Abstract

Sulfur mustard (SM), a vesicating agent first used during World War I, remains a potent threat as a chemical weapon to cause intentional/accidental chemical emergencies. Eyes are extremely susceptible to SM toxicity. Nitrogen mustard (NM), a bifunctional alkylating agent and potent analog of SM, is used in laboratories to study mustard vesicant-induced ocular toxicity. Previously, we showed that SM-/NM-induced injuries (in vivo and ex vivo rabbit corneas) are reversed upon dexamethasone (DEX) treatment, an FDA approved, steroidal anti-inflammatory drug. Here, we optimized NM injuries in ex vivo human corneas and assessed DEX efficacy. For injury optimization, one cornea (randomly selected from paired eyes) was exposed to NM: 100 nmoles for 2 h or 4 h, and 200 nmoles for 2 h, and the other cornea served as a control. Injuries were assessed 24 h post NM-exposure. NM 100 nmoles exposure for 2 h was found to cause optimal corneal injury (epithelial thinning [~69%]; epithelial-stromal separation [6-fold increase]). In protein arrays studies, 24 proteins displayed {greater than or equal to}40% change in their expression in NM exposed corneas compared to controls. DEX administration initiated 2 h post NM exposure and every 8 h thereafter until 24 h post-exposure reversed NM-induced corneal epithelial-stromal separation [2-fold decrease]). Of the 24 proteins dysregulated upon NM exposure, 6 proteins (DLL1, FGFbasic, CD54, CCL7, endostatin, ERBB4) associated with angiogenesis, immune/inflammatory responses, and cell differentiation/proliferation, showed significant reversal upon DEX treatment (Student's t-test; p{less than or equal to}0.05). Complementing our animal model studies, DEX was shown to mitigate vesicant-induced toxicities in ex vivo human corneas.

Significance Statement NM exposure-induced injuries were optimized in an ex vivo human cornea culture model and studies were carried out at 24 h post 100 nmoles NM exposure. DEX administration (started 2 h post NM exposure and every 8 h thereafter) reversed NM-induced corneal injuries. Molecular mediators of DEX action were associated with angiogenesis, immune/inflammatory responses, and cell differentiation/proliferation, indicating DEX aids wound healing via reversing vesicant-induced neovascularization (DLL1 and FGF basic) and leukocyte infiltration (CD54 and CCL7).