刘姿麟,林慕之,况春燕,刘兴德
LIU Zilin,LIN Muzhi,KUANG Chunyan,LIU Xingde

• 1:贵州医科大学附属人民医院
• 2:贵州医科大学
• 3:贵州省人民医院心内科

摘要(Abstract)：

目的:建立一种可行的原代血管平滑肌细胞(VSMC)培养方法。方法:严格无菌操作环境下取大鼠胸主动脉及腹主动脉中层组织,通过组织块贴壁法进行原代培养,胰酶消化法进行传代,应用倒置相差显微镜对不同培养时间的细胞进行形态学观察,使用免疫荧光法对培养细胞进行鉴定。结果:原代培养第4~6天时,组织块周围开始有长梭形细胞爬出;第7~9天时,细胞爬出较少的呈网状生长,组织块周围细胞爬出较多的呈漩涡状生长,致密排列呈束状;第10~14天时,细胞呈典型的"峰—谷"样结构,相互融合达80%,此时传代细胞生长速度增快,细胞变大,折光性减弱,免疫荧光染色鉴定可见VSMC特异性的α-平滑肌肌动蛋白(α-SM-actin)表达,表达的阳性细胞在95%以上。结论:成功建立一种可行的原代VSMC培养方法。
Objective:To establish a convenient and efficient method for culturing primary generation of aortic vascular smooth muscle cells(VSMC) of rat.Methods:The aorta was isolated from SD rats under strict sterile conditions.The primary culture and subculture were obtained by tissue-piece inoculation and trypsin respectively.The cellular morphology was observed with inverted phase contrast microscope and identified by immunofluorescence.Results:The VSMC grew out on days 4 ~ 6 and showed fusiform; On days 7 ~ 9,some cells grew to form mesh,and more cells grew in whorled pattern and form compact sarciniform; On days 10 ~ 14,cells appeared typical " peak-valley" structure,and became nearly 80% integrated.At this time,passage cells grew faster,and cells became larger,and their refractivity weakened.The typical α-Smooth muscle actin(α-SM-actin) in SMCS could be found by immunofluorescent staining,and the positive rate of α-SM-actin in SMCS was more than 95%.Conclusion:A reliable and feasible culture method for primary generation of VSMC has been established successfully.

关键词(KeyWords)： 血管平滑肌细胞;原代培养;组织块贴壁法;免疫荧光;大鼠,Sprague-Dawley
vascular smooth muscle cell;primary culture;adherent culture;immunofluorescence;rats,Sprague-Dawley

Abstract:

Keywords:

基金项目(Foundation): 国家自然科学基金(81360034,81560056)

作者(Author): 刘姿麟,林慕之,况春燕,刘兴德
LIU Zilin,LIN Muzhi,KUANG Chunyan,LIU Xingde

DOI: 10.19367/j.cnki.1000-2707.2017.02.001

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