Hoeben E, Briers T, Vanderstichele H, De Smet W, Heyns W, Deboel L, Vanderhoydonck F, Verhoeven G
Laboratory for Experimental Medicine and Endocrinology, Onderwijs en Navorsing, Gasthuisberg, Catholic University of Leuven, Belgium.
Endocrinology. 1995 Jul;136(7):2862-73. doi: 10.1210/endo.136.7.7789311.
Testicular peritubular and prostatic stromal cells produce extracellular matrix elements and paracrine factors that modulate the cytodifferentiation and function of the corresponding epithelial cells. The present paper describes the establishment and characterization of five rat testicular cell lines with peritubular characteristics and one prostatic stromal cell line. Four peritubular cell lines were isolated after transfection of a mixed peritubular-Sertoli cell culture with a v-myc-containing plasmid. The same immortalization procedure applied to prostatic stromal cells yielded one cell line. An additional testicular cell line arose by spontaneous immortalization during serial subculture. Except for one testicular cell line (RTC-8T1), the morphology of all of the immortalized cell lines strongly resembled that of primary cultures of peritubular and stromal cells. Flow cytometric analysis demonstrated that all cell lines scored positive for alpha-smooth muscle isoactin and negative for cytokeratins, confirming their myofibroblast-like nature. None of the cell lines, however, stained positive for alkaline phosphatase, and androgen receptor expression was also lost. Typical Leydig cell characteristics, such as steroidogenesis, and Sertoli cell markers, such as transferrin secretion, were absent. Coculture of the cell lines with Sertoli cells resulted in the formation of tubular structures. A cell attachment assay and an enzyme-linked immunosorbent assay for fibronectin confirmed the production of extracellular matrix elements by all of the established cell lines. Media conditioned by the cell lines stimulated Sertoli cell transferrin production. The active principle was partially purified and resembled the P-MOD-S-like factors produced by primary cultures of peritubular and stromal cells. It is concluded that the immortalized cell lines have retained several of the characteristics of primary cultures of peritubular and stromal cells and may be useful for further studies on mesenchymal-epithelial interactions in testis and prostate.
睾丸生精小管和前列腺基质细胞可产生细胞外基质成分和旁分泌因子，这些物质可调节相应上皮细胞的细胞分化和功能。本文描述了五种具有生精小管特征的大鼠睾丸细胞系和一种前列腺基质细胞系的建立及特性。在将含有v-myc的质粒转染至混合的生精小管-支持细胞培养物后，分离出了四种生精小管细胞系。将相同的永生化程序应用于前列腺基质细胞，得到了一种细胞系。另外一种睾丸细胞系是在连续传代培养过程中自发永生化产生的。除了一种睾丸细胞系（RTC-8T1）外，所有永生化细胞系的形态都与原代培养的生精小管和基质细胞非常相似。流式细胞术分析表明，所有细胞系的α-平滑肌肌动蛋白呈阳性，细胞角蛋白呈阴性，证实了它们类似肌成纤维细胞的性质。然而，所有细胞系的碱性磷酸酶染色均为阴性，雄激素受体表达也丧失了。典型的睾丸间质细胞特征，如类固醇生成，以及支持细胞标志物，如转铁蛋白分泌，均不存在。将这些细胞系与支持细胞共培养可导致管状结构的形成。细胞黏附试验和针对纤连蛋白的酶联免疫吸附试验证实了所有已建立的细胞系均可产生细胞外基质成分。这些细胞系的条件培养基可刺激支持细胞转铁蛋白的产生。活性成分经过部分纯化，类似于原代培养的生精小管和基质细胞产生的P-MOD-S样因子。结论是，永生化细胞系保留了原代培养的生精小管和基质细胞的一些特征，可能有助于进一步研究睾丸和前列腺中的间充质-上皮相互作用。
