Notch信号通路对慢性丙型肝炎患者CD8^+T淋巴细胞功能的影响  

Influence of the Notch signaling pathway on CD8^+T lymphocyte function in patients with chronic hepatitis C

作　　者：李彧[1] 张鸿[1] 李竹[1] 刘清娥[1] 王璐 张丹[1] 曹宁家[1] 苏莉[2] 

机构地区：[1]陕西省人民医院感染性疾病科,西安710068 [2]兰州大学公共卫生学院,兰州730000

出　　处：《临床肝胆病杂志》2018年第2期287-291,共5页Journal of Clinical Hepatology

基　　金：陕西省科学技术研究发展计划项目(2012SF2-01)

摘　　要：目的观察Notch信号通路对慢性丙型肝炎患者CD8^+T淋巴细胞功能的影响,初步阐释Notch信号通路在慢性丙型肝炎发病中的作用。方法收集2016年1月-8月在陕西省人民医院就诊的慢性丙型肝炎初治患者38例和健康志愿者17例,分选CD8^+T淋巴细胞,应用实时定量PCR法检测CD8^+T淋巴细胞中Notch1和Notch2 mRNA的表达水平。分别应用直接接触培养系统和间接接触培养系统将从HLA-A2限制性丙型肝炎患者分选的CD8^+T淋巴细胞与HCV感染的Huh7.5细胞共培养,加入Notch信号通路抑制剂DAPT,通过检测乳酸脱氢酶和细胞因子水平分析CD8^+T淋巴细胞的细胞毒性和非细胞毒性功能的变化。2组间计量资料的比较采用t检验。结果 Notch1和Notch2 mRNA在慢性丙型肝炎患者CD8^+T淋巴细胞中的表达水平较健康志愿者均显著升高,分别升高约10倍和4倍,差异均有统计学意义(t值分别为6.63、18.88,P值分别为0.003、<0.001)。抑制Notch信号通路可增加直接接触培养系统中细胞死亡的比例[(26.41±4.76)%vs(13.65±4.65)%,t=3.32,P=0.029];但抑制Notch信号通路对间接接触培养细胞中CD8^+T淋巴细胞毒性无明显影响。抑制Notch信号通路后,IFNγ的表达在直接接触培养系统[(52.19±12.38)pg/ml vs(18.62±5.66)pg/ml,t=4.27,P=0.013]和间接接触培养系统[(18.86±3.09)pg/ml vs(5.52±2.52)pg/ml,t=5.79,P=0.004]中均显著升高。TNFα的表达在抑制Notch信号通路后仅在直接接触培养系统中显著升高[(1296.0±293.3)pg/ml vs(585.3±32.62)pg/ml,t=4.17,P=0.014],而在间接接触培养系统中无明显变化[(297.3±175.4)pg/ml vs(273.7±105.9)pg/ml,t=0.20,P=0.851]。结论抑制Notch信号通路可增强慢性丙型肝炎患者中CD8^+T淋巴细胞的细胞杀伤和非细胞杀伤功能,Notch信号通路在慢性HCV感染中可能通过抑制CD8^+T淋巴细胞功能发挥诱导免疫耐受的作用。Objective To investigate the influence of the Notch signaling pathway on CD8~＋T lymphocyte function in patients with chronic hepatitis C（CHC） and the role of the Notch signaling pathway in the pathogenesis of CHC.Methods A total of 38 treatment-na6 ve CHC patients who visited Shaanxi Provincial People＇s Hospital from January to August,2016 and 17 healthy volunteers were enrolled in this study.CD8~＋T lymphocytes were isolated,and quantitative real-time PCR was used to measure the mRNA expression of Notch1 and Notch2 in CD8~＋T lymphocytes.CD8~＋T lymphocytes,which were isolated from HLA-A2-restricted CHC patients,were co-cultured with hepatitis C virus（HCV）-infected Huh7.5 cells in both direct and indirect contact culture systems.DAPT,a Notch signaling pathway inhibitor,was added,and the changes in cytotoxic and non-cytotoxic functions of CD8~＋T lymphocytes were analyzed by measuring the levels of lactate dehydrogenase level and cytokines.The t-test was used for comparison of continuous data between groups.Results The mRNA expression of Notch1 and Notch2 in CD8~＋T lymphocytes in CHC patients was 10 and 4 times,respectively,that in healthy volunteers（t = 6.63,P = 0.003; t = 18.88,P〈0.001）.Inhibition of the Notch signaling pathway significantly increased the percentage of dead cells in the direct contact culture system（26.41 ± 4.76% vs 13.65 ± 4.65%,t = 3.32,P = 0.029）,while it did not influence the cytotoxity of CD8~＋T lymphocytes in the indirect contact culture system.After inhibition of the Notch signaling pathway,there was a significant increase in the expression of interferon-γ in the direct（52.19 ± 12.38 pg/ml vs 18.62 ± 5.66 pg/ml,t = 4.27,P = 0.013） and indirect contact culture systems（18.86 ± 3.09 pg/ml vs 5.52 ± 2.52 pg/ml,t = 5.79,P = 0.004）.After inhibition of the Notch signaling pathway,there was a significant increase in the expression of tumor necrosis factor-α in the direct contact culture system（1296.0 ± 293.3 pg/ml vs 585.3 ± 32.62 pg/ml,t = 4.17,

关 键 词：肝炎 丙型 慢性 CD8阳性T淋巴细胞 信号传导 

分 类 号：R512.63[医药卫生—内科学]