Welsh M J, Wu W, Parvinen M, Gilmont R R
Department of Anatomy and Cell Biology, University of Michigan Medical School, Ann Arbor 48109, USA.
Biol Reprod. 1996 Jul;55(1):141-51. doi: 10.1095/biolreprod55.1.141.
The purpose of these studies was to define expression of hsp27 mRNA during the cycle of the seminiferous epithelium and to determine the distribution of hsp27 protein in the rat testis. To study hsp27 mRNA expression, a rat hsp27 cDNA was isolated and sequenced (GenBank no. M86389). The cDNA was used in Northern blot analysis to estimate the relative levels of hsp27 mRNA in rat seminiferous tubule segments selected for different stages of the cycle of the seminiferous epithelium. The level of hsp27 mRNA was low during stages IX-XII of the cycle of the seminiferous epithelium. Approximately 15-fold higher levels of hsp27 mRNA were expressed during stages II-VI, with intermediate levels being expressed during stages XIII-I and VII-VIII. No effect of FSH on hsp27 mRNA expression was detected in cultured Sertoli cells, suggesting that hsp27 synthesis in Sertoli cells is not directly regulated by FSH. The distribution of hsp27 was also studied by use of immunofluorescence in frozen sections of rat testis, in isolated seminiferous tubules, in primary cultures of Sertoli cells isolated from 19-26-day-old rats, in peritubular myoid cells from 26-day-old rats, and in several cell lines. Microfilaments were localized in similar preparations by using rhodamine-phalloidin or BODIPY-phallicidin (Molecular Probes, Eugene, OR). The hsp27 was co-localized with micro-filaments in Sertoli cells from 20-day-old and older rats, but not in Sertoli cells from younger rats. In other cell types, hsp27 was diffusely distributed throughout the cytoplasm. These results demonstrate that hsp27 expression varies with the cycle of the seminiferous epithelium and provide the first direct morphological evidence that hsp27 is associated with microfilaments in a normal, intact tissue. They also suggest that Sertoli cell micro-filaments, by virtue of their associated hsp27, may be different in composition and function from microfilaments of peritubular myoid cells and many other cell types.
这些研究的目的是确定生精上皮周期中hsp27 mRNA的表达情况，并确定hsp27蛋白在大鼠睾丸中的分布。为了研究hsp27 mRNA的表达，分离并测序了大鼠hsp27 cDNA（GenBank编号：M86389）。该cDNA用于Northern印迹分析，以估计在为生精上皮周期不同阶段选择的大鼠生精小管片段中hsp27 mRNA的相对水平。在生精上皮周期的IX-XII阶段，hsp27 mRNA水平较低。在II-VI阶段，hsp27 mRNA的表达水平大约高15倍，在XIII-I阶段和VII-VIII阶段表达水平处于中间值。在培养的支持细胞中未检测到FSH对hsp27 mRNA表达的影响，这表明支持细胞中hsp27的合成不受FSH的直接调节。还通过免疫荧光法研究了hsp27在大鼠睾丸冰冻切片、分离的生精小管、从19-26日龄大鼠分离的支持细胞原代培养物、26日龄大鼠的睾丸肌样细胞以及几种细胞系中的分布。通过使用罗丹明-鬼笔环肽或BODIPY-鬼笔环肽（Molecular Probes，俄勒冈州尤金市）在类似的标本中定位微丝。在20日龄及以上大鼠的支持细胞中，hsp27与微丝共定位，但在较年幼大鼠的支持细胞中未共定位。在其他细胞类型中，hsp27弥漫分布于整个细胞质中。这些结果表明，hsp27的表达随生精上皮周期而变化，并提供了首个直接的形态学证据，证明hsp27在正常完整组织中与微丝相关。它们还表明，支持细胞的微丝由于其相关的hsp27，在组成和功能上可能与睾丸肌样细胞和许多其他细胞类型的微丝不同。
