摘要
目的探讨骨形态形成蛋白2(bone morphogenetic protein2,BMP2)-Smad1/5及p38MAPK信号通路在胰岛素样生长因子1(insulin-like growth factor 1,IGF1)介导的促犬上颌窦黏膜干细胞(maxillary sinus membrane stem cells,MSMSCs)成骨分化中的作用。方法构建表达胰岛素样生长因子1(insulin-like growth factor1,IGF1)基因的重组腺病毒载体(recombinant adenovirus,rAdv)Ad-IGF1。感染Ad-IGF1的犬上颌窦黏膜干细胞,经成骨诱导培养后,qRT-PCR和Western blot检测BMP-Smads信号通路中重要信号蛋白Smad1/5的磷酸化水平和BMP2蛋白的表达;免疫组化观察磷酸化Smad1/5核转位情况;qRT-PCR及Western blot检测BMP-Smads通路抑制剂Noggin和p38MAPK信号通路抑制剂SB203580对IGF1介导的促犬MSMSCs成骨分化的影响。结果成功构建IGF1基因表达重组腺病毒载体Ad-IGF1;感染Ad-IGF1的犬MSMSCs,经成骨诱导培养后,Smad1/5的磷酸化水平和BMP2蛋白的表达升高,IGF1可促使Smad1/5核转位;BMP-Smads信号通路抑制剂Noggin可抑制Smad1/5的磷酸化,降低成骨标志物Runx2、OPN和ALP mRNA的表达,钙结节形成减少。p38MAPK信号通路抑制剂SB203580不能降低Ad-IGF1犬MSMSCs的p38磷酸化水平,亦不能降低成骨标志物Runx2、OPN和ALP mRNA的表达。结论犬MSMSCs成骨分化过程中,IGF1通过经典的Smads蛋白依赖性信号转导通路BMP2-Smad1/5促进成骨,而Smads蛋白非依赖性信号转导通路p38MAPK在IGF1介导的犬MSMSCs成骨过程中可能并不发挥作用。
ObjectiveTo investigate the role of the bone morphogenetic protein 2(BMP2)-Smad1/5 and p38 MAPK signaling pathways in the osteogenic differentiation of MSMSCs by insulin-like growth factor 1(IGF1).MethodsA recombinant adenovirus(RAD) and IGF1 expressing IGF1 gene were constructed.After osteogenic induction,qRT-PCR and Western blot were used to detect the phosphorylation level of Smad1/5 and the expression of the BMP-2 protein in the BMP-Smad signaling pathway;immunohistochemistry was used to observe the nuclear translocation of Smad1/5;qRTPCR and Western blot were used to detect IGF with Noggin and SB203580,inhibitors of the p38 MAPK signaling pathway 1-mediated osteogenic differentiation of MSMSCs.ResultsThe recombinant IGF1 adenovirus was constructed successfully.MSMSCs were cultured in inductive medium after infection with different concentrations of Ad-IGF1,and then,the protein levels of BMP2 and p-Smad1/5 increased.IGF1 can also induce nuclear translocation of Smad1/5.In addition,Noggin significantly reduced the phosphorylation level of Smad1/5 and the formation of mineralized nodules in the MSMSCs.The mRNA levels of Runx2,OPN and ALP also decreased.In contrast,SB203580 decreased neither the phosphorylation level of p38 nor the mRNA expression of Runx2,OPN and ALP in the Ad-IGF1 MSMSCs.Conclusion IGF1 can promote the osteogenic differentiation of MSMSCs via the BMP2-Smad1/5 signaling pathway.In contrast,IGF1 may not promote the osteogenic differentiation of MSMSCs via the p38 MAPK signaling pathway.
作者
廖春晖
李明飞
叶金梅
彭伟
陈松龄
LIAO Chunhui;LI Mingfei;YE Jinmei;PENG Wei;CHEN Songling(Department of Or⁃thodontics,Guangzhou Women and Children′s Medical Center,Guangzhou 510000,China;Department of Stomatol⁃ogy,The First Affiliated Hospital,SunYat⁃sen University,Guangzhou 510080,China)
出处
《口腔疾病防治》
2020年第1期16-23,共8页
Journal of Prevention and Treatment for Stomatological Diseases
基金
广东省自然科学基金项目(2018A030313173)
国家自然科学基金项目(81371111)
广州市妇女儿童医疗中心儿科研究所内部科研基金项目(YIP-2018-017)
