MICROVASCULAR OXIDATIVE STRESS PRECEDING LEUKOCYTE ACTIVATION ELICITED BY IN-VIVO NITRIC-OXIDE SUPPRESSION

This study was aimed to determine the mechanism by which endogenous ni tric oxide supression promotes leukocyte adhesion in vivo. The rat mes enteric microcirculation was superfused with NG-nitro-Larginine methyl ester (L-NAME; 100 mu M), and intracellular oxidant formation in seve ral microcirculatory cellular components such as arteriolar and venula r endothelium and mast cells was visually monitored by digital microfl uorography assisted by carboxydichlorofluorescein (CDCF), a hydroperox ide-sensitive fluorogenic probe. Adherent leukocyte density was measur ed simultaneously. L-NAME induced a significant time-dependent increas e in CDCF fluorescence in arteriolar and venular endothelium and mast cells followed by firm adhesion of leukocytes. L-NAME-induced CDCF ele vation showed a different spatial distribution compared with that evok ed by N-formylmethionyl-leucyl-phenylalanine, in which only local venu lar segments with adhering leukocytes exhibited CDCF fluorescence enha ncement. The level of hydroperoxide formation in arterioles and venule s evoked by 60-min L-NAME superfusion was equivalent to that induced b y the superfusion of similar to 880 mu M tert-butyl hydroperoxide for 10 min. Pretreatment with anti-intracellular adhesion molecule-1, anti -P-selectin, or anti-CD18 monoclonal antibody attenuated L-NAME-elicit ed venular leukocyte adhesion without abolishing CDCF fluorescence in situ. Pretreatment with desferioxamine (50 mg/kg iv; 1 h before L-NAME superfusion) significantly diminished the iron-catalyzed hydroperoxid e formation in arterioles and venules, but not in interstitial mast ce lls, as well as subsequent venular leukocyte adhesion. These findings indicate that endogenous nitric oxide may modulate oxidative stress in mast cells, arteriolar and venular microvascular endothelium and ther eby can play a crucial role in leukocyte recruitment in venules.