DNA Methylation-Based Estimates of Circulating Leukocyte Composition for Predicting Colorectal Cancer Survival: A Prospective Cohort Study.

Leukocytes are involved in the progression of colorectal cancer (CRC). The proportion of six major leukocyte subtypes can be estimated using epigenome-wide DNA methylation (DNAm) data from stored blood samples. Whether the composition of circulating leukocytes can be used as a prognostic factor is unclear. DNAm-based leukocyte proportions were obtained from a prospective cohort of 2206 CRC patients. Multivariate Cox regression models and survival curves were applied to assess associations between leukocyte composition and survival outcomes. A higher proportion of lymphocytes, including CD4+ T cells, CD8+ T cells, B cells, and NK cells, was associated with better survival, while a higher proportion of neutrophils was associated with poorer survival. CD4+ T cells outperformed other leukocytes in estimating the patients' prognosis. Comparing the highest quantile to the lowest quantile of CD4+ T cells, hazard ratios (95% confidence intervals) of all-cause and CRC-specific mortality were 0.59 (0.48, 0.72) and 0.59 (0.45, 0.77), respectively. Furthermore, the association of CD4+ T cells and prognosis was stronger among patients with early or intermediate CRC or patients with colon cancer. In conclusion, the composition of circulating leukocytes estimated from DNAm, particularly the proportions of CD4+ T cells, could be used as promising independent predictors of CRC survival.

Sarcoma classification by DNA methylation profiling.

Sarcomas are malignant soft tissue and bone tumours affecting adults, adolescents and children. They represent a morphologically heterogeneous class of tumours and some entities lack defining histopathological features. Therefore, the diagnosis of sarcomas is burdened with a high inter-observer variability and misclassification rate. Here, we demonstrate classification of soft tissue and bone tumours using a machine learning classifier algorithm based on array-generated DNA methylation data. This sarcoma classifier is trained using a dataset of 1077 methylation profiles from comprehensively pre-characterized cases comprising 62 tumour methylation classes constituting a broad range of soft tissue and bone sarcoma subtypes across the entire age spectrum. The performance is validated in a cohort of 428 sarcomatous tumours, of which 322 cases were classified by the sarcoma classifier. Our results demonstrate the potential of the DNA methylation-based sarcoma classification for research and future diagnostic applications.

Blood-derived DNA methylation predictors of mortality discriminate tumor and healthy tissue in multiple organs.

Evidence has shown that certain methylation markers derived from blood can mirror corresponding methylation signatures in internal tissues. In the current study, we aimed to investigate two strong epigenetic predictors for life span, derived from blood DNA methylation data, in tissue samples of solid cancer patients. Using data from the Cancer Genome Atlas (TCGA) and the German DACHS study, we compared a mortality risk score (MRscore) and DNAmPhenoAge in paired tumor and adjacent normal tissue samples of patients with lung (N = 69), colorectal (n = 299), breast (n = 90), head/neck (n = 50), prostate (n = 50), and liver (n = 50) cancer. To explore the concordance across tissue and blood, we additionally assessed the two markers in blood samples of colorectal cancer (CRC) cases and matched controls (n = 93) in the DACHS+ study. The MRscore was significantly elevated in tumor tissues compared to normal tissues of all cancers except prostate cancer, for which an opposite pattern was observed. DNAmPhenoAge was consistently higher in all tumor tissues. The MRscore discriminated lung, colorectal, and prostate tumor tissues from normal tissues with very high accuracy [AUCs of 0.87, 0.99 (TCGA) /0.94 (DACHS), and 0.92, respectively]. DNAmPhenoAge accurately discriminated five types of tumor tissues from normal tissues (except prostate cancer), with AUCs of 0.82-0.93. The MRscore was also significantly higher in blood samples of CRC cases than in controls, with areas under the curve (AUC) of 0.74, whereas DNAmPhenoAge did not distinguish cases from controls, with AUC of 0.54. This study provides compelling evidence that blood-derived DNAm markers could reflect methylation changes in less accessible tissues. Further research should explore the potential use of these findings for cancer diagnosis and early detection.

Mutational patterns and regulatory networks in epigenetic subgroups of meningioma.

DNA methylation patterns delineate clinically relevant subgroups of meningioma. We previously established the six meningioma methylation classes (MC) benign 1-3, intermediate A and B, and malignant. Here, we set out to identify subgroup-specific mutational patterns and gene regulation. Whole genome sequencing was performed on 62 samples across all MCs and WHO grades from 62 patients with matched blood control, including 40 sporadic meningiomas and 22 meningiomas arising after radiation (Mrad). RNA sequencing was added for 18 of these cases and chromatin-immunoprecipitation for histone H3 lysine 27 acetylation (H3K27ac) followed by sequencing (ChIP-seq) for 16 samples. Besides the known mutations in meningioma, structural variants were found as the mechanism of NF2 inactivation in a small subset (5%) of sporadic meningiomas, similar to previous reports for Mrad. Aberrations of DMD were found to be enriched in MCs with NF2 mutations, and DMD was among the most differentially upregulated genes in NF2 mutant compared to NF2 wild-type cases. The mutational signature AC3, which has been associated with defects in homologous recombination repair (HRR), was detected in both sporadic meningioma and Mrad, but widely distributed across the genome in sporadic cases and enriched near genomic breakpoints in Mrad. Compared to the other MCs, the number of single nucleotide variants matching the AC3 pattern was significantly higher in the malignant MC, which also exhibited higher genomic instability, determined by the numbers of both large segments affected by copy number alterations and breakpoints between large segments. ChIP-seq analysis for H3K27ac revealed a specific activation of genes regulated by the transcription factor FOXM1 in the malignant MC. This analysis also revealed a super enhancer near the HOXD gene cluster in this MC, which, together with general upregulation of HOX genes in the malignant MC, indicates a role of HOX genes in meningioma aggressiveness. This data elucidates the biological mechanisms rendering different epigenetic subgroups of meningiomas, and suggests leveraging HRR as a novel therapeutic target.

Effect of milking stall dimensions on upper limb and shoulder muscle activity in milkers.

Increasing societal awareness for animal welfare can promote changes in legislation. Some of these changes may also affect the person that interacts with the animal in a shared workspace, such as in milking stalls. Swiss milking stalls were designed many years ago, when cows were smaller than they are today. A recent animal-based study indicated that welfare decreased in cows exposed to restricted space allowance in milking stalls, which had resulted from increasing body size without adjustment of milking stall dimensions. However, changing the milking stall dimensions without considering the milker may be detrimental. For many years, health issues, particularly of the upper limb and shoulders, have affected milking personnel. The current study investigated the effect of large and standard milking stall dimensions on muscle activity in milkers (as a measure of workload) during milking. This assessment is fundamental to ensure that legislation improving animal welfare does not jeopardize human health. The study took place in an experimental milking parlor that allowed for size adjustment of the individual milking stall. Nine milkers performed 2 shifts of milking in a herringbone and 2 shifts in a side-by-side milking parlor. The milking stall dimensions were large on one side and standard on the other side of the parlor; the 2 sides were switched between milking shifts. We used surface electromyography to monitor bilateral muscle activity of forearm (flexor carpi ulnaris), arm (biceps brachii), and shoulder (deltoideus anterior; upper trapezius) muscles. Statistical analysis was performed separately for the herringbone and the side-by-side parlor for each muscle using mean and maximum muscle activity as the target variables in a linear mixed-effects model. The analysis showed that the different milking stall dimensions did not consistently affect activity of the measured muscles. Our results suggest that milking stall dimensions are not a primary risk factor for poor ergonomics in parlor workers.

Long intergenic noncoding RNA 299 methylation in peripheral blood is a biomarker for triple-negative breast cancer.

AIM: To identify DNA methylation biomarkers in peripheral blood samples from triple-negative breast cancer (TNBC) patients. MATERIALS & METHODS: We conducted an epigenome-wide association study (EWAS): the most promising markers were identified in 233 TNBC case-control pairs (discovery set) and subsequently validated in an independent validation set (57 TNBC patients and 124 controls). RESULTS: cg06588802 (LINC00299/ID2) showed a higher methylation in TNBC patients compared with controls (discovery set: 3% increase, p-value = 0.0009; validation set: 2% increase, p-value = 0.01). Consistent results at four neighboring methylation probes and the strong negative correlation (rho = -0.93) with LINC00299 expression add plausibility to this result. CONCLUSION: Hypermethylation of LINC00299 in peripheral blood may constitute a useful circulating biomarker for TNBC.

MEST mediates the impact of prenatal bisphenol A exposure on long-term body weight development.

Background: Exposure to endocrine-disrupting chemicals can alter normal physiology and increase susceptibility to non-communicable diseases like obesity. Especially the prenatal and early postnatal period is highly vulnerable to adverse effects by environmental exposure, promoting developmental reprogramming by epigenetic alterations. To obtain a deeper insight into the role of prenatal bisphenol A (BPA) exposure in children's overweight development, we combine epidemiological data with experimental models and BPA-dependent DNA methylation changes. Methods: BPA concentrations were measured in maternal urine samples of the LINA mother-child-study obtained during pregnancy (n = 552), and BPA-associated changes in cord blood DNA methylation were analyzed by Illumina Infinium HumanMethylation450 BeadChip arrays (n = 472). Methylation changes were verified by targeted MassARRAY analyses, assessed for their functional translation by qPCR and correlated with children's body mass index (BMI) z scores at the age of 1 and 6 years. Further, female BALB/c mice were exposed to BPA from 1 week before mating until delivery, and weight development of their pups was monitored (n ≥ 8/group). Additionally, human adipose-derived mesenchymal stem cells were treated with BPA during the adipocyte differentiation period and assessed for exposure-related epigenetic, transcriptional and morphological changes (n = 4). Results: In prenatally BPA-exposed children two CpG sites with deviating cord blood DNA-methylation profiles were identified, among them a hypo-methylated CpG in the promoter of the obesity-associated mesoderm-specific transcript (MEST). A mediator analysis suggested that prenatal BPA exposure was connected to cord blood MEST promoter methylation and MEST expression as well as BMI z scores in early infancy. This effect could be confirmed in mice in which prenatal BPA exposure altered Mest promoter methylation and transcription with a concomitant increase in the body weight of the juvenile offspring. An experimental model of in vitro differentiated human mesenchymal stem cells also revealed an epigenetically induced MEST expression and enhanced adipogenesis following BPA exposure. Conclusions: Our study provides evidence that MEST mediates the impact of prenatal BPA exposure on long-term body weight development in offspring by triggering adipocyte differentiation.

DNA methylation-based classification of central nervous system tumours.

Accurate pathological diagnosis is crucial for optimal management of patients with cancer. For the approximately 100 known tumour types of the central nervous system, standardization of the diagnostic process has been shown to be particularly challenging-with substantial inter-observer variability in the histopathological diagnosis of many tumour types. Here we present a comprehensive approach for the DNA methylation-based classification of central nervous system tumours across all entities and age groups, and demonstrate its application in a routine diagnostic setting. We show that the availability of this method may have a substantial impact on diagnostic precision compared to standard methods, resulting in a change of diagnosis in up to 12% of prospective cases. For broader accessibility, we have designed a free online classifier tool, the use of which does not require any additional onsite data processing. Our results provide a blueprint for the generation of machine-learning-based tumour classifiers across other cancer entities, with the potential to fundamentally transform tumour pathology.

Clinical impact of KMT2C and SPRY4 expression levels in intensively treated younger adult acute myeloid leukemia patients.

OBJECTIVE: To evaluate the prognostic impact of gene expression levels (ELs) of two tumor suppressor genes, sprouty 4 (SPRY4, located on 5q) and lysine methyltransferase 2C (KMT2C, located on 7q) in correlation with clinical characteristics and genetic abnormalities assessed at initial diagnosis in acute myeloid leukemia (AML). METHOD: Gene expression levels were measured on cDNA by RT-qPCR from diagnostic bone marrow samples of 275 intensively treated adult AML patients (median age, 48 years). RESULTS: KMT2C ELs were significantly lower in abn7q/-7 (P = .001), whereas SPRY4 ELs were not associated with abn5q/-5. Higher KMT2C and SPRY4 ELs were significantly associated with lower genetic risk groups as defined by the European LeukemiaNet classification. Additionally, KMT2C ELs were lower in cytogenetically normal patients with DNMT3A (P = .01) or FLT3-ITD mutations (P = .05). KMT2C ELs were not associated with prognosis, whereas higher SPRY4 ELs showed a favorable impact on event-free (EFS, P = .01), relapse-free (RFS, P = .01) and in-trend on overall survival (P = .06) for cytogenetically abnormal patients, which was confirmed in multivariable analysis for EFS (HR, 0.84; 95%-CI, 0.73-0.97; P = .02) and RFS (HR, 0.85; 95%-CI, 0.73-0.98; P = .02). CONCLUSION: Our data indicate that KMT2C ELs are associated with specific genetic features and that SPRY4 ELs may add prognostic information.

Leukocyte Telomere Length and All-Cause, Cardiovascular Disease, and Cancer Mortality: Results From Individual-Participant-Data Meta-Analysis of 2 Large Prospective Cohort Studies.

We studied the associations of leukocyte telomere length (LTL) with all-cause, cardiovascular disease, and cancer mortality in 12,199 adults participating in 2 population-based prospective cohort studies from Europe (ESTHER) and the United States (Nurses' Health Study). Blood samples were collected in 1989-1990 (Nurses' Health Study) and 2000-2002 (ESTHER). LTL was measured by quantitative polymerase chain reaction. We calculated z scores for LTL to standardize LTL measurements across the cohorts. Cox proportional hazards regression models were used to calculate relative mortality according to continuous levels and quintiles of LTL z scores. The hazard ratios obtained from each cohort were subsequently pooled by meta-analysis. Overall, 2,882 deaths were recorded during follow-up (Nurses' Health Study, 1989-2010; ESTHER, 2000-2015). LTL was inversely associated with age in both cohorts. After adjustment for age, a significant inverse trend of LTL with all-cause mortality was observed in both cohorts. In random-effects meta-analysis, age-adjusted hazard ratios for the shortest LTL quintile compared with the longest were 1.23 (95% confidence interval (CI): 1.04, 1.46) for all-cause mortality, 1.29 (95% CI: 0.83, 2.00) for cardiovascular mortality, and 1.10 (95% CI: 0.88, 1.37) for cancer mortality. In this study population with an age range of 43-75 years, we corroborated previous evidence suggesting that LTL predicts all-cause mortality beyond its association with age.

Lower working heights decrease contraction intensity of shoulder muscles in a herringbone 30° milking parlor.

Musculoskeletal disorders have been a main concern in milkers for many years. To improve posture, a formula was developed in a previous study to calculate ergonomically optimal working heights for various milking parlor types. However, the working height recommendations based on the formula for the herringbone 30° parlor were broad. To clarify the recommendations for the optimal working height, we investigated the effect of working height on upper limb and shoulder muscle contraction intensities. We evaluated 60 milking cluster attachment procedures in a herringbone 30° milking parlor in 7 men and 9 women. Specifically, we examined the effect of working height on muscle contraction intensity of 4 arm and shoulder muscles bilaterally (flexor carpi ulnaris, biceps brachii, deltoideus anterior, and upper trapezius) by using surface electromyography. The working heights (low, medium, and high), which reflect the ratio of the subject's height to the height of the udder base, were used in the milking health formula to determine and fit individual depth of pits. Data were evaluated for each muscle and arm side in the functions holding and attaching. Statistical analysis was performed using linear mixed effects models, where muscle contraction intensity served as a target variable, whereas working height coefficient, sex, subject height, and repetition were treated as fixed effects, and repetition group nested in working height nested in subject was considered a random effect. Contraction intensities decreased with decreasing working height for the deltoideus anterior and upper trapezius, but not for the flexor carpi ulnaris or the biceps brachii muscles in both holding and attaching arm functions. We found that milking at a lower working height reduced muscle contraction intensities of the shoulder muscles. Women showed higher contraction intensities than men, whereas subject height had no effect. The study demonstrated that a lower working height decreased muscular load during milking. These lower working heights should be used within the recommendations made by the milking health formula for the herringbone 30°. Working heights could be adjusted effectively for milkers of varying body height. Future studies should therefore use the milking health formula as a tool to objectively compare and improve the accuracy of the working height coefficients.

DNA methylation-based classification and grading system for meningioma: a multicentre, retrospective analysis.

BACKGROUND: The WHO classification of brain tumours describes 15 subtypes of meningioma. Nine of these subtypes are allotted to WHO grade I, and three each to grade II and grade III. Grading is based solely on histology, with an absence of molecular markers. Although the existing classification and grading approach is of prognostic value, it harbours shortcomings such as ill-defined parameters for subtypes and grading criteria prone to arbitrary judgment. In this study, we aimed for a comprehensive characterisation of the entire molecular genetic landscape of meningioma to identify biologically and clinically relevant subgroups. METHODS: In this multicentre, retrospective analysis, we investigated genome-wide DNA methylation patterns of meningiomas from ten European academic neuro-oncology centres to identify distinct methylation classes of meningiomas. The methylation classes were further characterised by DNA copy number analysis, mutational profiling, and RNA sequencing. Methylation classes were analysed for progression-free survival outcomes by the Kaplan-Meier method. The DNA methylation-based and WHO classification schema were compared using the Brier prediction score, analysed in an independent cohort with WHO grading, progression-free survival, and disease-specific survival data available, collected at the Medical University Vienna (Vienna, Austria), assessing methylation patterns with an alternative methylation chip. FINDINGS: We retrospectively collected 497 meningiomas along with 309 samples of other extra-axial skull tumours that might histologically mimic meningioma variants. Unsupervised clustering of DNA methylation data clearly segregated all meningiomas from other skull tumours. We generated genome-wide DNA methylation profiles from all 497 meningioma samples. DNA methylation profiling distinguished six distinct clinically relevant methylation classes associated with typical mutational, cytogenetic, and gene expression patterns. Compared with WHO grading, classification by individual and combined methylation classes more accurately identifies patients at high risk of disease progression in tumours with WHO grade I histology, and patients at lower risk of recurrence among WHO grade II tumours (p=0·0096) from the Brier prediction test). We validated this finding in our independent cohort of 140 patients with meningioma. INTERPRETATION: DNA methylation-based meningioma classification captures clinically more homogenous groups and has a higher power for predicting tumour recurrence and prognosis than the WHO classification. The approach presented here is potentially very useful for stratifying meningioma patients to observation-only or adjuvant treatment groups. We consider methylation-based tumour classification highly relevant for the future diagnosis and treatment of meningioma. FUNDING: German Cancer Aid, Else Kröner-Fresenius Foundation, and DKFZ/Heidelberg Institute of Personalized Oncology/Precision Oncology Program.

Next-generation personalised medicine for high-risk paediatric cancer patients - The INFORM pilot study.

The 'Individualized Therapy for Relapsed Malignancies in Childhood' (INFORM) precision medicine study is a nationwide German program for children with high-risk relapsed/refractory malignancies, which aims to identify therapeutic targets on an individualised basis. In a pilot phase, reported here, we developed the logistical and analytical pipelines necessary for rapid and comprehensive molecular profiling in a clinical setting. Fifty-seven patients from 20 centers were prospectively recruited. Malignancies investigated included sarcomas (n = 25), brain tumours (n = 23), and others (n = 9). Whole-exome, low-coverage whole-genome, and RNA sequencing were complemented with methylation and expression microarray analyses. Alterations were assessed for potential targetability according to a customised prioritisation algorithm and subsequently discussed in an interdisciplinary molecular tumour board. Next-generation sequencing data were generated for 52 patients, with the full analysis possible in 46 of 52. Turnaround time from sample receipt until first report averaged 28 d. Twenty-six patients (50%) harbored a potentially druggable alteration with a prioritisation score of 'intermediate' or higher (level 4 of 7). Common targets included receptor tyrosine kinases, phosphoinositide 3-kinase-mammalian target of rapamycin pathway, mitogen-activated protein kinase pathway, and cell cycle control. Ten patients received a targeted therapy based on these findings, with responses observed in some previously treatment-refractory tumours. Comparative primary relapse analysis revealed substantial tumour evolution as well as one case of unsuspected secondary malignancy, highlighting the importance of re-biopsy at relapse. This study demonstrates the feasibility of comprehensive, real-time molecular profiling for high-risk paediatric cancer patients. This extended proof-of-concept, with examples of treatment consequences, expands upon previous personalised oncology endeavors, and presents a model with considerable interest and practical relevance in the burgeoning era of personalised medicine.

Body mass index and leukocyte telomere length dynamics among older adults: Results from the ESTHER cohort.

OBJECTIVE: Telomere length (TL) has been proposed as a biomarker of ageing, which might be used to identify individuals at higher risk of age-related diseases. Obesity is a well-known risk factor for several diseases. This study aims to analyse the associations of BMI with TL and the rate of TL change in older adults. METHODS: Leukocyte TL (LTL) was measured by quantitative PCR in blood samples of 3600 older adults aged 50-75 years obtained at the baseline examination of a population-based cohort study in Germany. For longitudinal analyses, measurements were repeated in blood samples obtained at 8-year follow-up from 1000 participants. Multivariate linear regression models were used to estimate associations of BMI with LTL and changes in LTL over time. RESULTS: LTL was inversely associated with age (r = -0.090, p < 0.0001). BMI and LTL associations varied according to age (p for interaction = 0.021). BMI was significantly inversely associated with LTL in those younger than 60 years (-6 basepairs per 1 kg/m(2) difference in BMI). In particular, weight gain during adulthood was inversely associated with LTL in a dose-response manner in this age group, with those having gained ≥ 30 kg having significantly shorter telomeres (-209 basepairs) than those who maintained their weight. No clear patterns were observed between any of BMI-related variables and the rate of LTL change. CONCLUSIONS: Our cross-sectional analysis supports suggestions that weight gain during adulthood and obesity may contribute to shorter telomere length below 60 years of age, but this relationship could not be shown longitudinally.

Epigenetics and Proteomics Join Transcriptomics in the Quest for Tuberculosis Biomarkers.

UNLABELLED: An estimated one-third of the world's population is currently latently infected with Mycobacterium tuberculosis. Latent M. tuberculosis infection (LTBI) progresses into active tuberculosis (TB) disease in ~5 to 10% of infected individuals. Diagnostic and prognostic biomarkers to monitor disease progression are urgently needed to ensure better care for TB patients and to decrease the spread of TB. Biomarker development is primarily based on transcriptomics. Our understanding of biology combined with evolving technical advances in high-throughput techniques led us to investigate the possibility of additional platforms (epigenetics and proteomics) in the quest to (i) understand the biology of the TB host response and (ii) search for multiplatform biosignatures in TB. We engaged in a pilot study to interrogate the DNA methylome, transcriptome, and proteome in selected monocytes and granulocytes from TB patients and healthy LTBI participants. Our study provides first insights into the levels and sources of diversity in the epigenome and proteome among TB patients and LTBI controls, despite limitations due to small sample size. Functionally the differences between the infection phenotypes (LTBI versus active TB) observed in the different platforms were congruent, thereby suggesting regulation of function not only at the transcriptional level but also by DNA methylation and microRNA. Thus, our data argue for the development of a large-scale study of the DNA methylome, with particular attention to study design in accounting for variation based on gender, age, and cell type. IMPORTANCE: DNA methylation modifies the transcriptional program of cells. We have focused on two major populations of leukocytes involved in immune response to infectious diseases, granulocytes and monocytes, both of which are professional phagocytes that engulf and kill bacteria. We have interrogated how DNA methylation, gene expression, and protein translation differ in these two cell populations between healthy individuals and patients suffering from TB. To better understand the underlying biologic mechanisms, we harnessed a statistical enrichment analysis, taking advantage of predefined and well-characterized gene sets. Not only were there clear differences on various levels between the two populations, but there were also differences between TB patients and healthy controls in the transcriptome, proteome, and, for the first time, DNA methylome in these cells. Our pilot study emphasizes the value of a large-scale study of the DNA methylome taking into account our findings.

Smoking habits and leukocyte telomere length dynamics among older adults: Results from the ESTHER cohort.

BACKGROUND & AIMS: Leukocyte telomere length (LTL) shortens with age and short LTL has been associated with increased mortality and increased risk for some age-related outcomes. This study aims to analyse the associations of smoking habits with LTL and rate of LTL change per year in older adults. METHODS: LTL was measured by quantitative PCR at baseline in 3600 older adults, who were enrolled in a population-based cohort study in Germany. For longitudinal analyses, measurements were repeated in blood samples obtained at 8-year follow-up from 1000 participants. Terminal Restriction Fragment analysis was additionally performed in a sub-sample to obtain absolute LTL in base pairs. Multivariate linear regression models were used to estimate associations of smoking habits with baseline LTL and changes in LTL over time. RESULTS: LTL was inversely associated with age (r=-0.090, p<0.0001). Women had longer LTL than men (p<0.0001). Smoking was inversely associated with LTL. On average, current smokers had 73 base pairs (BP) shorter LTL compared to never smokers. Smoking intensity and pack-years of smoking were also inversely associated with LTL, and a positive association was observed with years since smoking cessation. Slower LTL attrition rates were observed in ever smokers over 8years of follow-up. CONCLUSIONS: Our cross-sectional analysis supports suggestions that smoking might contribute to shortening of LTL but this relationship could not be shown longitudinally. The overall rather small effect sizes observed for smoking-related variables suggest that LTL reflects smoking-related health hazards only to a very limited extent.

Frailty and telomere length: cross-sectional analysis in 3537 older adults from the ESTHER cohort.

Both telomere length and frailty were observed to be associated with aging. Whether and to what extent telomere length is related to frailty is essentially unknown. In this cross-sectional analysis of baseline data of 3537 community-dwelling adults aged 50 to 75 years of a large German cohort study, we assessed the hypothesis that shorter telomere length might be a biological marker for frailty. Using whole blood DNA we examined mean telomere repeat copy to single gene copy number (T/S ratio) using quantitative PCR. Construction of a frailty index (FI) was based on a deficit accumulation approach, which quantifies frailty as ratio of the deficits present divided by the total number of deficits considered. Mean FI was determined according to age by tertiles of T/S ratio. Furthermore, we used correlation analyses stratified for gender and age groups to examine the association of the T/S ratio with frailty. Mean FI value was similar across tertiles of the T/S ratio (0.24±0.14, 0.24±0.14 and 0.23±0.14, respectively (p=0.09)), and FI and the T/S ratio were uncorrelated in gender- and age-specific analyses. In conclusion, T/S ratio and frailty were unrelated in this large sample of older adults. T/S ratio may therefore not be a meaningful biological marker for frailty.