PURIFICATION OF FSH BINDING INHIBITORS

从猪卵泡膜中分离纯化了两种FSH受体结合抑制剂 卵泡液 有不同的物质， 对FSH反应细胞的影响;一种（58，000 MW蛋白质）是 刺激性（FSH激动剂），其他（复合糖肽）抑制性 (FSH拮抗剂）。该项目的具体目标是：1）获得 足够量的高度纯化的化学抑制剂， 生物学研究，2）化学鉴定两种抑制剂，3） 开发用于研究抑制剂的特异性探针，4）鉴定 这两种抑制剂的作用机制，以及5）鉴定它们的 体内细胞定位。 提出的纯化方法是基于以前的经验， 两种抑制剂均来自猪卵泡液，化学成分研究 低分子量抑制剂的抗原性和高分子量抑制剂的抗原性。 作用机制研究包括体外受体结合（Sertoli 细胞培养）和体内实验，旨在确定这些 抑制剂与FSH受体本身相互作用，如果是这样， 这些抑制剂与FSH所涉及的相同受体表位不相互作用， 约束力FSH β亚基或高分子量FSH的合成肽片段 结合抑制剂和针对这些合成肽的抗体将被 作为特异性探针产生，以促进作用机制研究。 免疫组织化学研究将用于定位抑制剂， vivo. 长期目标包括评估的生理意义， 抑制剂的这些抑制剂可能被证明是代表当地 （旁分泌/自分泌）调节剂在受体作用水平上发挥作用 调节FSH对性腺功能的控制。这种行动意味着， 生理学研究将因局部的代偿性变化而复杂化。 调节剂作为浓度的任何一个因素是实验 被操纵了实现本提案的目标将有助于 这一长期目标通过提供有关蜂窝的信息 定位抑制剂和开发特异性探针。获得的数据 通过这个项目，有望提高我们对FSH的理解 结构/功能关系。此外，该项目可能导致 提高对正常性腺功能的理解（例如，生育率）。以来 局部调节因子生物合成的异常控制可能是 某些形式的人类不育（例如，高促性腺激素性腺功能减退症），新 还可以获得对这些疾病的病因学的了解。

Two inhibitors of FSH binding to receptor have been purified from porcine follicular fluid. There are distinct substances which have opposite effects on FSH responsive cells; one (a 58,000 MW protein) being stimulatory (FSH agonist), the other (a complex glycopeptide) inhibitory (FSH antagonist). The specific aims of this project are: 1) to obtain sufficient quantities of highly purified inhibitors for chemical and biological study, 2) to chemically identify the two inhibitors, 3) to develop specific probes for use in studying the inhibitors, 4) to identify the mechanism of action of both inhibitors, and 5) to identify their cellular localization in vivo. Purification methods proposed are based on previous experience isolating both inhibitors from porcine follicular fluid, chemical composition studies of low MW inhibitor and antigenic characterization of high MW inhibitor. Mechanism of action studies include receptor binding, in vitro (Sertoli cell culture) and in vivo experiments designed to determine if these inhibitors interact with receptor for FSH per se, and, if so, whether or not the inhibitors interact with the same receptor epitopes involved in FSH binding. Synthetic peptide fragments of FSH beta subunit or high MW FSH binding inhibitor and antibodies against these synthetic peptides will be generated as specific probes to facilitate mechanism of action studies. Immunohistochemical studies will be used to localize the inhibitors in vivo. Long term goals include evaluation of the physiological significance of the inhibitors. These inhibitors may prove to represent local (paracrine/autocrine) regulators which act at the level of receptor action to modulate FSH control of gonadal function. Such action implies that physiological studies will be complicated by compensatory changes in local modulators as concentrations of any one factor are experimentally manipulated. Achieving the goals of the present proposal will facilitate this long term goal by providing information regarding the cellular localization of inhibitors and by developing specific probes. Data obtained by this project is expected to improve our understanding of FSH structure/function relationships. Furthermore, this project may lead to improved understanding of normal gonadal function (e.g., fertility). Since aberrant control of the biosynthesis of local regulators might underlie some forms of human infertility (e.g., hypergonadotropic hypogonadism), new insights into the etiology of these diseases may also be obtained.