间充质干细胞抗肝脏纤维化作用及其机制的探索研究
发布时间：2018-08-13 21:04
【摘要】：间充质干细胞(Mesenchymal stem cells, MSCs)由于其多向分化潜能、免疫调节和多因子分泌功能等诸多特性而成为干细胞研究领域的热点。近年来,越来越多研究显示了MSCs在组织修复和疾病治疗方面的应用潜能。本研究重点探讨了小鼠骨髓MSCs (mBM-MSCs)的抗肝脏纤维化作用,以及其中的细胞和分子机制。 首先我们证实了mBM-MSCs移植能向损伤肝脏迁移和整合,并且通过抑制纤维化过程中肝星状细胞活化,有效降低小鼠肝脏纤维化程度。随后在全基因芯片鉴定肝脏损伤相关因子时,我们发现Delta-likel (Dlkl)在损伤肝脏中急剧表达,提示其在肝脏损伤过程中可能发挥一定作用。因此我们对Dlkl在肝脏纤维化过程中的作用和机制做了进一步探讨。体内外实验证实了Dlkl促肝脏纤维化的重要作用。研究结果提示肝损伤过程中实质细胞表达Dlkl并通过旁分泌的方式作用于肝星状细胞,促进肝星状细胞活化,其中50kDa的游离片段起主要作用,而非25kDa的小片段。 进一步研究发现mBM-MSCs移植显著抑制肝纤维化过程中Dlkl的表达,而且主要在mRNA水平。免疫荧光双标以及后续"rescue"实验进一步证实了mBM-MSCs对Dlk1表达的抑制在其抗纤维化过程中起关键性作用。随后体外共培养等一系列实验表明,mBM-MSCs通过分泌因子调控Dlk1表达,而成纤维细胞生长因子2(FGF2)可能是其中重要因子之一。单独使用18kDaFGF2在体内外也能有效抑制Dlk1表达和肝星状细胞活化、降低肝脏纤维化,从侧面验证了FGF2在mBM-MSCs调控Dlk1过程中的作用,并提示了该因子抗肝脏纤维化新功能。 综上所述,本研究证实了小鼠骨髓间充质干细胞移植的抗肝脏纤维化效果,并展示了其中相关细胞和分子机制：肝实质细胞接受损伤信号刺激表达Dlkl并通过旁分泌方式促进肝星状细胞活化,参与肝脏纤维化发生；mBM-MSCs移植以Dlk1为靶点,通过分泌FGF2抑制其表达而降低肝星状细胞活化水平,最终发挥抗肝脏纤维化功效。
[Abstract]:Mesenchymal stem cell (Mesenchymal stem cells, MSCs) has become a hotspot in stem cell research due to its multi-differentiation potential, immunomodulation and multi-factor secretory function. In recent years, more and more studies have shown the potential of MSCs in tissue repair and disease treatment. The aim of this study was to investigate the anti-hepatic fibrosis effect of mouse bone marrow MSCs (mBM-MSCs) and its cellular and molecular mechanisms. First, we confirmed that mBM-MSCs transplantation can migrate and integrate into the injured liver, and reduce the degree of liver fibrosis by inhibiting the activation of hepatic stellate cells during the process of fibrosis in mice. Subsequently, in the identification of liver injury related factors by whole gene chip, we found that Delta-likel (Dlkl) was expressed rapidly in the injured liver, suggesting that it may play a role in the process of liver injury. Therefore, we further explore the role and mechanism of Dlkl in the process of liver fibrosis. In vivo and in vitro experiments confirmed the important role of Dlkl in promoting liver fibrosis. The results suggest that parenchymal cells express Dlkl and act on hepatic stellate cells in paracrine manner during liver injury, and promote the activation of hepatic stellate cells. Free segments of 50kDa play a major role, not small fragments of 25kDa. Further studies showed that mBM-MSCs transplantation significantly inhibited the expression of Dlkl during hepatic fibrosis, and mainly at the level of mRNA. Immunofluorescence double labeling and subsequent "rescue" experiments further confirmed that the inhibition of Dlk1 expression by mBM-MSCs plays a key role in the process of anti-fibrosis. Subsequently, a series of experiments showed that mBM-MSCs regulated the expression of Dlk1 by secretory factor, and fibroblast growth factor 2 (FGF2) may be one of the important factors. 18kDaFGF2 alone also inhibited the expression of Dlk1 and the activation of hepatic stellate cells in vivo and in vitro, and decreased hepatic fibrosis. The role of FGF2 in the regulation of Dlk1 by mBM-MSCs in vitro and in vivo was verified, and the new function of FGF2 in anti-hepatic fibrosis was suggested. In conclusion, this study confirmed the anti-fibrosis effect of bone marrow mesenchymal stem cell transplantation in mice. The related cellular and molecular mechanisms were also demonstrated: hepatic parenchymal cells were stimulated by injury signal to express Dlkl and promoted hepatic stellate cell activation through paracrine, which was involved in hepatic fibrosis. Dlk1 was used as the target for transplantation of mBM-MSCs. By secreting FGF2 to inhibit its expression, the activation of hepatic stellate cells was reduced, and the effect of anti-hepatic fibrosis was finally played.
【学位授予单位】：浙江大学
【学位级别】：博士
【学位授予年份】：2011
【分类号】：R329
本文编号：2182183
[Abstract]:Mesenchymal stem cell (Mesenchymal stem cells, MSCs) has become a hotspot in stem cell research due to its multi-differentiation potential, immunomodulation and multi-factor secretory function. In recent years, more and more studies have shown the potential of MSCs in tissue repair and disease treatment. The aim of this study was to investigate the anti-hepatic fibrosis effect of mouse bone marrow MSCs (mBM-MSCs) and its cellular and molecular mechanisms. First, we confirmed that mBM-MSCs transplantation can migrate and integrate into the injured liver, and reduce the degree of liver fibrosis by inhibiting the activation of hepatic stellate cells during the process of fibrosis in mice. Subsequently, in the identification of liver injury related factors by whole gene chip, we found that Delta-likel (Dlkl) was expressed rapidly in the injured liver, suggesting that it may play a role in the process of liver injury. Therefore, we further explore the role and mechanism of Dlkl in the process of liver fibrosis. In vivo and in vitro experiments confirmed the important role of Dlkl in promoting liver fibrosis. The results suggest that parenchymal cells express Dlkl and act on hepatic stellate cells in paracrine manner during liver injury, and promote the activation of hepatic stellate cells. Free segments of 50kDa play a major role, not small fragments of 25kDa. Further studies showed that mBM-MSCs transplantation significantly inhibited the expression of Dlkl during hepatic fibrosis, and mainly at the level of mRNA. Immunofluorescence double labeling and subsequent "rescue" experiments further confirmed that the inhibition of Dlk1 expression by mBM-MSCs plays a key role in the process of anti-fibrosis. Subsequently, a series of experiments showed that mBM-MSCs regulated the expression of Dlk1 by secretory factor, and fibroblast growth factor 2 (FGF2) may be one of the important factors. 18kDaFGF2 alone also inhibited the expression of Dlk1 and the activation of hepatic stellate cells in vivo and in vitro, and decreased hepatic fibrosis. The role of FGF2 in the regulation of Dlk1 by mBM-MSCs in vitro and in vivo was verified, and the new function of FGF2 in anti-hepatic fibrosis was suggested. In conclusion, this study confirmed the anti-fibrosis effect of bone marrow mesenchymal stem cell transplantation in mice. The related cellular and molecular mechanisms were also demonstrated: hepatic parenchymal cells were stimulated by injury signal to express Dlkl and promoted hepatic stellate cell activation through paracrine, which was involved in hepatic fibrosis. Dlk1 was used as the target for transplantation of mBM-MSCs. By secreting FGF2 to inhibit its expression, the activation of hepatic stellate cells was reduced, and the effect of anti-hepatic fibrosis was finally played.
【学位授予单位】：浙江大学
【学位级别】：博士
【学位授予年份】：2011
【分类号】：R329
【参考文献】
相关期刊论文 前1条
1 ;Rat bone marrow mesenchymal stem cells differentiate into hepatocytes in vitro[J];World Journal of Gastroenterology;2005年22期
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