Application of weighted gene co-expression network analysis to explore the targets of treadmill training in rats with spinal cord injury

Background Spinal cord injury (SCI) describes any damage to the spinal cord that results in a loss of function. With these injuries being commonly treated with subsequent treadmill training (TM), this study aimed to reveal the mechanisms facilitating TM-mediated rehabilitation following a SCI. Methods Differentially expressed genes (DEGs) were identified and then subjected to a weighted gene co-expression network analysis to identify TM-associated modules. Co-expression network construction, functional annotation, protein–protein interaction (PPI) network, and transcription factor (TF)-miRNA-target regulatory network were then conducted. We established an SCI rat model and validated key RNAs using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Predicted interactions between miR-135a-5p and JAK2 and STAT1 were tested using dual-luciferase reporter assay. Results Total 640 and 877 genes separately differentially expressed in SCI 8d + TM vs. SCI 8d and SCI 14d + TM vs. SCI 14d comparison groups were screened and merged as the DEGs, which were further narrowed to four TM-associated modules linked to the glutamatergic synapse and ribosome biogenesis pathways. PPI network evaluations identified v-Akt murine thymoma viral oncogene homolog 1 (AKT1), Janus kinase 2 (JAK2), tumor necrosis factor (TNF), and signal transducer and activator of transcription 1 (STAT1) as hub nodes. JAK2 and STAT1 were also shown to have interactions in module b. TF-miRNA-target regulatory network revealed that both STAT1 and JAK2 were targets of miR-135a. RT-qPCR results confirmed upregulation of AKT1 and STAT1 and downregulation of JAK2 , TNF and miR-135a after TM. Dual-luciferase reporter assays revealed the direct binding of miR-135a-5p to JAK2 and STAT1 . Conclusion These findings suggest that TM facilitates SCI recovery by regulating AKT1 , JAK2 , TNF , STAT1 and miR-135a.