CAJ | 학술논문

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-2 mRNA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、120 μmol/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低剂量组=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高剂删量组=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.
목적 연구고량강소대인간암Hep3B세포조망적영향병탐토상관궤제. 방법 체외배양인간암Hep3B세포,경고량강소처리후,RT-PCR화Western blot검측Bcl-2 mRNA화단백적표체수평,MTT법측정세포생장억제솔,Annexin V-FITC/PI쌍표기법측정세포조망솔,류식세포의분석선립체막전위,Western blot검측Caspase-3、9급Cytochrome C단백적표체수평. 결과 고량강소사Hep3B세포중Bcl-2 mRNA화단백적표체수평명현하강.불동제량적고량강소(40、80、120 μmol/L)대세포생장억제솔분별위6.38％±1.32％,21.58％±1.97％화43.18％±3.89％,명현고우대조조(t저제량조=13.01,t중제량조=15.12,t고제량조=14.79,균P＜0.01);세포조망솔분별위21.58％±1.97％,34.18％±3.89％화43.18％±3.89％,명현고우대조조(분별t저제량조=24.67,t중제량조=32.35,t고제량조=25.89,균P ＜0.01);선립체막전위록색형광단백표체백분비분별위18.93％±2.3％,31.11％±2.67％화46.06％±2.95％,명현고우대조조(분별t저제량조=16.70,t중제량조=31.38,t고제량조=48.15,균P ＜0.01);Caspase-3、9화포질내Cytochrome C표체수평명현고우대조조(Caspase-3:분별t저제량조 =11.94,t중제량조=10.18,t고제산량조=18.82,균P＜0.01;Caspase-9:분별t저제량조=15.11,t중제량조=20.41,t고제량조=21.25,균P＜0.01;Cytochrome C:t저제량조=15.11,t중제량조=28.47,t고제량조=16.01,P＜0.01),이선립체중Cytochrome C표체수평명현저우대조조(분별t저제량조=16.70,t중제량조=12.00,t고제량조=27.61,균P＜0.01). 결론 고량강소가하조Bcl-2 mRNA화단백표체촉진인간암Hep3B세포조망,기작용궤제가능여선립체도경상관.
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; Mitochondrial membrane potential was analyzed by flow cytometry; Western blot was used to detect protein expression of Caspase-3,9 and Cytochrome 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 alpinetin.After treated with different dosages of alpinetin (40,80,120 μmol/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 (tlowdose =13.01,tmiddle dose =15.12,thighdose =14.79,average P ＜ 0.01) ; the expression of mitochondrial membrane potential green fluorescence protein (GFP) were 18.93％ ± 2.3％,31.11％ ± 2.67％ and46.06％ ± 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 andCytochrome C in cytoplasm significantly was higher than those in control group(Caspase-3:llow 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,thish dose =21.25,average P ＜0.01; Cytochrome C:tlow dose =15.11,tmiddle dose =28.47,thigh dose =16.01,average P ＜ 0.01).while that Cytochrome C in mitochondria significantly lower than those in control group (tlow dose =16.70,tmiddle dose =12.00,thighdose =27.61,average P ＜ 0.01).Conclusions Alpinetin promotes apoptosis of human hepatic cancer cells Hep3B by down-regulating Bcl-2,probably through mitochondrial pathway.