Immunomodulation and Neuroprotection in Multiple Sclerosis

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Multiple sclerosis (MS), a disabling autoimmune disease affecting almost 2.5 million people around the world, affects individuals in their most productive years and is prevalent among US Veterans. MS is caused by abnormal activation of myelin-specific autoreactive lymphocytes and their CNS infiltration across the blood- brain barrier (BBB). Present day FDA approved MS drugs, targeting immune system, have limited efficacies as disease progression continues. In an effort to improve MS therapy, the goal of this proposal is to evaluate the feasibility of nitric oxide synthase (NOS) and nitric oxide (NO) metabolites as potential therapeutic target(s) for immune as well as BBB pathogenesis, two critical disease mechanisms of MS. Recent studies from our laboratory and others have documented the roles of NOS and NO metabolites (S- nitrosoglutathione; GSNO vs. peroxynitrite; ONOO¯) in physiological vs. pathological regulation of experimental autoimmune encephalomyelitis (EAE; a model for MS). Asymmetric dimethylarginine (ADMA), a cellular metabolite of L-arginine, shifts the NOS activity from NO production to superoxide (O2·¯) production, thus leading to increased toxic ONOO¯ formation. Therefore, elevation of ADMA levels in EAE and MS could potentially shift the NOS activity for increased production of ONOO¯ and thus cause related severe pathologies. The pathological importance of ADMA in MS was recognized first by the reported elevation of blood ADMA in MS patients. Next, our studies document the elevated blood ADMA in EAE. We further observed that treatment of EAE animals with exogenous ADMA aggravates the disease severity by increasing TH1/TH17 mediated pro- inflammatory immune responses. In addition, ADMA induced ONOO¯ synthesis in the brain microvascular endothelial cells and caused BBB dysfunction/disruption for CNS infiltration of immunocytes even in the absence of pertussis toxin. These findings, for the first time, document the participation of ADMA-mediated mechanisms in immune pathogenesis as well as BBB dysfunction in EAE. Our laboratory previously reported that NO and its physiological carrier molecule GSNO play critical roles in autoimmune responses as well as in maintenance of endothelial barrier integrity in EAE. These observations lead us to hypothesize that elevation of ADMA as a result of its defective metabolism, during the course of EAE, induces NOS dysfunction leading to inhibition of NO/GSNO-mediated anti-inflammatory and vaso-protective activity, while inducing the O2·¯/ONOO¯- mediated pro-inflammatory and vaso-disruptive activity in EAE. Therefore, ADMA-mediated mechanisms are novel and a potential therapeutic target for immune and BBB pathogenesis of MS/EAE. Based on this hypothesis, the proposed studies are to investigate the temporal relationship between dysfunctional ADMA and NO (ONOO¯ vs. GSNO) metabolism and immunological, vascular, neurological, and clinical pathologies of EAE (AIM 1), mechanism underlying impaired ADMA catabolism by DDAH-1/2 in EAE (AIM 2) and to evaluate potential therapeutics targeting ADMA and NO metabolism in EAE (AIM 3). The proposed studies are based on our original findings of pathological significance of ADMA in NOS/NO- mediated immune and vascular disease mechanisms of EAE. Results from these studies are expected to provide novel insights into disease mechanisms and to identify novel potential targets and therapeutics for MS. Translational potential of this study is high as GSNO mediated mechanisms target immune modulation (TH1/TH17 < TH2/Treg), rather than global immune suppression, as well as BBB pathologies of EAE.