At present, the predictability of cell culture-based methods to assess the effects of substances on the human body is limited, as they fail to emulate organ complexity and cross-talk. Biology-inspired microphysiological systems, such as TissUse’s Multi-Organ-Chip (MOC) platform, provide pre-clinical insights into absorption, distribution, metabolism, and toxicity of substances on a systemic level using human tissues. Here we describe a co-culture of a human liver equivalent based on the HepaRG™ cell line, combined with human stellate cells and a bronchial equivalent based on the MucilAir™ model. The coculture has been designed to elucidate the toxicity of inhaled compounds and to predict their effects and metabolism in a trans-organ environment. To address this challenge, a new MOC design and adequate co-culture conditions were established. Compared with an earlier version, the MOC was redesigned to optimize medium supply as well as to allow better oxygenation of the organ models. The tissue constructs were integrated in separate culture compartments of the closed circulatory perfusion system, interconnected by microfluidic channels, for up to 14 days. Tissue viability and homeostasis could be demonstrated by adenosine triphosphate-based cell viability assay, lactate dehydrogenase release, and metabolic profiling. Oxygenation was monitored over the culture period using an oxygen sensor (PreSens). Integrity and function of MucilAir™ tissues were additionally evaluated by histological analysis and measurements of trans-epithelial electrical resistance and cilia beat frequency. Furthermore, immunohistochemistry, gene expression analysis, and albumin secretion quantification verified liver tissue function in the course of the co-culture. In summary, the new MOC setup enables exposure studies of inhaled substances in order to investigate their toxic effects on a trans-organ level, emulating systemic substance effects on the human body.