Ferroptosis In Spinal Cord Injury And SRS 16-86 Inhibition

Object:Spinal cord injury(SCI)is a severe traumatic disease with high incidence rate and cast high pressure to patients’ family and the whole society.Due to the lack of effective treatment and less than 1% of SCI patients have complete neurological recovery,It has been recognized as the world medical problems for a long time.At present,the major difficult of spinal cord injury repair is how to effectively deal with the large number of cell death caused by the secondary injury.Controlling secondary injury and protecting neuron cells become the main process to treat SCI.Among the mechanisms of secondary injury,oxidative stress induced by free radicals is considered to be one of the most important factors.The overload of intracellular iron after secondary spinal cord injury also has an effect on lipid peroxidation.In recent years,the traumatic injury mechanism of brain,heart,kidney and other tissues have shown that the increased free iron level after injury can stimulate the generation of free radicals,which leading to lipid peroxidation and cell dysfunction.Iron chelating agents and antioxidant drugs have a certain effect on repairing SCI.However,the underlining mechanism is not clear,which limits the application in SCI.Recent studies has proposed a new mechanism of programmed cell death –Ferroptosis,which may exist in spinal cord injury and ferroptosis inhibitors may have treatment effects on SCI.Ferroptosis is found in ischemia-reperfusion injury,and ferroptosis inhibitors can apparently relieve tissue damage.This study aims to find out whether ferroptosis was involved in spinal cord injury and the treatment effect of ferroptosis inhibitor on SCI.This study designs the following experiments to verify the hypothesis:(1)ferroptosis exists in SCI and neural cell death in vivo.(2)ferroptosis inhibitors have protective effects on SCI and neuronal cell death.Method:In vivo experiments showed that Ferroptosis exists in spinal cord injury and can be abolished by ferroptosis inhibitor(SRS16-86).BBB scale was used to evaluate the locomotor function after spinal cord injury.Immunohistochemical staining was used to evaluate the pathological changes including neurons,glial scars and oligodendrocytes.The detection of ferroptosis markers proved its presence in SCI.Wistar rats were divided into three groups: SCI+ferroptosis inhibitor group,injury group and Sham group.The ultrastructural changes of spinal cord were observed by electron microscopy,and the characteristic changes of ferroptosis were also observed.The levels of ROS in different groups were observed by ROS fluorescent probe staining.The iron level in the spinal cord was measured.The apoptosis profile of spinal cord was observed by TUNEL staining.Immunofluorescence staining was used to observe the number and proportion of astrocytes,neurons and oligodendrocytes in injured spinal cord.Results:Rats SCI model was successfully produced verified by histopathological and functional scoring.The SRS16-86,inhibitor of Ferroptosis was successfully synthesized and verified by mass spectrometry.After treated with SRS16-86 in spinal cord injury rats,the content of ROS metabolite 4HNE and iron ion level reduced and the total glutathione increased dramatically in the injured area.And the application of SRS16-86 can improve the BBB scores,ameliorate the histopathological changes,inhibit the proliferation of astrocytes,reduce the formation of glial scar,increase the numbers of neurons,increase the area of CNPase positive area;and SRS16-86 can also reduce the expression of TNF-α,IL-1β and ICAM1 in injured spinal cord and inhibit the inflammatory response after SCI.Conclusion:In this study,we confirmed the presence of ferroptosis in spinal cord injury.As an inhibitor of Ferroptosis,SRS16-86 significantly improved hindlimb motor function,histopathological changes,protect neurons and oligodendrocytes,reduce the proliferation of astrocytes after spinal cord injury,thereby reducing the formation of glial scar in the late stage.