Sulfur mustard (SM) is a vesicant and pulmonary damaging agent that is still a threat in war and terroristic attacks. Although it was used as chemical warfare agent in world war I., a specific therapeutic against it is a still lacking. Models for investigation of the mechanisms of damage of the agent include animals and cell cultures. While animal lungs are less appropriate to follow biochemical changes because of complexity of the tissues, in monotypic cell cultures interactions of different cell types may be overseen. Therefore, a human alveolar co-culture model based on a human lung cell line with characteristics of type II pneumocytes and Clara cells (NCI H441) with human microvascular endothelial cell line (ISO-HAS-1) was developed. The in vitro bilayer models consist of NCI H441 in co-culture with ISO-HAS-1 on opposite sides of a permeable 24-well filter plate (Polycarbonate, Costar). To generate a polarized epithelial cell monolayer with typical junctional structures co-cultures were treated with dexamethasone starting at day 3 of co-cultivation. Functional barrier properties were investigated by measuring trans-bilayer electrical resistance (TER) and paracellular transport of sodium-fluorescein. Cell-cell contacts were characterized immunocytochemically using VE-cadherin and PECAM-1 as a marker for EC and ZO-1 and E-cadherin for epithelial cells. Immunofluorescence was performed using standard techniques. In co-culture of ISO-HAS-1 and NCI H441 the latter cells established contact inhibited monolayers, showing a continuous, circumferential immunostaining of the tight junctional protein, ZO-1 and the adherens junction protein, E-Cadherin. A phenotypically stable co-culture could be maintained for up to 14 days. This time period should permit examination of the effect of toxic compounds and related metabolites on barrier properties. We are currently investigating the influence of SM on NCI H441 and ISO-HAS-1 cell cultures and co-cultures. SM induced a concentration dependent apoptosis in both cell lines, which was confirmed by PARP cleavage and TUNEL staining. SM concentrations > 300 μM induced a time dependent increase of permeability. Conclusion. Our co-culture systems of NCI H441 with ISO-HAS-1 should provide a suitable in vitro alveolar model to examine the influence of highly toxic compounds such as chemical warfare agents on tight junction opening or cleavage in the distal lung.