鸢尾素缓解棕榈酸对骨髓间充质干细胞的成骨抑制

Irisin alleviates palmitic acid-induced osteogenic inhibition in bone marrow mesenchymal stem cells

背景:鸢尾素是肌肉细胞在运动时从跨膜蛋白FNDC5中分离出来的肌因子,具有诱导白色脂肪组织褐变的功能,但是其对脂毒性引起的成骨分化的影响及机制尚不清楚。目的:探究鸢尾素对棕榈酸诱导的骨髓间充质干细胞成骨能力的影响及作用机制。方法:CCK-8检测不同浓度棕榈酸对小鼠骨髓间充质干细胞增殖的影响,以及鸢尾素对棕榈酸存在条件下骨髓间充质干细胞增殖的影响;鸢尾素和棕榈酸预处理24 h后,对小鼠骨髓间充质干细胞成骨诱导分化,在成骨诱导培养第7天进行碱性磷酸酶染色,qRT-PCR检测成骨相关基因的表达,Western blot检测AMPK/BMP2/SMAD信号通路相关蛋白的表达,第21天进行茜素红染色,检测成骨差异。结果与结论:①CCK-8结果提示骨髓间充质干细胞的扩增与棕榈酸的浓度呈反比,但在0.02 mmol/L浓度时,棕榈酸对骨髓间充质干细胞扩增无显著影响,鸢尾素质量浓度在0.1-20μg/L范围内时,不影响骨髓间充质干细胞增殖;②碱性磷酸酶染色及茜素红染色结果显示,棕榈酸抑制骨髓间充质干细胞成骨分化能力,鸢尾素能改善棕榈酸诱导的骨髓间充质干细胞成骨抑制,qRT-PCR结果显示,棕榈酸可以引起成骨相关基因下调,而鸢尾素可以抑制这一趋势;③Western bolt结果显示,相较于单纯棕榈酸干预组,鸢尾素处理后增强了骨髓间充质干细胞中AMPK/BMP2/SMAD信号通路传导。该研究发现了鸢尾素能够改善棕榈酸预处理骨髓间充质干细胞的成骨分化能力并提出了具体的机制可能是由AMPK/BMP/SMAD信号通路介导的。

BACKGROUND:Irisin,a myokine isolated from the transmembrane protein FNDC5 by muscle cells during exercise,has the function of inducing the browning of white adipose tissue,but its effect on lipotoxicity-induced osteogenic differentiation and the mechanism is unclear.OBJECTIVE:To investigate the effect of irisin on the osteogenic ability of palmitic acid-induced bone marrow mesenchymal stem cells and the mechanism of action.METHODS:CCK-8 assay was used to detect the effect of different concentrations of palmitic acid on the proliferation of mouse bone marrow mesenchymal stem cells and the effect of irisin on the proliferation of mouse bone marrow mesenchymal stem cells in the presence of palmitic acid.After pretreatment with irisin and palmitic acid for 24 hours,osteogenic differentiation of mouse bone marrow mesenchymal stem cells was induced by alkaline phosphatase staining as well as qRT-PCR was performed to detect the expression of osteogenesis-related genes on day 7 of osteogenic induction culture.The expression of proteins related to the AMPK/BMP2/SMAD signaling pathway was detected by western blot assay.Alizarin red staining was conducted on day 21 to detect osteogenic differences.RESULTS AND CONCLUSION:(1)The CCK-8 assay results suggested that the amplification of bone marrow mesenchymal stem cells was inversely proportional to the concentration of palmitic acid,but at 0.02 mmol/L concentration,palmitic acid had no significant effect on the amplification of bone marrow mesenchymal stem cells,and irisin did not affect the proliferation of bone marrow mesenchymal stem cells when its mass concentration was in the range of 0.1-20μg/L.(2)Alkaline phosphatase staining and alizarin red staining showed that palmitic acid inhibited the osteogenic differentiation ability of bone marrow mesenchymal stem cells.Irisin improved palmitic acid-induced osteogenic inhibition of bone marrow mesenchymal stem cells.qRT-PCR results showed that palmitic acid could cause the downregulation of osteogenic-related genes,and irisin could inhibit this trend.(3)Western blot assay results showed that compared with the palmitic acid intervention group,irisin treatment enhanced AMPK/BMP2/SMAD signal transduction in bone marrow mesenchymal stem cells.It is found that irisin can improve the osteogenic differentiation ability of bone marrow mesenchymal stem cells pretreated with palmitic acid,and proposed that the specific mechanism might be mediated by AMPK/BMP/SMAD signaling pathway.