兔脂肪来源间充质干细胞的生物表型及其心肌样细胞的分化潜能

Biological phenotypes of rabbit adipose-derived mesenchymal stem cells and their differentiation potentiality for myocardiocyte-like cells

目的鉴定兔脂肪来源间充质干细胞(ADMSCs)的生物学表型及其向心肌细胞分化的潜能。方法成年兔肩胛区皮下脂肪组织经消化、培养,观察其形态变化。第3代细胞进行CD29、CD34、CD44、CD105免疫细胞化学显色、免疫荧光单标、双标及三标显色。5-氮杂胞苷(5-aza)诱导ADMSCs 7d,免疫组织化学和免疫荧光技术显示肌钙蛋白-I(Troponin-I)和肌球蛋白重链(MHC)表达。光学显微镜和激光扫描共焦显微镜观察并计数。结果ADMSCs开始为圆形,后逐渐伸展,呈现形态多样性。90%以上的第3代细胞阳性表达CD29、CD44、CD105,三者共表达于胞质中。CD34阴性表达。CD分子表达阳性的细胞70.59%±1.26%表达Troponin-I,71.98%±1.13%表达MHC。结论 90%以上的ADMSCs共表达干细胞表面标记CD29、CD44、CD105,它们具有很强的分化为心肌样细胞的潜能。

Objective To identify biological phenotypes of adipose-derived mesenchymal stem cells （ADMSCs）of rabbits and their differentiation potentiality for myocardiocyte-like ceils. Methods ADMSCs were isolated, digested and cultured from scapular subcutaneous adipose tissue of adult rabbits and their morphology was observed. The third passage of ADMSCs were stained by surface markers CD29, CD34, CD44 as well as CD105 using immunocytochemistry and single-, double-,and tri-labelled immunofluorescence. ADMSCs were induced by 5-aza for seven days, and then the Troponin-I and myosin heavy chain （ MHC ） expressions were examined with immunocytochemistry and immunofluorescence simultaneously. The expression cells were observed and counted under a light microscope and a laser confocal scanning microscopy. Results ADMSCs were round at beginning, and gradually extended, and showed multiplicity at the end. Over 90% cells were positive for CD29, CD44 and CD105 markers. Furthermore, those three markers were co-expressed in cytoplasm of ADMSCs. CD34 was negative in all cells. The Troponin-I （70.59% ± 1.26% ） and MHC （71.98% ±1. 13% ）were expressed in the positive ADMSCs of three CD molecules after induced by 5-aza. Conclusion Over 90% ADMSCs co-express CD29, CD44 and CD105 molecules which are three kinds of stem cell markers. All co-expression cells are of strongly differentiated potentiality for myocardiocyte-like cells.