血管内皮生长因子对内皮生长晕细胞冻存后复苏率及增殖、迁移能力的影响

目的研究血管内皮生长因子对冻存后内皮生长晕细胞复苏率及其增殖、迁移能力的影响。方法密度梯度离心法分离脐带血中单个核细胞,体外扩增培养内皮生长晕细胞,免疫组织化学法、荧光染色法鉴定内皮细胞特性。分别采用不含血管内皮生长因子和含50μg/L血管内皮生长因子的冻存液对内皮生长晕细胞进行冻存,分为正常组、对照组、50μg/L血管内皮生长因子组,24 h后复苏并观察细胞的复苏率及其增殖、迁移能力。结果50μg/L血管内皮生长因子组可提高冻存复苏后内皮生长晕细胞的复苏率。24 h内两组细胞增殖、迁移能力均较正常组减弱(P<0.01),但50μg/L血管内皮生长因子组显示对内皮生长晕细胞增殖、迁移能力的保护作用。48 h后50μg/L血管内皮生长因子组细胞迁移能力较正常组增强(P<0.01),增殖能力接近正常组(P>0.05),但对照组在观察期间内细胞增殖和迁移能力均较正常组与50μg/L血管内皮生长因子组减弱(P<0.01)。结论血管内皮生长因子可以提高冻存后内皮生长晕细胞的复苏率以及复苏后细胞的增殖、迁移能力。

The Effect of VEGF on the Resuscitation Rate and Proliferation and Migration Abilities of Endothelial Outgrowth Cell After Cryopreservation

Aim To investigate the effect of vascular endothelial growth factor(VEGF) on the resuscitation rate,proliferation and migration abilities of endothelial outgrowth cells(EOC) which were cryopreserved. Methods The mononuclear cells were separated from umbilical cord blood by density gradient centrifugation,followed by induction into EOC and expansion in vitro.The endothelial characteristics of EOC were identified by immunohistochemisty and fluorescent staining.Then EOC were cryopreserved at 4℃ for 4 h,-20℃ for 1 h,-70℃ for 24 h using culture medium with or without 50 μg/L VEGF.Cryopreserved EOC were resuscitated after 24 h,and the resuscitation rate,proliferation and migration abilities were measured by Annexin V/PI,CCK-8 and transwell assay respectively. Results EOC possessed many endothelial cell characteristics.The resuscitation rate of the 50 μg/L VEGF group was upregulated compared to the control group(P<0.01),and so were the proliferation and migration abilities(P<0.01).The proliferation and migration abilities of the two cryopreserved groups were both attenuated compared to normal group(P<0.01),while the migration ability of 50 μg/L group was higher than the normal group after 48 h(P<0.01).Moreover,the proliferation ability of 50 μg/L group was the same as the normal group(P>0.05).On the contrary,the proliferation and migration ability of the control group were both weaker than the normal and 50 μg/L group during the observation period(P<0.01). Conclusions VEGF can increase the resuscitation rate and enhance the proliferation and migration abilities of EOC post-cryopreservation.