Sirois J, Doré M
Centre de Recherche en Reproduction Animale, Faculté de Médecine Vétérinaire, Université de Montréal, Saint-Hyacinthe, Québec, Canada.
Endocrinology. 1997 Oct;138(10):4427-34. doi: 10.1210/endo.138.10.5462.
PGs are important mediators of the ovulatory process and prostaglandin G/H synthase-2 (PGHS-2) is a key rate-limiting enzyme in the PG biosynthetic pathway. To determine whether PGHS-2 is regulated in equine follicles before ovulation and, if so, to characterize its time course of induction, preovulatory follicles were isolated during estrus, 0, 12, 24, 30, 33, 36, and 39 h after an ovulatory dose of hCG (n = 5 follicles/time point). Cellular extracts were obtained from preparations of follicle wall (theca interna with attached granulosa cells), isolated granulosa cells, and theca interna and were analyzed by Western blot using specific anti-PGHS antibodies. Immunohistochemistry was used to characterize the in situ localization of PGHS-2 protein in preovulatory follicles, and follicular fluid concentrations of PGE2 and PGF were determined. The results showed the induction of PGHS-2, but not PGHS-1, in equine follicles before ovulation. The PGHS-2 protein (72,000 mol wt) was undetectable 0, 12, and 24 h post-hCG, first became apparent at 30 h, and reached maximal levels 39 h after hCG treatment. The induction of follicular PGHS-2 was localized exclusively in granulosa cells, and a pronounced staining was observed in the perinuclear region. Follicular fluid concentrations of PGE2 and PGF were low and not different between 0-33 h, but levels were increased at 36 and 39 h post-hCG (P < 0.01). Thus, the time course of PGHS-2 induction in equine follicles (30 h post-hCG) is clearly distinct from those previously observed in rat (4 h post-hCG) and bovine (18 h post-hCG) preovulatory follicles. Interestingly, in all three species, the interval from PGHS-2 induction to follicular rupture is highly conserved (approximately 10 h). Therefore, the progressively delayed expression of PGHS-2 in species with longer ovulatory processes supports its role as a molecular determinant of the species-specific length of the ovulatory process.
前列腺素（PGs）是排卵过程的重要介质，前列腺素G/H合酶-2（PGHS-2）是PG生物合成途径中的关键限速酶。为了确定PGHS-2在马卵泡排卵前是否受到调控，若受调控，则描述其诱导的时间进程，在发情期、给予促排卵剂量的人绒毛膜促性腺激素（hCG）后0、12、24、30、33、36和39小时分离排卵前卵泡（每个时间点n = 5个卵泡）。从卵泡壁（附着颗粒细胞的内膜）、分离的颗粒细胞和内膜制备物中获取细胞提取物，并用特异性抗PGHS抗体通过蛋白质免疫印迹法进行分析。免疫组织化学用于表征PGHS-2蛋白在排卵前卵泡中的原位定位，并测定卵泡液中前列腺素E2（PGE2）和前列腺素F（PGF）的浓度。结果显示，马卵泡在排卵前诱导了PGHS-2，但未诱导PGHS-1。PGHS-2蛋白（分子量72,000）在hCG后0、12和24小时检测不到，在30小时首次出现，并在hCG处理后39小时达到最高水平。卵泡PGHS-2的诱导仅定位于颗粒细胞，并且在核周区域观察到明显的染色。卵泡液中PGE2和PGF的浓度较低，在0至33小时之间无差异，但在hCG后36和39小时水平升高（P < 0.01）。因此，马卵泡中PGHS-2诱导的时间进程（hCG后30小时）与先前在大鼠（hCG后4小时）和牛（hCG后18小时）排卵前卵泡中观察到的明显不同。有趣的是，在所有三个物种中，从PGHS-2诱导到卵泡破裂的间隔高度保守（约10小时）。因此，在排卵过程较长的物种中PGHS-2表达的逐渐延迟支持了其作为排卵过程物种特异性长度的分子决定因素的作用。
