Acute Cutaneous Nitrogen Mustard Exposure Induces Persistent Fibrosis, Inflammation, and Psoriatic-Lesions in Mice Skin

Abstract ID 101872 Poster Board 214 Exposure to mustard vesicating, or blister, agents can lead to severe and acute long-term injuries such as blistering, systemic toxicity, secondary infections, and chronic inflammation. Sulfur mustard (SM) exposure has caused long-lasting cutaneous and respiratory complications in Iranian veterans of the Iran-Iraq War and may be a contributing factor in the development of Gulf War Illness in veterans of the Gulf War. SM was first used as a weapon during World War I (WWI) and has been used in a number of conflicts since, possibly including the recent Syrian Civil War. Accidental exposure to SM from stockpiles and its potential utility as a weapon of warfare and terror are the primary causes of concern. This is compounded by the fact that SM's long-term pathophysiology from cutaneous exposure and related mechanisms remain poorly understood and we thus lack effective therapies. This study aims to establish a mouse model of long-term toxic effects arising from acute cutaneous exposure to nitrogen mustard (NM, an analog of SM) and identify key biomarkers and mechanism(s) of long-lasting skin effects to inform future studies. The dorsal skin of male C57BL/6 mice was topically exposed to 0.5 mg NM dissolved in 100 mL of acetone. Mice were sacrificed and tissues collected at various timepoints during a 3-month study period. Histological assessments of injury progression were carried out by employing hematoxylin & eosin, toluidine blue, and Gomori's Trichrome staining of skin tissue sections. Expression of inflammatory, oxidative stress, and fibrotic markers were measured via qPCR analysis. A strong resemblance was seen between NM-induced skin lesions at days 29 and 84 post-exposure and lesions seen in mouse skin models of psoriasis. These similarities included epidermal acanthosis, pronounced rete ridges, hyperkeratosis, and dermal fibrosis in NM-exposed mice. Staining with Gomori's Trichrome, which stains collagen blue, showed a significant (p<0.05) increase in the levels of collagen staining in NM exposed skin as compared to control group at days 14 and 29 post-exposure and a trend (0.05<p<0.10) for increased collagen at day 84 post-exposure, indicating fibrosis. NM exposure also caused an increase in the expression of TGF-b (which is a marker of fibrosis) and IL-17 and IL-23 (which are the markers of psoriasis) in the skin of mice. The expression of inflammatory markers, such as TNF-a, IL-1b, MMP-9, IL-6, and COX-2, and the oxidative stress marker HO-1 were also seen to have significantly increased in the skin of NM-exposed mice. Increased mast cell degranulation was also seen in the skin of NM-exposed mice, with a bimodal pattern of an initial, statistically significant (p<0.05) increase at days 1 and 7 post-exposure followed by a decrease at day 14 post exposure, and becoming significantly increased (p<0.05) again at day 84post-exposure. Histological and molecular analyses have revealed that a single acute, cutaneous NM exposure in mice caused persistent inflammation and fibrosis with long-term effects including the development of psoriatic lesions . Such findings of persistent injury and the development of an inflammatory skin disease from NM exposure in mice are novel. Further studies to elucidate the role of mast cells in the long-term injury from mustard vesicants are underway which could be helpful in understanding the effects in veterans exposed to SM and in identifying effective medical countermeasures.