Systemic lupus erythematosus, in both animal models and in humans, is characterized by autoantibody production followed by immune complex deposition in target tissues. Ensuing target organ damage is modulated by reactive intermediates, including reactive nitrogen and oxygen species, through as of now incompletely understood mechanisms. Endothelial nitric oxide synthase is known to impact vascular reactivity; however its impact on reactive intermediate production and inflammatory renal disease is less well defined. In this study, we assessed the impact of endothelial nitric oxide synthase (eNOS) on disease in lupus prone MRL/lpr mice. Mice lacking eNOS developed earlier more severe disease with decreased survival. eNOS deficient mice died sooner and developed significantly more glomerular crescents, necrosis, inflammatory infiltrates and vasculitis, indicating a role for eNOS in modulating these renal lesions. Immune complex deposition was similar between groups, indicating the impact of eNOS is distal to antibody/complement glomerular deposition. Urinary nitric oxide production was decreased in the eNOS deficient mice, while proteinuria was increased. Urinary monocyte chemotactic protein-1 was also increased in the knockout mice. CD4+ T cells from MRL/lpr mice demonstrated mitochondrial hyperpolarization, increased nitric oxide and superoxide production and increased calcium flux compared to B6 control mice. Deficiency of eNOS resulted in decreased nitric oxide and mitochondrial calcium levels but had no effect on mitochondrial hyperpolarization. Renal cortices from MRL/lpr mice that are eNOS deficient demonstrated increased superoxide production, which was blocked by both nitric oxide synthase and NADPH oxidase inhibitors. These studies thus demonstrate a key role for eNOS in modulating renal disease in lupus prone MRL/lpr mice. The impact appears to be mediated by effects on superoxide production in the kidney, impacting downstream mediators such as monocyte chemotactic protein-1. These results suggest that modulation of eNOS may be a novel therapeutic approach to treating lupus nephritis.