Quantitative Proteomics Of Cerebrospinal Fluid(CSF) ITRAQ For The Improvement Of Spinal Shortening In Canine Cervical Syringomyelia

Spinal syringomyelia is a chronic progressive spinal cord disease.With the increase of the syringomyelia,it will cause damage to the surrounding spinal cord tissue and cause a series of complications such as sensory dysfunction,movement disorders,skin and neurotrophic disorders.Previous studies have suggested that cerebrospinal fluid circulation and decreased spinal cord tension are important factors affecting SM.At present,there is still a lack of a systematic intervention in clinical practice.In the previous clinical research,it has been found that spine shortening after phase I total spinal resection not only reduces the tension of the spinal cord and guarantees the safety of the spinal cord,but also effectively corrects the spinal deformity.Improve the CSF circulation disorder,and then achieve the control and improvement of postoperative SM.However,at present,a series of biological changes caused by the shortened spinal cord tension caused by the shortened spinal cord tension of PVCR surgery are not clear.Objective:On the basis of the animal model of non-communicating canine cervical SM,the animal model of PVCR spinal shortening for the treatment of canine cervical SM was further established,and the canine CSF samples were dynamically extracted.The DEPs in cerebrospinal fluid at different time points during SM reduction induced by spinal cord tension reduction were selected by using iTRAQ techniques of proteomics combined with LC-MS/MS.Furthermore,The DAVID was used to inquire and analyze the cellular signal transduction pathways involved in DEPs,and related signal transduction pathways and protein-protein interaction analysis were used to understand the pathogenesis of SM to screen out potential protein biomarkers of SM,so as to provide a new direction for the further study of proteins related to the pathogenesis of SM.Methods:The cervical SM model of non-communicating dogs was established by injecting trace Kaolin into the parenchyma of the corresponding spinal cord in the 7th cervical vertebra of beagle dogs.Before and 6 weeks after modeling,these dogs with SM were selected by MRI.According to the standard technical process of PVCR,PVCR of the first lumbar vertebra was performed,and the pedicle screw-rod system was used to compress the space of PVCR with a quantifiable spinal displacement controller.Cerebrospinal fluid samples were obtained from I(before cervical SM),I 2W(1 week after cervical SM),?(1 week after PVCR spinal shortening)and ? 1W(2 weeks after PVCR spinal shortening).The DEPs in cerebrospinal fluid at different time points during SM reduction induced by spinal cord tension reduction were screened by iTRAQ technique.The DEPs were annotated by GO,KEGG pathway annotation and enrichment analysis.Combined with the study of STRING protein interaction network and ELISA verification technique,the core regulatory pathway and related proteins in the biological process of SM reduction induced by spinal cord tension reduction were screened.Results:A total of 442 proteins were identified in group ?2W/?,of which 152 were DEPs(P<0.05),including 28 up-regulated proteins and 124 down-regulated proteins.A total of 562 proteins were identified in group ? 1 W/?,of which 133 were DEPs,including 40 up-regulated proteins and 93 down-regulated proteins.This study focused on the DEPs in group ? 1W/?,and the GO enrichment analysis was carried out on these DEPs.The molecular functional annotation analysis showed that the extracellular matrix tissue components and the activity of cell adhesion molecules were the main components.Cell component annotation analysis showed that extracellular matrix and foreign bodies were the main components.Biological process annotation analysis showed that DEPs were involved in ten biological processes,including protein metabolism,cell growth/repair,immune response,metabolism and so on.The proportion of DEPs involved in protein metabolism and cell growth/repair was 16.8%and 16.2%,respectively.GO and KEGG enrichment analysis of DEPs in DAVID showed that PI3K/AKT signal pathway played a dominant role in the whole process of SM,which included ten key proteins(VWF,CSF1,COL6A3,YWHAB,COL1A2,RELN,VTN,COL1A1,KIT and SPP1).Among them,the expression of COL1A1,COL1A2,SPP1 and CSF1 was verified by ELISA.Combined with the results of HPLC-MS protein detection,the expression trend of COL1A1,COL1A2 and SPP1 was consistent with that of iTRAQ.Conclusion:1.PI3K/AKT signaling pathway-mediated apoptosis plays an important role in the formation of kaolin-induced SM.Spinal shortening and spinal cord hypotension can activate the PI3K/AKT signaling pathway to inhibit apoptosis and play a repairing role in SM.2.Differentially expressed proteins such as SPPI are closely related to SM formation and improvement after intervention,and can be used as an in-depth study of potential biomarkers for SM.