高良姜素下调Bcl-2通过线粒体途径促进人肝癌Hep3B细胞凋亡  

Alpinetin down-regulating Bcl-2 promotes apoptosis of human hepatic cancer Hep3B cells

作　　者：唐博[1] 李扬[2] 唐芳[3] 王振冉[4] 聂容荣[5] 余水平[1] 李铂[1] 

机构地区：[1]桂林医学院附属医院肝胆胰外科,广西壮族自治区541001 [2]桂林医学院附属医院肿瘤科,广西壮族自治区541001 [3]桂林医学院附属医院病理科,广西壮族自治区541001 [4]桂林医学院附属医院普外科,广西壮族自治区541001 [5]桂林医学院附属医院中医科,广西壮族自治区541001

出　　处：《中华普通外科杂志》2013年第7期542-545,共4页Chinese Journal of General Surgery

摘　　要：目的研究高良姜素对人肝癌Hep3B细胞凋亡的影响并探讨相关机制。方法体外培养人肝癌Hep3B细胞，经高良姜素处理后，RT—PCR和Western blot检测Bcl-2mRNA和蛋白的表达水平，MTT法测定细胞生长抑制率，Annexin V—FITC／PI双标记法测定细胞凋亡率，流式细胞仪分析线粒体膜电位，Western blot检测Caspase-3、9及Cytochrome C蛋白的表达水平。结果高良姜素使Hep3B细胞中Bcl-2 mRNA和蛋白的表达水平明显下降。不同剂量的高良姜素（40、80、120μmol／L）对细胞生长抑制率分别为6．38％±1．32％，21．58％±1．97％和43．18％±3．89％，明显高于对照组（t低剂器组=13．01，t中剂量组=15．12，t高剂量组=14．79，均P〈0．01）；细胞凋亡率分别为21，58％±1．97％，34．18％±3．89％和43．18％±3．89％，明显高于对照组（分别t低剂量组=24．67，t中剂量组=32．35，t高剂量组=25．89，均P〈0．01）；线粒体膜电位绿色荧光蛋白表达百分比分别为18．93％±2．3％，31．11％±2．67％和46，06％±2．95％，明显高于对照组（分别t低剂量组=16．70，t中剂量组=31．38，t高剂量组=48．15，均P〈0．01）；Caspase-3、9和胞质内Cytochrome C表达水平明显高于对照组（Caspase-3：分别t低剂量组=11．94，t中剂量组=10．18，t高剂量组=18．82，均P〈0．01；Caspase-9：分别t低剂量组=15．11，t中剂量组=20．41，t高剂量组=21．25，均P〈0．01；Cytochrome C：t低剂量组=15．11，t中剂量组=28．47，t高剂量组=16．01，P〈0．01），而线粒体中Cytochrome C表达水平明显低于对照组（分别t低剂量组16．70，t中剂量组=12．00，t高剂量组=27．61，均P〈0．01）。结论高良姜素可下调Bcl-2 mRNA和蛋白表达促进人肝癌Hep3B细胞凋亡，其作用机制可能与线粒体途径相关。Objective To study the effects of alpinetin on apoptosis of Hep3B cells and explore the related mechanism. Methods Hep3B cells were cultured in vitro, treated with alpinetin; RT-PCR and Western blot was used to detect the mRNA and protein levels of Bcl-2; MTT assay was used to detect the cellular growth inhibitory rate; Annexin V-FITC/PI double staining was used to detect the apoptosis rate of cells; Mitochondria] membrane potential was analyzed by flow cytometry; Western blot was used to detect protein expression of Caspase-3, 9 and Cytoehrome C ; the experiment was carried out in four groups ： control group, high dosage of alpinetin group, middle dosage of alpinetin group and low dosage of alpinetin group. Results The expression of Bcl-2 in Hep3B cells were decreased by alpinefin. After treated with different dosages of alpinefin（40,80,120μmoL/L） , the apoptotic inhibitory rate detected by MTT were 6. 38% ±1.32% ,21.58% ±1.97% and 43.18% ±3.89% , significantly higher than those in control group （tlow dose = 13. 01 , tmiddle dose = 15. 12, thigh dose = 14.79, average P 〈 0. 01 ） ; the expression of mitochondria] membrane potential green fluorescence protein （GFP） were 18.93% ± 2. 3% , 31.11% ± 2. 67% and 46.06% ± 2.95% ,significantly higher than those in control group （tlow dose = 16.70, tmiddle dose = 31.38, thigh dose = 48. 15, average P 〈 0. 01 ）. Western blot analysis showed that the expression of Caspase-3, 9 and Cytoehrome C in cytoplasm significantly was higher than those in control group（ Caspase-3 ： tlow dose = 11.94, tmiddle dose = 10. 18, thigh dose = 18. 82, average P 〈 0. 01; Caspase-9： tlow dose = 15. 11, tmiddle dose= 20. 41,thigh dose= 21. 25, average P 〈 0. 01 ; Cytoehrome C ： tlow dose = 15. 11 , tmiddle dose= 28. 47, thigh dose = 16. 01 , average P 〈0.01 ）. while that Cytoehrome C in mitochondria significantly lower than those in control group （tlow dose = 16.70, tmiddle dose = 12. 00, thigh dose = 27. 61 . average P 〈 0.01 ）. Conclusions Alpi

关 键 词：肝肿瘤 高良姜素 细胞凋亡 基因 BCL-2 线粒体 肝 

分 类 号：R735.7[医药卫生—肿瘤]