摘要
目的观察梓醇对骨髓间充质干细胞(bone marrow mesenchymal stem cells,BMSCs)向成骨细胞分化的影响,并从Hedgehog信号角度探讨其可能的作用机制。方法体外分离培养BMSCs,并通过流式细胞仪对细胞加以鉴定。pNPP法筛选梓醇最佳促BMSCs骨向分化的浓度,以最佳梓醇浓度干预BMSCs的成骨性分化。后续实验分4组:对照组、梓醇组、经典组和联合组(梓醇组+经典组)。酶标法检测碱性磷酸酶(ALP)活性,茜素红染色法检测矿化结节数目,QPCR法检测Shh mRNA、Ihh m RNA水平,Western blot法检测Ptch1、Smo及Gli1蛋白的表达。结果梓醇促BMSCs骨向分化的最佳浓度为1×10-4mol/L;与对照组比较,梓醇组的ALP活性显著增强,矿化结节数目显著升高,Shh mRNA水平显著上升,Ptch1、Smo及Gli1蛋白表达均显著升高(P<0.05);与经典组比较,梓醇组的ALP活性、矿化结节数目、Shh mRNA水平、Ptch1、Smo和Gli1蛋白表达均低于经典组(P<0.05);各组中Ihh mRNA水平无明显变化。结论梓醇能够促进BMSC向成骨分化,可能与上调Hedgehog信号传导通路相关分子有关。
This study was performed to evaluate the effects of Catalpol on osteogenic differentiation of bone marrow mesenchymal stem cells(BMSCs)and its mechanisms via the hedgehog signaling pathway.BMSCs was isolated by vitro separation and cell surface antigens were detected by flow cytometry.pNPP method was used to measure the Alkaline phosphatase(ALP)activity for confirmation of the best effective concentration of catalpol.The cells in next experiment was divided into four groups:control group,catalpol group,classic group and combination group.The activity of ALP was detected by enzyme linked immunosorbent assay;the number of calcium nodes was detected by alizarin red staining;the mRNA expression levels of Shh and Ihh were detected by Real-time PCR;the protein expressions of Ptch1,Smo and Gli1 were detected by Western blotting.Data showed that the most effective concentration of catalpol on the osteogenic differentiation of BMSCs was 1×10-4 mol/L.Compared with control group,the ALP activity,calcium nodes,Shh mRNA expression,the protein expressions of Ptch1,Smo and Gli1 were significantly increased in catalpol group(P<0.05).Compared with classic group,the ALP activity,calcium nodes,Shh mRNA expression,the protein expressions of Ptch1,Smo and Gli1 were decreased in catalpol group(P<0.05).Ihh mRNA expression showed no different in each groups.Taken together,catalpol can promote osteogenic differentiation of BMSCs,which may due to upregulation of Hedgehog signaling pathway molecule.
作者
赖满香
廖利平
林基伟
谭玮璐
孙晓生
LAI Manxiang;LIAO Liping;LIN Jiwei;TAN Weilu;SUN Xiaosheng(Basic Medical College,Guangzhou University of Chinese Medicine,Guangzhou 510006,China;Guangdong Food and Drug Vocational College,Guangzhou 510520,China;Shenzhen Municipal Health Commission,Shenzhen 518020,China;Department of Health Construction Administration Center,Shenzhen Traditional Chinese Medicine Hospital,Shenzhen 518000,China)
出处
《免疫学杂志》
CAS
CSCD
北大核心
2019年第11期921-926,933,共7页
Immunological Journal
基金
广东省医学科学技术研究基金(B2015063)
