Scarpino S, Morena A R, Petersen C, Fröysa B, Söder O, Boitani C
Department of Histology and Medical Embryology, University of Rome La Sapienza, Italy.
Mol Cell Endocrinol. 1998 Nov 25;146(1-2):121-7. doi: 10.1016/s0303-7207(98)00190-7.
We have developed a rapid and convenient method of Sertoli cell preparation for studying the growth kinetics of these cells in in vitro culture. Datura Stramonium agglutinin (DSA)-coated dishes were used to rapidly purify single Sertoli cells from immature rat testis. We have monitored by immunohistochemical markers the degree of contamination of our Sertoli cell preparation by other cell types. The cell preparation is essentially free of germ cells and interstitial cells and contains a minimal percentage of myoid cells. Sertoli cells isolated with this method retain functional activities such as the FSH responsiveness in terms of cAMP production. In addition, we have studied the proliferative activity of Sertoli cells isolated by lectin binding from rats of different ages. Sertoli cells exhibited a characteristic pattern of proliferation which was a function of the donor animal age. The proliferative activity of isolated Sertoli cells decreased with age, being much higher in 3 day-old rats than in older animals. A similar pattern was observed when the mitotic activity of Sertoli cells in response to mitogens present in the testicular extracts from 5 day-old rats was evaluated. The method described here reduces or eliminates many of the drawbacks of the conventional procedures used to isolate Sertoli cells, thus providing a useful tool in studies of growth kinetics and regulation of cell proliferation in vitro.
我们开发了一种快速便捷的支持细胞制备方法，用于研究这些细胞在体外培养中的生长动力学。用曼陀罗凝集素（DSA）包被的培养皿从未成熟大鼠睾丸中快速纯化单个支持细胞。我们通过免疫组织化学标记物监测了我们制备的支持细胞被其他细胞类型污染的程度。该细胞制剂基本不含生殖细胞和间质细胞，且含有最低比例的肌样细胞。用这种方法分离的支持细胞保留了功能活性，如在产生环磷酸腺苷方面对促卵泡激素的反应性。此外，我们研究了通过凝集素结合从不同年龄大鼠中分离的支持细胞的增殖活性。支持细胞表现出一种特征性的增殖模式，这是供体动物年龄的函数。分离的支持细胞的增殖活性随年龄下降，在3日龄大鼠中比在年长动物中高得多。当评估支持细胞对5日龄大鼠睾丸提取物中存在的有丝分裂原的有丝分裂活性时，观察到了类似的模式。这里描述的方法减少或消除了用于分离支持细胞的传统程序的许多缺点，从而为体外生长动力学和细胞增殖调节的研究提供了一个有用的工具。
