Laila Bendix

The load of short telomeres, estimated by a new method, Universal STELA, correlates with number of senescent cells

Short telomeres are thought to trigger senescence, most likely through a single – or a group of few – critically shortened telomeres. Such short telomeres are thought to result from a combination of gradual linear shortening resulting from the end replication problem, reflecting the division history of the cell, superimposed by a more stochastic mechanism, suddenly causing a significant shortening of a single telomere. Previously, studies that have tried to explore the role of critically shortened telomeres have been hampered by methodological problems. With the method presented here, Universal STELA, we have a tool that can directly investigate the relationship between senescence and the load of short telomeres. The method is a variant of the chromosome‐specific STELA method but has the advantage that it can demonstrate short telomeres regardless of chromosome. With Universal STELA, we find a strong correlation between the load of short telomeres and cellular senescence. Further we show that the load of short telomeres is higher in senescent cells compared to proliferating cells at the same passage, offering an explanation of premature cell senescence. This new method, Universal STELA, offers some advantages compared to existing methods and can be used to explore many of the unanswered questions in telomere biology including the role that telomeres play in cancer and aging.

Longitudinal Changes in Leukocyte Telomere Length and Mortality in Humans

BACKGROUND Leukocyte telomere length (LTL) ostensibly shortens with age and has been moderately associated with mortality. In humans, these findings have come almost solely from cross-sectional studies. Only recently has LTL shortening within individuals been analyzed in longitudinal studies. Such studies are relevant to establish LTL dynamics as biomarkers of mortality as well as to disentangle the causality of telomeres on aging. METHODS We present a large longitudinal study on LTL and human mortality, where the 10-year change of LTL is analyzed in 1,356 individuals aged 30-70 years. RESULTS We find age, smoking status, and alcohol consumption to be associated with LTL attrition and confirm a strong association with baseline LTL. The latter association might be an epiphenomenon of regression to the mean. We do not find an association of mortality with either absolute LTL or LTL attrition. Further, we show that DNA quality has an impact on TS ratios. CONCLUSIONS This study establishes that certain lifestyle factors influence LTL dynamics. However, it questions the applicability of LTL dynamics as a predictor of mortality. We suggest cautiousness when assessing actual LTL attrition due to the need for high-quality DNA and the phenomena of regression to the mean.

Leukocyte telomere length and physical ability among Danish Twins age 70

Leukocyte telomere length (LTL) shortens with age and is potentially a biomarker of human aging. We examined the relation of LTL with physical ability and cognitive function in 548 same-sex twins from the Longitudinal Study of Aging Danish Twins. LTL was measured by Southern blots of the terminal restriction fragments (TRF). Physical ability was evaluated using a self reported scale of 11 questions, while cognitive function was scored by MMSE and a cognitive composite score sensitive to age-related decline. A random intercept model revealed a positive, significant association between LTL and physical ability. For every unit increase in physical ability score, LTL increased by 0.066 kb (p = 0.01), equal to approximately three years of age-dependent LTL shortening. A matched case-co-twin design showed that the group consisting of the twins from each pair with the longer LTL also displayed better physical ability (p < 0.01). Moreover, the intra-pair difference in LTL was associated with intra-pair difference in physical ability (p < 0.01), confirming the association. However, we found no association between cognitive function and LTL. The LTL-physical ability association in the elderly provides further support to the premise that LTL is an index of somatic fitness in the narrow context of human physical health.

Stressful life events and leucocyte telomere length: Do lifestyle factors, somatic and mental health, or low grade inflammation mediate this relationship? Results from a cohort of Danish men born in 1953

Exposure to psychosocial stress is associated with increased risk of a number of somatic and mental disorders with relation to immune system functioning. We aimed to explore whether stressful events in early and recent life was associated with leucocyte telomere length (TL), which is assumed to reflect the accumulated burden of inflammation and oxidative stress occurring during the life course. We specifically aimed to address whether childhood constitutes a sensitive period and how much of the relation between stressful life events and TL is mediated through somatic and mental health, lifestyle, and markers of low-grade inflammation. A cohort of Danish men born in 1953 has been followed since birth in the Metropolit Cohort. These men underwent a health examination including blood sampling in 2010 and a subset of 324 also had a quantitative PCR-based measurement of TL. The relation between stressful life events and TL was analysed using structural equation modelling, which also provided an estimate of the proportion of the total effect mediated by somatic and mental health (cardiovascular disease, body mass and depressive mood), lifestyle factors, and low grade inflammation (C-reactive protein (CRP), interleukin (IL)-6 and IL-10). Total number of stressful events experienced during the life course was not associated with TL. In terms of sensitive periods, we found that number of stressful events in childhood was associated with shorter TL (βper number stressful events in childhood=-0.02(SE=-0.02); P=0.05). This relation was particularly strong for being placed away from home (β=-0.16; P<0.000). Thirty percent of the total effect of stressful events in childhood on TL was mediated by the included variables, with the largest proportion being mediated through depressive mood (16%) and CRP (9%). This study suggests that stressful events in childhood are associated with shorter TL in middle-aged men and that part of this relation is explained by depressive mood and low grade inflammation.

Telomere dynamics in human mesenchymal stem cells after exposure to acute oxidative stress.

A gradual shortening of telomeres due to replication can be measured using the standard telomere restriction fragments (TRF) assay and other methods by measuring the mean length of all the telomeres in a cell. In contrast, stress-induced telomere shortening, which is believed to be just as important for causing cellular senescence, cannot be measured properly using these methods. Stress-induced telomere shortening caused by, e.g. oxidative damage happens in a stochastic manner leaving just a few single telomeres critically short. It is now possible to visualize these few ultra-short telomeres due to the advantages of the newly developed Universal single telomere length assay (STELA), and we therefore believe that this method should be considered the method of choice when measuring the length of telomeres after exposure to oxidative stress. In order to test our hypothesis, cultured human mesenchymal stem cells, either primary or hTERT immortalized, were exposed to sub-lethal doses of hydrogen peroxide, and the short term effect on telomere dynamics was monitored by Universal STELA and TRF measurements. Both telomere measures were then correlated with the percentage of senescent cells estimated by senescence-associated β-galactosidase staining. The exposure to acute oxidative stress resulted in an increased number of ultra-short telomeres, which correlated strongly with the percentage of senescent cells, whereas a correlation between mean telomere length and the percentage of senescent cells was absent. Based on the findings in the present study, it seems reasonable to conclude that Universal STELA is superior to TRF in detecting telomere damage caused by exposure to oxidative stress. The choice of method should therefore be considered carefully in studies examining stress-related telomere shortening as well as in the emerging field of lifestyle studies involving telomere length measurements.

Associations of subjective vitality with DNA damage, cardiovascular risk factors and physical performance

To examine associations of DNA damage, cardiovascular risk factors and physical performance with vitality, in middle‐aged men. We also sought to elucidate underlying factors of physical performance by comparing physical performance parameters to DNA damage parameters and cardiovascular risk factors.

The relationship between ultra-short telomeres, aging of articular cartilage and the development of human hip osteoarthritis

INTRODUCTION Ultra-short telomeres caused by stress-induced telomere shortening are suggested to induce chondrocyte senescence in human osteoarthritic knees. Here we have further investigated the role of ultra-short telomeres in the development of osteoarthritis (OA) and in aging of articular cartilage in human hips. MATERIALS AND METHODS Cartilage was obtained from four different distances of the central weight-bearing area in human femoral heads (14 OA and 9 non-OA). Samples were split into three: one for quantification of ultra-short single telomeres by Universal STELA and mean telomere length measurement by Q-PCR; one for histological grading of OA, and one for immunohistochemical staining. RESULTS Load of ultra-short telomeres increased closer to the central weight-bearing area and correlated with cartilage degradation in both OA and non-OA samples. Mean telomere length decreased with decreasing distance to the central weight-bearing area, however, unexpectedly increased in the most central zone. This increase was associated with immunohistochemical findings of cells expressing markers characteristic of progenitor-like cells. CONCLUSION These findings suggest a role of short telomeres in the development of OA and in aging of articular cartilage. Furthermore, progenitor-like cells with long telomeres may be recruited to the most damaged areas of the cartilage.

Reduced telomere length is not associated with early signs of vascular aging in young men born after intrauterine growth restriction: a paradox?

Objective: The mechanisms that increase cardiovascular risk in individuals born small for gestational age (SGA) are not well understood. Telomere shortening has been suggested to be a predictor of disease onset. Our aim was to determine whether impaired intrauterine growth is associated with early signs of vascular aging and whether telomere length could be a biomarker of this pathway. Methods: One hundred and fourteen healthy young men born SGA or after normal pregnancy [appropriate for gestational age (AGA)] were enrolled. Patient data were gathered from questionnaires and clinical exams, including blood pressure (BP) measurement routine laboratory analyses, and carotid intima–media thickness (cIMT). Leukocyte telomere length (LTL) was assessed by quantitative PCR. Birth data were obtained from medical records. Results: The SGA group had significantly higher pulse pressure and cIMT, and a trend to increased SBP and heart rate in comparison to the AGA group. Interestingly, SGA men exhibited a 42% longer LTL than the AGA group. LTL was inversely associated with age, BMI, BP and birth parameters. In multiple regression analysis, BMI was the key determinant of SBP and cIMT. Conclusion: Young men born SGA show early signs of vascular aging. Unexpectedly, in our cohort, the SGA group had longer telomeres than the normal controls. Although longer telomeres are predictive of better health in the future, our findings could indicate a faster telomere attrition rate and probable early onset of cardiovascular risk in SGA participants. Follow-up of this cohort will clarify hypothesis and validate telomere dynamics as indicators of future health risks.

Leucocyte Telomere Length and Risk of Cardiovascular Disease in a Cohort of 1,397 Danish Men and Women

Objectives: Short leucocyte telomere length (LTL) might be a risk factor for cardiovascular diseases (CVD). The present study examines the relation between LTL and incident fatal or non-fatal CVD, ischaemic heart disease (IHD) and stroke in a Danish cohort followed for 29 years. Methods: In total, 1,397 men and women who participated in health examinations with blood sampling in 1981-1984 were followed for CVD outcomes until the end of 2012 by linkage to national registers. Cox proportional hazard regression models were used to analyse the relation between LTL and CVD adjusting for potential confounding CVD risk factors. Results: During the follow-up, 603 participants experienced an incident fatal or non-fatal CVD. The survival analysis showed that baseline LTL was not associated with CVD outcomes. In the subanalysis with IHD as outcome, those with middle and short LTL had an increased hazard rate ratio of 1.97 (95% CI 1.31-2.93) and 1.55 (95% CI 1.02-2.35), respectively, which was attenuated when confounding factors were adjusted for. For stroke, the pattern of associations was similar but less precisely estimated. Conclusions: In this study short, LTL was not associated with an increased risk of CVD, but modestly associated with an increased risk of IHD.

Association between leukocyte telomere length and bone mineral density in women 25-93 years of age

Leukocyte telomere length (LTL) and bone mineral density (BMD) are associated with health and mortality. Because osteoporosis is an age-related condition and LTL is considered to be a biomarker of aging, we hypothesized that shorter LTL could predict lower BMD. The aim of our study was to assess whether there is an association of LTL with BMD and to determine whether this possible association is independent of age. The BMDs of the lumbar spine (LS), femoral neck (FN) and total hip (TH) were evaluated in 460 women using DXA. LTL was analyzed using quantitative polymerase chain reaction. The women completed a health and lifestyle questionnaire. The associations were estimated by regression models that considered age, body mass index (BMI), menopause, physical activity, alcohol consumption and smoking habits. We found a statistically significant unadjusted association between LTL and age (estimate and 95% confidence interval (CI): -0.003 (-0.005; -0.002)); and between BMI adjusted age and logarithmic transformed BMD. Estimates and 95% CI were as follows: LS: -0.13 (-0.26; -0.01); right TH: -0.44 (-0.53; -0.34); left TH: -0.38 (-0.48; -0.28); right FN: -0.57 (-0.67; -0.46) and left FN: -0.51 (-0.62; -0.40). There were no statistically significant associations between BMD and LTL (both logarithmically transformed) with or without age adjustments. The age-adjusted estimates and CI were as follows: LS: -0.10 (-0.71; 0.52); right TH: -0.13 (-0.66; 0.41); left TH: -0.13 (-0.67; 0.42); right FN: -0.03 (-0.58; 0.52) and left FN: 0.09 (-0.47; 0.66). In conclusion, we found no statistically significant associations between BMD and LTL, although the estimates of the crude associations were all positive, indicating hypothesis consistency; that shorter LTL predict lower BMD values. This positive association was no longer apparent after adjusting for age. As expected, age was statistically significantly associated with both telomere length and BMI adjusted BMD.