摘要
目的考察昆布多糖硫酸酯LAMS-2对人结肠癌细胞LOVO凋亡的影响及作用途径研究。方法采用四甲基偶氮唑蓝(MTT)法检测LOVO细胞的增殖,Hoechst 33258荧光染色观察细胞形态,流式细胞仪检测细胞内活性氧水平、线粒体通透性转换孔活性和线粒体膜电位,激光共聚焦显微镜观察细胞内钙离子水平,Western blot法检测凋亡蛋白Cyt-C、caspase-9和caspase-3的表达,比色法检测caspase-9和caspase-3的活性。结果昆布多糖硫酸酯LAMS-2作用于LOVO细胞后,能显著抑制细胞的增殖,诱导细胞凋亡,细胞出现明显的凋亡特征形态,引起细胞内活性氧增多,钙离子含量增加,细胞线粒体通透性转变孔开放,线粒体膜电位降低,胞质内Cyt-C蛋白表达量增加,caspase-9和caspase-3的蛋白表达和活性都显著升高,较阴性对照组和昆布多糖组都显著增加。结论昆布多糖硫酸酯LAMS-2可显著抑制人结肠癌LOVO细胞的增殖,并可通过线粒体途径诱导LOVO细胞发生凋亡。
OBJECTIVE To investigate the effects and mechanisms of LAMS-2 on the proliferation and apoptosis of human colon cancer cell line LOVO. METHODS LOVO cells were treated with different concentrations of LAMS-2,3-（ 4,5-Dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide（ MTT） assay and morphology observation were performed with Hoechst 33258 dye to determine the effects of LAMS-2 on the proliferation and apoptosis of LOVO cells. Flow cytometry（ FCM） was used to detect the level of reactive oxygen species（ ROS）,mitochondrion permeability transition pore（ MPTP） and mitochondrial membrane potential（ △ψm）. Laser scanning confocal microscope（ LSCM） was used to analyze intracellular calcium ion concentration,Western blot was performed to analyze the expressions of Cyt-C,caspase-9 and caspase-3. Spectrophotometer was applied to quantify the activity of caspase-9 and caspase-3. RESULTS LAMS-2 inhibited LOVO cell proliferation and induced apoptosis,the apoptosis morphology was obvious.LAMS-2 treatment increased the intracellular level of ROS and Ca^（2 ＋）; activated intracellular MPTP and decreased △ψm; it also induced the release of Cyt-C and the activation of caspase-9 and caspase-3,increased the activity of caspase-9 and caspase-3. CONCLUSION LAMS-2 can effectively inhibit the proliferation of LOVO cells and induce apoptosis through a mitochondria-mediated pathway.
作者
汲晨锋
季宇彬
JI Chen-feng;JI Yu-bin(Engineering Research Center of Natural Anticancer Drugs,Ministry of Education;Center of Research on Life Science and Environmental Science,Harbin University of Commerce,Harbin 150076,China)
出处
《中国药学杂志》
CAS
CSCD
北大核心
2018年第16期1373-1380,共8页
Chinese Pharmaceutical Journal
基金
黑龙江省自然科学基金项目资助(H2015088)
黑龙江省博士后特别资助项目资助(LBH-TZ0516)
哈尔滨市科技局科技创新人才项目资助(2016RQYXJ014)
哈尔滨商业大学科研团队计划项目资助(2016TD002)
