组织贴块法培养大鼠主动脉平滑肌细胞及鉴定

Identification and Culture of Rat AO Vascular Smooth Muscle Cells by Tissue - piece Inoculation

目的探讨以原代培养的方法获得活力稳定、高度纯化的大鼠血管平滑肌细胞的体外模型,并为相关研究提供所需的试验材料。方法采用组织贴块法进行细胞原代培养,胰酶消化传代,液氮冻存,并应用倒置相差显微镜和SP法免疫组化染色对细胞进行鉴定。结果 85%的组织块接种存活,传代后90%以上的平滑肌细胞重新贴壁生长,冻存后细胞再行1～2次传代后可恢复增生活力。显微镜下细胞呈典型的"谷峰状"生长,SP法免疫组化染色显示胞质内α平滑肌肌动蛋白表达阳性。结论组织块贴壁法简便易行,短期内可获大量符合后续试验要求的血管平滑肌细胞。

Objective To obtain a lasting and purified cell line of vascular smooth muscle cells （VSMCs） by primary culture,and to provide these test materials related research. Methods The primary and transfer culture was done by tissue - piece inoculationand trypsin digestion,respectively. The cells were frozen in liquid nitrogen,and cultured cells were identified by phase contrast microscopy and immu- nohistochemical SP kit. Results 85% of inoculated tissue pieces survived. More than 90% of VSMCs rcgrew in transfered culture. The frozen cells could recover their proliferation by several culture transfers. The VSMCs showed the typical ＂peak and valley＂ characteristics under microscopy. Immunohistochemical staining with antibody against SM - ct - actin expression in these cells was positive. Conclusion The tissue - piece inoculation is a simple method for obtaining satisfactory VSMCs in short term.