Fig 2: PGAM5 is up-regulated in response to TBI A C57BL/6 J male mice were anesthetized and the dura mater of left hemisphere was exposed by a craniectomy. The sensorimotor cortex was impacted by CCI device at different velocity and deformation depth to mimic mild, moderate and severe TBI. B TTC staining and immunostaining of NeuN proteins of CCI mouse brains on 4 dpi. White dashed lines indicate the boundaries of the impacted lesion. Scale bar, 2 mm (upper) and 0.5 mm (bottom). C Immunoblot of PGAM5 in tissue lysate collected from left hemisphere on 4 dpi. D Quantification of relative PGAM5 levels normalized to GAPDH levels. Data are presented as mean ± SEM (n = 6). * p < 0.05, one-way ANOVA with Tukey’s multiple comparisons. E Rat cortical neurons, hippocampal neurons and glia cells were isolated and cultured in vitro on DIV0. Primary neurons were scratch-injured with a pipette on DIV 8. Injured area was between the two black dashed lines. F Immunoblot of PGAM5 proteins in primary cortical neurons and glia cells. The numbers indicate the relative PGAM5 level normalized to GAPDH. Asterisk indicates the non-specific bands

Fig 3: miR-26a-5p directly targets Wnt5a and the cellular localization of Wnt5a in the dorsal horn of the spinal cord. (A) Venn diagram showing target mRNAs for miR-26a-5p from four databases. (B) Gene ontology enrichment analysis of target genes for miR-26a-5p. (C) mRNA level of gene associated with inflammation in Sham, CFA/NC, and CFA/miR-26a-5p groups. (D) Relative expression of miR-26a-5p in Sham and CFA groups. (E) Protein level of Wnt5a in Sham, CFA/NC, and CFA/miR-26a-5p groups. The CFA/miR-26a-5p group decreased significantly compared with CFA/NC group, n = 6 mice/group. Data are represented as mean ± sem. **p < 0.01. ##p < 0.01. (F) Immunofluorescent study revealed Wnt5a (red) localized within GFAP astrocyte/IBA1 microglia/NeuN neuron (green) in the L4-5 spinal dorsal horn of CFA-induced inflammatory pain mice. Wnt5a co-localized mostly with NeuN neuron. Some appeared in the IBA1 microglia. Few could be detected in the GFAP astrocyte. The blue spots are DAPI nuclear staining (scale bar: 50 µm).