In
glomerulonephritis, there is intraglomerular activation of
inducible nitric oxide synthase (iNOS) leading to high output production of
nitric oxide (NO). This can result in supraphysiologic amounts of NO and cause oxidative injury. It is unknown whether mechanisms of cellular defense against NO-mediated injury exist. Induction of the
heme catabolizing
enzyme heme oxygenase-1 (HO-1), which generates
biliverdin,
carbon monoxide (CO), and
iron (Fe), may provide such a mechanism, as CO and Fe are two negative modulators of iNOS activity and expression. This study assessed whether upregulation of HO-1 by a specific inducer,
hemin, negatively modulates iNOS expression and activity in anti-glomerular basement membrane antibody-mediated
glomerulonephritis. Glomerular HO-1 expression in nephritic animals was upregulated by treatment with
hemin (30 micromol/kg body wt). iNOS and HO-1
mRNA expression were assessed by reverse transcription-PCR of glomerular total
RNA from nephritic animals or nephritic animals pretreated with
hemin. iNOS activity in glomeruli was measured by assessing conversion of [14C]
L-arginine to [14C] L-
citrulline. HO-1
protein levels in glomeruli were assessed by Western blot analysis. The effect of
hemin treatment on monocyte/macrophage infiltration was assessed by enumeration of ED-1-positive cells in nephritic glomeruli. iNOS and HO-1 were coinduced in nephritic glomeruli.
Hemin treatment of nephritic animals resulted in upregulation of glomerular HO-1 levels and a two- to threefold reduction in glomerular iNOS
mRNA levels. iNOS activity in glomeruli was significantly reduced in
hemin-treated nephritic animals in which
proteinuria was also attenuated without a change in monocyte/macrophage infiltration.
Hemin (100 to 200 microM) also reduced iNOS
protein levels and
enzyme activity in cultured mesangial cells stimulated with
cytokines. These studies demonstrate that in glomerular immune injury,
hemin treatment upregulates glomerular HO-1 with an attendant downregulation of iNOS expression, and thus points to regulatory interaction between the two systems. The beneficial effect of
hemin treatment on
proteinuria could be linked to downregulation of iNOS.