Macrophage activation syndrome triggered by active systemic lupus erythematosus : Successful treatment by interleukin-1 inhibition (anakinra).

This article presents a case of fulminant macrophage activation syndrome (MAS) as a rare complication of active systemic lupus erythematosus in a 33-year-old female patient. Initial presentation showed severe lupus disease exacerbation with renal involvement, hemolytic anemia, and neuropsychiatric changes. Early therapy focused on broad immunosuppression (high-dose corticosteroids and cyclophosphamide); however, disease remission could not be achieved. After an additional inflammatory focus and underlying malignancy were excluded, the triplet of pancytopenia, fever, and high ferritin levels indicated MAS, a bone marrow biopsy confirmed secondary hemophagocytic histiocytosis. Treatment with an interleukin­1 antagonist (anakinra) induced a fast, effective therapeutic success.

Adherence and health literacy as related to outcome of patients treated for rheumatoid arthritis : Analyses of a large-scale observational study.

BACKGROUND: Disabilities in daily living and quality of life are key endpoints for evaluating the treatment outcome for rheumatoid arthritis (RA). Factors possibly contributing to good outcome are adherence and health literacy. METHODS: The survey included a representative nationwide sample of German rheumatologists and their patients with RA. The physician questionnaire included the disease activity score (DAS28) and medical prescriptions. The patient questionnaire included fatigue (EORTC QLQ-FA13), health assessment questionnaire (HAQ), quality of life (SF-12), health literacy (HELP), and patients' listings of their medications. Adherence was operationalized as follows: patient-reported (CQR5), behavioral (concordance between physicians' and patients' listings of medications), physician-assessed, and a combined measure of physician rating (1 = very adherent, 0 = less adherent) and the match between physicians' prescriptions and patients' accounts of their medications (1 = perfect match, 0 = no perfect match) that yielded three categories of adherence: high, medium, and low. Simple and multiple linear regressions (controlling for age, sex, smoking, drinking alcohol, and sport) were calculated using adherence and health literacy as predictor variables, and disease activity and patient-reported outcomes as dependent variables. RESULTS: 708 pairs of patient and physician questionnaires were analyzed. The mean patient age (73% women) was 60 years (SD = 12). Multiple regression analyses showed that high adherence was significantly associated with 5/7 outcome variables and health literacy with 7/7 outcome variables. CONCLUSION: Adherence and health literacy had weak but consistent effects on most outcomes. Thus, enhancing adherence and understanding of medical information could improve outcome, which should be investigated in future interventional studies.

Optimized testing for C. trachomatis DNA in synovial fluid samples in clinical practice.

AIM: No standardized polymerase chain reaction (PCR) assay is available for detection of Chlamydia trachomatis (C. tr.) in synovial fluid (SF) for diagnostic use in clinical practice. This study tested the performance of two optimized molecular biology methods, to determine which is best suited for detecting C. tr. in SF clinical samples from patients with various rheumatologic diseases. METHODS: Two DNA extraction methods, i.e., (1) alkaline lysis and (2) QIAEX II Gel Extraction Kit® + cetyltrimethylammonium bromide (CTAB; Qiagen, Hilden, Germany), and C. tr.-omp1-152 bp PCR were tested in SF samples from a total of 329 patients with the following diagnoses: reactive arthritis (ReA; n = 10, 4 patients had posturethritic ReA), undifferentiated arthritis (UA; n = 66), rheumatoid arthritis (RA; n = 169), psoriatic arthritis (PSA; n = 12), and osteoarthritis (OA) n = 72. RESULTS: In SF samples, C. tr.-omp1-152 bp PCR in combination with alkaline lysis DNA extraction allowed detection of more C. tr.-positive samples: 3/10 (30%) ReA patients (all with posturethritic ReA) and 20/66 (38%) UA patients were positive, compared to the 0/10 (0%) patients with ReA and 1/66 (2%) with UA detected using the QIAEX II Gel Extraction Kit® + CTAB. Moreover, 2/12 (17%) SF samples from PSA patients tested positive with alkaline lysis. All samples from patients with OA and RA tested negative. CONCLUSION: Alkaline lysis in combination with C. tr.-omp1-152 bp PCR emerged as the most sensitive method for identification of C. tr. in clinical SF samples.

Involvement of neurotrophins and their receptors in spondyloarthritis synovitis: relation to inflammation and response to treatment.

OBJECTIVE: To investigate whether expression of the four members of the neurotrophin (NT) family and their four corresponding receptors is related to synovial inflammation in patients with spondyloarthritis (SpA). MATERIAL AND METHODS: Synovial fluid (SF) and serum NTs and their receptors were measured by ELISA. Immunohistochemistry was used for synovial tissue biopsy specimens from patients with SpA, rheumatoid arthritis, and osteoarthritis (OA). In SpA synovium, immunoreactivity of the receptors trkA and NGFRp75 was also assessed before and after 12 weeks of treatment with the monoclonal anti-tumour necrosis factor alpha antibody, infliximab. RESULTS: mRNA transcripts of all NTs and receptors were expressed in the inflamed synovium. At the protein level, brain derived neurotrophic factor and NT-3 were significantly higher in the SF of patients with SpA than in those with OA. In contrast, ELISA of serum samples showed that the highest member in SpA was NT-4. Immunohistochemistry demonstrated that the NT receptors trkA and NGFRp75 were highly expressed in the inflamed synovium of patients with SpA, correlating with vascularity and lymphoid aggregates, respectively. Additionally, immunoreactivity of both receptors was significantly decreased after infliximab treatment. CONCLUSIONS: NTs and their receptors are expressed in inflamed peripheral joints of patients with SpA. Their expression is not constitutive but related to inflammation and they may be involved in the local disease processes.

Alpha beta but not gamma delta T cell clones in synovial fluids of patients with reactive arthritis show active transcription of tumour necrosis factor alpha and interferon gamma.

OBJECTIVE: To compare the cytokine expression profile of three CD8+, three CD4+, and three gammadelta+ T cell clones all derived from the synovial fluids of three patients with reactive arthritis (ReA). METHODS: Complementary DNA based microarrays containing the specific sequence of 56 cytokine transcripts were used for screening. Selected genes were confirmed by reverse transcriptase-polymerase chain reaction assay. RESULTS: Microarray showed that transcripts encoding for interferon gamma and tumour necrosis factor alpha were expressed by all CD8+ and CD4+ T cell clones. However, gammadelta+ T cells predominantly expressed transforming growth factor beta2 and granulocyte monocyte-colony stimulating factor. CONCLUSION: T lymphocyte clones from the joint of patients with ReA exhibit differential cytokine expression profiles. CD8+ and CD4+ T cells demonstrate a Th1 mediated profile, whereas gammadelta+ T cells show a more heterogeneous and less proinflammatory Th3 driven pattern.

Technical validation of cDNA based microarray as screening technique to identify candidate genes in synovial tissue biopsy specimens from patients with spondyloarthropathy.

OBJECTIVES: To validate the use of cDNA based microarray on synovial biopsies by analysing the experimental variability due to amplification of RNA, reproducibility of the assay, heterogeneity of the tissue, and statistical analysis. METHODS: Total RNA was extracted from three spondyloarthropathy (SpA) and three osteoarthritis (OA) synovial tissue biopsy specimens and from the peripheral blood mononuclear cells (PBMC) of four healthy donors. Exponential RNA amplification by SMART-PCR was compared with linear amplification. Reproducibility was tested by comparing different microarray systems and by performing duplicate experiments. Sample heterogeneity was assessed by comparing overall gene expression profiles, histopathology, and analysis of genes expressed in the synovium and normal PBMC. Statistical analysis using t test and Bonferroni adjustment was verified by permutation of class labels. RESULTS: Gene expression was concordant in 12/14 (86%) cytokine/chemokine genes between both microarrays and different RNA amplification systems. When one microarray system was used, expressed genes were 78-95% concordant in duplicate experiments. Gene expression profiles had a higher degree of similarity between SpA synovium than between PBMC or OA synovium despite clear histopathological differences between synovial samples. Comparison of SpA synovium with OA synovium and with PBMC yielded 11 and 18 expressed transcripts, respectively; six were shared in both comparisons. Permutations of SpA and OA samples yielded only one expressed gene in 19 comparisons. CONCLUSIONS: These data provide evidence that microarrays can be used for analysis of synovial tissue biopsies with high reproducibility and low variability of the generated gene expression profiles.

Anti-tumour necrosis factor (TNF)-alpha therapy in undifferentiated spondyloarthropathy.

The cytokine tumour necrosis factor (TNF)-alpha plays a major role in the spinal inflammatory process of spondyloarthropathy. In contrast to rheumatoid arthritis, disease modifying antirheumatic drugs have not been proved effective against inflammation and progressive ankylosis. Initial studies on TNFalpha inhibitors in ankylosing spondylitis are promising and raise the question as to whether early stages of the disease, mostly classified as "undifferentiated spondyloarthropathy" (uSpA), should also be treated with TNFalpha inhibitors. This article summarises the preliminary results of 11 uSpA patients in 4 different trials treated with TNFalpha inhibitors.

A 588-gene microarray analysis of the peripheral blood mononuclear cells of spondyloarthropathy patients.

OBJECTIVES: To identify genes which are more highly expressed in the peripheral blood mononuclear cells (PBMC) of patients with spondyloarthropathy (SpA), rheumatoid arthritis (RA) and psoriatic arthritis (PsA), in comparison to normal subjects. METHODS: A 588-gene microarray was used as a screening tool to select a panel of such genes from PBMC of these subjects and of normal subjects. Results were then validated by reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: The following genes were more highly expressed in arthritis patients than in normal subjects: macrophage differentiation marker MNDA (myeloid nuclear differentiation antigen), MRP8 and MRP14 (migratory inhibitory factor-related proteins); signalling molecules JAK3 (janus kinase 3) and MAP kinase p38 (mitogen-activated protein kinase); receptors TNFR2/p75, C-C-chemokine receptor type 1 (CCR1), C-X-C-chemokine receptor type 4 (CXCR4) and integrin beta1; and the cytokines/chemokines interleukin (IL) 1beta and IL-8. Expression of CXCR4 was unexpectedly high among all arthritis subjects. Using RT-PCR, ELISA and immunohistology, expression of stromal cell-derived factor 1 (SDF-1) was demonstrated in arthritis joints. CONCLUSIONS: The CXCR4/SDF-1 is a potential pro-inflammatory axis for RA, PsA and SpA.

Expression of host defense scavenger receptors in spondylarthropathy.

OBJECTIVE: Reactive arthritis (ReA) is postulated to be caused by a defective host defense against gram-negative bacteria. HLA-B27 could play a role in this process, but does not account for the many HLA-B27 negative patients. The objective of this study was to test the expression of 3 macrophage scavenger receptors (SRs) that are responsible for innate immunity against gram-negative bacteria: SR class A type I (SR-AI), SR-AII, and the macrophage receptor with collagenous structure (MARCO). We postulate that defects in such receptors might also contribute to the host risk factors that increase the predisposition to ReA and perhaps other subtypes of spondylarthropathy (SpA). METHODS: Peripheral blood, synovial fluid, and synovial tissue samples were obtained from patients with recent Salmonella infection, ReA, other SpA, and rheumatoid arthritis (RA). The expression of SRs receptors was assessed by semiquantitative reverse transcriptase-polymerase chain reaction. RESULTS: Evaluation of the peripheral blood mononuclear cells (PBMC) from 4 patients who were recently infected with Salmonella, showed that PBMC from 2 patients who developed ReA expressed positive levels of MARCO, while PBMC from 2 patients who recovered from infection without sequelae did not. The synovial fluid mononuclear cells (SFMC) from some ReA patients expressed MARCO, but the levels were only moderate. The level of MARCO in the SFMC from the SpA patient group was low. In marked contrast, MARCO expression was high in almost all samples of RA SFMC. These findings also extended to synovial tissues. CONCLUSION: Expression of the host defense gene MARCO was susceptible to modulation, not only during infections, but also in the inflammatory arthritis conditions RA and SpA. MARCO is a variable to be considered as a candidate factor that might contribute to ReA.

Infection of human monocyte-derived macrophages with Chlamydia trachomatis induces apoptosis of T cells: a potential mechanism for persistent infection.

Viruses can escape T-cell surveillance by infecting macrophages and thereby induce apoptosis of noninfected T cells. This ability had not been demonstrated for bacteria. We investigated whether infection of macrophages with the important human pathogen Chlamydia trachomatis can induce T-cell apoptosis. Because Chlamydia-Mycoplasma coinfection is a frequent event, the ability of Mycoplasma fermentans-infected macrophages to induce T-cell apoptosis was also studied. Infected macrophages were cocultivated with autologous T cells in different activation states. Propidium iodide-based fluorescence-activated cell sorter analysis demonstrated that macrophages infected with viable chlamydiae induced T-cell death. Apoptosis was identified as the mode of death induction by using a terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling assay. Induction of T-cell death was macrophage dependent. Incubation of T cells with infectious chlamydiae in the absence of macrophages did not lead to T-cell apoptosis. UV irradiation of chlamydiae diminished the ability to induce death. T-cell death was induced by a cell-free supernatant of infected macrophages. Not only phytohemagglutinin-preactivated T cells but also non-mitogen-preactivated T cells were susceptible to C. trachomatis-induced apoptosis. In contrast, M. fermentans infection of macrophages did not induce T-cell death. Coinfection had no additional effect. In summary, intracellular chlamydial infection of macrophages can induce T-cell apoptosis. Apoptosis induction by chlamydiae possibly explains how persistently infected macrophages escape T-cell surveillance and why the Chlamydia-specific T-cell response is diminished during persistent chlamydial infection.

Requirements for HLA-B*2705-binding peptides with special regard to the transporter associated with antigen processing (TAP).

OBJECTIVE: To refine the algorithms governing peptide presentation by HLA-B*2705 by analyzing: (i) the specificity of the human transporter associated with antigen processing (TAP) for HLA-B27 binding peptides; and (ii) the peptide binding affinity to HLA-B*2705. METHODS: TAP-translocation was measured with a labeled reporter peptide containing an N-linked glycosylation acceptor site in Streptolysin O-permeabilized cells for a panel of HLA-B27 binding peptides. Peptide binding affinity was determined by peptide-induced stabilization of empty HLA-B*2705 expressed by the TAP-deficient cell line T2-B27. RESULTS: Human TAP preferentially translocated analogues with residues leucine, isoleucine, methionine and arginine as the carboxy-terminal amino acids, whereas analogues with aspartic acid and serine were translocated poorly. The binding affinity to HLA-B*2705 of the poorly translocated aspartic acid and serine analogues was about 100-fold less compared to the parent HLA-B27 binding peptide. CONCLUSIONS: Human TAP shows considerable specificity for the C-terminus of potential HLA-B27 ligands. Nonamer peptides with aspartic acid and serine at the C-terminus are poorly translocated by the TAP and have low binding affinity for HLA-B*2705, and are therefore unlikely to become presented by HLA-B*2705.

A patient-derived cytotoxic T-lymphocyte clone and two peptide-dependent monoclonal antibodies recognize HLA-B27-peptide complexes with low stringency for peptide sequences.

HLA-B27 molecules expressed on the T2 mutant cell line do not have peptides. Such empty HLA-B27 molecules were not recognized by an HLA-B27-restricted cytotoxic T-lymphocyte (CTL) clone (auto-1) derived from synovial fluid. To test for peptide dependency of the clone, B27-T2 cells were incubated with a panel of 48 different peptides. This lack of stringency was compared with that of a peptide-dependent monoclonal antibody, B27.M2. Positive B27.M2 reactivity resulted when the B27-T2 cells were incubated with two peptides: RRKAMFEDI and RRMGPPVGHR, derived from Chlamydia HSP60 and human ribonucleoprotein, respectively. Because of the limited availability of CTL versus monoclonal antibody, the specificity of B27.M2 was studied in greater detail. The importance of the HLA-B27 heavy chain in antibody recognition of class I-peptide complexes was demonstrated by site-directed mutagenesis. The stringency of the peptide residues was tested by making analogs of each of the nine residues in RRKAMFEDI, creating a panel of 180 analogs. Although stringency was highest for the sixth position, as many as six different amino acids provided positive reactivity. These results indicate that immune recognition of HLA-B27-peptide complexes might have rather low stringency for the peptide sequences. In theory, then, pathogen-derived peptides which induce autoimmunity by generating autoreactive CTL might not share much sequence similarity with the responsible self peptides.

Sensitivities of PCR, MicroTrak, ChlamydiaEIA, IDEIA, and PACE 2 for purified Chlamydia trachomatis elementary bodies in urine, peripheral blood, peripheral blood leukocytes, and synovial fluid.

Routine microbiological diagnosis of Chlamydia-induced reactive arthritis is based mainly on the detection of Chlamydia trachomatis with urogenital swabs or in urine. Because chlamydial antigen, rRNA, and DNA are present in low quantities in the inflamed joint, highly sensitive assays are needed to detect C. trachomatis not only at the primary site of infection but also in peripheral blood and peripheral blood leukocytes, which are suspected carriers for dissemination, and in synovial fluid. To evaluate possible tools for this purpose, the sensitivities of PCR, MicroTrak, Chlamydia EIA, IDEIA, and PACE 2 for the detection of defined numbers of purified C. trachomatis elementary bodies (EB) in urine, peripheral blood, peripheral blood leukocytes, and synovial fluid were determined. In urine, PCR detected 2, MicroTrak and ChlamydiaEIA detected 2 x 10(3), and PACE 2 and IDEIA detected 2 x 10(4) EB per ml. In peripheral blood, only PCR and MicroTrak detected C. trachomatis, with detection limits of 100 and 2 x 10(7) EB per ml, respectively. For peripheral blood leukocytes, the detection limits were 2 EB per ml for PCR and 2 x 10(4) EB per ml for MicroTrak, ChlamydiaEIA, IDEIA, and PACE 2. In synovial fluid, PCR detected 200, MicroTrak and IDEIA detected 2 x 10(5), and PACE 2 detected 10(6) EB per ml. ChlamydiaEIA was unable to detect 2 x 10(6) EB per ml in synovial fluid. In summary, PCR was found to be the most sensitive method. The sensitivities of the other methods tested were at least 1,000 times lower than that of PCR. PCR should therefore be considered a most promising tool for routine diagnosis of Chlamydia-induced arthritis.

TCR1+ large granular lymphocyte proliferation in rheumatoid arthritis.

The T gamma-lymphoproliferative syndrome is characterized by a proliferation of large granular lymphocytes (LGL). It is often associated with neutropenia, and in 30% of cases with rheumatoid arthritis (RA). Phenotypic analysis has demonstrated that in most cases of RA with T gamma-proliferative disease, the LGL represent T cells with a clonal rearrangement of the alpha/beta T cell receptor (TCR2). Here, three patients with gamma/delta TCR1+ LGL proliferation suffering from long-standing arthritis and neutropenia are described. The first patient with RA showed an expansion of a heterogeneous CD2+ CD16+ CD56- LGL population, of which 30% coexpressed TCR1 with V delta 1 rearrangement. The second patient with ankylosing spondylitis and RA was suffering from proliferation of TCR1+ (V gamma 9-, V delta 1-), CD2+ CD16- CD56- LGL with low coexpression of CD8. The third patient with RA was suffering from a proliferation of TCR1+ (V delta 1+, V gamma 9-) CD4- CD8- CD16- CD56- lymphocytes. On the basis of these unusual findings, the pathogenetic role of TCR1+ T cells in RA is discussed.

[Syndrome of T gamma lymphoproliferation of Fc gamma RIII- positive lymphocytes in rheumatoid arthritis]. / Syndrom einer T gamma-Lymphoproliferation Fc gamma RIII-positiver Lymphozyten bei rheumatoider Arthritis.

A T gamma-lymphoproliferative syndrome is described in a 69-year-old woman with seropositive rheumatoid arthritis. The phenotype of this non-clonally increased large granular lymphocyte (LGL) population is unusual: LGL are CD56- Fc gamma RIII+(CD16+) and 30% coexpress gamma/delta-T-cell receptor.

Activation of CD16+ effector cells by rheumatoid factor complex. Role of natural killer cells in rheumatoid arthritis.

The role of natural killer (NK) cells in rheumatoid arthritis (RA) remains unclear. A pathogenetic function of rheumatoid factors (RF) also has not been defined. In the present studies, natural killer (NK) cells were examined as a model for FC gamma receptor type III-positive (FC gamma RIII+) cells, with regard to their interaction with RF. NK cell antigen CD16 (FC gamma RIII) and CD56 expression and functional NK and antibody-dependent cell-mediated cytotoxicity (ADCC) activity were compared in peripheral blood lymphocytes and autologous synovial fluid lymphocytes (SFL) of RA patients. Peripheral blood lymphocytes and SFL showed normal CD56 expression. In contrast, both the frequency and the density of CD16 antigen were decreased in SFL. Furthermore, diminished NK cytotoxicity and a significant decrease in ADCC were observed in SF NK cells. In subsequent in vitro studies with normal fresh NK cells, it was demonstrated that IgG-containing RF complexes from RA patients induced a modulation of FC gamma RIII structure from the NK cell surface, a decrease in NK activity, and a complete loss of ADCC. When purified RF was incubated with NK-enriched cell lines from RA patients, increased transcription and subsequent production of interferon-gamma and tumor necrosis factor alpha were observed. These data suggest a direct involvement of RF complexes in the pathogenetic process of chronic inflammation in RA.

[Activation of Fc gamma receptor (CD16) positive natural killer cells by rheumatoid factors in rheumatoid arthritis]. / Aktivierung von Fc gamma-Rezeptor-(CD16-)positiven natürlichen Killerzellen durch Rheumafaktoren im Rahmen der rheumatoiden Arthritis.

Fc gamma-receptor-(CD16-)positive natural killer (NK) cells are activated by rheumatoid factor in the synovial fluid of patients with rheumatoid arthritis. This activation induces modulation of the CD16 molecule and release of IFN gamma and TNF alpha by NK cells.