INTRAFOLLICULAR REGULATION OF OVARIAN TGF-B1 EXPRESSION

转化生长因子β1(TGF-β1)是一种潜在的重要的卵巢内生长因子，由于其对卵泡膜和颗粒细胞功能的显著影响以及与卵泡发生有关的生长作用的局限性，我们对其在细胞水平的调控及其与卵巢功能的关系仍存在着显著的认识不足。解决转化生长因子-β1的功能和调控之谜代表了该项目的长期目标。利用猪卵巢的培养细胞和完整的半卵泡，建立了一个调控转化生长因子-β1表达的模型。数据表明，尽管这两种细胞都表达转化生长因子-β1mRNA，但只有膜细胞表达和分泌生长因子。FSH、LH和cAMP都不能刺激培养的两种分离细胞中的转化生长因子-β部分。然而，在小半卵泡中，FSH而不是黄体生成素像cAMP一样刺激转化生长因子-β1的分泌。我们推测卵泡膜细胞在卵泡发育过程中表达和分泌转化生长因子-β1，但它是在FSH作用于颗粒细胞的指导下进行的。我们进一步假设，卵泡刺激素刺激的颗粒细胞产物通过激活PLA/PKC信号转导系统来调节膜细胞转化生长因子-β的表达。为了进一步阐明转化生长因子-β1在卵泡间的调节，我们将确定转化生长因子-β1的自身表达是否发生在颗粒细胞而不是膜细胞，这是否通过选择性地表达转化生长因子-βI和Ⅱ型受体，以及自身表达或受体表达是否随着卵泡的发育而改变。卵泡刺激素和转化生长因子-β1在多大程度上相互调节颗粒细胞中另一种受体的表达将被确定。颗粒细胞可以通过分泌因子直接调节卵泡膜细胞转化生长因子-β1表达的程度将通过共培养实验进行测试。随着卵泡的发育，卵泡转化生长因子-β1的表达是否从促卵泡激素转变为促黄体生成素的调控将被确定。最后，将研究一种胞浆蛋白在颗粒细胞中的选择性表达，该蛋白能够与抑制翻译的转化生长因子-β1 mRNA5‘-非翻译区的调节茎环结构结合。这可能是颗粒细胞差异表达转化生长因子-β1mRNA而不是生长因子的机制。相信本文所述研究的完成将提供关于转化生长因子-β1在卵泡间、旁分泌调节的新的重要信息，这将为深入了解其在卵泡发育中的作用提供依据。

Transforming growth factor beta1 (TGF-beta1) has been identified as a potentially important intraovarian growth factor due to its pronounced effects upon theca and granulosa cell function and the localization of the growth of work regarding TGF-beta1 and folliculogenesis, there remains a notable deficit in our knowledge regarding its regulation at the cellular level and how this relates to ovarian function. Solving the riddle regarding TGF- beta1 function and regulation represents the long term goals of this project. Using both cultured cells and intact hemi-follicles from the porcine ovary, a model for the regulation of TGF-beta1 expression has been formulated. Data suggest that although both cell types express TGF-beta1mRNA, only theca cells express and secrete the growth factor. Neither FSH, LH, nor cAMP stimulates TGF-beta section in either isolated cell type in culture. In small hemi-follicles, however, FSH but not LH stimulates TGF-beta1 secretion as does cAMP. We hypothesize that theca cells express and secrete TGF-beta1 during follicle development but that it is under the direction of FSH acting upon granulosa cells. We further hypothesize that a FSH-stimulated granulosa cell product regulates theca cell TGF-beta expression by activating the PLA/PKC signal transduction system. To further delineate the inter-follicular regulation of TGF-beta1 it will be established whether TGF- beta1 auto expression occurs in granulosa but not theca cells, whether this is mediated by selective expression of TGF-beta type I and II receptors, and whether auto-expression or receptor expression change with follicle development. The extent to which FSH and TGF-beta1 reciprocally regulate the expression of the other receptor in granulosa cells will be determined. The extent to which granulosa cells can directly regulate theca cell TGF- beta1 expression via secretory factors will be tested by co-culture experiments. Whether follicular TGF-beta1 expression switches from FSH to LH governance with follicle development will be determined. Lastly, the selective granulosa cell expression of a cytosolic protein capable of binding to a regulatory stem-loop structure of the TGF-beta1 mRNA 5'-untranslated region which inhibits translation will be investigated. This may be the mechanism by which granulosa cells differentially express TGF-beta1 mRNA but not the growth factor. It is believed that completion of the studies described herein will provide novel and important information regarding inter-follicular, paracrine regulation of TGF-beta1, which will provide insight into its role during follicle development.