This study examined the effects of nitrogen dioxide (NO₂) exposure on airway inflammation, blood cells, and antiviral respiratory defense. Twenty-one healthy volunteers were exposed on separate occasions to air and 0.6 and 1.5 ppm NO₂ for 3 h with intermittent moderate exercise. Phlebotomy and bronchoscopy were performed 3.5 h after each exposure, and recovered cells were challenged with respiratory viruses in vitro. Blood studies revealed a 4.1% NO₂ dose-related decrease in hematocrit (P = 0.003). Circulating total lymphocytes (P = 0.024) and T lymphocytes (P = 0.049) decreased with NO₂ exposure. Exposure to NO₂ increased the blood lymphocyte CD4⁺-to-CD8⁺ ratio from 1.74 ± 0.11 to 1.85 ± 0.12 in males but decreased it from 1.88 ± 0.19 to 1.78 ± 0.19 in females (P < 0.001 for gender difference). Polymorphonuclear leukocytes in bronchial lavage increased with NO₂ exposure (P = 0.003). Bronchial epithelial cells obtained after exposure to 1.5 ppm NO₂ released 40% more lactate dehydrogenase after challenge with respiratory syncytial virus than with air exposure (P = 0.024). In healthy subjects, exposures to NO₂ at levels found indoors cause mild airway inflammation, effects on blood cells, and increased susceptibility of airway epithelial cells to injury from respiratory viruses.