Identification and Characterization of a Novel Multipotent Sub-Population of Sca-1+ Cardiac Progenitor Cells for Myocardial Regeneration

Methods and ResultsThe cardiac stem/progenitor cells from adult mice were seeded at low density in serum-free medium. The colonies thus obtained were expanded separately and assessed for expression of stem cell antigen-1 (Sca-1). Two colonies each with high Sca-1 (CSH1; 95.9%; CSH2; 90.6%) and low Sca-1 (CSL1; 37.1%; CSL2; 17.4%) expressing cells were selected for further studies. Sca-1+ cells (98.4%) isolated using Magnetic Cell Sorting System (MACS) from the hearts were used as a control. Although the selected populations were similar in surface marker expression (low in c-kit, CD45, CD34, CD31 and high in CD29), these cells exhibited diverse differentiation potential. Unlike CSH1, CSH2 expressed Nanog, TERT, Bcrp1, Nestin, Musashi1 and Isl-1, and also showed differentiation into osteogenic, chondrogenic, smooth muscle, endothelial and cardiac lineages. MACS sorted cells exhibited similar tendency albeit with relatively weaker differentiation potential. Transplantation of CSH2 cells into infarcted heart showed attenuated infarction size, significantly preserved left ventricular function and anterior wall thickness, and increased capillary density. We also observed direct differentiation of transplanted cells into endothelium and cardiomyocytes. ConclusionsThe cardiac stem/progenitor cells isolated by a combined clonal selection and surface marker approach possessed multiple stem cell features important for cardiac regeneration.

方法与结果
从成年小鼠体内分离得到的心脏干/祖细胞，以低密度接种于无血清培养基中。将所获细胞集落分别进行扩增，并检测其干细胞抗原-1（Sca-1）的表达水平。分别选取两株高表达Sca-1（CSH1：95.9%；CSH2：90.6%）与两株低表达Sca-1（CSL1：37.1%；CSL2：17.4%）的细胞集落用于后续实验。采用磁激活细胞分选系统（Magnetic Cell Sorting System, MACS）从心脏组织中分离得到的Sca-1阳性细胞（占比98.4%）作为对照。
尽管上述筛选得到的细胞群表面标志物表达模式相似（均低表达c-kit、CD45、CD34、CD31，高表达CD29），但其分化潜能却存在显著差异。与CSH1不同，CSH2可表达Nanog、TERT、Bcrp1、Nestin、Musashi1及Isl-1，且能够向成骨细胞、软骨细胞、平滑肌细胞、内皮细胞及心肌细胞谱系分化。经MACS分选得到的细胞虽呈现相似的分化趋势，但其分化潜能相对较弱。
将CSH2细胞移植至心肌梗死模型小鼠的心脏后，可缩小梗死面积，显著保留左心室功能与前壁厚度，并提高毛细血管密度。同时还观察到移植细胞可直接分化为内皮细胞与心肌细胞。
结论
采用克隆筛选联合表面标志物分选策略分离得到的心脏干/祖细胞，具备多种与心脏再生相关的干细胞特性。