实验性精索静脉曲张对青春期大鼠睾丸、附睾中CRES蛋白表达的影响

Effects of Experimental Varicocele on CRES Protein in the Testis and Epididymis of Adolescent Rats

目的：研究青春期大鼠实验性左侧精索静脉曲张（ELV）对睾丸和附睾中胱蛋白酶抑制剂相关的附睾精子发生（CRES）蛋白表达的影响，探索精索静脉曲张导致不育的机制。方法：建立青春期雄性SD大鼠ELV模型，采用免疫组化及蛋白印迹方法检测CRES蛋白在ELV2周组（n=8）、4周组（n=8）及其相应的对照组（n均=5）大鼠睾丸和附睾头、体、尾中的表达变化。结果：免疫组化显示，CRES蛋白在ELV实验组和对照组大鼠睾丸和附睾中均有表达。在睾丸中，CRES蛋白主要定位于圆形和长形精子细胞的胞质、精子的顶体以及残余体中，其表达与生精周期密切相关，I～Ⅲ和Ⅸ～XⅣ期表达最强，Ⅶ～Ⅷ期表达次之，Ⅳ～Ⅵ期表达减弱。在附睾中，CRES蛋白主要表达于从附睾起始段到尾部的上皮主细胞的胞质中，且以体、尾部表达最强，头部次之，管腔分泌物也呈阳性表达。实验组比对照组CRES蛋白表达增强。Western印迹显示：实验组和对照组大鼠睾丸和附睾在相对分子质量（Mr）为19000和14000处均可见CRES蛋白条带，其中以M．19000处表达更为明显。实验组CRES蛋白的表达比对照组明显增强。对免疫组化和Western印迹结果进行灰度值和积分吸光度值（IA）测定，并经统计学分析显示：ELV2周组和4周组比其相对应的对照组表达增强且差异有显著性（P〈0．05或P〈0．01），而ELV各组问未见CRES蛋白表达有明显差异（P〉0．05）。结论：CRES蛋白在大鼠睾丸和附睾中均有表达，睾丸中表达呈现生精周期特异性和细胞特异性，附睾中表达呈现附睾上皮区段和细胞特异性；CRES蛋白在青春期ELV大鼠睾丸和附睾中的表达比对照组明显增强。这些结果提示CRES蛋白在精子发生和精子成熟过程中可能起着重要的作用，并且可能与ELV引起的不育或生育力低下有关。

Objective： To investigate the effects of experimental left varicocele （ELV） on the cystatin-related epididymal spermatogenic （CRES） protein in the testis and epididymis of adolescent rats. Methods ： The ELV model of Sprague-Dawley （SD） male adolescent rats was established, and the expression of CRES protein in the testis and epididymis was detected by immunohistochemistry and Western-blot at 2 and 4 weeks after surgery. Results ： Immunohistochemistry and Western-blot detected CRES protein in both the testis and the epididymis of the ELV rats and the control rats. Immunohistochemistry showed that within the testis, CRES protein was expressed mainly in the cytoplasm of round spermatids and elongating spermatids, sperm acrosomes and residual bodies. The expression was most intensive at Stages Ⅰ～Ⅲ and Ⅸ -ⅩⅣ, and then decreased gradually at Stages Ⅶ - Ⅷ and Ⅳ -Ⅵ. Within the epididymis, CRES protein was expressed mainly in the cytoplasm of the principal cells of epididymal epithelia. Western-blot detected CRES protein in Mr 19 000 and 14 000, stronger in the former than in the latter. Image and statistical analyses showed that the expression of CRES protein in the 2-week and 4-week ELV groups was significantly higher than in the control group（ P 〈 0.05, or P 〈 0.01 ）. Conclusion： CRES protein expressed in both the testis and epididymis of adolescent rats and the expression is stage-specific and cellspecific in the testis and segment-specific and cell-specific in the epididymis. The expression of CRES protein in the ELV rats is much stronger than in their corresponding controls. It is suggested that CRES protein may be significantly involved in the regulation of spermatogenesis and sperm maturation, and possibly associated with varicocele-related male infertility or subfertility.