摘要
目的研究miR-485-5p和脱氢酶/还原酶3(DHRS3)在骨髓间充质干细胞(BMSCs)增殖和分化中的表达和作用。方法将细胞分为第1转染组、第2转染组、第3转染组和第4转染组,分别用anti-miR-con、anti-miR-485-5p、si-con和si-DHRS3分别转染/共转染BMSCs细胞。转染9 d后,用四甲基偶氮唑盐(MTT)法检测细胞存活率,实时荧光定量-聚合酶链式反应检测各组细胞中miR-485-5p和DHRS3 mRNA的表达,Western blot检测DHRS3、成骨分化蛋白标志物核心结合蛋白因子2(RUNX2)、骨钙蛋白(OCN)和骨形态发生蛋白2(BMP2),增殖蛋白周期蛋白依赖性激酶4(CDK4)和细胞周期蛋白D1(Cyclin D1),Notch受体1(Notch 1)、Notch配体(Jagged1)和Split多毛增强子1(Hes1)蛋白的表达。结果第1转染组、第2转染组、第3转染组和第4转染组的细胞存活率分别为(100.58±10.09)%,(132.50±11.45)%,(135.62±12.66)%和(110.96±9.58)%;DHR3蛋白相对表达量分别为0.31±0.03,0.59±0.05,0.62±0.06和0.43±0.05;RUNX2蛋白相对表达量分别为0.33±0.04,0.49±0.03,0.52±0.06和0.37±0.07;OCN蛋白相对表达量分别为0.35±0.04,0.56±0.06,0.62±0.04和0.39±0.05;BMP2蛋白相对表达量分别为0.32±0.04,0.65±0.06,0.75±0.07和0.41±0.05;和Notch1蛋白相对表达量分别为0.45±0.04,0.74±0.07,0.77±0.06和0.55±0.05。第2转染组与第1转染组比较、第4转染组与第3转染组比较,差异均有统计学意义(均P<0.05)。结论miR-485-5p靶向DHRS3通过Notch信号调控BMSCs细胞增殖和成骨分化。
Objective To investigate the expression and effects of microRNA(miR)-485-5 p and dehydrogenase reductase 3(DHRS3)on proliferation and differentiation of bone marrow mesenchymal stem cells(BMSCs).Methods BMSCs were transfected or co-transfected with anti-miR-con,anti-miR-485-5 p,si-con and si-DHRS3,respectively.The BMSCs were divided into transfection-1 group,transfection-2 group,transfection-3 group and transfection-4 group.On day 9 of directional differentiation culture after transfection,the cell survival rates were determinded by MTT(Thiazolyl Blue Tetrazolium Bromide)assay.Expression levels of miR-485-5 p and DHRS3 mRNA in BMSCs of each group were detected by real-time fluorescence quantitative polymerase chain reaction.The expression levels of bone morphogenic protein markers runt-related transcription factor 2(RUNX2),osteocalcein(OCN)and bone morphogenetic protein 2(BMP2),proliferation protein cycle dependent kinase 4(CDK4)and cell cycle protein D1(Cyclin D1),the Notch receptor 1(Notch1),Notch ligand(Jagged1)and hairy and enhancer of split 1(Hes)1 were measured by Western blot.Results The cell survival rates of transfection-1 group,transfection-2 group,transfection-3 group and transfection-4 group were(100.58±10.09)%,(132.50±11.45)%,(135.62±12.66)%and(110.96±9.58)%,respectively.The relative expression of DHR3 protein of each group were 0.31±0.03,0.59±0.05,0.62±0.06 and 0.43±0.05,respectively.The relative expression of RUNX2 protein was 0.33±0.04,0.49±0.03,0.52±0.06 and 0.37±0.07,respectively.The relative expression of OCN protein was 0.35±0.04,0.56±0.06,0.62±0.04 and 0.39±0.05,respectively.The relative expression of BMP2 protein was 0.32±0.04,0.65±0.06,0.75±0.07 and 0.41±0.05,respectively.The relative expressions of Notch1 and Notch1 were 0.45±0.04,0.74±0.07,0.77±0.06 and0.55±0.05,respectively.There were statistically significant differences between transfection-2 group and transfection-1 group(P<0.05).There were statistically significant differences between transfection-4 group and
作者
丁亚文
冯刚
费雁
王晓娟
答邦明
DING Ya-wen;FENG Gang;FEI Yan;WANG Xiao-juan;DA Bang-ming(Department of Oncology,Wuhan Fourth Hospital,Puai Hospital,Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430033,Hubei Province,China)
出处
《中国临床药理学杂志》
CAS
CSCD
北大核心
2020年第13期1853-1856,共4页
The Chinese Journal of Clinical Pharmacology
基金
国家自然科学基金资助项目(81601934)
武汉市卫生计生科研基金资助项目(WX12C64)。
