RESUMEN

OBJECTIVES: The molecular mechanisms underlying the regulation of the CD152 (CTLA-4) gene are largely unknown. Two single nucleotide polymorphisms (SNPs) located in the promoter region are suspected to contribute to the pathogenesis of myasthenia gravis (MG) through regulation of gene expression. SETTING, SUBJECTS AND DESIGN: One hundred and sixty-five unrelated Swedish-Caucasian patients with MG (103 females and 62 males, age 17 to 92 years) and 148 ethnically matched healthy individuals were studied. Gene typing of two SNPs (T/C(-1772) and A/G(-1661)) and quantification of soluble CD152 were performed in the patients. Besides the association studies, the function of these two SNPs is characterized. RESULTS: We present new genetic associations of two SNPs in the CD152 gene with human MG. These SNPs located in the promoter region are involved in transcriptional binding activity for Nuclear Factor I (NF-1) and c/EBPbeta, as demonstrated using chromatin immunoprecipitation and electromobility shift assay. MG patients with the T/C(-1772) polymorphism have elevated levels of sCD152 in sera. CONCLUSIONS: The two SNPs in the promoter region are associated with MG and might cause abnormal alternative splicing and affect the expression of CD152, thereby contributing to the pathogenesis of MG.

Asunto(s)

Antígenos CD/genética , Antígenos de Diferenciación/genética , Miastenia Gravis/genética , Polimorfismo de Nucleótido Simple/genética , Factores de Transcripción/metabolismo , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Antígenos CD/metabolismo , Antígenos de Diferenciación/metabolismo , Antígeno CTLA-4 , Ensayo de Inmunoadsorción Enzimática , Femenino , Genética de Población , Humanos , Inmunosupresores , Masculino , Persona de Mediana Edad , Miastenia Gravis/etiología , Miastenia Gravis/metabolismo , Polimorfismo de Longitud del Fragmento de Restricción , Regiones Promotoras Genéticas , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa

RESUMEN

CD80 is a costimulatory factor mainly expressed on the surface of activated monocytes, B cells and dendritic cells. In this study, we demonstrate that 24% of healthy individuals have soluble forms of CD80, sCD80, in their serum. The concentration of sCD80 ranged from 0 to 1 mg/l. At the mRNA level, we detected a spliced form s1CD80 (771 bp), in unstimulated monocytes and B cells, while another form named s2CD80 (489 bp) was expressed in activated T cells as well as in freshly isolated and activated monocytes. s1CD80 lacks the transmembrane domain, and the IgC-like domain plus the transmembrane domain are spliced out of s2CD80. We also present data demonstrating that recombinant s1CD80 binds to recombinant CD152-Ig and CD28-Ig. It can also bind to T cells, preferentially to activated T cells. Recombinant sCD80 had immunomodulatory effects shown by its inhibition of the mixed lymphocyte reaction and inhibition of T-cell proliferation. sCD80 in human serum adds a new member to the family of soluble receptors, implying a network of soluble costimulatory factors with functional relevance. The inhibitory effect of the recombinant protein on T-cell activation makes it a possible candidate for treatment of diseases associated with hyperactivated T cells.

Asunto(s)

Antígenos CD/metabolismo , Antígenos de Diferenciación/metabolismo , Antígeno B7-1/sangre , Antígeno B7-1/genética , Antígenos CD28/metabolismo , Activación de Linfocitos/inmunología , Linfocitos T/inmunología , Empalme Alternativo , Antígeno B7-1/inmunología , Secuencia de Bases , Western Blotting , Antígeno CTLA-4 , Ensayo de Inmunoadsorción Enzimática , Citometría de Flujo , Humanos , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Isoformas de Proteínas/genética , Isoformas de Proteínas/inmunología , Isoformas de Proteínas/metabolismo , ARN Mensajero , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Linfocitos T/metabolismo

RESUMEN

OBJECTIVES: Wegener's granulomatosis (WG) is a chronic inflammatory disease characterized by granulomatosis inflammation, systemic vasculitis and glomerulonephritis. In patients, the peripheral T cells are characterized by mono/oligoclonal CD4+/CD8+ T-cell AV/BV receptor expansions, with aberrant expression of activation markers. This study was designed to characterize the phenotypic differences between the expanded and nonexpanded T-cell populations. Expression of markers for activation, costimulation and adhesion molecules was examined. As earlier studies have shown aberrant expression of CD28/CD152, we also analysed the expression of another costimulatory system, the tumour necrotic factor receptor (TNFR) superfamily proteins. DESIGN: Fluorocrome-conjugated monoclonal antibodies and flow cytometry was used to analyse the expression of the different markers on the surface of the expanded and nonexpanded subsets of T cells. SETTING: The Karolinska Hospital and Karolinska Institutet in Stockholm, Sweden. SUBJECTS: Nine patients with WG (six men and three women) had 16 TCRAV/BV CD4+/CD8+ expanded populations that were characterized. RESULTS: The expanded TCRA/BV CD4+ and CD8+ cells had lower percentages of cells expressing CD28 and higher of those expressing CD152 (CTLA-4). The expanded CD4+ population had more cells expressing HLA-DR, CD57 and CCR5 (CD195), whilst the expression of CD25 was present on fewer of the expanded cells. The expanded CD8+ population contained more cells expressing CD137 (4-1BB), CD137 (4-1BBL), CD30 (Ki-1), CD40 and CD134 (OX40). CONCLUSIONS: There were marked differences in the phenotypes of expanded and nonexpanded T-cell populations.

Asunto(s)

Granulomatosis con Poliangitis/inmunología , Activación de Linfocitos , Subgrupos de Linfocitos T/inmunología , Adulto , Anciano , Antígenos CD/análisis , Biomarcadores/análisis , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Femenino , Antígenos HLA-DR/análisis , Humanos , Inmunofenotipificación , Masculino , Persona de Mediana Edad , Receptores de Antígenos de Linfocitos T alfa-beta/análisis , Receptores CCR5/análisis

RESUMEN

Studies in experimental animal models of human autoimmune diseases have revealed that CD4(+)CD25(+) T regulatory (Tr) cells are of thymic origin and have potentials in preventing auto-aggressive immunity. Myasthenia gravis (MG) is the best-characterized autoimmune disease. Changes in the thymus are found in a majority of patients with MG. Thymectomy has beneficial effects on the disease severity and course in a substantial proportion of MG patients. But the occurrence and characteristics of Tr cells have not yet been defined in MG. We determined the frequencies and properties of circulating CD4(+)CD25(+) versus CD4(+)CD25(-) cells in MG patients and healthy controls (HCs), with special focus on the effect of thymectomy on CD4(+)CD25(+) cells. CD4(+)CD25(high) cells comprise only about 2% of blood lymphocytes in both MG patients and HCs. Frequencies of CD4(+)CD25(high) cells were similar in MG patients irrespective of treatment with thymectomy. CD4(+)CD25(+) cells in both MG patients and HCs are mainly memory T cells and are activated to a greater extent than CD4(+)CD25(-) cells, as reflected by high levels of CD45RO and human leucocyte antigen (HLA)-DR-positive cells. In both MG patients and HCs, CD4(+)CD25(+) cells also contained a high proportion of CD95-expressing cells as possible evidence of apoptosis-proneness. Upon stimulation with anti-CD3/CD28 monoclonal antibodies, CD4(+)CD25(+) cells responded more vigorously than CD4(+)CD25(-) cells in MG, irrespective of treatment with thymectomy, as well as in HCs. Although CD4(+)CD25(-) cells are mainly naïve T cells, in non-thymectomized MG patients, they are activated to a greater extent as reflected by higher expression of HLA-DR and CD95 on the surface compared to HCs. The data thus show that there is no deficiency of CD4(+)CD25(+) cells in MG, nor is the proportion of CD4(+)CD25(+) cells influenced by thymectomy.

Asunto(s)

Antígenos CD4/inmunología , Miastenia Gravis/inmunología , Receptores de Interleucina-2/inmunología , Linfocitos T/inmunología , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Antígenos CD4/sangre , Femenino , Antígenos HLA-DR/sangre , Antígenos HLA-DR/inmunología , Humanos , Antígenos Comunes de Leucocito/sangre , Antígenos Comunes de Leucocito/inmunología , Masculino , Persona de Mediana Edad , Receptores de Interleucina-2/sangre , Timectomía , Receptor fas/sangre , Receptor fas/inmunología

RESUMEN

Two pairs of monozygotic twins, discordant for myasthenia gravis (MG) for more than 30 years, were studied regarding T cell and antibody reactivity against disease related autoantigens, the acetylcholine receptor, one idiotypic and one anti-idiotypic human monoclonal antibody. The healthy and myasthenic twins had very similar autoantibody repertoires. IgG fractions from both healthy and myasthenic twins had the same capacity to decrease the free acetylcholine receptor content in mice after passive transfer. In comparison with their myasthenic sisters, the healthy twins had lower T cell responses against the acetylcholine receptor.

Asunto(s)

Autoinmunidad/fisiología , Linfocitos B/inmunología , Miastenia Gravis/inmunología , Linfocitos T/inmunología , Adulto , Animales , Anticuerpos Antiidiotipos/sangre , Anticuerpos Antiidiotipos/inmunología , Anticuerpos Monoclonales/sangre , Anticuerpos Monoclonales/inmunología , Autoantígenos/metabolismo , Autoinmunidad/genética , Linfocitos B/virología , Células Sanguíneas/inmunología , Linfocitos T CD4-Positivos/metabolismo , Linfocitos T CD8-positivos/metabolismo , Línea Celular , Transformación Celular Viral/inmunología , Citocinas/metabolismo , Femenino , Estudios de Seguimiento , Antígenos HLA-DR/metabolismo , Herpesvirus Humano 4 , Humanos , Antígenos Comunes de Leucocito , Estudios Longitudinales , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , Miastenia Gravis/patología , Miastenia Gravis/virología , Receptores de Antígenos de Linfocitos T/genética , Receptores de Antígenos de Linfocitos T/inmunología , Receptores Colinérgicos/sangre , Receptores Colinérgicos/inmunología , Linfocitos T/virología , Estudios en Gemelos como Asunto , Gemelos Monocigóticos

RESUMEN

CTLA-4 is an important negative regulator of the immune system. The regulation of the CTLA-4 gene (Ctla-4) transcription is poorly understood. A single nucleotide polymorphism (SNP) at -318 in the Ctla-4 promoter region is associated with certain autoimmune diseases. Since the -318 SNP occurs in a potential regulatory region, it is conceivable that the C' T transition may affect the expression of Ctla-4. In the present study, we show that the -318T allele is associated with a higher promoter activity than the -318C allele (8.13 +/- 0.46 vs 6.87 +/- 0.49). The presence of the -318T allele may thus contribute to up regulation of the expression of CTLA-4, and consequently represent one mechanism to inhibit exaggerated immune activity.

Asunto(s)

Antígenos de Diferenciación/genética , Regulación de la Expresión Génica , Inmunoconjugados , Polimorfismo Genético , Regiones Promotoras Genéticas , Abatacept , Antígenos CD , Antígeno CTLA-4 , Humanos , Transcripción Genética

RESUMEN

Cytotoxic T lymphocyte-associated molecule-4 (CTLA-4) is a receptor present on T cells that plays a critical role in the downregulation of antigen-activated immune responses. CTLA-4 interacts with the ligands CD80 and CD86 on antigen-presenting cells (APC), and also directs the assembly of inhibitory signalling complexes that lead to quiescence or anergy. In this study, we show that human monocytes constitutively express CTLA-4. About 3% of monocytes expressed CTLA-4 on the cell surface, whereas the intracellular expression was higher and present in about 20% of the monocytes. The sequences of the cDNAs from human monocytes were identical to the sequences of CTLA-4 from T cells. Expression of CTLA-4 was also confirmed in the activated myelomonocytic cell lines U937 and THP-1. Monocytes, but not T cells, activated by interferon (IFN)-gamma also secreted soluble CTLA-4 in vitro. The CTLA-4 expression was upregulated upon treatment with phorbol 12-myristate 13-acetate (PMA) and IFN-gamma. This increased expression could be partially abolished by staurosporine, an inhibitor of protein kinase C (PKC). Ligation of CTLA-4 in the monocyte-like cell-line U937 with antibodies against CTLA-4 partially inhibited the proliferation of cells and the upregulation of cell-surface markers CD86, CD54, HLA-DR and HLA-DQ induced by IFN-gamma and Staphylococcus aureus, Cowan I strain (SAC). Ligation of CTLA-4 suppressed the PMA-stimulated activation of transcription activator protein 1 (AP-1) and nuclear factor (NF)-kappaB in the U937 cell line, indicating the involvement of an inhibitory signal transduction. These data provide the first evidence that CTLA-4 is constitutively expressed by monocytes and thus might be important for the regulation of immune mechanisms associated with monocytes.

Asunto(s)

Antígenos de Diferenciación/metabolismo , Inmunoconjugados , Monocitos/inmunología , Abatacept , Adulto , Antígenos CD , Antígenos de Diferenciación/química , Antígenos de Diferenciación/genética , Secuencia de Bases , Antígeno CTLA-4 , Línea Celular , Membrana Celular/inmunología , Reactivos de Enlaces Cruzados , ADN/genética , ADN/metabolismo , Expresión Génica , Humanos , Técnicas In Vitro , Interferón gamma/farmacología , Líquido Intracelular/inmunología , Monocitos/efectos de los fármacos , Monocitos/metabolismo , FN-kappa B/metabolismo , Proteína Quinasa C/metabolismo , Proteínas Recombinantes , Factor de Transcripción AP-1/metabolismo , Células U937

RESUMEN

The cytotoxic T lymphocyte associated protein 4 (CTLA-4) gene (Ctla-4) is a candidate gene for autoimmune disease. We here report results of two single nucleotide polymorphisms (SNPs) in the Ctla-4, a +49 A/G SNP in CDS1 and a C/T promoter SNP at position -318. There were no differences in these two SNPs between patients and healthy individuals. The frequency of allele G and genotype G/G at position +49 in CDS1 was increased in patients with thymoma when compared with patients with normal and hyperplastic thymic histopathology. Patients with the G/G genotype had signs of immune activation manifested as higher levels of serum IL-1beta and higher percentage of CD28(+) T lymphocytes. There was a strong linkage between the 86bp allele in the 3'-UTR and the A(+49) allele in CDS1. Our results suggest that the SNP at position +49 in CDS1 might be associated with the manifestations of MG.

Asunto(s)

Antígenos de Diferenciación/genética , Inmunoconjugados , Miastenia Gravis/genética , Polimorfismo de Nucleótido Simple , Regiones Promotoras Genéticas , Regiones no Traducidas 3' , Abatacept , Antígenos CD , Antígenos CD28/análisis , Antígeno CTLA-4 , Femenino , Frecuencia de los Genes , Genotipo , Humanos , Interleucina-1/sangre , Interleucina-12/sangre , Desequilibrio de Ligamiento , Masculino , Miastenia Gravis/inmunología , Suecia , Linfocitos T/inmunología , Timoma/genética , Hiperplasia del Timo/genética , Neoplasias del Timo/genética

RESUMEN

Cytotoxic T-lymphocyte-associated antigen (CTLA-4) is an important downregulator of T-cell activation. In order to analyze the expression and regulation of CTLA-4 on human peripheral T cells, CTLA-4 mRNA and protein expression were determined using analysis by reverse transcription-polymerase chain reaction (RT-PCR) and FACs, respectively. Intracellular CTLA-4 was constitutively expressed in unstimulated CD4+ and CD8+ T cells. Interleukin (IL)-2 induced a dose-dependent increase of both intracellular and surface expression of CTLA-4 (CD152). Most of the CD4+ and CD8+ cells expressing CTLA-4 also expressed CD25. Interferon (IFN)-gamma induced the upregulation of CTLA-4 expression via antigen-presenting cells (APC) activation. The CTLA-4delTM mRNA (550 bp) had a shorter half-life than the full length CTLA-4 mRNA and the expression was downregulated upon activation of the cells by treatment with IL-2. Given an inhibitory role of CTLA-4 and CD4+ CD25+ T cells in immune responses, the present findings suggest that IL-2-induced immunosuppression may result from its stimulatory effect of the CTLA-4 expression.

Asunto(s)

Antígenos de Diferenciación/genética , Antígenos de Diferenciación/metabolismo , Inmunoconjugados , Linfocitos T/inmunología , Abatacept , Antígenos CD , Antígenos de Diferenciación/farmacología , Secuencia de Bases , Linfocitos T CD4-Positivos/efectos de los fármacos , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/efectos de los fármacos , Linfocitos T CD8-positivos/inmunología , Antígeno CTLA-4 , Membrana Celular/inmunología , Concanavalina A/farmacología , Cartilla de ADN/genética , Citometría de Flujo , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Técnicas In Vitro , Interferón gamma/farmacología , Interleucina-2/farmacología , Líquido Intracelular/inmunología , Activación de Linfocitos , Receptores de Interleucina-2/metabolismo , Proteínas Recombinantes , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Solubilidad , Linfocitos T/efectos de los fármacos

RESUMEN

Human autoimmune myasthenia gravis (MG) is associated with the IL-1beta TaqI RFLP allele 2. Individuals positive for this allele have high levels of inducible IL-1beta in their peripheral blood. Here, we have characterized MG induction and the immune response elicited by Torpedo acetylcholine receptor (AChR) immunization in wild-type and IL-1beta deficient (-/-) mice. Compared with wild-type mice, IL-1beta-/- mice were relatively resistant to induction of clinical experimental autoimmune myasthenia gravis (EAMG). Draining lymph node cells from IL-1beta-/- mice showed poor proliferative capacity upon AChR stimulation in vitro. Both Th1 (IFN-gamma, IL-2) and Th2 (IL-4) cytokine responses were reduced and levels of serum anti-AChR antibodies decreased in IL-1beta-/- mice compared to wild-type mice. Taken together, these results reveal a critical role for IL-1beta in the induction of MG in mice, and support a role for IL-1beta in the pathogenesis of MG in man.

Asunto(s)

Interleucina-1/fisiología , Miastenia Gravis/etiología , Receptores Colinérgicos/inmunología , Animales , Autoanticuerpos/sangre , Citocinas/biosíntesis , Inmunización , Interleucina-1/genética , Activación de Linfocitos , Ratones , Ratones Endogámicos C57BL , Miastenia Gravis/inmunología

RESUMEN

CD28 is required to promote T cell proliferation and cytokine production, while the cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) functions as a negative modulator for T cell activation. We previously reported that alleles with longer PCR products (designated as allele xx) in an (AT)n polymorphism in Ctla-4 are associated with myasthenia gravis with thymoma, while the shortest allele, 86, is negatively associated with the disease. Here, we demonstrate that serum IL-2 sRalpha increases parallel to the length of (AT)n in Ctla-4. Periphereal blood mononuclear cells (PBMC) from patients with Ctla-4 xx/xx contained higher activity of telomerase than patients bearing Ctla-4 86/86. Blockade of CTLA-4 increased the telomerase activity in PBMC stimulated by acetylcholine receptor in vitro. There was a positive correlation between the expression of CD28 and CTLA-4 on anti-CD3 activated PBMC, suggesting a balance between CD28 and CTLA-4. Cells from patients with Ctla-4 xx/xx had the highest level of T cell proliferative responses upon the addition of anti-CD28 antibodies to the anti-CD3 containing culture system while cells from patients with Ctla-4 86/xx had an intermediate and cells from patients with Ctla-4 86/86 the lowest increase. The current results point to the (AT)n in Ctla-4 as a myasthenia gravis facilitating mutation under certain permissive environments by influencing the T cell reactivity via the CD28 pathway.

Asunto(s)

Antígenos de Diferenciación/genética , Antígenos CD28/fisiología , Repeticiones de Dinucleótido , Inmunoconjugados , Activación de Linfocitos , Miastenia Gravis/inmunología , Linfocitos T/inmunología , Abatacept , Antígenos CD , Complejo CD3/inmunología , Antígeno CTLA-4 , Células Cultivadas , Humanos , Receptores de Interleucina-2/análisis , Telomerasa/metabolismo

RESUMEN

OBJECTIVE: The genetic factors predisposing to Wegener's granulomatosis (WG) are largely unknown. T cells are clearly involved in the disease, as are the proinflammatory cytokines tumor necrosis factor-alpha (TNF-alpha) and interleukin 1beta (IL-1beta). The cytotoxic T lymphocyte associated antigen-4 (CTLA-4) suppresses antigen-specific immune responses by opposing the CD28 pathway, and is crucial for a balanced T cell activation. Genetic variations in the TNF-alpha, IL-1beta, and CTLA-4 genes could thus be important in WG. METHODS: Polymorphisms in the genes coding for TNF-alpha, IL-1beta, and CTLA-4 were analyzed in 32 Swedish Caucasian patients and 109 ethnically matched controls. Results. A strong association of Ctla-4 (AT)n microsatellite to WG contrasts to the negative finding of associations between TNF-alpha NcoI, IL-1beta TaqI restriction fragment length polymorphism, and WG. The prevalence of the shortest Ctla-4 allele was decreased in patients with WG compared with healthy individuals (p < 0.0001, pc < 0.0016). CONCLUSIONS: This is the first report of a T cell related gene in association with WG. The Ctla-4 itself, or a gene close to Ctla-4, may thus contribute to the pathogenesis of WG by allowing an increased T cell activation by antigen.

Asunto(s)

Antígenos de Diferenciación/genética , Granulomatosis con Poliangitis/genética , Inmunoconjugados , Interleucina-1/genética , Polimorfismo Genético , Factor de Necrosis Tumoral alfa/genética , Abatacept , Adulto , Anciano , Anciano de 80 o más Años , Alelos , Animales , Antígenos CD , Antígeno CTLA-4 , Femenino , Humanos , Masculino , Persona de Mediana Edad , Ratas

RESUMEN

We examined the bi-allelic polymorphism at - 174 in the promoter region and the polymorphism in the 3' flanking AT rich region of the interleukin-6 (IL-6) gene in Swedish patients with myasthenia gravis (MG) and ethnically matched healthy individuals. There was no association between the polymorphisms and the disease. There was no relation of the polymorphisms to the clinical variables, the thymic histopathologies, the level of serum acetylcholine receptor antibodies or the concentrations of IgG and its subclasses. Our data yield no evidence for the IL-6 gene contributing to the disease susceptibility.

Asunto(s)

Interleucina-6/genética , Miastenia Gravis/genética , Polimorfismo Genético , Secuencia Rica en At , Femenino , Humanos , Inmunoglobulina G/sangre , Inmunoglobulina G/clasificación , Masculino , Miastenia Gravis/inmunología

RESUMEN

Patients with Wegener's granulomatosis have a high prevalence of expanded populations of CD4+ and CD8+ T cells bearing different alpha/beta T cell receptors. To elucidate the role of these populations, we studied the phenotypic and functional characteristics of 13 expanded T cell populations in four patients for a period of 35-51 months. The expanded populations generally showed a persistently high expression of the activation markers HLA-DR and CD25. This expression was independent of the activity of the disease. The expanded populations also expressed CD45RO and/or CD45RA and most of them expressed CD57 but not CD28. Analysis of intracellular presence and secretion of IFN-gamma, IL-2, and IL-4 showed that most of the expanded cell populations contained and/or secreted more of these cytokines than the nonexpanded populations, with an especially high expression/secretion of IFN-gamma and IL-2. The expanded populations showed little proliferative response to Con A and OKT3. The proliferative response of the cells was partly restored after preincubation in medium alone. Some of the expanded populations were associated with disease activity, thus suggesting a link between expanded T cells and the disease. The activated status of the expanded populations and the tendency for certain populations to correlate in magnitude with disease activity suggest their involvement in the disease process. The relative stability of these cell populations indicates that the stimulus driving them is persistent, in agreement with the chronicity of the disease.

Asunto(s)

Granulomatosis con Poliangitis/inmunología , Granulomatosis con Poliangitis/fisiopatología , Subgrupos de Linfocitos T/inmunología , Anciano , Concanavalina A/farmacología , Femenino , Humanos , Inmunofenotipificación , Interferón gamma/análisis , Interleucina-2/análisis , Interleucina-4/análisis , Estudios Longitudinales , Activación de Linfocitos , Recuento de Linfocitos , Masculino , Persona de Mediana Edad , Muromonab-CD3/farmacología , Receptores de Antígenos de Linfocitos T/sangre , Subgrupos de Linfocitos T/metabolismo

RESUMEN

T lymphocytes are implicated in the pathogenesis of systemic vasculitis such as Wegener's granulomatosis (WG) and polyarteritis nodosa (PAN). In the present study, we have characterized in detail the T-cell receptor (TCR) of peripheral blood T cells from eight vasculitis patients of known HLA class II genotypes. We used flow cytometry to outline the exact TCR V gene expression, complementarity determining region 3 (CDR3) fragment analysis to estimate the degree of clonality and cDNA sequencing to define the exact TCR or beta chain sequences. The TCR CDR3 region interacts with antigenic peptides presented by HLA molecules, and it is normally immensely diverse. It was therefore of particular interest to identify a common dominating TCR BV8-F/L-G-G-A/Q-G-J2S3 beta chain sequence in the CD4(+) T cells of four unrelated vasculitis patients. Furthermore, this BV8-associated CDR3 motif was linked to the HLA-DRB1*0401 allele, as well as to active disease and/or an established BV8(+) CD4(+) T-cell expansion. In contrast, age- and HLA-matched patients with rheumatoid arthritis did not harbor the described BV8 motif. These results strongly suggest that BV8(+) CD4(+) T cells with the described CDR3 motif recognize a specific antigen presented by DR4 molecules, indicating the existence of a common vasculitis-associated antigen.

Asunto(s)

Alelos , Autoantígenos/inmunología , Enfermedades Autoinmunes/inmunología , Linfocitos T CD4-Positivos/inmunología , Reordenamiento Génico de la Cadena beta de los Receptores de Antígenos de los Linfocitos T , Granulomatosis con Poliangitis/inmunología , Antígenos HLA-DR/genética , Poliarteritis Nudosa/inmunología , Receptores de Antígenos de Linfocitos T alfa-beta/genética , Adulto , Anciano , Secuencia de Aminoácidos , Enfermedades Autoinmunes/sangre , Enfermedades Autoinmunes/genética , Células Clonales/inmunología , Femenino , Genotipo , Granulomatosis con Poliangitis/sangre , Granulomatosis con Poliangitis/genética , Antígenos HLA-DR/inmunología , Cadenas HLA-DRB1 , Humanos , Activación de Linfocitos , Recuento de Linfocitos , Masculino , Persona de Mediana Edad , Poliarteritis Nudosa/sangre , Poliarteritis Nudosa/genética , Reacción en Cadena de la Polimerasa , Alineación de Secuencia

RESUMEN

The usage of T cell receptor (TCR) Valpha/Vbeta chains on cells from 38 patients with myasthenia gravis (MG) was determined by flow cytometry. There was a decreased number of cells expressing Vbeta2 in CD8+ and Vbeta3 in CD4+ cells in patients compared with healthy individuals. Abnormal expansions of T cells using particular TCR Valpha/Vbeta gene products were found in 18/38 patients. A significantly higher usage of Vbeta13 was observed but there was no restriction with regard to other TCR Valpha/Vbeta. Expanded cells belonging to both CD4+ and CD8+ were present in MG patients while restricted to the CD8+ population in healthy individuals. To elucidate the role of the expanded populations, we studied characteristics of the expanded and non-expanded T cells from MG patients who had persistent T cell expansions over more than 2 years. The cells were analysed with regard to phenotype, cytokine secretion, cytokine mRNA expression and reactivity with the autoantigen, the acetylcholine receptor. The characteristics of the expanded populations in MG clearly differed from those found in healthy individuals. More cells in the CD4+ expanded populations expressed HLA-DR and there was also a tendency for higher expression of CD25, CD28 and CD57. The number of cells spontaneously secreting cytokines was higher in the expanded populations. A dominant Th1-type cytokine secretion and mRNA expression was noted. Autoantigen-reactive CD4+ T cells were largely restricted to the expanded populations.

Asunto(s)

Miastenia Gravis/inmunología , Receptores de Antígenos de Linfocitos T alfa-beta/genética , Linfocitos T/fisiología , Adulto , Anciano , Anciano de 80 o más Años , Citocinas/genética , Femenino , Humanos , Interferón gamma/biosíntesis , Interleucina-4/biosíntesis , Masculino , Persona de Mediana Edad , ARN Mensajero/análisis

RESUMEN

The aim of this study was to analyze associations between myasthenia gravis (MG) and polymorphisms in the tumor necrosis factor (TNF) region in 79 Swedish patients and 155 unrelated controls. The frequency of the TNFa2 allele of a microsatellite located 3.5 kb upstream of the lymphotoxin alpha (LT-alpha) gene in the TNF region was found to be increased in overall MG patients compared to controls. The frequency of the short 5.5 kb fragment (TNFB * 1) of a bi-allelic NcoI RFLP polymorphism located at the first intron of the LT-alpha gene was increased in patients with an early onset of disease compared to patients with a later onset.

Asunto(s)

Miastenia Gravis/genética , Polimorfismo Genético/genética , Factor de Necrosis Tumoral alfa/genética , Edad de Inicio , Alelos , Humanos , Intrones/genética , Linfotoxina-alfa/genética , Repeticiones de Microsatélite/genética , Miastenia Gravis/epidemiología , Fragmentos de Péptidos/genética , Polimorfismo de Longitud del Fragmento de Restricción , Valores de Referencia

Asunto(s)

Miastenia Gravis/genética , Miastenia Gravis/inmunología , Polimorfismo de Longitud del Fragmento de Restricción , Timo/patología , Factor de Necrosis Tumoral alfa/genética , Miembro 3 de la Subfamilia B de Transportadores de Casetes de Unión a ATP , Transportadoras de Casetes de Unión a ATP/genética , Adulto , Alelos , Femenino , Antígenos HLA-DQ/genética , Cadenas beta de HLA-DQ , Antígeno HLA-DR3/genética , Haplotipos , Humanos , Hiperplasia , Linfotoxina-alfa/genética , Complejo Mayor de Histocompatibilidad , Masculino , Repeticiones de Microsatélite , Miastenia Gravis/patología , Valores de Referencia

RESUMEN

To characterize better the functional aspects of the HLA class II associations with myasthenia gravis (MG), T-cell receptor (TCR) V alpha/beta elements were studied in peripheral blood in 29 Swedish MG patients. HLA typing had previously been done using polymerase chain reaction with sequence-specific primers (PCR-SSP) or combined with sequence-specific oligonucleotide probes (PCR-SSO). The TCR V gene expression was determined by fluorescence-activated cell sorter (FACS) analysis using 12 monoclonal antibodies (mAb) that detected 30-40% of CD4+ and CD8+ T cells. No correlation between HLA-DQ genotype and TCR V elements could be found, nor was any restricted V gene usage seen. Fourteen (48%) of the patients had T cells showing signs of abnormal expansion in peripheral blood. There was an increased expression of TCR V gene elements in CD8+ T cells in patients (13/29) compared with CD4+ T cells in patients (5/29) (P < 0.05) and in unthymectomized patients compared with controls (14/56) (P < 0.005). TCR V gene expression was also increased in the CD8+ population in unthymectomized (7/8) compared with thymectomized patients (6/21) (P < 0.01). There was an increased expression in both CD4+ and CD8+ populations in unthymectomized patients (7/8, 88%), compared with thymectomized patients (7/21) (P < 0.05). We conclude that the abnormal T-cell expansion in peripheral blood could be a reflection of non-specific pathogenic processes in the muscle and thymus.

Asunto(s)

Antígenos HLA-DQ , Miastenia Gravis/inmunología , Receptores de Antígenos de Linfocitos T alfa-beta/inmunología , Adolescente , Adulto , Edad de Inicio , Anciano , Anticuerpos Monoclonales , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Niño , Femenino , Frecuencia de los Genes , Prueba de Histocompatibilidad/métodos , Humanos , Masculino , Persona de Mediana Edad , Estadísticas no Paramétricas , Suecia , Subgrupos de Linfocitos T , Población Blanca/genética

RESUMEN

The association between myasthenia gravis (MG) and TAP polymorphisms was studied in 79 Swedish patients and 155 unrelated controls. TAP typing was performed by ARMS-PCR technique and stratification analysis was used to determine if the TAP associations were independent or secondary to linkage disequilibrium with DQ2 and DR3. TAP1 and TAP2 alleles did not confer independent risk for MG. TAP2*0101 was, however, positively associated with MG in patients with an early onset of disease compared to patients with a late onset of disease. TAP1 and TAP2 alleles did not confer risk in MG patients negative for DQ2. In conclusion, susceptibility to MG is not primarily conferred by TAP alleles in the extended DR3 haplotype.

Asunto(s)

Transportadoras de Casetes de Unión a ATP/genética , Antígenos HLA-DQ/sangre , Antígeno HLA-DR3/sangre , Complejo Mayor de Histocompatibilidad/genética , Miastenia Gravis/genética , Miastenia Gravis/inmunología , Transportador de Casetes de Unión a ATP, Subfamilia B, Miembro 2 , Miembro 3 de la Subfamilia B de Transportadores de Casetes de Unión a ATP , Alelos , Susceptibilidad a Enfermedades , Frecuencia de los Genes , Humanos , Suecia