摘要
目的 探讨组织工程心脏瓣叶种子细胞———内皮细胞的体外分离、培养及鉴定。方法 杂种猪 4头 ,取整段主动脉。使用 0 .2 5 %Ⅱ型胶原酶消化分离血管内皮细胞。使用M199培养、扩增内皮细胞。使用倒置显微镜、免疫组化方法及透射电子显微镜对内皮细胞进行鉴定。结果 倒置显微镜结果显示 :原代细胞及经传代培养后的细胞 ,呈现典型的内皮细胞特征 ,呈铺路石样改变。免疫组化检测示内皮细胞Ⅷ因子相关抗原呈阳性反应。透射电镜显示 :细胞间存在上皮细胞特有紧密连接 ,未发现内皮细胞Webel Palade小体结构。内皮细胞经 3～ 4次传代 ,细胞数量可增殖达到 6× 10 6。结论 使用胶原酶消化法可有效分离内皮细胞 ,种子细胞经体外培养扩增达到一定数量 。
Objectives The experiment was designed to evaluate methods of isolation,culture and identification of the endothelial cell(EC) for tissue engineering of heart leaflets.Methods aortic arteries were harvested from 4 hybrid young pigs.The endothelial cell was isolated by use of 0.25% collagenase Ⅱ and cultured,expanded with medium 199.The endothelial cell was identified under inverted microscope,with immunohistochemical assay and transmission electron microscope(TEM).Results Inverted microscope revealed that the cell isolated had a typical 'cobblestone' morphology of endothelial cell.Assay for EC vwF Ⅷ was positive.TEM shows that there is tight and gap junction between cells,which is typical structure of epithelial cells.No Weibel Palade body was found.After three to four passages,6×10 6 endothelial cell were obtained.Conclusions By use of 0.25%collagenaseⅡ,endotheilial cells can be effectively isolated from aortic artery.After expanded,the endothelial cell is ready for seeding.
出处
《江西医学院学报》
2003年第3期15-18,共4页
Acta Academiae Medicinae Jiangxi
基金
广东省自然科学基金重点项目 (C 0 10 177)
