miR-1290对肺腺癌细胞增殖、迁移和侵袭能力的影响

Effects of miR-1290 on proliferation,migration and invasion abilities of lung adenocarcinoma cells

目的探究miR-1290对肺腺癌细胞增殖、迁移和侵袭能力的影响。方法回顾性收集2018年3月至2019年6月海南省人民医院收治的45例患者,取肺腺癌组织及癌旁组织,采用荧光定量聚合酶链式反应(PCR)检测组织中miR-1290的表达。将肺腺癌细胞随机分为对照组、miR-1290 mimic组和miR-1290 inhibitor组,通过脂质体2000试剂盒,分别以无意义序列、miR-1290 mimic、miR-1290 inhibitor转染细胞。分别采用细胞计数试剂盒(CCK-8)、Transwell小室、划痕实验检测细胞增殖、迁移和侵袭能力。蛋白质印迹法检测Cyclin D1、p27、基质金属蛋白酶(MMP-2和MMP-9)蛋白的表达。经在线生物信息学和双荧光素酶报告基因检测miR-1290的靶基因,荧光定量PCR和蛋白质印迹法检测miR-1290靶基因的表达。结果与癌旁组织比较,肺腺癌组织中miR-1290表达明显增加,差异有统计学意义(P<0.05)。与对照组比较,miR-1290 mimic组miR-1290表达明显增加,miR-1290 inhibitor组miR-1290表达明显降低,差异有统计学意义(P<0.05)。培养48 h时,与对照组比较,miR-1290 mimic组细胞吸光度值、侵袭细胞数及划痕“愈合”率明显增加,miR-1290 inhibitor组细胞吸光度值、侵袭细胞数及划痕“愈合”率明显降低,差异有统计学意义(P<0.05)。与对照组比较,miR-1290 mimic组细胞周期蛋白Cyclin D1、MMP-2和MMP-9蛋白表达量明显增加、p27蛋白表达明显降低,miR-1290 inhibitor组Cyclin D1、MMP-2和MMP-9蛋白表达量明显降低、p27蛋白表达明显增加,差异有统计学意义(P<0.05)。双荧光素酶报告基因检测显示,与对照组比较,共转染INPP4B-WT、miR-1290 mimic细胞中荧光表达量明显降低(P<0.05);与对照组比较,miR-1290 mimic组二型磷脂酰肌醇4磷酸酶(INPP4B)mRNA和蛋白表达量明显降低,miR-1290 inhibitor组INPP4B mRNA和蛋白表达量明显增加(P<0.05)。结论miR-1290在肺腺癌组织中表达上调,miR-1290可能靶向抑制INPP4B水平,促进肺腺癌细胞的增殖、侵袭和迁移。

Objective To explore the effects of miR-1290 on the proliferation,migration and invasion abilities of lung adenocarcinoma cells.Methods From March 2018 to June 2019,45 patients admitted to Hainan Provincial People's Hospital were retrospectively collected,and lung adenocarcinoma tissues and paracancerous tissues were collected.The expression of miR-1290 in the tissues was detected by fluorescence quantitative polymerase chain reaction(PCR).Lung adenocarcinoma cells were randomly divided into control group,miR-1290 mimic group and miR-1290 inhibitor group.The cells were transfected with nonsense sequence,miR-1290 mimic and miR-1290 inhibitor through liposome 2000 kit.The proliferation,migration and invasion abilities of cells were detected by cell counting kit(CCK-8),Transwell chamber and scratch test.Cyclin D1,p27,matrix metalloproteinase(MMP-2 and MMP-9)protein expression was detected by Western blotting.The target gene of miR-1290 was detected by online bioinformatics and dual luciferase reporter gene.The expression of miR-1290 target gene was detected by fluorescence quantitative PCR and Western blotting.Results Compared with para-carcinomatissues,the expression of miR-1290 in lung adenocarcinoma tissues was significantly increased(P<0.05).Compared with control group,the expression of miR-1290 in miR-1290 mimic group was significantly increased,while in miR-1290 inhibitor group was significantly decreased(P<0.05).After 48 h of culture,compared with control group,cell absorbancy,number of invaded cells and scratch healingrate in miR-1290 mimic group were significantly increased,while while in miR-1290 inhibitor group were significantly decreased(P<0.05).Compared with control group,expression levels of Cyclin D1 and matrix metalloproteinases(MMP-2,MMP-9)in miR-1290 mimic group were significantly increased,while expression of p27 protein was significantly decreased.The expression levels of Cyclin D1,MMP-2 and MMP-9 in miR-1290 inhibitor group were significantly decreased,while expression of p27 protein was significantly increased(P<0.05).The dual luciferase reporter gene test showed that compared with control group,expression level of fluorescence in cells co-transfected with INPP4B-WT and miR-1290 mimic was significantly decreased(P<0.05).Compared with control group,expression levels of inositol polyphosphate 4-phosphatase type II(INPP4B)mRNA and protein in miR-1290 mimic group were significantly decreased,while which were significantly increased in miR-1290 inhibitor group(P<0.05).Conclusion The expression of miR-1290 is up-regulated in lung adenocarcinoma tissues.The miR-1290 may promote the proliferation,invasion and migration of lung adenocarcinoma cells by inhibiting INPP4B level.