组织块贴壁法和混合酶消化法分离原代血管平滑肌细胞的表型特征比较

Comparison of Phenotypic Characteristics of Primary Vascular Smooth Muscle Cells Isolated by Tissue Block Adhesion Method and Mixed Enzyme Digestion Method

目的:探究组织块贴壁法、混合酶消化法分离培养的原代大鼠血管平滑肌细胞(VSMCs)在形态、生长、表型特征蛋白表达方面的不同,为合理使用VSMCs提供参考。方法:分别使用组织块贴壁法和混合酶消化法分离获得原代VSMCs,进行体外传代培养,观察细胞形态及细胞表型特征蛋白免疫荧光亮度。结果:混合酶消化法获得的原代VSMCs特征蛋白SMα-actin表达明显,表明在收缩表型特征方面优于组织块贴壁法,随着传代次数的增加,混合酶消化法在第7代仍部分保留收缩表型特性,而组织块贴壁法在第7代已丧失收缩表型特性,转变为合成型。结论:混合酶消化法在获得较稳定的收缩表型VSMCs方面比组织块贴壁法更具有优势,各实验室应根据自身的实验条件和实验设计准确选择。

Objective : To investigate the differences in morphology, growth and phenotypic characteristic protein expression of primary rat vascular smooth muscle cells (VSMCs) isolated and cultured by tissue block adhesion method and mixed enzyme digestion method, and provide reference for rational use of VSMCs. Methods : Primary VSMCs were isolated by tissue block adhesion method and mixed enzyme digestion method. The cell morphology and immunophenotypic expression of cell phenotypic protein were observed. Results : The expression of the characteristic protein SM α-actin of the primary VSMCs obtained by the mixed enzyme digestion method was obvious, indicating that the contractile phenotypic characteristics were significantly better than the tissue block adhesion method. With the increase of the number of passages, the mixed enzyme digestion method was still in the 7th generation. Partially retained contractile phenotypic properties, and the tissue block adhesion method lost its contractile phenotypic properties in the 7th generation and turned into a synthetic type. Conclusion : The enzymatic digestion method has more advantages than the tissue block adhesion method in obtaining stable phenotype VSMCs. Each laboratory should be accurately selected according to experimental conditions and experimental design.