The gastrointestinal epithelium functions as physical and innate immune barriers against commensal or pathogenic microbes. NADPH oxidase 1 (Nox1) and dual oxidase 2 (Duox2), highly expressed in the colon, are suggested to play a potential role in host defense. Guinea-pig gastric pit cells and human colonic epithelial cells (T84 cells) express Nox1. With regard to activation of Nox1, the gastric epithelial cells are primed with Helicobacter pylori lipopolysaccharide, whereas T84 cells preferentially use the Toll-like receptor (TLR) 5, rather than TLR4, against Salmonella enteritidis infection. Thus, gastric and colonic epithelial cells may use different TLR members to discern pathogenicities among bacteria, depending on their environments and to activate Nox1 appropriately for host defense. Nox1-derived reactive oxygen species (ROS) have been implicated in the pathogenesis of inflammation-associated tumor development. The human stomach does not express Nox1. Helicobacter pylori infection alone does not induce it, whereas Nox1 is specifically expressed in gastric adenocarcinomas. In the human colon, Nox1 is differentiation-dependently expressed, and its expression is upregulated in adenomas and well-differentiated adenocarcinomas. Although Nox1 expression may not be directly linked to mitogenic activity, Nox1-derived ROS may exert a cancer-promoting effect by increasing resistance to programmed cell death of tumor cells.