EXPRESSION OF INOS, ENOS, AND PEROXYNITRITE-MODIFIED PROTEINS IN EXPERIMENTAL ANTIMYELOPEROXIDASE ASSOCIATED CRESCENTIC GLOMERULONEPHRITIS

Nitric oxide radicals are recognized as important mediators in various physiological and pathophysiological processes. During inflammation, increased amounts of nitric oxide (NO) are produced, but it is unclear whether NO radicals are either protective or harmful. To obtain more insight into the role of NO in glomerular inflammation, we studied the temporal expression of endothelial NO synthase (eNOS) and inducible N OS (iNOS) in conjunction with platelet aggregation, inflammatory cell influx, superoxide anion production cells, and nitrotyrosine formation in an experimental model of anti-myeloperoxidase (MPO) associated nec rotizing crescentic glomerulonephritis (NCGN). Brown Norway rats were immunized with MPO in complete Freund's adjuvant (CFA) or CFA alone. A fter two weeks, the left kidney was perfused with a neutrophil lysosom al extract and H2O2. Rats were sacrificed at 24 hours, four days, and 10 days after perfusion. Kidney sections were stained by immunohistoch emistry for eNOS, iNOS, platelets, nitrotyrosines, polymorphonuclear c ells (PMN), monocytes, and T-cells. Superoxide anion producing cells w ere identified by enzyme cytochemistry using diaminobenzidine. Strong staining for eNOS was found in glomerular capillaries and interstitial tubular capillaries and larger vessels from non-perfused kidneys. At 24 hours after perfusion, glomerular and interstitial eNOS staining wa s greatly reduced, which was associated with massive platelet aggregat ion. At later time points, eNOS expression was absent in severely dama ged glomeruli. Inducible NOS expression was found at all time points i n infiltrating inflammatory cells, which by double labeling studies we re identified as PMNs and monocytes. The peak in iNOS expression was o bserved at four days after perfusion but declined thereafter. Superoxi de anion and nitrotyrosine generating cells were also found at all tim e points, bur were most abundantly present at four days after perfusio n, coinciding with the peak in iNOS expression. Double labeling experi ments revealed that most nitrotyrosine generating cells also produced superoxide anions and expressed iNOS. In conclusion, these studies sug gest that during the course of anti-MPO associated NCGN, loss of NO pr oduction by eNOS in conjunction with NO radical production by iNOS con tribute to tissue injury. This is compatible with a protective role fo r eNOS contrasting with the possibly harmful effects of iNOS in anti-M PO associated NCGN.