertension, cancer, type 2 diabetes mellitus, and COVID-19 with regard to sub-optimal 25(OH)D concentrations. Evidence for the beneficial effects comes from a variety of approaches including ecological and observational studies, studies of mechanisms, and Mendelian randomization studies. Although randomized controlled trials (RCTs) are generally considered the strongest form of evidence for pharmaceutical drugs, the study designs and the conduct of RCTs performed for vitamin D have mostly been flawed for the following reasons: they have been based on vitamin D dose rather than on baseline and achieved 25(OH)D concentrations; they have involved participants with 25(OH)D concentrations above the population mean; they have given low vitamin D doses; and they have permitted other sources of vitamin D. Thus, the strongest evidence generally comes from the other types of studies. The general finding is that optimal 25(OH)D concentrations to support health and wellbeing are above 30 ng/mL (75 nmol/L) for cardiovascular disease and all-cause mortality rate, whereas the thresholds for several other outcomes appear to range up to 40 or 50 ng/mL. The most efficient way to achieve these concentrations is through vitamin D supplementation. Although additional studies are warranted, raising serum 25(OH)D concentrations to optimal concentrations will result in a significant reduction in preventable illness and death.

Vitamin D is a crucial micronutrient, critical to human health, and influences many physiological processes. Oral and skin-derived vitamin D is hydroxylated to form calcifediol (25(OH)D) in the liver, then to 1,25(OH)2D (calcitriol) in the kidney. Alongside the parathyroid hormone, calcitriol regulates neuro-musculoskeletal activities by tightly controlling blood-ionized calcium concentrations through intestinal calcium absorption, renal tubular reabsorption, and skeletal mineralization. Beyond its classical roles, evidence underscores the impact of vitamin D on the prevention and reduction of the severity of diverse conditions such as cardiovascular and metabolic diseases, autoimmune disorders, infection, and cancer. Peripheral target cells, like immune cells, obtain vitamin D and 25(OH)D through concentration-dependent diffusion from the circulation. Calcitriol is synthesized intracellularly in these cells from these precursors, which is crucial for their protective physiological actions. Its deficiency exacerbates inflammation, oxidative stress, and increased susceptibility to metabolic disorders and infections; deficiency also causes premature deaths. Thus, maintaining optimal serum levels above 40 ng/mL is vital for health and disease prevention. However, achieving it requires several times more than the government's recommended vitamin D doses. Despite extensive published research, recommended daily intake and therapeutic serum 25(OH)D concentrations have lagged and are outdated, preventing people from benefiting. Evidence suggests that maintaining the 25(OH)D concentrations above 40 ng/mL with a range of 40-80 ng/mL in the population is optimal for disease prevention and reducing morbidities and mortality without adverse effects. The recommendation for individuals is to maintain serum 25(OH)D concentrations above 50 ng/mL (125 nmol/L) for optimal clinical outcomes. Insights from metabolomics, transcriptomics, and epigenetics offer promise for better clinical outcomes from vitamin D sufficiency. Given its broader positive impact on human health with minimal cost and little adverse effects, proactively integrating vitamin D assessment and supplementation into clinical practice promises significant benefits, including reduced healthcare costs. This review synthesized recent novel findings related to the physiology of vitamin D that have significant implications for disease prevention.

Context: Although an inverse association between vitamin D status and mortality has been reported in observational studies, the precise association shape and optimal vitamin D status remain undetermined.

Objective: To investigate the association between vitamin D status and risk of all-cause and cause-specific mortality and estimate optimal serum 25-hydroxyvitamin D [25(OH)D] concentrations.

Design: Prospective cohort study.

Setting: UK Biobank.

Participants: 365 530 participants who had serum 25(OH)D measurements and no history of cardiovascular disease (CVD), cancer, or diabetes at baseline (2006-2010).

Main Outcome Measures: All-cause and cause-specific mortality.

Results: During a median follow-up of 8.9 (interquartile range: 8.3-9.5) years, 10 175 deaths occurred, including 1841 (18.1%) due to CVD and 5737 (56.4%) due to cancer. The multivariate analyses revealed nonlinear inverse associations, with a decrease in mortality risk appearing to level off at 60 nmol/L of 25(OH)D for all-cause and CVD deaths and at 45 nmol/L for cancer deaths. Compared to participants with 25(OH)D concentrations below the cutoffs, those with higher concentrations had a 17% lower risk for all-cause mortality (hazard ratio [HR]: 0.83, 95% confidence interval [CI]: 0.79-0.86), 23% lower risk for CVD mortality (

Hr: 0.77, 95%

Ci: 0.68-0.86), and 11% lower risk for cancer mortality (

Hr: 0.89, 95%

Ci: 0.84-0.95).

Conclusions: Higher 25(OH)D concentrations are nonlinearly associated with lower risk of all-cause, CVD, and cancer mortality. The thresholds of 45 to 60 nmol/L might represent an intervention target to reduce the overall risk of premature death, which needs further confirmation in large clinical trials.

Background And Objectives: Measurement of vitamin B-12 deficiency using different methods may cause diagnostic difficulties. In order to rapidly and safely diagnose vitamin B-12 deficiency, it is important to determine the reference ranges of serum B-12 and its related biomarkers such as homocysteine, holotranscobalamine (holo-TC) and methylmalonic acid (MMA). This study aimed to determine reference interval (RI)s for serum vitamin B-12 and related markers.

Methods And Study Design: Samples were collected from 404 young-to-middle-aged healthy adults aged 18-65 years. Vitamin B-12, homocysteine, holotranscobalamin, folate were analyzed using the Arcitect i2000 device. Plasma MMA was analyzed by LC/MS. RIs were then evaluated accordingly.

Results: Vitamin B-12, folate, homocysteine, holotranscobalamin and plasma MMA were 139-619 pg/mL, 3.0-14.7 ng/mL, 5.6-18.4 μmol/L, 10.7-101 pmol/L, and 0.01-0.8 μmol/L, respectively. Age group-specific RIs were also generated.

Conclusions: This study revealed that the diagnosis of vitamin B-12 deficiency should not only be based on serum vitamin B-12 levels, but also of folate, homocysteine, holotranscobalamin and MMA levels; all which are related to vitamin B-12 metabolism.

In recent years there has been growing interest in the vitamins folic acid and vitamin B12 because of the realization that the status of these vitamins in populations is less than adequate, and that such inadequacy may be linked to adverse public health outcomes. This concern has prompted the United States, Canada, and other countries to fortify grain products with folic acid, while additional countries are considering doing so in the near future. This presentation provides a new approach which relies on the combination of the concentrations in blood of vitamins and their respective functional indicators to establish cutoff points for assessing folate and vitamin B12 status in populations. The premise is based on the fact that the relationship between plasma vitamin concentrations and their respective functional indicators is inverse and biphasic, with a steep slope at low concentrations of the vitamin and a more moderate slope at higher plasma vitamin concentrations. We propose that the intersection of these two slopes be used as a guideline for assessing the status of these vitamins and the adequacy of fortification programs. The cutoff would be 10 nmol/L for serum folate and 340 nmol/L for red blood cell folate, based on lowest plasma homocysteine. For serum vitamin B12, the cutoff would be 150 pmol/L based on lowest methylmalonic acid and 300 pmol/L based on lowest homocysteine.

Aging is an independent risk factor for the development of various diseases associated, among others, with detrimental blood levels of fat- and water-soluble vitamins. Thus, the objective of this study is to investigate age-related changes in blood levels of vitamin A, B12, C, D, and E. Subject serum vitamin levels were obtained from the combined National Health and Nutrition Examination Surveys (NHANES). NHANESIII and NHANES 1999-2000, 2001-2002, 2003-2004, and 2005-2006. The raw data set was stratified into five age groups G1- G5: 20 ≤ G1 < 30, 30 ≤ G2 < 40, 40 ≤ G3 < 50, 50 ≤ G4 < 60, and 60 ≤ G5 < 70 years of age. Age stratified data was cleaned using the modified Horn algorithm. The reference range for the vitamin level of a specific age group was defined as data between the first and third quartile of the subject defined by normal blood pressure and normal bone density. Age-dependent changes in serum/plasma vitamin levels were assessed using the bootstrap technique with 10,000 repeats and Bonferroni adjustment. There was a continuous increase in vitamin A, B12, D, and E levels in the blood. However, the vitamin C concentration remained virtually constant in all age groups. There was a lack of cross-correlations between lipid and water-soluble vitamin levels and blood pressure and bone health. The following reference levels for vitamin A, B12, C, D, and E in subjects older than 20 years of age were established: vitamin

A: 1.32-2.8 mmol/L, vitamin B12: 257.94-498.33 pmol/L, vitamin

C: 38.18-79.2 mmol/L, vitamin

D: 76.33-199.36 nmol/L and vitamin

E: 3.65-41.12 μmol/L.

Vitamin B12 (B12) is necessary for the proper functioning of the central and peripheral nervous systems. Although there is no exact definition for B12 levels, a value of 200 pg/mL is compatible with deficiency, 200-299 pg/mL is considered borderline, and 300 pg/mL is considered normal. In population studies, the prevalence of B12 deficiency ranges between 2.9% and 35%. Furthermore, many medications, such as metformin [for type 2 diabetes mellitus (T2DM)], can cause B12 deficiency. The objectives of this study were to determine the population status of B12 in southwestern Colombia (and the status of B12 in subjects with T2DM). In the total population (participants with and without T2DM), the prevalence of B12 deficiency was 17.8%; that of borderline was 19.3%; and that of normal levels was 62.9%. The prevalence of deficiency increased with age and was significantly higher in those aged ≥60 years (p = 0.000). In T2DM subjects, the prevalence of deficiency was significantly higher concerning those without T2DM (p = 0.002) and was significantly higher in those who received >1 gm/day of metformin (p = 0.001). Thus, the prevalence of deficiency and borderline levels of B12 in our population was high, particularly in those >60 years of age. B12 deficiency was significantly higher in individuals with T2DM than in individuals without T2DM, especially among those receiving high doses of metformin.

Background: Enriched cereal-grain products have been fortified in the United States for >20 y to improve folate status in women of reproductive age and reduce the risk of folic acid-responsive neural tube birth defects (NTDs).

Objectives: Our objectives were to assess postfortification changes in folate status in the overall US population and in women aged 12-49 y and to characterize recent folate status by demographic group and use of folic acid-containing supplements.

Methods: We examined cross-sectional serum and RBC folate data from the NHANES 1999-2016.

Results: Serum folate geometric means increased from 2007-2010 to 2011-2016 in persons aged ≥1 y (38.7 compared with 40.6 nmol/L) and in women (35.3 compared with 37.0 nmol/L), whereas RBC folate showed no significant change. Younger age groups, men, and Hispanic persons showed increased serum and RBC folate concentrations, whereas non-Hispanic black persons and supplement nonusers showed increased serum folate concentrations. The folate insufficiency prevalence (RBC folate <748 nmol/L; NTD risk) in women decreased from 2007-2010 (23.2%) to 2011-2016 (18.6%) overall and in some subgroups (e.g., women aged 20-39 y, Hispanic and non-Hispanic black women, and supplement nonusers). After covariate adjustment, RBC folate was significantly lower in all age groups (by ∼10-20%) compared with persons aged ≥60 y and in Hispanic (by 8.2%), non-Hispanic Asian (by 12.1%), and non-Hispanic black (by 20.5%) compared with non-Hispanic white women (2011-2016). The 90th percentile for serum (∼70 nmol/L) and RBC (∼1800 nmol/L) folate in supplement nonusers aged ≥60 y was similar to the geometric mean in users (2011-2014).

Conclusions: Blood folate concentrations in the US population overall and in women have not decreased recently, and folate insufficiency rates are ∼20%. Continued monitoring of all age groups is advisable given the high folate status particularly in older supplement users.

Background: Folate and vitamin B-12 are essential nutrients for normal cell growth and replication, but the association of serum folate and vitamin B-12 concentrations with mortality risk remains uncertain.

Objective: This study was performed to investigate the associations of serum folate and vitamin B-12 concentrations with mortality risk and test whether the methylenetetrahydrofolate reductase (MTHFR) C677T polymorphism modifies these associations.

Methods: A total of 3050 Japanese community residents aged ≥40 y were prospectively followed-up for mortality between 2002 and 2012. Cox proportional hazards models and restricted cubic splines were used to estimate HRs and 95% CIs of mortality.

Results: During a median follow-up period of 10.2 y, 336 participants died. Higher serum folate concentrations were associated with lower risks of all-cause mortality [multivariable-adjusted

Hr: 0.73; 95%

Ci: 0.56, 0.96 for the second tertile (8.8-12.2 nmol/L; median 10.4 nmol/L) and

Hr: 0.61; 95%

Ci: 0.46, 0.80 for the third tertile (≥12.5 nmol/L; median 15.6 nmol/L) serum folate concentrations compared with the first tertile (≤8.6 nmol/L; median 7.0 nmol/L)]. This association remained significant in all sensitivity analyses. Spline analyses showed a steady decline in all-cause mortality risk with increasing serum folate concentrations up to 20-25 nmol/L. This association persisted regardless of the MTHFR C677T genotypes. For serum vitamin B-12, the multivariable-adjusted HR of 1.32 (95%

Ci: 0.97, 1.79) of all-cause mortality was marginally significantly greater in the first tertile compared with the second tertile. This association was attenuated and nonsignificant after the exclusion of participants with a history of cardiovascular disease or cancer, or participants aged ≥85 y at baseline, or deaths in the first 3 y of follow-up.

Conclusions: Serum folate concentrations were inversely associated with the risk of all-cause mortality in Japanese adults. Serum vitamin B-12 concentrations were not consistently associated with all-cause mortality risk after accounting for reverse-causation bias.

Although some individual studies showed conflicting results, the limited overall evidence suggests that selenium supplementation has no effect on all-cause mortality, cardiovascular disease mortality, cardiovascular disease events, or cancer mortality.

The USPSTF also reviewed the evidence on the harms of vitamin and mineral supplements. For many supplements there was little to no evidence of serious harms.

The most serious harm identified was increased cardiovascular disease mortality and increased risk of lung cancer in persons who smoke or had workplace asbestos exposure, associated with beta carotene supplementation at doses of 30 and 20 mg/d. One of these trials also co-administered vitamin A at a dose of 25 000 IU/d, which exceeds the current tolerable upper intake level for vitamin A in adults. A minor harm of beta carotene was orange discoloration of the skin. Two cohort studies in women showed a statistically nonsignificant increased risk of hip fracture associated with vitamin A supplementation. Two trials showed an increased risk of hemorrhagic stroke associated with vitamin E supplementation at doses of 111 and 200 IU daily, and 1 cohort study found that a high intake of vitamin B 6 (≥35 mg/d) was associated with an increased risk of hip fracture compared with a low intake (<2 mg/d).

One trial and 2 cohort studies reported an increased risk of kidney stones in persons taking vitamin D. In the cohort studies, this risk was only associated with vitamin D doses of 1000 IU/d or more. Two cohort studies in men suggest an association between vitamin C supplementation and kidney stones. The evidence on an association between calcium use and kidney stones was mixed.

A draft version of this recommendation statement was posted for public comment on the USPSTF website from May 4 to June 1, 2021. In response to comments, the USPSTF specified the doses of beta carotene, vitamin A, and vitamin E used in trials reporting harms from these vitamins. Some comments requested that persons with chronic conditions or who are hospitalized be included in this recommendation. In response, the USPSTF wants to clarify that its recommendations apply to community-dwelling individuals without known underlying health conditions.

A total of 84 studies (N = 739 803) (eTable 3 in the Supplement) were included, comprising 78 RCTs (n = 324 837) and 6 cohort studies (n = 390 689), after review of 17 459 unique citations and 379 full-text articles (Figure 2). Fifty-two of the included studies were newly identified since the last review. The included studies addressed multivitamins; vitamins A, B 3 , B 6 , B 12 , C, D, and E; beta carotene; folic acid; calcium; magnesium; selenium; and zinc. The evidence for the B and C vitamins, folic acid, magnesium, selenium, and zinc was low, insufficient, or absent for all outcomes. Results for these supplements can be found in the full evidence report.

The mean age across all included studies was 61.0 years. An estimated 65.1% of all participants were women. Most participants were White in studies conducted in the US (with data from 22 of 36 studies). An estimated 19.6% of participants were Black, among studies reporting race and ethnicity. The vitamin D trials had, on average, greater representation of Black participants. Other racial and ethnic groups had minimal representation for all supplements.

Key Question 1. What is the efficacy of multivitamin supplementation for reducing cardiovascular disease, cancer, and mortality in the general adult population?

Nine RCTs addressed KQ1 (n = 51 945) (eTable 4 in the Supplement). Three large studies had primary aims of cardiovascular disease and cancer prevention, were all rated as good quality, and comprised most of the evidence for this KQ.

Figure 2:
Caption: Literature Search Flow Diagram: Vitamin and Mineral Supplements for Primary Prevention of Cardiovascular Disease and Cancer
Description: aReasons for exclusion: Intervention relevance: Study used an excluded intervention. Outcomes: Study did not report relevant outcomes. Design: Study did not use an included design. Setting: Study not conducted in a country relevant to US practice. Population: Study not conducted in community-dwelling adults without chronic disease and without nutritional deficiencies. Language: Not available in English. Publication type: Conference abstract. Study relevance: Study aim not relevant. Harms study design: Short-term nonserious harms reported, but lack of evidence of benefit. High dose: Supplement dose greater than the tolerable upper intake level as determined by the National Academies of Sciences, Engineering, and Medicine Food and Nutrition Board. Quality: Study did not meet criteria for fair or good quality.bStudies could be included for more than 1 key question (KQ).

Micronutrients are essential for maintaining physiological homeostasis and optimizing healthspan, defined as the years lived in good health without chronic diseases or disabilities. Despite increasing global life expectancy, improvements in healthspan have not kept pace, partly due to subclinical micronutrient deficiencies that often precede clinical symptoms. The triage theory highlights how micronutrient insufficiencies compromise long-term health by prioritizing critical metabolic functions. Micronutrients such as Vitamins B6, B9, B12, D, and K are particularly crucial to optimizing healthspan, by influencing energy metabolism, neurological health, immune regulation, and bone integrity. Traditional tools like Dietary Reference Intakes (DRIs) provide population-level guidelines but fail to account for individual factors such as genetics, lifestyle, and nutrient interactions. Quantitative assessment of micronutrient concentrations using biomarkers offers a more precise approach but faces challenges, including high costs and limited accessibility. National nutrition surveys demonstrate potential in addressing population-level deficiencies and form the basis for advancing precision supplementation strategies to improve health outcomes and extend healthspan by defining optimal micronutrient concentrations. Future efforts should aim to establish evidence-based thresholds for optimal micronutrient concentrations by integrating biomarker data with clinical outcomes, genetic profiles, and lifestyle factors, providing a framework to guide personalized and population-level supplementation strategies.

A well-functioning immune system is essential for human health and well-being. Micronutrients such as vitamins A, C, D, E, and zinc have several functions throughout the immune system, yet inadequate nutrient intakes are pervasive in the US population. A large body of research shows that nutrient inadequacies can impair immune function and weaken the immune response. Here, we present a new analysis of micronutrient usual intake estimates based on nationally representative data in 26,282 adults (>19 years) from the 2005-2016 National Health and Nutrition Examination Surveys (NHANES). Overall, the prevalence of inadequacy (% of population below estimated average requirement [EAR]) in four out of five key immune nutrients is substantial. Specifically, 45% of the U.S. population had a prevalence of inadequacy for vitamin A, 46% for vitamin C, 95% for vitamin D, 84% for vitamin E, and 15% for zinc. Dietary supplements can help address nutrient inadequacy for these immune-support nutrients, demonstrated by a lower prevalence of individuals below the EAR. Given the long-term presence and widening of nutrient gaps in the U.S.-specifically in critical nutrients that support immune health-public health measures should adopt guidelines to ensure an adequate intake of these micronutrients. Future research is needed to better understand the interactions and complexities of multiple nutrient shortfalls on immune health and assess and identify optimal levels of intake in at-risk populations.

Many Americans have inadequate intakes of several nutrients. The Dietary Guidelines for Americans 2015-2020 specifically identified vitamins A, C, D and E, calcium, magnesium, iron, potassium, choline and fiber as "underconsumed nutrients". Based on nationally representative data in 10,698 adults from National Health and Nutrition Examination Surveys (NHANES), 2009-2012, assessments were made of age-group differences in the impact of dietary supplements on nutrient intake and inadequacies. Compared to food alone, use of any dietary supplement plus food was associated with significantly ( < 0.01) higher intakes of 15-16 of 19 nutrients examined in all age groups; and significantly reduced rates of inadequacy for 10/17, 8/17 and 6/17 nutrients examined among individuals age ≥71, 51-70 and 19-50 years, respectively. Compared to the other age groups, older adults (≥71 years) had lower rates of inadequacy for iron and vitamins A, C, D and E, but higher rates for calcium. An increased prevalence of intakes above the Tolerable Upper Intake Level was seen for 8-9 of 13 nutrients, but were mostly less than 5% of the population. In conclusion, dietary supplement use is associated with increased micronutrient intake, decreased inadequacies, and slight increases in prevalence above the UL, with greater benefits seen among older adults.

Background And Aims: In view of the wide variations in reference intervals (RIs) for copper and zinc concentrations in serum, this study was designed to derive appropriate RIs that could be routinely adopted in other laboratories, so allowing better interpretation of results. A secondary aim was to assess the effects of several variables on serum concentrations of copper and zinc.

Materials And Methods: Data from over 21,000 individuals were downloaded from four National Health and Nutrition Examination Surveys conducted by the US Centers for Disease Control and Prevention. After controlling for the effects of several variables, age and gender-related RIs were calculated.

Results: A comprehensive list of reference intervals of copper and zinc in serum is provided. Serum copper concentrations were affected by age, gender, ethnicity, pregnancy, use of oral contraceptive pill, health status and smoking (in males). Serum zinc concentrations were affected by gender, age, fasting, ethnicity, serum albumin concentration, health status (in males), time of venipuncture and pregnancy.

Conclusion: Reliable RIs for copper and zinc in serum were derived. As analytical accuracy and precision were assured and optimal sample collection protocols were followed, these RIs can be transferred for routine use in other clinical biochemistry laboratories with good analytical performance in external quality assurance schemes.

Copper : For the metabolically stable adult receiving TPN, the suggested additive dosage level is 0.5 to 1.5 mg copper/day. For pediatric patients, the suggested additive dosage level is 20 mcg copper/kg/day. The normal plasma range for copper is approximately 80 to 160 mcg/100 mL.

Manganese : For the metabolically stable adult receiving TPN, the suggested additive dosage level for manganese is 0.15 mg to 0.8 mg/day. For pediatric patients, a dosage level of 2 to 10 mcg manganese/kg/day is recommended.

Chromium : For the metabolically stable adult receiving TPN, the suggested additive dosage level is 10 to 15 mcg chromium/day. The metabolically stable adult with intestinal fluid loss may require 20 mcg chromium/day, with frequent monitoring of blood levels as a guideline for subsequent administration. For pediatric patients, the suggested additive dosage level is 0.14 to 0.20 mcg/kg/day.

Selenium : For metabolically stable adults receiving TPN, the suggested additive dosage level is 20 to 40 mcg selenium/day. For pediatric patients, the suggested additive dosage level is 3 mcg/kg/day.

CONTEXT.—: Clinical testing for Wilson disease (WD) is potentially challenging. Measuring the fraction of labile bound copper (LBC) to total copper may be a promising alternative diagnostic tool with better sensitivity and specificity than some current biomarker approaches. A dual filtration-based inductively coupled mass spectrometry (ICP-MS) assay to measure LBC in serum was developed. OBJECTIVE.—: To establish a reference interval for LBC and LBC to total copper (LBC fraction) in a healthy adult population, and to examine associations between total copper, LBC, and LBC fraction with age, sex, menopausal status, hormone replacement therapy, and supplement use. DESIGN.—: Serum samples were collected from healthy male (n = 110) and female (n = 104) patients between the ages of 19 and 80 years. Total copper and LBC were analyzed using ICP-MS. Results were used to calculate the LBC fraction. Reference intervals were calculated for the 2.5th and 97.5th percentiles for both LBC and LBC fraction. RESULTS.—: The reference intervals for LBC were determined to be 13 to 105 ng/mL and 12 to 107 ng/mL for female and male patients, respectively. The reference intervals for the LBC fraction were 1.0% to 8.1% and 1.2% to 10.5% for female and male patients, respectively. No significant associations were found regarding age, menopausal status, hormone replacement therapy, or vitamin and supplement use. CONCLUSIONS.—: Sex-specific reference intervals have now been established for LBC and LBC fraction. These data in conjunction with further testing of WD populations can be used to assess the sensitivity and specificity of LBC fraction in screening, monitoring, and diagnosis.

Background: Copper is an essential trace element in the human body; its relationship with cardio-cerebrovascular diseases (CCVDs) remains unclear. This study aimed to comprehensively investigate the association between circulating copper concentrations and CCVD risk and mortality.

Methods: We searched the PubMed, Cochrane Library, Embase, Scopus, and Web of Knowledge databases for cohort studies reporting associations between circulating copper concentrations and cardiovascular diseases and mortality published up to August 23, 2023. Effect sizes were pooled using random-effects models. We graded the certainty of the evidence by the GRADE (Grading of Recommendations Assessment, Development, and Evaluation) framework.

Results: Our analysis included 47,813 patients across 17 cohort studies. Elevated circulating copper levels were linked to the risk of stroke (OR = 1.52; 95% CI 1.30-1.78), coronary artery disease mortality (RR = 2.77; 95% CI 1.82-4.19), cardiovascular mortality (RR = 1.79; 95% CI 1.52-2.11), and all-cause mortality (RR = 1.56; 95% CI 1.35-1.79) but not the risk of acute myocardial infarction (RR = 2.01; 95% CI 0.63-6.47). Continuous analysis (per 20 μg/dl increase) showed consistent results regarding the association between copper levels and stroke incidence (OR = 1.23; 95% CI 1.14-1.33), cardiovascular mortality (HR = 1.28; 95% CI 1.07-1.53) and all-cause mortality (HR = 1.22; 95% CI 1.04-1.44). Except for the low certainty of evidence of acute myocardial infarction incidence, all outcomes had moderate certainty of evidence.

Conclusion: Excessive circulating copper levels are associated with stroke, coronary artery disease mortality, cardiovascular mortality, and all-cause mortality with moderate certainty.

Manganese (Mn) is an essential element that participates in several biological processes. Mn serves as a cofactor for several enzymes, such as glutamine synthetase and oxidoreductases, that have an important role in the defense of the organisms against oxidative stress. The diet is the main source of Mn intake for humans, and adequate daily intake levels for this metal change with age. Moreover, in higher amounts, Mn may be toxic, mainly to the brain. Here, we provide an overview of Mn occurrence in food, addressing its bioaccessibility and discussing the dietary standard and recommended intake of Mn consumption. In addition, we review some mechanisms underlying Mn-induced neurotoxicity.

Background: The study aimed to explore the association between manganese concentration and all-cause, cardiovascular disease (CVD)-related, and cancer-related mortality in the general population of the United States.

Methods: We integrated the data from the National Health and Nutrition Examination Survey from 2011 to 2018. A total of 9,207 subjects were selected based on the inclusion and exclusion criteria. The relationship between manganese concentration and all-cause, CVD-related, and cancer-related mortality was analyzed by constructing a Cox proportional hazard regression model and a restricted cubic spline (RCS) plot. Additionally, subgroup analyses stratified by age, sex, race/ethnicity, hypertension, diabetes mellitus (DM), chronic heart disease, chronic heart failure, angina pectoris, heart attack, stroke, and BMI were further performed.

Results: In the full adjusted model, compared with the lowest quartile, the adjusted hazard ratios with 95% confidence intervals (CIs) for all-cause, CVD-related, and cancer-related mortality across manganese quartiles were (1.11 (0.87,1.41), 0.96 (0.74, 1.23), and 1.23 (0.96, 1.59); P-value for trend =0.041), (0.86 (0.54, 1.37), 0.87 (0.55, 1.40), and 1.07 (0.67, 1.72); P-value for trend =0.906), and (1.45 (0.92, 2.29), 1.14 (0.70, 1.88), and 1.26 (0.75, 2.11); P-value for trend =0.526), respectively. The RCS curve shown a U-shaped association between manganese concentration and all-cause mortality and CVD-related mortality (P-value for nonlinear <0.05). However, there was an increase and then a decrease in the link between manganese concentration and cancer-related mortality (P-value for nonlinear <0.05). Manganese exposure was positively correlated with sex (correlation coefficient, r =0.19, P-value <0.001) and negatively correlated with age (correlation coefficient, r =-0.11, P-value <0.001) and serum creatinine (correlation coefficient, r =-0.12, P-value <0.001), respectively.

Conclusions: Our findings suggest that elevated serum manganese concentrations are associated with all-cause and CVD-related mortality in the U.S. population and that maintenance of serum manganese between 8.67-9.23 µg/L may promote public health.

Manganese (Mn) is a known toxicant and an essential trace element, and it plays an important role in various mechanisms in relation to cardiovascular health. However, epidemiological studies of the association between blood Mn and cardiovascular diseases (CVD) among U.S. adults are rare. A cross-sectional study of 12,061 participants aged ≥ 20 was conducted using data from the National Health and Nutrition Examination Survey 2011-2018. Logistic regression and restricted cubic spline were used to examine the relationship between blood Mn levels and total CVD risk and specific CVD subtypes. Bayesian kernel-machine regression (BKMR) and weighted quantile sum (WQS) analyses were performed to explore the joint effects of Mn with other metals on CVD. The results showed that individuals with the third quartile group of blood Mn levels had significantly lower risks of CVD, displaying a non-linear U-shaped dose-response relationship. A significant interaction of age on this association was observed. No significant associations were found between Mn levels and specific CVD subtypes. BKMR and WQS analyses showed a positive association between heavy metal mixtures and CVD risks, with no interaction between Mn and other metals. In conclusion, blood Mn levels were significantly associated with CVD risks with a U-shaped relationship in U.S. adults, with possible age-specific differences. Future larger prospective studies are warranted to validate these findings.

Iodine nutrition is an important public health issue. Trends in iodine status over time among U.S. schoolchildren and adults and factors mediating changes of iodine status were examined. In this cross-sectional study of National Health and Nutrition Examination Survey (NHANES) data, we estimated trends in the U.S. population using linear regression analyses. Representative samples of U.S. children and adults were enrolled in NHANES 2001-2020. The NHANES cycles were categorized into 5 four-year periods: 2001-2004, 2005-2008, 2009-2012, 2013-2016, and 2017-2020. The final sample sizes of children and adults for analysis were 4288 and 19,661, respectively. The estimated average requirement (EAR) (based on guidelines from the Institute of Medicine), was used to estimate the prevalence rate of inadequate iodine intake. Binary logistic regression analyses were used to investigate the association between iodine status and contributing factors. From 2001-2004 to 2017-2020, among children, urinary iodine concentration (UIC) decreased from 243 to 166 μg/L ( = 0.0057) and prevalence of iodine intake below the EAR rose from 15.4% to 27.6%. In adults, the UIC decreased from 153 to 116 μg/L ( < 0.001) and prevalence of iodine intake below the EAR rose from 15.0% to 17.9%. A higher prevalence rate of iodine intake below the EAR was observed in females compared with males (children, 24.0% vs. 16.5%,  < 0.001; adults, 20.0% vs. 11.1%,  < 0.001). Inadequate iodine intake was less frequent among non-Hispanic White and Hispanic compared with non-Hispanic Black in children and adults. Adults without thyroid problems had a higher prevalence of inadequate iodine intake than those with thyroid problems (16.0% vs. 13.0%,  = 0.001). Inadequate iodine intake was less likely in the children who "sometimes" and "often" consumed milk products compared with children who "never or rarely" consumed milk products (OR = 0.60 [CI 0.30-1.21] and OR = 0.24 [CI 0.13-0.43], respectively). The prevalence of inadequate iodine intake among adults reporting "sometimes" (OR = 0.70 [CI 0.58-0.83]) and "often" consuming milk products was lower than those who "never or rarely" consumed them (OR = 0.36 [CI 0.30-0.44]). In this weighted survey, the prevalence of inadequate iodine intake increased from 2001-2004 to 2017-2020 among U.S. school-age children and adults. Sex, race, thyroid problems, and a decreased intake of milk products were significantly associated with iodine intake below the EAR.

The thyroid gland requires iodine to synthesize thyroid hormones, and iodine deficiency results in the inadequate production of thyroxine and related thyroid, metabolic, developmental, and reproductive disorders. Iodine requirements are higher in infants, children, and during pregnancy and lactation than in adult men and non-pregnant women. Iodine is available in a wide range of foods and water and is susceptible to almost complete gastric and duodenal absorption as an iodide ion. A healthy diet usually provides a daily iodine consumption not exceeding 50% of the recommended intake. Iodine supplementation is usually necessary to prevent iodine deficiency disorders (IDDs), especially in endemic areas. The community-based strategy of iodine fortification in salt has eradicated IDDs, such as endemic goiter and cretinism, in countries providing adequate measures of iodine prophylaxis over several decades in the 20th century. Iodized salt is the cornerstone of iodine prophylaxis in endemic areas, and the continuous monitoring of community iodine intake and its related clinical outcomes is essential. Despite the relevant improvement in clinical outcomes, subclinical iodine deficiency persists even in Western Europe, especially among girls and women, being an issue in certain physiological conditions, such as pregnancy and lactation, and in people consuming unbalanced vegetable-based or salt-restricted diets. Detailed strategies to implement iodine intake (supplementation) could be considered for specific population groups when iodized salt alone is insufficient to provide adequate requirements.

Currently, there is no global consensus about the essentiality of dietary chromium. To provide evidence to this debate, an examination of blood chromium levels and common chronic health conditions was undertaken. Using a subsample from the 2015−2016 US National Health and Nutrition Examination Survey (n = 2894; 40 years+), chi-square and binary logistic regression analyses were conducted to examine blood chromium levels (0.7−28.0 vs. <0.7 µg/L) and their associations with cardiovascular diseases (CVDs; self-report), diabetes mellitus (DM; glycohemoglobin ≥5.7%), and depression (Patient Health Questionnaire-9 score ≥5), while controlling for socio-demographic (age/sex/income/education/relationship status) and health-related (red blood cell folate/medications/co-morbidities/body mass index (BMI)/substance use) factors. The sample was almost evenly distributed between men and women (n = 1391, 48.1% (men); n = 1503, 51.9% (women)). The prevalence estimates of low blood chromium levels tended to be higher among those with CVDs (47.4−47.6%) and DM (50.0−51.6%). Comparisons between those with low vs. normal blood chromium levels indicate men have increased odds of CVDs (adjusted odds ratio (aOR) = 1.86, 95% confidence interval (CI): 1.22−2.85, p < 0.001) and DM (aOR = 1.93, 95%

Ci: 1.32−2.83, p < 0.001) and lower odds of depression (aOR = 0.42, 95%

Ci: 0.22−0.77, p < 0.05). Dietary chromium may be important in the prevention and management of CVDs and DM for men. Continued exploration of chromium’s role in chronic diseases, including differences by biological factors, is needed.

Diabetes is a global public health concern with increasing prevalence worldwide. Chromium (Cr), a trace element found in soil, water, and food, has been proposed to have a possible positive effect in glucose metabolism and diabetes mellitus prevention. However, the relationship between trivalent chromium [Cr(III)] exposure, mainly through the consumption of diet supplements, and type 2 diabetes mellitus (T2DM) remains controversial. An extensive systematic review of the current literature on randomized controlled studies (RCTs) was conducted from 1 January 2000, to January 2024 using the databases PubMed, Scopus, ScienceDirect, and Cochrane, with specific keywords and inclusion as well as exclusion criteria. After close screening of the research studies retrieved from the mentioned websites was conducted, the most related studies were included in the final systematic review. The studies were evaluated for the degree of relevance, quality, and risk bias, using appropriate quality assessment tools. Several of the included RCT studies reported possible benefits of Cr(III) supplementation, mainly in the form of chromium picolinate (CrPic), chromium yeast (CY), chromium chloride (CrCl3), and chromium nicotinate (CrN). The dosage of chromium was between 50 and 1000 μg/day and it was consumed from 2 to 6 months. Glycemic control markers, including FPG, insulin, HbA1C, and HOMA-IR levels, significantly decrease following chromium supplementation, mainly in studies with a longer intervention period. Supplementing with chromium (Cr) indicated that could significantly improve lipid profile by raising high-density lipoprotein and lowering triglyceride and total cholesterol while having little effect on low-density lipoprotein. However, most research findings include significant limitations, such as inconsistent dosage and type of chromium, formulation of supplements, and study duration. Further well-designed and high-quality research is needed to fully understand the role of chromium dietary supplementation and the potential risks related to its mechanisms of action, type, and dose, in the prevention and treatment of type 2 diabetes mellitus.

Background: Systolic blood pressure (SBP) lowering reduces major cardiovascular disease (CVD) and all-cause mortality. However, the optimal target for SBP lowering remains controversial.

Methods: We included trials with random allocation to an SBP <130 mm Hg treatment target and CVD as the primary outcome. Data were extracted from each study independently and in duplicate using a standardized protocol. Random-effects meta-analysis was used to obtain pooled hazard ratios (HRs) and 95% CIs for CVD and all-cause mortality comparing SBP <130 and ≥130 mm Hg treatment targets. A secondary analysis compared the same outcomes for randomization to an SBP target of <120 or <140 mm Hg.

Results: Seven trials, including 72 138 participants, met the eligibility criteria. Compared with an SBP target of ≥130 mm Hg, an SBP target of <130 mm Hg significantly reduced major CVD (HR, 0.78 [95% CI, 0.70-0.87]) and all-cause mortality (HR, 0.89 [95% CI, 0.79-0.99]). Compared with an SBP target of <140 mm Hg, an intensive SBP target of <120 mm Hg significantly reduced major CVD (HR, 0.82 [95% CI, 0.74-0.91]), but all-cause mortality was marginally insignificant (HR, 0.85 [95% CI, 0.71-1.01]). Adverse events were significantly more likely in the intensive SBP target groups, but the absolute risks were low.

Conclusions: This study suggests targeting an SBP <130 mm Hg significantly reduces the risks of major CVD and all-cause mortality. The findings also support an SBP target of <120 mm Hg, based on a smaller number of trials.

Registration:

Url: https://www.crd.york.ac.uk/PROSPERO/; Unique identifier: CRD42023490693.

The optimal target blood pressure in the treatment of hypertension is undefined. Whether more intense therapy is better than standard, typically <140/90 mm Hg, is controversial. The most recent American guidelines recommend ≤130/80 mm Hg for essentially all adults. There have been at least 28 trials targeting more versus less intensive therapy, including 13 aimed at reducing cardiovascular events and mortality, 11 restricted to patients with chronic kidney disease, and 4 with surrogate endpoints. We review these trials in a narrative fashion due to significant heterogeneity in targets chosen, populations studied, and primary endpoints. Most were negative, although some showed significant benefit to more intense therapy. When determining the optimal pressure for an individual patient, additional factors should be considered, including age, frailty, polypharmacy, baseline blood pressure, and the diastolic blood pressure J-curve. We discuss these modifying factors in detail. Whereas the tenet "lower is better" is generally true, one size does not fit all, and blood pressure control must be individualized.

Humans age at different rates and families with exceptional longevity provide an opportunity to understand why some people age slower than others. Unique features exhibited by centenarians include a family history of extended life span, compression of morbidity with resultant extension of health span, and longevity-associated biomarker profiles. These biomarkers, including low-circulating insulin-like growth factor 1 (IGF-1) and elevated high-density lipoprotein (HDL) cholesterol levels, are associated with functional genotypes that are enriched in centenarians, suggesting that they may be causative for longevity. While not all genetic discoveries from centenarians have been validated, in part due to exceptional life span being a rare phenotype in the general population, the APOE2 and FOXO3a genotypes have been confirmed in a number of populations with exceptional longevity. However, life span is now recognized as a complex trait and genetic research methods to study longevity are rapidly extending beyond classical Mendelian genetics to polygenic inheritance methodologies. Moreover, newer approaches are suggesting that pathways that have been recognized for decades to control life span in animals may also regulate life span in humans. These discoveries led to strategic development of therapeutics that may delay aging and prolong health span.

Background: Many factors contribute to exceptional longevity, with genetics playing a significant role. However, to date, genetic studies examining exceptional longevity have been inconclusive. This comprehensive review seeks to determine the genetic variants associated with exceptional longevity by undertaking meta-analyses.

Methods: Meta-analyses of genetic polymorphisms previously associated with exceptional longevity (85+) were undertaken. For each variant, meta-analyses were performed if there were data from at least three independent studies available, including two unpublished additional cohorts.

Results: Five polymorphisms, ACE rs4340, APOE ε2/3/4, FOXO3A rs2802292, KLOTHO KL-VS and IL6 rs1800795 were significantly associated with exceptional longevity, with the pooled effect sizes (odds ratios) ranging from 0.42 (APOE ε4) to 1.45 (FOXO3A males).

Conclusion: In general, the observed modest effect sizes of the significant variants suggest many genes of small influence play a role in exceptional longevity, which is consistent with results for other polygenic traits. Our results also suggest that genes related to cardiovascular health may be implicated in exceptional longevity. Future studies should examine the roles of gender and ethnicity and carefully consider study design, including the selection of appropriate controls.

Genomic analysis of longevity offers the potential to illuminate the biology of human aging. Here, using genome-wide association meta-analysis of 606,059 parents' survival, we discover two regions associated with longevity (HLA-DQA1/DRB1 and LPA). We also validate previous suggestions that APOE, CHRNA3/5, CDKN2A/B, SH2B3 and FOXO3A influence longevity. Next we show that giving up smoking, educational attainment, openness to new experience and high-density lipoprotein (HDL) cholesterol levels are most positively genetically correlated with lifespan while susceptibility to coronary artery disease (CAD), cigarettes smoked per day, lung cancer, insulin resistance and body fat are most negatively correlated. We suggest that the effect of education on lifespan is principally mediated through smoking while the effect of obesity appears to act via CAD. Using instrumental variables, we suggest that an increase of one body mass index unit reduces lifespan by 7 months while 1 year of education adds 11 months to expected lifespan.Variability in human longevity is genetically influenced. Using genetic data of parental lifespan, the authors identify associations at HLA-DQA/DRB1 and LPA and find that genetic variants that increase educational attainment have a positive effect on lifespan whereas increasing BMI negatively affects lifespan.

Human longevity is heritable, but genome-wide association (GWA) studies have had limited success. Here, we perform two meta-analyses of GWA studies of a rigorous longevity phenotype definition including 11,262/3484 cases surviving at or beyond the age corresponding to the 90th/99th survival percentile, respectively, and 25,483 controls whose age at death or at last contact was at or below the age corresponding to the 60th survival percentile. Consistent with previous reports, rs429358 (apolipoprotein E (ApoE) ε4) is associated with lower odds of surviving to the 90th and 99th percentile age, while rs7412 (ApoE ε2) shows the opposite. Moreover, rs7676745, located near GPR78, associates with lower odds of surviving to the 90th percentile age. Gene-level association analysis reveals a role for tissue-specific expression of multiple genes in longevity. Finally, genetic correlation of the longevity GWA results with that of several disease-related phenotypes points to a shared genetic architecture between health and longevity.

Background: Genetic background of healthy or pathological styles of aging and human lifespan is determined by joint gene interactions. Lucky combinations of antioxidant gene polymorphisms can result in a highly adaptive phenotype, providing a successful way to interact with external triggers. Our purpose was to identify the polygenic markers of survival and longevity in the antioxidant genes among elderly people with physiological and pathological aging.

Methods: In a 20-year follow-up study of 2350 individuals aged 18-114 years residing in the Volga-Ural region of Russia, sex-adjusted association analyses of MTHFR rs1801133, MSRA rs10098474, PON1 rs662, PON2 rs7493, SOD1 rs2070424, NQO1 rs1131341 and CAT rs1001179 polymorphic loci with longevity were carried out. Survival analysis was subsequently performed using the established single genes and gene-gene combinations as cofactors.

Results: The PON1 rs662*G allele was defined as the main longevity marker in women (OR = 1.44, p = 3E-04 in the log-additive model; HR = 0.77, p = 1.9E-04 in the Cox-survival model). The polymorphisms in the MTHFR, MSRA, PON2, SOD1, and CAT genes had an additive effect on longevity. A strong protective effect of combined MTHFR rs1801133*C, MSRA rs10098474*T, PON1 rs662*G, and PON2 rs7493*C alleles against mortality was obtained in women (HR = 0.81, p = 5E-03). The PON1 rs662*A allele had a meaningful impact on mortality for both long-lived men with cerebrovascular accidents (HR = 1.76, p = 0.027 for the PON1 rs662*AG genotype) and women with cardiovascular diseases (HR = 1.43, p = 0.002 for PON1 rs662*AA genotype). The MTHFR rs1801133*TT (HR = 1.91, p = 0.036), CAT rs1001179*TT (HR = 2.83, p = 0.031) and SOD1 rs2070424*AG (HR = 1.58, p = 0.018) genotypes were associated with the cancer mortality.

Conclusion: In our longitudinal 20-year study, we found the combinations of functional polymorphisms in antioxidant genes involved in longevity and survival in certain clinical phenotypes in the advanced age.

Background: Biomarker-disease relationships are extensively investigated. However, associations between common clinical biomarkers and healthspan, the disease-free lifespan, are largely unknown. We aimed to explore the predictive values of ten biomarkers on healthspan and lifespan, and to identify putative causal mechanisms.

Methods: Using data from 12,098 Swedish individuals aged 47-94 years, we examined both serum concentrations and genetically predicted levels of ten glycemic, lipid-, inflammatory, and hematological biomarkers. During a follow-up period of up to 16 years, 3681 incident cases of any chronic disease (i.e., end of healthspan) and 2674 deaths (i.e., end of lifespan) were documented. Cox regression models were applied to estimate the associations of a one standard deviation increase in biomarkers with healthspan and lifespan.

Findings: Seven out of ten serum biomarkers were significantly associated with risks of any chronic disease and death; elevated glycemic biomarkers and high-density lipoprotein-related biomarkers showed the strongest detrimental (hazard ratio [HR] 1·29 [95% CI 1·24-1·34]) and protective effects (HR 0·92 [95% CI 0·89-0·96]), respectively. Genetic predisposition to elevated fasting blood glucose (FBG) was associated with increased risks of any chronic disease (HR 1·05 [95% CI 1·02-1·09]); genetically determined higher C-reactive protein correlated with lower death risks (HR 0·91 [95% CI 0·87-0·95]). Notably, the genetically proxied FBG-healthspan association was largely explained by serum FBG concentration.

Interpretation: Circulating concentrations of glycemic, lipid-, and inflammatory biomarkers are predictive of healthspan and lifespan. Glucose control is a putative causal mechanism and a potential intervention target for healthspan maintenance.

Funding: This study was supported by the Swedish Research Council (2015-03,255, 2018-02,077), FORTE (2013-2292), the Loo & Hans Osterman Foundation, the Foundation for Geriatric Diseases, the Magnus Bergwall Foundation, the Strategic Research Program in Epidemiology at Karolinska Institutet (SH, JJ), the China Scholarship Council, and the Swedish National Graduate School for Competitive Science on Ageing and Health. The Swedish Twin Registry is managed by Karolinska Institutet and receives funding as an infrastructure through the Swedish Research Council, 2017-00,641.

Genetic studies have thus far identified a limited number of loci associated with human longevity by applying age at death or survival up to advanced ages as phenotype. As an alternative approach, one could first try to identify biomarkers of healthy ageing and the genetic variants associated with these traits and subsequently determine the association of these variants with human longevity. In the present study, we used this approach by testing whether the 35 baseline serum parameters measured in the Leiden Longevity Study (LLS) meet the proposed criteria for a biomarker of healthy ageing. The LLS consists of 421 families with long-lived siblings of European descent, who were recruited together with their offspring and the spouses of the offspring (controls). To test the four criteria for a biomarker of healthy ageing in the LLS, we determined the association of the serum parameters with chronological age, familial longevity, general practitioner-reported general health, and mortality. Out of the 35 serum parameters, we identified glucose, insulin, and triglycerides as biomarkers of healthy ageing, meeting all four criteria in the LLS. We subsequently showed that the genetic variants previously associated with these parameters are significantly enriched in the largest genome-wide association study for human longevity. In conclusion, we showed that biomarkers of healthy ageing can be used to leverage genetic studies into human longevity. We identified several genetic variants influencing the variation in glucose, insulin and triglycerides that contribute to human longevity.

Background: A variety of factors, including diet and lifestyle, obesity, physiology, metabolism, hormone levels, psychology, and inflammation, have been associated with longevity. The specific influences of these factors, however, are poorly understood. Here, possible causal relationships between putative modifiable risk factors and longevity are investigated.

Methods: A random effects model was used to investigate the association between 25 putative risk factors and longevity. The study population comprised 11,262 long-lived subjects (≥90 years old, including 3484 individuals ≥99 years old) and 25,483 controls (≤60 years old), all of European ancestry. The data were obtained from the UK Biobank database. Genetic variations were used as instruments in two-sample Mendelian randomization to reduce bias. The odds ratios for genetically predicted SD unit increases were calculated for each putative risk factor. Egger regression was used to determine possible violations of the Mendelian randomization model.

Results: Thirteen potential risk factors showed significant associations with longevity (≥90th) after correction for multiple testing. These included smoking initiation (OR:1.606;

Ci: 1.112-2.319) and educational attainment (OR:2.538,

Ci: 1.685-3.823) in the diet and lifestyle category, systolic and diastolic blood pressure (OR per SD increase: 0.518;

Ci: 0.438-0.614 for SBP and 0.620; CI 0.514-0.748 for DBP) and venous thromboembolism (OR:0.002;

Ci: 0.000-0.047) in the physiology category, obesity (

Or: 0.874;

Ci: 0.796-0.960), BMI (OR per 1-SD increase: 0.691;

Ci: 0.628-0.760), and body size at age 10 (OR per 1-SD increase:0.728;

Ci: 0.595-0.890) in the obesity category, type 2 diabetes (T2D) (OR:0.854;

Ci: 0.816-0.894), LDL cholesterol (OR per 1-SD increase: 0.743;

Ci: 0.668-0.826), HDL cholesterol (OR per 1-SD increase: 1.243;

Ci: 1.112-1.390), total cholesterol (TC) (OR per 1-SD increase: 0.786;

Ci: 0.702-0.881), and triglycerides (TG) (OR per 1-SD increase: 0.865;

Ci: 0.749-0.998) in the metabolism category. Both longevity (≥90th) and super-longevity (≥99th), smoking initiation, body size at age 10, BMI, obesity, DBP, SBP, T2D, HDL, LDL, and TC were consistently associated with outcomes. The examination of underlying pathways found that BMI indirectly affected longevity through three pathways, namely, SBP, plasma lipids (HDL/TC/LDL), and T2D (p < 0.05).

Conclusion: BMI was found to significantly affect longevity through SBP, plasma lipid (HDL/TC/LDL), and T2D. Future strategies should focus on modifying BMI to improve health and longevity.

Biological aging is accompanied by increasing morbidity, mortality, and healthcare costs; however, its molecular mechanisms are poorly understood. Here, we use multi-omic methods to integrate genomic, transcriptomic, and metabolomic data and identify biological associations with four measures of epigenetic age acceleration and a human longevity phenotype comprising healthspan, lifespan, and exceptional longevity (multivariate longevity). Using transcriptomic imputation, fine-mapping, and conditional analysis, we identify 22 high confidence associations with epigenetic age acceleration and seven with multivariate longevity. FLOT1, KPNA4, and TMX2 are novel, high confidence genes associated with epigenetic age acceleration. In parallel, cis-instrument Mendelian randomization of the druggable genome associates TPMT and NHLRC1 with epigenetic aging, supporting transcriptomic imputation findings. Metabolomics Mendelian randomization identifies a negative effect of non-high-density lipoprotein cholesterol and associated lipoproteins on multivariate longevity, but not epigenetic age acceleration. Finally, cell-type enrichment analysis implicates immune cells and precursors in epigenetic age acceleration and, more modestly, multivariate longevity. Follow-up Mendelian randomization of immune cell traits suggests lymphocyte subpopulations and lymphocytic surface molecules affect multivariate longevity and epigenetic age acceleration. Our results highlight druggable targets and biological pathways involved in aging and facilitate multi-omic comparisons of epigenetic clocks and human longevity.

The study of parental lifespan has emerged as an innovative tool to advance aging biology and our understanding of the genetic architecture of human longevity and aging-associated diseases. Here, we leveraged summary statistics of a genome-wide association study including over one million parental lifespans to identify genetically regulated genes from the Genotype-Tissue Expression project. Through a combination of multi-tissue transcriptome-wide association analyses and genetic colocalization, we identified novel genes that may be associated with parental lifespan. Mendelian randomization (MR) analyses also identified circulating proteins and metabolites causally associated with parental lifespan and chronic diseases offering new drug repositioning opportunities such as those targeting apolipoprotein-B-containing lipoproteins. Liver expression of HP, the gene encoding haptoglobin, and plasma haptoglobin levels were causally linked with parental lifespan. Phenome-wide MR analyses were used to map genetically regulated genes, proteins and metabolites with other human traits as well as the disease-related phenome in the FinnGen cohorts (n = 135,638). Altogether, this study identified new candidate genes, circulating proteins and metabolites that may influence human aging as well as potential therapeutic targets for chronic diseases that warrant further investigation.

People who have a balanced diet and engage in more physical activity live longer, healthier lives. This study aimed to test the hypothesis that these associations reflect a slowing of biological processes of aging. We analyzed data from 42 625 participants (aged 20-84 years, 51% female participants) from the National Health and Nutrition Examination Surveys (NHANES), 1999-2018. We calculated adherence to a Mediterranean diet (MeDi) and level of leisure time physical activity (LTPA) using standard methods. We measured biological aging by applying the PhenoAge algorithm, developed using clinical and mortality data from NHANES-III (1988-94), to clinical chemistries measured from a blood draw at the time of the survey. We tested the associations of diet and physical activity measures with biological aging, explored synergies between these health behaviors, and tested heterogeneity in their associations across strata of age, sex, and body mass index. Participants who adhered to the MeDi and who did more LTPA had younger biological ages compared with those who had less-healthy lifestyles (high vs low MeDi tertiles: β = 0.14 standard deviation [SD] [95% confidence interval {CI}: -0.18, -0.11]; high vs sedentary LTPA, β = 0.12 SD [-0.15, -0.09]), in models controlled for demographic and socioeconomic characteristics. Healthy diet and regular physical activity were independently associated with lower clinically defined biological aging, regardless of age, sex, and BMI category.

In recent decades, global life expectancies have risen significantly, accompanied by a marked increase in chronic diseases and population aging. This narrative review aims to summarize recent findings on the dietary factors influencing chronic diseases and longevity, primarily from large cohort studies. First, maintaining a healthy weight throughout life is pivotal for healthy aging and longevity, mirroring the benefits of lifelong, moderate calorie restriction in today's obesogenic food environment. Second, the specific types or food sources of dietary fat, protein, and carbohydrates are more important in influencing chronic disease risk and mortality than their quantity. Third, some traditional diets (e.g., the Mediterranean, Nordic, and Okinawa) and contemporary dietary patterns, such as healthy plant-based diet index, the DASH (dietary approaches to stop hypertension) diet, and alternate healthy eating index, have been associated with lower mortality and healthy longevity. These patterns share many common components (e.g., a predominance of nutrient-rich plant foods; limited red and processed meats; culinary herbs and spices prevalent in global cuisines) while embracing distinct elements from different cultures. Fourth, combining a healthy diet with other lifestyle factors could extend disease-free life expectancies by 8-10 years. While adhering to core principles of healthy diets, it is crucial to adapt dietary recommendations to individual preferences and cultures as well as nutritional needs of aging populations. Public health strategies should aim to create a healthier food environment where nutritious options are readily accessible, especially in public institutions and care facilities for the elderly. Although further mechanistic studies and human trials are needed to better understand molecular effects of diet on aging, there is a pressing need to establish and maintain long-term cohorts studying diet and aging in culturally diverse populations.

Introduction: Previous studies have examined the impact of healthy lifestyle choices on health-related outcomes; however, given their fragmented, often cross-sectional nature, assessing the relative impact of daily modifiable behaviors on overall long-term outcomes, particularly for a diverse working adult population, remains challenging.

Methods: Relationships between ten self-reported healthy lifestyle behaviors and health outcomes during the subsequent 9 years in a cohort of 10,248 participants enrolled during 2003 in a voluntary workplace wellness program were assessed. Cox proportional-hazards models computed hazard ratios (HRs) for lifestyle characteristics associated with time to one of seven self-reported chronic diseases or death. Data were collected between 2003 and 2012 and analyzed between 2014 and 2016.

Results: Behaviors that most significantly affected future outcomes were low-fat diet, aerobic exercise, nonsmoking, and adequate sleep. A dose-response effect was seen between dietary fat intake and hypertension, obesity, diabetes, heart disease, and hypercholesterolemia. After dietary fat intake, aerobic exercise was the next most significant behavior associated with development of outcomes. Compared with sedentary participants, those who exercised 4 days per week were less likely to develop new-onset diabetes (HR=0.31, 95% CI=0.20, 0.48); heart disease (HR=0.46, 95% CI=0.27, 0.80); and hypercholesterolemia (HR=0.61, 95% CI=0.50, 0.74). Low-fat diet and adequate sleep were more significant than commonly promoted healthy behaviors, such as eating a daily breakfast.

Conclusions: Modifiable lifestyle behaviors targeted in health promotion programs should be prioritized in an evidence-based manner. Top priorities for workplace health promotion should include low-fat diet, aerobic exercise, nonsmoking, and adequate sleep.

Biological age uses biophysiological information to capture a person's age-related risk of adverse outcomes. MetaboAge and MetaboHealth are metabolomics-based biomarkers of biological age trained on chronological age and mortality risk, respectively. Lifestyle factors contribute to the extent chronological and biological age differ. The association of lifestyle factors with MetaboAge and MetaboHealth, potential sex differences in these associations, and MetaboAge's and MetaboHealth's sensitivity to lifestyle changes have not been studied yet. Linear regression analyses and mixed-effect models were used to examine the cross-sectional and longitudinal associations of scaled lifestyle factors with scaled MetaboAge and MetaboHealth in 24,332 middle-aged participants from the Doetinchem Cohort Study, Rotterdam Study, and UK Biobank. Random-effect meta-analyses were performed across cohorts. Repeated metabolomics measurements had a ten-year interval in the Doetinchem Cohort Study and a five-year interval in the UK Biobank. In the first study incorporating longitudinal information on MetaboAge and MetaboHealth, we demonstrate associations between current smoking, sleeping ≥8 hours/day, higher BMI, and larger waist circumference were associated with higher MetaboHealth, the latter two also with higher MetaboAge. Furthermore, adhering to the dietary and physical activity guidelines were inversely associated with MetaboHealth. Lastly, we observed sex differences in the associations between alcohol use and MetaboHealth.

The main finding of this study was that a high overall healthy lifestyle score and various lifestyle profiles characterized by 4 optimal lifestyle factors were associated with significant gains in years lived without major noncommunicable diseases between ages 40 and 75 years in both sexes. Comparing the best with the worst lifestyle score was associated with approximately 9 additional years without chronic diseases. A 1-point advantage in healthy lifestyle score was associated with an almost 1-year increase in years spent without type 2 diabetes, coronary heart disease, stroke, cancer, asthma, and COPD. Of the 16 different lifestyle profiles studied, all 4 that were associated with the longest disease-free life span included a BMI less than 25 and at least 2 of the following health behaviors: never smoking, physical activity, and moderate alcohol consumption. The results were essentially the same when heart failure and dementia—2 further common conditions of older age—were considered in addition to the other 6 diseases.

We are not aware of other large-scale investigations on the different combinations of common lifestyle factors and disease-free life-years. Our findings suggest that normal weight is a particularly important component of the lifestyle profiles, although a greater total number of optimal lifestyle factors also characterized individuals who achieved a higher age without chronic disease. Our findings do not support a synergistic role for any specific combination of lifestyle factors; rather, a normal BMI, never smoking, physical activity, and moderate alcohol consumption appear to be associated with health span in a way that is consistent with an additive effect.

Our results regarding overall lifestyle score are comparable to those reported in previous studies on disease-free years and healthy lifestyle factors using heterogeneous operationalizations of the exposure and outcome. For example, a study of 33 000 men and women aged 20 to 70 years found approximately 2 extra disease-free years from chronic diseases in participants with all vs none of nonsmoking status, BMI less than 25, physical activity, and adherence to a Mediterranean-style diet (excluding alcohol). In a pooled analysis of 4 cohort studies, participants who were not smokers, physically inactive, or obese lived several years longer without 4 chronic diseases than those with at least 2 of these risk factors.

Background: Americans have a shorter life expectancy compared with residents of almost all other high-income countries. We aim to estimate the impact of lifestyle factors on premature mortality and life expectancy in the US population.

Methods: Using data from the Nurses' Health Study (1980-2014; n=78 865) and the Health Professionals Follow-up Study (1986-2014, n=44 354), we defined 5 low-risk lifestyle factors as never smoking, body mass index of 18.5 to 24.9 kg/m2, ≥30 min/d of moderate to vigorous physical activity, moderate alcohol intake, and a high diet quality score (upper 40%), and estimated hazard ratios for the association of total lifestyle score (0-5 scale) with mortality. We used data from the NHANES (National Health and Nutrition Examination Surveys; 2013-2014) to estimate the distribution of the lifestyle score and the US Centers for Disease Control and Prevention WONDER database to derive the age-specific death rates of Americans. We applied the life table method to estimate life expectancy by levels of the lifestyle score.

Results: During up to 34 years of follow-up, we documented 42 167 deaths. The multivariable-adjusted hazard ratios for mortality in adults with 5 compared with zero low-risk factors were 0.26 (95% confidence interval [CI], 0.22-0.31) for all-cause mortality, 0.35 (95% CI, 0.27-0.45) for cancer mortality, and 0.18 (95% CI, 0.12-0.26) for cardiovascular disease mortality. The population-attributable risk of nonadherence to 5 low-risk factors was 60.7% (95% CI, 53.6-66.7) for all-cause mortality, 51.7% (95% CI, 37.1-62.9) for cancer mortality, and 71.7% (95% CI, 58.1-81.0) for cardiovascular disease mortality. We estimated that the life expectancy at age 50 years was 29.0 years (95% CI, 28.3-29.8) for women and 25.5 years (95% CI, 24.7-26.2) for men who adopted zero low-risk lifestyle factors. In contrast, for those who adopted all 5 low-risk factors, we projected a life expectancy at age 50 years of 43.1 years (95% CI, 41.3-44.9) for women and 37.6 years (95% CI, 35.8-39.4) for men. The projected life expectancy at age 50 years was on average 14.0 years (95% CI, 11.8-16.2) longer among female Americans with 5 low-risk factors compared with those with zero low-risk factors; for men, the difference was 12.2 years (95% CI, 10.1-14.2).

Conclusions: Adopting a healthy lifestyle could substantially reduce premature mortality and prolong life expectancy in US adults.

Introduction: Lifestyle-related habits have a strong influence on morbidity and mortality worldwide. This study investigates the association between a multidimensional healthy lifestyle score and all-cause mortality risk, including in the score some less-studied lifestyle-related factors.

Methods: Participants (n=20,094) of the Seguimiento Universidad de Navarra cohort were followed up from 1999 to 2018. The analysis was conducted in 2019. A 10-point healthy lifestyle score previously associated with a lower risk of major cardiovascular events was applied, assigning 1 point to each of the following items: never smoking, moderate-to-high physical activity, moderate-to-high Mediterranean diet adherence, healthy BMI, moderate alcohol consumption, avoidance of binge drinking, low TV exposure, short afternoon nap, time spent with friends, and working ≥40 hours per week.

Results: During a median follow-up of 10.8 years, 407 deaths were documented. In the multivariable adjusted analysis, the highest category of adherence to the score (7-10 points) showed a 60% lower risk of all-cause mortality than the lowest category (0-3 points) (hazard ratio=0.40, 95% CI=0.27, 0.60, p<0.001 for trend). In analyses of the healthy lifestyle score as a continuous variable, for each additional point in the score, a 18% relatively lower risk of all-cause mortality was observed (adjusted hazard ratio=0.82, 95% CI=0.76, 0.88).

Conclusions: Adherence to a healthy lifestyle score, including some less-studied lifestyle-related factors, was longitudinally associated with a substantially lower mortality rate in a Mediterranean cohort. Comprehensive health promotion should be a public health priority.

As evidence that healthy lifestyle change is both necessary and attainable, we highlight the substantial bodies of research that link health behaviors with cardiovascular health and establish that health behavior change is feasible, improves health outcomes, and lowers healthcare costs.
The strong links between individual health behaviors and cardiovascular health were summarized in detail in the AHA’s 2020 strategic impact goals statement. Subsequent prospective studies have demonstrated that the presence of ideal cardiovascular health behaviors and biomarkers is associated with longevity and freedom from cardiovascular and all-cause morbidity and mortality. Ford and colleagues observed that adults who had ≥5 of the 7 metrics at ideal levels, compared with 0 metrics, were at 78% lower risk for all-cause mortality and 88% lower risk for cardiovascular disease mortality over 5.8 years of follow-up. Similarly, over 14.8 years of follow-up, Yang and colleagues observed a graded reduction in all-cause, cardiovascular disease, and ischemic heart disease mortality in conjunction with having greater numbers of metrics at ideal levels at baseline, which suggests that substantial proportions of deaths could be prevented or postponed with greater achievement of ideal cardiovascular health. Higher numbers of ideal health metrics were also associated with lower cancer mortality. Likewise, over 20 years of follow-up, adults in the Atherosclerosis Risk in Communities Study who had higher numbers of either health behaviors or biomarkers at ideal levels had dramatically lower cardiovascular disease event rates, and results were similar for blacks and whites.
Findings from the Coronary Artery Risk Development in Young Adults (CARDIA) study demonstrate that young adults aged 18 to 30 years with healthy behaviors grow up to become older adults with lower biological risk. Among those who practiced all of 5 healthy lifestyle behaviors in young adulthood (nonsmoking, low or no alcohol use, healthy diet, physically active, maintenance of normal weight), 60% had all ideal cardiovascular biomarkers in middle age. In contrast, of those who practiced none of these healthy behaviors in young adulthood, fewer than 5% reached middle age with all cardiovascular biomarkers at ideal levels.

Background: The impact of multiple healthy lifestyle factors on survival time is unclear.

Objective: The aim of this study was to examine differences in survival time associated with a healthy lifestyle versus a less healthy lifestyle.

Methods: This study consisted of 33 454 men (Cohort of Swedish Men) and 30 639 women (Swedish Mammography Cohort) aged 45-83 years and free of cancer and cardiovascular disease at baseline. The healthy lifestyle factors included the following: (i) nonsmoking; (ii) physical activity at least 150 min per week; (iii) alcohol consumption of 0-14 drinks per week; (iv) and healthy diet defined as a modified Dietary Approaches to Stop Hypertension Diet score above the median. Cox proportional hazards regression models and Laplace regression were used to estimate, respectively, hazard ratios of all-cause mortality and differences in survival time.

Results: During follow-up from 1998 through 2014, 8630 deaths amongst men and 6730 deaths amongst women were ascertained through linkage to the Swedish Cause of Death Register. Each of the four healthy lifestyle factors was inversely associated with all-cause mortality and increased survival time. Compared with individuals with no or one healthy lifestyle factor, the multivariable hazard ratios of all-cause mortality for individuals with all four health behaviours were 0.47 (95% 95% confidence interval [CI]: 0.44-0.51) in men and 0.39 (95%

Ci: 0.35-0.44) in women. This corresponded to a difference in survival time of 4.1 (95%

Ci: 3.6-4.6) years in men and 4.9 (95%

Ci: 4.3-5.6) years in women.

Conclusion: Adopting healthy lifestyle behaviours may markedly increase lifespan.

Background: There is poor knowledge on the association between combined lifestyles with mortality risk among individuals at high risk, and little is known on the biological mechanisms that could be on the pathway.

Methods: Longitudinal analysis on 22 839 individuals from the Moli-sani Study (Italy, 2005-2010). Among them, we identified 5200 elderly individuals (≥65 year), 2127 subjects with diabetes and 1180 with cardiovascular disease (CVD) at baseline. A healthy lifestyle score (HLS) was calculated, allocating 1 point for each of the following: abstention from smoking; adherence to Mediterranean diet; physical activity; absence of abdominal obesity. Hazard ratios (HR) with 95% confidence intervals (95%CI) were calculated by multivariable Cox regression and competing risk models.

Results: During 8.2 years of follow-up, 1237 deaths occurred. In the general population, adherence to all four healthy lifestyles, compared with none or 1, was associated with lower risk of all-cause (HR = 0.53; 95%CI:0.39-0.72), CVD (HR = 0.54; 0.32-0.91), cancer (HR = 0.62; 0.39-1.00) and mortality from other causes (HR = 0.39; 0.19-0.81). A 1-point increase in HLS was associated with 20%, 22% and 24% lower risk of total mortality among the elderly, in subjects with diabetes or CVD, respectively. Traditional (e.g. blood lipids), inflammatory (e.g. C-reactive protein) and novel biomarkers (e.g. markers of cardiac damage) accounted for up to 24% of the association of HLS with all-cause mortality risk in the general population.

Conclusions: The impact of combined four healthy lifestyles on survival was considerable, both in the general population and among high-risk subgroups. Inflammatory and novel biomarkers of CVD risk explained a substantial proportion of this association.

Importance 
General health checks, also known as general medical examinations, periodic health evaluations, checkups, routine visits, or wellness visits, are commonly performed in adult primary care to identify and prevent disease. Although general health checks are often expected and advocated by patients, clinicians, insurers, and health systems, others question their value.
Observations 
Randomized trials and observational studies with control groups reported in prior systematic reviews and an updated literature review through March 2021 were included. Among 19 randomized trials (906 to 59 616 participants; follow-up, 1 to 30 years), 5 evaluated a single general health check, 7 evaluated annual health checks, 1 evaluated biannual checks, and 6 evaluated health checks delivered at other frequencies. Twelve of 13 observational studies (240 to 471 415 participants; follow-up, cross-sectional to 5 years) evaluated a single general health check. General health checks were generally not associated with decreased mortality, cardiovascular events, or cardiovascular disease incidence. For example, in the South-East London Screening Study (n = 7229), adults aged 40 to 64 years who were invited to 2 health checks over 2 years, compared with adults not invited to screening, experienced no 8-year mortality benefit (6% vs 5%). General health checks were associated with increased detection of chronic diseases, such as depression and hypertension; moderate improvements in controlling risk factors, such as blood pressure and cholesterol; increased clinical preventive service uptake, such as colorectal and cervical cancer screening; and improvements in patient-reported outcomes, such as quality of life and self-rated health. In the Danish Check-In Study (n = 1104), more patients randomized to receive to a single health check, compared with those randomized to receive usual care, received a new antidepressant prescription over 1 year (5% vs 2%; P = .007). In a propensity score–matched analysis (n = 8917), a higher percentage of patients who attended a Medicare Annual Wellness Visit, compared with those who did not, underwent colorectal cancer screening (69% vs 60%; P < .01). General health checks were sometimes associated with modest improvements in health behaviors such as physical activity and diet. In the OXCHECK trial (n = 4121), fewer patients randomized to receive annual health checks, compared with those not randomized to receive health checks, exercised less than once per month (68% vs 71%; difference, 3.3% [95% CI, 0.5%-6.1%]). Potential adverse effects in individual studies included an increased risk of stroke and increased mortality attributed to increased completion of advance directives.
Conclusions and Relevance 
General health checks were not associated with reduced mortality or cardiovascular events, but were associated with increased chronic disease recognition and treatment, risk factor control, preventive service uptake, and improved patient-reported outcomes. Primary care teams may reasonably offer general health checks, especially for groups at high risk of overdue preventive services, uncontrolled risk factors, low self-rated health, or poor connection or inadequate access to primary care.

In general, a comprehensive assessment of risk factors should be performed at least every 5 years starting at 18 years of age, and a global risk score should be calculated at least every 5 years starting at the age of 35 years for men and 45 years for women. Those with increased cardiovascular risk, for example, those with diabetes, cigarette smokers, or those with obesity, should have their risk factors and cardiovascular risk assessed more frequently.

Peripheral immune markers are widely used to predict risk for inflammatory disease. However, whether single assessments of inflammatory biomarkers represent stable individual differences remains unclear. We reviewed 50 studies (N = 48,674; 57 % male; mean age 54 (range 13-79) years) that assessed markers of inflammation on >1 occasion, with time between measures ranging from 24 h to 7+ years. Separate random effects meta-analyses were conducted for each inflammatory marker and time interval. Markers that had broad coverage across most time intervals included C-reactive protein (CRP; k = 37), interleukin (IL)-6 (k = 22), TNF-α (k = 10), and fibrinogen (Fg; k = 9). For CRP, IL-6, and TNF-α, stability estimates generally decreased with time, with strong to moderate stability over intervals <6 months (r's = 0.80-0.61), modest to moderate stability over 6 months - 3 years (r's = 0.60-0.51), and low stability for >3 years (r's = 0.39-0.30). Estimates were less reliable for Fg for time intervals ≤ 3 years although they generally followed the same pattern; more reliable findings suggested greater stability for Fg than other markers for intervals >3 years (r = 0.53). These findings suggest that single measures of inflammatory biomarkers may be an adequate index of stable individual differences in the short term (<6 months), with repeated measures of inflammatory biomarkers recommended over intervals ≥ 6 months to 3 years, and absolutely necessary over intervals >3 years to reliably identify stable individual differences in health risk. These findings are consistent with stability estimates and clinical recommendations for repeated measurement of other cardiovascular measures of risk (e.g., blood lipids, blood pressure).

Blood biomarkers can be instrumental for understanding biologic processes and for targeting cardiovascular interventions, as has been shown with the measurement and pharmacologic reduction of low-density lipoprotein (LDL) cholesterol. At a time when the clinical community is moving beyond the evaluation and reduction of LDL cholesterol alone, high-sensitivity C-reactive protein (CRP), a biomarker of low-grade vascular inflammation, and lipoprotein(a), a genetically determined lipid fraction, have also become important for targeting cardiovascular interventions. To date, three randomized, placebo-controlled trials have shown that reducing inflammation can significantly decrease the incidence of cardiovascular events, and one antiinflammatory agent, low-dose colchicine, was recently approved by the Food and Drug Administration (FDA) for reduction of atherosclerotic events. Several outcome trials of agents that lower lipoprotein(a) levels are ongoing.

Current guidelines for the primary and secondary prevention of atherosclerotic disease are shifting to include a broader assessment that includes measurement of high-sensitivity CRP and lipoprotein(a) levels in addition to LDL cholesterol levels, a shift that is consistent with findings from contemporary studies that included short-term (3-to-5-year) follow-up among persons receiving and those not receiving statin therapy. However, data are scarce with respect to the long-term (25-to-30-year) risks associated with these biomarkers alone and in combination. Because atherosclerotic disease develops over decades, yet early-life interventions represent an important method for risk reduction, these long-term risks are a major concern particularly among women, for whom cardiovascular disease remains underdiagnosed and undertreated.

Because lipoprotein(a) levels are determined genetically and are stable over time, measurement is recommended once without the need for repeat evaluation. Consequently, we hypothesized that measuring high-sensitivity CRP, LDL cholesterol, and lipoprotein(a) together at a single time point might provide a useful method for the assessment of lifetime cardiovascular risk.

Aims: We conducted a cohort study to determine the screening intervals of metabolic disorders.

Method: Participants without diabetes mellitus (DM), hypertension (HTN), dyslipidemia, and abdominal obesity who underwent health examinations (2005-2019) in Korea were included. Participants were grouped according to baseline fasting glucose, LDL-C level, blood pressure (BP), and waist circumference (WC). The time to develop metabolic disorders and the percentile of survival time was assessed in each group.

Result: The median follow-up duration was 4.94 years (n=222,413; mean age 37.13 ± 7.49 years). After 8.32(95 %CI 8.22-8.41), 3.01(2.89-3.31), and 1.11(1.03-1.25) years, 10 % of participants developed DM in fasting glucose levels of 100-110, 110-120, and 120-125 mg/dL, respectively. After 8.40(8.33-8.45), 6.33(6.20-6.47), and 1.99(1.97-2.00) years, 10 % developed HTN in BP 120/70, 120/70-130/80, and 130/80-140/90 mmHg, respectively. After 5.99(5.94-6.04), 2.84(2.77-2.90), and 1.36(1.30-1.44) years, 10 % developed dyslipidemia in LDL-C 100-120, 120-140, and 140-160 mg/dL, respectively. After 4.62(4.41-4.80) and 1.67(1.64-1.69) years, 10 % developed abdominal obesity in baseline WC < 80(Women;W)/85(Men;M) and < 85(W)/90(M) cm, respectively.

Conclusion: In adults aged 30-40, the screening interval of metabolic disorders should be individualized based on the baseline metabolic derangement. An individual with borderline values may need an annual screening.

Multi-marker approaches are well suited for untangling the intrinsic complexity of aging and related conditions. Herein, we quantified (1) baseline concentrations of a panel of geroscience biomarkers pertaining to four biological domains (i.e., metabolism, inflammation, vascular/organ dysfunction and cellular senescence, and neurodegeneration) in individuals aged ≥60 years; (2) investigated linear and non-linear changes in biomarker levels over a 6-year period according to age and sex; and (3) described the relationships among geroscience biomarkers at baseline and follow-up. We found that repeated measures of age-dependent changes of 47 blood-borne biomarkers over 6 years had differential associations depending on the biological domains. The most relevant biomolecules in the associations between age and repeated assessments were (1) adiponectin, C-peptide, renin (metabolism), (2) CXCL10, IL-1α, IL-1β, IL-6, IL-10, IL-12p70, MPO (inflammation), (3) cystatin C, MMP7, MMP12, VCAM-1 (vascular/organ dysfunction and cellular senescence), and (4) S100B and Tau protein (neurodegeneration). Among these molecules, a negative association with increasing age was found for IL-1α, IL-1β, IL-12p70, S100B, and Tau protein. Non-linear relationships were also identified with age for IGFBP-1, leptin, β2M, TNFRSF1B, fibrinogen, GDF-15, N-cadherin, and BDNF. Our results indicate that inflammatory and metabolic biomolecules are strongly associated with aging over 6 years of follow-up. Whether the biological pathways reflected by these biomarkers contribute to the aging process or are associated with negative health-related events needs to be explored through comprehensive multi-omics longitudinal analysis in larger cohorts.

Background: Defining laboratory biomarker reference values in a healthy population and understanding the fluctuations in biomarker concentrations throughout life and between sexes are critical to clinical interpretation of laboratory test results in different disease states. The Canadian Health Measures Survey (CHMS) has collected blood samples and health information from the Canadian household population. In collaboration with the Canadian Laboratory Initiative on Pediatric Reference Intervals (CALIPER), the data have been analyzed to determine reference value distributions and reference intervals for several endocrine and special chemistry biomarkers in pediatric, adult, and geriatric age groups.

Methods: CHMS collected data and blood samples from thousands of community participants aged 3 to 79 years. We used serum samples to measure 13 immunoassay-based special chemistry and endocrine markers. We assessed reference value distributions and, after excluding outliers, calculated age- and sex-specific reference intervals, along with corresponding 90% CIs, according to CLSI C28-A3 guidelines.

Results: We observed fluctuations in biomarker reference values across the pediatric, adult, and geriatric age range, with stratification required on the basis of age for all analytes. Additional sex partitions were required for apolipoprotein AI, homocysteine, ferritin, and high sensitivity C-reactive protein.

Conclusions: The unique collaboration between CALIPER and CHMS has enabled, for the first time, a detailed examination of the changes in various immunochemical markers that occur in healthy individuals of different ages. The robust age- and sex-specific reference intervals established in this study provide insight into the complex biological changes that take place throughout development and aging and will contribute to improved clinical test interpretation.

Background: Biological covariates such as age and sex can markedly influence biochemical marker reference values, but no comprehensive study has examined such changes across pediatric, adult, and geriatric ages. The Canadian Health Measures Survey (CHMS) collected comprehensive nationwide health information and blood samples from children and adults in the household population and, in collaboration with the Canadian Laboratory Initiative on Pediatric Reference Intervals (CALIPER), examined biological changes in biochemical markers from pediatric to geriatric age, establishing a comprehensive reference interval database for routine disease biomarkers.

Methods: The CHMS collected health information, physical measurements, and biosamples (blood and urine) from approximately 12 000 Canadians aged 3-79 years and measured 24 biochemical markers with the Ortho Vitros 5600 FS analyzer or a manual microplate. By use of CLSI C28-A3 guidelines, we determined age- and sex-specific reference intervals, including corresponding 90% CIs, on the basis of specific exclusion criteria.

Results: Biochemical marker reference values exhibited dynamic changes from pediatric to geriatric age. Most biochemical markers required some combination of age and/or sex partitioning. Two or more age partitions were required for all analytes except bicarbonate, which remained constant throughout life. Additional sex partitioning was required for most biomarkers, except bicarbonate, total cholesterol, total protein, urine iodine, and potassium.

Conclusions: Understanding the fluctuations in biochemical markers over a wide age range provides important insight into biological processes and facilitates clinical application of biochemical markers to monitor manifestation of various disease states. The CHMS-CALIPER collaboration addresses this important evidence gap and allows the establishment of robust pediatric and adult reference intervals.

A review of meta-analyses and
systematic reviews published before September 19, 2013 was conducted in 2 stages. In the
first stage, meta-analyses and systematic reviews published before April 2011 were
identified and reviewed. In a second stage, conducted to update the evidence base before
publication, additional meta-analyses and systematic reviews published before September
19, 2013 were identified and reviewed using the same criteria applied in the first stage.
The reliance on published meta-analyses to evaluate novel biomarkers is a conservative
approach that helps avoid the influence of positive publication bias that can occur early
in the evaluation of a novel association and assures that we relied on a mature body of
evidence ( 39 ).
Members of the Work Group proposed an initial list of novel risk markers for
inclusion in CQ1 which was then prioritized during several rounds of discussion. In
selecting the final list, the Work Group gave priority to factors that have engendered
substantial discussion in the scientific community and that could be reasonably considered
as potentially feasible for widespread population use by primary care providers in routine
clinical settings in the United States. These deliberations considered availability, cost,
assay reliability, and risks of the test or downstream testing. The final list of new risk
markers to be evaluated included several blood and urine biomarkers (hs-CRP, ApoB,
creatinine [or estimated GFR], and microalbuminuria), several measures of subclinical CV
disease (CAC, CIMT, and ABI), family history, and cardiorespiratory fitness. Other novel
potential screening tools may be the subject of future guideline updates. When considering
the utility of incorporating these new risk factors into routine risk assessment, guidance
published by Hlatky et al ( 40 ) was considered.
Special attention was given to the additional value these markers contributed to risk
assessment in terms of discrimination, calibration, reclassification, and

Early identification of individuals in the general population at high risk for atherosclerotic cardiovascular disease shapes primary preventive strategies to reduce the risk of developing atherosclerotic cardiovascular disease. Risk scores based on traditional risk factors for atherosclerotic cardiovascular disease (eg, the European Society of Cardiology Systematic Coronary Risk Evaluation 2 [SCORE2], the American Heart Association/American College of Cardiology Pooled Cohort Equations, and the American Heart Association Predicting Risk of Cardiovascular Disease Events [PREVENT] equations) are widely available to estimate an individual’s risk for future cardiovascular events.

Cardiovascular biomarkers, such as cardiac troponin, natriuretic peptides, and C-reactive protein (CRP), are established in clinical care. Using newer, high-sensitivity cardiac troponin assays, concentrations became measurable in the general population, opening up the prospects for a broader application of this biomarker. Several studies have reported (1) strong associations of these biomarkers with incident atherosclerotic cardiovascular disease events in individuals with known atherosclerotic cardiovascular disease, but also, and most importantly, in apparently healthy individuals and (2) an improvement in risk stratification when these biomarkers were added to established risk prediction models.

Notwithstanding the achievements of earlier studies, the actual application of routinely available cardiovascular biomarkers for risk stratification in primary prevention has not become routine clinical practice. In addition, it remains uncertain which of the established biomarkers might be best suited to predict each outcome and how such associations are influenced by age.

Therefore, this study brings together the largest multinational individual-level dataset, to date, to investigate the comparative predictive value of cardiovascular biomarkers for incident atherosclerotic cardiovascular disease events in the general population and to elucidate their differential effects according to age.

Many different biomarkers of ASCVD and/or T2DM risk have been identified in addition to the five “classic” components of metabolic syndrome, including uric acid, apolipoprotein B, lipoprotein(a), adiponectin, leptin, fasting insulin or proinsulin, free fatty acids, homocysteine, the PAI-1 gene, fibrinogen, alanine aminotransferase as a marker of liver fat, hsCRP, inflammatory cytokines ( e.g., IL-6), homocysteine, liver or myocellular fat content by magnetic resonance spectroscopy, and microalbuminuria (in individuals without diabetes). Some of these have also been identified as markers of high diabetes risk ( 26 ). Emerging literature also suggests genomic markers to predict T2DM and ASCVD. However, the Writing Committee cannot recommend the measurement of these markers for routine clinical practice until further research has been completed (see “4. Further Research” below).
One example of a widely debated marker is hsCRP ( 27 ). A high hsCRP level is indicative of a high ASCVD risk. The therapeutic consequence may be that general therapy to lower ASCVD risk should be initiated earlier than would be done without an elevated hsCRP level for a given Framingham risk score. In that case, measures might need to be taken to decrease LDL-C and BP to lower targets, but evidence to support specific lower targets has not yet been identified.
Studies have addressed the issue of whether hsCRP and other markers enhance the risk estimates of the well-known risk scores/engines ( 28 ). They conclude that adding hsCRP, or other novel risk markers, to more basic risk models does not improve the prediction of ASCVD risk, because most risk factors are interrelated and, by themselves, do not have good predictive value. Therefore, in a clinical setting, health care providers can rely on simple, less expensive measures, such as asking about family history, cigarette smoking, and measurements of BP and serum lipids.

The mean of these 3 measurements was assigned as participants’ blood pressure. Hemoglobin A 1c and total cholesterol were obtained by nonfasting laboratory measurement. Body mass index was obtained by measurement of weight and standing height and calculated as weight in kilograms divided by height in meters squared. Definitions for each cardiovascular risk factor were based on prior US Department of Health and Human Services and CDC reports to ensure consistency with prior health surveillance reports. Hypertension was defined as a systolic blood pressure of 140 mm Hg or greater, diastolic blood pressure of 90 mm Hg or greater, or a prescription for antihypertensive therapy. Diabetes was defined as a hemoglobin A 1c of 6.5% or greater or a health care diagnosis of diabetes. Hyperlipidemia was defined as a total cholesterol of 200 mg/dL or greater or a health care diagnosis of high cholesterol. Obesity was defined as a body mass index of 30 or greater. Smoking history was defined as self-report of smoking at least 100 cigarettes ever. Uniform definitions for all outcomes were used over the study period.

We also examined cardiovascular risk factor treatment and control rates. Treatment in adults with hypertension was defined as a self-report of taking antihypertensive medications. Treatment in adults with diabetes was defined as a self-report of taking pills for diabetes or insulin. Control was assessed among adults receiving treatment for each respective condition, consistent with prior reports. Hypertension was considered controlled if systolic blood pressure was less than 140 mm Hg and diastolic blood pressure was less than 90 mm Hg. Because the 2017 ACC/AHA guidelines recently lowered blood pressure targets for young adults from less than 140/90 mm Hg to less than 130/80 mm Hg, we also assessed control rates using this more stringent goal. Diabetes was considered controlled if hemoglobin A 1c was less than 7%. We did not assess treatment (or control) in young adults with hyperlipidemia given major changes in guideline recommendations over the study period that shifted away from lipid-level targets for treatment.

110 However, challenges remain in the implementation of widespread biomarker screening due to cost and clinical actionability when elevated levels are identified. In the current paradigm, they may be more appropriately considered for use as sequential diagnostic tests to evaluate for subclinical CVD and reclassify risk in selected patients. This would be analogous to diagnostic testing with CT for coronary artery calcium (CAC) measurement as recommended by the 2019 ACC/AHA primary prevention guideline for patients with borderline or intermediate 10-year risk for ASCVD when there is clinical uncertainty or patient indecision regarding drug therapy. 28
Other biomarkers, such as high-sensitivity C-reactive protein, carotid intima media thickness, and ankle brachial index were also reviewed; given the lack of routine clinical measurement in asymptomatic individuals, they were not included in the current models. Family history of premature CVD was discussed given the strong heritable component of CVD 111 but was deemed to be inconsistently ascertained in most clinical settings, and data from previous cohort studies also demonstrated that it did not significantly improve model performance. Last, emergence of data on the association of “OMIC” markers (eg, proteomics, metabolomics, genomics) with risk for incident CVD has yielded great enthusiasm for the potential of precision medicine approaches in risk prediction. 103 , 104 Although substantive advances in the mechanistic pathways of disease have been borne out by these cutting-edge investigations, the available data do not support the utility of large-scale genomic and proteomic scores for risk prediction in the general population at this time. 112 , 113 For example, polygenic risk scores for CHD do not clinically meaningfully improve risk discrimination when added to traditional risk factors in middle-aged to older adults. 114–116 Furthermore, when CAC and polygenic risk scores were compared directly, only CAC improved risk discrimination in 2 population-based cohorts of middle-aged to older adults.

Background: Physicians sometimes consider whether or not to perform diagnostic testing in healthy people, but it is unknown whether nonextreme values of diagnostic tests typically encountered in such populations have any predictive ability, in particular for risk of death. The goal of this study was to quantify the associations among population reference intervals of 152 common biomarkers with all-cause mortality in a representative, nondiseased sample of adults in the United States.

Methods: The study used an observational cohort derived from the National Health and Nutrition Examination Survey (NHANES), a representative sample of the United States population consisting of 6 survey waves from 1999 to 2010 with linked mortality data (unweighted N = 30 651) and a median followup of 6.1 years. We deployed an X-wide association study (XWAS) approach to systematically perform association testing of 152 diagnostic tests with all-cause mortality.

Results: After controlling for multiple hypotheses, we found that the values within reference intervals (10-90th percentiles) of 20 common biomarkers used as diagnostic tests or clinical measures were associated with all-cause mortality, including serum albumin, red cell distribution width, serum alkaline phosphatase, and others after adjusting for age (linear and quadratic terms), sex, race, income, chronic illness, and prior-year healthcare utilization. All biomarkers combined, however, explained only an additional 0.8% of the variance of mortality risk. We found modest year-to-year changes, or changes in association from survey wave to survey wave from 1999 to 2010 in the association sizes of biomarkers.

Conclusions: Reference and nonoutlying variation in common biomarkers are consistently associated with mortality risk in the US population, but their additive contribution in explaining mortality risk is minor.

The adult well-male examination should provide evidence-based guidance toward the promotion of optimal health and well-being. The medical history should focus on tobacco and alcohol use, risk of human immunodeficiency virus and other sexually transmitted infections, and diet and exercise habits. The physical examination should include blood pressure screening, and height and weight measurements to calculate body mass index. Lipid screening is performed in men 40 to 75 years of age; there is insufficient evidence for screening younger men. One-time screening ultrasonography for detection of abdominal aortic aneurysm is recommended in men 65 to 75 years of age who have ever smoked. Screening for prostate cancer using prostate-specific antigen testing in men 55 to 69 years of age should be individualized using shared decision making. Screening for colorectal cancer should begin at 50 years of age for average-risk men and continue until at least 75 years of age. Screening options include fecal immunochemical testing, colonoscopy, or computed tomography colonography. Lung cancer screening using low-dose computed tomography is recommended in men 55 to 80 years of age who have at least a 30-pack-year smoking history and currently smoke or have quit within the past 15 years. Immunizations should be updated according to guidelines from the Advisory Committee on Immunization Practices.

Laboratory test results drive about 70% of clinical decisions and are important in making diagnosis, prognosis, ruling out conditions, testing for propensity to disease and monitoring the course of illnesses. The requirement for releasing laboratory results to patients has altered the dynamics of doctor patient interaction. Minor variations in laboratory test results that are labeled by the laboratory as low/high/abnormal may cause unwarranted worry to the patients. The number of laboratory results that are outside the "normal range" far exceeds the clinically meaningful abnormal results due to the usually accepted methodology for ascertaining "normal values"/reference ranges, variations in methods of testing at different laboratories, variations due to age, gender, ethnicity, seasonality, and random variations. The usual process for establishing "normal values/reference ranges" entails testing at least 120 healthy individuals in a given age-group, gender, ethnicity, testing method and related health issues. The central 95% of the values is usually adopted as the normal range. This practice, by definition, labels 5% of healthy individuals as having abnormal laboratory results. This review addresses various issues that affect laboratory test results and interpretation of such results. It also addresses doctor and patient concerns about assessing and reporting laboratory results. In addition to reporting normal values along with patient results, it may be useful to include clinical significance of the findings, in simple terms, such as, no immediate concern, warrants discussion with doctor at the next visit, recommend contacting your doctor for further action.

Reference limits (RLs) are required to evaluate laboratory results for medical decisions. The establishment of RL depends on the pre-analytical and the analytical conditions. Furthermore, biological characteristics of the sub-population chosen to provide the reference samples may influence the RL. The most important biological preconditions are gender, age, chronobiological influences, posture, regional and ethnic effects. The influence of these components varies and is often neglected. Therefore, a list of biological variables is collected from the literature and their influence on the estimation of RL is discussed. Biological preconditions must be specified if RL are reported as well for directly as for indirectly estimated RL. The influence of biological variables is especially important if RL established by direct methods are compared with those derived from indirect techniques. Even if these factors are not incorporated into the estimation of RL, their understanding can assist the interpretation of laboratory results of an individual.

Reference intervals (RIs) for biochemical and hematological markers determined using healthy adult and/or pediatric populations are vital for clinical interpretation of laboratory test results. Most clinical laboratories commonly use age- and sex-specific RIs, but the effect of ethnicity as a covariate is often overlooked. Ethnic differences in serum biomarker concentrations can occur as a result of genetic and environmental factors, while the degree to which each factor influences serum levels depends on the specific biomarker. Numerous studies have investigated ethnic differences in routine chemistry, fertility, endocrine, cancer, and hematological markers, as well as in vitamins and carotenoids, in children, adolescents and adults. In the present review, we summarize and discuss ethnic-specific differences observed for these laboratory markers and their potential impact on the clinical interpretation of laboratory test results. We categorized the available data into seven major ethnic groups (i.e. Black, Caucasian, East Asian, Hispanic, South Asian, South East Asian, and West Asian) for ease of comparison. While certain biomarkers could not be compared between ethnic groups because of insufficient information or contradictory results between studies, significant differences between ethnic groups were reported by one or more studies for most of the biomarkers included in this review. The clinical significance of these differences and the potential need for ethnic-specific RIs for certain biochemical markers are also discussed.

The interpretation of laboratory data is a comparative procedure. Physicians typically need reference values to compare patients' laboratory data for clinical decisions. Therefore, establishing reliable reference data is essential for accurate diagnosis and patient monitoring. Human metabolism is a dynamic process. Various types of systematic and random fluctuations in the concentration/activity of biomolecules are observed in response to internal and external factors. In the human body, several biomolecules are under the influence of physiological rhythms and are therefore subject to ultradian, circadian and infradian fluctuations. In addition, most biomolecules are also characterized by random biological variations, which are referred to as biological fluctuations between subjects and within subjects/individuals. In routine practice, reference intervals based on population data are used, which by nature are not designed to capture physiological rhythms and random biological variations. To ensure safe and appropriate interpretation of patient laboratory data, reference intervals should be personalized and estimated using individual data in accordance with systematic and random variations. In this opinion paper, we outline (i) the main variations that contribute to the generation of personalized reference intervals (prRIs), (ii) the theoretical background of prRIs and (iii) propose new methods on how to harmonize prRIs with the systematic and random variations observed in metabolic activity, based on individuals' demography.

Clinical laboratory test results alone are of little value in diagnosing, treating, and monitoring health conditions; as such, a clinically actionable cutoff or reference interval is required to provide context for result interpretation. Healthcare practitioners base their diagnoses, follow-up treatments, and subsequent testing on these reference points. However, they may not be aware of inherent limitations related to the definition and derivation of reference intervals. Laboratorians are responsible for providing the reference intervals they report with results. Yet, the establishment and verification of reference intervals using conventional direct methods are complicated by resource constraints or unique patient demographics. To facilitate standardized reference interval best practices, multiple global scientific societies are actively drafting guidelines and seeking funding to promote these initiatives. Numerous national and international multicenter collaborations demonstrate the ability to leverage combined resources to conduct large reference interval studies by direct methods. However, not all demographics are equally accessible. Novel indirect methods are attractive solutions that utilize computational methods to define reference distributions and reference intervals from mixed data sets of pathologic and non-pathologic patient test results. In an effort to make reference intervals more accurate and personalized, individual-based reference intervals are shown to be more useful than population-based reference intervals in detecting clinically significant analyte changes in a patient that might otherwise go unrecognized when using wider, population-based reference intervals. Additionally, continuous reference intervals can provide more accurate ranges as compared to age-based partitions for individuals that are near the ends of the age partition. The advantages and disadvantages of different reference interval approaches as well as the advancement of non-conventional reference interval studies are discussed in this review.

The 5th and 95th percentile limits (−1.645 and +1.645 z-score) of the healthy population can be used to identify individuals with unusually low or high results, respectively.
Ideally, limits of normal ought to be based on an individual's pre-disease measure or baseline. Further clinical decision making requires relevant thresholds based on prognosis or clinical risk of adverse outcomes. To date, no satisfactory outcome-based thresholds for lung function have been defined; therefore, careful consideration of the medical and exposure history of an individual is necessary when interpreting lung function results when using the limits of normal. Further research to establish a comprehensive disease-specific clinical approach to interpretation (not simply relying on whether results are within or outside the normal range) is necessary. It is the consensus of the Task Force that the percentile limits represent a standardised and unbiased approached to identify values outside the range of expected results from a normal population.
A reference range represents the distribution of values that are expected in a healthy population and the lower limit of normal (LLN) represents a cut-off to define results that are outside the range of values typically observed in health. This approach is used for many clinical outcomes in medicine [ 47 – 49 ]. Population-defined z-scores or percentile values describe the chance the observed result falls within the distribution of values in healthy individuals ( figure 2 ). At the 5th percentile (corresponding to a z-score of −1.645), there is a 5% chance that the results in a healthy individual would be at or below this level, as shown in figure 2 . At the 1st percentile, there would be a 1% chance. Since typically for spirometry, low values are considered abnormal, it has become standard to define the LLN as the 5th percentile, accepting that this will result in 5% of healthy individuals having a false-positive result (i.e. being incorrectly classified as having an abnormal result). The 5th percentile represents a trade-off between incorrectly classifying a low value in a healthy individual and missing a clinically significant reduction in lung function (i.e. increased sensitivity for less specificity compared with using a lower percentile).

Hypothyroidism has been associated with atherosclerosis, and evidence from large observational studies suggests that individuals with hypothyroidism may be at increased risk of coronary heart disease (CHD) events and CHD mortality. Among people with apparently normal thyroid function, thyrotropin levels in the upper part of the reference range may indicate an early stage of hypothyroidism. In addition, such thyrotropin levels have been associated with an adverse cardiovascular risk profile that includes high levels of non–high-density lipoprotein cholesterol, triglycerides, blood pressure, and body mass index, as well as low levels of high-density lipoprotein cholesterol. Therefore, it seems plausible that differences in thyroid function within the population reference range may be differentially associated with CHD risk. In support of this hypothesis, the results of one cohort study suggested that thyrotropin levels within the reference range may be positively associated with CHD mortality in women, but not in men. In contrast, the results of other studies do not suggest any association between thyrotropin levels within the reference range and the risk of CHD events, vascular mortality, or need for coronary revascularization.

The definition of the clinically normal range for thyroid function is controversial. Some experts have suggested that the upper thyrotropin reference limit should be lowered from approximately 4.5 to 2.5 or 3.0 mIU/L since higher thyrotropin levels may indicate early-stage hypothyroidism. Others disagree, because many people with a healthy thyroid gland would thereby be classified with abnormal thyroid function, and there is no firm evidence that thyrotropin levels in the upper part of the reference range are associated with health consequences that would benefit from early recognition and treatment of hypothyroidism. Most research on such potential consequences has focused on CHD and its risk factors, and a recent review concluded that there was good evidence for associations between higher thyrotropin levels within the reference range and cardiovascular risk factors and events. However, most of this evidence comes from cross-sectional studies, which are inferior to prospective studies for causal inference.