AIM: To prepare an improved medium for culturing human cardiac stem cells. METHODS: The heart samples of the right auricle obtained from the patients after cardiac surgery were minced into pieces (about 1 mm X 1 mm × 1 mm) , digested and cultured. The primary cells obtained were cultured with improved cardiosphere-growing medium (CGM) for proliferation, and the cells were identified by flow cytometry. Finally, purer c-kit + cells were obtained by the method of magnetic bead sorting. RESULTS: After about 2 weeks of culture, small, round and phase-bright cells migrated from the well-adherent explants over a layer of fibroblast-like cells. These cells were collected by a brief digestion with Ac-cutase, washed and cultured with improved CGM. No significant difference of the proliferative capacity between using traditional CGM and improved CGM was observed. After subculture and proliferation, the identification result by flow cytometry showed that the positive rate of c-Kit surface marker on these cells was (6.8 ±2. 1)%. By the method of anti-c-Kit magnetic bead sorting, purer c-Kit+ cardiac stem cells were obtained and differentiated into cardiomyocytes. CONCLUSION: Purer c-Kit+ cardiac stem cells are isolated with the improved CGM culture.%目的:用改良的培养基培养人心肌干细胞,并筛选鉴定.方法:通过心脏外科手术取下右心耳组织,用组织块培养法来获得原代心肌干细胞,用改良的心肌干细胞培养液培养,增殖传代后进行心肌干细胞表面标志物的鉴定,再用免疫磁珠筛选以获得较纯的目的心肌干细胞.结果:培养约2周后,贴壁良好的心肌组织块周围可见有小、圆、亮的细胞爬出,用Accutase(细胞消化液)短时间消化后冲洗细胞,培养增殖传代,其增殖能力与传统心肌干细胞培养液培养的细胞比较无显著差异,用流式细胞术鉴定c-Kit(干细胞表面标志物),其阳性率(6.8±2.1)%,之后用含抗c-Kit抗体(anti-c-Kit)的磁珠进行分选,即得较纯的c-Kit阳性(c-Kit+)心肌干细胞,它们可以分化为心肌细胞.结论:通过心脏组织块贴壁培养法,用改良后的心肌干细胞培养基培养,再借助免疫磁珠的分选,同样可得到较纯的c-Kit+心肌干细胞.
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