微小RNA-17在糖基化终末产物刺激下人牙周膜干细胞骨向分化过程中的调控作用

摘要

Objective This study aims to detect microRNA-17(mir-17) expression on the osteogenic differentiation of advanced glycation end products (AGEs)-stimulated hunman periodontal ligament stem cells (HPDLSCs) and to analyze the influence of these cells on this process. Methods HPDLSCs were isolated using limited dilution technique. After osteogenic differentiation occurred, different time points of mir-17 expression in the experimental groups were detected by real time polymerase chain reaction (PCR). The mir-17 overexpression and inhibition were evaluated using cell transfection technique. Differences in gene expressions were detected by real time PCR; differences in protein expressions were analyzed by Western blot. Results The mir-17 expression was reduced after osteogenic differentiation occurred at 3, 7, and 14 d compared with that in the control group (P<0.05). The expression levels of bone sialoprotein(BSP), Runt-related transcription factor-2 (Runx-2) and alkaline phosphatase (ALP) in the experimental groups were lower than those in the mimic control group when mir-17 expression increased. In addition, the protein expression levels of Runx-2 in the experimental groups were lower than those in the control group. The expression levels of BSP, Runx-2 and ALP in the experimental groups were higher than those in the inhibitor control group when mir-17 expression decreased. Likewise, the protein expression levels of Runx-2 in the expe-rimental groups were higher than those in the control group. Conclusion AGEs inhibit the osteogenic differentiation of HPDLSCs by affecting mir-17 expression.%目的：检测微小RNA-17（mir-17）在糖基化终末产物（AGEs）刺激下人牙周膜干细胞（HPDLSCs）骨向分化过程中的表达，并分析其对该过程的影响。方法体外组织块法和有限稀释法克隆化培养HPDLSCs。实时定量聚合酶链反应（real time PCR）检测实验组细胞在骨向分化过程中不同时间点mir-17的表达；采用细胞转染技术过表达和抑制mir-17的表达，real time PCR和Western blot分别检测转染前后其成骨基因mRNA水平和蛋白水平的表达情况。结果成骨诱导3、7、14 d后，对照组和实验组mir-17的表达均下调，差异有统计学意义（P<0.05）。上调mir-17后，与对照组相比，实验组骨涎蛋白（BSP）、碱性磷酸酶（ALP）、Runt相关转录因子-2（Runx-2）mRNA表达水平以及Runx-2蛋白水平均明显降低；下调mir-17后，实验组BSP、ALP、Runx-2 mRNA表达水平以及Runx-2蛋白水平均高于对照组。结论 AGEs通过影响HPDLSCs骨向分化过程中mir-17的表达从而抑制了HPDLSCs的骨向分化。