Alpha-lipoic acid (alpha-LA) is a neuroprotective metabolic antioxidant that has been shown to cross the blood brain barrier. We tested whether alpha-LA is capable to prevent MOG35-55-induced experimental autoimmune encephalomyelitis (EAE), an established model of multiple sclerosis (MS). Daily oral administration of alpha-LA, starting at the time of immunization, significantly prevented EAE progression as compared to control mice. This was associated with a reduction of CNS infiltrating T cells and macrophages as well as decreased demyelination. We then tested alpha-LA in a therapeutic protocol aimed at suppressing EAE after its onset. Intraperitoneal (i.p.), but not oral, administration of alpha-LA significantly prevented disease progression when compared to vehicle-treated controls. Similarly, we observed significant reduction of demyelination and inflammatory infiltration. This clinical effect was not due to an impairment of MOG35-55 recognition by encephalitogenic T cells. In contrast, MOG-specific T cells showed a decreased production of IFNgamma and IL-4, suggesting an immunosuppressive activity on both Th1 and Th2 cytokines. In addition, alpha-LA inhibited the proteolytic activity of MMP2 and MMP9 only at very high doses. Our data indicate that alpha-LA can effectively interfere with the autoimmune reaction associated with EAE through mechanisms other than its antioxidant activity and supports further studies on the use of alpha-LA as a potential therapy for MS.

To perform their distinct effector functions, pathogen-specific T cells have to migrate to target tissue where they recognize antigens and produce cytokines that elicit appropriate types of protective responses. Similarly, migration of pathogenic self-reactive T cells to target organs is an essential step required for tissue-specific autoimmunity. In this article, we review data from our laboratory as well as other laboratories that have established that effector function and migratory capacity are coordinately regulated in different T-cell subsets. We then describe how pathogenic T cells can enter into intact or inflamed central nervous system (CNS) to cause experimental autoimmune encephalomyelitis or multiple sclerosis. In particular, we elaborate on the role of CCR6/CCL20 axis in migration through the choroid plexus and the involvement of this pathway in immune surveillance of and autoimmunity in the CNS.

Despite some advances in the understanding of amyotrophic lateral sclerosis (ALS) pathogenesis, significant achievements in treating this disease are still lacking. Mesenchymal stromal (stem) cells (MSCs) have been shown to be effective in several models of neurological disease. To determine the effects of the intravenous injection of MSCs in an ALS mouse model during the symptomatic stage of disease, MSCs (1 × 10⁶) were intravenously injected in mice expressing human superoxide dismutase 1 (SOD1) carrying the G93A mutation (SOD1/G93A) presenting with experimental ALS. Survival, motor abilities, histology, oxidative stress markers and [³H]D-aspartate release in the spinal cord were investigated. MSC injection in SOD1/G93A mice improved survival and motor functions compared with saline-injected controls. Injected MSCs scantly home to the central nervous system and poorly engraft. We observed a reduced accumulation of ubiquitin agglomerates and of activated astrocytes and microglia in the spinal cord of MSC-treated SOD1/G93A mice, with no changes in the number of choline acetyltransferase- and glutamate transporter type 1-positive cells. MSC administration turned around the upregulation of metallothionein mRNA expression and of the activity of the antioxidant enzyme glutathione S-transferase, both associated with disease progression. Last, we observed that MSCs reverted both spontaneous and stimulus-evoked neuronal release of [³H]D-aspartate, a marker of endogenous glutamate, which is upregulated in SOD1/G93A mice. These findings suggest that intravenous administration of MSCs significantly improves the clinical outcome and pathological scores of mutant SOD1/G93A mice, thus providing the rationale for their exploitation for the treatment of ALS.

Mesenchymal stem cells (MSC) display a remarkable ability to modulate the immune response and protect the central nervous system mainly through the release of soluble factors in a paracrine fashion, affecting the functional behavior of cells in the tissues. Here we investigated the effect of the interaction between MSC and microglia in vitro, and we dissected the molecular and cellular mechanisms of this crosstalk. We demonstrated that MSC impair microglia activation by inflammatory cues through the inhibition of the expression and release of inflammatory molecules and stress-associated proteins. We showed that MSC significantly increase microglial expression and release of molecules associated with a neuroprotective phenotype such as CX3CR1, nuclear receptor 4 family, CD200 receptor, and insulin growth factor 1. Interestingly, MSC can enhance functional changes on microglia as depicted by the increase of intracellular calcium concentration and phagocytic activity. This last event is associated with an increased expression of triggering receptor expressed on myeloid cells-2, an innate immune receptor involved in phagocytosis in the absence of inflammation. The observed effects on CX3CR1-expressing microglia are due to the release of CX3CL1 by MSC, driven by inflammatory signals, as demonstrated by the reversal of the observed results when CX3CL1 expression was silenced in MSC or its release was blocked. Finally, we showed that exogenous CX3CL1 induce phenotypic and functional changes of microglia similar to those induced by MSC. These findings demonstrate that MSC instruct, through the release of CX3CL1, microglia responsiveness to proinflammatory signals by modulating constitutive "calming" receptors, typically expressed by "steady-state microglia" thus switching microglia from a detrimental phenotype to a neuroprotective one.

Dimethyl fumarate (DMF), recently approved as an oral immunomodulatory treatment for relapsing-remitting multiple sclerosis (MS), metabolizes to monomethyl fumarate (MMF) which crosses the blood-brain barrier and has demonstrated neuroprotective effects in experimental studies. We postulated that MMF exerts neuroprotective effects through modulation of microglia activation, a critical component of the neuroinflammatory cascade that occurs in neurodegenerative diseases such as MS. To ascertain our hypothesis and define the mechanistic pathways involved in the modulating effect of fumarates, we used real-time PCR and biochemical assays to assess changes in the molecular and functional phenotype of microglia, quantitative Western blotting to monitor activation of postulated pathway components, and ex vivo whole-cell patch clamp recording of excitatory post-synaptic currents in corticostriatal slices from mice with experimental autoimmune encephalomyelitis (EAE), a model for MS, to study synaptic transmission. We show that exposure to MMF switches the molecular and functional phenotype of activated microglia from classically activated, pro-inflammatory type to alternatively activated, neuroprotective one, through activation of the hydroxycarboxylic acid receptor 2 (HCAR2). We validate a downstream pathway mediated through the AMPK-Sirt1 axis resulting in deacetylation, and thereby inhibition, of NF-κB and, consequently, of secretion of pro-inflammatory molecules. We demonstrate through ex vivo monitoring of spontaneous glutamate-mediated excitatory post-synaptic currents of single neurons in corticostriatal slices from EAE mice that the neuroprotective effect of DMF was exerted on neurons at pre-synaptic terminals by modulating glutamate release. By exposing control slices to untreated and MMF-treated activated microglia, we confirm the modulating effect of MMF on microglia function and, thereby, its indirect neuroprotective effect at post-synaptic level. These findings, whereby DMF-induced activation of a new HCAR2-dependent pathway on microglia leads to the modulation of neuroinflammation and restores synaptic alterations occurring in EAE, represent a possible novel mechanism of action for DMF in MS.

Microglia cells, the resident innate immune cells in the brain, are highly active, extending and retracting highly motile processes through which they continuously survey their microenvironment for 'danger signals' and interact dynamically with surrounding cells. Upon sensing changes in their central nervous system microenvironment, microglia become activated, undergoing morphological and functional changes. Microglia activation is not an 'all-or-none' process, but rather a continuum depending on encountered stimuli, which is expressed through a spectrum of molecular and functional phenotypes ranging from so-called 'classically activated', with a highly pro-inflammatory profile, to 'alternatively activated' associated with a beneficial, less inflammatory, neuroprotective profile. Microglia activation has been demonstrated in most neurological diseases of diverse aetiology and has been implicated as a contributor to neurodegeneration. The possibility to promote microglia's neuroprotective phenotype has therefore become a therapeutic goal. We have focused our discussion on the role of microglia in multiple sclerosis, a prototype of inflammatory, demyelinating, neurodegenerative disease, and on the effect of currently approved or on-trial anti-inflammatory therapeutic strategies that might mediate neuroprotection at least in part through their effect on microglia by modifying their behaviour via a switch of their functional phenotype from a detrimental to a protective one. In addition to pharmaceutical approaches, such as treatment with glatiramer acetate, interferon-β, fingolimod or dimethyl fumarate, we address the alternative therapeutic approach of treatment with mesenchymal stem cells and their potential role in neuroprotection through their 'calming' effect on microglia.

The unmet need for therapies capable of repairing the central nervous system (CNS) damage occurring in many diseases including multiple sclerosis (MS) has sparked the interest of the neurological community for stem cell-based therapies. An exhaustive amount of preclinical data has shown that the intravenous administration of mesenchymal stem cells (MSC), a subset of progenitor cells isolated from many mesodermal tissues, effectively ameliorates experimental autoimmune encephalomyelitis (EAE), a model of MS, through the release of anti-inflammatory and neuroprotective molecules. Based on these results, several small pilot clinical trials in subjects with advanced MS have demonstrated that MSC administration is safe and provided an early signal of clinical effectiveness. The current aim of clinicians and scientists interested in the development of MSC-based strategies for the treatment of MS is to have the ultimate demonstration in large clinical trials that MSC can inhibit CNS inflammation and foster tissue repair as realized clinically, with functional recovery, or visualized by magnetic resonance imaging (MRI).

BACKGROUND

The hematopoietic stem cells (HSCs) niche of the bone marrow is comprised of HSCs, osteoblasts, endothelial cells and a stromal component of non-hematopoietic multipotent cells of mesenchymal origin named "mesenchymal stem cells" (MSCs).

RESULTS

Here we studied the global transcriptional profile of murine MSCs with immuno-therapeutic potential and compared it with that of 486 publicly available microarray datasets from 12 other mouse tissues or cell types. Principal component analysis and hierarchical clustering identified a unique pattern of gene expression capable of distinctively classifying MSCs from other tissues and cells. We then performed an analysis aimed to identify absolute and relative abundance of transcripts in all cell types. We found that the set of transcripts uniquely expressed by MSCs is enriched in transcription factors and components of the Wnt signaling pathway. The analysis of differentially expressed genes also identified a set of genes specifically involved in the HSC niche and is complemented by functional studies that confirm the findings. Interestingly, some of these genes play a role in the maintenance of HSCs in a quiescent state supporting their survival and preventing them from proliferating and differentiating. We also show that MSCs modulate T cell functions in vitro and, upon in vivo administration, ameliorate experimental autoimmune encephalomyelitis (EAE).

CONCLUSIONS

Altogether, these findings provide novel and important insights on the mechanisms of T cell function regulation by MSCs and help to cement the rationale for their application in the treatment of autoimmune diseases.

Stem cells are currently seen as a treatment for tissue regeneration in neurological diseases such as multiple sclerosis, anticipating that they integrate and differentiate into neural cells. Mesenchymal stem cells (MSCs), a subset of adult progenitor cells, differentiate into cells of the mesodermal lineage but also, under certain experimental circumstances, into cells of the neuronal and glial lineage. Their clinical development, however, has been significantly boosted by the demonstration that MSCs display significant therapeutic plasticity mainly occurring through bystander mechanisms. These features have been exploited in the effective treatment of experimental autoimmune encephalomyelitis, an animal model of multiple sclerosis where the inhibition of the autoimmune response resulted in a significant amelioration of disease and decrease of demyelination, immune infiltrates and axonal loss. Surprisingly, these effects do not require MSCs to engraft in the central nervous system but depend on the cells' ability to inhibit pathogenic immune responses both in the periphery and inside the central nervous system and to release neuroprotective and pro-oligodendrogenic molecules favoring tissue repair. These results paved the road for the utilization of MSCs for the treatment of multiple sclerosis.

The diagnosis of a neurological disease of the central nervous system (CNS) is often associated with the anticipation of an irreversible and untreatable disability. This is the case also of multiple sclerosis (MS) where approved treatments effectively modulate the autoimmune attack to myelin antigens, but poorly affect neurodegeneration and do not promote tissue repair. Thus, stem cell-based therapies are increasingly being considered a possible strategy for diseases of the CNS. Mesenchymal stem cells (MSC), the safety of which has been demonstrated in the last 20 years through clinical trials and case studies, are of particular interest in view not only of their neuroprotective, but also of their immunomodulatory properties. Here, we review the therapeutic features of MSC that make them relevant in the treatment of CNS illnesses and discuss the pioneer clinical experience with MSC-based therapy in neurological diseases.

In several cell types, a regulated efflux of NAD(+) across Connexin 43 hemichannels (Cx43 HC) can occur, and extracellular NAD(+) (NAD(+)(e)) affects cell-specific functions. We studied the capability of bone marrow-derived human mesenchymal stem cells (MSC) to release intracellular NAD(+) through Cx43 HC. NAD(+) efflux, quantified by a sensitive enzymatic cycling assay, was significantly upregulated by low extracellular Ca(2+) (5-6-fold), by shear stress (13-fold), and by inflammatory conditions (3.1- and 2.5-fold in cells incubated with lipopolysaccharide (LPS) or at 39°C, respectively), as compared with untreated cells, whereas it was downregulated in Cx43-siRNA-transfected MSC (by 53%) and by cell-to-cell contact (by 45%). Further, we show that NAD(+)(e) activates the purinergic receptor P2Y(11) and a cyclic adenosin monophosphate (cAMP)/cyclic ADP-ribose/[Ca(2+)](i) signaling cascade, involving the opening, unique to MSC, of L-type Ca(2+) channels. Extracellular NAD(+) enhanced nuclear translocation of cAMP/Ca(2+)-dependent transcription factors. Moreover, NAD(+), either extracellularly added or autocrinally released, resulted in stimulation of MSC functions, including proliferation, migration, release of prostaglandin E(2) and cytokines, and downregulation of T lymphocyte proliferation compared with controls. No detectable modifications of MSC markers and of adipocyte or osteocyte differentiation were induced by NAD(+)(e). Controls included Cx43-siRNA transfected and/or NAD(+)-glycohydrolase-treated MSC (autocrine effects), and NAD(+)-untreated or P2Y(11)-siRNA-transfected MSC (exogenous NAD(+)). These findings suggest a potential beneficial role of NAD(+)(e) in modulating MSC functions relevant to MSC-based cell therapies.

The pathophysiology of cerebral cortical lesions in multiple sclerosis (MS) is not understood. We investigated cerebral cortex microvessels during immune-mediated demyelination in the MS model chronic murine experimental autoimmune encephalomyelitis (EAE) by immunolocalization of the endothelial cell tight junction (TJ) integral proteins claudin-5 and occludin, a structural protein of caveolae, caveolin-1, and the blood-brain barrier-specific endothelial transporter, Glut 1. In EAE-affected mice, there were areas of extensive subpial demyelination and well-demarcated lesions that extended to deeper cortical layers. Activation of microglia and absence of perivascular inflammatory infiltrates were common in these areas. Microvascular endothelial cells showed increased expression of caveolin-1 and a coincident loss of both claudin-5 and occludin normal junctional staining patterns. At a very early disease stage, claudin-5 molecules tended to cluster and form vacuoles that were also Glut 1 positive; the initially preserved occludin pattern became diffusely cytoplasmic at more advanced stages. Possible internalization of claudin-5 on TJ dismantling was suggested by its coexpression with the autophagosomal marker MAP1LC3A. Loss of TJ integrity was confirmed by fluorescein isothiocyanate-dextran experiments that showed leakage of the tracer into the perivascular neuropil. These observations indicate that, in the cerebral cortex of EAE-affected mice, there is a microvascular disease that differentially targets claudin-5 and occludin during ongoing demyelination despite only minimal inflammation.

Recent reports have highlighted that adult stem cells are granted with yet poorly understood properties other than multipotentiality. In particular, mesenchymal stem cells (MSCs) represent a subset of adult stromal cells that can down-regulate several functions of the immune cells. In addition, MSCs may promote survival of damaged cells and tissues through paracrine mechanisms, possibly under the guidance of environmental cues. Thus, MSCs clinical application in autoimmune diseases seems an appealing opportunity and preclinical results in different experimental models of autoimmunity further support this strategy. Despite the absolute need for caution related to several clinical and technical issues, MSCs are now on the edge of a new era of clinical applications.

Malignant forms of multiple sclerosis (MS) represent a limited group of very aggressive demyelinating diseases, which rapidly progress to severe disability leading often to life-threatening conditions. On these clinical entities, currently available therapies for MS are not very effective. Recently, it has been demonstrated that intense immunosuppression followed by autologous stem cell transplantation (ASCT) can affect the clinical course of individuals with severe MS and completely abrogate the inflammatory activity detected by magnetic resonance imaging (MRI). We report on the treatment with intense immune ablation followed by ASCT of three patients with malignant MS whose clinical course indicated a dramatically poor prognosis. This procedure succeeded in halting the rapidly worsening course of disease. The effect was long lasting, as demonstrated by a sustained efficacy over a two-year period in two subjects and 12 months in the third case. In addition, a striking effect on inflammation-related MRI findings was obtained. These results support a role for intense immunosuppression followed by ASCT as treatment in rapidly evolving malignant MS cases unresponsive to conventional therapies.

Microglia are highly plastic immune cells which exist in a continuum of activation states. By shaping the function of oligodendrocyte precursor cells (OPCs), the brain cells which differentiate to myelin-forming cells, microglia participate in both myelin injury and remyelination during multiple sclerosis. However, the mode(s) of action of microglia in supporting or inhibiting myelin repair is still largely unclear. Here, we analysed the effects of extracellular vesicles (EVs) produced in vitro by either pro-inflammatory or pro-regenerative microglia on OPCs at demyelinated lesions caused by lysolecithin injection in the mouse corpus callosum. Immunolabelling for myelin proteins and electron microscopy showed that EVs released by pro-inflammatory microglia blocked remyelination, whereas EVs produced by microglia co-cultured with immunosuppressive mesenchymal stem cells promoted OPC recruitment and myelin repair. The molecular mechanisms responsible for the harmful and beneficial EV actions were dissected in primary OPC cultures. By exposing OPCs, cultured either alone or with astrocytes, to inflammatory EVs, we observed a blockade of OPC maturation only in the presence of astrocytes, implicating these cells in remyelination failure. Biochemical fractionation revealed that astrocytes may be converted into harmful cells by the inflammatory EV cargo, as indicated by immunohistochemical and qPCR analyses, whereas surface lipid components of EVs promote OPC migration and/or differentiation, linking EV lipids to myelin repair. Although the mechanisms through which the lipid species enhance OPC maturation still remain to be fully defined, we provide the first demonstration that vesicular sphingosine 1 phosphate stimulates OPC migration, the first fundamental step in myelin repair. From this study, microglial EVs emerge as multimodal and multitarget signalling mediators able to influence both OPCs and astrocytes around myelin lesions, which may be exploited to develop novel approaches for myelin repair not only in multiple sclerosis, but also in neurological and neuropsychiatric diseases characterized by demyelination.

The burden of neurological diseases in western societies has accentuated the need to develop effective therapies to stop the progression of chronic neurological diseases. Recent discoveries regarding the role of the immune system in brain damage coupled with the development of new technologies to manipulate the immune response make immunotherapies an attractive possibility to treat neurological diseases. The wide repertoire of immune responses and the possibility to engineer such responses, as well as their capacity to promote tissue repair, indicates that immunotherapy might offer benefits in the treatment of neurological diseases, similar to the benefits that are being associated with the treatment of cancer and autoimmune diseases. However, before applying such strategies to patients it is necessary to better understand the pathologies to be targeted, as well as how individual subjects may respond to immunotherapies, either in isolation or in combination. Due to the powerful effects of the immune system, one priority is to avoid tissue damage due to the activity of the immune system, particularly considering that the nervous system does not tolerate even the smallest amount of tissue damage.

This Opinion deals with the apparent paradox between the 'immune privileged' status of the central nervous system (CNS) and its propensity to act as a B-cell fostering environment in a variety of neurological disorders. Evidence will be reviewed that: (i) molecules regulating B-cell homing and survival are produced in the CNS, (ii) in different neuroinflammatory diseases, B cells can undergo a local recapitulation of the differentiation occurring in secondary lymphoid organs and (iii) ectopic lymphoid follicles develop in the meninges of multiple sclerosis (MS) patients.

In recent years much excitement has been generated over the possibility that adult stem cells may attempt repair of the injured central nervous system (CNS), thus setting the stage for their utilisation in the treatment of neurodegenerative disorders. Recent studies have shown that some subsets of stem cells can also modulate the (auto)immune response, thus providing a rationale for their use as therapy for experimental autoimmune encephalomyelitis (EAE), an animal model for multiple sclerosis (MS). This article reviews the scientific evidence supporting the possible use of neural stem cells (NSCs) and mesenchymal stem cells (MSCs) for the treatment of MS. In addition, possible mechanisms sustaining the beneficial mode of action of haematopoietic stem cells (HSCs) following transplantation in MS individuals are discussed. Overall, it is proposed that limited subsets of adult stem cells may have a dual function that may be effective for the treatment of MS, an autoimmune disease of the CNS where degeneration of neural cells follows inflammation.

Multiple sclerosis (MS) is a complex neurological disease where, in genetically predisposed individuals, the unbalanced interplay between pathogenic and regulatory T cells will result in the progression of the autoimmune assault to neural antigens. Fingolimod (FTY720), an oral sphingosine 1-phosphate modulator recently approved for the treatment of MS, inhibits the egress of T cells from lymph nodes acting specifically on naïve and memory T cells and sparing effector T cells. Here we characterized IL-17 and IFNγ producing effector CD4 and CD8 positive T cells as well as CD4 positive CD25(high)CD127(low) regulatory T cells in MS patients before and 1 month after treatment was started. We observed that fingolimod did not significantly affect the percentage of CCR6 and CD161 positive T cells in both CD4 and CD8 compartments. In contrast, it significantly reduced the levels of both CD4+ CCR6+ CD161+ and CD8+ CCR6+ CD161+ producing IFNγ alone or in combination with IL-17. The percentage of IL-17 secreting cells in both subsets was affected by the treatment to a lesser extent. Finally, we observed that CD4+ CD25(high)CD127(low) regulatory T cells were decreased in MS patients compared to healthy controls and fingolimod significantly increased their frequencies. All together these findings demonstrate that fingolimod functionally modulates the ability of potentially pathogenic effector cells to produce relevant pro-inflammatory cytokines and increases the number of circulating regulatory T cells possibly contributing in restoring a balance between these populations.

: Human mesenchymal stem cells (hMSCs) are being increasingly pursued as potential therapies for immune-mediated conditions, including multiple sclerosis. Although they can suppress human Th1 responses, they reportedly can reciprocally enhance human Th17 responses. Here, we investigated the mechanisms underlying the capacity of hMSCs to modulate human Th1 and Th17 responses. Human adult bone marrow-derived MSCs were isolated, and their purity and differentiation capacity were confirmed. Human venous peripheral blood mononuclear cells (PBMC) were activated, alone, together with hMSC, or in the presence of hMSC-derived supernatants (sups). Cytokine expression by CD4+ T-cell subsets (intracellular staining by fluorescence-activated cell sorting) and secreted cytokines (enzyme-linked immunosorbent assay) were then quantified. The contribution of prostaglandin E2 (PGE2) as well as of myeloid cells to the hMSC-mediated regulation of T-cell responses was investigated by selective depletion of PGE2 from the hMSC sups (anti-PGE2 beads) and by the selective removal of CD14+ cells from the PBMC (magnetic-activated cell sorting separation). Human MSC-secreted products could reciprocally induce interleukin-17 expression while decreasing interferon-γ expression by human CD4+ T cells, both in coculture and through soluble products. Pre-exposure of hMSCs to IL-1β accentuated their capacity to reciprocally regulate Th1 and Th17 responses. Human MSCs secreted high levels of PGE2, which correlated with their capacity to regulate the T-cell responses. Selective removal of PGE2 from the hMSC supernatants abrogated the impact of hMSC on the T cells. Selective removal of CD14+ cells from the PBMCs also limited the capacity of hMSC-secreted PGE2 to affect T-cell responses. Our discovery of a novel PGE2-dependent and myeloid cell-mediated mechanism by which human MSCs can reciprocally induce human Th17 while suppressing Th1 responses has implications for the use of, as well as monitoring of, MSCs as a potential therapeutic for patients with multiple sclerosis and other immune-mediated diseases.

CONCLUSIONS

Although animal studies have generated a growing interest in the anti-inflammatory potential of mesenchymal stem cells (MSCs) for the treatment of autoimmune diseases, MSCs possess the capacity to both limit and promote immune responses. Yet relatively little is known about human-MSC modulation of human disease-implicated T-cell responses, or the mechanisms underlying such modulation. The current study reveals a novel prostaglandin E2-dependent and myeloid cell-mediated mechanism by which human MSCs can reciprocally regulate human Th17 and Th1 responses, with implications for the use of MSCs as a potential therapeutic for patients with multiple sclerosis and other immune-mediated diseases.

The aim of the study was to characterise CCR7+ and CCR7- memory T cells infiltrating the inflamed joints of patients with juvenile idiopathic arthritis (JIA) and to investigate the functional and anatomical heterogeneity of these cell subsets in relation to the expression of the inflammatory chemokine receptors CXCR3 and CCR5. Memory T cells freshly isolated from the peripheral blood and synovial fluid (SF) of 25 patients with JIA were tested for the expression of CCR7, CCR5, CXCR3 and interferon-gamma by flow cytometry. The chemotactic activity of CD4 SF memory T cells from eight patients with JIA to inflammatory (CXCL11 and CCL3) and homeostatic (CCL19, CCL21) chemokines was also evaluated. Paired serum and SF samples from 28 patients with JIA were tested for CCL21 concentrations. CCR7, CXCR3, CCR5 and CCL21 expression in synovial tissue from six patients with JIA was investigated by immunohistochemistry. Enrichment of CD4+, CCR7- memory T cells was demonstrated in SF in comparison with paired blood from patients with JIA. SF CD4+CCR7- memory T cells were enriched for CCR5+ and interferon-gamma+ cells, whereas CD4+CCR7+ memory T cells showed higher coexpression of CXCR3. Expression of CCL21 was detected in both SF and synovial membranes. SF CD4+ memory T cells displayed significant migration to both inflammatory and homeostatic chemokines. CCR7+ T cells were detected in the synovial tissue in either diffuse perivascular lymphocytic infiltrates or organised lymphoid aggregates. In synovial tissue, a large fraction of CCR7+ cells co-localised with CXCR3, especially inside lymphoid aggregates, whereas CCR5+ cells were enriched in the sublining of the superficial subintima. In conclusion, CCR7 may have a role in the synovial recruitment of memory T cells in JIA, irrespective of the pattern of lymphoid organisation. Moreover, discrete patterns of chemokine receptor expression are detected in the synovial tissue.

Nicotinamide adenine dinucleotide (NAD+) biosynthesis from nicotinamide is used by mammalian cells to replenish their NAD+ stores and to avoid unwanted nicotinamide accumulation. Pharmacological inhibition of nicotinamide phosphoribosyltransferase (NAMPT), the key enzyme in this biosynthetic pathway, almost invariably leads to intracellular NAD+ depletion and, when protracted, to ATP shortage and cell demise. Cancer cells and activated immune cells express high levels of NAMPT and are highly susceptible to NAMPT inhibitors, as shown by the activity of these agents in models of malignant and inflammatory disorders. As the spectrum of conditions which could benefit from pharmacological NAMPT inhibition becomes broader, the mechanisms accounting for their activity are also eventually becoming apparent, including the induction of autophagy and the impairment of Ca2+--and NF-κB-dependent signaling. Here, we discuss the rationales for exploiting NAMPT inhibitors in cancer and inflammatory diseases and provide an overview of the preclinical and clinical studies in which these agents have been evaluated.

BACKGROUND

Achieving good adherence to self-injected treatments for multiple sclerosis can be difficult. Injection devices may help to overcome some of the injection-related barriers to adherence that can be experienced by patients. We sought to assess short-term adherence to, and tolerability of, interferon (IFN) β-1a administered via electronic autoinjection device in patients with relapsing-remitting multiple sclerosis (RRMS).

METHODS

BRIDGE (RebiSmart to self-inject Rebif serum-free formulation in a multidose cartridge) was a 12-week, multicentre, open-label, single-arm, observational, Phase IV study in which patients self-administered IFN β-1a (titrated to 44 μg), subcutaneously (sc), three times weekly, via electronic autoinjection device. Patients were assessed at baseline and 4-weekly intervals to Week 12 or early termination (ET) for: physical examinations; diary card completion (baseline, Weeks 4, 8 only); neurological examinations (baseline, Week 12/ET only); MS Treatment Concern Questionnaire (MSTCQ; Weeks 4, 8, 12 only); Convenience Questionnaire (Week 12 only); Hospital Anxiety and Depression Scale (HADS); and Paced Auditory Serial Addition Task (PASAT; baseline only). Adherence was defined as administration of ≥ 80% of scheduled injections, recorded by the autoinjection device.

RESULTS

Overall, 88.2% (105/119; intent-to-treat population) of patients were adherent; 67.2% (80/119) administered all scheduled injections. Medical reasons accounted for 35.6% (31/87) of missed injections, forgetfulness for 20.6% (18/87). Adherence did not correlate with baseline Expanded Disability Status Scale (P = 0.821) or PASAT (P = 0.952) scores, or pre-study therapy (P = 0.303). No significant changes (baseline-Week 12) in mean HADS depression (P = 0.482) or anxiety (P = 0.156) scores were observed. 'Overall convenience' was the most important reported benefit of the autoinjection device. Device features associated with handling and ease of use were highly rated. Mean MSTCQ scores for 'flu-like' symptoms (P = 0.022) and global side effects (P = 0.002) significantly improved from Week 4-12. Mean MSTCQ scores for pain at injection site and injection pain increased from Week 4-12 (P < 0.001). Adverse events were mild/moderate. No new safety signals were identified.

CONCLUSIONS

Convenience and ease of use of the autoinjection device may improve adherence and, therefore, outcomes, in patients with RRMS receiving sc IFN β-1a.

BACKGROUND

EU Clinical Trials Register (EU-CTR; http://www.clinicaltrialsregister.eu): 2009-013333-24.

Recent evidence has shown that CD56(bright) NK cells, a subset of NK cells abundant in lymph nodes, may have an immunoregulatory function. In multiple sclerosis (MS), expansion of CD56(bright) NK cells has been associated to successful response to different treatments and to remission of disease during pregnancy; how whether they exert immunoregulation in physiologic conditions and whether this is impaired in MS is not known. We dissected the immunoregulatory role of CD56(bright) NK cells function in healthy subjects (HS) and compared it with that of untreated MS subjects or patients with clinically isolated syndrome suggestive of MS (CIS). We found that CD56(bright) NK cells from HS acquire, upon inflammatory cues, the capability of suppressing autologous CD4+T cell proliferation through direct cytotoxicity requiring engagement of natural cytotoxicity receptors (NCRs) and secretion of granzyme B. CD56(bright) NK cells from patients with MS/CIS did not differ in frequency and share a similar phenotype but displayed a significantly lower ability to inhibit autologous T cell proliferation. This impairment was not related to deficient expression of NCRs or granzyme B by CD56(bright) NK cells, but to increased HLA-E expression on T cells from MS/CIS subjects, which could enhance the inhibitory effect mediated by NKG2A that is homogeneously expressed on CD56(bright) NK cells. The defect in controlling autologous T cells by CD56(bright) NK cells in MS/CIS might contribute to the excess of autoimmune response that is associated to disease development.