Silva C C, Groome N P, Knight P G
School of Animal and Microbial Sciences, University of Reading, Whiteknights, UK.
J Reprod Fertil. 1999 Mar;115(2):381-8. doi: 10.1530/jrf.0.1150381.
Changes in intrafollicular concentrations of different forms of inhibin (free alpha-subunits and alpha beta dimers) occur during follicle development and may influence the oocyte maturation process. The aim of this study was to investigate the effects of inhibin A and free alpha-subunit (pro-alpha C) isolated from bovine follicular fluid on maturation of bovine cumulus-oocyte complexes, as reflected by their competence for embryo development after in vitro fertilization. Bovine cumulus-oocyte complexes were isolated from ovaries obtained from an abattoir and were cultured for 22-24 h at 38.5 degrees C in TCM-199 medium supplemented with 10% oestrous cow serum, pregnant mares' serum gonadotrophin (2.5 iu ml-1) and either inhibin A (0, 0.2 and 1.0 microgram ml-1) or pro-alpha C (0, 2 and 10 micrograms ml-1). Neither inhibin A nor free alpha-subunit affected the cleavage rate of cumulus-oocyte complexes after fertilization (approximately 60%). Inhibin A reduced the proportion of cleaved oocytes reaching the eight-cell stage by 19% (P < 0.05), but did not affect the yield of blastocysts. However, pro-alpha C decreased the proportion of cleaved oocytes that reached the eight-cell (25%; P < 0.05) and blastocyst (28%; P < 0.05) stages. In addition, a negative correlation (r = -0.55, P < 0.001) was found between concentrations of total immunoreactive (ir) alpha-inhibin (measured by radioimmunoassay) produced by untreated control cumulus-oocyte complexes and their post-cleavage development to the blastocyst stage. In a second experiment, mouse monoclonal antibodies (20 micrograms ml-1) against two different regions of the inhibin alpha-subunit precursor (pro-region and alpha C fragment) were tested for their ability to neutralize endogenous inhibin alpha-subunit-related molecules produced by cumulus cells; control cumulus-oocyte complexes were treated with normal mouse IgG (20 micrograms ml-1). Although the cleavage rate was not affected, the yield of blastocysts was significantly higher in the presence of mouse monoclonal antibodies to both pro-alpha (77% increase; P < 0.05) and alpha C (48% increase; P < 0.05). None of the treatments tested affected endogenous production of activin-A or follistatin by cumulus-oocyte complexes. Overall, these results indicate that the inhibin alpha-subunit (pro-alpha C) has an inhibitory role in oocyte maturation which is independent of the modulatory effects of activin and follistatin.
在卵泡发育过程中，不同形式抑制素（游离α亚基和αβ二聚体）的卵泡内浓度会发生变化，这可能会影响卵母细胞的成熟过程。本研究的目的是探讨从牛卵泡液中分离出的抑制素A和游离α亚基（前αC）对牛卵丘-卵母细胞复合体成熟的影响，这可通过体外受精后它们发育成胚胎的能力来反映。从屠宰场获取的卵巢中分离出牛卵丘-卵母细胞复合体，并在补充有10%发情母牛血清、孕马血清促性腺激素（2.5国际单位/毫升）以及抑制素A（0、0.2和1.0微克/毫升）或前αC（0、2和10微克/毫升）的TCM-199培养基中于38.5℃培养22 - 24小时。抑制素A和游离α亚基均未影响受精后卵丘-卵母细胞复合体的分裂率（约60%）。抑制素A使分裂后的卵母细胞发育到八细胞阶段的比例降低了19%（P < 0.05），但不影响囊胚的产出率。然而，前αC使分裂后的卵母细胞发育到八细胞阶段（降低25%；P < 0.05）和囊胚阶段（降低28%；P < 0.05）的比例均下降。此外，在未处理的对照卵丘-卵母细胞复合体产生的总免疫反应性（ir）α抑制素浓度（通过放射免疫测定）与它们分裂后发育到囊胚阶段之间发现了负相关（r = -0.55，P < 0.001）。在第二个实验中，测试了针对抑制素α亚基前体两个不同区域（前区域和αC片段）的小鼠单克隆抗体（20微克/毫升）中和卵丘细胞产生的内源性抑制素α亚基相关分子的能力；对照卵丘-卵母细胞复合体用正常小鼠IgG（20微克/毫升）处理。虽然分裂率未受影响，但在存在针对前α的小鼠单克隆抗体（增加77%；P < 0.05）和αC的小鼠单克隆抗体（增加48%；P < 0.05）时，囊胚的产出率显著更高。所测试的任何处理均未影响卵丘-卵母细胞复合体中激活素A或卵泡抑素的内源性产生。总体而言，这些结果表明抑制素α亚基（前αC）在卵母细胞成熟中具有抑制作用，这独立于激活素和卵泡抑素的调节作用。
