Construction Of Natural Composite Hydrogel For Angiogenesis

Repair of large segmental bone defect is always a difficult problem to clinical treatment.In recent years,the tissue engineering scaffold has brought new insight for the repair of bone defects since it can mimic the structure and function of native bone.Bone regeneration is a complex process,including the ingrowth and proliferation of osteoblast,transport of nutrients and oxygen,and the elimination of the degradation products.Blood vessels were required to be involved in the process,and early vascularization of bone implant is of great significance.At present,the strategies for promoting the vascularization in bone tissue engineering included: introduction of vascular factors,co-culture of osteoblasts and vascular endothelial cells and bone scaffold,loading of related genes,microsurgical reconstruction in vitro and so on.Among them,vascular factors loading in the scaffold for promoting the vascularization is considered as a simple and effective method.In this work,mesoporous silica nanoparticle(MSNs),as favorable carriers,were doped into an in situ crosslinked hydrogel which was prepared through the Schiff's base reaction of aldehyde hyaluronic acid(A-HA)and N,O-carboxymethyl chitosan(NOCC).The functional drug Sphingosine 1-Phosphate(S1P)was loaded into MSN to investigate the potential of this hybrid hydrogel for promoting angiogenesis.Firstly,the pure hydrogel with different concentrations of NOCC(4,6 and 8 wt%)were prepared.Then the physicochemical properties and cytocompatibility of the hydrogels were studied to select an optimal NOCC concentration for the subsequent research.The MSNs were synthesized and modified with alginate/chitosan multilayers to improve the drug loading capacity and biocompatibility.The hybrid hydrogels with different mass ratio of MSNs-ALG/CHI were fabricated,followed by investigating the effects of the addition of MSNs-ALG/CHI on the physicochemical properties and drug release behavior.Besides,the bone marrow mesenchymal stem cells(BMSCs)adhesion and proliferation on the surface of hybrid hydrogels and the cell viability of BMSCs encapsulated in hybrid hydrogels were studied.Furthermore,the S1P-loaded hybrid hydrogels were used to investigate the ability to enhance endothelial cells migration in vitro and promote angiogenesis in vivo.The result showed that the mechanical properties and cross-linking degree of pure hydrogels were enhanced with the increase of NOCC concentration,while the cell proliferation was decreased.Therefore,the pure hydrogel with 6% NOCC and 6% A-HA was selected for further studies.The addition of nanoparticles significantly enhance the mechanical properties of hybrid hydrogel.In addition,the drug release rate was decreased with the increased content of nanoparticles.Moreover,the cell experiments showed that the hybrid hydrogels exhibited good biocompatibility and could well support BMSCs spreading after adding with(0.5 wt%)bovine serum albumin(BSA).The transwell assay showed that the S1P-loaded hybrid hydrogel could successfully induce directional migration of human umbilical vein endothelial cells(HUVEC).The chicken chorioallantoic membrance(CAM)assay and subcutaneous implanting in mice confirmed that the S1P-loaded hybrid hydrogel could significantly enhance the angiogenesis.More importantly,compared with the control group,the S1P-loaded hybrid hydrogel could obviously induce the new blood vessels after 3 weeks implantation.In conclusion,all the results suggested that the S1P-loaded hybrid hydrogel would be a great potential candidate for promoting angiogenesis in bone tissue engineering.