| Background:Spinal cord injury(SCI)is not rare and is difficult to treat,which may result in permanent loss or reduction of bodily function below the injury site.SCI usually causes enormous physical and mental suffering to patients for a long time with high treatment costs,and the morbidity of SCI is constantly increasing,these bring a heavy burden to society and families.The therapeutic effects of current treatment methods are very limited.Methylprednisolone(MP)is currently the most recognized therapeutic drug for SCI,but its clinical application is still controversial due to insufficient clinical evidence supporting the benefits of MP for SCI patients and the side effects of high-dose MP.Artemisinin(ART)is currently the most effective antimalarial drug.In recent years,studies have found that ART has a wide range of pharmacological effects on different diseases,such as neoplasm,viral infections,parasitic infections,autoimmune diseases and nervous system diseases,with low-toxicity.Mechanistic studies have shown that ART can alleviate inflammation and immune reactions in the anterior horn of the spinal cord and brain tissue,improve the survival rate of sensory and motor neurons,restores the continuity of damaged axons,and has a unique protective effect on nerve injuries.Objective:To explore whether ART can promote the repair and functional reconstruction of SCI in rats,and the pharmacology network and molecular mechanisms involved in this process.The results provided new methods and a basis for the clinical treatment of SCI.Method:Ninety-six specific pathogen-free(SPF)female rats were randomly divided into four groups:sham-operation group(Sham),spinal cord injury group(SCI),artemisinin treatment group(ART),and methylprednisolone treatment group(MP),with 24 rats in each group.According to different sampling time points,each group was randomly divided into 4 subgroups(n=6):1 day,3 days,7 days and 14 days.Three rats of each subgroup were used for pathological detection,and the other rats were used for protein detection.Modified Allen method was used to construct the T9 segmental SCI model.Corresponding intervention measures were given to different groups.Before being sacrificed,rats of each subgroup were tested by Basso-Beattie-Bresnahan(BBB)scores at 1,3,7,and 14 days.The spinal cord tissues were assessed for hematoxylin and eosin(HE)staining,Nissl body staining,TdT-mediated dUTP nick-end labeling(TUNEL)staining,and immunohistochemical staining for neurofilament-200(NF-200)and glial fibrillary acidic protein(GFAP).Using monomer drug network pharmacology and online databases,key targets for ART treatment of SCI were screened to analyze the molecular mechanism of action of ART treatment of SCI.Western blotting(WB)was performed to test the expression of proteins in the signaling pathways associated with the screened targets.Results:The BBB scores showed that ART could restore the motor function of rats after SCI significantly,and the effect of ART is similar to MP.HE-staining showed that ART significantly improved the tissue structure of the damaged area,reduced the proportion of the lesion area,and alleviated inflammation,necrosis,and cavitation.Nissl staining showed a remarkable increase was occurred in the number of Nissl-positive neurons in the spinal cord of SCI rats.ART treatment significantly improved the morphology and increased the number of Nissl bodies with stained darkly Nissl bodies and more concentrated and uniform array.TUNEL staining showed that ATR markedly decreased the number of TUNEL-positive cells.NF-200 immunohistochemistry showed that ART repaired bundle-like structures such as spinal axons and nerve fibers in SCI rats.GFAP immunohistochemistry showed that astrocyte activation and glial scar formation were both inhibited by ART.At 14 days after model construction,the pathological changes of ART group and MP group were not obvious.In network pharmacology analysis,a total of 93 key targets were taken by intersecting the ART targets and SCI-Targets.The protein-protein interactions(PPI)network of key genes was constructed,and the topological structure of the network was analyzed.The results showed that there were hydrogen bonds and hydrophobic bonds between several mitogen-activated protein kinase(MAPK)pathway proteins and ART.WB detection showed that ART could promote the regeneration of axons after SCI by activating MAPK signaling pathways.The effect of ART on this pathway at 14 days after model construction was not significantly different from that of MP.Conclusion:ART has a significant therapeutic effect on acute SCI,promoting restoration and functional reconstruction of the injured spinal cord.And various mechanism were involved,which included promoting the survival of motor neurons,inducing tissue repair,inhibiting apoptosis of spinal motor cytons,facilitating the repair of bundle-like structures in the spinal cord such as axons and nerve fibers and inhibiting glial scar formation.This effect may be achieved by activating the MAPK signaling pathway. |
PDF Full Text Request