Gail Marracci

Cyclophilin D inactivation protects axons in experimental autoimmune encephalomyelitis, an animal model of multiple sclerosis

Multiple sclerosis (MS) is the leading cause of neurological disability in young adults, affecting some two million people worldwide. Traditionally, MS has been considered a chronic, inflammatory disorder of the central white matter in which ensuing demyelination results in physical disability [Frohman EM, Racke MK, Raine CS (2006) N Engl J Med 354:942–955]. More recently, MS has become increasingly viewed as a neurodegenerative disorder in which neuronal loss, axonal injury, and atrophy of the CNS lead to permanent neurological and clinical disability. Although axonal pathology and loss in MS has been recognized for >100 years, very little is known about the underlying molecular mechanisms. Progressive axonal loss in MS may stem from a cascade of ionic imbalances initiated by inflammation, leading to mitochondrial dysfunction and energetic deficits that result in mitochondrial and cellular Ca2+ overload. In a murine disease model, experimental autoimmune encephalomyelitis (EAE) mice lacking cyclophilin D (CyPD), a key regulator of the mitochondrial permeability transition pore (PTP), developed EAE, but unlike WT mice, they partially recovered. Examination of the spinal cords of CyPD-knockout mice revealed a striking preservation of axons, despite a similar extent of inflammation. Furthermore, neurons prepared from CyPD-knockout animals were resistant to reactive oxygen and nitrogen species thought to mediate axonal damage in EAE and MS, and brain mitochondria lacking CyPD sequestered substantially higher levels of Ca2+. Our results directly implicate pathological activation of the mitochondrial PTP in the axonal damage occurring during MS and identify CyPD, as well as the PTP, as a potential target for MS neuroprotective therapies.

Alpha lipoic acid inhibits T cell migration into the spinal cord and suppresses and treats experimental autoimmune encephalomyelitis

Oxidative injury may be important to the pathogenesis of multiple sclerosis (MS). We tested the antioxidant alpha lipoic acid (ALA) in an experimental murine model of MS, experimental autoimmune encephalomyelitis (EAE). ALA was administered to SJL mice 7 days after immunization with proteolipid protein (PLP) 139-151 peptide. Mice that received 5-100 mg/kg/day of ALA had dose-dependent reductions in their 10-Day Cumulative Disease Scores (10-Day CDS) by 23-100%. Minimal inflammation, demyelination and axonal loss occurred in the spinal cords (SC) of ALA-suppressed mice, and there was a marked reduction in CD3+ T cells and CD11b+ monocyte/macrophage cells within the SC. Mice treated with ALA (100 mg/kg/day) commencing on the first day of clinical EAE had a significant reduction in 10-Day CDS. SC of ALA-treated mice had reduced demyelination and axonal loss and a rapid reduction in CD3+ T cells. In vitro, ALA and its reduced form, dihydrolipoic acid, inhibited the activity of matrix metalloproteinase-9 (MMP-9) in a dose-dependent fashion. ALA is highly effective at suppressing and treating EAE and does so by inhibiting T cell trafficking into the SC, perhaps by acting as a matrix metalloproteinase inhibitor.

Lipoic acid in multiple sclerosis: a pilot study

Lipoic acid (LA) is an antioxidant that suppresses and treats an animal model of multiple sclerosis (MS), experimental autoimmune encephalomyelitis. The purpose of this study was to determine the pharmacokinetics (PK), tolerability and effects on matrix metalloproteinase-9 (MMP-9) and soluble intercellular adhesion molecule-1 (sICAM-1) of oral LA in patients with MS. Thirty-seven MS subjects were randomly assigned to one of four groups: placebo, LA 600 mg twice a day, LA 1200 mg once a day and LA 1200 mg twice a day. Subjects took study capsules for 14 days. We found that subjects taking 1200 mg LA had substantially higher peak serum LA levels than those taking 600 mg and that peak levels varied considerably among subjects. We also found a significant negative correlation between peak serum LA levels and mean changes in serum MMP-9 levels (τ=-0.263, P=0.04). There was a significant dose response relationship between LA and mean change in serum sICAM-1 levels (P=0.03). We conclude that oral LA is generally well tolerated and appears capable of reducing serum MMP-9 and sICAM-1 levels. LA may prove useful in treating MS by inhibiting MMP-9 activity and interfering with T-cell migration into the CNS.

Lipoic acid inhibits expression of ICAM-1 and VCAM-1 by CNS endothelial cells and T cell migration into the spinal cord in experimental autoimmune encephalomyelitis

Lipoic acid (LA) suppresses and treats murine experimental autoimmune encephalomyelitis (EAE), which models multiple sclerosis. However, the mechanisms by which LA mediates its effects in EAE are only partially known. In the present study, LA (25, 50 and 100 microg/ml) inhibited upregulation of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in tumor necrosis factor-alpha (TNF-alpha) stimulated cultured brain endothelial cells. Immunohistochemical analysis of spinal cords from SJL mice that had received LA (100 mg/kg/day) following immunization to induce EAE exhibited markedly reduced expression of ICAM-1 and VCAM-1 compared with that of EAE mice receiving saline. Co-localization analysis showed that ICAM-1 and VCAM-1 expression increased over endothelial cells (staining positive for von Willebrand factor, vWF) in EAE and that LA decreased the expression levels to that observed in naïve mice. Spinal cords from mice receiving LA had significantly reduced inflammation (decreased CD4 and CD11b staining) as compared to EAE mice that received saline. Overall, our data suggest that the anti-inflammatory effects of LA in EAE may be partly due to inhibition of ICAM-1 and VCAM-1 expression by central nervous system (CNS) endothelial cells.

α lipoic acid inhibits human T-cell migration: Implications for multiple sclerosis

We have demonstrated previously the ability of the antioxidant α lipoic acid (ALA) to suppress and treat a model of multiple sclerosis (MS), relapsing experimental autoimmune encephalomyelitis (EAE). We describe the effects of ALA and its reduced form, dihydrolipoic acid (DHLA), on the transmigration of human Jurkat T cells across a fibronectin barrier in a transwell system. ALA and DHLA inhibited migration of Jurkat cells in a dose‐dependent fashion by 16–75%. ALA and DHLA reduced matrix metalloproteinase‐9 (MMP‐9) activity by 18–90% in Jurkat cell supernatants. GM6001, a synthetic inhibitor of MMP, reduced Jurkat cell migration, but not as effectively as ALA and DHLA did. Both ALA and DHLA downmodulated the surface expression of the α4β1 integrin (very late activation‐4 antigen; VLA‐4), which binds fibronectin and its endothelial cell ligand vascular cell adhesion molecule‐1 (VCAM‐1). Moreover, ALA, but not DHLA, reduced MMP‐9‐specific mRNA and extracellular MMP‐9 from Jurkat cells and their culture supernatants as detected by relative reverse transcriptase‐polymerase chain reaction (RT‐PCR) and enzyme‐linked immunosorbent assay (ELISA), respectively. ALA and DHLA inhibited Jurkat cell migration and have different mechanisms for inhibiting MMP‐9 activity. These data, coupled with its ability to treat relapsing EAE, suggest that ALA warrants investigation as a therapy for MS.

Omega-3 fatty acid supplementation decreases matrix metalloproteinase-9 production in relapsing-remitting multiple sclerosis

OBJECTIVES The primary objective was to evaluate the effect of omega-3 fatty acids (omega-3 FA) on matrix metalloproteinase-9 (MMP-9) production by immune cells in multiple sclerosis (MS). Quality of life, fatty acid levels, and safety were also evaluated. MATERIALS AND METHODS Ten participants with relapsing-remitting MS (RRMS) received omega-3 FA supplementation (9.6g/day fish oil) in an open-label study. Participants were evaluated at four time points, baseline, after 1 month of omega-3 FA supplementation, after 3 months of omega-3 FA supplementation, and after a 3-month wash out. RESULTS Immune cell secretion of MMP-9 decreased by 58% after 3 months of omega-3 FA supplementation when compared with baseline levels (p<0.01). This effect was coupled with a significant increase in omega-3 FA levels in red blood cell membranes. CONCLUSIONS Omega-3 FA significantly decreased MMP-9 levels in RRMS and may act as an immune-modulator that has potential therapeutic benefit in MS patients.

Lipoic acid decreases inflammation and confers neuroprotection in experimental autoimmune optic neuritis

Lipoic acid (LA) is an antioxidant that is effective in treating experimental autoimmune encephalomyelitis (EAE), a model for multiple sclerosis (MS). C57BL/6 mice with EAE develop experimental autoimmune optic neuritis (EAON), which models acute optic neuritis in humans. Here we determined whether LA is therapeutically effective in EAON. We immunized C57BL/6 mice with MOG 35-55 peptide. Mice received either daily subcutaneous injections of LA (100mg/kg) or saline in early or late suppression paradigms. In the early suppression paradigm, optic nerve cross-sections showed 14.9±3.8% (mean±SEM) damage in mice receiving saline (n=7) and 2.0±0.4% damage in mice given LA (n=7, p=0.001). In the late suppression paradigm, optic nerve sections showed 24.6±3.5% damage in mice treated with saline (n=7) and 8.4±2.5% in mice treated with LA (n=7, p=0.004). Thus a dramatic reduction in axonal injury was seen after LA administration in both experimental paradigms. Compared with saline treated mice with EAON, optic nerves from mice receiving LA had significantly fewer CD4+ and CD11b+ cells in both paradigms. This study provides a rationale for investigating the therapeutic efficacy of LA in acute optic neuritis in humans.

Lipoic acid stimulates cAMP production via the EP2 and EP4 prostanoid receptors and inhibits IFN gamma synthesis and cellular cytotoxicity in NK cells.

The antioxidant lipoic acid (LA) treats and prevents the animal model of multiple sclerosis (MS), experimental autoimmune encephalomyelitis (EAE). In an effort to understand the therapeutic potential of LA in MS, we sought to define the cellular mechanisms that mediate the effects of LA on human natural killer (NK) cells, which are important in innate immunity as the first line of defense against invading pathogens and tumor cells. We discovered that LA stimulates cAMP production in NK cells in a dose-dependent manner. Studies using pharmacological inhibitors and receptor transfection experiments indicate that LA stimulates cAMP production via activation of the EP2 and EP4 prostanoid receptors and adenylyl cyclase. In addition, LA suppressed interleukin (IL)-12/IL-18 induced IFNgamma secretion and cytotoxicity in NK cells. These novel findings suggest that LA may inhibit NK cell function via the cAMP signaling pathway.

Pharmacokinetic study of lipoic acid in multiple sclerosis: Comparing mice and human pharmacokinetic parameters

Lipoic acid is a natural anti-oxidant available as an oral supplement from a number of different manufacturers. Lipoic acid administered subcutaneously is an effective therapy for murine experimental autoimmune encephalomyelitis, a model of multiple sclerosis. The aim of this study was to compare serum lipoic acid levels with oral dosing in patients with multiple sclerosis with serum levels in mice receiving subcutaneous doses of lipoic acid. We performed serum pharmacokinetic studies in patients with multiple sclerosis after a single oral dose of 1200 mg lipoic acid. Patients received one of the three different racemic formulations randomly: tablet (Formulation A) and capsules (Formulations B and C). Mice pharmacokinetic studies were performed with three different subcutaneous doses (20, 50 and 100 mg/kg racemic lipoic acid). The pharmacokinetic parameters included Maximum Serum Concentrations (Cmax in μg/ml) and area under the curve 0—infinity (AUC 0—infinity in μg*min/ml). We found mean Cmax and AUC 0—infinity in patients with multiple sclerosis as follows: group A (N = 7) 3.8 ± 2.6 and 443.1 ± 283.9; group B (N = 8) 9.9 ± 4.5 and 745.2 ± 308.7 and group C (N = 8) 10.3 ± 3.8 and 848.8 ± 360.5, respectively. Mean Cmax and AUC 0—infinity in the mice were: 100 mg/kg lipoic acid: 30.9 ± 2.9 and 998 ± 245; 50 mg/kg lipoic acid: 7.6 ± 1.4 and 223 ± 20; 20 mg/kg lipoic acid: 2.7 ± 0.7 and 119 ± 33. We conclude that patients taking 1200 mg of lipoic acid from two of the three oral formulations achieved serum Cmax and AUC levels comparable to that observed in mice receiving 50 mg/kg subcutaneous dose of lipoic acid, which is a highly therapeutic dose in experimental autoimmune encephalomyelitis. A dose of 1200 mg oral lipoic acid can achieve therapeutic serum levels in patients with multiple sclerosis.

Japanese macaque encephalomyelitis: a spontaneous multiple sclerosis-like disease in a nonhuman primate.

To describe Japanese macaque encephalomyelitis (JME), a spontaneous inflammatory demyelinating disease occurring in the Oregon National Primate Research Centers (ONPRC) colony of Japanese macaques (JMs, Macaca fuscata).