家兔动脉血管平滑肌细胞培养方法的改进

目的提高家兔血管平滑肌细胞(vascular smooth mus-cle cell,VSMC)体外培养的成功率。方法以传统的主动脉平滑肌细胞培养方法中的组织块贴壁法为基础,从动物选择、培养基配制、血管取材、组织块的大小和接种间距、培养液的量及换液量、细胞传代的时机等多个重要环节进行改进;用倒置显微镜、电镜对培养细胞进行形态学观察,并用平滑肌细胞特异性α-肌动蛋白(α-actin)单克隆抗体免疫组织化学方法进行鉴定。结果台盼蓝检查传代细胞的存活率大于97%;光镜下见培养细胞平行排列呈长梭形及"峰、谷"样结构特征;免疫组织化学染色鉴定培养细胞中VSMC的纯度为97%;电镜技术观察到VSMC完整的超微结构。结论此培养方法能提高动脉平滑肌细胞原代培养的成功率,传代后血管平滑肌细胞生物学特征的稳定性好,可作为研究血管平滑肌细胞生物学行为的有效模型。

Methodological improvement of culturing vascular smooth muscle cells derived from rabbit aorta

Aim To improve the conditions of culturing vascular smooth muscle cells(VSMC)derived from rabbit aorta for increasing the survival rate of the passage cells.Methods Based on the traditional explant-attached method for VSMC culture,several key processes were improved,such as the selection of body weight of animal,preparation of culture medium,size and incubated distance of the tissue bits,the time and volume of exchanging medium,and the well-timed point for the primary VSMC passage.Then the VSMC culture was successfully established in vitro.The cultured cells were observed by inverted microscope and identified by immunohistochemical method(α-actin).Results The survival rate of passage cell observed by Typanblue assay was more than 97 percent;the primary VSMCs grew well with a long spindle-shape and showed a typical "hill and valley" pattern under the light microscope;the positive reaction rate of VSMCs in immunohistochemistry stain and the ultramicrostructures of the cell was observed clearly under the trainsmission electron microscope.Conclusions The improved method could enhance success rate of the primary culture of VSMC and keep its biological characteristics unchanged.It would provide a good cell model for the studies of cardiovascular diseases and pharmacology.