The Primary Study Of Influencing Factors On Tensile Strength And Angiogenic Activity Of Composite Chitosan Tissue Engineered Dermal Quivalents

Objective: 1. To measure the tensile strength of tissue engineered dermal equivalents composed of different quantities of fibroblasts and different scaffold by high-sensitive and micro-tension mechanical property device. 2. To investigate the isolation and cryopreservation of human umbilical vein endothelial cells (HUVECs) in vitro in order to discuss the possibility of providing source cells to reconstruct tissue engineered vascular equivalents. 3. To detect the quantities of vascular endothelial growth factor (VEGF) in the medium secreted by tissue engineered skin equivalents. 4. To measure the activity of conditioned medium of composite chitosan dermal equivalents on the proliferation of human umbilical vein endothelial cells.Methods: 1. Collagen dermal equivalents and composite chitosan dermal equivalents with different quantities of fibroblasts were cultured for fifteen days and then were measured their tensile strength. 2. Endothelial cells were acquired by filling collagen enzyme solution into the lumen of umbilical veins and then cultured in M199 with 20% fetal bovine serum. Cells were identified by their morphology andâ…§factor immunostaining. The endothelial cells were suspended in cryopreserving solution containing with 10% glycerol and cryopreserved in liquid nitrogen for one or four weeks. The proliferation of post-thawed cells and non-frozen cells were evaluated by cell-counting. 3. The VEGF in the cultured medium was detected by ELISA (antibody-VEGF-antibody) at different time point secreted by monolayer fibroblasts (FBs), collagen gel dermal equivalents, collagen gel skin equivalents, composite chitosan dermal equivalents or composite chitosan skin equivalents. 4. Endothelial serum-free medium is supplemented with 10% fibroblast culture medium or medium conditioned by monolayer or three-dimensional fibroblast cultures. The DNA synthesis of HUVEC was determined by...