Inhibiting pollen reduced nicotinamide adenine dinucleotide phosphate oxidase-induced signal by intrapulmonary administration of antioxidants blocks allergic airway inflammation.

BACKGROUND: Ragweed extract (RWE) contains NADPH oxidases that induce oxidative stress in the airways independent of adaptive immunity (signal 1) and augment antigen (signal 2)-induced allergic airway inflammation. OBJECTIVE: To test whether inhibiting signal 1 by administering antioxidants inhibits allergic airway inflammation in mice. METHODS: The ability of ascorbic acid (AA), N-acetyl cystenine (NAC), and tocopherol to scavenge pollen NADPH oxidase-generated reactive oxygen species (ROS) was measured. These antioxidants were administered locally to inhibit signal 1 in the airways of RWE-sensitized mice. Recruitment of inflammatory cells, mucin production, calcium-activated chloride channel 3, IL-4, and IL-13 mRNA expression was quantified in the lungs. RESULTS: Antioxidants inhibited ROS generation by pollen NADPH oxidases and intracellular ROS generation in cultured epithelial cells. AA in combination with NAC or Tocopherol decreased RWE-induced ROS levels in cultured bronchial epithelial cells. Coadministration of antioxidants with RWE challenge inhibited 4-hydroxynonenal adduct formation, upregulation of Clca3 and IL-4 in lungs, mucin production, recruitment of eosinophils, and total inflammatory cells into the airways. Administration of antioxidants with a second RWE challenge also inhibited airway inflammation. However, administration of AA+NAC 4 or 24 hours after RWE challenge failed to inhibit allergic inflammation. CONCLUSION: Signal 1 plays a proinflammatory role during repeated exposure to pollen extract. We propose that inhibiting signal 1 by increasing antioxidant potential in the airways may be a novel therapeutic strategy to attenuate pollen-induced allergic airway inflammation. CLINICAL IMPLICATIONS: Administration of antioxidants in the airways may constitute a novel therapeutic strategy to prevent pollen induced allergic airway inflammation.

ROS generated by pollen NADPH oxidase provide a signal that augments antigen-induced allergic airway inflammation.

Pollen exposure induces allergic airway inflammation in sensitized subjects. The role of antigenic pollen proteins in the induction of allergic airway inflammation is well characterized, but the contribution of other constituents in pollen grains to this process is unknown. Here we show that pollen grains and their extracts contain intrinsic NADPH oxidases. The pollen NADPH oxidases rapidly increased the levels of ROS in lung epithelium as well as the amount of oxidized glutathione (GSSG) and 4-hydroxynonenal (4-HNE) in airway-lining fluid. These oxidases, as well as products of oxidative stress (such as GSSG and 4-HNE) generated by these enzymes, induced neutrophil recruitment to the airways independent of the adaptive immune response. Removal of pollen NADPH oxidase activity from the challenge material reduced antigen-induced allergic airway inflammation, the number of mucin-containing cells in airway epithelium, and antigen-specific IgE levels in sensitized mice. Furthermore, challenge with Amb a 1, the major antigen in ragweed pollen extract that does not possess NADPH oxidase activity, induced low-grade allergic airway inflammation. Addition of GSSG or 4-HNE to Amb a 1 challenge material boosted allergic airway inflammation. We propose that oxidative stress generated by pollen NADPH oxidases (signal 1) augments allergic airway inflammation induced by pollen antigen (signal 2).