Average Telomere Length Research Articles

Leukocyte Telomere Length in Patients with Multiple Sclerosis and Its Association with Clinical Phenotypes

Aging is a significant factor influencing the course of multiple sclerosis (MS). Accelerated telomere attrition is an indicator of premature biological aging and a potential contributor to various chronic diseases, including neurological disorders. However, there is currently a lack of studies focusing on telomere lengths in patients with MS. We measured the average leukocyte telomere length (LTL) in biobanked DNA samples of 40 relapsing-remitting MS patients (RRMS), 20 primary progressive MS patients (PPMS), and 60 healthy controls using a multiplex quantitative polymerase chain reaction method. Changes in LTL over a period of >10 years were evaluated in a subset of 10 patients. Association analyses of baseline LTL with the long-term clinical profiles of the patients were performed using inferential statistical tests and regression models adjusted for age and sex. The cross-sectional analysis revealed that the RRMS group was characterized by a significantly shorter relative LTL, on average, as compared to the PPMS group and controls. Shorter telomeres at baseline were also associated with a higher conversion rate from RRMS to secondary progressive MS (SPMS) in the 10-year follow-up. The LTL decrease over time was similar in RRMS patients and PPMS patients in the longitudinal analysis. Our data suggest a possible contributory role of accelerated telomere shortening in the pathobiology of MS. The interplay between disease-related immune system alterations, immunosenescence, and telomere dynamics deserves further investigation. New insights into the mechanisms of disease might be obtained, e.g., by exploring the distribution of telomere lengths in specific blood cell populations.

Identification and characterization of novel ACD variants: modulation of TPP1 protein level offsets the impact of germline loss-of-function variants on telomere length.

Telomere biology disorders, largely characterized by telomere lengths below the first centile for age, are caused by variants in genes associated with telomere replication, structure, or function. One of these genes, ACD, which encodes the shelterin protein TPP1, is associated with both autosomal dominantly and autosomal recessively inherited telomere biology disorders. TPP1 recruits telomerase to telomeres and stimulates telomerase processivity. Several studies probing the effect of various synthetic or patient-derived variants have mapped specific residues and regions of TPP1 that are important for interaction with TERT, the catalytic component of telomerase. However, these studies have come to differing conclusions regarding ACD haploinsufficiency. Here, we report a proband with compound heterozygous novel variants in ACD (NM_001082486.1)—c.505_507delGAG, p.(Glu169del); and c.619delG, p.(Asp207Thrfs*22)—and a second proband with a heterozygous chromosomal deletion encompassing ACD: arr[hg19] 16q22.1(67,628,846-67,813,408)x1. Clinical data, including symptoms and telomere length within the pedigrees, suggested that loss of one ACD allele was insufficient to induce telomere shortening or confer clinical features. Further analyses of lymphoblastoid cell lines showed decreased nascent ACD RNA and steady-state mRNA, but normal TPP1 protein levels, in cells containing heterozygous ACD c.619delG, p.(Asp207Thrfs*22), or the ACD-encompassing chromosomal deletion compared to controls. Based on our results, we conclude that cells are able to compensate for loss of one ACD allele by activating a mechanism to maintain TPP1 protein levels, thus maintaining normal telomere length.

Implication of therapeutic outcomes associated with molecular characterization of paediatric aplastic anaemia

ObjectivesSevere aplastic anemia is characterized by a hypocellular bone marrow and peripheral cytopenia. Mesenchymal stem cells (MSCs) play a crucial role in haematopoietic stem cells (HSCs) development and the development of microenvironment suitable for hematopoiesis. Molecular characterization of telomere maintenance pathway and gene expression profiling of MSCs can be important for the therapeutic interventions among paediatric aplastic anaemia patients. MethodsThe study involved paediatric aplastic anaemia patients (n = 10) and age matched paediatric healthy donors (n = 8). Peripheral blood samples were collected from the individuals. Average leucocyte telomere length and gene expression of the telomere maintenance genes were determined by quantitative real time PCR. Microarray based gene expression profiles (GSE33812) of MSCs for five paediatric aplastic anaemia patients were analyzed compared to five healthy controls and the data was downloaded from the GEO database. ResultsThe telomere length was significantly shorter among paediatric AA patients compared to age matched healthy donors. Interestingly, one subgroup (n = 2) of paediatric AA patients has moderate telomere length comparable to age matched healthy donors. Based on the gene expression analysis of telomere maintenance pathway, TERF2 was significantly downregulated among paediatric patients with shorter telomere length but not among paediatric patients with moderate telomere length. Gene expression profiling of MSCs revealed three differentially expressed genes (GAS2L3, MK167 and TMSB15A) among the patients and was associated with therapeutic outcome. ConclusionTelomere length estimation and gene expression patterns of the MSCs and telomere length maintenance pathway may serve as a potential biomarker and could be associated with therapeutic choice of paediatric aplastic anaemia patients.

Systematic Review of Studies on Telomere Length in Patients with Multiple Sclerosis.

Telomeres are protective cap structures at the end of chromosomes that are essential for maintaining genomic stability. Accelerated telomere shortening is related to premature cellular senescence. Shortened telomere lengths (TL) have been implicated in the pathogenesis of various chronic immune-mediated and neurological diseases. We aimed to systematically review the current literature on the association of TL as a measure of biological age and multiple sclerosis (MS). A comprehensive literature search was conducted to identify original studies that presented data on TL in samples from persons with MS. Quantitative and qualitative information was extracted from the articles to summarize and compare the studies. A total of 51 articles were screened, and 7 of them were included in this review. In 6 studies, average TL were analyzed in peripheral blood cells, whereas in one study, bone marrow-derived cells were used. Four of the studies reported significantly shorter leukocyte TL in at least one MS subtype in comparison to healthy controls (p=0.003 in meta-analysis). Shorter telomeres in patients with MS were found to be associated, independently of age, with greater disability, lower brain volume, increased relapse rate and more rapid conversion from relapsing to progressive MS. However, it remains unclear how telomere attrition in MS may be linked to oxidative stress, inflammation and age-related disease processes. Despite few studies in this field, there is substantial evidence on the association of TL and MS. Variability in TL appears to reflect heterogeneity in clinical presentation and course. Further investigations in large and well-characterized cohorts are warranted. More detailed studies on TL of individual chromosomes in specific cell types may help to gain new insights into the pathomechanisms of MS.

Study of the possible relations between vitamin D, telomere length, and high-sensitivity C-reactive protein in older people

Background Ageing is a multifactorial process, and telomere shortening is one of the possible mechanisms of cellular ageing. Many factors can affect telomere length (TL). Our study investigated the effect of vitamin D and high-sensitivity C-reactive protein (hs-CRP), as one of the inflammatory markers, on TL in older people. Patients and methods This was a cross-sectional study on 100 older people who did not have malignancy, autoimmune diseases, or renal disease and did not take vitamin D supplementation. Measurement of TL was done using real-time PCR, and for each sample, calculation of the T/S ratio was done. Telomere (T) signals in experimental DNA samples and single-copy gene (S) signals were measured in separate wells, in comparison with a reference DNA, to yield relative T/S ratios that are proportional to average TL. Results The results showed that vitamin D was significantly lower in females than males, and there was a significant positive correlation between vitamin D and the level of education of our participants. We found a highly significant negative correlation between vitamin D and hs-CRP. However, there was no significant correlation between T/S ratio and either vitamin D or hs-CRP. Conclusion In conclusion, vitamin D has an anti-inflammatory effect that decreased the hs-CRP level in our study. However, factors other than vitamin D and hs-CRP may be responsible for shortening telomere in elderly patients.

Single-molecule telomere length characterization by optical mapping in nano-channel array: Perspective and review on telomere length measurement

In humans, the telomere consists of tandem 5′TTAGGG3′ DNA repeats on both ends of all 46 chromosomes. Telomere shortening has been linked to aging and age-related diseases. Similarly, telomere length changes have been associated with chemical exposure, molecular-level DNA damage, and tumor development. Telomere elongation has been associated to tumor development, caused due to chemical exposure and molecular-level DNA damage. The methods used to study these effects mostly rely on average telomere length as a biomarker. The mechanisms regulating subtelomere-specific and haplotype-specific telomere lengths in humans remain understudied and poorly understood, primarily because of technical limitations in obtaining these data for all chromosomes. Recent studies have shown that it is the short telomeres that are crucial in preserving chromosome stability. The identity and frequency of specific critically short telomeres potentially is a useful biomarker for studying aging, age-related diseases, and cancer. Here, we will briefly review the role of telomere length, its measurement, and our recent single-molecule telomere length measurement assay. With this assay, one can measure individual telomere lengths as well as identify their physically linked subtelomeric DNA. This assay can also positively detect telomere loss, characterize novel subtelomeric variants, haplotypes, and previously uncharacterized recombined subtelomeres. We will also discuss its applications in aging cells and cancer cells, highlighting the utility of the single molecule telomere length assay.

Довжина теломер у хворих на цукровий діабет 2-го типу та її зв’язок із метаболічним профілем

To study leukocyte telomere length and its relationship with metabolic profile 35 patients with metabolic syndrome and type 2 diabetes mellitus (T2DM) and 21 healthy people of middle age (35-59 years) were examined. The anthropometric characteristics of obesity, indicators of lipid and glucose metabolism, alanin aminotransferase (ALT) and high sensitive C-reactive protein levels were studied. The relative average telomere length was determined by the method of monochrome multiplex quantitative real time polymerase chain reaction. Patients with T2DM had higher BMI, waist circumference, higher high sensitive C-reactive protein, ALT and glucose levels and a worse lipid profile (p <0.05). At the same time, the median telomere length did not differ between groups. Nevertheless, in the T2DM group the telomere length inversely correlated with body weight (r = –0.35; p < 0.05), BMI (r = –0.36; p < 0.05), waist circumference (r = –0.34; p < 0.05) and ALT level (r = –0,44; p<0,05) in contrast to healthy subjects. No relationship was found between the telomere length and the level of fasting glycemia, as well as the age of the participants of both groups. Thus, in T2DM patients increase in BMI, waist circumference and ALT level were associated with a shorter leukocyte telomere length. Despite the worse metabolic profile, the telomere length in middle-aged T2DM patients did not differ from that in the control group. This indicates that the leukocyte telomere length is influenced not only by the presence of T2DM and the metabolic profile indicators, but, obviously, by other factors as well.

Photobiomodulation with 590 nm Wavelength Delays the Telomere Shortening and Replicative Senescence of Human Dermal Fibroblasts In Vitro.

Background: Cellular senescence is one of the major factors contributing to the aging process. Photobiomodulation (PBM) is known to trigger an array of cellular responses, but there are no data on how it affects the process of cellular senescence. In this study, we analyze the effect of PBM on the cellular senescence and telomere dynamics. Methods: Human dermal fibroblasts were irradiated by a panel of light-emitting diodes with 590 nm and dose 30 J/cm2 accumulated over 1200 sec repeated in 4-day cycle within 40 days. After the last cycle of PBM treatment, the difference in number of senescent cells between PBM treated groups end nontreated control groups was measured by senescent sensitive β-galactosidase assay, and the difference in average telomere length between the experimental end control groups was analyzed using relative human telomere length quantitative Polymerase Chain Reaction (qPCR) assay. Results: After 10 cycles of irradiation, the percentage of senescent cells in PBM-treated cultures was 19.7% ± 4.5%, p < 0.05 smaller than the percentage of senescent cells in the control group, and their relative telomere length was 1.19 ± 0.09-fold, p < 0.05 greater than nontreated controls. Conclusions: Our study demonstrates for the first time that PBM with appropriate parameters can delay the attrition of the telomeres and the entry of cells into senescence, suggesting a potential involvement of telomerase reactivation. A hypothetical mechanism for this light-induced antiaging effect is discussed.

Replicative senescence and arteriosclerosis after kidney transplantation.

Replicative senescence is associated with telomere shortening. In native kidneys, obtained prior to transplantation, we recently described and validated a significant association between shorter intrarenal telomere length and renal arteriosclerosis. After renal transplantation, animal experiments suggested that ischaemia-reperfusion injury, acute rejection episodes and cytomegalovirus disease associate with accelerated renal allograft senescence. The association between post-transplant events and replicative senescence has not yet been evaluated in a human setting. In a cohort of 134 kidney allograft recipients, we performed protocol-specified renal allograft biopsies at 3 months, 1 year, 2 years and 5 years after transplantation (n = 579 biopsies). We used quantitative real-time polymerase chain reaction to measure intrarenal relative average telomere length (T/S ratio). The association between donor and recipient demographic factors, post-transplant clinical/histological events, renal allograft histological evolution by 5 years post-transplant and intrarenal telomere length at 5 years after transplantation was studied using multiple regression models. At 5 years after transplantation, shorter intrarenal telomere length was associated with male donor gender, older donor age, donor history of hypertension and donor cardiovascular risk, which confirms the associations observed in native kidneys. Recipient characteristics and post-transplant events like delayed graft function, acute rejection episodes, presence of donor-specific antibodies, cytomegalovirus disease and immunosuppressive regimen did not associate with alterations of intrarenal telomere length at 5 years. Independent of donor age and donor cardiovascular risk, intrarenal arteriosclerosis in protocol biopsies obtained at 5 years after transplantation and progressive arteriosclerosis over time after transplantation associated with shorter telomere length, while this was not the case for other histological lesions. Moreover, telomere attrition augments the association between older donor age and the presence of severe arteriosclerosis. In the group with the oldest donor age and shortest telomere length, there was significantly more severe arteriosclerosis (43%) in protocol biopsies at 5 years after transplantation, compared with other combinations (13-28%) (P = 0.001). Intrarenal arteriosclerosis at 5 years after transplantation did not associate with post-transplant clinical events. We demonstrate that intrarenal telomere length at 5 years after transplantation, as a marker for replicative senescence, associates with renal arteriosclerosis and reflects kidney donor characteristics, but not post-transplant events.

Telomere biology disorder prevalence and phenotypes in adults with familial hematologic and/or pulmonary presentations

AbstractTelomere biology disorders (TBDs) present heterogeneously, ranging from infantile bone marrow failure associated with very short telomeres to adult-onset interstitial lung disease (ILD) with normal telomere length. Yield of genetic testing and phenotypic spectra for TBDs caused by the expanding list of telomere genes in adults remain understudied. Thus, we screened adults aged ≥18 years with a personal and/or family history clustering hematologic disorders and/or ILD enrolled on The University of Chicago Inherited Hematologic Disorders Registry for causative variants in 13 TBD genes. Sixteen (10%) of 153 probands carried causative variants distributed among TERT (n = 6), TERC (n = 4), PARN (n = 5), or RTEL1 (n = 1), of which 19% were copy number variants. The highest yield (9 of 22 [41%]) was in families with mixed hematologic and ILD presentations, suggesting that ILD in hematology populations and hematologic abnormalities in ILD populations warrant TBD genetic testing. Four (3%) of 117 familial hematologic disorder families without ILD carried TBD variants, making TBD second to only DDX41 in frequency for genetic diagnoses in this population. Phenotypes of 17 carriers with heterozygous PARN variants included 4 (24%) with hematologic abnormalities, 67% with lymphocyte telomere lengths measured by flow cytometry and fluorescence in situ hybridization at or above the 10th percentile, and a high penetrance for ILD. Alternative etiologies for cytopenias and/or ILD such as autoimmune features were noted in multiple TBD families, emphasizing the need to maintain clinical suspicion for a TBD despite the presence of alternative explanations.

Telomere attrition and dysfunction: a potential trigger of the progeroid phenotype in nijmegen breakage syndrome.

Background: Nibrin, as part of the NBN/MRE11/RAD50 complex, is mutated in Nijmegen breakage syndrome (NBS), which leads to impaired DNA damage response and lymphoid malignancy.Results: Telomere length (TL) was markedly reduced in homozygous patients (and comparably so in all chromosomes) by ~40% (qPCR) and was slightly reduced in NBS heterozygotes older than 30 years (~25% in qPCR), in accordance with the respective cancer rates. Humanized cancer-free NBS mice had normal TL. Telomere elongation was inducible by telomerase and/or alternative telomere lengthening but was associated with abnormal expression of telomeric genes involved in aging and/or cell growth. Lymphoblastoid cells from NBS patients with long survival times (>12 years) displayed the shortest telomeres and low caspase 7 activity.Conclusions: NBS is a secondary telomeropathy. The two-edged sword of telomere attrition enhances the cancer-prone situation in NBS but can also lead to a relatively stable cellular phenotype in tumor survivors. Results suggest a modular model for progeroid syndromes with abnormal expression of telomeric genes as a molecular basis.Methods: We studied TL and function in 38 homozygous individuals, 27 heterozygotes, one homozygous fetus, six NBS lymphoblastoid cell lines, and humanized NBS mice, all with the same founder NBN mutation: c.657_661del5.

144 Increased normal tissue telomere length is associated with decreased survival in melanoma patients

Normal tissue telomere length (nTL) varies within the population, affected by inherited genetics and environmental risk factors. Previous work has investigated whether telomere length interplays with overall cancer survival. We aim to determine whether nTL was predictive of melanoma patient survival. Both normal and tumor tissue telomere lengths from The Cancer Genome Atlas (TCGA) skin melanoma dataset, along with other cancers, were obtained from supplementary data of the Barthel et al. study (Nat Genet 49, 349–357 (2017)), which were estimated from whole genome/whole exome sequencing data. Univariate and multivariate Cox proportional hazard regression models were applied to identify variables predicting survival, adjusting for age, gender and stage. nTL from individuals with melanoma did not demonstrate a normal distribution. The majority of patients (56%) had short nTL, defined as lower than one-third of the range, while only 8% of patients demonstrated nTL in the intermediate range. Analyzed as a continuous variable, increased nTL was negatively correlated with both overall (OS) and progression free survival (PFS) for melanoma patients (p=0.005, p=0.042), even after adjusting for age, gender and stage. In contrast, tumor telomere length did not demonstrate an association with survival. When stratified into groups, those with longer than median nTL showed worse OS and PFS compared to those with shorter nTL (p<0.001, p=0.007). We then expanded this analysis to 30 other cancers in TCGA. When stratified into groups, longer nTL correlated with worse OS within bladder and colorectal cancers. However, unlike melanoma, no other cancer demonstrated a significant correlation between nTL and OS when nTL was treated as a continuous variable. In conclusion, normal tissue telomere length may represent a prognostic marker for melanoma survival.

381 - Comparison of hair cortisol concentrations between depressed patients, suicide completers and control subjects: A predictive biochemical indicator for suicide risk in depression?

A model of telomere dynamics is proposed and examined. Our model, which extends a previously introduced model that incorporates stem cells as progenitors of new cells, imposes the Hayflick limit, the maximum number of cell divisions that are possible. This new model leads to cell populations for which the average telomere length is not necessarily a monotonically decreasing function of time, in contrast to previously published models. We provide a phase diagram indicating where such results would be expected via the introduction of scaled populations, rate constants and time. The application of this model to available leukocyte baboon data is discussed.

ORCA/LRWD1 Regulates Homologous Recombination at ALT-Telomeres by Modulating Heterochromatin Organization.

SummaryTelomeres are maintained by telomerase or in a subset of cancer cells by a homologous recombination (HR)-based mechanism, Alternative Lengthening of Telomeres (ALT). The mechanisms regulating telomere-homeostasis in ALT cells remain unclear. We report that a replication initiator protein, Origin Recognition Complex-Associated (ORCA/LRWD1), by localizing at the ALT-telomeres, modulates HR activity. ORCA's localization to the ALT-telomeres is facilitated by its interaction to SUMOylated shelterin components. The loss of ORCA in ALT-positive cells elevates the levels of two mediators of HR, RPA and RAD51, and consistent with this, we observe increased ALT-associated promyelocytic leukemia body formation and telomere sister chromatid exchange. ORCA binds to RPA and modulates the association of RPA to telomeres. Finally, the loss of ORCA causes global chromatin decondensation, including at the telomeres. Our results demonstrate that ORCA acts as an inhibitor of HR by modulating RPA binding to ssDNA and inducing chromatin compaction.

Telomere length alterations and ATRX/DAXX loss in pituitary adenomas.

Telomeres are nucleoprotein complexes located at the termini of eukaryotic chromosomes that prevent exonucleolytic degradation and end-to-end chromosomal fusions. Cancers often have critically shortened, dysfunctional telomeres contributing to genomic instability. Telomere shortening has been reported in a wide range of precancerous lesions and invasive carcinomas. However, the role of telomere alterations, including the presence of alternative lengthening of telomeres (ALT), has not been studied in pituitary adenomas. Telomere length and the presence of ALT were assessed directly at the single cell level using a telomere-specific fluorescence in situ hybridization assay in tissue microarrays. Tumors were characterized as either ALT-positive or having short, normal, or long telomere lengths and then these categories were compared with clinicopathological characteristics. ATRX and DAXX expression was studied through immunohistochemistry. We characterized a discovery set of 106 pituitary adenomas including both functional and nonfunctional subsets (88 primary, 18 recurrent). Telomere lengths were estimated and we observed 64 (59.4%) cases with short, 39 (36.8%) cases with normal, and 0 (0%) cases with long telomeres. We did not observe significant differences in the clinicopathological characteristics of the group with abnormally shortened telomeres compared to the group with normal telomeres. However, three pituitary adenomas were identified as ALT-positive of which two were recurrent tumors. Two of these three ALT-positive cases had alterations in either of the chromatin remodeling proteins, ATRX and DAXX, which are routinely altered in other ALT-positive tumor subtypes. In a second cohort of 32 recurrent pituitary adenomas from 22 patients, we found that the tumors from 36% of patients (n = 8) were ALT-positive. This study demonstrates that short telomere lengths are prevalent in pituitary adenomas and that ALT-positive pituitary adenomas are enriched in recurrent disease.

Antioxidant supplementation slows telomere shortening in free-living white stork chicks.

Telomere length (TL) and shortening is increasingly shown to predict variation in survival and lifespan, raising the question of what causes variation in these traits. Oxidative stress is well known to accelerate telomere attrition in vitro, but its importance in vivo is largely hypothetical. We tested this hypothesis experimentally by supplementing white stork (Ciconia ciconia) chicks with antioxidants. Individuals received either a control treatment, or a supply of tocopherol (vitamin E) and selenium, which both have antioxidant properties. The antioxidant treatment increased the concentration of tocopherol for up to two weeks after treatment but did not affect growth. Using the telomere restriction fragment technique, we evaluated erythrocyte TL and its dynamics. Telomeres shortened significantly over the 21 days between the baseline and final sample, independent of sex, mass, size and hatching order. The antioxidant treatment significantly mitigated shortening rate of average TL (-31% in shorter telomeres; percentiles 10th, 20th and 30th). Thus, our results support the hypothesis that oxidative stress shortens telomeres in vivo.

From organ to cell: Multi-level telomere length assessment in patients with idiopathic pulmonary fibrosis.

RationaleA subset of patients with idiopathic pulmonary fibrosis (IPF) contains short leukocyte telomeres or telomere related mutations. We previously showed that alveolar type 2 cells have short telomeres in fibrotic lesions. Our objectives were to better understand how telomere shortening associates with fibrosis in IPF lung and identify a subset of patients with telomere-related disease.MethodsAverage telomere length was determined in multiple organs, basal and apical lung, and diagnostic and end-stage fibrotic lung biopsies. Alveolar type 2 cells telomere length was determined in different areas of IPF lungs.ResultsIn IPF but not in controls, telomere length in lung was shorter than in other organs, providing rationale to focus on telomere length in lung. Telomere length did not correlate with age and no difference in telomere length was found between diagnostic and explant lung or between basal and apical lung, irrespective of the presence of a radiological apicobasal gradient or fibrosis. Fifteen out of 28 IPF patients had average lung telomere length in the range of patients with a telomerase (TERT) mutation, and formed the IPFshort group. Only in this IPFshort and TERT group telomeres of alveolar type 2 cells were extremely short in fibrotic areas. Additionally, whole exome sequencing of IPF patients revealed two genetic variations in RTEL1 and one in PARN in the IPFshort group.ConclusionsAverage lung tissue telomere shortening does not associated with fibrotic patterns in IPF, however, approximately half of IPF patients show excessive lung telomere shortening that is associated with pulmonary fibrosis driven by telomere attrition.

PSXIII-13 Comparison of telomere length in age-matched primiparous and multiparous Brahman cows

Abstract Telomeres are comprised of G-rich nucleotide sequences (5’-TTAGGG-3’) at the chromosome termini that are responsible for protecting chromosomes; however, attrition of these sequences has been observed in conditions of physiological and psychological stress. The purpose of this study was to compare telomere length (TL) in 4-yr-old Brahman cows grouped by first parity (n = 8) and second parity (n = 11). Cows were bled by jugular venipuncture, weighed, and had BCS recorded d+28 prior to calving and d-7 and d-28 post calving. Cows were observed for duration of labor (Tlabor) and calving ease (CE) at the time of parturition. Calf birth weight (CBW) and gender (CG) were recorded. Peripheral leukocytes were isolated, complete blood counts (CBC) were recorded, and genomic DNA was extracted utilizing silicone membrane spin column kits. The relative quantity of telomere products, which is proportional to the average TL, was determined by multiplex quantitative PCR analysis using the ratio of bovine telomere and β-globulin DNA. An absolute standard of bovine telomere (1012–107 dilution series) and β-globulin (109-104 dilution series) genes was utilized to produce relative copy number. All samples were run in triplicate and samples were included if triplicate Cq difference was less than 0.25 cycles. Parity was the fixed effect of interest and random effects included sire and day repeated with cow as the subject. No differences in CBC were seen. Tlabor, CE, CG, and CBW did not impact TL (P &amp;gt; 0.1). A trend was observed for day-parity interaction (P = 0.0918). TL between parity differed most on d-28 (P = 0.1046; parity one 127292 ± 6483; parity two 111045 ± 5376). The stress of parturition and raising the first calf of a cow’s life may be responsible for slight attenuation in TL.

This Is a Title in Title Case: Prediction of CML Evolution By Telomere Length Analysis at the Single-Chromosome Level

Telomeresplay a vital role in DNA repair activities and protecting chromosomes from degradation[1]. Telomeres are shortened during each cell division because of the end replication problem. After several cell division, telomeres are shortened to a critical length (&lt; 3 kb), and eventually lead to overall genomic instability and triggering the DNA damage response[2-3], which is related to the cancerization of numerous cancers. The commonly used method to estimate telomere length isterminal restriction fragment (TRF) basedon southern blot, which requires thousands of cells and provides only a crude estimate of the average telomere length of all cells analyzed. However, it is believed that the frequency of critically short telomeres, rather than the mean telomere length, is a crucial factor for telomere dysfunction. Therefore, analysis of telomere length at the single-chromosome level is necessary to determine the frequency of critically short telomeres. Here, we describe the development of a high-throughput method for telomere length analysis at the single-chromosome level by using a laboratory-built high-sensitivity flow cytometer (HSFCM)[4] combined with targeted fluorescent peptide nucleic acid (PNA) probes. The unambiguous detection of the telomere signalsfrom a single chromosome was achieved via HSFCM analysis. The fluorescence intensity of single chromosome was converted to the probe number by a calibration curve, and was further transformed to the base pair number of the telomere. Five representative cell lines were analyzed to compare their telomere length and the ratio of critically short telomeres at the single-chromosome level. The potential of using frequency of short telomere for disease treatment monitoring is examined by analyzing the telomere length in lymphocyte of leukemia patients. The abundance of short telomeres was compared between healthy donors and patients with chronic myeloid leukemia (CML) to see whether it can be used to predict the efficacy of therapeutics. Moreover, the quantity of short telomeres was compared among patients with acute leukemia, patients in different phases of CML and healthy donors to see whether it can be used as a marker for disease progression prediction in CML. Reference [1] Blackburn E. H., Epel E. S., Lin J., Science,2015, 350, 1193-1198. [2]Collado M., Blasco M. A., Serrano M., Cell, 2007, 130, 223-233. [3] Deng Y., Chan S. S., Chang S., Nat. Rev. Cancer, 2008, 8, 450-458. [4] Yang LL, Zhu SB, Hang W, Wu LN, Yan XM, Anal. Chem., 2009, 81, 2555-2563. Figure 1 Disclosures No relevant conflicts of interest to declare.

Predicted Leukocyte Telomere Length and Risk of Myeloid Neoplasms

Background Maintenance of telomere length has long been established to play a role in the biology of cancer, and several lines of evidence suggest that it may be especially important in myeloid malignancy. Measuring leukocyte telomere length (LTL) directly in case control studies is problematic because telomere length is likely to be influenced by the disease process. Recent studies have led to the identification of a robust genetic predictor of LTL, which we have used to compare differences in predicted telomere length in case control studies of myeloid leukemia and myelodysplastic syndromes (MDS) using Mendelian randomization (MR). MR is a method used to measure the association between a trait (e.g. telomere length) and an outcome (e.g. myeloid malignancy) that can't be measured directly by utilizing genetic variants known to be associated with the trait as instrumental variables. The use of genetic predictors of telomere length reduces the possibility that the association is explained by reverse causation and confounding likely to be relevant in case-control studies where telomere length is measured directly because genotypes in the general population should be randomly distributed. Additionally, predicted telomere length has the added advantage of representing telomere length over the lifetime of the individual while measuring LTL directly will represent only one time point in an individual's life. Methods Myeloid leukemia (AML and CML) and MDS cases were identified by rapid case ascertainment through the Minnesota Cancer Surveillance System (MCSS). Centralized pathology and cytogenetics reviews were conducted to confirm diagnosis and classify by subtypes. Controls were identified through the Minnesota State driver's license/identification card list. We used the Sequenom platform to genotype seven SNPs that were validated predictors of LTL in a recently published meta-analysis. We estimated associations between individual SNPs and myeloid malignancy using multivariable logistic regression. Mendelian randomization was used to evaluate the association between predicted LTL and risk of myeloid malignancy. Results We included 253 AML cases, 353 MDS cases, 145 CML cases, and 1,042 controls. We observed a significant association between longer predicted telomere length and AML in analyses adjusted for sex and age (OR 4.82 per additional kilobase of average telomere length, 95% CI 1.32-17.63). The individual SNP analyses suggest that the association is largely driven by rs10936599, located in TERC (OR 1.47 per allele, 95% CI 1.14-1.90; Figure). We did not observe statistically significant associations between predicted LTL and MDS or CML. Discussion In this analysis of predicted telomere length and myeloid malignancy, we identified a significant association between longer predicted leukocyte telomere length and AML. In contrast, no significant association was observed for MDS or CML although we did observe an elevated OR for longer predicted telomere length and MDS that did not reach statistical significance. Figure Disclosures No relevant conflicts of interest to declare.