Fig 1.
Ocular Sulfur Mustard (SM) exposure causes increase in corneal thickness and causes corneal opacity, corneal ulceration, and corneal neovascularization.
The right eyes of New Zealand white rabbits were exposed to SM (~0.4 mg/L) vapor either for 5 min (shorter duration) or 7 min (longer duration) and the clinical progression of ocular injury was observed starting from day 1 (6 h) up to 28 days post-exposure, as detailed in the materials and methods section. The left eye was designated as the control eye and was exposed only to the dilution air. Corneal thickness (A), corneal opacity and quantification (B), corneal ulceration and quantification (C), and corneal neovascularization and quantification (D). Representative pictures are from the 7 min SM exposure group. Green arrows, corneal-stromal injury (opacity); red arrows, corneal ulceration; and purple arrows, corneal neovascularization. Data presented are mean±SEM (n = 5-7/group). *p<0.05 for both 5 min and 7 min exposure.
Fig 2.
Ocular SM exposure causes increased corneal thickness and leads to epithelial degradation and epithelial-stromal separation.
The right eyes of New Zealand white rabbits were exposed to SM (~0.4 mg/L) vapor either for 5 min (shorter duration) or 7 min (longer duration) and the clinical progression of ocular injury was observed starting from day 1 (6 h) up to 28 days post-exposure, as detailed in the materials and methods section. The left eye was designated as the control eye and was exposed only to the dilution air. Cornea was dissected post euthanasia at day 1, day 3, day 7, day 14, and day 28 post-exposure and histopathological evaluation. Corneal thickness and quantification (A), epithelial degradation and quantification (B), epithelial-stromal separation and quantification (C) in the H&E-stained control and SM-exposed sections was carried out as detailed under the materials and methods. Representative images are from the 7 min SM exposure. Data presented are mean±SEM (n = 3–5). *, p<0.05 compared to the control group; e, epithelium; s, stroma; red arrows, epithelial degradation/epithelial-stromal separation; size bar in representative images, 50 μm.
Fig 3.
Ocular SM exposure causes keratocyte cell death, influx of inflammatory cells and increase in the number of blood vessels in the stroma.
The right eyes of New Zealand white rabbits were exposed to SM (~0.4 mg/L) vapor either for 5 min (shorter duration) or 7 min (longer duration) and the clinical progression of ocular injury was observed starting from day 1 (6 h) up to 28 days post-exposure, as detailed in the materials and methods section. The left eye was designated as the control eye and was exposed only to the dilution air. Cornea was dissected post euthanasia at day 1, day 3, day 7, day 14, and day 28 post-exposure and histopathological evaluation was carried out. Number of keratocytes (A), inflammatory cells (B) and blood vessels (C) were quantified in H&E-stained corneal sections as detailed under the materials and methods. Data presented are mean±SEM (n = 3–5). *, p<0.05 compared to the control group; red arrows, keratocytes/inflammatory cells/blood vessels; size bar in representative images, 50 μm.
Fig 4.
Ocular SM exposure causes increased expression of COX-2, VEGF, MMP-9, and IL-8.
The right eyes of New Zealand white rabbits were exposed to SM (~0.4 mg/L) vapor either for 5 min (shorter duration) or 7 min (longer duration) and the clinical progression of ocular injury was observed starting from day 1 (6 h) up to 28 days post-exposure, as detailed in the materials and methods section. The left eye was designated as the control eye and was exposed only to the dilution air. Corneas were dissected post euthanasia at day 1, day 3, day 7, day 14, and day 28 post-exposure and fixed for IHC evaluation or frozen for cytokine array analysis. The corneal sections were IHC stained and COX-2 (A), MMP-9 (B), and VEGF (C) expression was quantified. Corneal lysates (7 min exposure group day 3 post-exposure) were prepared and subjected to cytokine array analysis (D) and relative fluorescence units for IL-8, MMP-9, IL-1α, IL-1β, IL-17A, IL-21, Macrophage Inflammatory Protein (MIP)-1β, TNF-α, and leptin were calculated. Only IL-8 and MMP-9 showed changes in expression levels upon SM-exposure; whereas, IL-1α, IL-1β, IL-17A, IL-21, MIP-1β, TNF-α, and leptin served as control cytokines, with similar expression in SM exposed and unexposed eyes. Data presented are mean±SEM (n = 3–5). *, p<0.05 compared to the control group; e, epithelium; s, stroma; size bar in representative images, 50 μm.