Abstract
Purpose :
We previously reported structural and functional sequelae 8 months following acoustic blast overpressure (ABO) exposure using a rodent model. Here we report quantitative analysis of retinal gliosis and microglia distribution under the same conditions, as well as cytokine expression 48 h post-blast (PB).
Methods :
Anesthetized adult male Long-Evans rats were exposed to a single blast (190 dB SPL, ~63 kPa) with a shock tube, as previously described (Allen et al., 2018). Eight months post-blast (PB), blast-exposed and age/sex-matched unblasted controls (UBC) were euthanized, eyes (N=7/group) were fixed, embedded, and cryosections were subjected to immunohistochemistry using polyclonal anti-GFAP (Müller glia) and IBA-1 Mab (microglia), with fluor-conjugated secondary antibodies, and imaged by confocal microscopy. Individual GFAP+ filaments were traced using Simple Neurite Tracer (Image-J®) in defined Z stack regions (0.5 mm/slice), then volume-filled (Djikstra’s algorithm, threshold 0.01), and corrected for total sample volume. For microglia distribution, IBA-1 signal was quantified in thresholded binary images in FIJI (B90-W255). Cytokine levels were assessed in pooled retina lysates at 48 h PB vs. UBC (3 tech. replicates/group; R&D systems Proteome Profiler Rat Cytokine Array and Bio-Rad ChemiDoc™). Student’s t-test was applied for statistical analyses.
Results :
At 8 mo. PB, GFAP levels in ipsilateral blast-exposed (IB) eyes were ≥1.5-fold greater than in UBC eyes (P=0.0025) with regional variability. Mean % total sample volume occupied by filled GFAP filaments were: UBC, 0.14%; IB, 0.28% (P=0.0039); mean volume-normalized filament lengths were: UBC, 0.0028; IB, 0.0043 (P=0.0019). IBA-1+ microglia in UBC retinas were predominantly localized in the ganglion cell layer-inner plexiform layer region, whereas IB retinas showed apparent shift of a subpopulation of IBA-1+ cells toward the inner nuclear layer/outer plexiform layer. Cytokine analysis suggested elevated platelet derived CXCL7 (Cys-X-Cys motif ligand 7), Fractalkine, and ICAM-1 levels in IB vs. UBC retinas.
Conclusions :
ABO exposure caused elevated regional gliosis and microglial activation and migration in the retina, which persisted for up to 8 months PB, compared to UBC. At 48 h PB, key pro-inflammatory cytokines also were elevated in this ABO model.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.