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CHAPTER 4 Approximate location of injury is indicated by the purple shaded area. (b) Example images of IL1β (red), IBA1 (green), and DAPI (blue) triple positive cells from spinal cord injured untreated and light-treated groups in the dorsal region, ipsilateral to the injury at 7-dpi. (c-d) Quantification of IL1β+IBA1+DAPI+ cells, expressed as triple positive cell density within the region of interest, in the dorsal region of the spinal cord, contralateral (c) and ipsilateral (d) to the injury of untreated and light-treated groups. (e-f) IL1β+IBA1+DAPI+ cell density in the lateral regions of interest contralateral (e) and ipsilateral (f) to the injury. (g-h) IL1β+IBA1+DAPI+ cell density in the ventral regions of interest contralateral (g) and ipsilateral (h) to the injury. n values indicated (legend) are for each time point. Data is expressed as mean ± SEM. See Figure 4.1 for abbreviations. 4.4.7 Red light reduces iNOS expressing microglia/macrophages at the injury zone Another pro-inflammatory molecule that is associated with pain following spinal cord injury is NO, which can is by iNOS in microglia/macrophages (Hulsebosch et al., 2009). As microglia/macrophages only produce the iNOS isoform, and UNOS detects all three nitric oxide synthases isoforms (eNOS, nNOS, and iNOS), six spinal cord regions (Figure 4.10a) were triple stained for UNOS/IBA1/DAPI to investigate the density of iNOS expressing microglia/macrophages in the spinal cord following injury and red light treatment. An example of staining is shown in Figure 4.10b. Throughout the 6 regions (Figure 4.10c-h), there were significantly greater UNOS+/IBA1+ cell density on the ipsilateral side (p = 1.2e-05, LMER), a strong effect of time (p = 0.0005), and an overall effect of treatment (p = 0.020). In the dorsal region of the spinal cord, UNOS+/IBA1+ cells were maintained below 50 per mm2 on the contralateral side throughout the recovery period in both groups (Figure 4.10c). However, on the ipsilateral side (Figure 4.10d), there was an increase compared to the contralateral side (p = 0.07). After 1-dpi, UNOS+/IBA1+ cell density increased on the ipsilateral side (p = 0.077, 1-dpi vs 3-dpi; p = 0.004, 1-dpi vs 5-dpi; LMER) before decreasing at 7-dpi (p = 0.015, 5-dpi vs 7-dpi). UNOS+/IBA1+ cell density in the ipsilateral dorsal region was significantly reduced by red light treatment (p = 0.008; LMER). Figure 4.10e-h also demonstrated similar time effects, but there were no significant effects of light treatment at these deeper levels. In summary, these results suggest that iNOS expressing IBA1 cells are present, predominantly on the ipsilateral to the injury following hemicontusion, and red light treatment reduces iNOS expression in the ipsilateral dorsal region. 102PDF Image | Effects of Red Light Treatment on Spinal Cord Injury
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