EFHD2蛋白的缺失对小鼠Sertoli细胞紧密连接蛋白的影响  

Effects of EFHD2 protein deletion on tight junction proteins in mouse Sertoli cells

作　　者：叶晓林 胥国麟 钱体军 秦凤 王云涛 程煜航 赵文珍 Ye Xiaolin;Xu Guolin;Qian Tijun;Qin Feng;Wang Yuntao;Cheng Yuhang;Zhao Wenzhen(Dept of Histology and Embryology,School of Basic Medical Sciences,Dali University,Dali 671000;Institute of Pathogens and Vectors,School of Basic Medical Sciences,Dali University,Dali 671000;Grade 2018 Clinical Medicine Major of Dali University,Dali 671000)

机构地区：[1]大理大学基础医学院组织学与胚胎学教研室,大理671000 [2]大理大学基础医学院病原与媒介生物研究所,大理671000 [3]大理大学,大理671000

出　　处：《安徽医科大学学报》2022年第11期1744-1749,共6页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金(编号:82160280);云南省科技厅科技计划项目(编号:202101BA070001-041、202101BA070001-108、202001BA070001-084)。

摘　　要：目的研究EF-手域蛋白D2(EFHD2)在小鼠睾丸内的定位及表达,及EFHD2蛋白在Sertoli细胞中的缺失对紧密连接蛋白组分Occludin的影响。方法取成年小鼠心、肝、脾、肺、肾、脑及睾丸组织提取总RNA和蛋白质,运用qRT-PCR和Western blot检测EFHD2在各器官组织中mRNA和蛋白质水平的差异性;运用免疫荧光和免疫组化技术,检测EFHD2在睾丸组织中的定位和表达。运用siRNA干扰技术降低Sertoli细胞中EFHD2来检测Occludin蛋白的表达。结果qRT-PCR显示EFHD2在睾丸中表达量最高;Western blot结果显示在睾丸组织中表达水平较高;间接免疫荧光和免疫组化结果显示该蛋白主要分布在Sertoli细胞内,与细胞骨架波形蛋白具有共定位,即明确该蛋白表达在Sertoli细胞。EFHD2蛋白表达降低后,Occludin蛋白表达也减少。结论EFHD2蛋白在睾丸组织中表达量相对较高,主要分布于睾丸Sertoli细胞中,与波形蛋白共定位,且能影响紧密连接蛋白Occludin的正常表达。提示EFHD2能促进和维持Sertoli细胞的细胞连接结构,为精子发生提供一个稳定的微环境。Objective To study the effect of EFHD2 protein deletion in Sertoli cells on Occludin,a component of tight junction protein and the localization and expression of EF-hand domain family member D2(EFHD2)in mouse testis.Methods Total RNA and protein were extracted from adult mice s heart,liver,spleen,lung,kidney,brain and testis tissues.The mRNA and protein levels of EFHD2 in each organ tissue were detected by qRT-PCR and Western blot.Immunofluorescence and immunohistochemistry detected the localization and expression of EFHD2 in testicular tissues.SiRNA interference was used to reduce EFHD2 in Sertoli cells to detect Occludin protein expression.Results qRT-PCR showed that the expression of EFHD2 was the highest in the testis.Western blot results showed that the expression level increased in testis tissue.Indirect immunofluorescence and immunohistochemistry results showed that the protein was mainly distributed in Sertoli cells and co-localized with cytoskeletal Vimentin,indicating that the protein was expressed in Sertoli cells.After the decrease of EFHD2 protein expression,Occludin protein expression also decreased.Conclusion The expression of EFHD2 protein in the testis is relatively high,mainly distributed in Sertoli cells of the testis,co-localized with Vimentin,and can affect the normal expression of tight junction protein Occludin.It is suggested that EFHD2 can promote and maintain the junction structure of Sertoli cells and provide a stable microenvironment for spermatogenesis.

关 键 词：EF-手域蛋白D2 睾丸 SERTOLI细胞 OCCLUDIN蛋白 波形蛋白 精子发生 

分 类 号：R321.1[医药卫生—人体解剖和组织胚胎学]