Increased percentage of cd3+, cd57+ lymphocytes in patients with rheumatoid arthritis. correlation with duration of disease

Objective. To determine whether a small CD3+ lymphocyte population expressing 110-kd CD57 antigens (HNK1) is expanded in patients with rheumatoid arthritis (RA), as it is in patients who have undergone bone marrow transplantation and patients with the acquired immunodeficiency syndrome, and to investigate whether it is involved in the pathogenesis of RA. Methods. The phenotype of CD3+, CD57+ lymphocytes was analyzed by flow cytometry, and correlations between the percentage of these cells in the blood and various clinical and biologic parameters were investigated. Results. The percentage of CD3+, CD57+ lymphocytes was increased in RA patients compared with controls. These lymphocytes expressed T cell receptor α/β. Eighty percent expressed the CD8 accessory molecule, and 20% expressed the CD4 accessory molecule. The leukocyte common antigen CD45RA isoform was expressed by these CD3+, CD57+ lymphocytes in blood. The HLA–DR antigen was expressed in synovial fluid but not in blood. Finally, the percentage of these lymphocytes in the blood correlated with the duration of RA. Conclusion. The expansion of the CD3+, CD57+ lymphocyte population and their activation in the synovial fluid of RA patients suggest that these cells are involved in the inflammatory process.     

Comparison of blood and synovial fluid lymphocyte subsets in rheumatoid arthritis and osteoarthritis

Summary Immunoregulatory T-cell deficiency is thought to underlie pathogenesis of rheumatoid arthritis (RA) as a systemic autoimmunopathy. The aim of this study was a simultaneous analysis of peripheral blood and synovial lymphocyte subsets (Ly-SS) of RA patients as compared to patients with locally active osteoarthritis (OA). Peripheral blood Ly-SS and paired synovial fluid Ly-SS from 87 RA patients were analysed by two dimensional flow cytometry (Simulset Becton Dickinson) as compared to 15 OA patients. The control group consisted of 32 healthy subjects. The peripheral blood analysis from RA and OA patients revealed a significant decrease of CD8+T-cells and increase of CD4+:CD8+ ratio when compared to the control group. The blood of RA patients showed a significant increase of HLA DR+ and IL 2R+T cells as compared to OA group. The synovial fluid from RA and OA patients showed a significant increase of CD3+, CD8+, HLA DR+ T-cells and decrease of CD4+:CD8+ ratio and CD19+ cells in comparison to the peripheral blood. This study shows, that the OA T-cell system seems not to be activated in peripheral blood in opposition to RA patients. Synovial fluid Ly-SS in OA, however, showed only quantitative but not qualitative differences. OA seems to be mainly a local inflammatory response depending on T-cells, when lymphocyte T activity in blood is diminished.     

Mycobacteria and human heat shock protein—specific cytotoxic t lymphocytes in rheumatoid synovial inflammation

Objective. To study the cytotoxic capacity of mycobacteria-specific T lymphocyte lines and clones from sites of inflammation in patients with rheumatoid arthritis (RA). We also studied antigen specificity, surface phenotype, expression of T cell receptors (TCR), and HLA restriction. Methods. Autologous macrophages (Mπ) from the synovial membrane (SM), synovial fluid (SF), or peripheral blood (PB) were used as target cells in cytotoxicity assays. Results. All SM and SF cell lines tested thus far have shown specific lysis of the autologous Mπ from SM or PB that had been pulsed with BCG (bacillus Calmette-Guerin), but no cytotoxicity when the targets were pulsed with irrelevant antigens such as tetanus toxoid and Chlamydia. Both CD4+ and CD8+ cells were shown to be involved in the specific cytolysis. The majority of the cytotoxic T lymphocyte (CTL) lines were TCRα/β+ cells. However, both TCRα/β+ and TCRγ/δ+ clones (TCR δ1+) from one RA patient showed antigen-specific lysis. Antigen-specific recognition by a number of CTL lines and clones generated from SF and SM was restricted by HLA—DR molecules. Two Mycobacterium bovis 65-kd heat shock protein (HSP)–specific TCRα/β+ SF T cell clones also lysed Mπ that had been pulsed with a recombinant human 65-kd HSP. Conclusion. Joint inflammation and destruction might be partly attributable to a cross-reaction of mycobacteria-induced cytotoxic T cells with self HSP.     

Lymphocyte subset abnormalities,autoantibodies and their relationship with HLA DR types in children with Type 1 (insulin-dependent) diabetes and their first degree relatives

Summary Type 1 (insulin-dependent) diabetes mellitus is associated with abnormalities of circulating lymphocyte subsets and autoantibodies. To investigate the prevalence of these in non-diabetic siblings and non-diabetic parents of children with Type 1 diabetes, we analysed T-cell subsets of function and activation in 31 families with an index case of Type 1 diabetes and related these to autoantibodies and HLA DR type. Using two and three colour cytofluorimetry, we studied total and activated (HLA-DR+) CD3+, CD4+, CD8+ lymphocytes and on CD4+ lymphocytes the CD45RA/RO “naive” and “memory” cell phenotypes. Diabetic children (mean duration of disease 3.1 years) had a reduced total lymphocyte count (p <0.05), their non-diabetic siblings a reduced CD4+ T-helper cell count (p <0.05), and their parents a reduced percentage and number of CD3+ T cells (p <0.01 and p <0.05) compared with age-matched control subjects. Diabetic children, their siblings and parents all had significantly increased levels of activated CD4+ T-helper cells (p <0.01, p <0.05 and p <0.01). In diabetic children and their siblings there was a significant over-expression of the CD45RO “memory” cell marker and significant under-expression of the CD45RA “naive” cell marker, whilst these were normal in the parents. Islet cell antibody positive diabetic children had significantly higher levels of CD45RO-expressing CD4+ lymphocytes than those who were islet cell antibody negative (p <0.05). Amongst the siblings and parents, possession of HLA-DR4 was associated with lower percentages of CD4+ and higher percentages of CD8+ T cells. These findings extend current knowledge about the role of immunoregulatory CD45RA/ RO cells in Type 1 diabetes. In addition, they demonstrate lymphocyte subset abnormalities in unaffected family members, some of which may be influenced by HLA DR alleles. [Diabetologia (1994) 37: 155–165] Received: 1 March 1993 and in final revised form: 16 August 1993     

Lymphocyte subpopulations in peripheral blood in primary sclerosing cholangitis

BACKGROUND/AIMS: Primary sclerosing cholangitis (PSC) is a chronic disease of autoimmunological etiology, leading to inflammation, destruction and atrophy of the bile ducts. The aim of the study was to determine peripheral lymphocyte B, T, and NK cells in PSC. METHODOLOGY: The estimation of peripheral blood lymphocytes in 17 patients (54+/-12 years old) with PSC was carried out; the control group consisted of 27 subjects (38+/-11 years). The following T lymphocyte subpopulations were determined using duo color flow cytometry: CD3+, CD4+, CD8+, CD3++HLA DR+, B cells CD19+, and NK cells CD16+ +CD56+. RESULTS: In PSC we observed doubled increase in activated T lymphocytes of CD3+ +HLA DR+ phenotype as compared to healthy subjects (7.9% vs. 2.7%, p<0.01) and NK cells (12.6% vs. 10.3%, respectively, p<0.05). There were no significant differences in the composition of CD3+, CD4+, and CD8+. In peripheral blood we noted, in patients with PSC, a decrease in B lymphocytes (11.2% vs. 12.3%, p<0.19). CONCLUSIONS: The examinations showed that activated T (HLA DR+) lymphocytes and NK cells played an important role in development of PSC.     

Reduction of leukocyte and interleukin-1β concentrations in the synovial fluid of rheumatoid arthritis patients treated with methotrexate

Objective. To examine the effect of methotrexate (MTX) on the numbers of leukocytes in the peripheral blood (PB) and synovial fluid (SF) of patients with active rheumatoid arthritis (RA). Methods. Twelve patients were treated with MTX; 5 patients not taking MTX served as controls. Samples of PB and SF were collected at 0, 1, 4, and 8 weeks of the study. Disease activity was scored, and total leukocytes, neutrophils, lymphocytes, and CD4+, CD8+, DR+, and CD25+ lymphocyte subsets were analyzed in PB and SF. Interleukin-1β (IL-1β) concentrations in SF were determined. Results. Patients treated with MTX showed significant clinical improvement. No change in PB leukocytes or lymphocyte subsets was observed in either patient group over the 8-week study period. In contrast, the number of leukocytes, the number and proportion of neutrophils, and the concentration of IL-1β in the SF of patients treated with MTX were reduced. In addition, in MTX-treated patients, there was an appreciable decrease in SF CD8+ lymphocytes, but not CD4+, DR+, or CD25+ lymphocytes. Conclusion. These findings suggest that in RA, MTX acts, at least in part, by reducing the migration of leukocytes into the inflamed synovium. Local reduction of IL-1β secretion may contribute to this effect.     

Cd28 expression on t cell subsets in vivo and cd28-mediated t cell response in vitro in patients with rheumatoid arthritis

Objective. In view of the critical importance of the CD28–CD80 (B7/BB1) costimulatory pathway in antigen-specific T cell activation and clonal expansion, we examined CD28 surface molecule expression in vivo, and T cell receptor/CD3-mediated and B7/BB1-costimulated T cell proliferation in vitro, in rheumatoid arthritis (RA). Methods. Two-color immunofluorescence analyses of peripheral blood and synovial fluid–derived T cells, as well as ³H-thymidine incorporation assays, were performed. Results. In the peripheral blood of 31 patients with active, untreated RA, a mean of 91% (range 48–100%) of CD4+ and 46% (range 13–82%) of CD8+ T cell subsets were CD28+, which was not significantly lower than normal. Although an overall decrease in the number of T cells was not observed, the numbers of CD28+CD8+ T cells were significantly lower in RA patients (mean 233/μl, versus 292/μl in controls), and this decrease was more pronounced in patients with severe disease (mean 172/μl). CD28 expression on peripheral CD8+ T cells in RA patients, but not in healthy individuals, correlated inversely with T cell activation as assessed by HLA–DR antigen expression. In contrast to the peripheral blood, RA synovial fluid T cells were almost exclusively CD28+, suggesting that migration of CD28+CD8+ T cells to active sites of inflammation may occur. In vitro proliferative responses of peripheral blood T cells to B7/BB1 costimulation in the presence of mitogenic doses of anti-CD3 monoclonal antibody were identical in patients with RA and healthy individuals. Conclusion. Functionally intact CD28+ T cells may contribute to the abnormal immunoregulation and joint inflammation in RA.     

The intrinsic migratory capacity of memory T cells contributes to their accumulation in rheumatoid synovium

Objective. Mechanisms controlling the infiltration of T cells into rheumatoid synovium have not been fully characterized. These studies were undertaken to investigate the relationship between T cell phenotype and migratory capacity, so as to elucidate mechanisms that might contribute to the accumulation of T cells at inflammatory sites. Methods. The characteristics of in vivo migrating cells were studied by dual-immunofluorescence FACS (fluorescence-activated cell sorter) analysis of rheumatoid synovial and peripheral blood T cells. Migratory cells were also characterized using a recently developed in vitro assay, wherein peripheral blood T lymphocytes (PBTL) with the capacity to migrate through endothelial cell monolayers were retrieved and assessed. Results. Migratory CD4+ T cells from rheumatoid arthritis (RA) and normal individuals were characterized as being CD45RA-, CD29^bright, CD11a^bright, L-selectin-, CD54+, and CD58+. Migrating RA PBTL (compared with normal PBTL), however, were significantly enriched in activated HLA—DR+ T cells. RA synovial tissue lymphocytes exhibited a similar phenotype, but with decreased surface density of CD4 and an increase in HLA-DR and VLA-1. RA synovial lymphocytes exhibited a 2–3-fold increase in migratory capacity over normal and RA PBTL. Conclusion. These studies demonstrate the inherent migratory proficiency of CD4+ T cells that express a memory phenotype (CD29^bright, CD11a^bright, and CD58+). In addition, enhanced transendothelial migration was observed for CD4+ T cells that were CD54+ and L-selectin—. These studies demonstrate that the migratory patterns of circulating lymphocytes may be correlated with their surface phenotype and that the intrinsic migratory capacity of memory T cells is one component contributing to their accumulation in the rheumatoid synovium.     

The large granular lymphocyte syndrome with rheumatoid arthritis. immunogenetic evidence for a broader definition of felty's syndrome

Objective. To assess whether the HLA—DR4 association found in rheumatoid arthritis (RA) is also seen in the large granular lymphocyte (LGL) syndrome. Methods. HLA—DR genotyping was performed using restriction fragment length polymorphism and polymerase chain reaction analysis. Results. LGL syndrome patients with RA showed the same HLA–DR4 association seen in RA/Felty's syndrome (FS), while LGL syndrome patients without arthritis did not. Conclusion. It is proposed that FS and the LGL syndrome represent different variants of a broader syndrome comprising RA, neutropenia, LGL expansions, HLA—DR4 positivity, and splenomegaly.     

Peripheral blood T,B, and NK cells in relation to histological hepatitis activity and fibrosis stage in chronic hepatitis C

BACKGROUND/AIMS: Many data on the pathogenesis of chronic hepatitis C have pointed to host's immune system disorders and a high variety of virus. However, there are no known criteria that could prognose the course of chronic hepatitis C infection. The analysis of T and B lymphocyte subpopulations in the peripheral blood was undertaken in patients with chronic hepatitis C of more than 6 months of duration. METHODOLOGY: Fluorescein isothiocyanate or phycoerythryne conjugated monoclonal antibodies for CD3+, CD4+, CD8+, CD19+, CD3++ HLA DR+, CD16++ CD56+ were used. The correlation between histological hepatitis activity and fibrosis (according Scheuer's scale) and the distribution of lymphocytes in the peripheral blood was sought. RESULTS: All patients with chronic hepatitis showed statistically significant increase in active lymphocytes CD3++ HLA DR+ and CD16++ CD56+ NK cells in peripheral blood. We observed the correlation between these cells and histological hepatitis activity and fibrosis. There was no correlation between the value of CD3+ and CD8+ cells and the stage of liver failure. In the early stage of chronic hepatitis C we noted decrease CD4+ cells with increase B cells CD19+. CD4+/CD8+ ratio was maintained as slightly decreased in chronic hepatitis C in favor of lymphocytes CD8+. CONCLUSIONS: The results show the correlation between peripheral blood value of activated T cell (HLA DR+) and NK cells with histological activity and fibrosis in chronic hepatitis C. Lymphocyte T (CD4+, CD8+) and B (CD19+) did not correlate with grade and stage of hepatitis C.     

Immunological response in chronic hepatitis C virus infection during interferon alpha therapy

BACKGROUND/AIMS: Chronic hepatitis C infection is a serious therapeutic problem. Interferon therapy is one of the possible methods leading to HCV infection elimination in some patients. The aim of the study was the estimation of administration of interferon alpha 2a and its effect on the lymphocyte subpopulations in peripheral blood in patients with chronic hepatitis C. METHODOLOGY: A cytometric analysis was completed concerning the level of CD19+, CD3+, CD4+, CD8+, CD4+ + HLA DR+, CD16+ + 56+ in 21 patients in the 0, 4, 24, 48 weeks of interferon alpha 2a treatment, with the dose of 18 MU/week/48 weeks. RESULTS: Virus elimination was obtained in 26% of patients (responders, R) and a higher CD8+ level and a decrease in CD4+ was observed during interferon administration in those patients. A slight increased NK cell level was noticed mainly in patients who did not eliminate HCV (non-responders, NR). In R patients, lower lymphocyte values of CD19+, NK, CD3+ + HLA DR+, and CD4+ were observed in comparison to NR, which could suggest that they play a role in the process of HCV elimination. Significant immunological changes in peripheral blood were observed mainly in the first 4 weeks of interferon alpha therapy. CONCLUSIONS: Our studies did not reveal whether any of examined lymphocyte subpopulations in peripheral blood could be considered as a predictive factor of a positive reaction to interferon alpha treatment. However, there are significant differences in the levels of lymphocytes in R and NR.     

类风湿关节炎患者滑膜树突状细胞的研究

目的　探讨滑膜树突状细胞在类风湿关节炎 (RA)中的作用。方法　采用免疫组织化学方法观察 2 5例RA滑膜中HLA DR抗原阳性表达细胞、S 10 0阳性CD1a阳性的树突状细胞(DC)的多少和分布情况。结果　 2 3例RA滑膜细胞HLA DR表达阳性 ,19例S 10 0表达阳性 ,16例CD1a表达阳性 ,其阳性例数的百分率均高于对照组。RA滑膜中HLA DR、S 10 0和CD1a标记阳性细胞数均明显高于对照组 ,其差异有非常显著意义 (P <0 0 1)。结论　RA滑膜中DC和被激活的DC及HLA DR阳性细胞的存在和增多可能在RA病变局部递呈自身抗原的过程中起重要的作用 ,是RA自身免疫异常的重要的始发因素     

Changes in CD4+ T Lymphocyte Subsets in Circulating Blood and Synovial Fluid Following Filtration Leukocytapheresis Therapy in Patients with Rheumatoid Arthritis

The purpose of this study is to determine the changes in CD4+ T lymphocyte subsets in the circulating blood and synovial fluid following filtration leukocytapheresis (LCP) therapy for patients with rheumatoid arthritis (RA). A Cellsorba column packed with polyester fibers was used for the removal of circulating leukocytes. For patients with RA, filtration LCP or sham procedures were performed 3 times with 1 week intervals between procedures. T lymphocyte surface markers in the peripheral blood and synovial fluid were measured by flow cytometry. The proportions of activated CD4+ T cells (CD4+DR+, CD4+CD25+, and CD4+CD71+) and CD4+CD29+ T cells increased significantly in the peripheral blood, but the counts of these cells were significantly reduced in the synovial fluid after 2 treatment sessions in the LCP group. No significant changes were observed in the proportion of these cells in the control group. Our findings suggest that filtration LCP may cause a redistribution of activated T cells from affected joints into the circulating blood.     

High percentage of CD8+, Leu-7+ cells in rheumatoid arthritis synovial fluid

Objective. While analyzing the phenotype of the synovial fluid mononuclear cells (SFMC) clustered about dendritic cells in rheumatoid arthritis (RA) joint effusions, it was noted that most of the clustering cells were CD8+ and coexpressed Leu-7. Therefore, the present study was conducted to investigate the frequency of CD8+, Leu-7+ cells in RA SF. Methods. SFMC from 13 patients with RA and from 12 patients with non-RA inflammatory arthritides were examined for CD8 and Leu-7 expression using 2-color immunofluorescence flow cytometry. Results. RA SFMC had statistically significantly greater percentages of total CD8+ cells, total Leu-7+ cells, and CD8+, Leu-7+ cells, compared with SFMC from the non-RA patients. These RA CD8+, Leu-7+ SFMC had a distinctive electron microscopic appearance compared with CD8+, Leu-7– SFMC. When peripheral blood mononuclear cells (PBMC) from 31 RA patients (including 7 from the SFMC group) were compared with PBMC from 15 normal controls, the percentage of CD8+, Leu-7+ cells was not significantly greater in the RA patients. However, the combination of a modest increase in CD8+, Leu-7+ cells and a decrease in total CD8 cells in RA PBMC altered the composition of the RA CD8 population compared with normal PBMC, such that over 40% of RA peripheral blood CD8 cells coexpressed Leu-7. Conclusion. The increased frequency of CD8+, Leu-7+ cells in RA SFMC may arise from the fact that a high percentage of the CD8+ PBMC in RA patients are also Leu-7+. This altered composition of CD8 cells in RA SF may have a role in the pathogenesis of the disease.     

Accelerated generation of CD14+ monocyte-lineage cells from the bone marrow of rheumatoid arthritis patients

Objective. To examine the capacity of bone marrow progenitor cells to generate CD14+ cells, in order to assess the role of bone marrow in the pathogenesis of rheumatoid arthritis (RA). Methods. CD14- cells purified from bone marrow specimens of 11 patients with active RA and 8 control patients (osteoarthritis or trauma) were cultured in the presence or absence of granulocyte–macrophage colony-stimulating factor (GM-CSF; 100 pg/ml). After incubation for various lengths of time, the cells were analyzed by flow cytometry for expression of CD14 and HLA-DR. Results. The spontaneous generation of CD14+ cells from bone marrow CD14- progenitor cells was accelerated in RA patients compared with control patients. Moreover, the expression of HLA–DR on the bone marrow–derived CD14+ cells was also accelerated in RA patients compared with controls. GM-CSF significantly enhanced the generation of CD14+ cells, as well as the expression of HLA–DR, on CD14+ cells of control patients, but not those of RA patients. GM-CSF levels in the culture supernatants of bone marrow CD14- cells were not significantly different between RA patients and control patients (undetectable in most cases). Conclusion. These observations strongly support the hypothesis that the accelerated generation of CD14+ cells from bone marrow progenitor cells and the accelerated maturation of such CD14+ cells into HLA–DR+ cells play an important role in the pathogenesis of RA. Moreover, the data suggest a functional alteration of RA bone marrow CD14- cells in their responsiveness to GM-CSF.     

CD7– T cells in rheumatoid arthritis

Objective. Rheumatoid arthritis (RA) is characterized by decreased expression of CD7 in the peripheral blood and in the synovium. The present study was designed to identify the basis for and functional consequences of this decreased expression. Methods. Peripheral blood lymphocytes from normal controls and from patients with RA or systemic lupus erythematosus (SLE), and T cell lines derived from rheumatoid synovium, were evaluated using 3-color fluorescence-activated cell sorter analysis. Results. Normal subjects and most SLE patients expressed homogeneous, bright CD7 on CD4+, CD45RA+ cells, whereas RA patients demonstrated a significantly increased proportion of CD7– cells. T cell lines derived from rheumatoid synovium demonstrated a striking deficiency of CD7 on CD4+, CD45RA– cells. CD4+, CD45RA+ cells from RA patients changed phenotype after in vitro activation to CD45RA negativity, with up-regulation of CD7. CD7–, CD4+, CD45RA– cells were assessed for their ability to induce pokeweed mitogen-driven IgM and IgM-rheumatoid factor synthesis, and they were found to be potent helper/inducer cells. An increased population of CD7-, CD4+ cells in peripheral blood was found to predict a low response to recall antigens. Conclusion. The low expression of CD7 in RA may explain some of the immune abnormalities which may contribute to the pathogenesis of this disease.     

Local anti—type ii collagen antibody production in rheumatoid arthritis synovial fluid

Objective. To investigate the production of type II collagen (CII) antibodies in the synovial fluid (SF) of rheumatoid arthritis (RA) patients, and to examine the HLA dependence of this local production. Methods. The ELISPOT method was used for enumerating anti-CII—reactive cells. Serologic tissue typing was performed. Results. Anti-CII—reactive cells were found in the SF of 16 of 31 patients, but not in any of the peripheral blood samples obtained in parallel. SF anti-CII antibody production showed no correlation with clinical parameters, but its frequency increased significantly with age. The IgG anti-CII response occurred exclusively in patients who were positive for HLA—DR4 and was significantly associated with DR4. Conclusion. Anti-CII production may be important in local immune complex formation. The indirect demonstration of a DR4-restricted T cell response to CII is an indication of a pathogenetic role of collagen autoimmunity in RA.     

Cytokines in chronic inflammatory arthritis. III. Rheumatoid arthritis monocytes are not unusually sensitive to γ-interferon,but have defective γ-interferon–mediated HLA–DQ and HLA–DR induction

Macrophages present in the synovium and synovial fluid of patients with rheumatoid arthritis (RA) express large amounts of HLA–DR molecules on their surface, despite low levels of γ-interferon (γ-IFN) in the joint. To determine whether this apparent paradox is the result of increased sensitivity to γ-IFN in RA, we compared concentrations of γ-IFN that induced HLA–DR and DQ on peripheral blood monocytes of RA patients and normal donors, using fluorescence-activated cell sorter analysis. Among normal donors, highly variable sensitivity to γ-IFN was observed. Higher amounts of γ-IFN were required to induce class II major histocompatibility complex molecules on RA monocytes versus normal monocytes. The maximum amount of HLA–DR that could be induced on RA and normal monocytes was similar; however, peak levels of HLA–DQ were significantly less in RA. Monocytes from patients with other forms of chronic inflammatory arthritis had intermediate HLA–DQ expression after γ-IFN treatment. These data suggest that an increased sensitivity to γ-IFN in RA does not account for the high level of HLA–DR expression in the joint. Also, a defect in HLA–DQ and HLA–DR induction by γ-IFN was observed.     

klebsiella pneumoniae-reactive t cells in blood and synovial fluid of patients with ankylosing spondylitis comparison with hla–b27 + healthy control subjects in a limiting dilution study and determination of the specificity of synovial fluid t cell clones

Objective. To study the frequency of Klebsiella pneumoniae–responsive T cells in the peripheral blood (PB) of ankylosing spondylitis (AS) patients compared with that in healthy HLA–B27+ donors, and to examine T lymphocyte clones (TLC) derived from AS patient synovial fluid (SF) for the presence of Klebsiella reactivity. Methods. Limiting dilution analysis of PB T cells in 8 patients with active AS and in 8 HLA–B27+ healthy subjects was used to determine the frequency of PB T cells responsive to K pneumoniae and Escherichia coli GroEL. SF T cells from a patient with active AS were cloned, and 125 TLC were characterized in proliferation assays. Results. There were fewer T cells in the PB of AS patients that reacted with K pneumoniae than in the PB of healthy HLA–B27+ subjects. The frequencies of E coli GroEL–responsive T cells were ∼5–10 times lower in all subjects tested (healthy donors and AS patients), but without significant differences between the 2 groups. Two CD4+ TLC that recognized K pneumoniae (1 cross-reactive with E coli) as well as 3 TLC that recognized GroEL (2 CD4+, 1 T cell receptor γ/δ+) were isolated from the SF of a patient with active AS. Conclusion. Our results indicate that there is a quantitative reduction of K pneumoniae-responsive T cells in the PB of AS patients as compared with healthy controls. This may reflect a defective peripheral T cell defense in the immune response to Klebsiella and may allow bacterial antigens to reach the synovium, where they initiate specific T cell responses.     

Proinflammatory mediator–induced reversal of CD4+,CD25+ regulatory T cell–mediated suppression in rheumatoid arthritis

Objective

We previously demonstrated that CD4+,CD25+ regulatory T (Treg) cells are present in increased numbers in the synovial fluid (SF) of rheumatoid arthritis (RA) patients and display enhanced suppressive activity as compared with their peripheral blood (PB) counterparts. Despite the presence of these immunoregulatory cells in RA, chronic inflammation persists. The purpose of the present study was to investigate whether particular proinflammatory mediators that are associated with RA could abrogate CD4+,CD25+ Treg–mediated suppression.

Methods

Monocyte phenotype was determined by flow cytometry and cytokine levels by enzyme‐linked immunosorbent assay. Magnetically sorted CD4+,CD25– and CD4+,CD25+ T cells derived from the PB and SF obtained from RA patients were stimulated alone or in coculture with anti‐CD3 monoclonal antibody (mAb) and autologous antigen‐presenting cells, in the absence or presence of anti‐CD28 mAb or the proinflammatory cytokines interleukin‐6 (IL‐6), tumor necrosis factor α (TNFα), or IL‐7.

Results

Monocytes from the SF of RA patients displayed increased expression of HLA class II molecules, CD80, CD86, and CD40 as compared with PB‐derived monocytes, indicating their activated status. Mimicking this increased costimulatory potential, addition of anti‐CD28 mAb to cocultures of CD4+,CD25– and CD4+,CD25+ T cells resulted in reduced CD4+,CD25+ Treg–mediated suppression in both PB and SF. Furthermore, IL‐7 and, to a limited extent, TNFα, both of which are produced by activated monocytes and were detected in SF, abrogated the CD4+,CD25+ Treg–mediated suppression. In contrast, IL‐6 did not influence Treg‐mediated suppression.

Conclusion

Our findings suggest that the interaction of CD4+,CD25+ Treg cells with activated monocytes in the joint might lead to diminished suppressive activity of CD4+,CD25+ Treg cells in vivo, thus contributing to the chronic inflammation in RA.