卡介苗感染人树突状细胞对miRNA-99b表达和Th17/Treg分化的影响

Influence of human dendritic cells with Bacillus Calmette Guerin infection on expression of miRNA-99b and differentiation of Th17/Treg

目的 探讨牛型分枝杆菌减毒活菌—卡介苗(BCG)感染对人树突状细胞(DCs)microRNA-99b(miRNA-99b)表达的影响及对新生儿脐血CD4+初始T淋巴细胞分化为Th17/Treg的作用。方法 BCG感染体外培养的人树突状细胞,然后与新生儿脐血CD4+初始T淋巴细胞混合培养,qRT-PCR观察人树突状细胞miRNA-99b的表达变化,qRT-PCR检测CD4+T淋巴细胞Foxp3、ROR-γt、γ干扰素(IFN-γ)、白细胞介素-10(IL-10)基因转录水平,观察CD4+T淋巴细胞分化亚型的改变;ELISA法检测培养上清INF-γ、IL-17和IL-23表达水平。结果 BCG感染后,人树突状细胞miRNA-99b基因表达增加(P<0.001);CD4+T淋巴细胞IFN-γ、IL-10基因表达增加(P<0.05),ROR-γt基因表达降低(P<0.05),Foxp3基因转录增加,但差异无统计学意义;感染组培养上清中INF-γ表达水平增加(P<0.001),IL-17和IL-23表达水平均有一定的增加,但差异无统计学意义。结论 结核感染可能通过影响树突状细胞miRNA-99b的表达进而诱导初始CD4+T淋巴细胞分化为Th17/Treg的失衡参与结核病的发病过程。

OBJECTIVE To investigate the effects of Bacillus Calmette Guerin vaccine(BCG)infection on microRNA-99 b(miRNA-99 b)expression in human dendritic cells and the differentiation of CD4+naive T cells in neonate cord blood.METHODS After infected by BCG,human dendritic cells cultured in vitro were mixed cultured with CD4+naive T cells of neonate cord blood.The change in expression of miRNA-99 b in human dendritic cells was observed by qRT-PCR.The transcriptional levels of Foxp3,ROR-γt,IFN-γand IL-10 genes of CD4+T cells were determined by qRT-PCR to observe the change in differentiation subtypes of T lymphocytes.The INF-γ,IL-17 and IL-23 levels in culture supernatant were analyzed by ELISA.RESULTS After infected by BCG,the miRNA-99 b expression was dramatically improved in human dendritic cells(P<0.001),the IFN-γand IL-10 gene expression levels of CD4+T cells significantly increased(P<0.05),ROR-γt gene expression of CD4+T cells significantly decreased(P<0.05).INF-γlevel increased significantly in culture supernatant(P<0.001).IL-17 and IL-23 expression levels also increased in culture supernatant,but the differences were not significant.CONCLUSION Tuberculosis infection may be involved in the pathogenesis of tuberculosis through controlling miRNA-99 b expression in human dendritic cells and inducing Th17/Treg differentiation imbalance.