Spinal Cord Injury Affects Gene Expression of Transmembrane Proteins in Tissue and Release of Extracellular Vesicle in Blood: In Silico and In Vivo Analysis

Objective: Spinal cord injury (SCI) can disrupt membrane transmission by affecting transmembrane channels orneurotransmitter release. This study aimed to explore gene expression changes of transmembrane proteins underlyingSCI through bioinformatics approaches and confirming in SCI model in rats.Materials and Methods: In this experimental study, the differentially expressed genes (DEGs) in acute and subacuteSCI were obtained based on microarray data downloaded from the gene expression omnibus (GEO). Transmembraneproteins of DEGs were recognized by using the UniProt annotation and transmembrane helices prediction (TMHMM)methods. The model of SCI was established through a weight-dropping procedure in rats. To confirm the SCI model,hematoxylin and eosin (H E) staining was performed. Total mRNA was extracted from spinal cord tissues, and the RNAexpression profile of some of the significantly changed genes in the previous part that has been confirmed by real-timepolymerase chain reaction (PCR). Blood was collected from rats before sacrificing. Extracellular vesicles (EVs) wereisolated by high-speed centrifugation from plasma. For the assessment of protein expression, western blotting wasused.Results: Based on bioinformatics analysis, we candidated a set of membrane proteins in SCI’s acute and sub-acutephases, and confirmed significant upregulation in Grm1, Nrg1, CD63, Enpp3, and Cxcr4 between the acute and controlgroups and downregulation in Enpp3 between acute and subacute groups at the RNA level. Considering CD63 as anEV marker, we examined the protein expression of CD9 and CD63 in the plasma-derived EVs, and CD9 has significantexpression between acute and control groups. We also demonstrate no significant CD63 and Cxcr4 expressionsbetween groups.Conclusion: Our results provide new insight into the relationship between candidate transmembrane protein expressionand different stages of SCI using in-silico approaches. Also, results show the release of EVs in blood in each group afterSCI helping enlarge strategies to enhance recovery following SCI.