Differential regulation of dual NADPH oxidases/peroxidases, Duox1 and Duox2, by Th1 and Th2 cytokines in respiratory tract epithelium

Abstract

Partially reduced metabolites of molecular oxygen, superoxide (O ÅÀ2 ) and hydrogen peroxide (H 2 O 2 ), are detected in respiratory tract lining fluid, and it is assumed that these are key components of innate immunity. Whether these reactive oxygen species (ROS) are produced specifically by the respiratory epithelium in response to infection, or are a non-specific by-product of oxidant-producing inflammatory cells is not well characterized. Increasing evidence supports the hypothesis that the dual function NAD(P)H oxidases/peroxidases, Duox1 and Duox2, are important sources of regulated H 2 O 2 production in respiratory tract epithelium. However, no studies to date have characterized the regulation of Duox gene expression. Accordingly, we examined Duox1 and Duox2 mRNA expression by real-time PCR in primary respiratory tract epithelial cultures after treatment with multiple cytokines. Herein, we determined that Duox1 expression was increased several-fold by treatment with the Th2 cytokines IL-4 and IL-13, whereas Duox2 expression was highly induced following treatment with the Th1 cytokine IFN-c. Duox2 expression was also elevated by polyinosine-polycytidylic acid (poly(I:C)) and rhinovirus infection. Diphenyleneiodonium (DPI)-inhibitable apical H 2 O 2 production was similarly increased by the addition of Th1 or Th2 cytokines. These results demonstrate for the first time the regulation of Duox expression by immunomodulatory Th1 and Th2 cytokines, and suggest a mechanism by which ROS production can be regulated in the respiratory tract as part of the host defense response.

NADPH oxidases in lung biology and pathology: Host defense enzymes, and more

“…The differences in DUOX1/2 promoter regions are consistent with the highly variable transcriptional regulation of airway epithelial DUOX1 and DUOX2 in response to pro-inflammatory cytokines or to bacterial or viral stimuli (36,83,96). For example, DUOX1 mRNA expression in primary airway epithelial cultures is induced 3-5 fold by the Th2 cytokines interleukin (IL)-4 and IL-13 (83), which induce ciliatedto-mucus cell transdifferentiation and mucus hyperplasia during allergic airway inflammation (97,98). DUOX1 expression is also dramatically enhanced after hormonal differentiation of fetal lung epithelial cells to alveolar type II cells (90).…”

“…Expression of DUOX1 and DUOX2 in thyrocytes was found to be inducible in response to cAMP stimulation by forskolin (12,92), but this was not observed in human epithelial cells ( (90) and unpublished results), and the human DUOX promoters does not appear to contain a cAMP-responsive element (94). The differences in DUOX1/2 promoter regions are consistent with the highly variable transcriptional regulation of airway epithelial DUOX1 and DUOX2 in response to pro-inflammatory cytokines or to bacterial or viral stimuli (36,83,96). For example, DUOX1 mRNA expression in primary airway epithelial cultures is induced 3-5 fold by the Th2 cytokines interleukin (IL)-4 and IL-13 (83), which induce ciliatedto-mucus cell transdifferentiation and mucus hyperplasia during allergic airway inflammation (97,98).…”

“…The fact that ROS production by polarized epithelia occurred only apically and could be stimulated by activators of protein kinase C (82), known to activate the phagocyte NADPH oxidase (5), suggested regulated and compartmentalized epithelial ROS production by an NADPH oxidase. The source of epithelial ROS was identified only a few years ago, after initial discovery of prominent expression of the NOX homologs DUOX1 and DUOX2 within the lung (35), and studies on lung tissues using in situ hybridization and immunohistochemistry, which revealed the presence of DUOX1 at the apical surface of tracheobronchial epithelial cells, and DUOX2 within salivary and submucosal glands (19,79,83,84 (88,89), which is due to the presence of DUOX1 (90). Although DUOX expression in the lung is primarily localized to airway or alveolar epithelial cells, recent studies have also suggest the presence of DUOX proteins in lymphocytes (51,91).…”

“…To date, relatively little is known about transcriptional regulation of DUOX (83,90,94). DUOX1 and its maturation factor DUOXA1 are both located on chromosome 15q15.3 in a complex locus in a head-to-head arrangement (92)(93)(94), and may share a bidirectional promoter.…”

“…DUOX1 expression is also dramatically enhanced after hormonal differentiation of fetal lung epithelial cells to alveolar type II cells (90). In contrast, airway epithelial expression of DUOX2, but not DUOX1, is dramatically upregulated by the Th1 cytokine IFN-γ, and after rhinovirus or bacterial infection (36,83,96). DUOX1/2 expression in human thyrocytes was recently found to be downregulated in the presence of IL-1α/IFN-γ (99).…”