Preliminary Study On The Effect Of Surface Modification And Morphology Of Gold Magnetic Nanoparticles On The Function Of Endothelial Cells

Gold magnetic nanoparticles are composite nanomaterials which come from gold and iron oxide and have great applications in multimodal imaging,drug delivery and tumor treatment.The vascular endothelial cells（VEC）are the inner-most structure that coats the blood vessels around the body,Vascular endothelial cells,flat cells on the inside of blood vessels,distribute all over the body.VEC only serve as a mere barrier between blood and tissues but also play critical role in inflammatory response.Additionally,this cell participate in vascular relaxation and constriction,angiogenesis and repair,growth of smooth muscle cells et.al.The interaction of VEC with gold magnetic nanoparticles lead to various cellular responses depending on the physicochemical properties of this nanoparticle,such as morphology,surface charge and chemical group.In this study,we investigate the effects of different modifications,different charges and different morphologies of magnetic nanoparticles（MNPs）on Vascular endothelial cells.These MNPs including core-shell Au/Fe₃O₄composite nanoparticles（CSNPs）,core-shell Au/Fe₃O₄ composite nanoparticles modified with polystyrene sulfonate（CSNPs-PSS）,core-shell Au/Fe₃O₄ composite nanoparticles modified with polyacrylic acid（CSNPs-PAA）,flower-like Au/Fe₃O₄ composite nanoparticles（FLNPs）and ferroferric oxide nanoparticles（Fe₃O₄-NPs）.FLNPs were first synthesized in this study.Then,the samples,including CSNPs,CSNPs-PSS,CSNPs-PAA,Fe₃O₄-NPs,FLNPs were characterized to confirm the properties of these five MNPs.The protein corona,collected from MNPs,were analyzed by SDS-PAGE.The results showed that the protein isolated from Fe₃O₄-NPs showed higher band intensity and complex band patterns when incubated in DMEM with 10%FBS,followed by FLNPs,CSNPs,CSNPs-PSS,CSNPs-PAA.When incubated in DMEM with protein from cells,protein isolated from CSNPs-PAA showed higher band intensity and complex band patterns,followed by CSNPs,CSNPs-PSS,Fe₃O₄-NPs,FLNPs.The results showed that the protein corona on the surface of MNPs only affected by surface modifications of MNPs but also the proteins types from the environment.Next,we studied the influence of different MNPs on VEC function.First,we examined the viability of cells after treating with MNPs.VEC did not significantly change the viability following 24 h treatment with all of the MNPs under the concentration up to 200μg/m L.It means all of the MNPs have good biocompatibility.Perls’Prussian blue staining was carried out to determine the adhesion and uptake of nanoparticles by VEC.When cultured in DMEM with 10%FBS,a considerable amount of blue staining CSNPs-PAA was present within VEC,followed by Fe₃O₄-NPs and FLNPs,and much fewer blue spots were observed in cells exposure to CSNPs and CSNPs-PSS.The results showed that different surface modifications on the MNPs could effect the absorption of nanoparticles by VEC.It means that a significantly increased MNPs attachment on VEC in DMEM without FBS compared with that of DMEM with 10%FBS.The adhesion of all MNPs was enhanced in DMEM without FBS.It confirmed that proteins in the environment could form protein corona on the surface of MNPs,which affected the absorption of MNPs by VEC.Moreover,proteins from cells form protein corona on the surface of MNPs could promote the absorption of MNPs by VEC.Cell migration effected by MNPs showed that CSNPs-PAA and Fe₃O₄-NPs inhibited the migration of VEC at 200μg/ml.This result was consistent with the adsorption experiment,and indicating that the adsorption of MNPs by VEC inhibits cell migration.After 24 h treatment of the nanoparticles,the m RNA levels of genes involved in inflammatory response and tight junction were assessed.A dramatic increase was found on m RNA levels of interleukin 6（IL-6）,Vascular cell adhesion molecule-1（VCAM-1）,Monocyte chemotactic protein 1（MCP-1）in FLNPs,Fe₃O₄-NPs and CSNPs-PAA treated b End.3 compared with negative control.While,cells exposure to CSNPs and CSNPs-PSS showed no significant difference on IL-6,VCAM-1 and MCP-1.However,cells exposure to MNPs showed a remarkable decrease on the transcript levels of ZO-1 and Claudin-5.The results showed that the morphology and modification of MNPs could effect the expression of inflammatory factors and intercellular tight junction proteins in VEC.Previous studies have shown that MCP-1 can inhibit the expression of tight junction,and affect the permeability of VEC layer.In this study,further verified that FLNPs could promote the expression of MCP-1 by ELISA,followed by Fe₃O₄-NPs and CSNPs-PAA.So it is hypothesis that FLNPs,Fe₃O₄-NPs and CSNPS-PAA could inhibit the expression of the intercellular tight junction and increase the endothelial leakiness by promoting the expression of MCP-1.