Thomas Häupl

Autoregulation of Th1-mediated inflammation by twist1

The basic helix-loop-helix transcriptional repressor twist1, as an antagonist of nuclear factor κB (NF-κB)–dependent cytokine expression, is involved in the regulation of inflammation-induced immunopathology. We show that twist1 is expressed by activated T helper (Th) 1 effector memory (EM) cells. Induction of twist1 in Th cells depended on NF-κB, nuclear factor of activated T cells (NFAT), and interleukin (IL)-12 signaling via signal transducer and activator of transcription (STAT) 4. Expression of twist1 was transient after T cell receptor engagement, and increased upon repeated stimulation of Th1 cells. Imprinting for enhanced twist1 expression was characteristic of repeatedly restimulated EM Th cells, and thus of the pathogenic memory Th cells characteristic of chronic inflammation. Th lymphocytes from the inflamed joint or gut tissue of patients with rheumatic diseases, Crohns disease or ulcerative colitis expressed high levels of twist1. Expression of twist1 in Th1 lymphocytes limited the expression of the cytokines interferon-γ, IL-2, and tumor necrosis factor-α, and ameliorated Th1-mediated immunopathology in delayed-type hypersensitivity and antigen-induced arthritis.

Quantitative determination of the diagnostic accuracy of the synovitis score and its components

Slansky E, Li J, Häupl T, Morawietz L, Krenn V & Pessler F
(2010) Histopathology 57, 436–443
Quantitative determination of the diagnostic accuracy of the synovitis score and its components

Influence of peptidylarginine deiminase type 4 genotype and shared epitope on clinical characteristics and autoantibody profile of rheumatoid arthritis

Background: Recent evidence suggests that distinction of subsets of rheumatoid arthritis (RA) depending on anti-cyclic citrullinated peptide antibody (anti-CCP) status may be helpful in distinguishing distinct aetiopathologies and in predicting the course of disease. HLA-DRB1 shared epitope (SE) and peptidylarginine deiminase type 4 (PADI4) genotype, both of which have been implicated in anti-CCP generation, are assumed to be associated with RA. Objectives: To elucidate whether PADI4 affects the clinical characteristics of RA, and whether it would modulate the effect of anti-CCPs on clinical course. The combined effect of SE and PADI4 on autoantibody profile was also analysed. Methods: 373 patients with RA were studied. SE, padi4_94C>T, rheumatoid factor, anti-CCPs and antinuclear antibodies (ANAs) were determined. Disease severity was characterised by cumulative therapy intensity classified into ordinal categories (CTI-1 to CTI-3) and by Steinbrocker score. Results: CTI was significantly associated with disease duration, erosive disease, disease activity score (DAS) 28 and anti-CCPs. The association of anti-CCPs with CTI was considerably influenced by padi4_94C>T genotype (C/C: ORadj = 0.93, padj = 0.92; C/T: ORadj = 2.92, padj = 0.093; T/T: ORadj = 15.3, padj = 0.002). Carriage of padi4_94T exhibited a significant trend towards higher Steinbrocker scores in univariate and multivariate analyses. An association of padi4_94C>T with ANAs was observed, with noteworthy differences depending on SE status (SE−: ORadj = 6.20, padj<0.04; SE+: ORadj = 0.36, padj = 0.02) and significant heterogeneity between the two SE strata (p = 0.006). Conclusions: PADI4 genotype in combination with anti-CCPs and SE modulates clinical and serological characteristics of RA.

Fibrinogen and factor XIII A-subunit genotypes interactively influence C-reactive protein levels during inflammation

Objective Fibrinogen is a target of autoimmune reactions in rheumatoid arthritis (RA). Fibrin(ogen) derivatives are involved in inflammatory processes and the generation of a stable fibrin network is necessary for sufficient inflammation control. As the density and stability of fibrin networks depend on complex interactions between factor XIIIA (F13A) and fibrinogen genotypes, the authors studied whether these genotypes were related to C-reactive protein (CRP) levels during acute-phase reactions. Methods Association between α-fibrinogen (FGA), β-fibrinogen (FGB) and F13A genotypes with CRP levels was tested in two cohorts with longitudinal CRP measurements. Discovery and replication cohorts consisted of 288 RA (913 observations) and 636 non-RA patients (2541 observations), respectively. Results Genotype FGB −455G>A (rs1800790) was associated with CRP elevations (≥10 mg/l) in both cohorts (RA, OR per allele 0.69, p=0.0007/Padj<0.015; non-RA, OR 0.70, p=0.0004/padj<0.02; combined, OR 0.69, p<10−5/padj=0.001). Genotype F13A 34VV (rs5985) was conditional for the association of FGB −455G>A with CRP as indicated by a clear restriction on F13A 34VV individuals and a highly significant heterogeneity between F13A 34VV and F13A 34L genotypes (p<10−5, padj=0.001). In both cohorts, mean CRP levels significantly declined with ascending numbers of FGB −455A alleles. Genotype FGA T312A (rs6050) exhibited opposite effects on CRP compared with FGB −455G>A. Again, this relation was dependent on F13A V34L genotype. Conclusion Novel genetic determinants of CRP completely unrelated to previously known CRP regulators were identified. Presumably, these haemostatic gene variants modulate inflammation by influencing fibrin crosslinking. These findings could give new perspectives on the genetic background of inflammation control.

Expression analysis of rheumatic diseases, prospects and problems

Gene expression profiling provides a completely new approach to rheumatology research. As an interdisciplinary technology, it has stimulated fruitful collaboration between experts in array technology, bioinformatics, immunology and rheumatology. The molecular overview given by genome-wide profiles has revealed that many problems arise and demand systematic and structured generation of expression data. This helps to dissect the complexity of cellular mixtures in clinical samples and may also contribute to identify functional components to enable comprehensive interpretation of profiles from each patient individually. This will provide a deeper understanding in the molecular mechanisms of rheumatic diseases and advance our effort in an optimised and individualised antirheumatic therapy.

Differential immunological response of patients with rheumatoid arthritis towards two different Epstein-Barr virus strains: inhibition of interleukin-1 release by the B95-8, but not the P3HR-1 virus strain.

SummaryAn abnormal immune response towards the Epstein-Barr virus (EBV) has been documented in patients with rheumatoid arthritis (RA). To investigate whether these findings are due to the transformation event caused by EBV, RA blood mononuclear cells and monocyte-enriched preparations were incubated with two different EBV strains: the transforming virus secreted by the cell line B95-8 and the virus released by the P3HR-1 cell line that is not able to transform due to a small deletion in the U2 region of the virus genome. Immunological response was determined by the production of interleukin-1 (IL-1) and tumor necrosis factor (TNF) using ELISA and bioassay systems. There was a striking difference in cytokine measurements with a strong inhibition of IL-1 and TNF production after incubation with the B95-8 virus, but not the P3HR-1 virus. These data indicate that the disturbed reaction of the immune system towards EBV is either dependent on the full transformation of B cells in RA patients or alternatively due to the secretion of a cytokine inhibitor by the B95-8 cell line.

Cellular Immune Reactions to Borrelia Burgdorferi — the T-Cell-Macrophage Axis

Even though Lyme borreliosis (LB) has been clearly identified as an infectious disease, there are still many riddles to be solved, especially with regard to the immune response to this organism. These include the question why only a small proportion of individuals originally exposed to B. burgdorferi progress to later disease stages, the genetic background and the apparent dissociation between the cellular and humoral immune reactivity with an extensive, but nevertheless restricted and apparently non-protective antibody repertoire.

Sectional Uptake and Cytosolic Processing of Borrelia Burgdorferi by Human Phagocytes

As soon there was compelling evidence that the spirochete Borrelia burgdorferi was the causative agent of Lyme disease (Burgdorfer et al. 1982), the first ultrastructural studies were performed on the non-specific cellular immune response to this pathogen. The examination of ultrathin sections with the electron microscope showed the presence of B. burgdorferi cells within phagolysosomes (Benach et al. 1984a,b; Peterson 1984).

05.08 Increased turnover of monocytes in patients with rheumatoid arthritis identified by transcriptome and cytometric profiling

Background Targeting molecules involved in monocyte activation is an important treatment strategy for RA. In this study we aimed to determine monocyte maturation and activation from bone marrow (BM) via blood into synovial fluid (SF) by investigating monocytes transcriptomes and by cytometric profiling of classical (CD14++CD16-), intermediated (CD14++CD16+) and non-classical (CD14+CD16+) monocytes. Materials and methods CD14+ cells from BM and blood of RA and osteoarthritis (OA) patients were profiled with Affymetrix microarrays. A detailed functional analysis was performed with reference transcriptomes of BM precursors, monocyte blood subsets, monocyte activation and egress from BM induced by G-CSF (granulocyte colony-stimulating factor). Cytometric profiling of CD14, CD16, HLA-DR and CD163 expression were used to determine monocyte subsets and to follow their activation and differentiation in BM, blood and SF. Results Transcriptomes of RA-BM monocytes exhibited i) pronounce gene pattern of early myeloid precursors from BM and ii) weak gene pattern of late myeloid precursors from BM. Transcriptomes of RA blood monocytes demonstrated i) pattern of late myeloid precursors from BM and ii) reduced pattern of terminally differentiated CD14+CD16+ monocytes from blood. Cytometric profiling of BM, blood and SF monocytes in RA and OA showed that all three body compartments have their own distribution of monocyte subsets. BM was characterised with classical and intermediate subsets and both subsets showed decreased CD16 expression in RA when compared to OA. As expected, blood was characterised with three subsets, and RA blood showed decreased CD14 and HLA-DR expression on classical monocytes and reduced frequency of non-classical subset. In RA-SF, classical monocytes were absent, intermediate were most dominant and cell-phenotype with low CD16 expression but similar to non-classical monocytes was related to macrophages. Cell frequency of intermediate subset in SF positively correlated with inflammation (ESR; R>0.85) and showed the highest expression of HLA-DR, CD14, CD163. Conclusions Monocyte turnover is increased in RA and characterised with accelerated monocytopoiesis, faster BM egress and migration into inflamed joints. Permanent monocyte activation in the joint and their role in linking innate and adaptive immunity, which is targeted by biologics, emphasises their high diagnostic value and relevance for therapeutic stratification.

Monocytes transcriptom from sle and ra patients revelaed the disease-specific imprint of TNFα, IFNα and IFNγ

Background and objectives Many cytokines are involved in the pathogenesis of chronic rheumatic diseases like systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA). To estimate the role of cytokines in these diseases is intriguing considering the fact that they act together within complex cytokine networks. Although various cell types produce cytokines, monocytes are considered as their principal source. Nevertheless, how functions of monocytes are altered in the course of different rheumatic diseases and how cytokines influence these alterations remained largely unknown. Materials and methods The global gene-expression profiling has been utilised for development and analyses of SLE and RA profiles (Affymetrix HG-arrays). To decipher the alterations in SLE and RA in more details, cytokine-specific signatures were developed following monocytes stimulation in vitro by tumour necrosis factor α (TNFα), IFNα2a and interferon γ (IFNγ). Ingenuity pathway analysis were utilised for identifying the molecular pathways and cytokine networks within SLE and RA profiles. Whole-genome rVISTA was applied for the promoter analyses of these profiles. Results Monocytes from SLE and RA patients revealed diseases-specific gene-expression profiles. Ingenuity pathway analysis of SLE and RA profiles identified cytokines as essential players in shaping profiles from diseases and the alterations within IFNα/IFNγ (IFNs) and TNFα signalling pathways. To decipher the SLE and RA profiles in more details, TNFα−, IFNα2a− and IFNγ−reference signatures were generated in vitro. Comparisons between disease-specific and the in vitro-generated reference signatures showed that the SLE profile was predominantly driven by IFNs, while the RA profile was primarily influenced by TNFα. The IFN-imprints in SLE was characterised by an activation of the transcription factor STAT1. Interestingly, the activation of STAT1 was found to be silenced by TNFα in patients with RA. However, the IFN-imprints were also identified in RA and the TNFα-imprint was evident in SLE. It was obvious that the responses to the same cytokines in SLE and RA were qualitatively and quantitatively different. Conclusions Altogether, this study has demonstrated that monocytes from RA and SLE patients exhibit disease-dependent gene-expression profiles, which can be molecularly dissected when compared to in vitro-generated cytokine-signatures. The IFNs- and TNFα-imprints were identified to be disease-dependent and principally they reflected the interplay of cytokines within various inflammatory milieus. The results from this study suggest that estimating the imprints of cytokines in rheumatic diseases would be indispensible for an improvement of diagnosis, proper selection of particular cytokine target(s) for therapeutic intervention and for following up and predicting the response to anticytokine drug(s).