[1]蒋琳,周鹏飞,王佳,等.BMPs-ERK5信号通路调控人牙周膜干细胞成骨分化的研究[J].陆军军医大学学报(原第三军医大学学报),2016,38(07):718-725.
 Jiang Lin,Zhou Pengfei,Wang Jia,et al.Bone morphogenetic proteins-ERK5 signaling pathway mediates osteogenic differentiation of human periodontal ligament stem cells[J].J Amry Med Univ (J Third Mil Med Univ),2016,38(07):718-725.
点击复制

BMPs-ERK5信号通路调控人牙周膜干细胞成骨分化的研究(/HTML )
分享到:

陆军军医大学学报(原第三军医大学学报)[ISSN:1000-5404/CN:51-1095/R]

卷:
38卷
期数:
2016年第07期
页码:
718-725
栏目:
基础医学
出版日期:
2016-04-15

文章信息/Info

Title:
Bone morphogenetic proteins-ERK5 signaling pathway mediates osteogenic differentiation of human periodontal ligament stem cells
作者:
蒋琳周鹏飞王佳张燕
重庆医科大学附属口腔医院:预防口腔科,重庆市高校市级口腔生物医学工程重点实验室,口腔疾病与生物医学重庆市重点实验室
Author(s):
Jiang Lin Zhou Pengfei Wang Jia Zhang Yan

Department of Preventive Dentistry, Key Laboratory of Oral Biomedical Engineering of Chongqing Higher Education, Chongqing Key Laboratory of Oral Diseases and Biomedical Sciences, Affiliated Stomatological Hospital of Chongqing Medical University, Chongqing, 401147, China

关键词:
骨形态发生蛋白牙周膜干细胞成骨分化细胞外信号调节激酶5
Keywords:
bone morphogenetic protein periodontal ligament stem cells osteogenic differentiation extracellular signal-regulated kinase 5
分类号:
R322.41; R329.21; R329.26
文献标志码:
A
摘要:

目的      比较骨形态发生蛋白(bone morphogenetic proteins, BMP)2、7、9对人牙周膜干细胞﹙human periodontal ligament stem cells, hPDLSCs﹚成骨分化的诱导作用,探讨BMPs-ERK5信号通路在成骨分化中的作用。      方法      首先采用组织块酶消化法分离培养原代hPDLSCs并鉴定,利用腺病毒载体介导的GFP、BMP2、BMP7和BMP9(Ad-GFP、Ad-BMP2、Ad-BMP7、Ad-BMP9)分别感染hPDLSCs,通过早期成骨分化指标碱性磷酸酶活性定性和定量检测、茜素红染色检测晚期成骨分化指标钙盐结节沉积情况、qRT-PCR检测细胞成骨分化相关基因骨钙蛋白和骨桥蛋白mRNA表达,比较发现BMP9对细胞成骨分化的作用最强;采用Ad-BMP9感染hPDLSCs 24 h后,检测BMP9作用PDLSCs后对ERK5磷酸化水平的影响和ERK5信号通路特异性抑制剂BIX02189预处理后BMP9-ERK5通路对hPDLSCs成骨分化的影响。      结果      ①分离培养的hPDLSCs为成纤维细胞样,呈漩涡状生长,抗波形蛋白表达阳性,抗角蛋白表达阴性,具有间充质属性和克隆形成能力。②BMP2、BMP7和BMP9均有促进细胞成骨分化作用,其中3组的碱性磷酸酶定量结果和成骨分化相关基因相对表达量与GFP组比较,差异具有统计学意义(P<0.05),且BMP9效力最强。③Western blot检测显示BMP9增强p-ERK5的表达。而抑制剂BIX02189呈剂量依赖性地抑制p-ERK5的表达。BIX02189预处理细胞后、感染Ad-BMP9的结果显示,BMP9对细胞的促成骨作用较未处理组明显降低,其中BMP9组的碱性磷酸酶定量分析结果与成骨分化相关基因相对表达量与BIX02189处理组比较,差异具有统计学意义(P<0.05)。      结论       BMP9具有较强的促hPDLSCs成骨分化作用,ERK5信号转导途径在BMP9诱导hPDLSCs成骨分化过程中发挥了正向调节作用。

Abstract:

Objective      To determine osteogenic differentiation effects of bone morphogenetic proteins (BMP 2, 7 and 9) on human periodontal ligament stem cells (hPDLSCs), and determine the effect of BMPs-extracellular signal-regulated kinase 5 (ERK5) signaling pathway on osteogenic differentiation.       Methods      hPDLSCs were isolated from fresh periapical tissues and cultured by tissue block and enzyme digestion, and then identified. After hPDLSCs were infected by adenovirus vectors carrying GFP, BMP2, BMP7 and BMP9 respectively (Ad-GFP, Ad-BMP2, Ad-BMP7 and Ad-BMP9), the activity and quantity of alkaline phosphatase were determined for early osteogenic differentiation, and calcium nodule deposition was studied by alizarin red staining for late osteogenic differentiation. And the expression levels of some genes related to osteogenic differentiation and osteopontin were detected by qRT-PCR. After BMP9 was determined as exerting strongest effect on osteogenic differentiation, the phosphorylation of ERK5 was detected after hPDLSCs were intervened with Ad-BMP9 and BIX02189 (inhibitor of ERK5 signaling pathway) for 24 h respectively for the effect of the signaling pathway on osteogenic differentiation.       Results      The isolated and cultured hPDLSCs were fibroblast-likes, in swirl-like arrangement, positive to vimentin while negative to keratin, and showing mesenchymal properties and clone forming ability. Adenovirus vectors Ad-BMP2, BMP7 and BMP9 all had inducing effects on osteogenic differentiation in hPDLSCs, and enhanced the activity of alkaline phosphatase and the mRNA expression of relative genes than Ad-GFP vector (P<0.05), what’s more, adenovirus vector BMP9 had strongest effects. Western blotting showed that BMP9 up-regulated the phosphorylation of ERK5, and BIXO2189 decreased the phosphorylation of ERK5 in a dose-dependent manner. And BIX02189 attenuated BMP9-induced osteogenic differentiation of hPDLSCs. The activity of alkaline phosphatase and mRNA expression levels of relative genes were significantly lower in the BIX02189 groups than the BMP9 group (P<0.05).       Conclusion      BMP9 has strong the osteogenesis induction effect on hPDLSCs. ERK5 signaling pathway exerts a positive regulatory effect on BMP9-induced osteogenic differentiation.

相似文献/References:

[1]刘一涵,刘洪臣,刘文佳.牙周膜干细胞对脐血单个核细胞分化为破骨样细胞的影响[J].陆军军医大学学报(原第三军医大学学报),2012,34(10):1011.
 Liu Yihan,Liu Hongchen,Liu Wenjia.Periodontal ligament stem cells improve peripheral blood mononuclear cells diffe-rentiation into osteoclast-like cells[J].J Amry Med Univ (J Third Mil Med Univ),2012,34(07):1011.
[2]王萍,周玥,王亚平,等.人参皂苷Rg1促进人牙周膜干细胞增殖与成骨分化[J].陆军军医大学学报(原第三军医大学学报),2013,35(15):1566.
 Wang Ping,Zhou Yue,Wang Yaping,et al.Promotion of ginsenoside Rg1 on proliferation and osteogenic potential of human periodontal ligament stem cells[J].J Amry Med Univ (J Third Mil Med Univ),2013,35(07):1566.

更新日期/Last Update: 2016-04-01