Tolerogenic Dendritic Cells Generated by In Vitro Treatment With SAHA Are Not Stable In Vivo.

The aim of this study is to examine whether the histone deacetylase inhibitor, suberoylanilide hydroxamic acid (SAHA), can generate dendritic cells (DCs) with a stable tolerogenic phenotype to counteract autoimmune responses in an animal model of multiple sclerosis. We investigated if the tolerogenic potency of DCs could be increased by continuous treatment during in vitro differentiation toward DCs compared to standard 24-h in vitro treatment of already terminally differentiated DCs. We show that in vitro treatment with SAHA reduces the generation of new CD11c(+) DCs out of mouse bone marrow. SAHA-generated DCs show reduced antigen-presenting function as evidenced by a reduction in myelin endocytosis, a decreased MHC II expression, and a failure to upregulate costimulatory molecules upon LPS challenge. In addition, SAHA-generated DCs display a reduction in proinflammatory cytokines and molecules involved in apoptosis induction, inflammatory migration, and TLR signaling, and they are less immunostimulatory compared to untreated DCs. We demonstrated that the underlying mechanism involves a diminished STAT1 phosphorylation and was independent of STAT6 activation. Although in vitro results were promising, SAHA-generated DCs were not able to alleviate the development of experimental autoimmune encephalomyelitis in mice. In vitro washout experiments demonstrated that the tolerogenic phenotype of SAHA-treated DCs is reversible. Taken together, while SAHA potently boosts tolerogenic properties in DCs during the differentiation process in vitro, SAHA-generated DCs were unable to reduce autoimmunity in vivo. Our results imply that caution needs to be taken when developing DC-based therapies to induce tolerance in the context of autoimmune disease.

Human Wharton's Jelly-Derived Stem Cells Display Immunomodulatory Properties and Transiently Improve Rat Experimental Autoimmune Encephalomyelitis.

Umbilical cord matrix or Wharton's jelly-derived stromal cells (WJ-MSCs) are an easily accessible source of mesenchymal-like stem cells. Recent studies describe a hypoimmunogenic phenotype, multipotent differentiation potential, and trophic support function for WJ-MSCs, with variable clinical benefit in degenerative disease models such as stroke, myocardial infarction, and Parkinson's disease. It remains unclear whether WJ-MSCs have therapeutic value for multiple sclerosis (MS), where autoimmune-mediated demyelination and neurodegeneration need to be halted. In this study, we investigated whether WJ-MSCs possess the required properties to effectively and durably reverse these pathological hallmarks and whether they survive in an inflammatory environment after transplantation. WJ-MSCs displayed a lowly immunogenic phenotype and showed intrinsic expression of neurotrophic factors and a variety of anti-inflammatory molecules. Furthermore, they dose-dependently suppressed proliferation of activated T cells using contact-dependent and paracrine mechanisms. Indoleamine 2,3-dioxygenase 1 was identified as one of the main effector molecules responsible for the observed T-cell suppression. The immune-modulatory phenotype of WJ-MSCs was further enhanced after proinflammatory cytokine treatment in vitro (licensing). In addition to their effect on adaptive immunity, WJ-MSCs interfered with dendritic cell differentiation and maturation, thus directly affecting antigen presentation and therefore T-cell priming. Systemically infused WJ-MSCs potently but transiently ameliorated experimental autoimmune encephalomyelitis (EAE), an animal model for MS, when injected at onset or during chronic disease. This protective effect was paralleled with a reduction in autoantigen-induced T-cell proliferation, confirming their immunomodulatory activity in vivo. Surprisingly, in vitro licensed WJ-MSCs did not ameliorate EAE, indicative of a fast rejection as a result of enhanced immunogenicity. Collectively, we show that WJ-MSCs have trophic support properties and effectively modulate immune cell functioning both in vitro and in the EAE model, suggesting WJ-MSC may hold promise for MS therapy. Future research is needed to optimize survival of stem cells and enhance clinical durability.

Circulating dendritic cells of multiple sclerosis patients are proinflammatory and their frequency is correlated with MS-associated genetic risk factors.

BACKGROUND: The role of the adaptive immune system and more specifically T cells in the pathogenesis of multiple sclerosis (MS) has been studied extensively. Emerging evidence suggests that dendritic cells (DCs), which are innate immune cells, also contribute to MS. OBJECTIVES: This study aimed to characterize circulating DC populations in MS and to investigate the contribution of MS-associated genetic risk factors to DCs. METHODS: Ex vivo analysis of conventional (cDCs) and plasmacytoid DCs (pDCs) was carried out on peripheral blood of MS patients (n = 110) and age- and gender-matched healthy controls (n = 112). RESULTS: Circulating pDCs were significantly decreased in patients with chronic progressive MS compared to relapsing-remitting MS and healthy controls. While no differences in cDCs frequency were found between the different study groups, HLA-DRB1*1501(+) MS patients and patients not carrying the protective IL-7Rα haplotype 2 have reduced frequencies of circulating cDCs and pDCs, respectively. MS-derived DCs showed enhanced IL-12p70 production upon TLR ligation and had an increased expression of the migratory molecules CCR5 and CCR7 as well as an enhanced in vitro chemotaxis. CONCLUSION: DCs in MS are in a pro-inflammatory state, have a migratory phenotype and are affected by genetic risk factors, thereby contributing to pathogenic responses.

Systemic treatment with the inhibitory neurotransmitter γ-aminobutyric acid aggravates experimental autoimmune encephalomyelitis by affecting proinflammatory immune responses.

Transcriptomic and proteomic analyses of multiple sclerosis (MS) lesions indicate alterations in the gamma-aminobutyric acid (GABA) inhibitory system, suggesting its involvement in the disease process. To further elucidate the role of GABA in central nervous system (CNS) inflammation in vivo, the chronic myelin oligodendrocyte glycoprotein (MOG)(35-55) experimental autoimmune encephalomyelitis (EAE) model was used. Daily GABA injections (200mg/kg) from day 3 onwards significantly augmented disease severity, which was associated with increased CNS mRNA expression levels of tumor necrosis factor alpha (TNF-α) and interleukin (IL)-6. GABA-treated mice showed enhanced MOG-dependent proliferation and were skewed towards a T helper 1 phenotype. Moreover, in vitro, the lipopolysaccharide (LPS)-induced increase in interleukin (IL)-6 production by macrophages was enhanced at low GABA concentrations (0.03-0.3mM). In sharp contrast to exogenous GABA administration, endogenous GABA increment by systemic treatment with the GABA-transaminase inhibitor vigabatrin (250mg/kg) had prophylactic as well as therapeutic potential in EAE. Together, these results indicate an immune amplifying role of GABA in neuroinflammatory diseases like MS.

The inhibitory neurotransmitter glycine modulates macrophage activity by activation of neutral amino acid transporters.

Glycine, an important inhibitory neurotransmitter in the mammalian central nervous system (CNS), has been shown to modulate peripheral immune cell responses. In that respect, glycine levels are increased in several neuroinflammatory disorders, such as amyotrophic lateral sclerosis (ALS) and multiple sclerosis (MS). In this study, we show that glycine modulates macrophage effector functions implicated in CNS inflammation and in other, related inflammatory conditions. We demonstrate that glycine does not affect the production of reactive oxygen species but stimulates myelin phagocytosis and the production of the proinflammatory mediators nitric oxide (NO) and tumor necrosis factor (TNF)-alpha by rat macrophages. These effects of glycine are not mediated by the glycine receptor (GlyR) or by glycine transporters (GlyTs), as neither the GlyR antagonist strychnine nor the antagonist of GlyT1 (ALX5407) reverses the observed effects. In contrast, 2-aminoisobutyric acid, a substrate of neutral amino acid transporters (NAATs), inhibits the glycine-mediated enhancement of myelin phagocytosis as well as of NO and TNF-alpha production. In conclusion, our findings demonstrate that glycine modulates macrophage function through activation of NAATs. Glycine may thereby influence immunological processes in inflammatory diseases involving macrophage activation and demyelination, including MS and related conditions associated with altered glycine levels.