白杨素对顺铂及喜树碱诱导肝肿瘤细胞HepG2凋亡的增敏作用  

Sensitization of Chrysin on the Apoptosis Induced by Cisplatin or Camptothecin in Hepatoma Cell Lines( Hep G2)

作　　者：王剑宁[1] 李欣[2] 陈美芬[2] 王凤岩[2] 熊习昆[2] 陈秀娟[2] 杨美玲[2] 黄俊明[2] WANG Jian-ming LI Xin CHEN Mei-fen WANG Feng-yan XIONG Xi-kun CHEN Xiu-juan YANG Mei-ling HUANG Jun-ming(Guanghua School of Stomatology, Hospital of Stomatology, Sun Yat-sen University, Guang- dong Provincial Key Laboratory of Stomatology, Guangzhou 510055, China Guangdong Provincial Center for Disease Control and Prevention, the Institute of Toxicology, Guangzhou 511430, China)

机构地区：[1]中山大学光华口腔医学院附属口腔医院口腔颌面外科,广东省口腔医学重点实验室,广州510055 [2]广东省疾病预防控制中心卫生毒理所,广州511430

出　　处：《中国药学杂志》2016年第24期2088-2093,共6页Chinese Pharmaceutical Journal

基　　金：国家自然科学基金资助项目(81202199);广东省科技计划项目资助(2013B021800044,2014A020212437);广州市科技计划项目(2014J4100090)

摘　　要：目的研究天然产物白杨素(chrysin)对损伤DNA的抗肿瘤药物顺铂和喜树碱诱导肝肿瘤细胞(Hep G2)死亡的作用及机制。方法白杨素以不同浓度单独/联合顺铂或喜树碱处理Hep G2,于倒置显微镜下观察肿瘤细胞形态变化,以四甲基偶氮唑蓝(MTT)法分析细胞活性,获得细胞死亡的定量资料;并以Western blot方法分析凋亡标志蛋白caspase-3和多聚(ADP-核糖)聚合酶(PARP)以及凋亡抑制蛋白Bcl-x L、x IAP、FLIP相应的变化情况。结果形态学观察可发现白杨素联合顺铂或喜树碱处理可明显增加Hep G2细胞的死亡,MTT法分析显示,白杨素联合抗肿瘤药物处理细胞,无论与未处理对照比较还是与单独白杨素、单独顺铂或单独喜树碱处理比较,均使细胞活性显著降低(P<0.05);Hochest 33342荧光染色表明,联合白杨素和抗肿瘤药物处理细胞可观察到明显的核固缩,而未处理对照及单独白杨素、单独顺铂或喜树碱处理则无此作用;Thermo高内涵筛选系统分析结果显示,联合白杨素和抗肿瘤药物处理可使凋亡细胞显著增加(P<0.05)。Western blot检测到凋亡标志蛋白caspase-3和PARP活化降解;全caspase酶抑制剂z-VAD-fmk可有效阻止凋亡标志蛋白caspase-3和PARP的活化降解。白杨素联合顺铂可抑制顺铂上调的凋亡抑制蛋白FLIP和x IAP表达,而白杨素联合喜树碱可降低喜树碱引起的凋亡抑制蛋白Bcl-x L的高表达。结论天然产物白杨素能有效促进DNA损伤类抗肿瘤药物诱导的Hep G2细胞凋亡,核因子(NF)-κB调节的凋亡抑制蛋白表达降低是其重要的机制。OBJECTIVE To investigate the effects of chrysin on the apoptosis induced by DNA damage antitumor drugs in hepato- ma （ Hep G2） cell lines, and its molecular mechanism. METHODS Hep （;2 cells were pretreated with chrysin for 2 h, then treated with cisplatin or camptothecin for 24 h. The morphologie changes were observed under inversed microscope and the cell viability was measured using MTT test. The proteins of caspase-3, PARP, Bcl-xL, xlAP and FLIP were determined by Western blot. RESULTS Increases of cell death were observed in the combination of chrysin and eisplatin or camptotheein. There were significant differences in the cell viability not only between the combined treatment and the untreated control, but also between the combined treatment and chry- sin alone, cisplatin alone or camptothecin alone. Chromatin condensation could be observed when the cells were stained by Hochest 33342 in the combination of ehrysin and DNA damage antitumor drugs, and the apoptotic cells showed significant increase in the com- bination group eompared with other groups（ P 〈 0. 05 ）. The proproteins of caspase-3 and PARP degraded. The pan-caspase inhibitor z- VAD-fmk could inhibit the activation of caspase-3 and PARP induced by the combination of chrysin and eisplatin or camptothecin. The apoptosis inhibitory proteins, FLIP and xlAP which upregulated by cisplatin alone, could be downregulated by the cotreatment of chrysin and cisplatin;and Bcl-xL upregulated by eamptothecin alone was downregulated by the combination of ehrysin and eamptothecin. CON- CLUSION Chrysin could sensitize the apoptosis induced by cisplatin and camptotheein, and the downregulation of apoptosis inhibitory proteins, which were regulated by NF-KB and augmented by cisplatin and camptothecin, played an important role in the sensitization.

关 键 词：白杨素 顺铂 喜树碱 凋亡 核因子-KB 

分 类 号：R286.91[医药卫生—中药学]