Biological Activity Study In Cryopreserving Vascular Endothelial Cell Of Composite Tissue With Two Different Freezing Methods

ObjectiveTo investigate the activity of vascular endothelial cell of composite tissuefollowing rapid-freezing and slow-freezing,to explore the less influence of twocryopreservation methods plays in vascular endothelial cell of composite tissue.Methods40New Zealand white rabbits were randomly divided into control group、cryopreservation for12hours group、3days group、7days group of10each.Control group without any treatment. Experimental groups were divided NewZealand white rabbits hind legs (20limbs in each group) into two subgroups（Rapid freezing and slow freezing）,10limbs per group. Then putting the Rapidfreezing group into liquid nitrogen for12h、3days、7days; In group slowfreezing，freezing was carried out to一80℃using programmed frozen，thenfrozen to一196℃.ALL groups need cryopreservation for12hours,3days,7days, and then rapid rewarming. After thawing，the artery was dissected and its endothelium were examined by optical microscope and immunohistochemistry.The results were analyzed statistically.Results①After examined by optical microscope,there were less endothelial cellloss in control group. The vascular endothelial cells of Experimental groupshave different levels of loss and damaged。 By statistical analysis Experimentalgroups were significant differences with the control group (P <0.05); but therewas no significant difference between Experimental groups (P>0.05);②After examined immunohistochemistry, there was patchy and mild expression ofVEGF and CD34in the vascular endothelial cells of all experimental groupsbefore and after culture. Compared with rapid-freezing groups，the expressionof VEGF and CD34in slow-freezing was closer to that in fresh control group.Slow-frozen groups was significantly better than rapid frozen groups (P <0.05).Conclusions1. The activity of vascular endothelial of the composite tissue show differentinjuries after programmed freezing-thawing process.2. Slow-freezing is superior to rapid-freezing to preserve vascularendothelial cells of composite tissue,and plays less influence in VEGF andCD34expression and vascular injury of composite tissue.