Laminin in the cutaneous basement membrane as a potential target in lewisite vesication

The epidermal-dermal junction has a complex molecular architecture, with numerous components playing key roles in adhesion of the epidermis to the dermis. The purpose of this study was to examine structural components of the epidermal-dermal junction as potential targets for toxicity by lewisite (dichloro(2-chlorovinyl)arsine). This was accomplished by (1) immunocytochemical mapping of laminin, type IV collagen, and bullous pemphigoid antigen (BPA) in lewisite-treated isolated perfused porcine skin flaps (IPPSF), (2) evaluation of protease activity in IPPSF blister fluid against laminin substrate from murine EHS tumor and human keratinocytes, and (3) examination of human keratinocyte laminin for direct chemical modification by lewisite. Lewisite-induced epidermal-dermal separation was localized to the lamina lucida. Localization of the separation suggested that laminin, a cysteine-rich and highly protease-sensitive adhesive glycoprotein, is a potential target for lewisite action. It was hypothesized that chemical modification of laminin directly (via chemical alkylation of laminin thiols by the arsenical) or indirectly (due to lewisite-induced cytotoxic release of proteases) could result in blister formation. Employing sensitive methodology, no evidence of proteolytic activity against EHS tumor laminin or human keratinocyte laminin was identified in the blister fluid. In addition, no evidence for direct chemical modification of laminin by lewisite was demonstrated. However, up to 36% of the thiol groups in human keratinocyte laminin immunoprecipitates was potentially available for reaction with alkylating agents. While these studies did not demonstrate a lewisite-induced chemical modification of laminin, they do not rule out the possibility that other adhesive molecules of the basement membrane are targets for lewisite action. Further evaluation of the molecular role that these binding modalities play in vesicant-induced separation may provide new insights into therapeutic and prophylactic strategies against the toxicity of such compounds and contribute to a better understanding of basement membrane biochemistry.