A Mechanism Study On Promoting Bladder Regeneration Angiogenesis By Adipose Stem Cells And Their Extracellular Vesicles With Scaffolds

Research purpose: This thesis aims to investigate the feasibility of acellular scaffold,adipose stem cells(ASCs)and their extracellular vesicles(EVs)combined with scaffolds in promoting bladder regeneration angiogenesis,and explore underlying mechanisms.Research contents: This thesis prepared and evaluated bi-layer chitosan scaffold,rat ASCs-seeded bladder acellular matrix graft(BAMG)-silk fibroin(SF)composite scaffold,and human ASCs-EVs-encapsulated BAMG-hydrogel-SF mesh composite scaffold.Bladder morphological,histological and functional recoveries were evaluated after augmenting rat bladder with these scaffolds,of which the angiogenetic mechanisms were explored in association with SDF-1α/ CXCR4 pathway.Research methods: Initially,mechanical properties of bi-layer chitosan scaffold were evaluated.Bladder morphological,histological and functional restorations were evaluated at 21 and 70 days after bladder augmentation with bi-layer chitosan scaffold.The association between its angiogenetic potential and SDF-1α/ CXCR4 pathway was explored as a pilot study.Afterwards,the mechanical properties and cytocompatibility of rat ASCs-seeded BAMG-SF composite scaffold were measured.Bladder morphological,histological and functional restorations were evaluated at 2,4 and 12 weeks after bladder augmentation with this scaffold.The association between its angiogenetic potential,SDF-1α/ CXCR4 pathway and downstream signalling molecules were futher explored.Finally,the angiogenetic potential and related mechanisms of human ASCs-EVs were validated in vitro.The feasibility of human ASCs-EVs-encapsulated BAMG-hydrogel-SF mesh composite scaffold in augmentation cystoplasty was evaluated as described above.Meanwhile,the role of SDF-1α/ CXCR4 pathway in promoting angiogenesis of regenerated bladder by this scaffold was verified in vivo.Research results: Bi-layer chitosan scaffold increased blood vessels density,promoted bladder smooth muscle regeneration and functional recovery at 70 days post-implantation.Its angiogenetic potential was related to SDF-1α/ CXCR4 pathway.Rat ASCs-seeded BAMG-SF composite scaffold increased blood vessels density,inhibit local fibrosis and imflammation,promoted functional recovery of regenerated bladder.It promoted VEGF-mediated angiogenesis by activating SDF-1α/ CXCR4 pathway through ASCs paracrine.Human ASCs-EVs-encapsulated BAMG-hydrogel-SF mesh composite scaffold improved blood vessels density and diameter,inhibit local fibrosis and inflammation of regenerated bladder,thus recovered bladder function to physiological level.Combined with results in vitro,this scaffold promoted VEGF-mediated angiogenesis by activating SDF-1α/ CXCR4 pathway and upregulating ERK 1/2 phosphorylation.Research conclusion: Bi-layer chitosan,BAMG-SF scaffold combined with rat ASCs and BAMG-hydrogel-SF scaffold combined with human ASCs-EVs promoted angiogenesis of bladder regeneration by activating SDF-1α/ CXCR4 pathway.