RESUMO
BACKGROUND: Imatinib, a tyrosine kinase inhibitor, has been shown to restore blood-brain barrier integrity and reduce infarct size, haemorrhagic transformation and cerebral oedema in stroke models treated with tissue plasminogen activator. We evaluated the safety of imatinib, based on clinical and neuroradiological data, and its potential influence on neurological and functional outcomes. METHODS: A phase II randomized trial was performed in patients with acute ischaemic stroke treated with intravenous thrombolysis. A total of 60 patients were randomly assigned to four groups [3 (active): 1 (control)]; the active treatment groups received oral imatinib for 6 days at three dose levels (400, 600 and 800 mg). Primary outcome was any adverse event; secondary outcomes were haemorrhagic transformation, cerebral oedema, neurological severity on the National Institutes of Health Stroke Scale (NIHSS) at 7 days and at 3 months and functional outcomes on the modified Rankin scale (mRS). RESULTS: Four serious adverse events were reported, which resulted in three deaths (one in the control group and two in the 400-mg dose group; one patient in the latter group did not receive active treatment and the other received two doses). Nonserious adverse events were mostly mild, resulting in full recovery. Imatinib ameliorated neurological outcomes with an improvement of 0.6 NIHSS points per 100 mg imatinib (P = 0.02). For the 800-mg group, the mean unadjusted and adjusted NIHSS improvements were 4 (P = 0.037) and 5 points (P = 0.012), respectively, versus controls. Functional independence (mRS 0-2) increased by 18% versus controls (61 vs. 79; P = 0.296). CONCLUSION: This phase II study showed that imatinib is safe and tolerable and may reduce neurological disability in patients treated with intravenous thrombolysis after ischaemic stroke. A confirmatory randomized trial is currently underway.
Assuntos
Isquemia Encefálica/tratamento farmacológico , Mesilato de Imatinib/uso terapêutico , Proteínas Tirosina Quinases/antagonistas & inibidores , Acidente Vascular Cerebral/tratamento farmacológico , Terapia Trombolítica , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Isquemia Encefálica/mortalidade , Esquema de Medicação , Feminino , Humanos , Mesilato de Imatinib/administração & dosagem , Mesilato de Imatinib/efeitos adversos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Acidente Vascular Cerebral/mortalidade , Resultado do Tratamento , Adulto JovemRESUMO
Multiple sclerosis (MS) is a disease with unknown cause characterized by inflammation and demyelination in the central nervous system. Although an autoimmune pathogenesis has been suggested, there are no conclusive data on the number of T cells autoreactive with myelin antigens in MS compared to controls. We showed that T lymphocytes secreting interferon-gamma in response to possible target autoantigens are severalfold more common among PBL mononuclear cells in patients with MS than in patients with aseptic meningitis and tension headache. On average T cells reactive with myelin basic protein (MBP), two different MBP peptides, or with proteolipid protein amounted to 2.7-5.2/10(5) PBL from MS patients. MBP-reactive T cells were still more frequent among mononuclear cells isolated from the cerebrospinal fluid (CSF; 185/10(5) CSF cells). We concluded that T cells reactive with myelin autoantigens are strongly increased in MS. This approach to detect them could allow definition of immunodominant T cell epitopes in individual MS patients, and thereby enable further development towards specific immunotherapy.
Assuntos
Doenças Autoimunes/imunologia , Interferon gama/biossíntese , Esclerose Múltipla/imunologia , Linfócitos T/imunologia , Adulto , Idoso , Autoantígenos/imunologia , Líquido Cefalorraquidiano/citologia , Humanos , Pessoa de Meia-Idade , Proteína Básica da Mielina/imunologia , Proteínas da Mielina/imunologia , Proteína Proteolipídica de MielinaRESUMO
(1) Nicotinamide nucleotide transhydrogenases in submitochondrial particles from beef heart mitochondria, chromatophores from Rhodospirillum rubrum and membrane preparations from Escherichia coli and Pseudomonas aeruginosa have been compared with respect to the following properties: stereospecificity for the 4-hydrogen of NADH, reactivity with 3'-NADP, inhibition by palmityl-CoA, sensitivity tot rypsin, and effects of Ca2+ and 2'-AMP on the reaction rates. (2) Transhydrogenases from submitochondrial particles, R. rubrum chromatophores and E. coli membrane preparations have A-side stereospecificity for NADH, do not react with 3'-NADP, are inhibited by palmityl-CoA and are extremely sensitive to trypsin treatment. No effects of Ca2+ or 2'-AMP on the reaction rates were observed. In R. rubrum chromatophores trypsin-sensitive sites are present both in the soluble transhydrogenase factor and in the membrane preparation devoid of transhydrogenase factor. (3) In contrast, P. aeruginosa transhydrogenase is allosterically regulated by Ca2+ and 2'-AMP, is reactive with 3'-NADP, has B-side stereospecificity for NADH and is insensitive to palmityl-CoA or trypsin treatment. (4) It is concluded that the properties characterizing the transhydrogenase in E. coli, R. rubrum chromatophores and submitochondrial particles are closely connected with the interaction of the enzyme with an energy-conserving membrane system.
Assuntos
Proteínas de Bactérias/metabolismo , Mitocôndrias Cardíacas/enzimologia , NADP Trans-Hidrogenases/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Cálcio/metabolismo , Bovinos , Mitocôndrias Cardíacas/ultraestrutura , NAD/metabolismo , NADP/metabolismo , Palmitoil Coenzima A/metabolismo , Rhodospirillum/metabolismo , Estereoisomerismo , Tripsina/metabolismoRESUMO
Glucose-induced insulin release is decreased in the spontaneously diabetic GK rat, a nonobese rodent model of type 2 diabetes. Forskolin restores the impaired insulin release in both the isolated perfused pancreas and isolated islets from these rats (Abdel-Halim et al., Diabetes 45:934-940, 1996). We demonstrate here that the insulinotropic effect of forskolin in the GK rat is due to increased generation of cAMP and that it is associated with overexpression of adenylyl cyclase (AC)-III mRNA and gene mutations. The AC-III mRNA overexpression was demonstrated by in situ hybridization using oligonucleotide probes binding to different regions of the rat AC-III mRNA. It was associated with the presence of two point mutations identified at positions -28 bp (A --> G) and -358 bp (A --> C) of the promoter region of the AC-III gene and was demonstrable in both GK rat islets and peripheral blood cells. Transfection of COS cells with a luciferase reporter gene system revealed up to 25-fold increased promoter activity of GK AC-III promoter when compared with normal rat promoter (P < 0.0001). In conclusion, forskolin restores the impaired insulin release in islets of the GK rat through enhanced cAMP generation. This is linked to overexpression of AC-III mRNA in GK islets due to two functional point mutations in the promoter region of the AC-III gene.
Assuntos
Adenilil Ciclases/genética , AMP Cíclico/biossíntese , Diabetes Mellitus Tipo 2/genética , Regulação Enzimológica da Expressão Gênica/genética , Insulina/metabolismo , Ilhotas Pancreáticas/química , Ilhotas Pancreáticas/metabolismo , Mutação/genética , Regiões Promotoras Genéticas/genética , Animais , Sequência de Bases , Estudos de Coortes , Primers do DNA/química , Diabetes Mellitus Tipo 2/enzimologia , Diabetes Mellitus Tipo 2/patologia , Modelos Animais de Doenças , Hibridização In Situ , Técnicas In Vitro , Secreção de Insulina , Ilhotas Pancreáticas/patologia , Masculino , Reação em Cadeia da Polimerase , RNA Mensageiro/análise , RNA Mensageiro/genética , Ratos , Ratos Endogâmicos , Ratos WistarRESUMO
A Trypanosoma brucei brucei-derived lymphocyte triggering factor (TLTF) induced CD8+ T cells to produce IFN-gamma, which in turn stimulates parasite growth. This parasite-host interaction was studied in mouse strains that are either relatively susceptible (C3H/He) or resistant (C57Bl/6J) to infection, as well as in athymic nude mice. In all mouse strains, T. b. brucei infection caused a strong induction of IFN-gamma production by spleen mononuclear cells (MNC). In vivo blocking of IFN-gamma by intraperitoneal injection of mouse monoclonal anti-IFN-gamma antibody suppressed parasite growth and increased survival of both C3H/H3 and C57Bl/6J animals, suggesting that, irrespective of strain-related disease susceptibility, IFN-gamma is a growth-promoting stimulus for T. b. brucei. Spleen MNC from noninfected mice of all strains were in vitro like-wise strongly induced to IFN-gamma production when exposed to TLTF. This suggests that CD8+ expressing T cell receptor (TCR) alpha/beta, gamma/delta-bearing T cells and NK cells may all be triggered to IFN-gamma production by TLTF. In all mouse strains, TLTF also caused an increase in the number of cells expressing mRNA for TGF-beta in vitro. However, significant triggering to IL-4 mRNA expression only occurred in the relatively disease-resistant C57Bl/6J strain. As IL-4 is required for the synthesis and class switches of immunoglobulins, which are essential host immune defenses against T. b. brucei, the degree of resistance may be related to inherent strain ability to produce IL-4 in response to TLTF.
Assuntos
Fatores Biológicos/farmacologia , Indutores de Interferon/farmacologia , Interferon gama/biossíntese , Interleucina-4/biossíntese , Fator de Crescimento Transformador beta/biossíntese , Trypanosoma brucei brucei/fisiologia , Animais , Suscetibilidade a Doenças , Feminino , Interações Hospedeiro-Parasita , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Camundongos Nus , Especificidade da EspécieRESUMO
An autoimmune response to myelin basic protein (MBP) has been proposed to participate in the development of the chronic neurologic manifestations that may accompany Borrelia burgdorferi-induced Lyme disease. Using an immunospot assay, we counted cells secreting antibodies to MBP. Anti-MBP IgG antibody-secreting cells were detected in CSF from eight of 13 consecutive patients with Lyme neuroborreliosis irrespective of stage of disease. The numbers were between 1/370 and 1/5,000 CSF cells (mean, 1/1,250 in the 13 patients). The highest numbers were encountered in two patients with severe signs of CNS involvement. The numbers decreased in parallel with clinical improvement after treatment. Anti-MBP IgG antibody-secreting cells were also observed in the CSF from patients with a variety of other inflammatory diseases of the nervous system, and their role in the development of tissue damage remains unsettled. Anti-MBP IgG antibody-secreting cells were not detected in the patients' blood, reflecting accumulation of this autoantibody response to CSF.
Assuntos
Células Produtoras de Anticorpos/citologia , Doença de Lyme/imunologia , Proteína Básica da Mielina/imunologia , Doenças do Sistema Nervoso/imunologia , Adulto , Idoso , Humanos , Imunoglobulina G/líquido cefalorraquidiano , Doença de Lyme/líquido cefalorraquidiano , Doença de Lyme/patologia , Pessoa de Meia-Idade , Doenças do Sistema Nervoso/líquido cefalorraquidiano , Doenças do Sistema Nervoso/patologiaRESUMO
Migration properties and occurrence of antibodies against myelin basic protein (MBP) in paired CSF and serum specimens from patients with multiple sclerosis (MS) were demonstrated after agarose isoelectric focusing, immunoblot transfer, and immunoperoxidase staining. Oligoclonal IgG antibody bands directed against MBP were found in the CSF of 9 of 28 patients with MS (32%), but not in the CSF of any of 34 patients with other neurologic diseases. No serum showed anti-MBP antibody bands. The CSF anti-MBP antibodies migrated to the anodal region of the IgG area in a different fashion from oligoclonal IgG and anti-measles IgG antibodies, which were detected in parallel. The anti-MBP bands were transient in three of seven patients whom we studied consecutively. Enzyme-linked immunosorbent assay (ELISA) of serum and CSF for detection of IgG reactivity against MBP showed absorbance values above 2 standard deviations of controls in 44% of the MS patients and in 21% of those with other neurologic diseases. Results of this assay correlated partly with those of the immunoblot assay. ELISA positive and immunoblot negative results might be due to a broad polyclonal anti-MBP antibody response.
Assuntos
Anticorpos/líquido cefalorraquidiano , Imunoglobulina G/líquido cefalorraquidiano , Esclerose Múltipla/imunologia , Proteína Básica da Mielina/líquido cefalorraquidiano , Adulto , Eletroforese em Gel de Ágar , Feminino , Humanos , Técnicas Imunológicas , Focalização Isoelétrica , Masculino , Pessoa de Meia-Idade , Proteína Básica da Mielina/imunologia , Doenças do Sistema Nervoso/imunologiaRESUMO
T cells recognizing the myelin components myelin basic protein (MBP) and proteolipid protein (PLP) are increased in multiple sclerosis (MS), and there are elevated numbers of T cells recognizing the nicotinic acetylcholine receptor (AChR) in myasthenia gravis (MG). However, the cytokine repertoires in these diseases are largely unknown. We adopted in situ hybridization with radiolabeled complementary DNA oligonucleotide probes to enumerate mononuclear cells that expressed the T-helper type 1 (Th1) cell-related interferon-gamma (IFN-gamma) and Th2-associated interleukin-4 (IL-4) after short-term culture in the presence of autoantigen. High numbers of IFN-gamma and IL-4 mRNA-expressing cells in response to MBP and PLP were detected in patients with untreated MS, and to AChR in MG. The levels of IFN-gamma and IL-4 mRNA-positive cells in MS after culture in the presence of AChR, and in MG after culture in the presence of MBP or PLP, did not differ from those detected after culture without antigen. The CSF of MS patients contained four- to eightfold more myelin protein-reactive IFN-gamma and IL-4 expressing cells. The findings imply that MS and MG are associated with mixed Th1- and Th2-like cell responses directed to organ-specific target antigens.
Assuntos
Expressão Gênica , Interferon gama/genética , Interleucina-4/genética , Esclerose Múltipla/genética , Miastenia Gravis/genética , Adulto , Autoantígenos/imunologia , Autoantígenos/fisiologia , Epitopos , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Monócitos/metabolismo , Esclerose Múltipla/imunologia , Miastenia Gravis/imunologia , Proteína Básica da Mielina/imunologia , Proteínas da Mielina/imunologia , Proteína Proteolipídica de Mielina , RNA Mensageiro/metabolismo , Receptores Colinérgicos/fisiologiaRESUMO
The kinetics of mRNA expression in the central nervous system (CNS) for a series of putatively disease-promoting and disease-limiting cytokines during the course of experimental autoimmune encephalomyelitis (EAE) in Lewis rats were studied. Cytokine mRNA-expressing cells were detected in cryosections of spinal cords using in situ hybridization technique with synthetic oligonucleotide probes. Three stages of cytokine mRNA expression could be distinguished: (i) interleukin (IL)-12, tumor necrosis factor (TNF)-beta (= lymphotoxin-alpha) and cytolysin appeared early and before onset of clinical signs of EAE; (ii) TNF-alpha peaked at height of clinical signs of EAE; (iii) IL-10 appeared increasingly at and after clinical recovery. The early expression of IL-12 prior to the expression of interferon-gamma (IFN-gamma) mRNA shown previously is consistent with a role of IL-12 in promoting proliferation and activation of T helper 1 (Th1) type cells producing IFN-gamma. The TNF-beta mRNA expression prior to onset of clinical signs favours a role for this cytokine in disease initiation. A pathogenic effector role of TNF-alpha was suggested from these observations that TNF-alpha mRNA expression roughly paralleled the clinical signs of EAE. This may be the case also for cytolysin. IL-10-expressing cells gradually increased to high levels in the recovery phase of EAE, consistent with a function in down-regulating the CNS inflammation. From these data we conclude that there is an ordered appearance of putative disease-promoting and -limiting cytokines in the CNS during acute monophasic EAE.
Assuntos
Encefalomielite Autoimune Experimental/genética , Interleucina-10/genética , Interleucina-12/genética , Linfotoxina-alfa/genética , Glicoproteínas de Membrana/genética , Medula Espinal/metabolismo , Fator de Necrose Tumoral alfa/genética , Animais , Expressão Gênica , Hibridização In Situ , Masculino , Perforina , Proteínas Citotóxicas Formadoras de Poros , RNA Mensageiro/genética , Ratos , Ratos Endogâmicos LewRESUMO
An immunospot assay that detects single secretory cells was used to enumerate interferon-gamma secreting cells (IFN-gamma-sc) in mononuclear cell suspensions from the central nervous system (CNS) and peripheral lymphoid organs after actively induced experimental allergic encephalomyelitis (EAE) in Lewis rats. In the CNS compartment there was a significant increase in the number of IFN-gamma-sc preceding the onset of the clinical signs of EAE. Both in rats with EAE and rats immunized with Freund's complete adjuvant (FCA) the number of IFN-gamma-sc increased in peripheral lymphoid organs, as compared to non-immunized controls. In view of the potent immunoregulatory effects of IFN-gamma, its intra-CNS secretion may play a crucial role for clinicopathological events in EAE. To study the numbers of primed T cells that in response to myelin antigens produced IFN-gamma, mononuclear cell suspensions from peripheral lymphoid organs were precultured to allow for antigen uptake, presentation and T cell triggering, followed by enumeration of IFN-gamma-sc. T cells responding to a peptide of myelin basic protein (MBP) that previously have been shown encephalitogenic in Lewis rats, appeared initially and were quantitatively dominant over the course of EAE. Later, T cell reactivities to multiple regions of MBP appeared, showing that the concept of immunodominance in EAE is non-absolute and time dependent. Splenocyte cultures from EAE rats exposed to the different antigens showed a reduced number of IFN-gamma-sc compared to cultures not exposed to antigen, suggesting an antigen-induced suppression of T cell effector molecules.
Assuntos
Encefalomielite Autoimune Experimental/imunologia , Imunidade , Interferon gama/metabolismo , Linfócitos T/imunologia , Animais , Antígenos/imunologia , Encefalomielite Autoimune Experimental/metabolismo , Encefalomielite Autoimune Experimental/patologia , Linfonodos/imunologia , Linfonodos/metabolismo , Linfonodos/patologia , Proteína Básica da Mielina/farmacologia , Fragmentos de Peptídeos/farmacologia , Ratos , Ratos Endogâmicos Lew , Baço/imunologia , Baço/metabolismo , Baço/patologia , Distribuição TecidualRESUMO
Studies on patients with monosymptomatic optic neuritis (ON) should give opportunities to identify features typical for early multiple sclerosis (MS). There are increased T and B cell responses to the myelin components myelin basic protein (MBP) and proteolipid protein (PLP) in both ON and MS, but there is little information on the types of cytokines produced by such cells. We describe the use of in situ hybridization with complementary DNA oligonucleotide probes to detect and enumerate mononuclear cells expressing mRNA for the cytokines interferon-gamma (IFN-gamma) which augments cell-mediated immunity; interleukin-4 (IL-4) which promotes the B cell response; and transforming growth factor beta (TGF-beta) that in many cases downregulates immune responses. Expression of these cytokines was studied in mononuclear cells from peripheral blood and cerebrospinal fluid (CSF) from patients with ON and MS after in vitro exposure to MBP and PLP, and in absence of antigen. There were elevated levels of cells that in response to MBP and PLP expressed IFN-gamma, IL-4 and TGF-beta mRNA in blood and further enriched in CSF in both ON and MS, compared to patients with other neurological diseases. The results suggest that IFN-gamma, IL-4 as well as TGF-beta are involved in both ON and MS, and that the cytokine profile in early MS as reflected by ON is not different from that in clinically definite MS.
Assuntos
Interferon gama/biossíntese , Interleucina-4/biossíntese , Proteína Básica da Mielina/farmacologia , Proteínas da Mielina/farmacologia , Neurite Óptica/metabolismo , Fator de Crescimento Transformador beta/biossíntese , Adulto , Feminino , Humanos , Interferon gama/genética , Interleucina-4/genética , Masculino , Pessoa de Meia-Idade , Esclerose Múltipla/metabolismo , Proteína Proteolipídica de Mielina , RNA Mensageiro/análise , Fator de Crescimento Transformador beta/genéticaRESUMO
Antibodies to myelin components, such as myelin basic protein (MBP), may play a role in pathogenesis of multiple sclerosis (MS) but results from determinations of anti-MBP antibodies are inconsistent. Enumeration of cells secreting antibodies represents a new approach to evaluate a specific antibody response regarding extent and localization, and reduces effects of e.g. antibody binding to target. Anti-MBP IgG antibody secreting cells were present in MS patients' cerebrospinal fluid (CSF) at a mean value of 1 per 833 cells, and they amounted to a mean value of about 2454 in the whole CSF compartment. Similar numbers were encountered in patients with other inflammatory neurological diseases (OIND). During follow-up, anti-MBP IgG antibody secreting cells persisted regarding frequency and numbers in MS, but decreased in OIND. Such cells were rarely detected in patients with tension headache. No correlations to clinical exacerbation of MS, disability or duration were discernable. In blood from MS and OIND patients, anti-MBP IgG antibody secreting cells were detected infrequently and at low numbers. The anti-MBP antibody response is strongly restricted to the IgG isotype. The anti-MBP IgG antibody response which is persistent and compartmentalized to the diseased organ, may be important for the development of MS.
Assuntos
Imunoglobulina G/líquido cefalorraquidiano , Esclerose Múltipla/imunologia , Proteína Básica da Mielina/imunologia , Adulto , Idoso , Células Produtoras de Anticorpos/imunologia , Feminino , Cefaleia/imunologia , Humanos , Masculino , Meningite/imunologia , Pessoa de Meia-Idade , Esclerose Múltipla/líquido cefalorraquidianoRESUMO
Sulfasalazine (SASP; 5-(p-(2-pyridylsulfamoyl)phenylazo)salicyclic acid) has beneficial effects on certain inflammatory diseases and has been proposed for clinical trials in multiple sclerosis (MS). We have explored the effects of SASP on actively induced experimental autoimmune encephalomyelitis (EAE) in Lewis rats. SASP was given orally at three different doses from the day of immunization to day 40 post-immunization (p.i.). All doses led to a clinically more protracted disease, increased numbers of T cells infiltrating into the central nervous system (CNS) and to increased numbers of interferon-gamma-secreting cells (IFN-gamma-sc) in the CNS. The effects of SASP treatment on T cell-mediated autoimmunity against CNS myelin and peptides of myelin basic protein (MBP) were measured by IFN-gamma secretion and proliferation by lymph node mononuclear cells in response to these antigens. In SASP-treated rats, increased numbers of IFN-gamma-sc appeared in response to myelin antigens, while the proliferative responses were decreased. We suggest that monitoring cell-mediated immunity with the IFN-gamma-sc method may be relevant for the evaluation of new immunotherapeutic strategies in inflammatory demyelinating diseases. Furthermore, our results demand caution as to clinical trials with SASP in MS.
Assuntos
Encefalomielite Autoimune Experimental/imunologia , Sulfassalazina/farmacologia , Linfócitos T/efeitos dos fármacos , Animais , Encéfalo/patologia , Antígenos CD4/análise , Antígenos CD8/análise , Encefalomielite Autoimune Experimental/etiologia , Encefalomielite Autoimune Experimental/patologia , Interferon gama/metabolismo , Ativação Linfocitária/efeitos dos fármacos , Ratos , Ratos Endogâmicos Lew , Linfócitos T/imunologiaRESUMO
Using 2- and 3-colour FACS analysis we found increased levels of fetal-type CD5+ B cells and CD4-8- T cells in cerebrospinal fluid (CSF) of patients with multiple sclerosis (MS) and aseptic meningitis (AM) compared to control probands with muscular tension headache (TH). Similar differences were found for CD5+ B cells in peripheral blood, but at lower levels. CD4-8- T cells in blood exceeded those in CSF in all patient groups, with the exception of relapsing remitting MS, revealing the highest values in AM. There was a positive correlation between CD4-8- T cells and T cell receptor (TCR) gamma delta bearing T cells in blood and CSF. The double-negative T cells exceeded the TCR gamma delta T cells by about 1%. A positive correlation between CD5+ B cells and CD4-8- T cell level in CSF was found in MS and AM, but not in TH, nor in blood of any patient group. HLA-DR expression was lower in CD5+ B cells than in CD5- B cells. We conclude that fetal-type lymphocytes are enriched in CSF compartment of patients with inflammatory diseases of the central nervous system, irrespective of autoimmune mechanisms involved, but the function of CD5+ B cells is mainly to produce the autoantibodies.
Assuntos
Antígenos CD/líquido cefalorraquidiano , Linfócitos B , Cefaleia/líquido cefalorraquidiano , Contagem de Leucócitos , Meningite Asséptica/líquido cefalorraquidiano , Doenças Neuromusculares/líquido cefalorraquidiano , Linfócitos T , Adolescente , Adulto , Idoso , Feminino , Cefaleia/imunologia , Cefaleia/patologia , Humanos , Masculino , Meningite Asséptica/imunologia , Meningite Asséptica/patologia , Pessoa de Meia-Idade , Doenças Neuromusculares/imunologia , Doenças Neuromusculares/patologia , Linfócitos T Auxiliares-Indutores , Linfócitos T ReguladoresRESUMO
Experimental autoimmune encephalomyelitis (EAE) in rats is typically a brief and monophasic disease with sparse demyelination. However, inbred DA rats develop a demyelinating, prolonged and relapsing encephalomyelitis after immunization with rat spinal cord in incomplete Freund's adjuvant. This model enables studies of mechanisms related to chronicity and demyelination, two hallmarks of multiple sclerosis (MS). Here we have investigated, in situ, the dynamics of cytokine mRNA expression in the central nervous system (CNS) and peripheral lymphoid organs (lymph node cells and splenocytes) of diseased DA rats. We demonstrate that peripheral lymphoid cells stimulated in vitro with encephalitogenic peptides 69-87 and 87-101 of myelin basic protein responded with high mRNA expression for proinflammatory cytokines; interferon-gamma, interleukin-12 (IL-12), tumour necrosis factors alpha and beta, IL-1 beta and cytolysin. A high expression of mRNA for these proinflammatory cytokines was also observed in the CNS where it was accompanied by classical signs of inflammation such as expression of major histocompatibility complex class I and II, CD4, CD8 and IL-2 receptor. The expression of mRNA for proinflammatory cytokines was remarkably long-lasting in DA rats as compared to LEW rats which display a brief and monophasic EAE. Furthermore, mRNAs for putative immunodownmodulatory cytokines, i.e. transforming growth factor-beta (TGF-beta), IL-10 and IL-4 were almost absent in DA rats, in both the CNS and in vitro stimulated peripheral lymphoid cells, while their levels were elevated in the CNS of LEW rats during the recovery phase. We conclude that the MS-like prolonged and relapsing EAE in DA rats is associated with a prolonged production of proinflammatory cytokines and/or low or absent production of immunodownmodulatory cytokines.
Assuntos
Citocinas/genética , Encefalomielite Autoimune Experimental/imunologia , Interleucina-10/genética , Medula Espinal/metabolismo , Fator de Crescimento Transformador beta/genética , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais , Citocinas/metabolismo , Expressão Gênica , Imuno-Histoquímica , Masculino , Dados de Sequência Molecular , Proteína Básica da Mielina/química , Proteína Básica da Mielina/imunologia , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/imunologia , RNA Mensageiro/genética , Ratos , Ratos Endogâmicos Lew , Ratos Endogâmicos , Fator de Crescimento Transformador beta/metabolismoRESUMO
The increased intrathecal production of immunoglobulins within the cerebrospinal fluid (CSF) compartment commonly observed in multiple sclerosis (MS) implicates participation of B cell activating factors. One effect of the cytokine interleukin (IL)-6 is induction of immunoglobulin production by activated B cells. Employing in situ hybridization (ISH) with synthetic oligonucleotide probes, we measured numbers of IL-6 mRNA-expressing mononuclear cells (MNC) in blood and CSF from patients with MS, aseptic meningo-encephalitis (AM), and in blood from patients with other neurological diseases (OND) and healthy subjects. Numbers of IL-6 mRNA-expressing MNC were elevated in blood (mean frequency 1 per 33,000 MNC) and even further enriched in the CSF (1 per 10,000 MNC) of MS patients, and to a similar extent in AM patients' blood. Cultivation in the presence of myelin basic protein and proteolipid protein revealed strong augmentation of IL-6 mRNA-positive cells in MS but not in OND. The results suggest that IL-6 is one of several cytokines which are upregulated in MS, in particular locally in the CSF. A role of IL-6 in MS, whether disease- promoting or protective, remains unclear.
Assuntos
Interleucina-6/genética , Leucócitos Mononucleares/metabolismo , Esclerose Múltipla/metabolismo , RNA Mensageiro/análise , Adulto , Idoso , Idoso de 80 Anos ou mais , Líquido Cefalorraquidiano/citologia , Líquido Cefalorraquidiano/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
Nervous tissue expression of immunological signal and recognition molecules, as well as lymphoid tissue immune responses after facial nerve trauma was studied in male rats of the Lewis and Brown Norway (BN) strains. In both rat strains nerve transection caused within four days the appearance of IFN-gamma-like immunoreactivity in the cytoplasm of axotomized motor neurons and an induction of MHC class I and II, and CD4 molecules on surrounding glial cells to a similar extent. T lymphocytes also infiltrated the facial nuclei ipsilateral to the axotomy in all animals. The number of autoreactive T cells in superficial cervical lymph nodes, which in response to whole myelin or peptides of myelin basic protein (MBP) secreted IFN-gamma increased markedly after axotomy. This response was more conspicuous in Lewis rats, which are susceptible to experimental allergic encephalomyelitis (EAE), than in BN rats, which are EAE resistant. A proportion of the axotomized Lewis rats also developed widespread perivascular infiltration of mononuclear cells in the CNS, reminiscent of EAE. Hypothetically, a strong expansion of myelin autoreactive IFN-gamma producing T cells secondary to nerve trauma may have immunopathological consequences in genetically predisposed individuals. It is also possible that myelin reactive T cells, whether recruited to the lesioned nerve, could have impact on macrophage function during Wallerian degeneration in the distal stump.
Assuntos
Autoimunidade , Traumatismos do Nervo Facial , Proteínas da Mielina/imunologia , Subpopulações de Linfócitos T/imunologia , Animais , Quimiotaxia de Leucócito , Suscetibilidade a Doenças , Encefalomielite Autoimune Experimental/imunologia , Nervo Facial/imunologia , Nervo Facial/patologia , Imunidade Celular , Interferon gama/metabolismo , Masculino , Proteína Básica da Mielina/imunologia , Pescoço , Ratos , Ratos Endogâmicos BN/imunologia , Ratos Endogâmicos Lew/imunologia , Subpopulações de Linfócitos T/metabolismoRESUMO
In this study, cytokine mRNA profiles in microglia from newborn rats were detected by in situ hybridization. Under natural culture conditions, microglia expressed the immunosuppressive transforming growth factor-beta 1 (TGF-beta 1) and interleukin (IL) 10 to a greater degree than the pro-inflammatory cytokines IL-1 beta, IL-6, IL-12, interferon-gamma (IFN-gamma) and TNF-alpha. High TGF-beta 1 and IL-10 levels could reflect one mechanism for immune privilege within the CNS under physiological conditions. Stimulation of microglia with LPS or IFN gamma resulted in strong up-regulation of proinflammatory cytokines, while TGF-beta 1 and IL-10 were down-regulated. These effects of LPS or IFN-gamma are anticipated to reflect immunopathogenic processes within the CNS.
Assuntos
Animais Recém-Nascidos/metabolismo , Contagem de Células/efeitos dos fármacos , Citocinas/metabolismo , Interferon gama/farmacologia , Lipopolissacarídeos/farmacologia , Microglia/efeitos dos fármacos , Animais , Relação Dose-Resposta a Droga , Hibridização In Situ , Microglia/metabolismo , Ratos , Ratos Endogâmicos LewRESUMO
Evidence from epidemiological, clinical and experimental studies favour the hypothesis that inflammatory events are part of the neuropathology in Alzheimer's disease. Proinflammatory cytokines such as interleukin-1 (IL-1), interleukin-6 (IL-6) and tumour necrosis factor-alpha (TNF-alpha) have been found in activated microglia in the vicinity of amyloid plaques in Alzheimer's disease brain. In the present study, the levels of soluble IL-1 receptor type II (sIL-1R type II), IL-1 receptor antagonist (IL-1ra), IL-1beta, IL-6 and TNF-alpha were analyzed in cerebrospinal fluid (CSF) samples from Alzheimer's disease patients and control subjects. The levels of sIL-1R type II were significantly higher in CSF from Alzheimer's disease patients than in CSF samples from control subjects (38.5+/-8 pg/ml (mean+/-S.E.M.) vs. 7.9+/-4 pg/ml, p<0.05). Measurements of the proinflammatory cytokines IL-6 and TNF-alpha showed no significant difference between the two groups, and the levels of IL-1beta and IL-1ra in the present material were too low to permit detection. The increased levels of sIL-1R type II may reflect a compensatory mechanism to balance an increased release of IL-1 receptor agonists in the Alzheimer's disease brain.
Assuntos
Doença de Alzheimer/líquido cefalorraquidiano , Doença de Alzheimer/diagnóstico , Receptores de Interleucina-1/metabolismo , Idoso , Idoso de 80 Anos ou mais , Biomarcadores , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Interleucina-6/líquido cefalorraquidiano , Masculino , Microglia/metabolismo , Pessoa de Meia-Idade , Solubilidade , Fator de Necrose Tumoral alfa/líquido cefalorraquidianoRESUMO
Antibodies against acetylcholine receptor (AChR), a protein that binds to alpha-bungarotoxin (alpha-BuTx), are characteristic for myasthenia gravis (MG) and are considered to be of importance in the pathogenesis of the disease. Antibodies against a beta-BuTx binding protein, presynaptic membrane receptor (PsmR), have also been reported in most MG patients. We have analysed the specificity and cross-reactivity of antibodies to bovine AChR and PsmR in sera from 11 patients with MG. More than 90% of antibodies to PsmR were adsorbed specifically by PsmR conjugated affinity chromatography. Similarly, more than 90% of anti-AChR antibodies were absorbed by AChR conjugated affinity chromatography. Specificities of antibodies from affinity chromatography were also confirmed by ELISA and agarose isoelectric focusing. However, the antibodies to PsmR and AChR from MG patients' sera showed about 45-55% cross-reactivity, and there was a high correlation between serum levels of both antibodies. The demonstration of anti-PsmR antibodies could be important in documenting presynaptic damage and understanding the pathogenetic process in MG.