摘要：
目的 观察肝硬化合并自发性细菌性腹膜炎患者腹水中白细胞介素-7(IL-7)的表达水平,并检测重组人IL-7对腹水中CD4+T淋巴细胞和CD8+T淋巴细胞功能的影响.方法 选择2017年8月至2018年4月住院治疗的肝硬化患者84例,其中肝硬化合并单纯腹水患者51例,肝硬化合并自发性细菌性腹膜炎患者33例,常规采集外周血和收集腹水,应用酶联免疫吸附法检测外周血和腹水中IL-7的水平.分选腹水中CD4+T淋巴细胞和CD8+T淋巴细胞,使用重组人IL-7刺激培养,检测CD4+T淋巴细胞增殖、关键转录因子mRNA以及分泌细胞因子的变化,检测CD8+T淋巴细胞中穿孔素、颗粒酶B和颗粒溶素mRNA和分泌细胞因子的变化,利用直接接触和间接接触共培养系统检测CD8+T淋巴细胞的细胞杀伤和非细胞杀伤功能的变化.符合正态分布的计量资料使用Student检验进行两组数据间比较,使用配对t检验进行刺激前后数据比较.不符合正态分布的计量资料使用Mann-Whitney检验进行两组间数据比较,使用Wilcoxon配对检验进行刺激前后数据比较.结果 肝硬化合并自发性细菌性腹膜炎组和肝硬化合并单纯腹水组患者血清IL-7水平的差异无统计学意义[(5001±1 458)pg/ml比(4768±1128)pg/ml,P=0.412),而肝硬化合并自发性细菌性腹膜炎患者腹水中IL-7的水平较肝硬化合并单纯腹水患者显著降低[(792.1±126.4)pg/ml比(966.4±155.8)pg/ml,P<0.01].重组人IL-7刺激可促进肝硬化合并自发性细菌性腹膜炎患者腹水中CD4+T淋巴细胞和CD8+T淋巴细胞的增殖,增加CD4+T淋巴细胞中转录因子T-bet mRNA的相对表达量和干扰素-γ的分泌,亦可增加CD8+T淋巴细胞中穿孔素、颗粒酶B和颗粒溶素mRNA的相对表达量以及CD8+T淋巴细胞分泌干扰素-γ和肿瘤坏死因子-α的水平.重组IL-7刺激可增加肝硬化合并自发性细菌性腹膜炎患者腹水中CD8+T淋巴细胞的细胞杀伤功能和非细胞杀伤功能,主要表现为IL-7刺激CD8+T淋巴细胞后直接接触和间接接触共培养系统中靶细胞死亡比例和干扰素-γ分泌水平的升高.结论 IL-7可增强肝硬化合并自发性细菌性腹膜炎患者腹水中T淋巴细胞的功能.%Objective To observe ascitic interleukin-7 expression level in cirrhotic patients complicated with spontaneous bacterial peritonitis,and to detect the effect of recombinant human IL-7 on CD4+ and CD8+T lymphocyte function.Methods A total of 84 patients with liver cirrhosis who were hospitalized from August 2017 to April 2018 were selected.Among them,51 cases were complicated with cirrhosis and untainted ascites,and 33 cases were cirrhosis complicated with spontaneous bacterial peritonitis.Peripheral blood and ascites were collected routinely.The levels of IL-7 in peripheral blood and ascites were measured by enzyme-linked immunosorbent assay.CD4+T cells and CD8+T cells were purified from ascites,and were stimulated with recombinant IL-7.Cellular proliferation,key transcription factors for mRNA,and cytokines production by CD4+T cells in response to IL-7 stimulation was measured.mRNA expression corresponding to perforin,granzyme B,and granulysin as well as cytokines production by CD8+T cells was also measured in response to IL-7 stimulation.Cytolytic and non-cytolytic activity of CD8+T cells in response to IL-7 stimulation was also investigated in both direct and indirect contact co-culture system.Measurement data of the normal distribution were compared between the two groups by Student's t-test and the data before and after stimulation were compared by paired t-test.Measurements that did not conform to normal distribution were compared between the two groups using Mann-Whitney U test,and data before and after stimulation were compared using Wilcoxon paired test.Results There was no significant statistical difference in serum IL-7 levels between the two groups [(5001±1458)pg/ml vs.(4768±1128)pg/ml,P=0,41].The level of ascitic IL-7 in cirrhotic patients complicated with SBP was significantly lower than cirrhosis patients with untainted ascites [(966.4 + 155.8)pg/ml vs.(792.1 + 126.4)pg/ml,P<0.01].Recombinant IL-7 stimulation promoted the proliferation of CD4+ and CD8+T cells from ascites in patients with liver cirrhosis complicated by SBP.T-bet mRNA relative expression and IFN-γ secretion in CD4+T cells was also elevated in response to IL-7 stimulation in vitro.Moreover,IL-7 stimulation also increased the mRNA expressions of perforin,granzyme B,and granulysin as well as productions of IFN-γand TNF-α by CD8+T cells.Recombinant IL-7 stimulation elevated cytolytic and non-cytolytic activity of CD8+T cells from ascites in patients with liver cirrohosis complicated by SBP,which manifested as increased target cell death and IFN-γproduction in both direct and indirect contact co-culture system.Conclusion Ascitic IL-7 promotes T lymphocyte function in patients with liver cirrhosis complicated with SBP.
