RESUMO

The present study examined whether male resveratrol intake affected mitochondrial DNA copy number (mt-cn) and telomere length (TL) in blastocysts fathered by young and aged male mice. C57BL/6N male mice supplied with water or water containing 0.1 mM resveratrol were used for embryo production at 14-23 and 48-58 weeks of age. Two-cell-stage embryos were collected from the oviducts of superovulated female mice (8-15 weeks old) and cultured for 3 days until the blastocyst stage. Mt-cn and TL levels were measured by real-time polymerase chain reaction. Resveratrol intake did not affect body weight or water consumption. Resveratrol intake increased the expression levels of SIRT1 in the liver, the antioxidative ability of serum, and extended TL in the heart, whereas there was no significant difference in mt-cn in the heart or TL in sperm. The rate of blastocyst development was significantly lower in aged male mice than in younger mice, and resveratrol intake increased the total number of blastocysts derived from both young and aged males. Resveratrol intake did not affect mt-cn or TL in blastomeres of blastocyst-stage embryos derived from young mice, but significantly increased both mt-cn and TL in blastomeres of blastocysts derived from aged fathers. In conclusion, resveratrol intake increased mt-cn and TL levels in blastocysts derived from aged male mice.

Assuntos

Blastocisto , DNA Mitocondrial , Camundongos Endogâmicos C57BL , Resveratrol , Telômero , Animais , Resveratrol/farmacologia , Masculino , Blastocisto/efeitos dos fármacos , Blastocisto/metabolismo , Feminino , Camundongos , DNA Mitocondrial/metabolismo , Telômero/efeitos dos fármacos , Telômero/metabolismo , Homeostase do Telômero/efeitos dos fármacos , Desenvolvimento Embrionário/efeitos dos fármacos , Espermatozoides/efeitos dos fármacos , Espermatozoides/metabolismo , Sirtuína 1/metabolismo , Sirtuína 1/genética , Variações do Número de Cópias de DNA/efeitos dos fármacos , Antioxidantes/farmacologia , Antioxidantes/metabolismo , Envelhecimento , Estilbenos/farmacologia , Idade Paterna

RESUMO

Ethanol affects pre-conceptional oocyte quality in women. In this study, we examined the effect of low ethanol concentrations on mouse oocytes. Oocytes were collected from the ovaries of 9-10 week old mice and allowed to mature in vitro in the presence of low concentrations of ethanol (0.1% and 0.2% v/v) for 24 h. Treatment of oocytes with ethanol (0.2%) during maturation decreased the mitochondrial DNA content and membrane potential compared to that in untreated ones, whereas the ATP content did not differ between the groups. Both 0.1% and 0.2% ethanol reduced the lipid content in the oocytes. In addition, immunostaining revealed that oocytes cultured in maturation medium containing ethanol (0.2%) had reduced levels of global DNA methylation and DNMT3A compared with untreated oocytes, and decreased rate of blastocyst development with low mitochondrial protein levels (TOMM40) in embryo. RNA-sequencing of the ethanol-treated (0.2%) and untreated oocytes revealed that mitochondria were a major target of ethanol. In conclusion, treatment of oocytes with low concentration of ethanol reduces the developmental rate to the blastocyst stage, with a lower total cell number and global DNA methylation. In addition, ethanol affected mitochondrial function and mitochondria-related gene expression.

Assuntos

Metilação de DNA , Etanol , Técnicas de Maturação in Vitro de Oócitos , Mitocôndrias , Oócitos , Animais , Oócitos/efeitos dos fármacos , Oócitos/metabolismo , Etanol/farmacologia , Camundongos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Feminino , Metilação de DNA/efeitos dos fármacos , Técnicas de Maturação in Vitro de Oócitos/veterinária , Desenvolvimento Embrionário/efeitos dos fármacos , Meios de Cultura/química , Blastocisto/efeitos dos fármacos , Blastocisto/metabolismo , DNA Mitocondrial/metabolismo , Transcriptoma/efeitos dos fármacos , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Potencial da Membrana Mitocondrial/efeitos dos fármacos

RESUMO

Telomere length (TL) and mitochondrial DNA copy number (mt-cn) are associated with embryonic development. Here, we investigated the correlation between TL and mt-cn in bovine embryos to determine whether TL regulates mt-cn. TL and mt-cn were closely correlated in embryos derived from six bulls. Treatment of embryos with a telomerase inhibitor (TMPyP) and siTERT shortened the TL and reduced mt-cn in blastocysts. RNA-sequencing of blastocysts developed with TMPyP revealed differentially expressed genes associated with transforming growth factor-ß1 signaling and inflammation. In conclusion, TL regulates mt-cn in embryos.

Assuntos

Blastocisto , Variações do Número de Cópias de DNA , Animais , Bovinos , Blastocisto/metabolismo , Blastocisto/efeitos dos fármacos , Telômero/metabolismo , DNA Mitocondrial/genética , DNA Mitocondrial/metabolismo , Masculino , Feminino , Homeostase do Telômero/efeitos dos fármacos , Mitocôndrias/metabolismo , Mitocôndrias/genética , Desenvolvimento Embrionário/efeitos dos fármacos , Desenvolvimento Embrionário/genética

RESUMO

In brief: This study shows that ageing affects miRNA profiles in follicular fluid, and an miRNA that is highly abundant in the follicular fluid of young cows supports the growth of oocytes derived from early antral follicles. Abstract: We examined age-associated changes in miRNA profiles in the follicular fluid (FF) of cows. The role of miR-19b, which is abundant in the FF of young cows, in in vitro growth of early antral follicles (EAFs)-derived oocytes was assessed. FF was collected from the antral follicles of young (20-40 months) and aged (>120 months) cows. The miRNA profiles were similar between the FF of both age groups, whereas the abundance of some miRNAs differed between these samples. The miRNA profiles in granulosa cells (GCs) and the spent culture medium of oocyte-GC complexes (OGCs) derived from EAFs were distinct. Some miRNA groups overlapped among the GCs, culture media, and FFs. miR-19b was highly abundant in the FF of young cows, GCs, and culture medium. The supplementation of OGC culture medium with miR-19b increased the diameter, acetylation levels, and fertilisation ability of the oocytes. To assess whether miR-19b was functional in the GCs, a dual-luciferase assay, suppression of target protein, and RNA-sequencing of the GCs followed by functional annotation of the differentially expressed genes (DEGs) were conducted. Functional annotation of the DEGs suggested that miR-19b influences genes associated with FoxO signalling, endocytosis, and NR3C1 in GCs. These results suggest that in FFs, ageing affects the abundance of miRNAs that have important roles in oocyte development.

Assuntos

Líquido Folicular , MicroRNAs , Animais , Bovinos , Meios de Cultura , Feminino , Líquido Folicular/metabolismo , Células da Granulosa/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Oócitos , Folículo Ovariano

RESUMO

PURPOSE: The present study examined the relationships among the amount of cell-free-DNA (cfDNA) in porcine follicular fluid (FF), the developmental ability of enclosed oocytes, and characteristics of granulosa cells and examined the effect of cfDNA content in maturation medium on the developmental ability of the oocytes. METHODS: Oocytes and FF were collected from individual gilts, and the gilts were rated based on the ability of their oocytes to develop to the blastocyst stage and the amount of cfDNA in the FF. The copy numbers of mitochondrial DNA (Mt-DNA) and nuclear DNA (N-DNA) were measured by real-time PCR and the DNA sequence. FF or cfDNA was added to the maturation medium, and the developmental ability of the oocytes was examined. RESULTS: The amount of cfDNA was associated with apoptosis of the granulosa cells, and high-cfDNA content in FF was associated with low developmental ability of oocytes. Supplementation of the maturation medium with FF containing high cf-Mt-DNA or with DNA extracted from the FF did not affect oocyte developmental competence. CONCLUSIONS: Cell-free DNA content in FF is a marker for oocyte competence, but cfDNA in the oocyte maturation environment did not affect oocyte developmental ability.