Onoda M, Djakiew D
Department of Anatomy and Cell Biology, Georgetown University School of Medicine, Washington, DC 20007.
Mol Cell Endocrinol. 1990 Oct 1;73(1):35-44. doi: 10.1016/0303-7207(90)90042-7.
The influence of rat round spermatid protein(s) (RSP) on protein synthesis and secretory function of Sertoli cells was used in the bicameral chamber system. Round spermatids (RS) were purified from 90-day-old rats by centrifugal elutriation. RS were incubated in a supplement-enriched culture medium that lacked exogenous proteins. The RS-conditioned media were dialysed and lyophilized to obtain RSP. Most de novo protein synthesized under basal conditions by Sertoli cells (18-day-old) was secreted into the apical chamber (apical/basal ratio: 3.42). Follicle-stimulating hormone (FSH, 100 ng/ml) stimulated total protein secretion from Sertoli cells by a factor of 1.54. The RSP (100 micrograms/ml) stimulated total protein secretion from Sertoli cells by a factor of 2.33. The enhancement of total Sertoli cell protein secretion by FSH and RSP additively increased by a factor of 2.82. The combined effect of FSH and RSP on total protein secretion from Sertoli cells was dose dependent and saturated at approximately 200 micrograms/ml of RSP. Polarity of total protein secretion from Sertoli cells (apical/basal ratio: 3.42) was stimulated by RSP predominantly in the apical direction (apical/basal ratio: 8.48). The modulation of radiolabeled Sertoli cell secretory proteins (ceruloplasmin, CP; sulfated glycoprotein-2, SGP-2; testins and transferrin, Tf) by cold (non-labeled) RSP was investigated by immunoprecipitation followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The secretion of CP, SGP-2 and Tf was stimulated in a dose-dependent manner by the addition of RSP up to a saturating concentration of between 200 and 300 micrograms/ml, whereas the secretion of Sertoli cell testins did not reach saturation at 300 micrograms/ml RSP. These results indicate that FSH and RSP independently modulate Sertoli cell protein secretion, and that Sertoli cell secretory proteins may differentially respond to RSP stimulation.
在双室系统中研究了大鼠圆形精子细胞蛋白（RSP）对支持细胞蛋白质合成和分泌功能的影响。通过离心淘析从90日龄大鼠中纯化圆形精子细胞（RS）。将RS在缺乏外源蛋白的富含补充剂的培养基中孵育。对RS条件培养基进行透析和冻干以获得RSP。基础条件下支持细胞（18日龄）合成的大多数新生蛋白分泌到顶室中（顶室/基底室比例：3.42）。促卵泡激素（FSH，100 ng/ml）使支持细胞的总蛋白分泌增加了1.54倍。RSP（100μg/ml）使支持细胞的总蛋白分泌增加了2.33倍。FSH和RSP对支持细胞总蛋白分泌的增强作用相加后增加了2.82倍。FSH和RSP对支持细胞总蛋白分泌的联合作用呈剂量依赖性，在约200μg/ml的RSP时达到饱和。RSP主要在顶向方向刺激支持细胞总蛋白分泌的极性（顶室/基底室比例：8.48）（顶室/基底室比例：3.42）。通过免疫沉淀随后进行十二烷基硫酸钠-聚丙烯酰胺凝胶电泳，研究了冷（未标记）RSP对放射性标记的支持细胞分泌蛋白（铜蓝蛋白，CP；硫酸化糖蛋白-2，SGP-2；睾丸特异性蛋白和转铁蛋白，Tf）的调节作用。添加RSP直至饱和浓度在200至300μg/ml之间时，CP、SGP-2和Tf的分泌呈剂量依赖性受到刺激，而支持细胞睾丸特异性蛋白的分泌在300μg/ml RSP时未达到饱和。这些结果表明FSH和RSP独立调节支持细胞蛋白分泌，并且支持细胞分泌蛋白可能对RSP刺激有不同反应。
