过表达CD59抑制补体对骨髓间充质干细胞的损伤作用

Overexpressed CD59 inhibits complement-induced damage to bone marrow mesenchymal stem cells

目的骨髓间充质干细胞(BMSCs)在移植过程中如何避免补体系统攻击的研究相对较少。文中主要探讨在自体移植过程中大鼠BMSCs(r BMSCs)表面CD59过表达对补体攻击r BMSCs的影响。方法分离培养SD大鼠BMSCs,电转化CD59过表达质粒后,分为r BMSCs空白组(r BMSCs不做任何处理),r BMSCs/CD59组(r BMSCs+CD59过表达质粒)和r BMSCs空载组(r BMSCs+空载质粒)。采用流式细胞仪检测转化后r BMSCs中CD59表达情况。抽取大鼠颈静脉血分离血清,采用自体血清处理DC59过表达后的r BMSCs,分为r BMSCs组(r BMSCs不与血清共孵育)、r BMSCs+ARS组[r BMSCs+大鼠自体血清(ARS)]、r BMSCs+i ARS组[r BMSCs+已灭活的大鼠自体血清(i ARS)]、r BMSCs/CD59+ARS组[r BMSCs/CD59+ARS]和r BMSCs/CD59+i ARS组。检测自体血清中补体对BMSCs的杀伤作用,补体形成情况以及细胞存活情况。结果与r BMSCs空白组细胞中CD59表达量比较,r BMSCs/CD59组明显升高(50.5%vs 74.9%,P<0.05)。与r BMSCs+ARS组细胞的凋亡率[(40.3±4.3)%]比较,r BMSCs组[(8.4±1.1)%]、r BMSCs/CD59+ARS组[(19.1±3.1)%]明显降低(P<0.05)。与r BMSCs+ARS组细胞中膜上补体膜攻击复合物(MAC)积累(99.7%)比较,r BMSCs组(50.1%)、r BMSCs/CD59+ARS组(71.0%)显著下降(P<0.05)。r BMSCs/CD59组各时期细胞凋亡率显著低于r BMSCs空白组(P<0.05)。结论 BMSCs CD59过表达可能通过减少补体攻膜复合物的形成,抑制自体移植过程中补体对r BMSCs的杀伤作用。

Objective Studies are rarely reported on how to avoid complement system attack during the transplantation of bone marrow mesenchymal stem cells( BMSCs). We explored the effect of the overexpression of CD59 on complement membrane attack-induced damage to rat BMSCs( r BMSCs) during autologous transplantation. Methods BMSCs from SD rats were cultured and treated with CD59 overexpression plasmid or r BMSC empty vector or left untreated,followed by detection of the expression of CD59 in the r BMSCs by flow cytometry. Then the r BMSCs were incubated with autologous rat serum( ARS),inactivated ARS( i ARS),CD59 + ARS,or CD59+i ARS or not incubated. The cytotoxicity of the serum complement on the r BMSCs was observed by PI staining and the apoptosis of the r BMSCs determined by flow cytometry. Results The expression of CD59 was significantly higher in the r BMSCs treated with CD59 than in those untreated( 74.9% vs 50.5%,P<0.05). The apoptosis rate was remarkably lower in the r BMSCs not incubated( [8.4±1.1]%) and those incubated with CD59+ARS( [19.1± 3.1]%)than in those incubated with ARS( [40.3±4.3]%)( P<0.05). The deposition of the complement membrane attack complex was significantly decreased in the r BMSCs not incubated( 50.1%) and those incubated with CD59+ARS( 71.0%) as compared with those incubated with ARS( 99.7%)( P<0.05). The apoptosis rate of the r BMSCs treated with CD59 was markedly lower than that of those left untreated( P<0.05). Conclusion The overexpression of CD59 inhibits the damage induced by complement to r BMSCs by reducing the formation of the complement membrane attack complex during autologous transplantation.