GITR阳性的效应T淋巴细胞参与AAF/PH小鼠肝脏再生调控  

GITR-positive effector T cells are involved in AAF/PH mouse liver regeneration

作　　者：李丽[1] 何宇 刘锴[1] 王萍[1] LI Li;HE Yu;LIU Kai;WANG Ping(Liver Disease Center,Beijing Friendship Hospital,Capital Medical University,Beijing 100050,China)

机构地区：[1]首都医科大学附属北京友谊医院肝病中心,100050

出　　处：《肝脏》2023年第1期112-116,共5页Chinese Hepatology

基　　金：国家自然科学基金(81570548);王宝恩肝纤维化基金(2019073);国家新药研发重大专项(2018ZX09201016)。

摘　　要：目的 分析2-乙酰氨基芴联合部分肝切除(AAF/PH)小鼠肝脏再生模型肝组织中浸润的效应T淋巴细胞的变化情况。方法 C57BL/6J小鼠每天应用AAF连续灌胃5 d后切除1/3肝组织,术后继续每天应用AAF灌胃至术后7 d、14 d。利用免疫荧光染色、蛋白免疫印迹检测肝脏前体细胞活化情况;采用多色免疫荧光染色与流式细胞术检测肝组织中效应T淋巴细胞及其活化标志物GITR阳性细胞比例。结果 AAF/PH组小鼠术后7 d肝组织重量即可恢复至对照小鼠水平。肝组织中炎症细胞浸润和纤维化不明显,但术后14 d时,肝脏汇管区中出现明显的炎症细胞浸润与细胞外基质沉积。随着术后时间延长,肝组织中Sox9+肝脏前体细胞数量逐渐增多,肝组织Sox9蛋白表达水平亦显著增加。流式细胞分析结果显示,随着术后时间延长肝组织中浸润的CD8+T淋巴细胞的比例显著增加,术后14 d为对照组的1.47倍(P=0.0289),而糖皮质激素诱导的肿瘤坏死因子受体(GITR)阳性的CD8+T淋巴细胞比例呈先升高再降低的变化趋势,术后7 d、14 d分别为对照组的1.5倍(P=0.0098)、0.65倍(P=0.0427)。尽管术后CD4+T淋巴细胞比例无明显变化,但术后14 d时GITR阳性的CD4+T淋巴细胞比例显著降低,为对照组的0.85倍(P=0.0225)。结论 AAF/PH小鼠肝再生模型中伴随肝脏前体细胞的活化,效应T淋巴细胞参与了小鼠的肝脏再生过程。Objective To observe the ratio variation of effector T lymphocytes during liver regeneration in 2-acetylaminofluorene/partial hepatectomy(AAF/PH) mouse. Methods After daily intragastric administration of AAF for 5 days, C57BL/6J mice were performed 1/3 hepatectomy followed by daily administration of AAF for 7 days or 14 days. Immune fluorescence staining and western blot were used to detect the activation of hepatic progenitor cells. Collagenase digestion and Percoll-density centrifugation were used to isolate liver non-parenchymal cells, and multi-color immune staining and flow cytometry were used to analyze the percentage of effector T lymphocytes and their glucocorticoid-induced TNF receptor(GITR)-positive ratio. Results On the 7th post-operative day, there was no significant difference with regard to liver weght between the AAF/PH and control groups. Although few inflammatory cells and no significant liver fibrosis were observed in the liver tissue of the AAF/PH group, the liver tissue showed significant inflammatory cell infiltration and extracellular matrix deposition at the portal area on the 14th post-operative day. After partial hepatectomy, the number of Sox9+ hepatic progenitor cells increased time-dependently, and the expression level of Sox9 also enhanced in the liver tissue. Flow cytometry analysis showed that the ratio of CD8+T lymphocytes to CD3+T lymphocytes increased after partial hepatectomy, while the ratio of CD8+GITR+T cells increased at 7 days post-operation, when compared to the control group, but reduced to the level of the control group at 14 days post-operation. Although the ratio of CD4+T lymphocytes to CD3+T lymphocytes did not change significantly, the ratio of CD4+GITR+T lymphocytes reduced significantly at 14 days post-operation compared to the control group. Conclusion AAF/PH activates hepatic progenitor cells in mouse liver regeneration model, and effector T cells are involved in mouse liver regeneration.

关 键 词：2-乙酰氨基芴联合部分肝切除 肝脏前体细胞 糖皮质激素诱导的肿瘤坏死因子受体 

分 类 号：R392[医药卫生—免疫学]