RESUMEN
Phosphosugars, such as mannose-6-phosphate (M6P), have been shown previously to display anti-inflammatory properties, notably inhibition of experimental autoimmune encephalomyelitis (EAE) and adjuvant-induced arthritis in rats. It has been proposed that M6P exerts its anti-inflammatory effect by displacing lysosomal enzymes, which are involved in T-cell extravasation into inflammatory sites, from the 300 kDa mannose-6- phosphate receptor (MPR-300) on the surface of T cells. If this model is correct MPR-300 should be selectively expressed on the surface of activated T cells, as T cell entry into the central nervous system in EAE depends on the T cells being in an activated state. Thus, the present study examines whether cell surface expression of MPR-300 by T lymphocytes correlates with their state of activation and whether T cells in inflammatory sites express the receptor. Flow cytometric studies showed MPR-300 to be absent from the surface of unstimulated rat T cells isolated from peripheral blood and lymphoid tissues, and T cells resident within the peritoneal cavity. In contrast, MPR-300 was expressed on activated T cells derived from an inflammatory peritoneal exudate. In vitro studies demonstrated transient expression of MPR-300 on the surface of splenic T cells following stimulation with Con A. MPR-300 was also induced on T-cell lines by antigen stimulation. These data demonstrate that T cells in inflammatory sites express MPR-300 on their surface and activation of T lymphocytes induces cell surface expression of MPR-300. Such findings are consistent with the hypothesis that cell surface MPR-300 is required for the entry of T cells into inflammatory sites.
Asunto(s)
Activación de Linfocitos , Receptor IGF Tipo 2/metabolismo , Linfocitos T/metabolismo , Animales , Líquido Ascítico/química , Líquido Ascítico/citología , Concanavalina A/farmacología , Femenino , Citometría de Flujo , Ratones , Ratones Endogámicos BALB C , Ratas , Ratas Endogámicas F344 , Ratas Endogámicas Lew , Receptor IGF Tipo 2/química , Linfocitos T/efectos de los fármacos , Linfocitos T/inmunología , Tioglicolatos/farmacologíaRESUMEN
NO and IFN-gamma have normally been considered cytotoxic and proinflammatory molecules, respectively, in the setting of the central nervous system inflammatory disease autoimmune encephalomyelitis (EAE). Using mice lacking the ligand binding chain of the IFN-gamma receptor (IFNgammaR-/-), we have previously shown that IFN-gamma is not essential for myelin oligodendrocyte glycoprotein peptide (MOG35-55) induced EAE expression but is in fact essential for its down-regulation. Here we examined the downstream molecular and cellular mechanism(s) of IFN-gamma regulation and demonstrate that neither IL-4 nor IL-10 appear to play a role in down-regulation nor do various lymphoid cell populations. Cells of the macrophage lineage are key to down-regulation as evidenced by the fact that peritoneal exudate cells from IFNgammaR+/+ mice inhibit Ag-driven proliferation of IFNgammaR-/- lymphocytes, whereas IFNgammaR-/- peritoneal exudate cells do not. High levels of reactive nitrogen intermediates are detected in the former cultures but not the latter, and the inhibition of proliferation is reversible with an inhibitor of inducible NO synthase, indicating a key role for NO in down-regulation. Studies with bone marrow chimeras indicate that down-regulation occurs not only systemically but also within the target tissue. These data suggest that IFN-gamma down-regulates EAE by inducing inducible NO synthase and subsequently NO production, both by macrophages in the periphery and, by inference, microglia and astrocytes in the target tissue.
Asunto(s)
Encefalomielitis Autoinmune Experimental/inmunología , Encefalomielitis Autoinmune Experimental/metabolismo , Interferón gamma/fisiología , Óxido Nítrico/fisiología , Secuencia de Aminoácidos , Animales , Líquido Ascítico/citología , Líquido Ascítico/inmunología , Linfocitos T CD8-positivos/inmunología , Células Cultivadas , Regulación hacia Abajo/genética , Regulación hacia Abajo/inmunología , Encefalomielitis Autoinmune Experimental/genética , Inhibidores Enzimáticos/farmacología , Femenino , Interleucina-10/fisiología , Interleucina-4/fisiología , Activación de Linfocitos/inmunología , Depleción Linfocítica , Transfusión de Linfocitos , Masculino , Ratones , Ratones Noqueados , Datos de Secuencia Molecular , Glicoproteína Asociada a Mielina/antagonistas & inhibidores , Glicoproteína Asociada a Mielina/inmunología , Óxido Nítrico Sintasa/antagonistas & inhibidores , Óxido Nítrico Sintasa de Tipo II , Especificidad de Órganos/genética , Especificidad de Órganos/inmunología , Quimera por Radiación/inmunología , Receptores de Interferón/biosíntesis , Receptores de Interferón/genética , Bazo/trasplante , Receptor de Interferón gammaRESUMEN
Nitric oxide was first described being produced in inflammatory cells involved in experimental autoimmune encephalomyelitis in 1992. Since then some 45 papers have appeared examining the role of NO in this central nervous system autoimmune inflammatory disease. Of the first 10 papers published all resulted in the interpretation that NO was a pathologic or "bad" molecule in the context of EAE. A few papers then began to appear suggesting that NO may not in fact always be a harmful molecule and by the end of 1997 early 1998, 22 papers suggested a destructive role for the molecule while three suggested it was protective. The past two years have seen a significant increase in reports supporting a protective mechanism for NO in EAE such that as of July 1999, 27 papers suggest a destructive and 15 a protective role for NO with a few uncommitted. This review sets out in a more or less chronological order the studies examining the role of NO in EAE and maps our changing understanding of the molecules role in this CNS inflammatory disease and by inference perhaps multiple sclerosis.
Asunto(s)
Encefalomielitis Autoinmune Experimental/metabolismo , Óxido Nítrico/fisiología , Animales , Sistema Nervioso Central/enzimología , Sistema Nervioso Central/metabolismo , Encefalomielitis Autoinmune Experimental/enzimología , Humanos , Ratones , Ratones Transgénicos , Óxido Nítrico/biosíntesis , Óxido Nítrico Sintasa/antagonistas & inhibidores , Óxido Nítrico Sintasa/biosíntesis , RatasRESUMEN
Emerging data suggest that polymorphonuclear leukocytes (PMNLs) can play an important role in Ag-dependent immune responses. Therefore, we have assessed the involvement of these cells in the development of an organ-specific autoimmune disease, experimental autoimmune encephalomyelitis (EAE), in the mouse. Depletion of peripheral blood PMNLs beginning day 8 after immunization significantly delayed and in some cases totally prevented the development of clinical EAE in mice. Depletion of PMNLs beginning 1 day before sensitization and continuing until day 7 postimmunization had no effect on the subsequent development of EAE, suggesting that depletion alters the efferent but not the afferent arm of the immune response. In vitro studies showed that lymphoid cells from mice protected from EAE by PMNL depletion beginning on day 8 postsensitization proliferated in response to specific Ag to a level equal to cells from sensitized animals treated with control serum, again indicating that treatment was not affecting the afferent limb of the immune response. Further evidence that PMNL may be necessary in initiating the pathology of EAE was seen in passive transfer experiments where PMNL-depleted recipients of MBP-specific lymphoid effector cells developed EAE much less effectively than did animals treated with control Ab. Taken together, these data indicate that PMNLs play a critical role in the effector phase of the development of the clinicopathologic expression of EAE in mice.
Asunto(s)
Autoanticuerpos/uso terapéutico , Encefalomielitis Autoinmune Experimental/inmunología , Encefalomielitis Autoinmune Experimental/prevención & control , Neutrófilos/inmunología , Animales , Movimiento Celular/inmunología , Modelos Animales de Enfermedad , Encefalomielitis Autoinmune Experimental/etiología , Femenino , Leucocitos Mononucleares/inmunología , Leucopenia/etiología , Leucopenia/inmunología , Ratones , Ratones Endogámicos , Médula Espinal/patologíaRESUMEN
Rat strains vary in their susceptibility to experimental autoimmune encephalomyelitis (EAE) and in many cases, factors other than MHC antigens are thought to play a role in this. We found that PVG rats, which have a very low susceptibility to EAE, were rendered highly susceptible to clinical disease when treated with N-methylarginine (NMA) an inhibitor of nitric oxide synthase (NOS). The clinical course of the ensuing disease in NMA-treated PVG rats was in most cases fulminating in nature and accompanied by some mortality. Following immunisation with myelin basic protein (MBP)-complete Freund's adjuvant (CFA), PVG rats developed higher serum levels of the surrogate markers of nitric oxide production, reactive nitrogen intermediates (RNI; nitrite and nitrate), than did their Lewis counterparts. This in vivo finding was reflected in vitro, where the levels of RNI produced in 24, 48 and 72 h IFN-gamma-stimulated spleen cell cultures for PVG rats were significantly higher than those for Lewis rats. A mechanism by which increased NO production might protect PVG rats against clinical EAE was suggested by the finding that lymph node cells, isolated from NMA-treated MBP-immunised PVG rats, proliferated in response to MBP at a rate approximately 3 x greater than those from MBP-immunised, saline treated rats. Thus, the greater number of MBP-specific T cells generated in the NOS inhibitor-treated vs. untreated rats could account for their increased susceptibility to developing clinical EAE. The findings in this study suggest that NO plays a role in protecting PVG rats against developing EAE.
Asunto(s)
Encefalomielitis Autoinmune Experimental/genética , Encefalomielitis Autoinmune Experimental/fisiopatología , Óxido Nítrico/fisiología , Ratas Mutantes/genética , Ratas Mutantes/fisiología , Animales , División Celular/efectos de los fármacos , Células Cultivadas , Encefalomielitis Autoinmune Experimental/sangre , Inhibidores Enzimáticos/farmacología , Predisposición Genética a la Enfermedad , Interferón gamma/farmacología , Ganglios Linfáticos/citología , Ganglios Linfáticos/efectos de los fármacos , Proteína Básica de Mielina/farmacología , Nitratos/sangre , Óxido Nítrico/antagonistas & inhibidores , Óxido Nítrico Sintasa/antagonistas & inhibidores , Nitritos/sangre , Ratas , Ratas Endogámicas Lew , Bazo/citología , Bazo/efectos de los fármacos , Bazo/metabolismo , omega-N-Metilarginina/farmacologíaRESUMEN
The ultimate aim in the treatment of autoimmune disease is to restore self-tolerance to the autoantigen(s) in question. In lieu of this ideal result, the conversion of a destructive or pathogenic autoimmune response into one of benign autoimmunity would also be highly desirable. In either case the use of the antigenic epitope, which is the target of the destructive immune response, would ideally be employed so as to give specificity to the protection without the need for long-term immunosuppression. This review describes a number of different approaches using various forms, doses, and routes of injection of specific neuroantigen to inhibit the different clinical varieties of autoimmune encephalomyelitis in a number of animal models; all done with the view to translating the findings into the clinic for the treatment of multiple sclerosis. We conclude that any treatment strategy for multiple sclerosis (MS) must have a number of features: it must be clinically acceptable, specific, long-lasting, require only short-term treatment, able to shunt off ongoing disease, and have the potential to prevent or deal with epitope spreading. Few of the approaches we describe fulfill all of these criteria. We suggest that investigations of new adjunctive agents to be used with a specific antigen be pursued, and that currently the use of chimeric proteins or DNA vaccination with or without the new adjunctives may hold the most hope for the future.
Asunto(s)
Autoantígenos/uso terapéutico , Enfermedades Autoinmunes/terapia , Enfermedades del Sistema Nervioso Central/terapia , Adyuvantes Inmunológicos , Animales , Encefalomielitis Autoinmune Experimental/terapia , Humanos , Inmunidad Innata , Inmunoterapia , Esclerosis Múltiple/terapiaRESUMEN
Currently available anti-inflammatory drugs for the treatment of multiple sclerosis (MS) and other inflammatory diseases are generally inadequate, with disease progression not being arrested by the treatments and undesirable side effects posing problems. In response to these deficiencies our laboratories have, over the past 10 years, been developing novel drugs that interfere with the entry of leucocytes into inflammatory sites by inhibiting their passage through the subendothelial basement membrane (BM). This review initially summarizes evidence supporting the hypothesis that the subendothelial BM is a major barrier to the accumulation of leucocytes in inflammatory sites. An important point that has emerged is that breaching of the BM is probably a cooperative process, involving activation- and cytokine-induced degradative enzymes contributed by leucocytes, endothelial cells and platelets. The review then discusses the properties of three separate classes of anti-inflammatory compounds we have developed, namely sulfated polysaccharides/oligosaccharides, phosphosugars, and castanospermine (CS), which inhibit the passage of leukocytes through BM. Each drug type appears to prevent BM degradation by a different mechanism. Sulfated polysaccharides/oligosaccharides mediate their anti-inflammatory effect by inhibiting the endoglycosidase, heparanase, which plays a key role in the solubilization of BM by invading leucocytes. In fact, our studies have highlighted the heparanase enzyme as a major target for future drug development. Phosphosugars probably inhibit inflammation by displacing lysosomal enzymes, which are involved in BM degradation, from cell surface mannose 6-phosphate receptors. This mechanism of expressing degradative enzymes on the cell surface is particularly evident with activated T lymphocytes. On the other hand, CS interferes with appropriate targeting of lysosomal enzymes involved in BM degradation. For reasons which are still unclear, CS specifically inhibits BM degradation by endothelial cells, which results in a characteristic perivascular arrest of leucocytes in inflammatory sites. Overall, our studies have established that inhibitors of subendothelial BM degradation represent viable anti-inflammatory agents. It is hoped that future work will result in the development of a totally new class of highly effective, subtle and non-toxic anti-inflammatory drugs for the treatment of MS and other inflammatory diseases.
Asunto(s)
Antiinflamatorios/uso terapéutico , Enfermedades del Sistema Nervioso Central/terapia , Animales , Autoantígenos/inmunología , Membrana Basal/fisiología , Movimiento Celular , Enfermedades del Sistema Nervioso Central/inmunología , Humanos , Inflamación , Leucocitos/fisiología , Linfocitos T/inmunologíaRESUMEN
Protection against experimental autoimmune encephalomyelitis (EAE) induced by s.c. infusion of myelin basic protein (MBP) alone is dose dependent and long lived. Protection is not effective against passively induced disease nor is it transferable with lymphoid cells. The proliferative response of lymph node cells to MBP following encephalitogenic challenge is decreased in the EAE-protected animals as is the production of IL-2 and IFN-gamma by these cells. Treatment with soluble MBP promed rats for antibody production is evidenced by the early appearance of anti-MBP antibody following encephalitogenic challenge. Determination of antibody isotype following challenge revealed a change in the ratio of IgG1 to IgG2a with a significant increase in the amount of IgG1 produced. These data suggest that infusion of high dose soluble neuroantigen primes the immune response such that subsequent challenge with an encephalitogenic inoculum pushes the response down a non-destructive Th2 autoimmune pathway.
Asunto(s)
Encefalomielitis Autoinmune Experimental/prevención & control , Proteínas Fimbrias , Proteína Básica de Mielina/farmacología , Animales , Formación de Anticuerpos/inmunología , Formación de Anticuerpos/fisiología , Antígenos Bacterianos/metabolismo , Antígenos Bacterianos/farmacología , Proteínas Bacterianas/metabolismo , Proteínas Bacterianas/farmacología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/fisiología , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/fisiología , Relación Dosis-Respuesta Inmunológica , Femenino , Tolerancia Inmunológica , Inmunización Pasiva , Inyecciones Subcutáneas , Interferón gamma/metabolismo , Interleucina-2/metabolismo , Ganglios Linfáticos/inmunología , Ganglios Linfáticos/fisiología , Proteína Básica de Mielina/inmunología , Ratas , Ratas Endogámicas Lew , Solubilidad , Bazo/citología , Bazo/efectos de los fármacos , Bazo/inmunología , Linfocitos T/inmunología , Linfocitos T/fisiologíaRESUMEN
Resistance to autoimmune encephalomyelitis was induced by s.c. infusion of myelin basic protein (MBP) in saline in combination with i.p. injections of anti-CD11a (LFA-1) mAbs. This treatment induces resistance to EAE induction, which appears early and persists for at least one month after treatment. Some MBP-CFA-challenged resistant rats showed minimal inflammation in the central nervous system, which was, however, confined to the meninges of the lower spinal cord. Examination of the immune status of MBP-anti-LFA-1 treated rats before encephalitogenic challenge failed to reveal any priming when assessed by Ag driven proliferation and cytokine production by lymphoid cells, and by circulating Ab production. Following challenge of protected rats, lymph node cell proliferation to MBP was unaltered, indicating that reactive cells had not been deleted or energized. Resistance could not be transferred with lymphoid cells from treated rats nor abrogated by cyclophosphamide treatment. In treated rats following challenge, there was a shift in the isotype of anti-MBP Ab produced, from an IgG2a:IgG1 ratio of 2:1 to 1:1, due to an increase in IgG1 production, indicating a possible bias towards a nonpathogenic Th2 CD4+ T cell response. The IgG1 Ab was detected early after challenge suggesting that pretreatment had indeed primed the animals, and had primed them to go down the Th2 pathway following encephalitogenic challenge. The ability to divert immune reactivity from a destructive to a nondestructive response could have important therapeutic implications for autoimmune disease.