Reich R, Daphna-Iken D, Chun S Y, Popliker M, Slager R, Adelmann-Grill B C, Tsafriri A
Department of Pharmacology, Hebrew University, Jerusalem, Israel.
Endocrinology. 1991 Oct;129(4):1869-75. doi: 10.1210/endo-129-4-1869.
The preovulatory surge of gonadotropins activates a cascade of proteolytic enzymes resulting in the rupture of the follicular wall and the release of a fertilizable ovum during ovulation. In the rat the process is initiated by a rise in follicular tissue-type plasminogen activator, produced predominantly in granulosa cells. Recent studies revealed a preovulatory increase in ovarian collagenolytic activity in vivo and an increase in activatable collagenase in vitro. In view of the complicated control of mammalian collagenase synthesis and activity by local inhibitors and activators, we examined the expression of ovarian interstitial and type IV collagenases and tissue inhibitor of metalloproteinase (TIMP) mRNA after an ovulatory stimulus. Ovarian mRNA was isolated from immature PMSG-treated rats 3, 6, and 9 h after hCG stimulation. Northern blot analyses revealed a mRNA of 1.7 kilobases (kb) hybridizing with the human interstitial collagenase cDNA probe. The levels of this mRNA showed a 25-fold increase between 3-6 h after hCG stimulation. The human cDNA probe of collagenase IV hybridized with a mRNA of 3.1 kb, which showed only a 4-fold increase 9 h after hCG treatment. The interstitial collagenase mRNA was expressed in both granulosa cells of preovulatory follicles and the residual ovarian tissue, whereas the expression of collagenase IV mRNA was limited to the residual tissue. Inhibitors of eicosanoid synthesis, previously shown to block ovulation and the LH/hCG-induced rise in ovarian collagenolysis, suppressed the gonadotropic stimulation of interstitial collagenase mRNA, but slightly stimulated that of collagenase IV. The mouse cDNA probe of TIMP hybridized with a 0.9-kb mRNA, which was stimulated by hCG to reach a maximum (7- to 8-fold increase) between 6-9 h after stimulation. TIMP was expressed and stimulated in both the granulosa cells and the residual tissue. Inhibitors of eicosanoid synthesis did not affect the gonadotropic stimulation of TIMP mRNA. These data support the suggested role of interstitial collagenase in follicle rupture and the essential role of eicosanoids in the mediation of gonadotropic stimulation of interstitial collagenase production and action. The observed stimulation of TIMP mRNA expression by the gonadotropin and the lack of any effect of eicosanoid synthesis inhibitors on this action of LH/hCG offer an additional mechanism by which these inhibitors may block ovulation. Thus, the suppression of ovulation by inhibitors of eicosanoid synthesis may result from selective inhibition of interstitial collagenase expression and undisturbed gonadotropin-stimulated TIMP expression.
促性腺激素的排卵前激增会激活一系列蛋白水解酶，导致卵泡壁破裂，并在排卵时释放出可受精的卵子。在大鼠中，这一过程由卵泡组织型纤溶酶原激活物的增加引发，该激活物主要由颗粒细胞产生。最近的研究表明，体内卵巢胶原酶活性在排卵前会增加，体外可激活胶原酶也会增加。鉴于哺乳动物胶原酶的合成和活性受到局部抑制剂和激活剂的复杂调控，我们研究了排卵刺激后卵巢间质胶原酶、IV型胶原酶以及金属蛋白酶组织抑制剂（TIMP）mRNA的表达情况。从经孕马血清促性腺激素（PMSG）处理的未成熟大鼠中，在人绒毛膜促性腺激素（hCG）刺激后3、6和9小时分离出卵巢mRNA。Northern印迹分析显示，一条1.7千碱基（kb）的mRNA与人类间质胶原酶cDNA探针杂交。该mRNA水平在hCG刺激后3至6小时内增加了25倍。IV型胶原酶的人类cDNA探针与一条3.1 kb的mRNA杂交，hCG处理9小时后该mRNA仅增加了4倍。间质胶原酶mRNA在排卵前卵泡的颗粒细胞和残留卵巢组织中均有表达，而IV型胶原酶mRNA的表达仅限于残留组织。此前已证明能阻断排卵以及LH/hCG诱导的卵巢胶原溶解增加的类花生酸合成抑制剂，可抑制促性腺激素对间质胶原酶mRNA的刺激，但对IV型胶原酶的刺激作用略有增强。TIMP的小鼠cDNA探针与一条0.9 kb的mRNA杂交，该mRNA在hCG刺激下，于刺激后6至9小时达到最大值（增加7至8倍）。TIMP在颗粒细胞和残留组织中均有表达且受到刺激。类花生酸合成抑制剂不影响促性腺激素对TIMP mRNA的刺激。这些数据支持了间质胶原酶在卵泡破裂中所起的作用，以及类花生酸在介导促性腺激素对间质胶原酶产生和作用的刺激方面的重要作用。观察到促性腺激素对TIMP mRNA表达的刺激以及类花生酸合成抑制剂对LH/hCG这一作用无任何影响，这提供了这些抑制剂可能阻断排卵的另一种机制。因此，类花生酸合成抑制剂对排卵的抑制作用可能是由于对间质胶原酶表达的选择性抑制以及促性腺激素刺激的TIMP表达未受干扰所致。
