Nitric oxide (NO•) is produced by many diverse cell types as a cellular or intracellular signaling molecule, by the activation of nitric oxide synthases (NOSs). All three known NOS isoforms are expressed within the respiratory tract and mediate various airway functional properties such as airway smooth muscle tone, ciliary function, epithelial electrolyte transport, and innate host defense. The respiratory epithelium is a major source of NO•, in which it regulates normal epithelial cell function and signaling as well as signaling pathways involved in airway inflammation. In addition to its normal physiological properties, increased airway NO• production in inflammatory respiratory tract diseases such as asthma may activate additional signaling mechanisms to regulate inflammatory-immune pathways, and epithelial barrier (dys)function or repair. The biological actions of NO• are controlled at various levels, including mechanisms that regulate NOS localization and activation, and variable oxidative metabolism of NO•, resulting in generation of bioactive reactive nitrogen species (RNS). Moreover, in addition to altered production of NO• or RNS, the presence of various target enzymes and/or metabolic regulators of NO•/RNS can be dramatically altered during airway inflammatory conditions, and contribute to alterations in NO•-mediated signaling pathways in disease. This review summarizes current knowledge regarding NO•-mediated epithelial signaling, as well as disease-related changes in airway NOS biology and target enzymes that affect NO•/RNS signaling mechanisms. A detailed understanding of these various changes and their impact on NO• signaling pathways are needed to fully appreciate the contributions of NO•/RNS to airway inflammation and to develop suitable therapeutic approaches based on regulating NO• function.