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目的:探讨PD-L1(Programed Death Ligand-1)表达的调控以及其对Jurkat细胞增殖分化及凋亡的影响。方法:通过IFN-γ在体外诱导Jurkat细胞使PD-L1表达上调,并设计针对PD-L1的siRNA对Jurkat细胞进行转染抑制PD-L1的表达,通过RT-PCR、PCR测定Jurkat细胞分泌IL-2的变化;流式细胞术测定Jurkat细胞的凋亡情况。结果:运用β-actin进行半定量后,根据相应的模板量进行PCR,在设计的3条针对PD-L1的siRNA(siRNA-A,siRNA-B,siRNA-C)中选出一条可以高效抑制PD-L1表达的即siRNA-A,siRNA-A终浓度80nM转染Jurkat细胞24小时后PD-L1的表达明显抑制。使用IFN-γ终浓度2000U/ml作用于Jurkat细胞24小时后PD-L1表达明显增高,通过RT-PCR、PCR观察到PD-L1表达高低与Jurkat细胞分泌IL-2水平呈负相关,流式细胞术显示PD-L1表达增强能降低Jurkat细胞凋亡。结论:体外合成的针对PD-L1基因特异性siRNA,转染Jurkat细胞后可特异性抑制PD-L1的表达。IFN-γ在体外可刺激Jurkat细胞PD-L1表达增高。PD-L1信号在体外对Jurkat细胞分泌IL-2呈负相关,并且PD-L1能抑制Jurkat细胞凋亡。
Objective: To investigate the regulation of PD-L1 expression and its effect on proliferation, differentiation and apoptosis of Jurkat cells. Methods: Jurkat cells were induced by IFN-γ to up-regulate the expression of PD-L1. SiRNA targeting PD-L1 was transfected into Jurkat cells to inhibit PD-L1 expression. Jurkat cells were secreted by RT-PCR and PCR -2 changes; flow cytometry Jurkat cell apoptosis. Results: After semi-quantification with β-actin, PCR was performed according to the amount of the corresponding template, and one of the three siRNAs designed for PD-L1 (siRNA-A, siRNA-B and siRNA-C) PD-L1 expression of siRNA-A, siRNA-A PD-L1 expression was significantly inhibited after transfection of Jurkat cells with a final concentration of 80nM for 24 hours. The expression of PD-L1 in Jurkat cells was significantly increased after treated with 2000U / ml IFN-γ for 24 hours. The level of PD-L1 was negatively correlated with the level of IL-2 secreted by Jurkat cells by RT-PCR and PCR. Cytometry showed that enhanced PD-L1 expression can reduce Jurkat cell apoptosis. Conclusion: PD-L1 gene-specific siRNA synthesized in vitro can specifically inhibit PD-L1 expression after Jurkat cells transfection. IFN-γ stimulates PD-L1 expression in Jurkat cells in vitro. PD-L1 signal was negatively correlated with Jurkat cells secreting IL-2 in vitro, and PD-L1 could inhibit Jurkat cell apoptosis.