miR-129-5p靶向HMGB1对胰腺癌细胞凋亡的作用研究

目的:研究miR-129-5p调控HMGB1表达在胰腺癌细胞凋亡中的作用。方法:以未处理的胰腺癌SW1990细胞作为对照组(Control组),用脂质体转染法将mimics-NC(空载体)、miR-129-5p mimics、inhibitor-NC(空载体)、miR-129-5p inhibitor分别转染SW1990细胞作为mimics-NC组、miR-129-5p过表达组(miR-129-5p mimics组)、inhibitor-NC组、miR-129-5p低表达组(miR-129-5p inhibitor组),通过在线靶基因预测网站Target genes预测miR-129-5p与HMGB1是否存在结合位点,双荧光素酶基因报告实验验证miR-129-5p与HMGB1的靶向关系。采用qRT-PCR检测各组细胞中miR-129-5p的表达水平,Western blot法检测HMGB1蛋白及凋亡相关蛋白Caspase-3、Bcl-2表达。Hoechst染色观察细胞凋亡变化。结果:与mimics-NC组及Control组比较,miR-129-5p mimics转染显著上调miR-129-5p水平(P<0.01),抑制HMGB1(P<0.01)、Bcl-2(P<0.05)蛋白表达,促进Caspase-3蛋白表达(P<0.05),促进细胞凋亡;与inhibitor-NC组及Control组比较,miR-129-5p inhibitor转染显著下调miR-129-5p水平(P<0.05),促进HMGB1、Bcl-2蛋白表达(均P<0.05),抑制Caspase-3蛋白表达(P<0.01),抑制细胞凋亡。双荧光素酶报告基因检测结果显示miR-129-5p可抑制野生型HMGB1细胞的荧光活性,靶向调控HMGB1的表达。结论:miR-129-5p通过靶向抑制HMGB1的表达进而促进胰腺癌SW1990细胞凋亡。

The effect of miR-129-5p in pancreatic cancer cells on apoptosis through targeted of HMGB1

Objective:To investigate the role of miR-129-5p in regulating HMGB1 expression in pancreatic cancer cell apoptosis.Methods:The untreated pancreatic cancer SW1990 cells were used as the control group.Mimics-NC(empty vector),miR-129-5p mimics,inhibitor-NC(empty vector)and miR-129-5p inhibitor were transfected into SW1990 cells by liposome transfection method as the mimics-NC group,miR-129-5p overexpression group(miR-129-5p mimics group),inhibitor-NC group and miR-129-5p low expression group(miR-129-5p inhibitor group).The binding site of miR-129-5p and HMGB1 was predicted by online target gene prediction website Target genes,and the targeting relationship between miR-129-5p and HMGB1 was verified by dual luciferase gene report experiment.The expression of miR-129-5p in each group was detected by qRT-PCR,and the expression of HMGB1 protein and apoptosis-related proteins Caspase 3 and Bcl-2 by Western blot.Hoechst staining was used to observe the changes of apoptosis.Results:Compared with the mimics-NC group and control group,miR-129-5p mimics transfection significantly up-regulated miR-129-5p level(P<0.01),inhibited HMGB1(P<0.01)and Bcl-2(P<0.05)protein expression,pro-moted Caspase 3 protein expression(P<0.05),and promoted apoptosis;compared with the inhibitor-NC group and control group,miR-129-5p inhibitor transfection significantly down-regulated miR-129-5p level(P<0.05),promoted HMGB1 and Bcl-2 protein expression(all P<0.05),inhibited Caspase 3protein expression(P<0.01),and inhibited apoptosis.The results of dual luciferase reporter gene assay showed that miR-129-5p could inhibit the fluorescence activity of wildtype HMGB1 cells and target the expression of HMGB1.Conclusion:miR-129-5p promotes the apoptosis of pancreatic cancer SW1990 cells by targeting inhibition of HMGB1 expression.

皂角籽的总生物碱含量测定及体外抑菌活性初步研究

目的:通过对皂角籽总生物碱的体外抑菌活性研究,为充分利用皂角籽资源提供理论依据。方法:料液比为(g:ml)1∶9采用酒精回流提取法。提取时间是1 h,提取两次。采用打孔法考察了皂角籽总生物碱提取物对金黄色葡萄球菌、铜绿假单胞菌、鲍曼不动杆菌、枯草芽孢杆菌、阴沟产气荚膜梭菌、炭疽芽孢杆菌的抑菌性能。结果:皂角籽提取物中总生物碱含量为2.06mg/ml,皂角籽总生物碱对金黄色葡萄球菌和枯草芽孢杆菌、炭疽芽孢杆菌抑菌作用较强,对铜绿假单胞菌和鲍曼不动杆菌抑菌作用较弱,对阴沟产气荚膜梭菌没有抑菌作用。皂角籽总生物碱提取物对金黄色葡萄球菌、铜绿假单胞菌、鲍曼不动杆菌、枯草芽孢杆菌、炭疽芽孢杆菌的最小抑菌浓度为200、400、400、200、200mg/ml。