BIOREGULATION OF SPERMATOGENESIS

Sertoli细胞在细胞凋亡的生物调节中起重要作用。 通过提供机械支持和化学信号， 生精细胞 该实验室的工作表明， 中间丝蛋白，波形蛋白是磷酸化的， FSH依赖的方式在培养的Sertoli细胞。 我们现在要确定 调节波形蛋白磷酸化的机制和其他 大鼠睾丸支持细胞不同发育阶段细胞骨架蛋白的研究 精子发生 双向聚丙烯酰胺凝胶电泳 用于解析[32 P]正磷酸盐和[γ-32 P] ATP标记的蛋白质 cAMP和Ca 2 +/钙调蛋白依赖性蛋白激活后 激酶。 初步研究表明支持细胞分泌 也是生精阶段依赖性的特异性蛋白质。 我们提出 确定控制这些蛋白质合成的因素 在大鼠精子发生过程中。 生精管节对应于 不同的精子发生阶段，通过它们的透射模式识别 组织学将用[35 S]蛋氨酸和分泌蛋白标记 通过凝胶电泳分离。 支持细胞特异性分泌蛋白 命名为S80、S45和S35的产物将在DEAE-纤维素柱上纯化 色谱分析并表征。 针对这些蛋白质的抗体将 制作。 生长因子对细胞生长调节的影响 支持细胞和管周细胞的促有丝分裂和非促有丝分裂功能将 使用Stiles等人的互补试验和凝胶电泳进行评价。 电泳以确定自分泌或旁分泌效应。 涉及的因素 在由管周细胞诱导的支持细胞功能的调节中 将使用共培养方法进行研究， 相互作用，但防止细胞与细胞接触。 我们将使用凝胶 电泳以确定肾小管周细胞特异性 蛋白质，指定P95，P40和P38，对合成和分泌 [35 S]蛋氨酸标记的支持细胞特异性蛋白，S80、S45和S35。 这些支持细胞特异性分泌蛋白是令人感兴趣的，因为它们 已显示以生精阶段依赖性方式表达 因此，它们可能在精子发生中起重要作用。 精子发生调节机制的知识与以下方面有关： 制定可接受的避孕方法和治疗 男性不育症。

Sertoli cells play an important role in the bioregulation of spermatogenesis by providing mechanical support and chemical signals to spermatogenic cells. Work of this laboratory has shown that the intermediate filament protein, vimentin is phosphorylated in a FSH-dependent manner in cultured Sertoli cells. We now want to determine the mechanisms regulating the phosphorylation of vimentin and other cytoskeletal proteins of Sertoli cells during different stages of rat spermatogenesis. Two-dimensional polyacrylamide gel electrophoresis will be used to resolve [32P]orthophosphate and [Gamma-32P]ATP-labeled proteins following activation of cAMP- and Ca2+/calmodulin-dependent protein kinases. Preliminary studies have shown that Sertoli cells secrete specific proteins that are also spermatogenic stage-dependent. We propose the identification of factors that control the synthesis of these proteins during rat spermatogenesis. Seminiferous tubular segments corresponding to distinct spermatogenic stages identified by their transillumination pattern and histology will be labeled with [35S]methionine and secretory proteins resolved by gel electrophoresis. Sertoli cell-specific secretory proteins designated S80, S45 and S35 will be purified on DEAE-cellulose column chromatography and characterized. Antibodies against these proteins will be produced. The effect of growth factors on the regulation of the mitogenic and non-mitogenic function of Sertoli and peritubular cells will be evaluated using the complementation test of Stiles et al. and gel electrophoresis to define autocrine or paracrine effects. Factors involved in the modulation of Sertoli cell function induced by peritubular cells will be studied using a coculture method that allows cell-cell functional interaction but prevents cell-cell contact. We will use gel electrophoresis to determine the effect of peritubular cell-specific proteins, designated P95, P40 and P38, on the synthesis and secretion of [35S]methionine-labeled Sertoli cell-specific proteins, S80, S45 and S35. These Sertoli cell-specific secretory proteins are of interest because they have been shown to be expressed in a spermatogenic stage-dependent manner and, therefore, they may play an important role in spermatogenesis. Knowledge of the regulatory mechanisms of spermatogenesis are relevant to the development of acceptable methods of contraception and treatment of infertility of the male.