Restoration of axon conduction and motor deficits by therapeutic treatment with glatiramer acetate.

Glatiramer acetate (GA; Copaxone) is an approved drug for the treatment of multiple sclerosis (MS). The underlying multifactorial anti-inflammatory, neuroprotective effect of GA is in the induction of reactive T cells that release immunomodulatory cytokines and neurotrophic factors at the injury site. These GA-induced cytokines and growth factors may have a direct effect on axon function. Building on previous findings that suggest a neuroprotective effect of GA, we assessed the therapeutic effects of GA on brain and spinal cord pathology and functional correlates using the chronic experimental autoimmune encephalomyelitis (EAE) mouse model of MS. Therapeutic regimens were utilized based on promising prophylactic efficacy. More specifically, C57BL/6 mice were treated with 2 mg/mouse/day GA for 8 days beginning at various time points after EAE post-induction day 15, yielding a thorough, clinically relevant assessment of GA efficacy within the context of severe progressive disease. Therapeutic treatment with GA significantly decreased clinical scores and improved rotorod motor performance in EAE mice. These functional improvements were supported by an increase in myelinated axons and fewer amyloid precursor protein-positive axons in the spinal cords of GA-treated EAE mice. Furthermore, therapeutic GA decreased microglia/macrophage and T cell infiltrates and increased oligodendrocyte numbers in both the spinal cord and corpus callosum of EAE mice. Finally, GA improved callosal axon conduction and nodal protein organization in EAE. Our results demonstrate that therapeutic GA treatment has significant beneficial effects in a chronic mouse model of MS, in which its positive effects on both myelinated and non-myelinated axons results in improved axon function

Therapeutic laquinimod treatment decreases inflammation, initiates axon remyelination, and improves motor deficit in a mouse model of multiple sclerosis.

BackgroundTherapeutic strategies that induce effective neuroprotection and enhance intrinsic repair mechanisms are central goals for future treatment of multiple sclerosis (MS), as well as other diseases. Laquinimod (LQ) is an orally administered, central nervous system (CNS)-active immunomodulator with demonstrated efficacy in MS clinical trials and a favorable safety and tolerability profile.AimsWe aimed to explore the pathological, functional, and behavioral consequences of prophylactic and therapeutic (after presentation of peak clinical disease) LQ treatment in the chronic experimental autoimmune encephalomyelitis (EAE) mouse model of MS.Materials and methodsActive EAE-induced 8-week-old C57BL/6 mice were treated with 5 or 25 mg/kg/day LQ via oral gavage beginning on EAE post-immunization day 0, 8, or 21. Clinical scores and rotorod motor performance were assessed throughout the disease course. Immune analysis of autoantigen-stimulated splenocytes, electrophysiological conduction of callosal axons, and immunohistochemistry of white matter-rich corpus callosum and spinal cord were performed.ResultsProphylactic and therapeutic treatment with LQ significantly decreased mean clinical disease scores, inhibited Th1 cytokine production, and decreased the CNS inflammatory response. LQ-induced improvement in axon myelination and integrity during EAE was functional, as evidenced by significant recovery of callosal axon conduction and axon refractoriness and pronounced improvement in rotorod motor performance. These improvements correlate with LQ-induced attenuation of EAE-induced demyelination and axon damage, and improved myelinated axon numbers.DiscussionEven when initiated at peak disease, LQ treatment has beneficial effects within the chronic EAE mouse model. In addition to its immunomodulatory effects, the positive effects of LQ treatment on oligodendrocyte numbers and myelin density are indicative of significant, functional neuroprotective and neurorestorative effects.ConclusionsOur results support a potential neuroprotective, in addition to immunomodulatory, effect of LQ treatment in inhibiting ongoing MS/EAE disease progression

Laquinimod dampens hyperactive cytokine production in Huntington's disease patient myeloid cells.

Huntington's disease (HD) is a neurodegenerative condition characterized by pathology in the brain and peripheral tissues. Hyperactivity of the innate immune system, due in part to NFκB pathway dysregulation, is an early and active component of HD. Evidence suggests targeting immune disruption may slow disease progression. Laquinimod is an orally active immunomodulator that down-regulates proinflammatory cytokine production in peripheral blood mononuclear cells, and in the brain down-regulates astrocytic and microglial activation by modulating NFκB signalling. Laquinimod had beneficial effects on inflammation, brain atrophy and disease progression in multiple sclerosis (MS) in two phase III clinical trials. This study investigated the effects of laquinimod on hyperactive proinflammatory cytokine release and NFκB signalling in HD patient myeloid cell cultures. Monocytes from manifest (manHD) and pre-manifest (preHD) HD gene carriers and healthy volunteers (HV) were treated with laquinimod and stimulated with lipopolysaccharide. After 24 h pre-treatment with 5 μM laquinimod, manHD monocytes released lower levels of IL-1β, IL-5, IL-8, IL-10, IL-13 and TNFα in response to stimulation. PreHD monocytes released lower levels of IL-8, IL-10 and IL-13, with no reduction observed in HV monocytes. The effects of laquinimod on dysfunctional NFκB signalling in HD was assessed by inhibitor of kappa B (IκB) degradation kinetics, nuclear translocation of NFκB and interactions between IκB kinase (IKK) and HTT, in HD myeloid cells. No differences were observed between laquinimod-treated and untreated conditions. These results provide evidence that laquinimod dampens hyper-reactive cytokine release from manHD and preHD monocytes, with a much reduced effect on HV monocytes. Evidence suggests targeting CNS and peripheral immune disruption may slow Huntington's disease (HD) neurodegenerative processes. The effects of laquinimod, an orally active immunomodulator, on hyperactive cytokine release and dysfunctional NFκB signalling in stimulated myeloid cell cultures from pre-manifest and manifest HD gene carriers and healthy volunteers were investigated. Laquinimod dampened cytokine release but did not impact NFκB signalling. Read the Editorial Highlight for this article on page 670

Arachidyl Amido Cholanoic Acid Improves Liver Glucose and Lipid Homeostasis in Nonalcoholic Steatohepatitis Via AMPK and mTOR Regulation

BACKGROUND Arachidyl amido cholanoic acid (Aramchol) is a potent downregulator of hepatic stearoyl-CoA desaturase 1 (SCD1) protein expression that reduces liver triglycerides and fibrosis in animal models of steatohepatitis. In a phase IIb clinical trial in patients with nonalcoholic steatohepatitis (NASH), 52 wk of treatment with Aramchol reduced blood levels of glycated hemoglobin A1c, an indicator of glycemic control. AIM To assess lipid and glucose metabolism in mouse hepatocytes and in a NASH mouse model [induced with a 0.1% methionine and choline deficient diet (0.1MCD)] after treatment with Aramchol. METHODS Isolated primary mouse hepatocytes were incubated with 20 mu mol/L Aramchol or vehicle for 48 h. Subsequently, analyses were performed including Western blot, proteomics by mass spectrometry, and fluxomic analysis with(13)C-uniformly labeled glucose. For thein vivopart of the study, male C57BL/6J mice were randomly fed a control or 0.1MCD for 4 wk and received 1 or 5 mg/kg/d Aramchol or vehicle by intragastric gavage for the last 2 wk. Liver metabolomics were assessed using ultra-high-performance liquid chromatography-time of flight-MS for the determination of glucose metabolism-related metabolites. RESULTS Combination of proteomics and Western blot analyses showed increased AMPK activity while the activity of nutrient sensor mTORC1 was decreased by Aramchol in hepatocytes. This translated into changes in the content of their downstream targets including proteins involved in fatty acid (FA) synthesis and oxidation [P-ACC alpha/beta(S79), SCD1, CPT1A/B, HADHA, and HADHB], oxidative phosphorylation (NDUFA9, NDUFB11, NDUFS1, NDUFV1, ETFDH, and UQCRC2), tricarboxylic acid (TCA) cycle (MDH2, SUCLA2, and SUCLG2), and ribosome (P-p70S6K[T389] and P-S6[S235/S236]). Flux experiments with(13)C-uniformely labeled glucose showed that TCA cycle cataplerosis was reduced by Aramchol in hepatocytes, as indicated by the increase in the number of rounds that malate remained in the TCA cycle. Finally, liver metabolomic analysis showed that glucose homeostasis was improved by Aramchol in 0.1MCD fed mice in a dose-dependent manner, showing normalization of glucose, G6P, F6P, UDP-glucose, and Rbl5P/Xyl5P. CONCLUSION Aramchol exerts its effect on glucose and lipid metabolism in NASH through activation of AMPK and inhibition of mTORC1, which in turn activate FA beta-oxidation and oxidative phosphorylation.Supported by the National Institutes of Health Grant, No. R01CA172086; Plan Nacional of I+D, No. SAF2017-88041-R; Ministerio de Economia y Competitividad de Espana, No. SAF2017-87301-R; Asociacion Espanola contra el Cancer, No. AECC17/302; Ayudas Fundacion BBVA a equipos de Investigacion Cientifica 2018; Fondo Europeo de Desarrollo Regional, Ministerio de Economia y Competitividad de Espana, No. PGC2018-099857-BI00; Basque Government Grants, No. IT1264-19; Ministerio de Economia y Competitividad de Espana for the Severo Ochoa Excellence Accreditation, No. SEV2016-0644. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Metabolic subtypes of patients with NAFLD exhibit distinctive cardiovascular risk profiles

Background and Aims We previously identified subsets of patients with NAFLD with different metabolic phenotypes. Here we align metabolomic signatures with cardiovascular disease (CVD) and genetic risk factors. Approach and Results We analyzed serum metabolome from 1154 individuals with biopsy-proven NAFLD, and from four mouse models of NAFLD with impaired VLDL-triglyceride (TG) secretion, and one with normal VLDL-TG secretion. We identified three metabolic subtypes: A (47%), B (27%), and C (26%). Subtype A phenocopied the metabolome of mice with impaired VLDL-TG secretion; subtype C phenocopied the metabolome of mice with normal VLDL-TG; and subtype B showed an intermediate signature. The percent of patients with NASH and fibrosis was comparable among subtypes, although subtypes B and C exhibited higher liver enzymes. Serum VLDL-TG levels and secretion rate were lower among subtype A compared with subtypes B and C. Subtype A VLDL-TG and VLDL-apolipoprotein B concentrations were independent of steatosis, whereas subtypes B and C showed an association with these parameters. Serum TG, cholesterol, VLDL, small dense LDL5,6, and remnant lipoprotein cholesterol were lower among subtype A compared with subtypes B and C. The 10-year high risk of CVD, measured with the Framingham risk score, and the frequency of patatin-like phospholipase domain-containing protein 3 NAFLD risk allele were lower in subtype A. Conclusions Metabolomic signatures identify three NAFLD subgroups, independent of histological disease severity. These signatures align with known CVD and genetic risk factors, with subtype A exhibiting a lower CVD risk profile. This may account for the variation in hepatic versus cardiovascular outcomes, offering clinically relevant risk stratification.National Institutes of Health (R01DK123763, R01DK119437, HL151328, P30DK52574, P30DK56341, and UL1TR002345); Ministerio de Economía y Competitividad de España (SAF2017-88041-R); Ministerio de Economía y Competitividad de España for the Severo Ochoa Excellence Accreditation (SEV-2016-0644); CIBERehd (Biomedical Research Center in Hepatic and Digestive Diseases) and Netherlands Organization for Applied Scientific Research Program (PMC13 and PMC15); Spanish Carlos III Health Institute (PI15/01132 and PI18/01075); Miguel Servet Program (CON14/00129 and CPII19/00008); Fondo Europeo de Desarrollo Regional, CIBERehd, Department of Industry of the Basque Country (Elkartek: KK-2020/00008); La Caixa Scientific Foundation (HR17-00601); Liver Investigation: Testing Marker Utility in Steatohepatitis consortium funded by the Innovative Medicines Initiative Program of the European Union (777377), which receives support from the European Union’s Horizon 2020 research and innovation programme and EFPIA; Newcastle NIHR Biomedical Research Center; Czech Ministry of Health (RVO-VFN64165/2020); Fondo Nacional De Ciencia y Tecnología de Chile (1191145); and the Comisión Nacional de Investigación, Ciencia y Tecnología (AFB170005, CARE Chile UC); Agencia Nacional de Investigación y Desarrollo (ANID ACE 210009); European Union's Horizon 2020 Research and Innovation Program (825510)

Therapeutic laquinimod treatment decreases inflammation, initiates axon remyelination, and improves motor deficit in a mouse model of multiple sclerosis.

BackgroundTherapeutic strategies that induce effective neuroprotection and enhance intrinsic repair mechanisms are central goals for future treatment of multiple sclerosis (MS), as well as other diseases. Laquinimod (LQ) is an orally administered, central nervous system (CNS)-active immunomodulator with demonstrated efficacy in MS clinical trials and a favorable safety and tolerability profile.AimsWe aimed to explore the pathological, functional, and behavioral consequences of prophylactic and therapeutic (after presentation of peak clinical disease) LQ treatment in the chronic experimental autoimmune encephalomyelitis (EAE) mouse model of MS.Materials and methodsActive EAE-induced 8-week-old C57BL/6 mice were treated with 5 or 25 mg/kg/day LQ via oral gavage beginning on EAE post-immunization day 0, 8, or 21. Clinical scores and rotorod motor performance were assessed throughout the disease course. Immune analysis of autoantigen-stimulated splenocytes, electrophysiological conduction of callosal axons, and immunohistochemistry of white matter-rich corpus callosum and spinal cord were performed.ResultsProphylactic and therapeutic treatment with LQ significantly decreased mean clinical disease scores, inhibited Th1 cytokine production, and decreased the CNS inflammatory response. LQ-induced improvement in axon myelination and integrity during EAE was functional, as evidenced by significant recovery of callosal axon conduction and axon refractoriness and pronounced improvement in rotorod motor performance. These improvements correlate with LQ-induced attenuation of EAE-induced demyelination and axon damage, and improved myelinated axon numbers.DiscussionEven when initiated at peak disease, LQ treatment has beneficial effects within the chronic EAE mouse model. In addition to its immunomodulatory effects, the positive effects of LQ treatment on oligodendrocyte numbers and myelin density are indicative of significant, functional neuroprotective and neurorestorative effects.ConclusionsOur results support a potential neuroprotective, in addition to immunomodulatory, effect of LQ treatment in inhibiting ongoing MS/EAE disease progression

Therapeutic laquinimod treatment decreases inflammation, initiates axon remyelination, and improves motor deficit in a mouse model of multiple sclerosis

BACKGROUND: Therapeutic strategies that induce effective neuroprotection and enhance intrinsic repair mechanisms are central goals for future treatment of multiple sclerosis (MS), as well as other diseases. Laquinimod (LQ) is an orally administered, central nervous system (CNS)-active immunomodulator with demonstrated efficacy in MS clinical trials and a favorable safety and tolerability profile. AIMS: We aimed to explore the pathological, functional, and behavioral consequences of prophylactic and therapeutic (after presentation of peak clinical disease) LQ treatment in the chronic experimental autoimmune encephalomyelitis (EAE) mouse model of MS. MATERIALS AND METHODS: Active EAE-induced 8-week-old C57BL/6 mice were treated with 5 or 25 mg/kg/day LQ via oral gavage beginning on EAE post-immunization day 0, 8, or 21. Clinical scores and rotorod motor performance were assessed throughout the disease course. Immune analysis of autoantigen-stimulated splenocytes, electrophysiological conduction of callosal axons, and immunohistochemistry of white matter-rich corpus callosum and spinal cord were performed. RESULTS: Prophylactic and therapeutic treatment with LQ significantly decreased mean clinical disease scores, inhibited Th1 cytokine production, and decreased the CNS inflammatory response. LQ-induced improvement in axon myelination and integrity during EAE was functional, as evidenced by significant recovery of callosal axon conduction and axon refractoriness and pronounced improvement in rotorod motor performance. These improvements correlate with LQ-induced attenuation of EAE-induced demyelination and axon damage, and improved myelinated axon numbers. DISCUSSION: Even when initiated at peak disease, LQ treatment has beneficial effects within the chronic EAE mouse model. In addition to its immunomodulatory effects, the positive effects of LQ treatment on oligodendrocyte numbers and myelin density are indicative of significant, functional neuroprotective and neurorestorative effects. CONCLUSIONS: Our results support a potential neuroprotective, in addition to immunomodulatory, effect of LQ treatment in inhibiting ongoing MS/EAE disease progression