Oxidative stress has been implicated in the pathogenesis of bronchial asthma. Besides granulocytes, the airway epithelium can produce large amounts of reactive oxygen species and can contribute to asthma-related oxidative stress. Histamine is a major inflammatory mediator present in large quantities in asthmatic airways. Whether histamine triggers epithelium-derived oxidative stress is unknown. We therefore aimed at characterizing human airway epithelial H2O2 production stimulated by histamine. We found that air-liquid interface cultures of primary human bronchial epithelial cells (BECs) and an immortalized BEC model (Cdk4/hTERT HBEC) produce H2O2 in response to histamine. The main source of airway epithelial H2O2 is an NADPH dual oxidase, Duox1. Out of the four histamine receptors (H1R-H4R), H1R has the highest expression in BECs and mediates the H2O2-producing effects of histamine. IL-4 induces Duox1 gene and protein expression levels and enhances histamine-induced H2O2 production by epithelial cells. Using HEK-293 cells expressing Duox1 or Duox2 and endogenous H1R, histamine triggers an immediate intracellular calcium signal and H2O2 release. Overexpression of H1R further increases the oxidative output of Duox-expressing HEK-293 cells. Our observations show that BECs respond to histamine with Duox-mediated H2O2 production. These findings reveal a mechanism that could be an important contributor to oxidative stress characteristic of asthmatic airways, suggesting novel therapeutic targets for treating asthmatic airway disease.