【摘要】 目的探讨程序性死亡因子配体1(PD-L1)和PD-L2在人胎盘源性间充质干细胞(hPMSCs)上表达对外周血T细胞分泌IL-17的影响。方法应用酶消化法分离人胎盘间充质干细胞(hPMSCs),并进行表型及分化鉴定;RT-PCR、激光扫描共聚焦显微镜技术(LSCM)及流式细胞术(FCM)检测hPMSCs上PD-L1及PD-L2的表达;应用化学合成的PD-L1 siRNA、PD-L2 siR-NA沉默hPMSCs上PD-L1及PD-L2表达;密度梯度离心法分离纯化人外周血T细胞;细胞胞内因子染色法分析沉默PD-L1或PD-L2后hPMSCs对PMA活化的T细胞分泌IL-17的影响。结果除PD-L1外,hPMSCs高表达PD-L2;siRNA能够有效阻断PD-L1和PD-L2在hPMSCs上的表达;胞内染色结果显示,hPMSCs能够上调T细胞IL-17的分泌,阻断PD-L1或PD-L2后,T细胞IL-17的分泌被进一步上调,且PD-L1和PD-L2具有叠加作用。结论 PD-L1和PD-L2在hPMSCs上表达,可拮抗hPMSCs刺激外周血T细胞分泌IL-17的作用。更多还原

【Abstract】 Objective To investigate the expressions of programmed death ligand 1(PD-L1) and PD-L2 in human placenta-derived mesenchymal stem cells(hPMSCs) and their mediated immunoregulation on the secretion of IL-17 of peripheral blood T cells.Methods hPMSCs were isolated from mature human placenta by the method of enzyme digestion.The cells were cultured and expanded in vitro,and after the third passage,they were used in phenotyping and differentiation experiments.The expressions of PD-L1 and PD-L2 were detected by RT-PCR,laser-scanning confocal microscopy(LSCM) and flow cytometry.Specific siRNAs were transfected into hPMSCs via cathodolyte liposome transfection method to silence the expressions of PD-L1 and PD-L2.T cells were sorted from healthy peripheral blood by gradient centrifugation.IL-17 secretion of T cells activated by PMA was detected by intracellular staining after the expressions of PD-L1 and PD-L2 were silenced.Results Besides PD-L1,hPMSCs also highly expressed PD-L2,which could be silenced effectively by specific siRNA.Intracellular staining showed that hPMSCs up-regulated the secretion of IL-17,which was further up-regulated after PD-L1 or/and PD-L2 had been silenced and there were overlapping roles of PD-L1 and PD-L2 in inhibiting IL-17 secretion by T cells.Conclusion PD-L1 and PD-L2 expressed on hPMSCs could inhibit the hPMSCs-mediated up-regulation on the expression of IL-17 secreted by peripheral blood T cells.更多还原