CAJ | 학술논문

目的：观察miR-335-5p在高糖状态下对成骨细胞增殖、凋亡的影响并探讨其分子机制。方法将MC3T3-E1成骨细胞分为正常对照组、高糖组、agomir-335-5p组及agomir阴性对照组( agomir NC组)。采用四甲基偶氮唑蓝( MTT)比色分析法测定细胞增殖,流式细胞术检测细胞凋亡率,实时定量PCR检测细胞miR-335-5p与DKK1 mRNA的表达,Western印迹检测DKK1及caspase-3的蛋白表达水平。结果与正常对照组相比,高糖组MC3T3-E1成骨细胞miR-335-5p mRNA的表达与细胞增殖明显降低(P<0.05), DKK1、caspase-3的蛋白表达及细胞凋亡显著增加(P<0.05)。与高糖组和agomir NC组相比,agomir-335-5p组miR-335-5p mRNA的表达与细胞增殖显著增加(P<0.05),DKK1、caspase-3的蛋白表达及细胞凋亡显著减少(P<0.05)。结论高糖通过下调miR-335-5p的表达,进而上调DKK1的表达,从而抑制MC3T3-E1成骨细胞的增殖,诱导细胞凋亡。
목적：관찰miR-335-5p재고당상태하대성골세포증식、조망적영향병탐토기분자궤제。방법장MC3T3-E1성골세포분위정상대조조、고당조、agomir-335-5p조급agomir음성대조조( agomir NC조)。채용사갑기우담서람( MTT)비색분석법측정세포증식,류식세포술검측세포조망솔,실시정량PCR검측세포miR-335-5p여DKK1 mRNA적표체,Western인적검측DKK1급caspase-3적단백표체수평。결과여정상대조조상비,고당조MC3T3-E1성골세포miR-335-5p mRNA적표체여세포증식명현강저(P<0.05), DKK1、caspase-3적단백표체급세포조망현저증가(P<0.05)。여고당조화agomir NC조상비,agomir-335-5p조miR-335-5p mRNA적표체여세포증식현저증가(P<0.05),DKK1、caspase-3적단백표체급세포조망현저감소(P<0.05)。결론고당통과하조miR-335-5p적표체,진이상조DKK1적표체,종이억제MC3T3-E1성골세포적증식,유도세포조망。
Objective To investigate the effects of miR-335-5p on the proliferation and apoptosis of osteoblasts which were exposed to high glucose condition, and explore its possible molecular mechanisms. Methods MC3T3-E1 osteoblasts were divided into four groups:control group(5. 5 mmol/L glucose), high glucose group(HG group, 22. 0 mmol/L glucose), agomir-335-5p group(transfected with agomir-335-5p and exposed to 22. 0 mmol/L glucose) , and agomir negative control group( agomir NC group, transfected with agomir negative control and exposed to 22. 0 mmol/L glucose), cultured for 7 days. Cell proliferaton, cell apoptosis, expressions of miR-335-5p and dickkopfhomolog1(DKK1)mRNA,proteinlevelsofDKK1andcysteinylaspartate-specificproteinase-3(caspase-3) were detected using MTT, flow cytometry, quantitative realtime PCR and western blot, respectively. Results Compared with control group, the expression of miR-335-5p mRNA and cell proliferation in HG group were significantly decreased(P<0. 05), while cell apoptosis and the protein levels of DKK1 and caspase-3 were increased significantly(P<0. 05), the expression of DKK1 mRNA did not change (P>0. 05). The miR-335-5p mRNA expression and cell proliferation in agomir-335-5p group were higher than those in HG group and agomir NC group(P<0. 05). However, Cell apoptosis and the protein levels of DKK1 and caspase-3 in agomir-335-5p group were lower than those in HG group(P<0. 05). Conclusion High glucose inhibits the proliferation and induce the apoptosis of MC3T3-E1 osteoblast through decreasing the expression of miR-335-5p and subsequently increasing the DKK1 expression.