血管内皮和平滑肌细胞联合培养构建组织工程化血管的方法

目的：探讨组织工程血管种子细胞原代培养、传代扩增、鉴定的方法，为组织工程血管的构建提供理想的种子细胞。方法：无菌条件下取正常产妇分娩的脐带动静脉，ROSS法（即组织贴块法）原代培养混合血管细胞。倒置相差显微镜、免疫组化法对细胞进行鉴定。结果：倒置显微镜显示原代细胞及经传代培养后的细胞，呈现内皮样细胞、平滑肌样细胞混合生长的特征。免疫组化法检测示内皮细胞Ⅷ因子、平滑肌细胞α-肌动蛋白呈阳性反应。细胞经3次～5次传代，数量可增殖达到8×10^6—10×10^6。结论：使用组织贴块法原代培养可获得内皮细胞、平滑肌细胞混合体，细胞经体外联合培养、传代可以达到组织工程血管种子细胞所需的数量。

Co-culture of Vascular Endothelial Cells and Smooth Muscle Cells and Tissue Engineered Vascularization

Objective：To explore the methods of primary culture,passage culture and identification of progenitor cells of tissue-engineered blood vessels.Methods：Umbilical cord artery and vein were collected from healthy lying-in women. Under axenic condition,vascular endothelial cells and smooth muscle cells were co-cultured by enzyme digestion and tissue plant method. Morphological characteristics of the cells were observed with inverted microscope.Expression of Ⅷ related antigen in endothelial cells and α-smooth muscle actin （α-SMA）in smooth muscle cells were detected by immunohistochemistry. Results：Under inverted microscope,the morphological characteristics of two kinds of cells accord with those of vascular endothelial cells and smooth muscle cells.Vascular endothelial cells and smooth muscle cells were stained with antibody of rabbit anti-human Ⅷ related antigen and rat anti-human α-SMA.Conclusion：The cultured cells may be as seed cells of vascular tissue engineering.