Effects of different doses of X-rays on cGAS-STING signaling pathway and tumor immune microenvironment
许明燕 1陈娴 1张华 1伊力亚尔·努尔如拉 1肖蕾 1李辰
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作者信息
1. 新疆医科大学第一附属医院肿瘤中心二科，乌鲁木齐 830054
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摘要
目的 研究不同剂量X射线照射对肝癌细胞免疫微环境及cGAS-STING信号通路的影响。 方法 C57BL/C小鼠右侧腋部皮下注射Hepa 1－6肝癌细胞，建立皮下成瘤肝癌模型，按顺序随机分为0、4、8、12 Gy照射组，每组10只，监测小鼠体重和肿瘤体积。照射后28 d收集标本，采用ELISA法和流式细胞术，比较各组肿瘤组织内巨噬细胞肿瘤坏死因子α(TNF-α)、干扰素γ(IFN-γ)、白介素6(IL-6)、趋化因子配体5(CCL5)、IL-10、IL-13、转化生长因子β(TGF-β)、IL-4和巨噬细胞M1、M2表型比例(M1/M2)的变化。采用实时荧光定量聚合酶链式反应(qRT-PCR)和免疫印迹实验检测肝癌细胞cGAS-STING信号通路相关基因与蛋白表达情况。 结果 随着照射剂量的增加，肿瘤体积明显减小(F＝8.42，P<0.05)，细胞坏死比例增加(F＝3.89，P<0.05)，除IL-4之外的巨噬细胞相关细胞因子含量增加(F＝6.32～15.50，P<0.05)，肝癌肿瘤免疫微环境中的M1、M2型巨噬细胞的比例升高(F＝5.46、5.14，P<0.05)。肝癌细胞内的cGAS-STING信号通路基因表达与蛋白表达水平升高(mRNA：F＝6.35、16.10，P<0.05；蛋白：F＝71.31、37.15，P<0.05)。 结论 X射线照射可激活肝癌细胞的cGAS-STING信号通路，促使肿瘤免疫微环境重塑。 Objective To study the effects of different doses of X-ray irradiation on the immune microenvironment and cGAS-STING signaling pathway of hepatocellular carcinoma cells. Methods C57BL/C mice were subcutaneously injected with Hepa 1-6 hepatocellular carcinoma cells in the right axilla to establish a subcutaneous tumor-forming hepatocellular carcinoma model. The mice were randomly divided into 0, 4, 8, 12 Gy irradiation groups, with 10 mice in each group. The body weights and tumor volumes were monitored. Specimens were collected 28 d after irradiation. The ELLSA and Flow Cytometry method was used to compare the macrophage-associated cytokines tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ), interleukin-6 (IL-6), chemokine ligand 5 (CCL5), IL-10, IL-13, transforming growth factor-β (TGF-β), IL-4 and macrophage M1, M2 phenotype ratio (M1/M2). Real-time fluorescence quantitative polymerase chain reaction (qRT-PCR) and immunoblotting assay were used to detect the expression of genes and proteins related to the cGAS-STING signaling pathway in hepatoma cells. Results With the increase of irradiation dose, the tumor volume was significantly reduced (F=8.42, P<0.05), the proportion of cell necrosis increased (F=3.89, P<0.05), the content of macrophage-associated cytokines other than IL-4 increased (F=6.32-15.50, P<0.05), and the proportion of M1 and M2 types of macrophage in the immune microenvironment of hepatocellular carcinoma tumors was elevated (F= 5.46, 5.14, P < 0.05).The gene expression and protein expression levels of cGAS-STING signaling pathway were elevated in hepatocellular carcinoma cells (mRNA expression of cGAS and STING: F=6.35, 16.10, P<0.05 protein expression of cGAS and STING:F=71.31, 37.15, P<0.05). Conclusions X-ray irradiation activates the cGAS-STING signaling pathway in hepatocellular carcinoma cells and contributes to the remodeling of the tumor immune microenvironment.
Abstract
Objective To study the effects of different doses of X-ray irradiation on the immune microenvironment and cGAS-STING signaling pathway of hepatocellular carcinoma cells. Methods C57BL/C mice were subcutaneously injected with Hepa 1-6 hepatocellular carcinoma cells in the right axilla to establish a subcutaneous tumor-forming hepatocellular carcinoma model. The mice were randomly divided into 0, 4, 8, 12 Gy irradiation groups, with 10 mice in each group. The body weights and tumor volumes were monitored. Specimens were collected 28 d after irradiation. The ELLSA and Flow Cytometry method was used to compare the macrophage-associated cytokines tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ), interleukin-6 (IL-6), chemokine ligand 5 (CCL5), IL-10, IL-13, transforming growth factor-β (TGF-β), IL-4 and macrophage M1, M2 phenotype ratio (M1/M2). Real-time fluorescence quantitative polymerase chain reaction (qRT-PCR) and immunoblotting assay were used to detect the expression of genes and proteins related to the cGAS-STING signaling pathway in hepatoma cells. Results With the increase of irradiation dose, the tumor volume was significantly reduced (F=8.42, P<0.05), the proportion of cell necrosis increased (F=3.89, P<0.05), the content of macrophage-associated cytokines other than IL-4 increased (F=6.32-15.50, P<0.05), and the proportion of M1 and M2 types of macrophage in the immune microenvironment of hepatocellular carcinoma tumors was elevated (F= 5.46, 5.14, P < 0.05).The gene expression and protein expression levels of cGAS-STING signaling pathway were elevated in hepatocellular carcinoma cells (mRNA expression of cGAS and STING: F=6.35, 16.10, P<0.05 protein expression of cGAS and STING:F=71.31, 37.15, P<0.05). Conclusions X-ray irradiation activates the cGAS-STING signaling pathway in hepatocellular carcinoma cells and contributes to the remodeling of the tumor immune microenvironment.
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