青蒿琥酯通过调控GADD45A、NACC1的表达抑制肝癌细胞的作用  

The inhibitory effect of artesunate on hepatocellular carcinoma cells by regulating expression of GADD45A and NACC1

作　　者：沈冠彤 董金垚 冯璟[3] 秦楠 杜根来 朱飞 连科 刘新宇 李清靓 张迅玮 师如意[1] SHEN Guan-tong;DONG Jin-yao;FENG Jing;QIN Nan;DU Gen-lai;ZHU Fei;LIAN Ke;LIU Xin-yu;LI Qing-liang;ZHANG Xun-wei;SHI Ru-yi(Dept of Medical Cell Biology and Genetics,Shanxi Medical University,Taiyuan 030001,China;Dept of Hepatobiliary,Pancreatic and Gastric Surgery,Shanxi Cancer Hospital,Taiyuan 030013,China;Dept of Gastroenterology,Shanxi Provincial People's Hospital,Taiyuan 030012,China;College of Pharmaceutical and Food Engineering,Shanxi University of Chinese Medicine,Jinzhong Shanxi 030619,China;Shanxi Bethune Hospital,Shanxi Academy of Medical Sciences,Third Hospital of Shanxi Medical University,Taiyuan 030032,China)

机构地区：[1]山西医科大学基础医学院医学细胞生物学与遗传学教研室,山西太原030001 [2]山西省肿瘤医院肝胆胰胃外科,山西太原030013 [3]山西省人民医院消化科,山西太原030012 [4]山西中医药大学中药与食品工程学院,山西晋中030619 [5]山西白求恩医院(山西医学科学院同济山西医院),山西医科大学第三医院,山西太原030032

出　　处：《中国药理学通报》2024年第6期1089-1097,共9页Chinese Pharmacological Bulletin

基　　金：山西省基础研究计划(No 202103021224369);山西省中医药管理局项目(No 2023ZYYC2043);山西省研究生教育创新计划项目(No 2023KY397)。

摘　　要：目的阐明青蒿琥酯(artesunate,ART)在肝细胞癌(hepatocellular carcinoma,HCC)病理进程中对肿瘤细胞的作用及潜在的机制。方法用含不同浓度ART的培养基处理两种HCC细胞系MHCC-97H、HCC-LM3,以0 mg·L-1 ART处理的细胞作为对照组,对比ART对HCC的作用。采用MTT和克隆形成实验测定ART对HCC细胞生长能力的影响;划痕实验和Transwell实验分别检测ART对HCC细胞迁移和侵袭能力的影响;流式细胞术验证ART对HCC细胞凋亡和细胞周期变化的影响;通过转录组测序分析以及qRT-PCR验证关键基因的表达情况探究ART在HCC细胞中的可能作用机制。结果与对照组相比,ART处理后的肿瘤细胞增殖、迁移、侵袭能力明显降低(P<0.01),而细胞凋亡率明显上升,且G_(2)/M期细胞比例明显增高,提示肿瘤细胞被阻滞于该期。使用RNA测序数据进行转录组学分析,确定生长停滞与DNA损伤诱导蛋白45A(growth arrest and DNA damage inducible alpha,GADD45A)是ART的潜在作用靶点,并且提示,ART可通过上调GADD45A的表达进而诱导细胞凋亡和细胞周期阻滞。此外,生信分析结果提示,GADD45A与其上游转录因子伏隔核相关蛋白1(nucleus accumbens associated 1,NACC1)有蛋白互作现象,并且经ART处理后,GADD45A与NACC1的mRNA水平和蛋白水平均发生明显变化。结论ART可能通过影响GADD45A及其潜在上游NACC1基因的表达抑制HCC的发生发展。通过探究ART作用于HCC的功能影响及发生机制,为临床治疗肝癌提供新药物、新方向和新思路。Aim To explore the effect and mechanism of the artesunate(ART)on hepatocellular carcinoma(HCC).Methods The cell lines MHCC-97H and HCC-LM3 were used to be detected.MTT and clone formation were used to determine the cell proliferation;Wound healing was used to detect the cell migration;Transwell was used to test the cell invasion.Flow-cytometry was used to detect cell apoptosis and cell cycle.RNA-seq and qRT-PCR was used to detect the genes expression.Results The proliferation,migration and invasion of treated cells were obviously inhibited(P<0.01).Moreover,the apoptosis rate increased significantly,so did the proportion of G_(2)/M cells.Transcriptomic analysis identified GADD45A as a potential target of ART through RNA-sequencing data,and suggested that ART might induce apoptosis and cell cycle arrest through regulating the expression of GADD45A.In addition,the results of mechanism studies and signaling analysis suggested that GADD45A had interaction with its upstream gene NACC1(nucleus accumbens associated 1).Moreover,after ART treatment,the expressions of GADD45A and NACC1 were changed significantly.Conclusion ART may be a potential drug to resist HCC by affecting the expression of GADD45A and its upstream gene NACC1,which provides a new drug,a new direction and a new method for the clinical treatment of HCC.

关 键 词：青蒿琥酯 肝细胞癌 抗癌作用机制 GADD45A NACC1 细胞凋亡 G_(2)/M 

分 类 号：R284.1[医药卫生—中药学] R329.25[医药卫生—中医学] R329.28R394.2R735.7