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OBJECTIVES: GMCSF+T-cells may be involved in pathogenesis of rheumatoid arthritis (RA), and polyfunctionality may be a marker of pathogenicity. Although, higher frequencies of CD4+GMCSF+ T-cells have been reported, there are no data on CD8+GMCSF+ T-cells or polyfunctionality.Our objective was to enumerate frequencies of CD8+GMCSF+ T cells in RA blood and synovial fluid (SF), and assess their polyfunctionality, memory phenotype and cytotoxic ability. METHODS: This study included RA patients (blood samples,in some with paired synovial fluid (SF)), healthy controls (HC) (blood) and SpA patients (SF). In some RA patients' blood was sampled twice, before and 16-24 weeks after methotrexate (MTX) treatment. After mononuclear cell isolation from blood and SF, ex-vivo stimulation using PMA/Ionomycin was done, and cells were stained (surface and intracellular after permeabilisation/fixation). Subsequently, frequencies of GMCSF+CD8+ and CD4+ T-cells, polyfunctionality (TNFα, IFNγ, IL-17), phenotype (memory) and perforin/granzyme expression were assessed by flowcytometry. RESULTS: There was no significant difference in frequencies of GMCSF+CD8+ (3.7, 4.1%, p=0.540) or GMCSF+CD4+ T-cells (4.5, 5.2%, p=0.450) inblood of RA and HC. However, there was significant enrichment of both CD8+GMCSF+ (5.8, 3.9%, p=0.0045) and CD4+GMCSF+ (8.5, 4.5%, p=0.0008) T-cells inSF compared to blood in RA patients. Polyfunctional triple cytokine positive TNFα+IFNγ+GMCSF+CD8+T-cells (81, 36%, p=0.049) and CD4+T-cells (48, 32%, p=0.010) was also higher in SF compared to blood in RA. CD8+ T cells showed higher frequency of effector-memory phenotype and granzyme-B expression in RA-SF. On longitudinal follow-up, blood CD4+GMCSF+ T-cells significantly declined (4.6, 2.9%, p=0.0014) post-MTX. CONCLUSIONS: We report a novel finding of enrichment of CD8+GMCSF+ in addition to CD4+GMCSF+ T-cells in RA-SF. These cells showed higher polyfunctionality for TNFα and IFNγ, and effector memory phenotype suggesting their involvement in RA pathogenesis.
Assuntos
Artrite Reumatoide , Linfócitos T CD4-Positivos , Linfócitos T CD8-Positivos , Fator Estimulador de Colônias de Granulócitos e Macrófagos , Interferon gama , Líquido Sinovial , Fator de Necrose Tumoral alfa , Humanos , Artrite Reumatoide/imunologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Líquido Sinovial/imunologia , Líquido Sinovial/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Pessoa de Meia-Idade , Masculino , Feminino , Interferon gama/metabolismo , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Adulto , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Estudos de Casos e Controles , Idoso , Fenótipo , Antirreumáticos/uso terapêutico , Memória Imunológica , Metotrexato/uso terapêutico , Granzimas/metabolismo , Interleucina-17/metabolismo , Perforina/metabolismo , Resultado do Tratamento , Células T de Memória/imunologia , Células T de Memória/metabolismo , Citotoxicidade ImunológicaRESUMO
AIM: This study was undertaken to explicate the shared and distinctive genetic susceptibility and immune dysfunction in patients with T1D alone and T1D with CD (T1D + CD). METHODS: A total of 100 T1D, 50 T1D + CD and 150 healthy controls were recruited. HLA-DRB1/DQB1 alleles were determined by PCR-sequence-specific primer method, SNP genotyping for CTLA-4 and PTPN22 was done by simple probe-based SNP-array and genotyping for INS-23 Hph1 A/T was done by RFLP. Autoantibodies and cytokine estimation was done by ELISA. Immune-regulation was analysed by flow-cytometry. Clustering of autoantigen epitopes was done by epitope cluster analytical tool. RESULTS: Both T1D alone and T1D + CD had a shared association of DRB1*03:01, DRB1*04, DRB3*01:07/15 and DQB1*02. DRB3*01:07/15 confers the highest risk for T1D with relative risk of 11.32 (5.74-22.31). Non-HLA gene polymorphisms PTPN22 and INS could discriminate between T1D and T1D + CD. T1D + CD have significantly higher titers of autoantibodies, expression of costimulatory molecules on CD4 and CD8 cells, and cytokine IL-17A and TGF-ß1 levels compared to T1D patients. Epitopes from immunodominant regions of autoantigens of T1D and CD clustered together with 40% homology. CONCLUSION: Same HLA genes provide susceptibility for both T1D and CD. Non-HLA genes CTLA4, PTPN22 and INS provide further susceptibility while different polymorphisms in PTPN22 and INS can discriminate between T1D and T1D + CD. Epitope homology between autoantigens of two diseases further encourages the two diseases to occur together. The T1D + CD being more common in females along with co-existence of thyroid autoimmunity, and have more immune dysregulated state than T1D alone.
Assuntos
Autoantígenos , Doença Celíaca , Diabetes Mellitus Tipo 1 , Predisposição Genética para Doença , Proteína Tirosina Fosfatase não Receptora Tipo 22 , Humanos , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/imunologia , Índia/epidemiologia , Doença Celíaca/genética , Doença Celíaca/imunologia , Feminino , Masculino , Autoantígenos/imunologia , Autoantígenos/genética , Criança , Adolescente , Proteína Tirosina Fosfatase não Receptora Tipo 22/genética , Adulto , Cadeias beta de HLA-DQ/genética , Autoanticorpos/imunologia , Autoanticorpos/sangue , Cadeias HLA-DRB1/genética , Adulto Jovem , Polimorfismo de Nucleotídeo Único , Pré-Escolar , Antígeno CTLA-4/genética , Genótipo , Estudos de Casos e ControlesRESUMO
Background Mucous membrane pemphigoid (MMP) is a rare subepidermal autoimmune blistering disorder. The clinical and demographic parameters of this disease in Indian patients have not yet been elucidated in detail. Objective We aimed to study the clinical and demographic characteristics, disease course, and treatment aspects of MMP patients. Methods The data for this study were obtained by reviewing the case record forms of patients registered in the Autoimmune Bullous Disease (AIBD) Clinic of the Department of Dermatology, Venereology & Leprology, Postgraduate Institute of Medical Education and Research, Chandigarh, a tertiary care centre in India. The diagnosis of MMP was established on the basis of clinical and immune-histopathological features which are consistent with standard diagnostic criteria for the disease. Results A total of 52 patients with MMP registered in the AIBD clinic were included. The mean age at disease onset was 50 years and the average age at presentation was 56 years. Females outnumbered males in the study with a ratio of 1.36:1. The oral and ocular mucosae were the most commonly affected sites (82.6% and 63.4% respectively). Visual difficulty was reported by half the patients (26 of 52 patients). IgG, C3, and IgA deposits were detected on direct immunofluorescence (DIF) in 29, 21, and 11 patients, respectively. Serologic analysis was performed in only 7 of the patients and of these, just 1 exhibited a positive result on multivariant ELISA and epidermal pattern of binding on salt split skin indirect immunofluorescence. Most patients were treated with prednisolone (44 of 52). Steroid-sparing adjuvants were used in combination including cyclophosphamide, azathioprine, methotrexate, dapsone, and colchicine. Rituximab was administered in 7 patients with severe or refractory disease. Limitations This is a retrospective analysis of data available from a clinic registry. In patients with negative direct immunofluorescence on biopsy, the diagnosis was based on clinico-pathologic consensus. Conclusion MMP is not as uncommon in India as the paucity of reports suggest. Visual complications are frequent in Indian MMP patients. A high index of suspicion is required for early diagnosis and appropriate treatment to prevent ocular complications.
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BACKGROUND AND OBJECTIVES: The long-term effect of regular plateletpheresis on donors has not been characterized. Hence, we planned to study the long-term alterations in hematological, biochemical, and immunological parameters in regular repeat platelet apheresis donors. MATERIALS AND METHODS: Thirty-three healthy voluntary regular repeat apheresis donors presenting for platelet donation, fulfilling the requisite donor selection criteria, underwent sequential analysis of the hematological, biochemical, and immunological parameters over 1 year. RESULTS: A total of 33 regular repeat donors were enrolled in the study; out of these, 22 could be followed up to 3 months, 12 up to 6 months, and 10 donors up to 12 months for their hematological, biochemical, and immunological parameters. Overall, there was no significant change in hematological profile except a rise in platelet count at 3 months (P = 0.023) with no significant difference at 6 and 12 months from the baseline. In addition, serum thrombopoietin levels at 3 months (P = 0.010) and serum erythropoietin at 6 months (P = 0.01) were significantly higher than baseline. Mean platelet volume was significantly higher from baseline at 12 months (P = 0.00). Serum protein, lymphocyte subpopulation, and serum ferritin did not show any significant change from baseline over 12 months of follow-up. However, there was a significant decline (P = 0.00) in serum calcium and an increase in serum magnesium from baseline (P = 0.03) at 12 months. INTERPRETATIONS AND CONCLUSIONS: To conclude, apheresis platelet donation is a safe procedure. However, a complete hematological, biochemical, immunological profile and bone marrow density at regular intervals (3-6 months) are recommended to ensure the safety of regular repeat plateletpheresis donors.