甘露聚糖结合凝集素对白假丝酵母菌感染鼠体内Treg/Th17细胞免疫平衡的调节作用

Mannan-binding lectin regulates Treg/Thl7 cell balance in mice with Candida albicans infection

目的研究甘露聚糖结合凝集素(mannan-binding lectin,MBL)对白假丝酵母菌感染鼠体内Th17、Treg细胞分化的调节。方法MBL基因敲除型(MBL^-/-)和野生型(WT)C57BL/6小鼠分4组,2×10^7CFU白假丝酵母菌(C.albicans)对感染组小鼠腹腔注射;感染3d后取小鼠肝脏和肾脏病理切片,HE染色和PAS染色观察小鼠病理组织学变化;7d后摘除小鼠眼球取静脉血流式细胞术分析小鼠体内Treg和Th17细胞;静脉抗凝血ELISA检测小鼠体内细胞因子IL-10和IL-17A的含量;颈椎脱臼处死小鼠后取脾脏细胞MACS磁性分选获得CD4^+T细胞,提取总RNA,qRT-PCR检测其体内Foxp3、RORγtmRNA的表达;取小鼠脾脏CD4^+T细胞,提取总蛋白,Western blot检测Foxp3、RORγt蛋白表达水平。结果成功建立了C.albicans感染鼠模型,和WT感染组相比较:流式分析显示MBL^-/-感染组小鼠体内Th17细胞亚型数目比例增高,Treg细胞亚型数目比例降低;ELISA结果显示MBL^-/-感染组小鼠体内IL-17A水平明显增高,IL-10水平降低;qRT-PCR结果显示MBL^-/-感染组小鼠体内转录因子RORγtmRNA表达升高,转录因子Foxp3mRNA表达降低;Western blot印迹结果表明MBL^-/-感染组小鼠体内RORγt蛋白表达量明显增多,而Foxp3蛋白表达量减少。结论在C.albicans感染中,MBL促进了CD4^+T细胞向Treg细胞亚型诱导分化,抑制了CD4^+T细胞向Th17细胞亚型的诱导分化,从而调节了C.albicans感染鼠体内Treg/Th17细胞的免疫平衡。

Objective To investigate the regulation of Treg/Thl7 axis by mannan-binding lectin (MBL) in mice with Candida albicans ( C. albicans) infection. Methods MBL gene-knockdown (MBL^-/-) C57BL/6 mice and wild-type ( WT) C57BL/6 mice were divided into four groups. C. albicans strains (2×10^7 CFU) were injected intraperitoneally into the mice of infection groups. Paraffin sections of mouse liver and kidney tissues were prepared on 3 d. Histopathological changes were observed with hematoxylin and eosin ( HE) and Periodic acid-Schiff ( PAS) staining. Flow cytometry was performed to analyze Th 17 and Treg cells in mice on 7 d. The levels of IL-10 and 1L-17A were measured by ELISA. CD4^+ T cells were obtained from spleen cells by magnetic sorting. Expression of Foxp3 and RORγi at mRNA and protein levels were detected by qRT-PCR and Western blot, respectively. Results The mouse model of C. albicans infection was established successfully. After infection, the MBL^-/- mice had higher percentages of Th 17 cells, but lower percentages of Freg cells than the WT mice. ELISA results showed that com pined with the WT mice with C. albicans infection, the MBL^-/- mice had significantly increased IL-17A and decreased IL-10 after infection. Moreover, the expression of R0Rγt at holh mRNA and protein levels was up-regulated ,while that of Foxp3 was down-regulated in the MBL^-/- mice than in the WT mice following infection. Conclusions MBL regulates the immune balance of Treg/Thl7 cells in mice infected with C. albicans through promoting the differenticition of CD4^+ T cells into Treg cell subsets and inhibiting the differentiation into Th 17 cell subsets.