Construction And Comparison Of Two Functional Microenvironments Space(SDF-1α/Zn²⁺、VEGF/zn²⁺) On The Surface Of Cardiovascular Materials

Stents are still the main treatment for cardiovascular diseases.Complications such as endometrial injury,late thrombosis,restenosis and new atherosclerosis caused by stent implantation still affect the clinical therapeutic effect seriously.Natural vascular endothelium is a natural barrier between blood and vascular wall,which can maintain blood flow and inhibit intima hyperplasia.Therefore,to obtain the complete endodermis,surface modification was used.Currently,many functional factors used to promote the biological behavior of endothelial cells have been used in stents modification widely.The cytokines can bind to receptors and stimulate a series of signaling pathways to promote cell activity,proliferation and migration.Meanwhile,the essential trace elements（metal ions）are the components of various enzymes,and the reaction center of enzyme activity,and also participate in the anti-oxidation and anti-apoptosis pathways of cells.The maintenance of cell biological behavior and function in vivo depends on the interaction of cytokines and metal ions in the extracellular matrix microenvironment.Therefore,in this study,introduced the important cytokines（VEGF,SDF-1α）and metal ions(Zn²⁺)that affect tissue repair in vivo were introduced into the surface of the material to explore the synergistic effect of the two functional factors in the surface modification of vascular scaffolds.The main work of this study is as follows:（1）Effects of Hep-PLL nanoparticles loaded with SDF-1αor VEGF on endothelial repair and intimal regeneration.In this study,SDF-1αor VEGF was loaded in Hep-PLL nanoparticle respectively,the morphology of endothelial cells from different sources were studied.Meanwhile,we evaluated the morphology,activity and differentiation of bone marrow mesenchymal stem cells（BMSCs）.The results showed that Hep-PLL nanoparticles loaded with cytokines could not only promote the surface endothelialization（regulating the morphology of endothelial cells）,but also promote the formation of new intima on the modified surface（promoting the differentiation of BMSC into endothelial cells）.（2）Effects of Hep-PLL nanoparticles loaded with SDF-1αand Zn²⁺（NPS-Zn）on cell biological behaviors were evaluated.Firstly,SDF-1α/Hep and PLL/Zn²⁺mixed suspensions were prepared.Then,the two suspensions were mixed in equal volume to prepare Hep-PLL nanoparticles loaded with SDF-1αand Zn²⁺.Subsequently,using the amine groups exposed on the surface of the nanoparticles to react with dopamine,the nanoparticles were grafted onto the stainless steel（SS）surface which was deposited dopamine.The results showed that the size and potential of the nanoparticles,the hydrophilicity of the modified surface,the exposure amount of functional groups and the release behavior of functional factors were affected.In vitro cytocompatibility evaluation showed that NPS-Zn can regulation the behavior of macrophages to a certain extent,and inhibit the proliferation of smooth muscle cells effectively.However,the influence of the endothelial cells behavior was not obvious.（3）The effect of Hep-PLL nanoparticles loading VEGF and Zn²⁺（NPV-Zn）on cell biological behaviors were evaluated.The preparation and grafting of NPV-Zn nanoparticles are similar to NPS-Zn.The results showed that the loading of VEGF and Zn²⁺had changed the nanoparticles’size,stability,fixed density and the release behavior of functional factors.Cell biology experiments showed that Hep-PLL nanoparticles loaded with VEGF and Zn²⁺could regulate cell inflammatory response effectively,including macrophage transformation into M2 type,inhibition of TNF-αsecretion,and promotion of IL-10 secretion.It also regulates cell biological behaviors selectively,such as promoting endothelial cell growth,preventing endothelial cell from oxidative damage and inhibiting smooth muscle cell proliferation.（4）Effects of NPS-Zn and NPV-Zn modified microenvironments on cell biological behaviors were studied in vitro.In order to explore a more effective and excellent way to construct a microenvironment for promoting intimal repair in surface modification,we conducted a comparative study on the biocompatibility of NPS-Zn and NPV-Zn in vitro and in vivo.It was found that NPV-Zn samples could release Zn²⁺more quickly than NPS-Zn samples,while heparin and cytokines could be released slowly,and functional factors maintained good activity.Cell biological behavior studies showed that NPV-Zn had better effective inflammatory regulation,anti-proliferation of smooth muscle cells and maintenance of endothelial cell function in vitro.（5）Effects of NPS-Zn and NPV-Zn on vascular intima repair in vivo and mechanism exploration.Compared with NPS-Zn,animal experiments showed that NPV-Zn modified samples could regulate the expression of macrophages to M2 type,promote the expression of smooth muscle cells to contractile type,and accelerate the process of endothelialization.The new intima was thin and smooth,realized the intima repair on the surface of stent effectively.Meanwhile,the mechanism of different functional factors was explored.The results showed that the simultaneous introduction of VEGF and Zn²⁺could promote the growth of endothelial cells and intima hyperplasia,inhibit the activation of endothelial cells and reduce the invasion of inflammatory cells simultaneously.（6）Expanded study of the synergistic effect of VEGF and Zn²⁺on the outside realm of vascular repair（skin repair）was performed.In order to demonstrated the synergistic effect of VEGF and Zn²⁺,they were applied to injured skin tissues simultaneously.VEGF and Zn²⁺were wrapped in fibrin gel firstly.And then the cell behavior in vitro and the effect of surface injury repair in vivo were evaluated.The results showed that the fibrin gel samples coated with VEGF and Zn²⁺promoted the activity and proliferation of various cells involved in skin repair.And promoted wound contraction,granulation tissue and new micro-vessels formation,and improve the quality of wound repair.In conclusion,this paper selected cytokines（SDF-1αand VEGF）and metal ions(Zn²⁺)to explore the synergistic effects between different factors.Compared to the SDF-1α,VEGF and Zn²⁺had better synergistic effects on intima repair and injured skin.This paper provides a new thought for repair of damaged microscopic-tissue（vascular atherosclerosis）and macroscopic-tissue（skin mechanical injury）rapidly.