| Nitric oxide (NO) is an important biological messenger in the regulation of tissue homeostasis. It exhibits a wide range of effects during physiological and pathophysiological processes. Typical beneficial properties of NO include the regulation of vascular tone, the protection of cells against apoptosis, themodulation of immune responses, and the killing of microbial pathogens. On the other hand, NO may cause severe vasodilation and myocardial depression during bacterial sepsis or act as a cytotoxic and tissue-damaging molecule in autoimmune diseases. NO is closely involved in mammalian reproductive system, such as sexual behavior, steriodogenesis, follicle and oocyte development, blastocyst implantation and embryo development. Such research is beneficial to animal and human nutrion, welfare, reproduction fitness, diseases and even xenotransplantation in a long run. As the result, this study was to investigate the functions of NO on porcine oocyte meiotic maturation and its mechanisms. It consists of 6 independent experiments.Experiment 1. The present investigation was undertaken to ascertain the effects of NO at high level on porcine oocyte meiotic development. Cumulus-enclosed oocytes (CEOs) and denuded oocytes (DOs) were obtained from the follicles (2-6 mm diameter). Exogenous NO was provided with sodium nitroprusside (SNP). 0 mM (Control), 0.1 mM, 1 mM or 10 mM SNP was introduced to the medium. Meanwhile, 0.1 mM, 1 mM and 10 mM preincubated SNP was also added to the medium, respectively. The cumulus expansion, cumulus cells DNA fragmentation, oocyte meiotic maturation and degeneration were determined 44 h after incubation. SNP inhibited cumulus expansion and cumulus cells DNA fragmentation in a dose-dependent manner. Significantly fewer CEOs treated by any level of SNP resumed meiosis than control (P<0.05) and more CEOs exposed to 0.1 mM or 1 mM SNP were arrested at GV stage. Moreover, SNP increased the percentage of CEOs at MI stage but decreased the percentage of CEOs at MII stage. 0.1 mM or 1 mM SNP showed no influence on the proportion of GVBD of DOs and only 10 mM SNP reduced the proportion. SNP accelerated degeneration of both CEOs and DOs, especially at the high concentrations. DOs were more sensitive to SNP toxicity. Preincubated SNP showed no effect on cumulus expansion, cumulus cells DNA fragmentation, oocyte meiotic maturation or degeneration at lower concentration, but higher concentrations had effect, especially on the viability of porcine oocytes. Taken together, high level of NO inhibits porcine oocyte meiotic maturation, especially the transition from Ml to M II, by mediating functions of cumulus cells.Experiment 2. This study was conducted to investigate the involvement of nitric oxide (NO) in cumulus expansion, oocyte mortality and meiotic maturation of porcine cumulus enclosed oocytes (CEOs) cultured in two different models when gonadotropins, including follicle-stimulating hormone (FSH) and human chorionic gonadotropin (hCG) were presented or not. And the interaction between NO and p-mercaptoethanol (p-ME), a free radical scavenger was also investigated. Two models refer to spontaneous maturation model and hypoxanthine (HX) medium model. All the 3433 eligible CEOs were incubated at 39C and the cumulus expansion, oocyte morphology and nuclear phase were evaluated 44 h after incubation. (1) In spontaneous maturation model, NO stimulates the cumulus expansion and P-ME delayed it. NO doesn't affect the oocyte meiotic resumption but inhibits the oocytes to develop to metaphase II. (2) In HX medium model, NO or P-ME doesn't affect the expansion in the absence of gonadotropins, but in the presence of gonadotropins, NO or P-ME inhibits the expansion. In the presence of gonadotropins, NO inhibits the oocyte meiotic resumption, and it especially inhibits the oocyte to develop to metaphase II, and P-ME reverses such inhibitory effects. The cooperation of gonadotropins and P-ME stimulates the meiotic resumption, and, especially, promotesthe CEOs to develop to metaphase II in both m...
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