Richardson L L, Kleinman H K, Dym M
Department of Cell Biology, Georgetown University Medical Center, Washington, District of Columbia 20007, USA.
Biol Reprod. 1995 Feb;52(2):320-30. doi: 10.1095/biolreprod52.2.320.
Both Sertoli and myoid cells have been shown to be required for the appropriate deposition of basement membrane in the testis. We sought to define the pattern of basement membrane gene expression in Sertoli and peritubular myoid cells in vitro in order to begin to understand the regulatory mechanisms involved in basement membrane synthesis. Sertoli and myoid cells cultured alone or together were examined for synthesis of basement membrane components. Immunocytochemical localization demonstrated that Sertoli cells alone produced laminin and collagen IV, but not fibronectin, while myoid cells produced all three proteins. In Sertoli:myoid cocultures, a sequential deposition of the components into extracellular fibers was noted during 5 days of culture. Northern blot analysis revealed that mRNA levels for the laminin B1 chain and collagen IV increased from Days 3 to 5 in Sertoli cell monocultures. By contrast, the levels of laminin B1, collagen IV, heparan sulfate proteoglycan, and fibronectin decreased in the cocultures. Transcripts for the laminin A chain were not detected in the myoid cells; instead these cells produced the mRNA for the laminin homologue, merosin. This observation was confirmed by immunolocalization of merosin to the tunica propria of the testis and in cultured myoid cells. These data describe the expression of the basement membrane genes by Sertoli and peritubular myoid cells and provide the basis for future studies to determine the mechanisms that regulate the expression of the basement membrane genes in the testis.
支持细胞和肌样细胞已被证明是睾丸中基底膜正常沉积所必需的。我们试图确定支持细胞和睾丸周肌样细胞在体外基底膜基因表达的模式，以便开始了解参与基底膜合成的调控机制。对单独培养或共同培养的支持细胞和肌样细胞进行基底膜成分合成检测。免疫细胞化学定位显示，单独的支持细胞产生层粘连蛋白和IV型胶原，但不产生纤连蛋白，而肌样细胞产生所有这三种蛋白质。在支持细胞与肌样细胞的共培养中，在5天的培养过程中，观察到这些成分依次沉积到细胞外纤维中。Northern印迹分析显示，在支持细胞单培养中，层粘连蛋白B1链和IV型胶原的mRNA水平从第3天到第5天增加。相比之下，在共培养中，层粘连蛋白B1、IV型胶原、硫酸乙酰肝素蛋白聚糖和纤连蛋白的水平降低。在肌样细胞中未检测到层粘连蛋白A链的转录本；相反，这些细胞产生层粘连蛋白同系物merosin的mRNA。通过merosin在睾丸固有膜和培养的肌样细胞中的免疫定位证实了这一观察结果。这些数据描述了支持细胞和睾丸周肌样细胞基底膜基因的表达情况，并为今后研究确定睾丸中基底膜基因表达调控机制提供了基础。
