摘要
目的 观察大鼠骨髓内皮祖细胞 (EPCs)体外诱导成内皮细胞的生长增殖规律。方法 密度梯度离心得到Wistar大鼠骨髓单个核细胞 ;以 10 μg/L血管内皮细胞生长因子 (VEGF)和2 μg/L碱性成纤维细胞生长因子 (bFGF)体外诱导 ,通过细胞形态、免疫荧光法观察内皮细胞表型 ;原代细胞克隆形成率、细胞计数和流式细胞术检测VEGFR 2、VE 钙黏蛋白 (cadherin)阳性表达率观察体外增殖能力与表型。结果 诱导后细胞呈铺路石样形态。免疫荧光法发现诱导细胞表达小鼠抗大鼠Ⅷ因子 (vWF)、VEGFR 2、VE cadherin和CD3 1。原代克隆形成率为 6.66± 0 .75 ;细胞随代次增加增殖能力逐渐下降 ;VEGFR 2和VE cadherin在P2与P5中的阳性率差异无显著性 (P >0 .0 5 )。结论 骨髓EPCs在体外可诱导为成内皮细胞 ,并可保持相对稳定的诱导阳性率。
Objective To investigate the growth dynamics of rat bone marrow derived endothelial progenitor cells (EPCs) in ex vivo expansion.Methods Bone marrow mononuclear cells were isolated by density gradient centrifugation and were induced in M199 medium containing 10% FBS,VEGF (10 μg/L) and bFGF (2 μg/L).Cellular morphology as well as immunofluorescence for vWF,VEGFR 2,VE cadherin and CD31 were used to observe cell phenotype.Proliferation and colony forming rate were calculated.FACS was used to examine expression rate of VEGFR 2 and VE cadherin in induced cells of different passages.Results The induced cells showed a typical cobblestone like appearance and a fibroblast like shape was observed in cells of P4.Immunofluorescence revealed the specific protein expression of endothelial cell in induced cells.The colony forming rate was 6.66±0.75 and 4.18±1.05 respectively in either induced or noninduced per 4×10 7 cells.With cell passages,proliferation rate of both cell types was decreased gradually.Importantly,the marker expression rate maintained relatively stable in cells from passage 2 to passage 5 (35% 40%).Conclusion Bone marrow derived EPCs can be induced into endothelial cells by VEGF and bFGF.Induced cells can maintain their endothelial phenotype during cell passage and thus may generate enough cells for endothelium engineering.
出处
《中华实验外科杂志》
CAS
CSCD
北大核心
2004年第6期652-653,i001,共3页
Chinese Journal of Experimental Surgery
基金
国家高技术研究发展计划 (863计划 )课题(2 0 0 1AA2 1 60 2 1
2 0 0 2AA2 0 50 1 1 )
