【摘要】 目的:探讨麦冬皂苷B(OP-B)对甲状腺癌TPC-1细胞凋亡、增殖和免疫逃逸的影响及其与环状GMP-AMP合成酶(cGAS)-干扰素基因刺激蛋白(STING)信号通路的调节机制。方法:将TPC-1细胞分为TPC-1组(未做任何处理的TPC-1细胞)、L-OP-B组(2.5μmol/L OP-B处理TPC-1细胞)、M-OP-B组(5μmol/L OP-B处理TPC-1细胞)、H-OP-B组(10μmol/L OP-B处理TPC-1细胞)、RU.521组(100μmol/L cGAS-STING信号通路抑制剂RU.521处理TPC-1细胞)、H-OP-B+RU.521组(100μmol/L RU.521和10μmol/L OP-B共同处理TPC-1细胞)。MTT法检测OP-B对正常人甲状腺细胞系NTHY ORI 3-1细胞和TPC-1细胞的毒性；MTT法检测TPC-1细胞增殖；流式细胞术检测TPC-1细胞凋亡；将CD8~+ T细胞与各组处理的TPC-1细胞共培养，台盼蓝染色及流式细胞术检测CD8~+ T细胞活力；酶联免疫吸附(ELISA)检测上清液中γ干扰素(IFN-γ)、肿瘤坏死因子-α(TNF-α)水平；Western blot检测程序性死亡配体1(PD-L1)、cGAS-STING通路蛋白水平。结果:与TPC-1组比较，L-OP-B组、M-OP-B组、H-OP-B组OD值和PD-L1蛋白水平显著降低，CD8~+T细胞比例、CD8~+ T细胞存活率、TPC-1凋亡率、IFN-γ和TNF-α水平、cGAS和STING蛋白水平显著升高，且OP-B的作用效果呈现剂量相关性，而RU.521组以上指标呈现相反的趋势(均P<0.05)。与H-OP-B组比较，H-OP-B+RU.521组OD值、PD-L1蛋白水平显著升高，TPC-1凋亡率、CD8~+T细胞比例、CD8~+ T细胞存活率、IFN-γ和TNF-α水平、cGAS和STING蛋白水平显著下降(均P<0.05)。结论:OP-B可诱导TPC-1细胞凋亡，抑制细胞增殖和免疫逃逸，其机制可能与激活cGAS-STING信号通路相关。更多还原

【Abstract】 Objective:To investigate the effect of Ophiopogonin B(OP-B) on the apoptosis, proliferation and immune escape of thyroid cancer TPC-1 cells and the regulatory mechanism of cyclic GMP-AMP synthase(cGAS)-stimulator of interferon gene(STING) signaling pathway.Methods:TPC-1 cells were grouped into TPC-1 group(untreated TPC-1 cells),L-OP-B group(2.5 μmol/L OP-B treated TPC-1 cells),M-OP-B group(5 μmol/L OP-B treated TPC-1 cells),H-OP-B group(10 μmol/L OP-B treated TPC-1 cells),RU.521 group(100 μmol/L cGAS-STING signaling pathway inhibitor RU.521 treated TPC-1 cells),and H-OP-B+RU.521 group(100 μmol/L RU.521 and 10 μmol/L OP-B co-treated TPC-1 cells).MTT method was applied to detect the toxicity of OP-B on normal human thyroid cell line NTHY ORI 3-1 cells and TPC-1 cells.MTT method was applied to detect TPC-1 cell proliferation.Flow cytometry was applied to detect apoptosis of TPC-1 cells.CD8~+ T cells were co-cultured with TPC-1 cells in each group, detection of CD8~+T cell viability was performed by trypan blue staining and flow cytometry.ELISA method was applied to detect the levels of IFN-γ and TNF-α in the supernatant.Western blot was applied to detect programmed death ligand 1(PD-L1) and cGAS-STING pathway protein levels.Results:Compared with the TPC-1 group, the OD value and PD-L1 protein level in the L-OP-B group, M-OP-B group, and H-OP-B group were obviously reduced, the apoptosis rate of TPC-1 cells, proportion of CD8~+T cells, survival rate of CD8~+ T cells, the IFN-γ and TNF-α levels, cGAS and STING protein levels were obviously increased, and the effect of OP-B showed a dose-dependent relationship, while the RU.521 group showed an opposite trend in the above indicators(all P<0.05).Compared with the H-OP-B group, the OD value and protein level of PD-L1 in the H-OP-B+RU.521 group were obviously increased, the apoptosis rate of TPC-1 cells, proportion of CD8~+T cells, survival rate of CD8~+ T cells, the IFN-γ and TNF-α levels, cGAS and STING protein levels were obviously reduced(all P<0.05).Conclusion:OP-B can induce apoptosis of TPC-1 cells, inhibit cell proliferation and immune escape, and the mechanism may be related to the activation of cGAS-STING signaling pathway.更多还原