中药复方制剂抑制CD133^+肝癌干细胞增殖及干性转录因子的表达  

Chinese medicine compound preparation inhibits proliferation of CD133^+liver cancer stem cells and the expression of stemness transcription factors

作　　者：覃艳春 荣震[2] 蒋锐沅 付彬 洪晓华 莫春梅[2] Qin Yanchun;Rong Zhen;Jiang Ruiyuan;Fu Bin;Hong Xiaohua;Mo Chunmei(Department of Graduate,Guangxi University of Chinese Medicine,Nanning 530000,Guangxi Zhuang Autonomous Region,China;First Affiliated Hospital of Guangxi University of Chinese Medicine,Nanning 530000,Guangxi Zhuang Autonomous Region,China)

机构地区：[1]广西中医药大学研究生学院,广西壮族自治区南宁市530000 [2]广西中医药大学第一附属医院,广西壮族自治区南宁市530000

出　　处：《中国组织工程研究》2021年第19期3016-3023,共8页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金(81760850),项目负责人:莫春梅;广西高校中青年教师基础能力提升项目(2018KY0282),项目负责人:莫春梅;广西医疗卫生适宜技术开发与推广应用项目(S2018046),项目负责人:莫春梅;广西中医药大学第一附属医院院内制剂研究与开发(2018ZJ004),项目负责人:莫春梅。

摘　　要：背景:癌症起源于干细胞的恶性转化越来越得到普遍认同。肝癌干细胞具有不断自我更新与多向分化潜能,可促进肝癌的发生发展、术后复发和耐药性的产生。目的:探讨中药复方制剂敷和备化方对CD133^+肝癌干细胞增殖及干性转录因子表达的影响。方法:采用免疫磁珠法分选出CD133^+HepG2肝癌干细胞,分别用2%,4%,8%,10%,12%,16%6个不同体积分数中药复方制剂敷和备化方含药血清干预CD133^+HepG2肝癌干细胞,通过CCK-8法检测细胞增殖抑制作用,筛选出最佳体积分数含药血清。然后将CD133^+HepG2肝癌干细胞分为二甲基亚砜组、敷和备化方组、血清对照组、空白对照组,其中二甲基亚砜组用0.05%二甲基亚砜干预,敷和备化方组用体积分数为16%敷和备化方含药血清干预,血清对照组用体积分数为16%正常大鼠血清干预,干预6 d后采用流式细胞术检测CD133^+HepG2肝癌干细胞百分比,RT-PCR以及Western blot检测干细胞转录因子SOX2、NANOG和OCT4的mRNA以及蛋白表达。结果与结论:①中药复方制剂敷和备化方含药血清对CD133^+HepG2细胞增殖有抑制作用,且与时间、体积分数相关,其中体积分数为16%敷和备化方含药血清干预72 h抑制作用最明显(P<0.05);②与血清对照组和空白对照组相比,敷和备化方组和二甲基亚砜组都可降低CD133^+HepG2细胞百分比(P<0.05),且敷和备化方组作用更明显;③与空白对照组和血清对照组相比,敷和备化方组和二甲基亚砜组都可下调干细胞转录因子SOX2、NANOG、OCT4 mRNA和蛋白表达(P<0.05),且敷和备化方组作用更明显;④结果表明,中药复方制剂敷和备化方含药血清能有效降低肝癌干细胞表面标记物CD133和干细胞转录因子SOX2、NANOG、OCT4的表达水平,降低肝癌干细胞恶性程度。BACKGROUND:The malignant transformation of cancer originating from stem cells is increasingly recognized.Liver cancer stem cells have the potential of continuous self-renewal and multi-directional differentiation,which can promote the occurrence and development of liver cancer,postoperative recurrence and the production of drug resistance.OBJECTIVE:To explore the effect of Fuhe preparation formula on proliferation of CD133^+liver cancer stem cells and stemness transcription factor expression.METHODS:CD133^+HepG2 liver cancer stem cells were selected by immunomagnetic bead method.CD133^+HepG2 cells were treated with serum containing 2%,4%,8%,10%,12%and 16%Fuhe preparation formula respectively.The inhibitory rate of CD133^+HepG2 cells was determined by CCK-8 method,and the optimal volume fraction of serum containing drugs was selected.The CD133^+HepG2 liver cancer stem cells were divided into dimethyl sulfoxide group,application and preparation group,serum control group and blank control group.Cells in the dimethyl sulfoxide group were intervened with 0.05%dimethyl sulfoxide.Cells in the application and preparation group were intervened with 16%application and preparation containing serum.Cells in the serum control group were intervened with 16%normal rat serum.After 6 days of intervention,the percentage of CD133^+HepG2 liver cancer stem cells was detected by flow cytometry.The mRNA and protein levels of transcription factors SOX2,NANOG,and OCT4 were detected by RT-PCR and western blot assay.RESULTS AND CONCLUSION:(1)The serum containing application and preparation formula inhibited the proliferation of CD133^+HepG2 cells,which was related to the time and concentration.The inhibitory effect of 16%application and preparation formula was the most obvious in 72 hours(P<0.05).(2)Compared with the serum control and blank control groups,the percentage of CD133^+HepG2 liver cancer stem cells was decreased in the application and preparation and dimethyl sulfoxide groups(P<0.05),and the effect was more obvious in the applic

关 键 词：干细胞 肝癌干细胞 敷和备化方 含药血清 CD133 干细胞转录因子 

分 类 号：R459.9[医药卫生—治疗学] R394.2[医药卫生—临床医学]