摘要
目的改进睾丸支持细胞(Sertoli 细胞)的分离方法及培养条件,探讨 Sertoli 细胞体外诱导异种淋巴细胞凋亡的作用机制。方法取2～4周龄的 SD 大鼠睾丸,采用 V 型胶原酶、胰蛋白酶及脱氧核糖核酸酶二步消化法制备 Sertoli 细胞。用免疫组织化学法(SABC 法)检测 Sertoli 细胞中Fas 配体(FasL)、转化生长因子β_1(TGF-β_1)、clusterin 的表达;流式细胞仪检测 Sertoli 细胞诱导异种淋巴细胞凋亡情况。结果在分离培养纯化过程中,Sertoli 细胞占培养细胞总数的90%以上,生长状况良好,能稳定表达 FasL、TGF-β_1和 clusterin,并且能够诱导异种淋巴细胞发生凋亡。结论应用本实验方法能够稳定获得大量高纯度的 Sertoli 细胞,并且能够发挥其免疫豁免的功能,从而用于细胞联合移植。
Objective To modify the isolation and culivation method of Sertoli cells and investigate its＇ effects on xeno-lymphocytes apeptesis. Methods Sertoli cells which was isolated from 2-4 week-old Sprague Dawley（SD） rats, were successfully prepared by collagenase type V ,trypsin and DNase I and then identified by electron microscope. Viability and apeptesis of cultured cells were measured by flow cytometry. The apeptesis rates of Balb/c mouse lymphocytes were examined which were co-cultured with Sertoli cells of SD rats by flow cytometry, too. The expression of FasL,TGF-β1 and clusterin on Sertoli cells were detected by immunocytochemistry. Results In the co-cultured system, Sertoli cells accounted for more than 90%. The viability of Sertoli cells was above 95% and the apeptesis rate was 10. 87% ± 3. 87% in this study. The lymphocytes apeptesis ratio was 15.52% ± 0. 17% （ P 〈 0. 01 ） . Streptavidin-biotin-peroxidase-complex immunochemistry staining showed that the Sertoli cells could express FasL, TGF-β1 and clusterin, respectively. Condusiom It indicates that the expression of FasL,TGF-β1 on the Sertoli cells might relate to the immune privilege, and it supposed to be benefit for xenotransplantation.
出处
《中华外科杂志》
CAS
CSCD
北大核心
2007年第5期331-334,共4页
Chinese Journal of Surgery
基金
国家高新技术发展研究计划("863"计划)基金(2003AA205009)
高等学校博士学科点专项基金(20040487077)
