TECs produce iNOS after activation.
(A) RT-PCR analysis of iNOS expression. TECs expressed iNOS only after stimulation by a cocktail of cytokines. Optimal expression was observed using IFN-γ, IL-1β, and TNF-α. In these optimal conditions, iNOS was barely detected after 3 hours and clearly observed after 18 hours. (B) Western blot analysis showed the presence of iNOS protein (130 kd) in TECs activated in optimal conditions but not in unstimulated cells. The positive control consisted of stimulated macrophages. (C) Immunoperoxidase analysis of iNOS expression in cultured TECs. TECs expressed significant levels of iNOS after 24 hours of optimal activation; unstimulated cells were unreactive. The scale bar represents 20 μm. (D) NO2 accumulation in TEC culture supernatants was measured by using the Griess reagent. The specificity of the reaction was checked by using L-NAME, an inhibitor of NO2 synthesis.
