成年犬骨骼肌成肌细胞培养方法改良及生长特性研究  

Modified Culture and Biological Characteritics of Adult Canine Skeletal Myoblasts

作　　者：窦克非[1] 杨跃进[1] 阮英茆[2] 王清峙[2] 陈曦[3] 陈纪林[1] 高润霖[1] 陈在嘉[1] 

机构地区：[1]中国医学科学院,中国协和医科大学心血管病研究所,阜外心血管病医院冠心病诊疗中心,北京市100037 [2]中国医学科学院,中国协和医科大学心血管病研究所,阜外心血管病医院病理科 [3]中国医学科学院,中国协和医科大学心血管病研究所,阜外心血管病医院生化研究室

出　　处：《中国循环杂志》2003年第5期382-384,共3页Chinese Circulation Journal

摘　　要：目的 :为细胞心肌成形治疗初步探讨较为简便、经济的成年犬骨骼肌成肌细胞体外培养方法并研究其生长特性。　　方法 :采用机械分解法与IA型胶原酶和胰蛋白酶的二步酶消化法相结合获取骨骼肌成肌细胞 ;通过细胞形态、生长曲线研究其生长特性 ;细胞组化染色鉴定 ,并比较差速贴壁纯化与否所得细胞组化染色阳性计数差异。　　结果 :改良方法适于获取成年犬骨骼肌成肌细胞 ;细胞生长旺盛 ,在细胞密集或分化培养基作用下可融合成肌管 ;α 横纹肌肌动蛋白、骨骼肌特异性肌球蛋白组化染色细胞为弱阳性 ,而肌管为强阳性。胚胎成肌细胞调节因子组化染色为阳性 ;各组细胞α 横纹肌肌动蛋白组化染色阳性数无显著差异。　　结论 :改良方法获得的骨骼肌成肌细胞具有良好的增殖、分化能力 ;差速贴壁纯化与否获得成肌细胞的阳性计数无显著差异。Objective: To establish a thrifty, convenient culture procedure and observe the biological characteristics of adult canine skeletal myoblasts for cellular cardi omyoplasty(CCM). Methods: Adult canine myoblasts were obtained using the method of two step digestion (inclduing collagenase IA and trypsin) combined wi th the method of mechanical cell dissociation;Morphorlogical characteristics and growth curve wer e observed;The myoblasts were identified by cellular immunochemical staining. We also calculated the positive count of anti α sarcometric actin stained cell s obtained with or without purificative procedure respectively. Results: Adult canine myoblasts obtained by the modified method replicated v ery fast and formed myotubes in differential medium. Though the myoblasts were w eakly positively stained by anti α sarcometric actin and anti myosin immunoc hemical staining, the formed myotubes were strongly positively stained. Anti my ogenin immunochemical staining showed strongly positive. The positive count of a nti sarcometric actin stained cells obtained with or without purificative proc edure had no significant differnece. Conclusion: The modified method can be used as a thrifty and convenient cho ice f or getting adult canine myoblasts. The purificative procedure utilizing differen t attatchment ability of cells can be skipped over.

关 键 词：成年犬 骨骼肌 成肌细胞 细胞培养 细胞生长特性 细胞形态 

分 类 号：R-332[医药卫生] Q2[生物学—细胞生物学]