Ethanol has been found to affect pulmonary cells by interfering with vitamin D metabolism and pulmonary defense mechanisms. The objective of this study was to understand the mechanisms of ethanol's disruptive influence on the vitamin D pathway and inhibition of anti-microbial peptide cathelicidin (LL-37). Bronchial epithelial cells (BEAS-2Bs), primary human bronchial epithelial cells (HBECs), primary human alveolar epithelial cells (HPAEpiCs), and human monocyte cells (THP-1s) were used in this study. These cells were cultured and exposed to different treatment groups: medium-only control, ethanol (70 mM) only, diallyl disulfide (DADS) (10 μM) -only, and a co-exposure of ethanol (70 mM) and DADS (10 μM) for 10 or 24 hours. Calcidiol (50 ng/mL) and calcitriol (0.05 ng/mL) dose-response studies were conducted for 48 hours. After incubation, cells were trypsinized, lysed, and centrifuged, and the cellular lysate was prepared for assay. Protein was quantified, and levels of inactive vitamin D [25(OH)D 3 ], active vitamin D [1, 25(OH) 2 D 3 ], and anti-microbial peptides (cathelicidin/ LL-37) in the samples were assayed using commercially available ELISA kits. In the ethanol-exposed group, cellular lysate concentrations of 25(OH)D 3 and LL-37 were significantly reduced by 30%, and 40% in BEAS-2B cells, and 35% and 80% in HPAEpi cells respectively. Overall 1, 25(OH) 2 D 3 cellular lysate levels were lower but followed a similar trend as the 25(OH)D 3 response. LL-37 levels in primary bronchial, alveolar cells, and ThP-1 cells were statistically reduced in ethanol-exposed groups (60%, 80%, and 65%, respectively) when compared with control. Following the addition of DADS, levels of LL-37 were recovered to within control levels for all three cell types.This study establishes two clinically relevant observations: that the exposure of pulmonary epithelial and monocyte cells to physiologically relevant levels of excessive ethanol selectively disrupts the activation of pulmonary vitamin D and inhibits the presence of antimicrobial peptide (LL-37) in vitro, and the co-exposure of DADS significantly attenuates ethanol-induced intracellular LL-37 depletion.