The airway epithelium is continuously subjected to environmental pollutants, airborne pathogens, and allergens and relies on several intrinsic mechanisms to maintain barrier integrity and to promote epithelial repair processes following injury. Here, we report a critical role for dual oxidase 1 (Duox1), a newly identified NADPH oxidase homolog within the tracheobronchial epithelium, in airway epithelial cell migration and repair following injury. Activation of Duox1 during epithelial injury is mediated by cellular release of ATP, which signals through purinergic receptors expressed on the epithelial cell surface. Purinergic receptor stimulation by extracellular ATP is a critical determinant of epithelial cell migration and repair following injury and is associated with activation of extracellular signalregulated kinases (ERK1/2) and matrix metalloproteinase-9 (MMP-9). Stimulation of these integral features of epithelial cell migration and repair processes was found to require the activation of Duox1. Our findings demonstrate a novel role for Duox1 in the tracheobronchial epithelium, in addition to its proposed role in antimicrobial host defense, by participating in epithelial repair processes to maintain epithelial integrity and barrier function in the face of environmental stress.One of the primary functions of the airway epithelium is to provide a protective barrier against inhaled environmental toxins and airborne pathogens, and airway epithelial cells express a number of intrinsic factors that serve to minimize invasion by infectious agents and to promote repair processes following injury (1, 2). A newly discovered NADPH oxidase homolog, dual oxidase (Duox), 3 has been recently identified within the tracheobronchial epithelium (3-6) and is thought to contribute to innate epithelial host defense based on structural similarities with the phagocyte NADPH oxidase system (7,8). Of the two known Duox isoforms, Duox1 is primarily expressed within the ciliated epithelium and believed to be responsible for epithelial H 2 O 2 production after cell stimulation (5), whereas Duox2 has been localized to salivary or submucosal glands, thus constituting a functional host defense system with co-localized lactoperoxidase (4, 5). Other than recent observations in Drosophila (9), a direct role for Duox1/2 in host defense has not yet been directly demonstrated, and other functions of airway epithelial Duox1 have been postulated (6, 10).Both Duox isozymes contain two EF-hand Ca 2ϩ -binding domains and are activated by Ca 2ϩ -mobilizing stimuli (11, 12). A critical mechanism of Ca 2ϩ -mediated signaling within epithelial cells after bacterial infection or mechanical or oxidative injury involves the activation of purinergic receptors at the cell surface by release of ATP (13-15). ATP-mediated autocrine or paracrine signaling is known to regulate diverse processes involved in host defense, including anion transport, ciliary function, and mucin expression (16 -19), and some of these events have recently been associated with Duox1 act...

The synthesis and reactivity of geometrically constrained tricoordinate phosphorus (σ(3)-P) compounds supported by tridentate triamide chelates (N[o-NR-C6H4]2(3-); R = Me or (i)Pr) are reported. Studies indicate that 2 (P{N[o-NMe-C6H4]2}) adopts a Cs-symmetric structure in the solid state. Variable-temperature NMR studies demonstrate a low-energy inversion at phosphorus in solution (ΔG(‡)(exptl)(298) = 10.7(5) kcal/mol), for which DFT calculations implicate an edge-inversion mechanism via a metastable C2-symmetric intermediate. In terms of reactivity, compound 2 exhibits poor nucleophilicity, but undergoes oxidative addition at ambient temperature of diverse O-H- and N-H-containing compounds (including alcohols, phenols, carboxylic acids, amines, and anilines). The resulting pentacoordinate adducts 2·[H][OR] and 2·[H][NHR] are characterized by multinuclear NMR spectroscopy and X-ray crystallography, and their structures (which span the pseudorotation coordinate between trigonal bipyramidal and square planar) are evaluated in terms of negative hyperconjugation. At elevated temperatures, the oxidative addition is shown to be reversible for volatile alcohols and amines.

Ammonia, alkyl amines, and aryl amines are found to undergo rapid intermolecular N-H oxidative addition to a planar mononuclear σ(3)-phosphorus compound (1). The pentacoordinate phosphorane products (1·[H][NHR]) are structurally robust, permitting full characterization by multinuclear NMR spectroscopy and single-crystal X-ray diffraction. Isothermal titration calorimetry was employed to quantify the enthalpy of the N-H oxidative addition of n-propylamine to 1 ((n)PrNH2 + 1 → 1·[H][NH(n)Pr], ΔHrxn(298) = -10.6 kcal/mol). The kinetics of n-propylamine N-H oxidative addition were monitored by in situ UV absorption spectroscopy and determination of the rate law showed an unusually large molecularity (ν = k[1][(n)PrNH2](3)). Kinetic experiments conducted over the temperature range of 10-70 °C revealed that the reaction rate decreased with increasing temperature. Activation parameters extracted from an Eyring analysis (ΔH(⧧) = -0.8 ± 0.4 kcal/mol, ΔS(⧧) = -72 ± 2 cal/(mol·K)) indicate that the cleavage of strong N-H bonds by 1 is entropy controlled due to a highly ordered, high molecularity transition state. Density functional calculations indicate that a concerted oxidative addition via a classical three-center transition structure is energetically inaccessible. Rather, a stepwise heterolytic pathway is preferred, proceeding by initial amine-assisted N-H heterolysis upon complexation to the electrophilic phosphorus center followed by rate-controlling N → P proton transfer.

Studies on the stoichiometric and catalytic reactivity of a geometrically constrained phosphorous triamide 1 with pinacolborane (HBpin) are reported. The addition of HBpin to phosphorous triamide 1 results in cleavage of the B-H bond of pinacolborane through addition across electrophilic phosphorus and nucleophilic N-methylanilide sites in a cooperative fashion. The kinetics of this process of were investigated by NMR spectroscopy, with the determined overall second order empirical rate law given by ν = − k[1][HBpin] where k = 4.76 × 10−5 M−1s−1 at 25 °C. The B–H bond activation process produces a P-hydrido-1,3,2-diazaphospholene intermediate 2, which exhibits hydridic reactivity capable of reacting with imines to give phosphorous triamide intermediates, as confirmed by independent synthesis. These phosphorous triamide intermediates are typically short-lived, evolving with elimination of the N-borylamine product of imine hydroboration with regeneration of the deformed phosphorous triamide 1. The kinetics of this latter process are shown to be first-order, indicative of a unimolecular mechanism. Consequently, catalytic hydroboration of a variety of imine substrates can be realized with 1 as catalyst and HBpin as terminal reagent. A mechanistic proposal implicating a P–N cooperative mechanism for catalysis that incorporates the various independently verified stoichiometric steps is presented and a comparison to related phosphorus-based systems is offered.

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