Khan S A, Teerds K, Dorrington J
Banting and Best Department of Medical Research, University of Toronto, Ontario, Canada.
Endocrinology. 1992 Feb;130(2):599-606. doi: 10.1210/endo.130.2.1733710.
A protein [steroidogenesis-inducing protein (SIP)] has been isolated from human ovarian follicular fluid and shown previously to stimulate steroidogenesis in Leydig cells, adrenal cells, and early luteal cells. Since proteins and peptides known to regulate steroidogenesis, such as gonadotropins and growth factors, also influence the growth of gonadal cells, the present study was designed to assess the effects of SIP on the synthesis of DNA by Leydig cells in vitro. Leydig cells were isolated from 10- and 20-day-old rats and cultured in serum-free medium for 48 h. The cells were then treated with the test materials for 18 h. Incorporation of [3H]thymidine into DNA was measured during the final 4 h of the culture. SIP significantly stimulated DNA synthesis in Leydig cells in a dose- and time-dependent manner, and the response to SIP was higher than that obtained with maximal concentrations of LH/hCG. The stimulatory effects of SIP were significantly enhanced when the cell cultures were preincubated in the presence of low levels of ovine LH (2 ng/ml). Cultures treated with SIP, followed by incubation with [3H]thymidine, contained 22 times as many labeled cells as control cultures, as assessed by autoradiography. The cells that were labeled were identified morphologically as Leydig cells. Insulin/insulin-like growth factor-I and/or transforming growth factor-alpha alone stimulated DNA synthesis and enhanced the effects of SIP on DNA synthesis. Dramatic changes in the morphology of cultured Leydig cells treated with SIP were observed; cells became flattened and developed extended projections which connected adjacent cells. LH/hCG, insulin, and transforming growth factor-alpha did not induce effects comparable to those of SIP on the morphology of Leydig cells. The effects of SIP on the synthesis of DNA and the morphology of Leydig cells were blocked in the presence of cycloheximide. It is concluded that SIP not only stimulates steroid production in Leydig cells, as shown previously, but also stimulates DNA synthesis and induces morphological changes in these cells. The latter properties of SIP combined with the magnitude of the responses elicited identify SIP as a unique gonadal protein.
一种蛋白质[类固醇生成诱导蛋白（SIP）]已从人卵巢卵泡液中分离出来，先前的研究表明它能刺激睾丸间质细胞、肾上腺细胞和早期黄体细胞的类固醇生成。由于已知调节类固醇生成的蛋白质和肽，如促性腺激素和生长因子，也会影响性腺细胞的生长，因此本研究旨在评估SIP对体外睾丸间质细胞DNA合成的影响。从10日龄和20日龄大鼠中分离出睾丸间质细胞，并在无血清培养基中培养48小时。然后用测试材料处理这些细胞18小时。在培养的最后4小时内测量[3H]胸腺嘧啶核苷掺入DNA的情况。SIP以剂量和时间依赖性方式显著刺激睾丸间质细胞中的DNA合成，并且对SIP的反应高于用最大浓度的LH/hCG获得的反应。当细胞培养物在低水平的羊LH（2 ng/ml）存在下预孵育时，SIP的刺激作用显著增强。通过放射自显影评估，用SIP处理后再与[3H]胸腺嘧啶核苷孵育的培养物中，标记细胞的数量是对照培养物的22倍。标记的细胞在形态上被鉴定为睾丸间质细胞。单独的胰岛素/胰岛素样生长因子-I和/或转化生长因子-α刺激DNA合成并增强SIP对DNA合成的作用。观察到用SIP处理的培养睾丸间质细胞的形态发生了显著变化；细胞变得扁平并形成连接相邻细胞的延伸突起。LH/hCG、胰岛素和转化生长因子-α对睾丸间质细胞形态的影响与SIP不同。在放线菌酮存在的情况下，SIP对DNA合成和睾丸间质细胞形态的影响被阻断。得出的结论是，如先前所示，SIP不仅刺激睾丸间质细胞中的类固醇生成，还刺激这些细胞中的DNA合成并诱导形态变化。SIP的后一种特性与所引发反应的程度相结合，将SIP确定为一种独特的性腺蛋白。
