Smoking represents a major risk factor for chronic obstructive pulmonary disease (COPD), but it is difficult to characterize smoke-induced injury responses under physiological breathing conditions in humans due to patient-to-patient variability. Here, we show that a small airway-on-a-chip device lined by living human bronchiolar epithelium from normal or COPD patients can be connected to an instrument that “breathes” whole cigarette smoke in and out of the chips to study smoke-induced pathophysiology in vitro. This technology enables true matched comparisons of biological responses by culturing cells from the same individual with or without smoke exposure. These studies led to identification of ciliary micropathologies, COPD-specific molecular signatures, and epithelial responses to smoke generated by electronic cigarettes. The smoking airway-on-a-chip represents a tool to study normal and disease-specific responses of the human lung to inhaled smoke across molecular, cellular and tissue-level responses in an organ-relevant context. Display omitted •Smoking lung airway chip recapitulated clinical oxidative stress molecular profiles•New smoke-induced ciliary micropathologies were identified•This technology supported study of potential toxic effects of electronic cigarettes•COPD-specific responses were reproduced in vitro and novel biomarkers were identified Benam et al. describe a microengineered in vitro model system that permits analysis of the effects of whole smoke, from both conventional tobacco and electronic cigarettes, delivered under physiologically relevant flow conditions that mimic breathing on the pathophysiology of differentiated human mucociliated bronchiolar epithelium cultured in a microfluidic small airway-on-a-chip.