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1.
Arthritis Res Ther ; 21(1): 43, 2019 01 31.
Artigo em Inglês | MEDLINE | ID: mdl-30704507

RESUMO

OBJECTIVES: An important limitation in granulomatosis with polyangiitis (GPA) is the lack of disease activity markers. Immunoglobulin G4-positive (IgG4+) B cells and plasma cells are implicated in the pathogenesis of GPA. We hypothesized that the presence of these cells in peripheral blood could serve as disease activity parameter in GPA. METHODS: We included 35 proteinase 3-antineutrophil cytoplasmic antibodies-positive patients with GPA in a cross-sectional study. Active disease was defined as Birmingham Vasculitis Activity Score (BVAS) ≥ 3 (n = 15), remission as BVAS of 0 (n = 17), and low disease activity (LDA) as BVAS of 1-2 and clinical remission (n = 3). Healthy subjects (n = 10), patients with systemic lupus erythematosus (n = 24), and patients with rheumatoid arthritis (n = 19) functioned as control subjects. An additional longitudinal study was performed in ten patients with GPA. Using a validated qPCR test, we measured the IgG4:IgG RNA ratio in all groups and compared the results with known biomarkers. RESULTS: The median qPCR score was higher in active GPA (21.4; IQR 12.1-29.6) than in remission/LDA (3.3; IQR 1.6-5.6) (Mann-Whitney U test, p < 0.0001) and outperformed other known disease activity parameters in detecting activity. A cutoff qPCR score of 11.2% differentiated active disease from remission/LDA accurately (AUC 0.993). The qPCR test correlated well with the BVAS (Spearman r = 0.77, p < 0.0001). In the longitudinal study, a decrease in BVAS correlated with qPCR score reduction (paired t test, p < 0.05). CONCLUSIONS: The IgG4:IgG RNA ratio in GPA accurately distinguishes active disease from remission and correlates well with disease activity in these single-center studies. If these results are confirmed in larger longitudinal studies, this test might help to steer treatment decisions in patients with GPA.


Assuntos
Anticorpos Anticitoplasma de Neutrófilos/imunologia , Linfócitos B/imunologia , Granulomatose com Poliangiite/diagnóstico , Imunoglobulina G/genética , Mieloblastina/imunologia , Plasmócitos/imunologia , RNA/genética , Adulto , Idoso , Anticorpos Anticitoplasma de Neutrófilos/sangue , Linfócitos B/metabolismo , Biomarcadores/sangue , Estudos Transversais , Diagnóstico Diferencial , Feminino , Granulomatose com Poliangiite/genética , Granulomatose com Poliangiite/imunologia , Humanos , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Mieloblastina/metabolismo , Plasmócitos/metabolismo , RNA/sangue , RNA/imunologia , Remissão Espontânea , Adulto Jovem
2.
J Crohns Colitis ; 11(7): 831-839, 2017 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-28158397

RESUMO

BACKGROUND AND AIM: T cells are key players in the chronic intestinal inflammation that characterises Crohn's disease. Here we aim to map the intestinal T-cell receptor [TCR] repertoire in patients with Crohn's disease, using next-generation sequencing technology to examine the clonality of the T-cell compartment in relation to mucosal inflammation and response to therapy. METHODS: Biopsies were taken from endoscopically inflamed and uninflamed ileum and colon of 19 patients with Crohn's disease. From this cohort, additional biopsies were taken after 8 weeks of remission induction therapy from eight responders and eight non-responders. Control biopsies from 11 patients without inflammatory bowel disease [IBD] were included. The TCRß repertoire was analysed by next-generation sequencing of biopsy RNA. RESULTS: Both in Crohn's disease patients and in non-IBD controls, a broad intestinal T-cell repertoire was found, with a considerable part consisting of expanded clones. Clones in Crohn's disease were more expanded [p = 0.008], with the largest clones representing up to as much as 58% of the total repertoire. There was a substantial overlap of the repertoire between inflamed and uninflamed tissue and between ileum and colon. Following therapy, responders showed larger changes in the T-cell repertoire than non-responders, although a considerable part of the repertoire remained unchanged in both groups. CONCLUSIONS: The intestinal T-cell repertoire distribution in Crohn's disease is different from that in the normal gut, containing profoundly expanded T-cell clones that take up a large part of the repertoire. The T-cell repertoire is fairly stable regardless of endoscopic mucosal inflammation or response to therapy.


Assuntos
Doença de Crohn/imunologia , Doença de Crohn/patologia , Receptores de Antígenos de Linfócitos T/metabolismo , Linfócitos T/metabolismo , Linfócitos T/patologia , Adalimumab/uso terapêutico , Adulto , Anti-Inflamatórios/uso terapêutico , Biópsia , Budesonida/uso terapêutico , Proteína C-Reativa/metabolismo , Estudos de Casos e Controles , Células Clonais/efeitos dos fármacos , Colo/patologia , Doença de Crohn/tratamento farmacológico , Feminino , Fármacos Gastrointestinais/uso terapêutico , Humanos , Íleo/patologia , Inflamação/imunologia , Inflamação/patologia , Infliximab/uso terapêutico , Masculino , Pessoa de Meia-Idade , Índice de Gravidade de Doença , Linfócitos T/efeitos dos fármacos , Adulto Jovem
3.
Ann Rheum Dis ; 73(4): 756-62, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23606709

RESUMO

OBJECTIVE: To identify potential autoreactive B-cell and plasma-cell clones by quantitatively analysing the complete human B-cell receptor (BCR) repertoire in synovium and peripheral blood in early and established rheumatoid arthritis (RA). METHODS: The BCR repertoire was screened in synovium and blood of six patients with early RA (ERA) (<6 months) and six with established RA (ESRA) (>20 months). In two patients, the repertoires in different joints were compared. Repertoires were analysed by next-generation sequencing from mRNA, generating >10 000 BCR heavy-chain sequence reads per sample. For each clone, the degree of expansion was calculated as the percentage of the total number of reads encoding the specific clonal sequence. Clones with a frequency ≥ 0.5% were considered dominant. RESULTS: Multiple dominant clones were found in inflamed synovium but hardly any in blood. Within an individual patient, the same dominant clones were detected in different joints. The majority of the synovial clones were class-switched; however, the fraction of clones that expressed IgM was higher in ESRA than ERA patients. Dominant synovial clones showed autoreactive features: in ERA in particular the clones were enriched for immunoglobulin heavy chain gene segment V4-34 (IGHV4-34) and showed longer CDR3 lengths. Dominant synovial clones that did not encode IGHV4-34 also had longer CDR3s than peripheral blood. CONCLUSIONS: In RA, the synovium forms a niche where expanded--potentially autoreactive--B cells and plasma cells reside. The inflamed target tissue, especially in the earliest phase of disease, seems to be the most promising compartment for studying autoreactive cells.


Assuntos
Artrite Reumatoide/imunologia , Autoimunidade/imunologia , Linfócitos B/imunologia , Membrana Sinovial/imunologia , Sequência de Aminoácidos , Artrite Reumatoide/genética , Células Clonais/imunologia , Regiões Determinantes de Complementaridade/genética , Regiões Determinantes de Complementaridade/imunologia , Feminino , Humanos , Switching de Imunoglobulina/imunologia , Cadeias Pesadas de Imunoglobulinas/genética , Região Variável de Imunoglobulina/genética , Ativação Linfocitária/imunologia , Masculino , Dados de Sequência Molecular , Plasmócitos/imunologia , Índice de Gravidade de Doença
4.
PLoS Pathog ; 8(9): e1002889, 2012 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-23028307

RESUMO

CD8(+) T-cell responses against latent viruses can cover considerable portions of the CD8(+) T-cell compartment for many decades, yet their initiation and maintenance remains poorly characterized in humans. A key question is whether the clonal repertoire that is raised during the initial antiviral response can be maintained over these long periods. To investigate this we combined next-generation sequencing of the T-cell receptor repertoire with tetramer-sorting to identify, quantify and longitudinally follow virus-specific clones within the CD8(+) T-cell compartment. Using this approach we studied primary infections of human cytomegalovirus (hCMV) and Epstein Barr virus (EBV) in renal transplant recipients. For both viruses we found that nearly all virus-specific CD8(+) T-cell clones that appeared during the early phase of infection were maintained at high frequencies during the 5-year follow-up and hardly any new anti-viral clones appeared. Both in transplant recipients and in healthy carriers the clones specific for these latent viruses were highly dominant within the CD8(+) T-cell receptor Vß repertoire. These findings suggest that the initial antiviral response in humans is maintained in a stable fashion without signs of contraction or changes of the clonal repertoire.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Infecções por Citomegalovirus/imunologia , Citomegalovirus/imunologia , Infecções por Vírus Epstein-Barr/imunologia , Herpesvirus Humano 4/imunologia , Receptores de Antígenos de Linfócitos T/genética , Antígenos Virais/imunologia , Citomegalovirus/genética , Herpesvirus Humano 4/genética , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Transplante de Rim/imunologia , Pessoa de Meia-Idade , Fatores de Tempo , Latência Viral , Adulto Jovem
5.
Ann Rheum Dis ; 71(6): 1088-93, 2012 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-22294635

RESUMO

OBJECTIVE: To profile quantitatively the T-cell repertoire in multiple joints and peripheral blood of patients with recent onset (early) or established rheumatoid arthritis (RA) using a novel next-generation sequencing protocol to identify potential autoreactive clones. METHODS: Synovium of patients with recent onset (early) RA (<6 months) (n=6) or established RA (>18 months) (n=6) was screened for T-cell clones by sequencing over 10 000 T-cell receptors (TCR) per sample. T cells from paired blood samples were analysed for comparison. From two patients synovial T cells were obtained from multiple inflamed joints. The degree of expansion of each individual clone was based on its unique CDR3 sequence frequency within a sample. Clones with a frequency of over 0.5% were considered to be highly expanded clones (HEC). RESULTS: In early RA synovium, the T-cell repertoire was dominated by 35 HEC (median, range 2-70) accounting for 56% of the TCR sequenced. The clonal dominance in the synovium was patient specific and significantly greater than in established RA (median of 11 HEC (range 5-24) in established RA synovium accounting for 9.8% of T cells; p<0.01). 34% (range 28-40%) of the most expanded T-cell clones were shared between different joints in the same patients, compared with only 4% (range 0-8%) between synovium and blood (p=0.01). CONCLUSIONS: In RA, a systemic autoimmune disease, the inflamed synovium forms a niche for specific expanded T-cell clones, especially in early disease. This suggests that, at least in RA, autoreactive T cells should be addressed specifically in the inflamed tissue, preferably in the early phase of the disease.


Assuntos
Artrite Reumatoide/imunologia , Artrite Reumatoide/patologia , Autoimunidade/imunologia , Linfócitos T/citologia , Linfócitos T/imunologia , Biópsia , Microambiente Celular/imunologia , Células Clonais/citologia , Células Clonais/imunologia , Progressão da Doença , Humanos , Membrana Sinovial/imunologia , Membrana Sinovial/patologia
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