心肌再生过程中心外膜分化方向的谱系追踪研究

发布时间：2018-07-31 09:57

【摘要】：目的:利用谱系追踪技术研究新生小鼠心肌再生过程中心外膜的分化方向。方法:对野生型小鼠进行心尖切除模型的构建,通过苏木精-伊红(HE)染色(n=6)观察再生情况。免疫荧光(n=6)和RT-PCR(n=4)检测Wt1(Wilm's Tumor Gene 1)心外膜激活情况。应用Wt1特异标记小鼠(Wt1GFPCre,n=6和Wt1CreERT2;Rosa26RFP,n=6)探究激活的心外膜在心肌再生过程中的激活分化情况。结果:野生型1天龄小鼠切除心尖(apex resection,AR)的10%-15%后,HE染色(n=6)结果证实新生小鼠AR后21天心脏可以完全再生,心脏组织的Wt1免疫荧光染色(n=6)和qRT-PCR检测结果(n=4)证明新生小鼠AR后7天的切口处心外膜细胞被大量激活。Wt1GFPCre小鼠(n=6)AR后7天的GFP信号(Wt1细胞特异表达)能与上皮细胞向间充质细胞转换(EMT)的标志物(Vimentin)共定位,提示AR后激活的心外膜细胞能够发生EMT。Wt1CreERT2;Rosa26RFP谱系追踪小鼠1天龄行AR手术,7天后RFP信号(Wt1来源的细胞特异表达)在切口附近能够分别与αActinin(心肌细胞标志物)、αSMA(血管平滑肌细胞标志物)和FSP1(成纤维细胞标志物)共定位,说明AR后Wt1来源的细胞可以分化为心肌细胞、血管平滑肌细胞和成纤维细胞,进而可能参与心肌再生修复过程。结论:在新生小鼠心肌再生过程中,心外膜细胞被激活,并可以分化为心肌细胞、血管平滑肌细胞以及成纤维细胞。这些发现为心脏医学中的心肌细胞以及血管的再生提示了可靠的细胞来源,也为临床研究心脏再生提供了新思路。
[Abstract]:Objective: To study the differentiation direction of the central outer membrane in the process of myocardial regeneration in neonatal mice. Methods: the apex resection model of wild type mice was constructed and the regeneration was observed by hematoxylin eosin (HE) staining (n=6). Immunofluorescence (n=6) and RT-PCR (n=4) were used to detect the activation of the epicardium in Wt1 (Wilm's Tumor Gene 1). Wt1 specific labelled mice (Wt1GFPCre, n=6 and Wt1CreERT2; Rosa26RFP, n=6) explored the activation and differentiation of the activated epicardium during cardiac regeneration. Results: the wild type 1 days old mice excised the 10%-15% of the apex (apex resection, AR), HE staining (n=6) could completely regenerate the heart 21 days after the neonatal mice. Immunofluorescence staining (n=6) and qRT-PCR test results (n=4) showed that the epicardial cells in the incisional incision at 7 days after AR were activated by a large number of.Wt1GFPCre mice (n=6) AR after 7 days of GFP signal (Wt1 cell specific expression) could be Co located with the marker of epithelial cells converting to mesenchymal cells (EMT), suggesting the epicardial cells activated after AR. EMT.Wt1CreERT2; Rosa26RFP pedigree traced mice at 1 days of age for AR surgery, and 7 days later, RFP signals (specific expression of Wt1 source cells) were able to co localize with alpha Actinin (myocardial cell marker), alpha SMA (vascular smooth muscle cell marker) and FSP1 (fibroblast cell marker) near the incision, indicating that the cells of Wt1 origin after AR can be found. It is differentiated into cardiomyocytes, vascular smooth muscle cells and fibroblasts, and may be involved in the process of regeneration and repair of myocardium. Conclusion: epicardial cells are activated during the regeneration of neonatal mice and can be differentiated into cardiomyocytes, vascular smooth muscle cells and fibroblasts. These findings are cardiac myocytes in cardiac medicine and The regeneration of blood vessels indicates reliable cell source and provides a new idea for clinical research on cardiac regeneration.
【学位授予单位】：北京协和医学院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R54

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