Sildenafil (Viagra) ameliorates clinical symptoms and neuropathology in a mouse model of multiple sclerosis

Abstract

Cyclic GMP (cGMP)-mediated pathways regulate inflammatory responses in immune and CNS cells. Recently, cGMP phosphodiesterase inhibitors such as sildenafil, commonly used to treat sexual dysfunction in humans including multiple sclerosis (MS) patients, have been reported to be neuroprotective in animal models of stroke, Alzheimer's disease, and focal brain lesion. In this work, we have examined if sildenafil ameliorates myelin oligodendrocyte glycoprotein peptide (MOG₃₅₋₅₅)-induced experimental autoimmune encephalomyelitis (EAE), a mouse model of MS. We show for the first time that treatment with sildenafil after disease onset markedly reduces the clinical signs of EAE by preventing axonal loss and promoting remyelination. Furthermore, sildenafil decreases CD3+ leukocyte infiltration and microglial/macrophage activation in the spinal cord, while increasing forkhead box transcription factor 3-expressing T regulatory cells (Foxp3 Tregs). However, sildenafil treatment did not significantly affect MOG₃₅₋₅₅-stimulated proliferation or release of Th1/Th2 cytokines in splenocytes but decreased ICAM-1 in spinal cord infiltrated cells. The presence of reactive astrocytes forming scar-like structures around infiltrates was enhanced by sildenafil suggesting a possible mechanism for restriction of leukocyte spread into healthy parenchyma. These results highlight novel actions of sildenafil that may contribute to its beneficial effects in EAE and suggest that treatment with this widely used and well-tolerated drug may be a useful therapeutic intervention to ameliorate MS neuropathology.

Mechanisms Involved in the Remyelinating Effect of Sildenafil

“…As above mentioned, sildenafil treatment after cuprizone induced demyelination preserved myelin and axon integrity (Nunes et al 2012, Raposo et al 2014) and previous studies developed in the laboratory where this thesis was developed have shown in significant amelioration of EAE clinical symptoms after sildenafil treatment, suggesting a possible role of sildenafil in remyelination due to an increase in myelin stainings the protection exerted against apoptotic death of oligodendrocytes, the decrease of reactive gliosis and increase in neurotrophic factor BDNF (Pifarre et al 2011, Pifarre et al 2014). …”

“…Studies conducted in the laboratory where this thesis was developed showed that in a model of multiple sclerosis (MS), MOG induced experimental autoimmune encephalomyelitis (EAE), that the administration of the PDE5 inhibitor sildenafil in the acute phase of the disease rapidly ameliorates clinical symptoms, prevents axonal, decreased CD3+-leukocyte infiltration and microglial/macrophage activation in the spinal cord and suggests to restore myelin content (Pifarre et al 2011). Additionally, early administration of sildenafil have shown to delay EAE progression, preserves axons and myelin sheath integrity, prevents the death of oligodendrocytes and enhances expression of BDNF neurotrofic factor in chronic EAE, it was also observed reduced microglia/macrophage reactivity while increased the expression of Ym-1 anti-inflammatory microglia/macrophage phenotype (M2) protein and modulating adaptative immune response (Pifarre et al 2014).…”

“…SC from EAE mice were homogenized in in 50 mM Tris-HCl and 1 mM EDTA, pH 7.4 containing RIPA with antiprotease and antiphosphatases cocktail at a 10% w/v proportion in a glass-teflon Potter-Elvehjeim homogenizer 52 mechanically with 20 strokes at 800 rpm as previously described (Pifarre et al 2011). …”

“…Samples from SC or organotypic cultures homogenates, containing 20 µg and 30 µg of protein respectively, were subjected to Novex NuPAGE® 4-12% Using MOG-induced EAE as a model of MS, the laboratory were this thesis was developed has previously shown that daily treatment with sildenafil at peak disease rapidly ameliorates clinical symptoms and neuropathology (Pifarre et al 2011). An increase in LFB staining of myelin after 8 days of treatment suggested that sildenafil was promoting remyelination.…”

“…an in vivo model that mimics neuropathological and neuroinflammatory features of MS, inducing almost full recovery after 8 days of sildenafil treatment starting at peak of disease, effect associated with an increase in myelin staining and reduced axonal damage (Pifarre et al 2011). In addition, sildenafil treatment after cuprizone induced demyelination in vivo, showed protective effects in cerebellum by preserving myelin and axons ultra-structure and reducing pro-inflammatory cytokines IFN-γ, TNF-α, IL-1β, IL-2 and COX-2 (Nunes et al 2012).…”