RNA干扰SOCS1对线粒体M2蛋白作用树突状细胞功能的影响

Study on influence of RNA interference SOCS1 on mitochondrial M2 protein for dendritic cell function

目的探讨细胞因子信号转导抑制分子1(SOCS1)干扰后,线粒体M2蛋白对外周血单个核细胞(PB-MC)来源的树突状细胞(DC)功能的影响。方法诱导和培养健康人PBMC来源DC,小干扰RNA(siRNA)抑制SOCS1的表达,用不同浓度的M2蛋白刺激DC,用流式细胞术(FCM)分析DC表型CD83、CD86的表达,用酶联免疫吸附试验(ELISA)检测DC培养上清液中白细胞介素10(IL-10)和白细胞介素12(IL-12)的变化。结果DC在M2蛋白浓度为70μg/mL刺激24 h,35μg/mL刺激24、48和72 h时,与对照组比较,CD83和CD86的表达率以及IL-10和IL-12的水平均明显升高(P<0.05)。M2蛋白刺激SOCS1干扰后的DC,在不同刺激浓度和作用时间,CD83和CD86表达率以及IL-12水平与单纯M2蛋白刺激组相比均明显升高(P<0.05),而IL-10水平在两者之间差异无统计学意义(P>0.05)。结论 DC在接受高浓度的M2蛋白刺激后,其成熟度、抗原递呈和Th1的极化能力增强;抑制SOCS1的表达,M2蛋白可进一步促进DC功能的增强,可能导致自身耐受的破坏。

Objective To study the influence of mitochondrial M2 protein on peripheral blood mononuclear cell （ PBMC ） -derived dendritic cells （DC） after silencing suppressor of cytokine signaling 1 （ SOCS1 ）. Methods DC from PBMC of healthy subjects were induced and cultured. The SOCS1 expression was silenced by small interfering RNA （siRNA）. DC were stimulated by various concentrations of M2 protein, and the expressions of CD83 and CD86 of DC were performed by flow cytometry （ FCM ）. The levels of interleukin-lO （ IL-10 ） and interleukin-12 （ IL-12 ） in culture supernatant of DC were measured by enzyme-linked immunosorbent assay （ELISA）. Results The expressions of CD83 and CD86 and the levels of IL-10 and IL-12 of DC under the stimulation of M2 protein at 70 μg/mL after 24 h and at 35 μg/mL after 24, 48 and 72 h were all significantly higher than those in the control group（P 〈0.05）. After silencing SOCS1 by siRNA of DC with various concentrations of M2 protein at different times, the expressions of CD83 and CD86 and the levels of IL-12 all increased more significantly than those in the M2 protein stimulating group （P 〈 0.05 ）. However, there was no significant difference for IL-10 levels of DC between the 2 groups （P 〉 0.05）. Conclusions DC after stimulation of M2 protein at high concentration has increased the capacity to activate Thl subset proliferation, maturation and antigen presentation. The functions are further enhanced after silencing SOCS1 of DC in the presence of M2 protein, which maybe contribute to the break of self-tolerance.