补体应答基因32在小鼠部分肝切除后肝再生过程中的表达及意义  

Expression and significance of response gene to complement 32 in liver regeneration after partial hepatectomy in mice

作　　者：李兴元 杨艳芳 陈琰 胡文慧 赵小英 唐俊明 孔德营 LI Xingyuan;YANG Yanfang;CHEN Yan;HU Wenhui;ZHAO Xiaoying;TANG Junming;KONG Deying(Department of Physiology,School of Basic Medical Sciences,Zunyi Medical University,Zunyi,Guizhou 563000,China;Department of Central Laboratory,Guizhou Provincial People’s Hospital,Guiyang 550002,China;Department of Physiology,School of Basic Medicine,Hubei University of Medicine,Shiyan,Hubei 442000,China)

机构地区：[1]遵义医科大学基础医学院生理学教研室,贵州遵义563000 [2]贵州省人民医院中心实验室,贵阳550002 [3]湖北医药学院基础医学院生理学教研室,湖北十堰442000

出　　处：《临床肝胆病杂志》2023年第10期2396-2405,共10页Journal of Clinical Hepatology

基　　金：国家自然科学基金(31760339);湖北省教育厅科学研究计划资助项目(Q20212107);贵州省科技计划项目(黔科合基础-ZK[2022]一般594)。

摘　　要：目的 探讨补体应答基因32(RGC32)在部分肝切除(PH)术后肝再生过程中的表达及作用。方法 42只10周龄雄性C57BL/6小鼠,随机分为对照组[切除小鼠完整的肝脏称重、拍照作为正常对照(sham组),进一步切除肝左叶和肝中叶后称重、拍照作为手术对照(0 d组),sham组和0 d组共用一组小鼠]、术后1天组(1 d)、术后2天组(2 d)、术后4天组(4 d)、术后6天组(6 d)、术后8天组(8 d)和术后10天组(10 d),每组6只;PH术建模成功后分别于术后1、2、4、6、8、10天处死小鼠,收集小鼠肝脏,检测肝脏大小变化。HE和油红O染色评估肝组织形态学变化,血清ALT、AST检测评价肝功能变化,免疫组化染色检测增殖细胞核抗原(PCNA)和Ki67表达并分析肝再生过程中细胞增殖变化,实时荧光定量PCR和免疫组化染色技术检测肝再生过程中RGC32的表达及其亚细胞分布。Ed U细胞增殖实验分析L02细胞过表达或者敲除RGC32对肝细胞增殖的影响。计量资料多组间比较采用方差分析,进一步两两比较采用LSD-t检验;两组间比较采用成组t检验。相关性分析采用Pearson相关分析法。结果 PH术后肝脏逐渐增大,第0~6天为肝体比(肝脏质量/体质量)上升的高峰期,不同时间点比较差异均有统计学意义(P值均<0.05);第6~10天肝脏大小变化不明显。PH术后肝脏脂滴显著增多,随着肝再生,脂滴减少,且呈现门静脉区和中央静脉区差异化(P值均<0.05)。与sham组比较,PH术后1天,血清ALT、AST水平明显升高(P值均<0.05),随后分别于术后第6天和术后第2天恢复至sham组水平(P值均>0.05)。免疫组化染色结果显示,PH术后PCNA和Ki67阳性肝实质细胞数迅速增多,第2天数目最多,分别为86±5和89±5,随后逐渐减少;而PCNA和Ki67阳性非实质细胞数逐渐增多,第6天才达到高峰,分别为34±5和25±3,随后逐渐减少。PH术后总RGC32表达在第2天升到最高,随后逐渐降低,细胞质RGC32表达变化趋势与之一Objective To investigate the expression and role of response gene to complement 32(RGC32)in liver regeneration after partial hepatectomy(PH).Methods A total of 42 male C57BL/6 mice,aged 10 weeks,were randomly divided into control group,postoperative day 1 group(1-d group),postoperative day 2 group(2-d group),postoperative day 4 group(4-d group),postoperative day 6 group(6-d group),postoperative day 8 group(8-d group),and postoperative day 10 group(10-d group),with 6 mice in each group.In the control group,the complete liver of the mice was resected for weighing and photography as the normal control group(sham group);further,the left and middle lobes of the liver were resected for weighing and photography as the surgical control group(0-day group);the sham group and the 0-day group shared the same group of mice.After successful modeling by PH,the mice were sacrificed on days 1,2,4,6,8,and 10 after surgery,and the liver was collected to measure the change in size.HE staining and oil red O staining were used to evaluate liver histomorphological changes;serum alanine aminotransferase(ALT)and aspartate aminotransferase(AST)were measured to evaluate the changes in liver function;immunohistochemical staining was used to measure the expression of proliferating cell nuclear antigen(PCNA)and Ki67 and analyze the change in cell proliferation during liver regeneration;quantitatie real-time PCR and immunohistochemical staining were uused to measure the expression and subcellular distribution of RGC32 during liver regeneration;EdU cell proliferation assay was used to analyze the effect of RGC32 overexpression or knocknout on hepatocyte proliferation in L02 cells.For continuous data,comparison between multiple groups was made by analysis of variance,and further pairwise comparisons were conducted using the LSD-t test.The independent samples t-test was used for comparison of continuous data between two groups.A Pearson correlation analysis was performed.Results The liver gradually enlarged after PH,and the liver/body weight rati

关 键 词：补体应答基因32 肝切除术 肝再生 小鼠 近交C57BL 

分 类 号：R657.3[医药卫生—外科学]