聚乙二醇干扰素-α-2b对HBeAg阴性慢性HBV感染者HBV特异性CD8+T淋巴细胞杀伤功能的影响  

Effect of pegylated interferon-α-2b therapy on cytotoxicity of virus-specific CD8+T cells in HBeAg-negative patients with chronic hepatitis B virus infection

作　　者：秦蕾[1] 李广鹏[2] 申培君 张兰芳[1] 杨晓飞 彭梅娟 张野 QIN Lei;LI Guangpeng;SHEN Peijun;ZHANG Lanfang;YANG Xiaofei;PENG Meijuan;ZHANG Ye(Department of Gastroenterology,The First Affiliated Hospital of Xinxiang Medical University,Xinxiang,Henan 453000,China;Department of Emergency Medicine,The First Affiliated Hospital of Xinxiang Medical University,Xinxiang,Henan 453000,China;Department of Infectious Diseases,Tangdu Hospital,Air Force Medical University,Xi’an 710038,China)

机构地区：[1]新乡医学院第一附属医院消化内科,河南新乡453000 [2]新乡医学院第一附属医院急诊科,河南新乡453000 [3]空军军医大学唐都医院传染病科,西安710038

出　　处：《临床肝胆病杂志》2025年第4期628-636,共9页Journal of Clinical Hepatology

基　　金：空军军医大学科技发展基金(2022XB031)。

摘　　要：目的观察经聚乙二醇干扰素-α-2b(PEG-IFN-α-2b)治疗后,HBeAg阴性慢性HBV感染者HBV特异性CD8+T淋巴细胞活性的变化。方法纳入2020年4月—2022年6月于新乡医学院第一附属医院、空军军医大学唐都医院就诊的HBe Ag阴性慢性HBV感染者53例,予以PEG-IFN-α-2b(180μg/周,皮下注射)抗病毒治疗,研究终点为HBsAg阴转(疗程<48周)或48周(疗程≥48周),分别选取基线和研究终点外周血单个核细胞,检测外周血T淋巴细胞计数,酶联免疫斑点试验检测分泌穿孔素、颗粒酶B、IFN-γ的HBV特异性CD8+T淋巴细胞频数。选择HLA-A^(*)02限制性患者17例,纯化CD8+T淋巴细胞,建立HBV特异性CD8+T淋巴细胞与Hep G2.2.15细胞直接接触和间接接触共培养系统,通过检测上清中乳酸脱氢酶水平计算Hep G2.2.15细胞死亡率,检测上清中HBV DNA水平、毒性分子和细胞因子分泌,流式细胞术检测凋亡配体表达,评估HBV特异性CD8+T淋巴细胞的杀伤功能。符合正态分布的计量资料2组间比较采用成组t检验或配对t检验,不符合正态分布的计量资料2组间比较采用Mann-Whitney U检验或Wilcoxon秩和检验。结果研究终点时,HBsAg阴转率为30.19%(16/53)。外周血T淋巴细胞计数(CD3+、CD4+、CD8+T淋巴细胞)在基线和研究终点时的差异均无统计学意义(P值均>0.05)。研究终点时,患者分泌穿孔素、颗粒酶B、IFN-γ的HBV特异性CD8+T淋巴细胞频数较基线显著升高(U=177.50,t=11.90,U=186.50,P值均<0.001),发生HBsAg阴转的患者分泌穿孔素、颗粒酶B、IFN-γ的HBV特异性CD8+T淋巴细胞频数亦显著高于未发生HBsAg阴转的患者(U=120.50,t=2.73,U=121.50,P值均<0.01)。在直接接触和间接接触共培养系统中,研究终点时HBV特异性CD8+T淋巴细胞均可诱导Hep G2.2.15细胞上清中HBV DNA显著下降(P值均<0.001),IFN-γ和TNF-α分泌水平显著升高(P值均<0.05),但仅在直接接触共培养系统中HBV特异性CD8+T淋巴细胞诱导Hep G2.2.15细胞死�Objective To investigate the change in the activity of hepatitis B virus(HBV)-specific CD8+T cells after pegylated interferon-α-2b(PEG-IFN-α-2b)therapy in HBeAg-negative patients with chronic HBV infection.Methods A total of 53 HBeAgnegative patients with chronic HBV infection who attended The First Affiliated Hospital of Xinxiang Medical University and Tangdu Hospital of Air Force Mdical University from April 2020 to June 2022 were enrolled and treated with PEG-IFN-α-2b(180μg/week,subcutaneous injection)antiviral therapy.The study endpoint was HBsAg clearance(course of treatment<48 weeks)or 48 weeks(course of treatment≥48 weeks).Peripheral blood mononuclear cells were isolated at baseline and study endpoint,and peripheral blood T cell counts were measured.Enzyme-linked immunospot assay was used to measure the frequency of HBV-specific CD8+T cells secreting perforin,granzyme B,and interferon-γ.A total of 17 HLA-A^(*)02-restricted patients were selected,and CD8+T cells were purified to establish direct-and indirect-contact co-culture systems for HBV-specific CD8+T cells and HepG2.2.15 cells.The level of lactate dehydrogenase in supernatant was measured to calculate the mortality rate of HepG2.2.15 cells,and the levels of HBV DNA,cytotoxic molecules,and cytokines in supernatant were also measured.Flow cytometry was used to measure the expression of apoptosis ligands,and the cytotoxicity of HBV-specific CD8+T cells was evaluated.The independent samples t-test or the paired t-test was used for comparison of normally distributed continuous data between two groups,and the Mann-Whitney U test or the Wilcoxon rank-sum test was used for comparison of non-normally distributed continuous data between two groups.Results The HBsAg clearance rate at study endpoint was 30.19%(16/53).There were no significant differences in peripheral blood T cell counts(CD3+,CD4+,and CD8+T cells)between baseline and study endpoint(P>0.05).At study endpoint,there was a significant increase in the frequency of HBV-specific CD8+T cells secr

关 键 词：聚乙二醇干扰素 乙型肝炎E抗原 乙型肝炎病毒 CD8阳性T淋巴细胞 

分 类 号：R51[医药卫生—内科学]