No abstract

C-type natriuretic peptide (CNP) has been considered as a physiological meiotic inhibitor that stimulates the cGMP production by cumulus cell natriuretic peptide receptor 2 (NPR2), which inhibits oocyte phosphodiesterase type 3 activity and increases cAMP. In this study, we explored the effect of CNP pretreatment on the in vitro maturation (IVM) of bovine oocytes by examining changes in cleavage rate, blastocyst formation, mitochondrial DNA (mtDNA) copy number, reactive oxygen species (ROS) level, glutathione (GSH) content, and redox state. Our results showed that 200 nM CNP could effectively maintain meiotic arrest of bovine oocytes in vitro within 6 h. The two-step IVM system in which oocytes were pretreated with 200 nM CNP for 6 h and then cultured IVM for 28 h yielded a significantly (P < 0.05) increased blastocyst rate and cell number after in vitro fertilization (IVF) while compared to the conventional one-step IVM method. In addition, in comparison with the conventional 24-h matured oocyte, oocytes pretreated with 200 nM CNP for 6 h followed by 28 h IVM resulted in significantly (P < 0.05) higher mtDNA copy number and ROS levels in oocytes, while GSH level significantly (P < 0.05) decreased. Remarkably, regardless of treatment, no changes were observed in FAD++, NAD(P)H autofluorescence intensity, and redox ratio (FAD++/NAD(P)H) within the oocytes, maintaining a healthy metabolic equilibrium of redox throughout the two-step IVM. In conclusion, these results indicate that CNP pretreatment could dramatically improve the quality of bovine oocytes during in vitro maturation.

No abstract

Metformin (MET) can effectively treat endometrial hyperplasia (EH), and the expression of glucose transporter type 4 insulin-responsive (GLUT4) is closely associated with the development of EH. The present study aimed to verify the effect of MET in functional EH and polycystic ovary syndrome (PCOS). H&E staining was performed to analyze the severity of EH, and immunohistochemistry was performed to evaluate the expression of GLUT4 in the endometrium of PCOS rats. Reverse transcription-quantitative PCR was used to calculate the expression of long non-coding (lnc)RNA-maternally expressed gene 3 (MEG3), lncRNA-small nucleolar RNA host gene 20 (SNHG20), GLUT4 mRNA, microRNA (miR)-223 and miR-4486. Sequence analysis and luciferase assays were performed to explore the regulatory relationship among certain lncRNAs, miRNAs and target genes. EH in PCOS rats was efficiently inhibited by MET administration. The increased expression of GLUT4 in PCOS rats was attenuated by MET treatment. Moreover, the expression levels of lncRNA-MEG3 and lncRNA-SNHG20 were significantly inhibited in the endometrium of PCOS rats. MET treatment also showed remarkable efficiency in restoring the expression of lncRNA-MEG3 and lncRNA-SNHG20. Meanwhile, the expression levels of miR-223 and miR-4486 were notably elevated in the endometrium of PCOS rats, while MET treatment reduced the expression of miR-223 and miR-4486 in PCOS rats. Furthermore, a luciferase assay confirmed the inhibitory relationship between miR-223 and lncRNA-MEG3/GLUT4 expression, as well as between miR-4486 and lncRNA-SNHG20/GLUT4 expression. GLUT4 knockdown restored the decreased viability of HCC-94 cells induced by overexpression of lncRNA-MEG3. To conclude, MET exhibited a therapeutic effect in the treatment of EH by modulating the lncRNA-MEG3/miR-223/GLUT4 and lncRNA-SNHG20/miR-4486/GLUT4 signaling pathways. This work provides mechanistic insight into the development of EH.

The low efficiency of in vitro embryo production is associated with oxidative stress induced by suboptimal culture conditions. p66Shc is a 66‐kDa protein of the ShcA (Src homologous‐collagen homolog) adaptor protein family, which is involved in signaling pathways involved in oxidative stress regulation, apoptosis induction, and aging. However, the functional role of p66Shc during the preimplantation development of sheep embryos is not understood. Our results showed that early‐cleavage (≤28 hr) embryos had a higher developmental potential than late‐cleavage (>28 hr) embryos. The poor quality of these late‐cleavage embryos was associated with increased the transcripts and protein of p66Shc and decreased mitochondrial activity. In addition, exogenous hydrogen peroxide‐induced oxidative stress significantly increased p66Shc protein abundance and suppressed embryonic development, which was ameliorated by antioxidant treatment. Notably, oxidative stress induced the nuclear localization of p66Shc and phosphorylated (Ser‐36) p66Shc. Collectively, these observations suggest that p66Shc may be playing an important role in the regulation of oxidative stress during the preimplantation development of sheep embryos.