Hou H R, Kang Y
Department of Infectious Diseases, Henan Provincial People's Hospital, Zhengzhou 450003, China.
Zhonghua Gan Zang Bing Za Zhi. 2023 Feb 20;31(2):138-146. doi: 10.3760/cma.j.cn501113-20220314-00108.
To observe the level and detection of ascites CD100 on the activity of CD4(+) and CD8(+) T lymphocytes in vitro in the peripheral blood of patients with liver cirrhosis combined with spontaneous bacterial peritonitis. Peripheral blood and ascites were collected from 77 cases of liver cirrhosis (49 patients with liver cirrhosis combined with simple ascites and 28 patients with liver cirrhosis combined with SBP), and peripheral blood was collected from 22 controls. Soluble CD100 (sCD100) in peripheral blood and ascites was detected by an enzyme-linked immunosorbent assay. Flow cytometry was used to detect membrane-bound CD100 (mCD100) on the surface of CD4(+) and CD8(+)T lymphocytes. CD4(+) and CD8(+)T lymphocytes in ascites were sorted. CD4(+)T lymphocyte proliferation, key transcription factor mRNA, and secreted cytokine changes, as well as CD8(+)T lymphocyte proliferation, important toxic molecule mRNA, and secreted cytokine changes, were detected after CD100 stimulation. The killing activity of CD8(+)T cells was detected by direct contact and indirect contact culture systems. Data conforming to normality were compared using one-way ANOVA, a student's t-test, or a paired t-test. Data that did not conform to a normal distribution were compared using either the Krusal-Willis test or the Mann-Whitney test. There was no statistically significant difference in plasma sCD100 level between patients with liver cirrhosis combined simple ascites (1 415 ± 434.1) pg/ml, patients with liver cirrhosis combined with SBP (1 465 ± 386.8) pg/ml, and controls (1 355 ± 428.0) pg/ml ( = 0.655). The ascites sCD100 level was lower in patients with liver cirrhosis combined with SBP than that of patients with simple ascites [(2 409 ± 743.0) pg/ml vs. (2825±664.2) pg/ml, P=0.014]. There was no statistically significant difference in the level of mCD100 in peripheral blood CD4(+) and CD8(+) T lymphocytes among the three groups ( > 0.05). The levels of mCD100 in ascites CD4(+) and CD8(+) T lymphocytes were higher in patients with liver cirrhosis combined with SBP than those in patients with simple ascites ( < 0.05). CD100 stimulation had no significant effect on the proliferation of CD4(+) and CD8(+)T lymphocytes in the ascites of patients with liver cirrhosis combined with SBP ( > 0.05). There were no significant effects on the expression of transcription factors in effector CD4(+)T lymphocytes (T-bet, retinoic acid associated solitary nuclear receptor γt, aromatic hydrocarbon receptor) or secretion of cytokines (interferon-γ, 17, and 22) ( > 0.05). CD100 stimulation had increased the relative expression of perforin, granzyme B, and granlysin mRNA and the levels of secreted interferon-γ and tumor necrosis factor-α, killing activity in ascites CD8+ T lymphocytes of patients with liver cirrhosis combined with SBP ( < 0.05). The active form of CD100 is sCD100 instead of mCD100. There is an imbalance between the expression of sCD100 and mCD100 in the ascites of patients with cirrhosis combined with SBP. sCD100 can enhance the function of CD8(+)T lymphocytes in the ascites of patients with cirrhosis combined with SBP and thus is one of the potential therapeutic targets.
观察肝硬化合并自发性细菌性腹膜炎患者外周血腹水中CD100水平及其检测对体外CD4(+)和CD8(+)T淋巴细胞活性的影响。收集77例肝硬化患者（49例肝硬化合并单纯腹水患者和28例肝硬化合并自发性细菌性腹膜炎患者）的外周血和腹水，并收集22例对照者的外周血。采用酶联免疫吸附测定法检测外周血和腹水中的可溶性CD100（sCD100）。采用流式细胞术检测CD4(+)和CD8(+)T淋巴细胞表面的膜结合CD100（mCD100）。分选腹水中的CD4(+)和CD8(+)T淋巴细胞。检测CD100刺激后CD4(+)T淋巴细胞增殖、关键转录因子mRNA及分泌细胞因子变化，以及CD8(+)T淋巴细胞增殖、重要毒性分子mRNA及分泌细胞因子变化。采用直接接触和间接接触培养系统检测CD8(+)T细胞的杀伤活性。符合正态分布的数据采用单因素方差分析、Student t检验或配对t检验进行比较。不符合正态分布的数据采用Kruskal-Willis检验或Mann-Whitney检验进行比较。肝硬化合并单纯腹水患者（1415±434.1）pg/ml、肝硬化合并自发性细菌性腹膜炎患者（1465±386.8）pg/ml和对照者（1355±428.0）pg/ml的血浆sCD100水平差异无统计学意义（P=0.655）。肝硬化合并自发性细菌性腹膜炎患者的腹水sCD100水平低于单纯腹水患者[（2409±743.0）pg/ml比（2825±664.2）pg/ml，P=0.014]。三组外周血CD4(+)和CD8(+)T淋巴细胞mCD100水平差异无统计学意义（P>0.05）。肝硬化合并自发性细菌性腹膜炎患者腹水中CD4(+)和CD8(+)T淋巴细胞mCD100水平高于单纯腹水患者（P<0.05）。CD100刺激对肝硬化合并自发性细菌性腹膜炎患者腹水中CD4(+)和CD8(+)T淋巴细胞增殖无显著影响（P>0.05）。对效应CD4(+)T淋巴细胞中转录因子（T-bet、视黄酸相关孤核受体γt、芳烃受体）的表达或细胞因子（干扰素-γ、白细胞介素-17和白细胞介素-2）的分泌无显著影响（P>0.05）。CD100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淋巴细胞穿孔素、颗粒酶B和颗粒溶素mRNA的相对表达及分泌的干扰素-γ和肿瘤坏死因子-α水平、杀伤活性（P<0.05）。CD100的活性形式是sCD100而非mCD100。肝硬化合并自发性细菌性腹膜炎患者腹水中sCD100和mCD100的表达存在失衡。sCD100可增强肝硬化合并自发性细菌性腹膜炎患者腹水中CD8(+)T淋巴细胞的功能，因此是潜在的治疗靶点之一。
