Exploring the Relationship of Relative Telomere Length and the Epigenetic Clock in the LipidCardio Cohort

Telomere length has been accepted widely as a biomarker of aging. Recently, a novel candidate biomarker has been suggested to predict an individual’s chronological age with high accuracy: The epigenetic clock is based on the weighted DNA methylation (DNAm) fraction of a number of cytosine-phosphate-guanine sites (CpGs) selected by penalized regression analysis. Here, an established methylation-sensitive single nucleotide primer extension method was adapted, to estimate the epigenetic age of the 1005 participants of the LipidCardio Study, a patient cohort characterised by high prevalence of cardiovascular disease, based on a seven CpGs epigenetic clock. Furthermore, we measured relative leukocyte telomere length (rLTL) to assess the relationship between the established and the promising new measure of biological age. Both rLTL (0.79 ± 0.14) and DNAm age (69.67 ± 7.27 years) were available for 773 subjects (31.6% female; mean chronological age= 69.68 ± 11.01 years; mean DNAm age acceleration = −0.01 ± 7.83 years). While we detected a significant correlation between chronological age and DNAm age (n = 779, R = 0.69), we found neither evidence of an association between rLTL and the DNAm age (β = 3.00, p = 0.18) nor rLTL and the DNAm age acceleration (β = 2.76, p = 0.22) in the studied cohort, suggesting that DNAm age and rLTL measure different aspects of biological age

DNA Damage in Nijmegen Breakage Syndrome Cells Leads to PARP Hyperactivation and Increased Oxidative Stress

Nijmegen Breakage Syndrome (NBS), an autosomal recessive genetic instability syndrome, is caused by hypomorphic mutation of the NBN gene, which codes for the protein nibrin. Nibrin is an integral member of the MRE11/RAD50/NBN (MRN) complex essential for processing DNA double-strand breaks. Cardinal features of NBS are immunodeficiency and an extremely high incidence of hematological malignancies. Recent studies in conditional null mutant mice have indicated disturbances in redox homeostasis due to impaired DSB processing. Clearly this could contribute to DNA damage, chromosomal instability, and cancer occurrence. Here we show, in the complete absence of nibrin in null mutant mouse cells, high levels of reactive oxygen species several hours after exposure to a mutagen. We show further that NBS patient cells, which unlike mouse null mutant cells have a truncated nibrin protein, also have high levels of reactive oxygen after DNA damage and that this increased oxidative stress is caused by depletion of NAD+ due to hyperactivation of the strand-break sensor, Poly(ADP-ribose) polymerase. Both hyperactivation of Poly(ADP-ribose) polymerase and increased ROS levels were reversed by use of a specific Poly(ADP-ribose) polymerase inhibitor. The extremely high incidence of malignancy among NBS patients is the result of the combination of a primary DSB repair deficiency with secondary oxidative DNA damage

Sports and Exercise at Different Ages and Leukocyte Telomere Length in Later Life – Data from the Berlin Aging Study II (BASE-II)

Saßenroth D, Meyer A, Salewsky B, et al. Sports and Exercise at Different Ages and Leukocyte Telomere Length in Later Life – Data from the Berlin Aging Study II (BASE-II). PLOS ONE. 2015;10(12): e0142131.Physical activity and sports have repeatedly been reported to be associated with telomere length. We studied the association of different types of sports across different stages of life on relative leukocyte telomere length (rLTL) in advanced age. 815 participants (397 men) from the Berlin Aging Study II aged over 61 years were included in the analysis. rLTL was measured by real time PCR and physical activity was determined retrospectively by questionnaire, assessing type and duration of sports in the past as well as currently. Five separate multiple linear regression models adjusted for various control variables were performed. 67.3% of participants exercised currently, whereas 19.4% performed sports only between the age of 20 and 30. rLTL was higher in subjects who stated to exercise currently (N = 456), and in subjects who engaged in endurance (N = 138) or intensive activity sports (N = 32). Current physical activity was positively associated with rLTL in the risk factor adjusted regression model (beta = 0.26, p < 0.001) and practicing sports for a minimum of 10 years preceding the assessment had a significant effect on rLTL (beta = 0.39, p = 0.011). The highest impact was seen for intensive activity sports (beta = 0.79, p < 0.001) and physical activity since at least 42 years (beta = 0.47, p = 0.001). However, physical activity only between 20 and 30 years of age did not affect rLTL in old age when compared to no sports at all (beta = -0.16, p = 0.21). Physical activity is clearly associated with longer rLTL. The effect is seen with longer periods of physical activity (at least 10 years), with intensive sports activities having the greatest impact on rLTL. Our data suggest that regular physical activity for at least 10 years is necessary to achieve a sustained effect on rLTL

Evaluation of the role of STAP1 in Familial Hypercholesterolemia

Familial hypercholesterolemia (FH) is characterised by elevated serum levels of low-density lipoprotein cholesterol (LDL-C) and a substantial risk for cardiovascular disease. The autosomal-dominant FH is mostly caused by mutations in LDLR (low density lipoprotein receptor), APOB (apolipoprotein B), and PCSK9 (proprotein convertase subtilisin/kexin). Recently, STAP1 has been suggested as a fourth causative gene. We analyzed STAP1 in 75 hypercholesterolemic patients from Berlin, Germany, who are negative for mutations in canonical FH genes. In 10 patients with negative family history, we additionally screened for disease causing variants in LDLRAP1 (low density lipoprotein receptor adaptor protein 1), associated with autosomal-recessive hypercholesterolemia. We identified one STAP1 variant predicted to be disease causing. To evaluate association of serum lipid levels and STAP1 carrier status, we analyzed 20 individuals from a population based cohort, the Cooperative Health Research in South Tyrol (CHRIS) study, carrying rare STAP1 variants. Out of the same cohort we randomly selected 100 non-carriers as control. In the Berlin FH cohort STAP1 variants were rare. In the CHRIS cohort, we obtained no statistically significant differences between carriers and non-carriers of STAP1 variants with respect to lipid traits. Until such an association has been verified in more individuals with genetic variants in STAP1, we cannot estimate whether STAP1 generally is a causative gene for FH

High levels of ROS in <i>Nbn</i> null mutant murine fibroblasts and NBS patient cells after DNA damage.

<p>(A) FACS profiles of ROS measurements in murine cells with the indicated genotypes with or without treatment with bleomycin. Cells were stained with CM-H₂DCFDA 12 hours after treatment with bleomycin. Fluorescence intensity is proportional to ROS. The experiment was repeated six times and the same profiles were obtained. (B) Western-blot demonstration of conditional <i>Nbn</i> null mutation in murine fibroblasts. Lysates from <i>Nbn</i>^Ins-6/lox-6 fibroblasts with and without treatment with HTNC were probed on immunoblots with anti-nibrin and anti-actin antibodies. (C) Representative FACS profiles of ROS measurements in LN9 wild type and GM7166VA7 NBS patient fibroblasts with or without treatment with bleomycin. Cells were stained with CM-H₂DCFDA 12 hours after treatment with bleomycin. Fluorescence intensity is proportional to ROS. The experiment was repeated more than five times and essentially the same profiles were obtained.</p

ROS levels in NBS patient fibroblasts after DNA damage are reduced by antioxidant scavengers but PARP remains hyperactivated.

<p>(A) FACS profiles of ROS measurements in NBS-1LBI patient cells 12 hours after treatment with bleomycin and in the presence or absence of the antioxidant TROLOX. Cells were stained with CM-H₂DCFDA for ROS detection. The data shown are from one of three experiments with essentially identical results. (B) Lysates from NBS-1LBI patient cells were harvested 15 minutes after DNA damage by bleomycin in the presence the PARP inhibitor KU-0058948 or the antioxidant TROLOX as indicated. Lysates were probed on immunoblots with antibodies directed against poly(ADP-ribose) and ß-actin.</p

ROS in <i>Nbn</i> null mutant murine fibroblasts and NBS patient cells after DNA damage.

<p>Relative levels of ROS after treatment with bleomycin are given for murine and human LN9, GM166VA7 and NBS-1LBI cells with the given genotypes and after the indicated treatments. ** p = 0.0095 in the Mann-Whitney U Test (two-tailed, n¹ = 4, n² = 6); * p = 0.017 in the Mann-Whitney U Test (two-tailed, n¹ = 3, n² = 12).</p

Increased PARP activity in <i>Nbn^−/−</i> murine fibroblasts and NBS patient fibroblasts after DNA damage.

<p>Lysates from mouse (A) and human LN9 and GM166VA7 fibroblasts (B) with the given genotypes were harvested at the indicated timepoints (minutes) after a bleomycin treatment and probed on immunoblots with antibodies directed against poly(ADP-ribose) and ß-actin.</p