Faculty Bibliography

PURPOSE: The effect of age on telomere length heterogeneity in men has not been studied previously. Our aims were to determine the relationship between variation in sperm telomere length (STL), men's age, and semen parameters in spermatozoa from men undergoing in vitro fertilization (IVF) treatment. METHODS: To perform this prospective cross-sectional pilot study, telomere length was estimated in 200 individual spermatozoa from men undergoing IVF treatment at the NYU Fertility Center. A novel single-cell telomere content assay (SCT-pqPCR) measured telomere length in individual spermatozoa. RESULTS: Telomere length among individual spermatozoa within an ejaculate varies markedly and increases with age. Older men not only have longer STL but also have more variable STL compared to younger men. STL from samples with normal semen parameters was significantly longer than that from samples with abnormal parameters, but STL did not differ between spermatozoa with normal versus abnormal morphology. CONCLUSION: The marked increase in STL heterogeneity as men age is consistent with a role for ALT during spermatogenesis. No data have yet reported the effect of age on STL heterogeneity. Based on these results, future studies should expand this modest sample size to search for molecular evidence of ALT in human testes during spermatogenesis.

PURPOSE: Ovarian aging is closely tied to the decline in ovarian follicular reserve and oocyte quality. During the prolonged reproductive lifespan of the female, granulosa cells connected with oocytes play critical roles in maintaining follicle reservoir, oocyte growth and follicular development. We tested whether double-strand breaks (DSBs) and repair in granulosa cells within the follicular reservoir are associated with ovarian aging. METHODS: Ovaries were sectioned and processed for epi-fluorescence microscopy, confocal microscopy, and immunohistochemistry. DNA damage was revealed by immunstaining of gammaH2AX foci and telomere damage by gammaH2AX foci co-localized with telomere associated protein TRF2. DNA repair was indicated by BRCA1 immunofluorescence. RESULTS: DSBs in granulosa cells increase and DSB repair ability, characterized by BRCA1 foci, decreases with advancing age. gammaH2AX foci increase in primordial, primary and secondary follicles with advancing age. Likewise, telomere damage increases with advancing age. In contrast, BRCA1 foci in granulosa cells of primordial, primary and secondary follicles decrease with monkey age. BRCA1 positive foci in the oocyte nuclei also decline with maternal age. CONCLUSIONS: Increased DSBs and reduced DNA repair in granulosa cells may contribute to ovarian aging. Discovery of therapeutics that targets these pathways might help maintain follicle reserve and postpone ovarian dysfunction with age.

OBJECTIVE: To analyze whether leukocyte telomere length (LTL) is impaired in women with polycystic ovary syndrome (PCOS). DESIGN: Case-control study. SETTING: Hospital. PATIENT(S): A total of 274 women, including 150 patients with PCOS and 124 controls. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Body mass index (BMI), waist circumference, systemic arterial pressure, lipid profile, E2, LH, T, androstenedione, PRL, TSH, sex hormone-binding globulin, C-reactive protein (CRP), homocysteine, free androgen index, and the homeostatic model of insulin sensitivity (HOMA-IR) index were analyzed. The LTL evaluation was measured by quantitative polymerase chain reaction. RESULT(S): The PCOS group had higher values for weight, BMI, waist circumference, systolic arterial pressure, triglycerides, LH, T, insulin, CRP, free androgen index, and HOMA-IR compared with the control group. Sex hormone-binding globulin and E2 levels were lower in the PCOS group than in the control group. The LTL did not differ between groups. Age, BMI, and HOMA-IR had no significant effect on LTL. The inflammatory biomarkers CRP and homocysteine were negatively correlated with LTL in patients with PCOS. CONCLUSION(S): Our results showed no differences in LTL between patients with PCOS and controls, but CRP and homocysteine biomarkers negatively correlated with LTL in the PCOS group.

The oocyte is the major determinant of embryo developmental competence in women. It delivers half the chromosomal complement to the embryo, but the maternal and paternal genomes are neither symmetrical nor equal in their contributions to embryo fate. Unlike the paternal genome, the maternal genome carries a heavy footprint of parental aging. Indeed, age is the single best predictor of reproductive outcome in women, and the oocyte is the locus of reproductive aging in women. The oocyte transmits not only the mother's nuclear but also her mitochondrial genome to the embryo, and mitochondrial DNA is known to be especially susceptible to aging. Morphological studies of the oocyte and its associated cumulus corona cells provide only marginal value in the assessment of embryo developmental potential. A number of novel technologies, however, have improved the noninvasive assessment of oocyte quality. Moreover, during maturation, the oocyte ejects half its homologous chromosomes into the first polar body and half its chromatids into the second polar body. Polar body DNA is remarkably similar to that of the oocyte, so analysis of polar body DNA provides a window into the oocyte's genome and telomeres, which may enhance prediction of embryo developmental competence.

Haplo-insufficiency of telomerase genes in humans leads to telomere syndromes such as dyskeratosis congenital and idiopathic pulmonary fibrosis. Generation of pluripotent stem cells from telomerase haplo-insufficient donor cells would provide unique opportunities toward the realization of patient-specific stem cell therapies. Recently, pluripotent human embryonic stem cells (ntESCs) have been efficiently achieved by somatic cell nuclear transfer (SCNT). We tested the hypothesis that SCNT could effectively elongate shortening telomeres of telomerase haplo-insufficient cells in the ntESCs with relevant mouse models. Indeed, telomeres of telomerase haplo-insufficient (Terc+/-) mouse cells are elongated in ntESCs. Moreover, ntESCs derived from Terc+/- cells exhibit naive pluripotency as evidenced by generation of Terc+/- ntESC clone pups by tetraploid embryo complementation, the most stringent test of naive pluripotency. These data suggest that SCNT could offer a powerful tool to reprogram telomeres and to discover the factors for robust restoration of telomeres and pluripotency of telomerase haplo-insufficient somatic cells.