Analysis On Process Of Neovascularization Of Porous Silk Fibroin Films

Silk fibroin has been widely applied in dermis, bone, vessel regeneration due to its biocompatibility, degradability and remarkable mechanical properties. Angiogenesis is a key step during wound healing in most tissues, so biomaterials aiding wound healing requires rapid and effective neovascularization of biomaterials after implantation. Process of neovascularization involved in interactions between cell-cell, cell-growth factors and silk fibroin–extracellular matrix (ECM). Although numbers of researches have focused on the design and preparation of porous biomaterials for excellent neovascularization, the analysis about processes and model on angiogenesis of biomaterial in vivo has been little reported so far.It is necessary to explore and analyze the characters and process of porous silk fibroin films( PSFFs )neovascularization. So this paper aims to study the dynamic process involved in the vascularization of PSFFs for dermis regeneration using immunochemical method, transmission electron microscopy (TEM), Reverse Transcription Polymerase Chain Reaction (RT-PCR) method. Angiogenic factors VEGF and its receptor, hypoxia inducible factor-1(HIF),integrinαvβ3, angiopoietin-1 and so on were observed for exploring a real and systemic process of PSFFs neovascularization.This paper includes three parts:The first part is to observe process and mode of angiogenesis in PSFFs by hematoxylin-eosin staining (HE) in histology and TEM in ultrastructure. The result showed capillaries formation in PSFFs and polyvinyl alcohol (PVA) sponges went through form avascular to vascular, and matured through remodeling into a complex vascular net. At early stage of wound healing capillaries were formed in PSFFs by sprouting, and intussusceptive angiogenesis were mainly present at late stage. The second part is to study vascular endothelial growth factor (VEGF), hypoxia inducible factor-1(HIF), integrinαvβ3, CD34, fibronectin (FN) and laminin(LN) expression in PSFFs and PVA through immunochemical method. The results show that mean value of MVD in PSFFs is 0, 3.54, 11.6, 24.5 respectively after 1, 7, 15, 25 days implantation. It is significant difference compared with that of PVA.VEGF expression in PSFFs is 3.4,1.9, 2.5, 2.6 respectively by day 1,7,15,25. Mean value of HIF expression in PSFFs was 12,4.8,8.6,9.0 by day1,7,15,25. Mean value of HIF and VEGF expression in PVA compared with those of PSFFs are significant difference only by day 7. Trends of HIF expression is consistent with that of VEGF in PSFFs, otherwise integrinαvβ3, FN and LN expression were discussed too. PSFFs have better neovascularization than that of PVA due to excellent characters in angiogenesis, degradation.The third part is to study the mRNA expression about VEGF and its receptor, angiopoietin-1(Ang-1), integrinαvβ3, HIF at different times. More proliferating cells in PSFFs were in the absence of oxygen after implanting to dermis defects. The potent angiogenic factors VEGF and its receptor,integrinαvβ3 and Ang-1 may be activated in response to hypoxia under the control of the transcription factor HIF-1.The results of VEGF, HIF -1 and integrinαvβ3 were consistent with that of immunochemical level.In all, Processes and model of capillaries formed in PSFFs were studies in immunochemical and mRNA level, PSFFs have better neovascularization than that of PVA due to excellent characters in degradation, angiogenesis. PSFFs as scaffolds can promote angiogenesis by sprouting at early stage and intussusception at late stage in wound healing; HIF, VEGF and integrinαvβ3, FN and LN expression in PSFFs and PVA were in a dynamic process in mRNA and immuochemical level in order to adapt environment.General regulations in angiogenesis were followed by neovascularization in PSFFs. Primary model for processes of neovascularization were put forward.