唐古特大黄多糖对过氧化氢诱导的肠上皮细胞凋亡的保护作用及其可能机制  

Protective effect of Rheum tanguticum polysaccharide against H_2O_2 induced IEC-6 cell apoptosis and its possible mechanism

作　　者：刘琳娜[1] 张峰[2] 梅其炳[2] 张琰[1] 刘新友[1] 

机构地区：[1]第四军医大学唐都医院药剂科,陕西西安710038 [2]第四军医大学药学院药理学教研室,陕西西安710032

出　　处：《中国药理学通报》2010年第7期877-881,共5页Chinese Pharmacological Bulletin

基　　金：国家自然科学基金青年科学基金资助项目(No30300450)

摘　　要：目的探讨唐古特大黄多糖(Rheum tanguticumpoly-saccharide,RTP)组分1(RTP1)对过氧化氢(H2O2)诱导的肠上皮细胞IEC-6凋亡的保护作用及其可能的机制。方法以不同浓度的RTP1(10、30、100 mg.L-1)预处理体外培养的IEC-6细胞后,加入H2O2诱导IEC-6细胞凋亡,MTT法检测细胞活力,激光共聚焦显微镜荧光染色法检测细胞内活性氧的产生,DNA琼脂糖凝胶电泳、吖啶橙染色及流式细胞仪检测细胞凋亡的发生,Western blot测定Caspase-3酶活性。结果 100μmol.L-1H2O2处理细胞2 h明显降低细胞存活率并诱导细胞凋亡,凋亡率为31.3%,细胞内活性氧水平及Caspase-3的活性明显升高,不同浓度(10、30、100 mg.L-1)RTP1预处理细胞24 h,可明显提高细胞存活率,有效抑制DNAladder的发生,流式细胞仪检测凋亡率(30、100 mg.L-1)分别降低至24.4%和21.5%,Caspase-3酶活性降低,并呈现一定的剂量依赖性。结论 RTP1可明显抑制H2O2诱导的IEC-6细胞凋亡,其细胞保护作用可能与降低细胞内活性氧水平和抑制Caspase-3活性相关。Aim To investigate the protective effect of RTP1,one of the polysaccharide isolated from Rheum tanguticum,on H2O2 induced apoptosis in IEC-6 cells and its possible mechanism.Methods H2O2（100 mmol·L-1）was used to induce IEC-6 cell apoptosis.Different doses（10,30,100 mg·L-1）of RTP1 were administrated before H2O2 was added into IEC-6 cell culture.Cell viability was observed by MTT assay.Reactive oxygen species were measured with laser scanning confocal microscopy（LSCM）.DNA content and percentage of apoptosis were assayed by DNA agarose gel electrophoresis,acridine orange staining and flow cytometry.The activation of Caspase-3 was detected with Western blot analysis.Results Following treatment with H2O2 for 2 h,H2O2 induced a significant decrease in cell viability,while DNA ladder was observed and apoptosis percentage was as high as 31.3%.Accumulation of intracellular ROS and increase in Caspase-3 activity were also detected.Pretreatment with RTP1 for 24 h exhibited cytoprotective effects in a dose-dependent manner.RTP1 obviously enhanced cell viability,reduced formation of DNA ladder and significantly reduced the number of cells labeled with Annexin V.The percentage of apoptosis/necrosis cells was markedly decreased to 24.4% and 21.5%,respectively.LSCM showed that RTP1 attenuated the accumulation of ROS.The significant decrease in Caspase-3 activity was detected.Conclusion RTP1 has cytoprotective capacity to antagonize H2O2-induced IEC-6 cell apoptosis and injury,and this effect may be related to decrease ROS and inhibit Caspase-3 activity

关 键 词：唐古特大黄多糖 细胞凋亡 过氧化氢 IEC-6细胞 活性氧 半胱氨酸天冬氨酸蛋白酶3 

分 类 号：R284.1[医药卫生—中药学] R329.25[医药卫生—中医学]