Smith G W, McCrone S, Petersen S L, Smith M F
Department of Animal Science, University of Missouri, Columbia 65211.
Endocrinology. 1995 Feb;136(2):570-6. doi: 10.1210/endo.136.2.7835290.
Metalloproteinase inhibitors, such as tissue inhibitors of metalloproteinases 1 and 2 (TIMP-1 and -2), play a key role in the regulation of metalloproteinases that modify extracellular matrix composition. Although expression of TIMP-1 within ovarian tissues has been well characterized, little information is available regarding expression of TIMP-2. The objective of the present studies was to characterize the ontogeny and localization of TIMP-2 messenger RNA (mRNA) within ovine preovulatory follicles and luteal tissue. Total cellular RNA was isolated from preovulatory follicles collected before (presurge; n = 3), or 12-14 h after (post surge; n = 4) an LHRH-induced gonadotropin surge, and from luteal tissue collected on days 3, 7, 10, 13, and 16 post estrus (n = 5, 5, 4, 5, and 5, respectively). TIMP-2 mRNA was expressed by both presurge and postsurge follicles, and expression did not increase after the gonadotropin surge (P = 0.44). In situ hybridization localized TIMP-2 mRNA primarily to the thecal layer of post-surge follicles (n = 3). TIMP-2 mRNA was also localized in a heterogeneous distribution within corpora lutea collected on days 3 and 10 post estrus (n = 3 each). Concentrations of TIMP-2 mRNA (picograms per microgram tissue DNA) were greater in corpora lutea collected during the early luteal phase (days 3 and 7) than the late luteal phase (day 16; P < 0.05). TIMP-2 mRNA was detected in purified populations of both small (n = 4) and large (n = 3) luteal cells, and mRNA concentrations (picograms per microgram DNA) were greater in the large luteal cells (P < or = 0.0002). In addition, immunoreactive TIMP-2 (approximately 21,000 M(r)) was detected by Western blot analysis of ovine luteal cell secreted proteins. We conclude that 1) TIMP-2 mRNA is expressed by the thecal layer of ovine preovulatory follicles and expression is not increased by the preovulatory gonadotropin surge; 2) expression of TIMP-2 mRNA is maximal during the early luteal phase; and 3) expression of TIMP-2 mRNA is greatest in large luteal cells.
金属蛋白酶抑制剂，如金属蛋白酶组织抑制剂1和2（TIMP - 1和 - 2），在调节改变细胞外基质组成的金属蛋白酶中起关键作用。尽管TIMP - 1在卵巢组织中的表达已得到充分表征，但关于TIMP - 2表达的信息却很少。本研究的目的是表征TIMP - 2信使核糖核酸（mRNA）在绵羊排卵前卵泡和黄体组织中的个体发生及定位。从促黄体生成素释放激素（LHRH）诱导的促性腺激素激增前（激增前；n = 3）或激增后12 - 14小时（激增后；n = 4）收集的排卵前卵泡，以及发情后第3、7、10、13和16天收集的黄体组织（分别为n = 5、5、4、5和5）中分离总细胞RNA。激增前和激增后的卵泡均表达TIMP - 2 mRNA，且促性腺激素激增后表达未增加（P = 0.44）。原位杂交将TIMP - 2 mRNA主要定位在激增后卵泡的卵泡膜层（n = 3）。TIMP - 2 mRNA在发情后第3天和第10天收集的黄体中也呈异质性分布（各n = 3）。黄体期早期（第3天和第7天）收集的黄体中TIMP - 2 mRNA浓度（皮克/微克组织DNA）高于黄体期晚期（第16天；P < 0.05）。在纯化的小黄体细胞群体（n = 4）和大黄体细胞群体（n = 3）中均检测到TIMP - 2 mRNA，且大黄体细胞中的mRNA浓度（皮克/微克DNA）更高（P ≤ 0.0002）。此外，通过对绵羊黄体细胞分泌蛋白的蛋白质印迹分析检测到免疫反应性TIMP - 2（约21,000 M(r)）。我们得出结论：1）TIMP - 2 mRNA由绵羊排卵前卵泡的卵泡膜层表达，且排卵前促性腺激素激增不会使其表达增加；2）TIMP - 2 mRNA的表达在黄体期早期最高；3）TIMP - 2 mRNA在大黄体细胞中的表达最高。
