成体小鼠心脏Sca-1^+细胞体外分化潜能的实验研究

Experimental study on the differentiation potential of Sca-1-positive cells from adult mouse heart in vitro

目的:研究成体小鼠心脏Sca-1^+细胞体外诱导分化的潜能。方法:利用免疫磁珠分选法分离成体小鼠心脏Sca-1^+细胞,通过BMP-2/FGF-4,TGF-β1及VEGF165等三种不同的体外诱导方法使其向心脏"三系"分化,并从细胞形态的变化、RT-PCR和免疫荧光染色等方面鉴定其分化潜能。结果:经BMP-2/FGF-4诱导,小鼠心脏Sca-1^+细胞发生形态改变,上调心肌细胞特征基因α-MHC、β-MHC、MLC-2a和MLC-2v的mRNA表达,同时表达心肌特征性蛋白cTNT和cMHC。经TGF-β1诱导,小鼠心脏Sca-1^+细胞亦发生形态改变,上调平滑肌特征基因α-SMA和Calponin的mRNA表达,同时表达平滑肌特征性蛋白SMA、sMHC和Calponin的表达。经VEGF165诱导,小鼠心脏Sca-1^+细胞同样发生了形态改变,上调内皮细胞特征基因CD31、vWF和VE-Cadherin的mRNA表达,同时表达内皮细胞特征性抗原CD31。结论:成体小鼠心脏Sca-1^+细胞在体外多种因素的分别作用下,能向心肌样细胞、平滑肌样细胞和内皮样细胞分化,具有心脏干细胞特性的多向分化潜能。

Objective：To study the differentiation potential of Sca-1~＋cells from adult mouse heart in vitro.Methods：We isolate Sca-1~＋cells derived from mouse heart by means of immunomagnetic cell sorting.By using three different kinds of differentiation inducing factors,including BMP-2/FGF-4,TGF-β1and VEGF165,to induce Sca-1~＋cells to differentiated into the heart ＂three lines＂ in vitro.The differentiation potential of the cells was identified from the changes of cell morphology,RT-PCR and immunofluorescence staining.Results：After BMP-2/FGF-4induction,the mouse cardiac Sca-1＋cells underwent morphological changes,up regulation of the mRNA expression ofα-MHC,β-MHC,MLC-2aand MLC-2v,and the protein expression of cTNT and cMHC,which were the specific markers of cardiomyocytes.After TGF-β1induction,these Sca-1~＋cells also had morphological changes,up regulation of mRNA expression ofα-SMA and Calponin,and the protein expression of SMA,sMHC,and Calponin,which were the specific markers of smooth muscle cells.After VEGF165 induction,the Sca-1~＋cells also underwent morphological changes,up regulation of endothelial cell specific gene CD31,vWF,and VE-Cadherin expression,and the expression of endothelial cell specific antigen CD31.Conclusions：Adult mouse cardiac Sca-1~＋cells can differentiate into cardiomyocyte-like cells,smooth muscle-like cells and endothelial-like cells under different inducing factors in vitro,which suggest these cells have multi-lineage differentiation potential as cardiac stem cells.