摘要
目的:探讨骨形成蛋白8B(BMP8B)对人牙周膜干细胞(PDLSCs)增殖、分化的影响。方法:体外分离培养人PDLSCs,免疫荧光染色检测PDLSCs细胞蛋白基质细胞抗原1(STRO-1)、CD146蛋白。将PDLSCs细胞分为对照组、阴性转染组、BMP8B沉默组和BMP8B过表达组。流式细胞术、茜素红染色和油红O染色对PDLSCs进行鉴定。CCK-8法检测细胞增殖情况。对PDLSCs进行成骨诱导,检测细胞碱性磷酸酶(ALP)活性。茜素红染色检测成骨细胞矿化结节。实时荧光定量聚合酶链反应(qRT-PCR)检测细胞中BMP8B、Osterix、OCN、Runx2 mRNA表达水平,蛋白免疫印迹(WB)检测细胞蛋白表达情况。结果:显微镜下观察PDLSCs细胞为单核、多边形或梭形,具有成骨潜能和成脂潜能。与对照组和阴性转染组相比,BMP8B沉默组细胞增殖情况、ALP活性、茜素红染色程度及BMP8B、Osterix、OCN、Runx2 mRNA和CyclinD1、CDK2、Osterix、OCN、Runx2蛋白表达显著降低(P<0.05)。与对照组和阴性转染组相比,BMP8B过表达组各项指标均明显升高(P<0.05)。对照组和阴性转染组差异无统计学意义(P>0.05)。结论:BMP8B影响PDLSCs细胞的增殖和成骨分化,表明BMP8B在成骨分化中的机制可能是牙周炎治疗研究的新方向。
Objective:To investigate the effects of bone morphogenetic protein 8B(BMP8B)on the proliferation and differentiation of human periodontal ligament stem cells(PDLSCs).Methods:Human PDLSCs were isolated and cultured in vitro,and immunofluorescence staining was used to detect PDLSCs cell protein stromal cell antigen 1(STRO-1)and CD146 protein.The PDLSCs cells were divided into control group,negative transfection group,BMP8B silencing group and BMP8B overexpression group.Flow cytometry,alizarin red staining and oil red O staining were used to identify PDLSCs.Cell proliferation was detected by CCK-8 method.The PDLSCs was osteogenesis inducted,cell alkaline phosphatase(ALP)activity was detected,mineralized nodules in osteoblasts were detected by alizarin red staining,the expression levels of BMP8B,Osterix,OCN,and Runx2 mRNA in cells were detected by real-time fluorescent quantitative polymerase chain reaction(qRT-PCR),and the cell protein expression was detected by western blot(WB).Results:Under the microscope,PDLSCs cells were mononuclear,polygonal or spindle shaped,with osteogenic potential and lipogenic potential.Compared with control group and negative transfection group,cell proliferation,ALP activity,alizarin red staining,mRNA expression of BMP8B,Osterix,OCN,Runx2 and CyclinD1,CDK2,Osterix,OCN,Runx2 were significantly decreased in BMP8B silencing group(P<0.05).Compared with the control group and the negative transfection group,all indexes of BMP8B overexpression group were significantly increased(P<0.05).There was no significant difference between control group and negative transfection group(P>0.05).Conclusion:BMP8B affects the proliferation and osteogenic differentiation of PDLSCs cells,suggesting that the mechanism of BMP8B in osteogenic differentiation may be a new direction for the treatment of periodontitis.
作者
吴帅楠
李召宝
王建琪
郭香君
曹素敏
林秀雅
褚亚辉
WU Shuainan;LI Zhaobao;WANG Jianqi(Cangzhou Central Hospital,Hebei Cangzhou 061000,China)
出处
《河北医学》
CAS
2023年第1期7-12,共6页
Hebei Medicine
基金
河北省卫生健康委医学科学研究课题,(编号:20210670)。
关键词
骨形成蛋白8B
成骨细胞
增殖
分化
Bone morphogenetic protein 8B
Osteoblasts
Proliferation
Differentiation
