Depuydt C E, Mahmoud A M, Dhooge W S, Schoonjans F A, Comhaire F H
Department of Internal Medicine, University Hospital Ghent, Belgium.
J Androl. 1999 Jan-Feb;20(1):54-62.
The influences of follicle-stimulating hormone (FSH), gonadal steroids, and culture time were studied in relation to inhibin B production by Sertoli cells of immature rats cultured in vitro. Sertoli cell-enriched cultures were established from 18-day-old rats and were maintained in medium supplemented with insulin, transferrin, and epidermal growth factor at 34 degrees C. A recently developed ELISA for the measurement of inhibin B was used to assess the effects of recombinant human FSH (rh FSH), testosterone (T), and estradiol (E2) on inhibin B production and accumulation in the culture media of Sertoli cell-enriched cultures and to optimize the cell culture system to serve as a bioassay for the detection and quantification of estrogens and estrogenlike substances. Prolonging the incubation time (24, 48, or 72 hours) of Sertoli cells with control medium without rh FSH, T, or E2 resulted in a time-dependent increase of inhibin B production. Incubation with rh FSH (1, 2.5, 5, or 10 U/L) caused a dose- and time-dependent increase of inhibin B production by Sertoli cells (but not by cultured Leydig cells), reaching a plateau at 5 U/L rh FSH. Addition of T in concentrations of 2.88, 5, or 50 ng/ml to medium without rh FSH and E2 significantly lowered the daily production rate of inhibin B (P < 0.05). In contrast, addition of E2 (0.01 and 0.1 ng/ml) caused a dose-responsive increase in inhibin B production after 24 and 48 hours. The relative increment of inhibin B production induced by E2 was maximal after 24 hours in the presence of 2.5 U/L rh FSH (acting synergistically) and in the absence of T. When these conditions are implemented, the Sertoli cell culture system may serve as a bioassay for estrogenic substances, and it may reflect the possibly harmful effect they may have on spermatogenesis.
研究了促卵泡激素（FSH）、性腺类固醇和培养时间对体外培养的未成熟大鼠支持细胞抑制素B产生的影响。从18日龄大鼠建立富含支持细胞的培养物，并在补充有胰岛素、转铁蛋白和表皮生长因子的培养基中于34℃下维持培养。使用最近开发的用于测量抑制素B的酶联免疫吸附测定法（ELISA）来评估重组人FSH（rh FSH）、睾酮（T）和雌二醇（E2）对富含支持细胞培养物的培养基中抑制素B产生和积累的影响，并优化细胞培养系统，以用作检测和定量雌激素及雌激素样物质的生物测定法。在不含rh FSH、T或E2的对照培养基中延长支持细胞的孵育时间（24、48或72小时）导致抑制素B产生呈时间依赖性增加。用rh FSH（1、2.5、5或10 U/L）孵育导致支持细胞（而非培养的睾丸间质细胞）产生抑制素B呈剂量和时间依赖性增加，在5 U/L rh FSH时达到平台期。在不含rh FSH和E2的培养基中添加浓度为2.88、5或50 ng/ml的T显著降低了抑制素B的日产生率（P<0.05）。相反，添加E2（0.01和0.1 ng/ml）在24和48小时后导致抑制素B产生呈剂量反应性增加。在存在2.5 U/L rh FSH（协同作用）且不存在T的情况下，E2诱导的抑制素B产生的相对增量在24小时后最大。当实施这些条件时，支持细胞培养系统可作为雌激素物质的生物测定法，并且它可能反映它们对精子发生可能产生的有害影响。
