Immunohistochemical demonstration of a paracrine role of nitric oxide in bronchial function

We addressed the controversial role of nitric oxide (NO) in bronchial function by an immunohistochemical study of the localization of NO synthase (NOS) and its effector protein, soluble guanylate cyclase, in rat bronchus. For this study, a monoclonal antibody to the bovine constitutive neuronal NOS was developed and characterized. In Western blot analysis, this monoclonal antibody (anti-NOS antibody) reacted with bovine cerebellum NOS (150 kDa) as well as with structurally different NOSs from cultured bovine aortic endothelial cells (130 kDa) and cultured RAW 264.7 macrophages (130 kDa). The reactivity of anti-NOS antibody was confirmed by immunohistochemical staining of rat cerebellum, arterial endothelial cells, and cultured stimulated macrophages. When the distribution of NOS in rat airway was characterized, the anti-NOS antibody showed immunoreactivity within respiratory epithelium but not in the bronchial smooth muscle. The NADPH-diaphorase staining correlated with the immunostaining. In contrast, a monoclonal antibody to the rat lung-soluble guanylate cyclase immunostained respiratory smooth muscle but not epithelium. This study suggests a paracrine role for NO in bronchial function analogous to the function of the NOS-soluble guanylate cyclase pathway in blood vessels.