Immune modulation in MS models by peptidoglycan, CD97 and CD44

Multipele sclerose (MS) wordt beschouwd als een chronische ziekte van het centrale zenuwstelsel (CZS: hersenen en ruggenmerg). Bij MS patiënten wordt gedacht dat het afweersysteem ontspoord is. Het afweersysteem is normaliter gericht tegen lichaamsvreemde, bedreigende componenten, zoals virussen en bacteriën. Bij MS patiënten worden door het afweersysteem ook lichaamseigen componenten in het CZS aangevallen, zoals de beschermende schede rondom de uitlopers van zenuwcellen (de zogeheten myelineschede). Schade aan de myelineschede en zenuwcellen verstoort de zenuwfunctie. Het klinische beeld wordt bepaald door de plaats van myeline afbraak. MS wordt gekarakteriseerd door ontstekingshaarden (in.ltraten) in het CZS. Deze in.ltraten worden gevormd door cellen van het afweersysteem, de leukocyten. Er bestaan verschillende typen leukocyten, zoals antigeen presenterende cellen (APC), T en B cellen. APC kunnen bacteriën of virussen, maar ook andere componenten opnemen. De APC herkent bepaalde structuren van bacteriën en virussen en worden hierdoor geactiveerd. In dit proefschrift hebben we onderzocht of een bacterieel celwand component, het peptidoglycaan, kan bijdragen aan MS. Verder hebben we onderzocht of de cel oppervlakte moleculen CD97 en CD44 betrokken zijn bij MS. Peptidoglycaan (PG) is een belangrijk bestanddeel van de celwand van bacteriën en werkt ontstekingsbevorderend via activering van speci.eke receptoren voor PG. Deze receptoren zijn aanwezig op en in APC. APC kunnen PG vervoeren van de slijmvliezen of van plaatsen van infectie naar de lymfoïde organen. Daarnaast worden PG-bevattende APC ook in hoge aantallen gevonden op plaatsen van chronische ontsteking, zoals in de gewrichten van patiënten met reuma. Eerder onderzoek heeft aangetoond dat ook in hersenweefsel van MS patiënten meer PG-bevattende APC aanwezig zijn in vergelijking met controle personen. Wij denken dat PG bijdraagt aan de chronische ontsteking bij patiënten met MS door activering van APC. Om deze hypothese te toetsen hebben we gebruik gemaakt van diermodellen voor MS. In die diermodellen wordt het op MS gelijkende ziektebeeld experimentele autoimmuun encefalomyelitis (EAE) genoemd. Lysozym en amidase zijn twee enzymen die geproduceerd worden door APC, en die PG kunnen afbreken. Hierdoor verliest PG zijn ontstekingsbevorderende werking. In hoofdstuk 2 hebben we de aanwezigheid van PG-bevattende cellen en PG-afbrekende enzymen bepaald in hersenweefsel van twee verschillende aapmodellen voor MS. Voor deze studie hebben we ingevroren weefsels gebruikt uit voorgaande experimenten. EAE in marmoset apen verloopt chronisch, terwijl resus apen een acuut ziekteverloop vertonen. In beide modellen werden bij apen met EAE meer PG-bevattende cellen in de hersenen gedetecteerd in vergelijking met controle dieren zonder EAE. Hersenweefsel van resus apen die een acuut ziektebeeld vertonen bevatte veel meer cellen met PG in vergelijking met marmoset apen waar het ziektebeeld chronisch verloopt. Verder zagen we dat amidase alleen aanwezig was in in.ltraten in het hersenweefsel van resus apen met EAE, en niet in marmoset apen. Lysozym daarentegen, was afwezig was in alle dieren (resus en marmoset), met of zonder EAE. Deze gegevens impliceren dat tijdens EAE cellen naar de hersenen migreren die PG bevatten, waarbij het aantal PG-bevattende cellen afhangt van het verloop van de ziekte. De afwezigheid van PG-afbrekende enzymen in de hersenen suggereert dat PG op deze plek niet afgebroken kan worden en mogelijk bijdraagt aan de chronische ontsteking. In hoofdstuk 3 hebben we aangetoond dat PG daadwerkelijk kan bijdragen aan EAE ontwikkeling in muizen. EAE treedt niet spontaan op, maar wordt geïnduceerd door een injectie van myeline antigenen in een emulsie van olie met ontstekingbevorderende componenten (gewoonlijk dode tuberculose bacteriën). Het huidige onderzoek toont aan dat PG, als een enkel component van de bacteriële celwand, de gehele tuberkel bacterie kan vervangen voor de inductie van EAE. Verder zagen we dat het ingespoten PG al na 4 uur aanwezig was in de lymfoïde organen in een gespecialiseerd type APC, de dendritische cel. APC kunnen andere cellen activeren, zoals T en B cellen die speci.ek gericht zijn tegen de antigenen (deeltjes) die de APC presenteren. Dit gebeurt onder normale omstandigheden in lymfoïde organen, zoals de lymfeklieren en de milt. In een kweeksysteem hebben we dit nagebootst. We hebben aangetoond dat PG de dendritische cel kan stimuleren tot een verhoogde opname van antigenen en productie van ontstekingsbevorderende eiwitten. PG kon via dendritische cellen ook T cellen laten delen en uitrijpen tot ontstekingsbevorderende T helper 1 cellen. Deze gegevens suggereren dat PG in lymfoïde organen of zelfs in het CZS kan bijdragen aan MS, door het geven van ontstekingssignalen aan APC. Activatie en migratie van cellen vanuit de lymfoïde organen naar plaatsen van ontsteking (bij MS patiënten het CZS) is een zorgvuldig gereguleerd proces. Dit proces wordt onder andere gecoördineerd door moleculen die aanwezig zijn op het oppervlak van zowel de migrerende cellen als de cellen van de bloedvaatjes op plaatsen waar een ontstekingsreactie is. De moleculen van de migrerende cellen en de receptor moleculen op andere cellen grijpen in elkaar als een sleutel en slot. Door deze signalen kunnen de cellen de bloedbaan uit migreren. In dit proefschrift hebben we tevens onderzocht of de cel oppervlakte moleculen CD97 en CD44 bijdragen aan het ziekteproces in MS, zoals hieronder beschreven. CD97 en CD44 zijn moleculen die voorkomen op het oppervlak van verschillende celtypen. Eerder onderzoek heeft aangetoond dat deze moleculen een belangrijke rol spelen bij reuma en chronische darmontsteking (colitis). Deze moleculen dragen mogelijk bij aan het ziekteproces in MS door coördinatie van verschillende cellulaire processen, zoals activatie, deling en migratie naar plaatsen van ontsteking. CD97 is betrokken bij de activatie, migratie en adhesie van cellen. CD55 was de enige ligand voor CD97 dat in de literatuur bekend was tijdens het onderzoek voor dit proefschrift. Inmiddels is bekend dat CD97 zich ook kan binden aan integrinen en componenten van de extracellulaire matrix (tussenstof). CD55 beschermt cellen tegen complement factoren. Het complement systeem bestaat uit moleculen die helpen bij het bestrijden van infecties. Doordat lichaamseigen cellen moleculen zoals CD55 op hun oppervlakte dragen worden zij niet aangevallen door het complement. Het onderzoek beschreven in hoofdstuk 4 toont aan dat CD97 en CD55 aanwezig zijn op plaatsen van ontsteking in het CZS van MS patiënten. CD97 kwam niet en CD55 kwam nauwelijks tot expressie op cellen in normaal hersenweefsel. Daarentegen werd in ontstekingsgebieden in MS hersenweefsel een verhoogde expressie van CD55 gevonden op endotheelcellen (cellen die de wand van bloedvaten aan de binnenzijde bekleden). Direct naast deze endotheelcellen kwam CD97 tot expressie op in.ltrerende APC, T en B cellen. Daarom veronderstellen wij dat CD97 op leukocyten bijdraagt aan de migratie van cellen uit de bloedbaan naar het CZS, door interactie met CD55 op endotheelcellen. Om te bepalen of CD97 interacties met CD55 van belang zijn voor MS, hebben we EAE in muizen behandeld met een blokkerende antistof die de interactie tussen CD97-CD55 voorkomt. Het gebruikte behandelingsschema resulteerde niet in een verminderde ziektelast. Dit kan betekenen dat 1) de CD97-CD55 interactie niet van belang is in muizen met EAE, of 2) de behandeling van te korte duur was om de ziektelast te verminderen. In vervolgonderzoek kunnen we gebruik maken van muizen die het gen voor CD97 ontberen, om zodoende een beter inzicht te krijgen in de functionele rol van CD97 in MS. CD44 en de variant isovormen van CD44 (CD44v1-v10) spelen een belangrijke rol bij allerlei biologische processen, zoals celactivatie, -costimulatie, -migratie, ­adhesie en geprogrammeerde celdood. Geprogrammeerde celdood is van belang bij het beëindigen van een ontstekingsreactie, b.v. als het infectiegevaar geweken is. Daarnaast kan CD44 als een magneet fungeren voor ontstekingsmediatoren. Zodoende kan CD44 ervoor zorgen dat er lokaal een verhoogde concentratie ontstaat van deze ontstekingsmediatoren. In hoofdstuk 5 hebben we aangetoond dat bepaalde CD44v isovormen speci.ek tot expressie komen in hersenweefsel van MS patiënten en muizen met EAE. In muizen met EAE hebben we op verschillende tijdstippen na ziekte inductie de expressie van CD44v10 bestudeerd in de hersenen. CD44v10 kwam tot expressie op verschillende CZS in.ltrerende cellen (APC, T en B cellen). Met name CD44v10+ T helper cellen konden zelfs nog op late tijdstippen na EAE inductie gedetecteerd worden, terwijl deze cellen afwezig waren in controlemuizen. Daarnaast laten we zien dat muizen, die het gen voor CD44v7 of CD44v10 missen, een verminderde ziektelast vertoonden na EAE inductie. Deze gegevens tonen aan dat CD44v isovormen een belangrijke bijdrage kunnen leveren aan EAE. Deze isovormen zijn mogelijk ook betrokken bij het ziekteproces in MS. Verder onderzoek moet aantonen op welke manier CD44v moleculen een rol spelen in de ziekte (b.v. bij adhesie of geprogrammeerde celdood). Kort samengevat heeft dit proefschrift de volgende inzichten opgeleverd. PG kan bijdragen aan MS door activatie van APC. PG-bevattende cellen kunnen mee migreren naar het CZS tijdens MS of EAE en dragen daar mogelijk bij aan een ontstekings-bevorderend milieu. Niet alleen in het CZS, maar ook in lymfoïde organen zou PG een bijdrage kunnen leveren aan MS en EAE. Hier kan PG bijdragen door stimulatie van T en B cellen die gericht zijn tegen componenten van het CZS (zoals myeline), via APC. CD97 en CD44v isovormen kunnen ervoor zorgen dat celactivatie, -migratie, -adhesie en -overleving gestimuleerd worden tijdens MS en EAE. We laten zien dat CD97 en CD44v isovormen tot expressie komen in ontstekingshaarden in het CZS van MS patiënten. EAE in muizen kan worden verminderd door afwezigheid van CD44v7 of CD44v10. Soortgelijke effecten worden verwacht bij afwezigheid van het CD97 molecuul. Dit proefschrift verschaft een verbeterd inzicht in de pathogenese van MS en draagt mogelijk bij in een rationele doelgerichte ontwikkeling van nieuwe therapeutische benaderingen.Multiple sclerosis (MS) is a chronic in.ammatory disease of the central nervous system (CNS). We are interested in the mechanisms involved in the immunopathogenesis of MS, especially in microbial signals able to break T cell tolerance, and in the function of molecules on the activated cells in costimulation, migration, adhesion and apoptosis. Peptidoglycan (PG) is a major cell wall component of Gram-positive bacteria, that activates the innate immune system by binding to TLR2/6, Nod1 (Card4) and Nod2 (Card 15) receptors. The enzymes lysozyme and NAMLAA are produced by phagocytic cells and can degrade PG. However, after bacterial phagocytosis, PG can persist intracellularly when the digestion is incomplete. During chronic in.ammation, PG-containing antigen presenting cells (APC) accumulate at in.ammatory sites, as previously demonstrated in patients with colitis, arthritis and MS. At these in.ammatory sites or in peripheral lymphoid tissues PG may serve as a costimulatory factor in autoimmune disease by overriding tolerance against self-antigens. To obtain more insight into the functional relevance of PG in MS, we compared the presence of PG and its degrading enzymes in the CNS of two distinct non­human primate EAE models (chapter 2). As in brain tissue of MS patients we here demonstrate that EAE-affected brain tissue from marmoset and rhesus monkeys contained elevated numbers of cells with PG, compared to control immunized animals. Interestingly, chronic EAE in marmoset monkeys was accompanied by a modest number of PG-containing cells in the brain, whereas brain tissue from rhesus monkeys that had developed acute EAE contained abundant numbers of PG-containing cells. Lysozyme was only sporadically expressed in EAE-affected brain tissue from both monkey species. In contrast, NAMLAA was expressed on many perivascular cells in EAE-affected brain tissue from rhesus monkeys, but only by few perivascular cells in marmoset monkeys. Double-labeling revealed that NAMLAA was mostly expressed by neutrophils. Thus, in EAE-affected brain tissue, PG was present within signi.cantly higher numbers of APC compared to controls. Conversely, lysozyme was mainly absent in normal and in.amed brain tissue, and NAMLAA was only abundantly expressed during acute CNS in.ammation in rhesus monkeys. These data suggest that PG persists inside APC in the CNS and may contribute to the in.ammation by stimulating TLR/Nod receptors. To further elucidate the functional contribution of PG to autoimmune disease we investigated the hypothesis that PG acts as a costimulatory factor for disease development in MS, by using mouse EAE (chapter 3). EAE is normally induced by s.c. immunization of autoantigens admixed in a strong adjuvant with attenuated Mycobacterium tuberculosis bacteria. In.ammatory Staphylococcus aureus PG could replace whole M. tuberculosis in EAE induction. We demonstrate that S. aureus PG was transported to the spleen and draining lymph nodes after immunization with S. aureus PG-containing adjuvant. In the draining lymph nodes S. aureus PG was localized within dendritic cell (DC) clusters. Clusters of PG-containing cells were also present in the spleen of rhesus monkeys that developed EAE (chapter 2). Compared to control mice that were immunized with autoantigen in incomplete Freund’s adjuvant (IFA), PG increased autoantigen­speci.c T cell proliferation and Th1 cell development, when the autoantigen-IFA mix was supplemented with PG. By using bone marrow-derived DC, S. aureus PG was shown to stimulate antigen uptake by DC, DC maturation, and subsequent differentiation and proliferation of Th1 cells. Taken together, proin.ammatory PG may stimulate autoimmune-mediated processes in MS, either in the CNS or in the periphery, by accelerating (auto)antigen uptake, initiating DC maturation, and polarizing and expanding (auto)antigen-speci.c T cells. CD97 and CD44 are membrane-expressed molecules required for cell-cell and cell-matrix interactions. These molecules contribute to disease development in experimental models for other chronic in.ammatory diseases, e.g. arthritis and colitis and may determine interactions between leukocytes, endothelium and the extracellular matrix (ECM) of the CNS during MS and EAE. Here we assessed whether CD97 and CD44v isoforms are involved in the immunopathogenesis of MS and EAE. CD97 is a member of the 7-span transmembrane receptor family that is expressed on leukocytes early after activation. CD97 is involved in migration and adhesion, by binding to CD55 (decay accelerating factor), a5/ß1 and avß3 integrins and ECM components. The CD97-ligand CD55 is expressed on several cell types, protecting them from complement-mediated damage. Here we examined the expression of CD97 and CD55 by immunohistochemistry in MS brain tissue. Chapter 4 shows that cells in normal white matter did not express CD97, whereas many MF or microglial cells and T cells in MS brain lesions expressed CD97. Endothelial cells in normal white matter only modestly expressed CD55. In contrast, in MS brain lesions, endothelial cells expressed high levels of CD55 and different leukocyte subsets also expressed CD55. In active lesions, expression of CD55 was predominantly detected on MF or microglial cells. In these lesions, a substantial proportion of cells also expressed CD97. The soluble form of CD97 was found in serum, but not in CSF of a signi.cant number of MS patients. These .ndings suggest that local CD97-CD55 interactions may contribute to the immunopathogenesis of MS. CD55 expressed on brain endothelial cells may serve to protect the vessels from complement-mediated damage. Additionally, CD97­expressing cells may utilize CD55 on activated brain endothelial cells to facilitate adhesion and transmigration into the CNS. To determine whether CD97-CD55 interactions are important in disease development, we blocked the interaction between CD97 and CD55 by CD97 Mab treatment in the priming phase of mouse EAE. This treatment regimen did not affect the onset and severity of disease. However, these data do not exclude the possibility that CD97-CD55 interactions are involved in EAE development, since the treatment duration might have been suboptimal. Alternatively, CD97 may participate in MS and EAE by binding to integrins or ECM components. By using mice with a genetic deletion for CD97, we can elucidate whether CD97 functionally contributes to EAE development. CD44 and its variant isoforms (CD44v1-v10) are required for many different biological processes such as lymphocyte activation, costimulation, adhesion/ extravasation into in.ammatory sites and apoptosis. The CD44 molecule also serves as a docking site for mediators of in.ammation by binding cytokines, growth factors and chemoattractants. One gene encodes different CD44v isoforms, which emerge as a result of complex RNA splicing. We determined whether CD44v isoforms are involved in the immunopathogenesis of MS and EAE (chapter 5). It appeared that only CD44v10 was expressed by normal white matter endothelium, whereas both CD44v3 and v10 were expressed by cells in perivascular in.ltrates and by microglia or MF in active lesions. Due to technical dif.culties, we could not assess expression of CD44v7. All other isoforms were not expressed in MS and control brain. Also in mouse EAE, brain-in.ltrating cells expressed CD44v10 and were already detected before clinical signs. The number of leukocytes that expressed CD44v10 paralleled the extent of clinical EAE symptoms. CD44v10-expressing CD4+ T cells persisted in EAE brain until late time points after immunization, whereas these cells were absent in control-immunized mice. Single deletion for either CD44v7 or CD44v10 resulted in reduced EAE, which was accompanied by reduced in.ltration of the spinal cord. Adoptive transfer experiments demonstrated that CD44v7 exerted its effect on the donor lymphocyte compartment as well as on the recipients APC. No clear differences were detected in autoantigen-speci.c T cell proliferation and cytokine production around the day of onset. In the remission phase of EAE, CD44v7-deleted lymph node cells showed a reduced proliferation compared to wild type cells. At present, it is unclear by which mechanism CD44v7 and CD44v10 reduced EAE. Currently we are exploring the possibility that de.ciency of CD44v7 or v10 promote clearance of in.ammation by the induction of apoptosis. Additionally, CD44v10 may also affect cell migration, as indicated in animal models for Th1-mediated DTH responses. It remains to be determined whether CD44v3 can functionally contribute to EAE. Taken together these data show that a single bacterial component, PG, can create an in.ammatory environment. During an autoimmune attack, PG-containing cells most likely enter the in.ammatory site and contribute to the pathological process. Intracellular PG may activate cells by engaging TLR or Nod receptors. PG promotes antigen uptake, DC maturation and proin.ammatory cytokine production. In the presence of autoantigens, PG induces autoantigen-speci.c Th1 cell priming and expansion. This is relevant in MS, since elevated numbers of PG-containing APC are found in the CNS, where autoantigens are released during in.ammatory attacks. Moreover, autoantigens and PG are present within APC of peripheral lymphoid organs, where similar events may take place. CD97 and CD44v isoforms may facilitate cellular activation, costimulation, adhesion, extravasation and survival. By their interactions with other cellular ligands and different ligands in ECM these molecules likely contribute to tissue damage and chronic in.ammation in MS. We here show that CD97 and CD44v molecules are expressed in MS brain lesions. EAE models demonstrated that by targeting CD44v7 or CD44v10 the clinical symptoms of the disease can be reduced. Total prevention of CD97 expression, by genetic deletion or long-lasting antibody treatment against ligand-binding CD97EGF domains may give similar effects. Eventually, altering the immune response by the modulation of PG, CD97 and CD44v isoforms in EAE will give more insights in the possibilities for therapeutic interventions in MS

Reduced experimental autoimmune encephalomyelitis after intranasal and oral administration of recombinant lactobacilli expressing myelin antigen

Oral administration of autoantigens is a safe and convenient way to induce peripheral T-cell tolerance in autoimmune diseases like multiple sclerosis (MS). To increase the efficacy of oral tolerance induction and obviate the need for large-scale purification of human myelin proteins, we use genetically modified lactobacilli expressing myelin antigens. A panel of recombinant lactobacilli was constructed producing myelin proteins and peptides, including human and guinea pig myelin basic protein (MBP) and proteolipid protein peptide 139-151 (PLP139-151). In this study we examined whether these Lactobacillus recombinants are able to induce oral and intranasal tolerance in an animal model for multiple sclerosis, experimental autoimmune encephalomyelitis (EAE). Lewis rats received soluble cell extracts of Lactobacillus transformants intranasally three times prior to induction of EAE. For the induction of oral tolerance, rats were fed live transformed lactobacilli for 20 days. Ten days after the first oral administration EAE was induced. Intranasal administration of extracts containing guinea pig MBP (gpMBP) or MBP72-85 significantly inhibited EAE in Lewis rats. Extracts of control transformants did not reduce EAE. Live lactobacilli expressing guinea pig MBP72-85 fused to the marker enzyme β-glucuronidase (β-gluc) were also able to significantly reduce disease when administered orally. In conclusion, these experiments provide proof of principle that lactobacilli expressing myelin antigens reduce EAE after mucosal (intranasal and oral) administration. This novel method of mucosal tolerance induction by mucosal administration of recombinant lactobacilli expressing relevant autoantigens could find applications in autoimmune disease in general, such as multiple sclerosis, rheumatoid arthritis and uveitis

Proinflammatory bacterial peptidoglycan as a cofactor for the development of central nervous system autoimmune disease

Upon stimulation by microbial products through TLR, dendritic cells (DC) acquire the capacity to prime naive T cells and to initiate a proinflammatory immune response. Recently, we have shown that APC within the CNS of multiple sclerosis (MS) patients contain peptidoglycan (PGN), a major cell wall component of Gram-positive bacteria, which signals through TLR and NOD. In this study, we report that Staphylococcus aureus PGN as a single component can support the induction of experimental autoimmune encephalomyelitis (EAE) in mice, an animal model for MS. Mice immunized with an encephalitogenic myelin oligodendrocyte glycoprotein peptide in IFA did not develop EAE. In contrast, addition of PGN to the emulsion was sufficient for priming of autoreactive Th1 cells and development of EAE. In vitro studies demonstrate that PGN stimulates DC-mediated processes, reflected by increased Ag uptake, DC maturation, Th1 cell expansion, activation, and proinflammatory cytokine production. These data indicate that PGN-mediated interactions result in proinflammatory stimulation of Ag-specific effector functions, which are important in the development of EAE. These PGN-mediated processes may occur both within the peripheral ly

The STAR Study: A Real-World, International, Observational Study of the Safety and Tolerability of, and Adherence to, Serum-Free Subcutaneous Interferon beta-1a in Patients With Relapsing Multiple Sclerosis

AbstractBackgroundAdverse reactions, particularly injection site reactions (ISRs), are common reasons for nonadherence to injectable multiple sclerosis (MS) treatments. Adherence to MS treatment is important to ensure good treatment outcomes.ObjectiveThe aim of this study was to assess the local tolerability of subcutaneous (SC) serum-free interferon (IFN) β-1a in patients with relapsing MS over 1 year in a real-life, international setting. The study also assessed safety, disease activity, and adherence.MethodsThis was a prospective, international, multicenter, observational study of 251 patients with relapsing-remitting MS treated with SC serum-free IFN β-1a 44 μg or 22 μg 3 times weekly for 12 months or until early discontinuation. The primary end point was the proportion of patients with ISRs. Secondary end points included proportion of patients with adverse events (AEs); annualized relapse rate (ARR); proportion of patients remaining relapse-free; and adherence to treatment.ResultsDuring the observation period, 27.5% (69 of 251) of patients experienced nonserious ISRs, which was consistent with the incidence reported in clinical studies. Five patients discontinued treatment and 2 patients suspended treatment because of ISRs. Mean age was 35.8 years; patients were predominantly white (94.8%), and two thirds (66.1%) were female. The overall incidence of AEs was 63.7% (160 of 251), and overall safety and tolerability were assessed as excellent, very good, or good in >85% of patients. More than 70% of patients remained relapse-free, and the mean ARR was 0.4. More than 90% of patients had very good or good adherence to treatment; a significantly greater proportion of these were relapse-free at 12 months compared with those with fair or poor adherence (77.6% vs 50.0%; P = 0.0107), and their ARR was significantly lower (0.3 vs 0.9; P = 0.0055). Patients with fair or poor adherence had 4.6 times higher odds of experiencing a relapse than those with very good or good adherence.ConclusionsThe incidence of ISRs and the overall safety profile in this observational study, in an international population in a real-life setting, confirm the good local tolerability of SC serum-free IFN β-1a reported in clinical studies. The association between good adherence and a lower ARR underlines the importance of good adherence. The good local and general tolerability of SC IFN β-1a may help ensure a high level of adherence, which is associated with better clinical outcomes. ClinicalTrials.gov identifier: NCT01080027

Reduced experimental autoimmune encephalomyelitis after intranasal and oral administration of recombinant lactobacilli expressing myelin antigen

Oral administration of autoantigens is a safe and convenient way to induce peripheral T-cell tolerance in autoimmune diseases like multiple sclerosis (MS). To increase the efficacy of oral tolerance induction and obviate the need for large-scale purification of human myelin proteins, we use genetically modified lactobacilli expressing myelin antigens. A panel of recombinant lactobacilli was constructed producing myelin proteins and peptides, including human and guinea pig myelin basic protein (MBP) and proteolipid protein peptide 139-151 (PLP139-151). In this study we examined whether these Lactobacillus recombinants are able to induce oral and intranasal tolerance in an animal model for multiple sclerosis, experimental autoimmune encephalomyelitis (EAE). Lewis rats received soluble cell extracts of Lactobacillus transformants intranasally three times prior to induction of EAE. For the induction of oral tolerance, rats were fed live transformed lactobacilli for 20 days. Ten days after the first oral administration EAE was induced. Intranasal administration of extracts containing guinea pig MBP (gpMBP) or MBP72-85 significantly inhibited EAE in Lewis rats. Extracts of control transformants did not reduce EAE. Live lactobacilli expressing guinea pig MBP72-85 fused to the marker enzyme β-glucuronidase (β-gluc) were also able to significantly reduce disease when administered orally. In conclusion, these experiments provide proof of principle that lactobacilli expressing myelin antigens reduce EAE after mucosal (intranasal and oral) administration. This novel method of mucosal tolerance induction by mucosal administration of recombinant lactobacilli expressing relevant autoantigens could find applications in autoimmune disease in general, such as multiple sclerosis, rheumatoid arthritis and uveitis

Expression of the EGF-TM7 receptor CD97 and its ligand CD55 (DAF) in multiple sclerosis

CD97 is a recently identified seven-span transmembrane (7-TM) protein that is expressed by leukocytes early after activation. CD97 binds to its cellular ligand CD55 (decay accelerating factor), which protects several cell types from complement-mediated damage. The functional consequences of CD97-CD55 binding are largely unknown, but previous data imply that CD97-CD55 interactions play a role in cellular activation, migration, and adhesion under inflammatory conditions. Here we examined the expression of CD97 and CD55 by immunohistochemistry in multiple sclerosis (MS). On the basis of established criteria for inflammation and demyelination, different lesion stages were distinguished in MS post-mortem brain tissue. In normal white matter, CD97 expression was not found, but CD55 was expressed with weak staining intensity on endothelial cells. In pre-active lesions, defined by abnormalities of the white matter, many infiltrating T cells, macrophages (MPhi) and microglia expressed CD97. CD55 was highly expressed by endothelial cells. In active lesions with myelin degradation, MPhi and microglia expressed both CD55 and CD97. Furthermore, a sandwich ELISA showed significantly (p <0.05) elevated levels of soluble CD97 in serum but not in cerebrospinal fluid of MS patients (37%) compared to healthy controls (8%).Collectively, these data suggest that CD97-CD55 interactions are involved in the inflammatory processes in MS. CD55, which is expressed in lesions by vessels to protect against complement-mediated damage, might bind to CD97 on infiltrating leukocytes. This interaction may facilitate cell activation and migration through the blood-brain barrier. In addition, CD97-CD55 interactions in the parenchyma of the brain may contribute to the inflammatio