GSNO attenuates EAE disease by S‐nitrosylation‐mediated modulation of endothelial‐monocyte interactions

Abstract

S-Nitrosoglutathione (GSNO) is an endogenous nitric oxide carrier and recently, has been documented for its anti-inflammatory effects in rat model of cerebral ischemia (Khan et al. (2005) J Cereb Blood Flow Metab 25:177-192). Here, we explored the neuroprotective effects mediated by GSNO in Lewis rat model of EAE and its mechanism of action using in vitro model of monocyte-endothelial cell interaction. Oral administration of GSNO attenuated the clinical disease course in EAE animals by inhibiting the infiltration of vascular immune cells in the CNS that subsequently led to the reduction in the expression of proinflammatory cytokines and consequently limited demyelination. Based on the inhibition in infiltration of immune cells, we hypothesized that GSNO modulated endothelial cell activation that led to reduce cellular infiltration in the CNS. Using an in vitro model, we established that GSNO inhibited monocyte adhesion to the activated endothelial cell, which was mediated by down regulation of endothelial cell adhesion molecules (CAMs). The mechanism by which GSNO modulated CAMs expression appeared to be via S-nitrosylation of p65, which consequently inhibited nuclear factor kappa B (NF-kappaB) activation in endothelial cells. These observations suggest that GSNO exerts its protective effects in EAE by inhibition of cellular infiltration into the CNS by S-nitrosylation of p65, thereby modulating NF-kappaB-CAMs pathway in endothelial cells.

S-Nitrosoglutathione Induces Ciliary Neurotrophic Factor Expression in Astrocytes, Which Has Implications to Protect the Central Nervous System under Pathological Conditions

“…Importantly, PPAR-␥ activators are documented to enhance anti-inflammatory activities in astrocytes and other cell types (64,65). In light of these studies, we conclude that NO signaling in astrocytes is crucial to provide anti-inflammatory and neuroprotective activities in experimental autoimmune encephalomyelitis and traumatic brain injury models (25,26,30,31).…”

“…GSNO has recently been reported to regulate various cellular functions (23,38). In addition, earlier we documented that exogenously administered GSNO restores blood brain barrier integrity and attenuates disease severity in experimental autoimmune encephalomyelitis (25,26) and traumatic brain injury (30,31) models. These studies provide evidence that regardless of the excessive expression of inducible nitric-oxide synthase/NO production by activated CNS glial cells in response to inflammatory insult, the exogenously administered GSNO has a tendency to limit the generation of circulating reactive nitrogen species NO/ONOO in vivo (39).…”

“…GSNO) has emerged as a regulator of various intracellular signaling mechanisms via protein S-nitrosylation (23,24). We recently documented that GSNO treatment attenuates experimental autoimmune encephalomyelitis in a murine model of MS via inhibition of mononuclear cell infiltration into the CNS and restoration of blood brain barrier integrity (25,26). GSNO treatment was also protective in other disease models such as stroke (27,28) and traumatic spine and brain injury (29 -31).…”

STAT3 Regulation by S-Nitrosylation: Implication for Inflammatory Disease

“…Protein S-nitrosylation was detected using the biotin switch method with slight modification as described previously (39). Cells were lysed in 250 mM HEPES, pH 7.7, 1 mM EDTA, 0.1 mM neocuproine, 1% Nonidet P-40, 150 mM NaCl, 1 mM PMSF, 20 lM methyl methanethiosulfonate (MMTS), 80 lM carmustine, protease inhibitor mixture (Sigma), and mixed with an equal volume of 25 mM HEPES, pH 7.7, 0.1 mM EDTA, 10 lM neocuproine, 5% SDS, 20 lM MMTS and incubated at 50°C for 20 min.…”

“…S-nitroso-L-cysteine and S-nitrosoglutathione (GSNO) are the most characterized and abundant endogenous RSNOs synthesized by reaction between NO and cysteine or NO and glutathione (GSH). Recently, these low molecular mass RSNOs have been implicated as a potent anti-inflammatory and antioxidant mediators, vasodilators, and inhibitors of platelet aggregation (6,12,27,32,39). However, the mechanism(s) underlying the role of NO and RSNO in the regulation of microglial proliferation is not understood well at present.…”