Epigenetic modifications alter telomere length during preimplantation embryo development

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OBJECTIVE: To study epigenetic effects on telomere length during preimplantation development and in embryonic stem cells (ESC's). DESIGN: Laboratory study of embryos and ESC's. MATERIALS AND METHODS: Telomere length affects the development, differentiation and longevity of cells. In oocytes telomeres shorten with age and reset during early embryo development (NatureCell Biol 9:1436-41, 2007). Epigenetic changes regulate telomere length in somatic cells, but their role in telomere length regulation during preimplantation embryo development and in ESC's have not been studied. We examined the effects of a histone deacetylase inhibitor, trichostatin A (TSA) on telomere length in embryos and ESC's. Eggs were collected from super ovulated C57BL6 mice and activated by strontium. Embryos were cultured in KSOM for 24 hours in the presence or absence of 0.3 μM TSA. After 24 hours, treated (N=62) and untreated (N=58) morphologically normal 2 cell embryos were frozen at – 20°C for later telomere length measurement by Real Time PCR as described (Nature Cell Biol 9:1436-41, 2007). Histone deacetylase inhibition was verified by immunostaining. ESC's, isolated from blastocysts of young C57BL6 mice, were cultured for 24 hours in the presence or absence of 0.3 μM TSA. Stem cell markers verified ESC's. Telomere length was measured in 0.3-0.6 ng DNA from embryos and 40 ng of stem cells. Experiments were run in duplicate for embryos and triplicate for stem cells. For each PCR reaction, a standard curve was made using serial dilutions of known concentration of genomic DNA. Data analysis was by the 2-ΔΔ Ct method. RESULTS: Inhibition of histone deactylation shortened telomeres over five fold in embryos (mean Ct 19.55±1.55 for TSA group and 17.5 ± 2.90 for controls). TSA did not alter telomere length in ESC's (mean Ct 9.85±0.29 for controls group and 9.76±0.35 for TSA treated). CONCLUSIONS: Epigenetic regulation has been shown to underlie the global changes in transcription during development. Here we show that inhibition of a key epigenetic modification, histone deacetylation, alters telomere length in preimplantation embryos. Telomere length in ESC's was not affected by inhibition of histone deacetylation, perhaps because of a different role for histone deacetylation in these cells. We do not know whether the epigenetic changes associated with in vitro culture also can alter telomere length in embryos.