β-谷甾醇靶向CDC25B抑制肝癌细胞增殖  

Mechanisms ofβ-sitosterol on Inhibiting the Proliferation of Hepatocellular Carcinoma Cells by Targeting CDC25B

作　　者：王凯 李卫 李志芳 纪会琼 郭志伟 曹俊秋 WANG Kai;LI Wei;LI Zhi-fang;JI Hui-qiong;GUO Zhi-wei;CAO Jun-qiu(Department of General Foundation Requisite Teaching,Dazhou Vocational College of Traditional Chinese Medicine,Dazhou 635000,China;Affiliated Hospital of Dazhou Center for Disease Control and Prevention,Dazhou 635000,China;Department of Medical Imaging,Nanchong Central Hospital,Nanchong 637000,China;Department of Hepatobili-ary Surgery,West China Hospital,Sichuan University,Chengdu 610000,China)

机构地区：[1]达州中医药职业学院公共基础部,四川达州635000 [2]达州市疾病预防控制中心附属医院,四川达州635000 [3]南充市中心医院影像科,四川南充637000 [4]四川大学华西医院肝胆外科,四川成都610000

出　　处：《中山大学学报（医学科学版）》2022年第4期675-684,共10页Journal of Sun Yat-Sen University:Medical Sciences

基　　金：四川省医学(青年创新)科研课题(Q20043)。

摘　　要：【目的】探讨β-谷甾醇对肝癌细胞增殖的影响及其可能作用机制。【方法】通过SwissTarget和TargetNet数据平台预测β-谷甾醇的靶点、GEO2R分析GSE101728肝癌数据集中表达上调基因并分析两者共同基因,通过下载TCGA_LIHC数据矩阵分析得到细胞分裂周期25B(CDC25B)与肝癌临床分期和患者生存预后存在关系。CCK-8、细胞集落形成实验、流式细胞法检测β-谷甾醇作用下肝癌细胞HepG2和Hep3B细胞活力、细胞增殖、细胞周期分布和细胞凋亡情况;构建HepG2皮下移植瘤裸鼠模型分为Control组和β-谷甾醇组,β-谷甾醇组腹腔注射β-谷甾醇(50 mg/kg·d),测量移植瘤体积和质量,免疫组化检测Ki67表达水平;Western blot检测β-谷甾醇不同作用浓度和不同作用时间HepG2和Hep3B细胞系中CDC25B蛋白水平,构建稳定过表达CDC25B的HepG2和Hep3B细胞系并给予β-谷甾醇处理,同上方法检测HepG2和Hep3B细胞系细胞活力、细胞增殖、细胞周期分布和细胞凋亡情况。【结果】CDC25B表达水平,肝癌组织高于其癌旁组织(P<0.05),TNM分期(T2-4)高于TNM分期(T1)(P<0.05),肝癌临床分期Ⅱ-Ⅳ高于临床分期I(P<0.05),肝癌G3-4分化高于G1-2分化(P<0.05),复发患者高于未复发者(P<0.05),预后不良者高于预后良好者(P<0.05);CDC25B高表达组患者的总生存率低于CDC25B低表达组(P<0.05)。Hep3B和HepG2的细胞活力随着β-谷甾醇作用剂量增加、作用时间延长而降低,Hep3B(β-谷甾醇15µmol/L)和HepG2(β-谷甾醇18µmol/L)的相对克隆数目、S期细胞百分比均显著低于Control组(P<0.05),而G0/G1期细胞百分比、细胞凋亡率均显著高于Control组(P<0.05)。β-谷甾醇组移植瘤的体积、质量和Ki67相对表达水平均显著低于Control组(P<0.05)。Hep3B和HepG2细胞中的CDC25B蛋白表达均随着β-谷甾醇浓度增加、作用时间延长而减低,Hep3B和HepG2细胞的β-谷甾醇+CDC25B组的细胞生长、相对克隆数目、S期细胞百分比均【Objective】To explore the effects and possible actions mechanisms ofβ-sitosterol on the proliferation of hepatocellular carcinoma(HCC)cells.【Methods】The target ofβ-sitosterol was predicted by SwissTarget and TargetNet data platforms.The up-regulated gene in GSE101728 HCC data set was analyzed by GEO2R,and the common gene be⁃tween the target and the up-regulated gene was analyzed.According to the analysis of TCGA_LIHC data matrix,the rela⁃tionship between cell division cyclin 25B(CDC25B)and clinical staging,prognosis of HCC was obtained.The activities,proliferation,cycles distribution and apoptosis of HCC cells(HepG2,Hep3B)treated withβ-sitosterol were detected by CCK-8,colony-forming assay and flow cytometry.The models of nude mice with subcutaneous xenografted tumors in HepG2 were constructed and divided into control group andβ-sitosterol group.Theβ-sitosterol group was intraperitoneal⁃ly injected withβ-sitosterol(50 mg/kg·d).The volume and mass of tumors were measured.The expression level of Ki67 was detected by immunohistochemistry.The levels of CDC25B protein in HepG2 and Hep3B cell lines treated with differ⁃ent concentrations ofβ-sitosterol for different action time were detected by Western blot.HepG2 and Hep3B cell lines with stable overexpression of CDC25B were constructed,and then which were treated withβ-sitosterol.The activities,prolifer⁃ation,cycles distribution and apoptosis of HepG2 and Hep3B cell lines were detected by the above-mentioned methods.【Results】The expression level of CDC25B in HCC tissues was higher than that in para-carcinoma tissues(P<0.05).The expression level of CDC25B in patients with TNM staging at stage T2-4,clinical staging at stageⅡ-Ⅳ,G3-4 differentia⁃tion,recurrence and poor prognosis was higher than that with TNM staging at stage T1,clinical staging at stage I,G1-2 differentiation,non-recurrence and good prognosis,respectively(P<0.05).The overall survival rate in high-expression CDC25B group was lower than that in low-expression group(P<0.0

关 键 词：肝癌 Β-谷甾醇 细胞分裂周期25B 细胞增殖 

分 类 号：R735.7[医药卫生—肿瘤]