N-nitrosodimethylamine induces cytochrome P450 activation but not epithelial remodeling in well-differentiated human airway epithelial cells.

N-nitrosodimethylamine (NDMA) is a volatile environmental contaminant formed from diverse precursor chemicals widely present in byproducts of industrial processes, personal care products, and pharmaceuticals. Therefore, NDMA is frequently detected in both indoor and outdoor air. However, its potential risk from inhalation exposure has been largely underestimated and the mechanisms underlying its toxicity remain insufficiently studied. To address this knowledge gap, we investigated the effects of NDMA on primary human bronchial epithelial (HBE) cells differentiated in air-liquid interface cultures, a physiologically relevant in vitro model that mimics the structure and function of the human airway epithelium. To determine time and dose-dependent responses, we exposed well-differentiated HBE cells to NDMA at increasing concentrations (0.05 - 5 mM) either basolaterally for up to 24 h or apically for 4 h daily over 1, 3, or 5 days. We then assessed cellular responses linked to epithelial cell functions, including cell viability, epithelial-to-mesenchymal transition (EMT), epithelial junction integrity, and xenobiotic metabolism, focusing on cytochrome P450 (CYP450) enzyme. NDMA exposure at concentrations up to 5 mM did not affect cell viability under any tested conditions. Similarly, we observed no evidence of EMT, as determined by mRNA expression of vimentin and ZEB1, and no disruption of epithelial junctions, as indicated by preserved expression and localization of E-cadherin and ZO-1. In contrast, 5 mM NDMA significantly increased mRNA expression and enzymatic activity of CYP450 in a time-dependent manner over 5 days. Together these data indicate that NDMA activates epithelial metabolic pathways in the absence of EMT or epithelial remodeling.