β-elemene Promotes Functional Recovery Of Locomotion In Rats With Spinal Cord Injury

ObjectiveTo explore whether β-elemene promote the recovery of motor function in rats after spinal cord injury via reducing endoplasmic reticulum stress and promoting the neurite growth of injured neurons through in vivo and in vitro experiments,providing a new clinical treatment strategy.MethodsThe effects of β-elemene on the viability and toxicity of anterior horn motor neuron cell line VSC4.1 cell under normal culture conditions were first examined by CCK-8 assay and LDH release assay.Subsequently,an injury model was induced by cobalt chloride constructed to simulate cell hypoxia injury,and CCK-8 assay and LDH release assay were used to detect the effect of P-elemene on the viability and toxicity of VSC4.1 cell after cobalt chloride injury.The effect of apoptosis on VSC4.1 cell after cobalt chloride injury was measured by FACS assays using Annexin/PI staining.Western blotting was used to detect the expression of endoplasmic reticulum stress-related protein(grp78,chop)and apoptosis-related proteins(Cleaved-caspase3)in the control group,β-elemene group,cobalt chloride injury group,β-elemene+cobalt chloride group.Subsequently,we used immunofluorescence staining to observe the growth of neurites in primary cortical neurons treated by P-elemene and Y27632.The pull-down assay was used to detect the activation of RhoA kinase.Western blotting was used to detect the phosphorylation of limk and confilin proteins.In vivo experiments,we used Allen’s method to construct the spinal cord injury model.TUNEL staining was used to detect the apoptosis of anterior horn motoneurons in the sham operation group,the injury group and the β-elemene treatment group.Growth-related protein GAP43 expression and BBB score were used to evaluate hindlimb motor function in each group.Resultβ-elemene below the concentration of 40 ug/ml promotes the proliferation of VSC4.1 cell under normal culture conditions and promotes the survival of VSC4.1 under the hypoxia injury of cobalt chloride.Compared with the cobalt chloride-injured group,the expression of grp78,chop,cleaved-caspase 12 and cleaved-caspase3 protein was significantly decreased in the β-elemene-treated group,and the apoptotic cells were decreased,β-elemene and Y27632 promotes neurites outgrowth in primary cortical neurons,β-elemene reduces RhoA kinase activation,β-elemene and Y27632 promotes GAP43 protein expression,and inhibites the phosphorylation of limk and confilin proteins.In rat experiments,β-elemene treatment attenuates spinal cord anterior horn motor neuronal apoptosis and up-regulates GAP43 protein expression,and BBB score is significantly increased.ConclusionLow concentration of β-elemene reduces apoptosis of motor neurons by reducing endoplasmic reticulum stress,promotes its survival,and promotes neuronal neurite outgrowth after spinal cord injury by inhibiting RhoA pathway,ultimately promoting motor function recovery in rats.