糖基化终末产物对人牙周膜干细胞和骨髓间充质干细胞骨向分化能力影响的比较研究

Effects of advanced glycation end products on the osteogenic differentiation of human bone marrow mesenchymal stem cells and human periodontal ligament stem cells

目的:观察糖基化终末产物(AGEs)对人牙周膜干细胞(hPDLSCs)及人颌骨骨髓间充质干细胞(hBMMSCs)骨向分化能力的影响。方法:组织块法和有限稀释法克隆化培养hPDLSCs及hBMMSCs,流式细胞仪检测两种细胞表型分子表达,取第3代细胞分别在10μg/mL AGEs刺激下进行矿化诱导,于诱导的21 d,茜素红染色、十六烷基吡啶定量观察钙结节形成情况、诱导7 d,碱性磷酸酶染色观察ALP活性、实时定量聚合酶链反应(real time PCR)和Western blot检测成骨相关基因及蛋白表达情况。结果:流式细胞仪显示两种细胞均阳性表达CD44、CD146、stro-1、CD90。成骨诱导21 d后茜素红染色和定量分析显示,hPDLSCs AGEs组矿化能力低于对照组;而hBMMSCs AGEs组矿化能力高于对照组(P<0.05)。成骨诱导7 d后,ALP染色显示,两种细胞ALP活性变化趋势与茜素红定量分析相同;RT-PCR检测hPDLSCs中ALP、Runx-2、Col-1、Runx-2mRNA表达水平均低于对照组,而hBMMSCs中各基因均高于对照组(P<0.05)。Western blot检测显示,各组Runx-2蛋白表达趋势与RT-PCR结果趋势相同。结论:AGEs抑制hPDLSC的骨向分化,促进hBMMSC骨向分化。

AIM： To investigate the effects of advanced glycation end products （AGEs） on the osteogenic differentiation of human periodontal ligament stem cells （hPDLSCs） and human bone marrow mesenchymal stem cells （hBMMSCs）. METHODS： PDLSCs and BMMSCs were cultured and cloned. Flow cytometery were employed to ex- amine the surface marker of the cells, hPDLSCs and hBMMSCs of passage3 were induced by 10 μg/mL AGEs. The os- teogenic differentiation capacity of the cells were evaluated by alizarin red staining, ALP staining and real-time RT - PCR. RESULTS： CD44, CD146, stro-1 and CI）90 were expressed in AGE-BSA induced hPDLSCs and hBMMSCs. After 21- day induction, the mineralization nodules were fewer in AGEs induced hPDLSCs than in the controls（P 〈 O. 05 ） , but were more in AGEs induced hBMMSCs than in the controls （P 〈 0.05）. After 7-day induction ALP, Runx-2 Col-1 and Runx-2 mRNAs were lower in AGEs induced hPDLSCs than in the controls（P 〈 0.05）, but were higher in hBMMSCs than in the controls（P 〈0.05）. CONCLUSION：AGEs may inhibite the osteogenic differentiation of hPDLSCs and promote the osteogenic differentiation of hBMMSCs.