Chitosan Coated Hollow Manganese Dioxide Nanoparticles Loaded With Resveratrol For The Treatment Of Spinal Cord Injury

ObjectiveThe aim of this study was to design a chitosan-coated hollow manganese dioxide nanoparticle（CM）with inherent anti-oxidative stress capacity as a delivery carrier for resveratrol（Res）,and studied the ability of this delivery system to cross the blood-spinal barrier（BSCB）to reach the injured site and the therapeutic effect in spinal cord injury（SCI）.MethodsIn this study,hollow manganese dioxide nanoparticles were prepared,chitosan was coated on the surface of hollow manganese dioxide nanoparticles by electrostatic adsorption,and then Res was loaded into CM by adsorption.Hollow manganese dioxide nanoparticles（HM）and CM were characterized by transmission electron microscopy（TEM）and Malvern laser granulometry（DLS）.The drug loading and release rate of Res were measured by ultraviolet spectrophotometry.The drug-loaded system was characterized by X-ray energy spectrometer（EDS）,Raman spectroscopy（Raman）,differential scanning calorimetry（DSC）and X-ray diffraction（PXRD）.In vitro cellular experiments,MTT assay was used to examine the cytotoxicity of CM.Cellular uptake assay was used to investigate the uptake ability of CM by PC12 cells and BV2 cells.OGD was used to establish oxidative stress injury models of PC12 cells and inflammation models of BV2 cells to investigate the neuroprotective and anti-inflammatory effects of CMR.In the in vivo animal experiments,the ability of CM to cross the BSCB to reach the injury site was investigated by small animal in vivo imaging technology,and the anti-inflammatory and anti-apoptotic effects of CMR were investigated by Western blotting.Immunofluorescence assay confirmed the anti-oxidative,anti-inflammatory and anti-apoptotic effects of CMR in vitro and in vivo.The recovery of motor function of mice in each group was evaluated by BMS score and footprint analysis.ResultsTEM and DLS results indicated that HM and CM were monodisperse regular hollow spheres.The particle size of HM was 112.5±12.35 nm,and that of CM was 132.3±13.28 nm.The zeta potential of HM was-35.2±2.35 m V,and that of CM was 22.8±1.23 mV.The drug loading of Res measured by UV spectrophotometry was 21.39±2.53%.In vitro drug release results showed that the Res release of CMR in 24 h was 85.6±5.05%.EDS results showed that HM contained manganese and oxygen elements.The Raman results showed a characteristic peak of MnO₂at 635 cm^-1,indicating the successful preparation of HM.DSC and XRD results indicated that Res exists in CMR in an amorphous state.MTT assay demonstrated good biosafety of CMR.In vitro cellular uptake experiments demonstrated that CMR could be efficiently taken up by PC12 cells and BV2 cells.In vivo animal experiments,in vivo imaging results demonstrated the ability of CM to cross BSCB to reach the injury site,and Western blot results showed that CMR had anti-inflammatory and anti-apoptotic effects.Immunofluorescence assay results demonstrated the anti-oxidative,anti-inflammatory and anti-apoptotic effects of CMR in vitro and in vivo.BMS score and footprint analysis showed that CMR could promote the recovery of motor function in SCI mice.ConclusionsIn conclusion,the present study successfully synthesized chitosan coated hollow manganese dioxide nanoparticles for the treatment of SCI.As a carrier of Res,CM has the advantages of high drug loading and good sustained release effect.In vitro and in vivo animal experiments have shown that CM could effectively transport Res to the injury site,inhibit oxidative stress,reduce inflammatory response and protect neurons,thereby promoting the recovery of motor function in SCI mice.Therefore,CMR could provide a new therapeutic strategy for the clinical treatment of SCI.