A Novel Mechanism For The Treatment Of Spinal Cord Injury By Targeting IL17-JAK/STAT-VEGF Axis And The Development Of A New Complement Inhibitory Protein CD59

Spinal cord injury is a serious neurological disorder that causes neuronal degeneration and permanent nerve paralysis.The current research hotspot is to explore the secondary injury after primary injury of spinal cord and the treatment strategy for this secondary injury.Our experiment mainly explores the pathogenesis of spinal cord injury and proposes new treatment methods from two aspects.On the one hand,it is generally known that spinal cord injury causes inflammatory reactions in local tissues,and inflammation in turn contributes to the progression of the spinal cord injury.The activation of astrocytes during inflammatory reaction promotes the progression of spinal cord injury by inducing the formation of glial scars.Interleukin 17(IL-17)is also upregulated in inflammation following spinal cord injury.In this study,we found that IL-17 can induce astrocyte activation,which is manifested by the increase in the well-known marker glial fibrillary acidic protein(GFAP),both in vivo and in vitro.In addition,we demonstrate that the up-regulation of VEGF in astrocytoma cells is induced by IL-17.In our further study,the expression of VEGF was induced by IL-17 through activating JAK/STAT signaling pathway.Moreover,we found that IL-17 significantly reduces spinal tissue integrity,increases urinary retention,and impairs the blood-spinal cord barrier in the rat spinal cord injury model.This newly discovered IL-17-JAK / STAT-VEGF axis enhances our understanding of the molecular mechanisms underlying the inflammatory response after spinal cord injury and provides another potential target for spinal cord injury.On the other hand,we are interested in complement activation after spinal cord injury.A growing number of studies have shown that complement activation is closely related to the immune-inflammatory response in the secondary injury of the spinal cord.The complex immune-inflammatory response eventually leads to the activation of the complement cascade and the membrane attack complex(MAC)deposition on neurons,astrocytes,oligodendrocytes and other surfaces,resulting in a large number of cell death,causing demyelination and nerve necrosis.Thus,it shows that this complement-dependent cell-mediated cytotoxicity(CDCC)may have a huge impact on the prognosis of spinal cord injury.At present,there are few inhibitors for complement activation in the market,so it is particularly urgent to develop a drug that can target complement activation in the treatment of spinal cord injury.The CD59 protein is a natural complement inhibitory protein that is anchored on the surface of cell membranes by glycosylphosphatidylinositol(GPI),and it can inhibit the binding of C8 and C9 to inhibit the formation of MAC,thereby inhibiting complement activation-mediated cytotoxicity.Therefore,we are committed to developing CD59 protein as a clinical drug for the treatment of spinal cord injury and complement-related diseases.In the study,we first verified that the local tissue after spinal cord injury does indeed have complement activation and MAC deposition.Subsequently,a large amount of CD59 protein was successfully expressed by E.coli expression system in vitro.At this time,CD59 has no biological activity.Previous investigation reported that GPI structure is essential for the activity of CD59 protein,however,the CD59 protein expressed by E.coli lacks GPI structure,so we need to find a substance that can replace GPI to help the CD59 protein anchor on the cell membrane surface.To overcome this problem,we used peptide solid-phase synthesis(SPPS)technology to synthesize polypeptide molecules that can not only insert into the cell membrane but also attach to the CD59 protein,thus solving the problem of concentrating CD59 on the cell membrane surface.Subsequently,the classical complement hemolysis test was used to demonstrate that the polypeptide-modified CD59 has the activity of inhibiting MAC formation in vitro.We will continue to optimize this technology to improve the activity of CD59 in animals and the application of CD59 in the treatment of spinal cord injury as soon as possible.