Włodzimierz Maśliński

Fibroblast-Like Synoviocytes from Rheumatoid Arthritis Patients Express Functional IL-15 Receptor Complex: Endogenous IL-15 in Autocrine Fashion Enhances Cell Proliferation and Expression of Bcl-x L and Bcl-2

The hallmarks of rheumatoid arthritis (RA) are leukocytic infiltration of the synovium and expansiveness of fibroblast-like synoviocytes (FLS). The abnormal proliferation of FLS and their resistance to apoptosis is mediated, at least in part, by present in RA joints proinflammatory cytokines and growth factors. Because IL-15 exerts properties of antiapoptotic and growth factors, and is produced by RA FLS, we hypothesized that IL-15 participates in RA FLS activation. To test this hypothesis, we first examined whether RA FLS express chains required for high affinity functional IL-15R. Indeed, RA FLS express IL-15Rα at mRNA and protein levels. Moreover, we confirmed the presence of IL-2Rβ and common γ-chains. Interestingly, TNF-α or IL-1β triggered significant elevation of IL-15Rα chain at mRNA and protein levels. Next, we investigated the effects of exogenous or endogenous IL-15 on Bcl-2 and Bcl-xL expression, FLS proliferation, and apoptosis. Exogenous IL-15 enhanced RA FLS proliferation and increased the level of mRNA-encoding Bcl-xL. To test the role of endogenous IL-15 in the activation of RA FLS, an IL-15 mutant/Fcγ2a protein exerting properties of specific antagonist to the IL-15Rα chain was used. We found that blocking IL-15 biological activities using this protein substantially reduced endogenous expression of Bcl-2 and Bcl-xL, and RA FLS proliferation that was reflected by increased apoptosis. Thus, we have demonstrated that a distinctive phenotype of RA FLS, i.e., persistent activation, proliferation, and resistance to apoptosis, is related to the autocrine activation of IL-15Rs by FLS-derived IL-15.

Rottlerin, a PKC isozyme-selective inhibitor, affects signaling events and cytokine production in human monocytes.

The implication of select protein kinase C (PKC) isoenzymes in cytokine production by human monocytes was investigated using an isozyme‐selective inhibitor of PKC, rottlerin. We found that lipopolysaccharide (LPS) triggers cytosol‐to‐membrane translocation of PKCα and δ isoenzymes, whereas phorbol ester (PMA) induces translocation of several PKC isoforms. Moreover, we show that in LPS‐ and PMA‐stimulated monocytes rottlerin affects several cellular responses. (1) At low (15 μM) concentration it blocks translocation of PKCδ, diminishes DNA binding activity of AP‐1 transcription factor, and attenuates cytokine production [tumor necrosis factor α (TNF‐α) > interleukin‐1β (IL‐1β)]. (2) At high (50 μM) concentration it prevents translocation of PKCα, and subsequently inhibits ERK1/ERK2 phosphorylation, DNA binding activities of AP‐1 and nuclear factor‐κB transcription factors, and the production of both tested cytokines. Thus, we propose that cytosol‐to‐membrane translocation of PKCα and PKδ isoenzymes may represent early steps in the signaling cascades that lead to TNF‐α and IL‐1β production in human monocytes. J. Leukoc. Biol. 67: 249–258; 2000.

Taurine chloramine inhibition of cell proliferation and cytokine production by rheumatoid arthritis fibroblast‐like synoviocytes

OBJECTIVE To examine whether taurine (Tau) or its physiologic chlorinated derivative, taurine chloramine (Tau-CI), affects proliferation of, and proinflammatory cytokine (interleukin-6 [IL-6] and IL-8) production by, fibroblast-like synoviocytes (FLS) isolated from rheumatoid arthritis (RA) patients. METHODS FLS, isolated from the synovial tissue of 19 RA patients and cultured in vitro for 3-6 passages, were stimulated with the recombinant human cytokines IL-1beta (1 ng/ml), tumor necrosis factor alpha (TNFalpha; 10 ng/ml), or IL-17 (10 ng/ml) in the presence of either Tau or Tau-Cl, which were added at concentrations of 50-500 microM. Tau and Tau-Cl were added simultaneously with, 2 hours before, or 24 hours after the stimuli. The concentrations of IL-6 and IL-8 were determined in culture supernatants using specific enzyme-linked immunosorbent assays. Proliferation of FLS was estimated on the basis of 3H-thymidine incorporation into the cells, which were cultured for 72 hours in the presence of recombinant human basic fibroblast growth factor (bFGF) (1 ng/ml) and Tau or Tau-Cl, which were added simultaneously at the beginning of the culture. RESULTS Cultured in vitro, RA FLS spontaneously secreted low levels of IL-6 and IL-8, but when RA FLS were stimulated with IL-1beta, TNFalpha, or IL-17, significantly higher amounts of IL-6 and IL-8 were produced. Tau-Cl, but not Tau, inhibited cytokine-triggered synthesis of IL-6 (50% inhibitory concentration [IC50] approximately 225 microM) and IL-8 (IC50 approximately 450 microM) when added simultaneously with the stimuli. However, IL-17-induced production of IL-8 was not affected by Tau-Cl. In the cells prestimulated with IL-1beta for 24 hours, Tau-Cl still inhibited synthesis of IL-6, but did not affect IL-8 production. Moreover, Tau-Cl inhibited spontaneous and bFGF-triggered proliferation of FLS in a dose-dependent manner. Neither Tau nor Tau-Cl affected cell viability. CONCLUSION The results of these studies demonstrate that Tau-Cl inhibits production of proinflammatory cytokines by RA FLS, as well as proliferation of these cells. Thus, Tau-Cl may act as a physiologic modulator of FLS functions related to their pathogenic role in RA.

Anti-Inflammatory Effects of Taurine Derivatives (Taurine Chloramine, Taurine Bromamine, and Taurolidine) are Mediated by Different Mechanisms

In this study, in an animal model of zymosan-induced peritonitis we have tested anti-inflammatory properties of Taurolidine (TRD), a synthetic derivative of taurine. In vitro, the effect of TRD and HOCl treated TRD on peritoneal macrophages was compared with that of TauCl. We report that locally administered TRD (Taurolin) shows strong anti-inflammatory properties. TRD inhibits vascular permeability increased by inflammatory stimuli; it also significantly attenuates the influx of neutrophils into the peritoneal cavity, as well as the production of pro-inflammatory cytokines (TNF-alpha, IL-6) by peritoneal exudate cells. Chlorination of TRD resulted in the formation of chloramine (TRD-Cl), as confirmed by characteristic UV spectra. Both TRD and TRD-Cl, more effectively than TauCl, inhibited the production of IL-6 by stimulated macrophages. The effect was not dependent on its well-known anti-endotoxin activity since TRD inhibited cytokine production by macrophages stimulated with either LPS or IFN-gamma. Finally, we report that anti-inflammatory activities of TRD and taurine haloamines are mediated by different mechanisms. TRD, in contrast to TauCl and TauBr, does not induce expression of HO-1, a stress inducible enzyme with strong anti-inflammatory properties.

Anti-inflammatory Activities of Taurine Chloramine

Neutrophils recruited into the site of inflammation generate a large number of highly reactive oxidants, including hypochlorous acid (HOCl), produced by the myeloperoxidase-catalyzed oxidation of C1 by hydrogen peroxide1. Hypochlorous acid is the major neutrophil microbicidal agent but its excessive production leads to tissue damage. Oxidative tissue damage is thought to be of pathogenic significance in a large number of diseasese.g.atherosclerosis, malignancy and rheumatoid arthritis. Taurine (Tau), a dominant free amino acid present in most mammalian tissues and in the cytosol of phagocytic cells at high (10–20 mM) concentration2, acts as a major trap for HOCl. The reaction of Tau with HOCl forms the long-lived but weaker oxidant taurine chloramine (Tau-Cl) and thus reduces HOCl toxicity. On the other hand, there is growing evidence that Tau-Cl: (i) down-regulates the production of pro-inflammatory mediators by macrophages, neutrophils and other cells engaged in inflammatory response3, (ii) regulates the function of dendritic cells4, and (iii) enhances protein immunogenecity by chlorination5. This implies Tau-Cl to act as an important physiologic immunoregulatory factor maintaining the delicate balance between mounting an effective immune response, and minimizing the destruction of the tissue by the inflammatory cells.

Taurine Haloamines and Heme Oxygenase-1 Cooperate in the Regulation of Inflammation and Attenuation of Oxidative Stress

Taurine chloramine (TauCl) and Taurine bromamine (TauBr), products of the neutrophil myeloperoxidase halide system, exert anti-inflammatory properties. They inhibit the production of a variety of inflammatory mediators, such as prostaglandin E2 (PGE2), nitric oxide (NO) and proinflammatory cytokines. Heme oxygenase-1 (HO-1), a stress inducible enzyme, degrades heme to biliverdin, free iron and carbon monoxide (CO), which are involved in the anti-inflammatory and antioxidant actions of HO-1. Recently we have demonstrated that taurine haloamines induce the expression of HO-1 in inflammatory cells. In this study we examined whether HO-1 participates in taurine haloamines-mediated suppression of proinflammatory cytokine production. We have shown that TauCl/TauBr and CO inhibit the production of TNF-alpha, IL-12 and IL-6, in a similar dose-dependent manner. However, the suppressor activity of TauCl was not altered in HO-1 deficient mice. Therefore, HO-1 and TauCl may independently regulate the production of proinflammatory cytokines. We suggest that TauCl and TauBr provide a link between the two antioxidant systems: the cysteine pathway and the heme oxygenase system.

Enthesopathies and enthesitis. Part 2: Imaging studies.

The pathologies of tendon and ligament attachments are called enthesopathies. Enthesitis is one of enthesopathies and it is considered a characteristic sign of rheumatic diseases from the spondyloarthritis group, including peripheral spondyloarthritis. Therefore, enthesitis has been included in a number of clinical classifications for diagnosing these diseases. Clinical diagnosis of enthesitis is based on rather non-specific clinical signs and results of laboratory tests. It is believed that imaging examinations might improve diagnosis, particularly because numerous papers prove that differentiating enthesitis from other enthesopathic processes is possible. On the other hand, a number of authors report the lack of specific signs in imaging as well as typical histological and immunological features that would enable confirmation of clinical diagnosis of enthesitis. The first part of the publication presented theories on the etiopathogenesis of enthesitis (inflammatory, mechanical, autoimmune and associated with the synovio-entheseal complex) as well as on the formation of enthesophytes (inflammatory, molecular and mechanical). This paper – the second part of the article, is a review of the state-of-the-art on the ability of imaging examinations to diagnose enthesitis. It turns out that none of the enthesitis criteria used in imaging examinations is specific for inflammation. As enthesitis may be the only symptom of early spondyloarthritis (particularly in patients with absent HLA-B27 antigen), the lack of its unambiguous picture in ultrasound and magnetic resonance imaging prompts the search for other signs characteristic of spondyloarthritis and more specific features in imaging in order to make a diagnosis as early as possible.

Role of inflammatory factors and adipose tissue in pathogenesis of rheumatoid arthritis and osteoarthritis. Part I: Rheumatoid adipose tissue.

For many years, it was thought that synovial cells and chondrocytes are the only sources of proinflammatory cytokines and growth factors found in the synovial fluid in patients suffering from osteoarthritis and rheumatoid arthritis. Currently, it is more and more frequently indicated that adipose tissue plays a significant role in the pathogenesis of these diseases as well as that a range of pathological processes that take place in the adipose tissue, synovial membrane and cartilage are interconnected. The adipose tissue is considered a specialized form of the connective tissue containing various types of cells which produce numerous biologically active factors. The latest studies reveal that, similarly to the synovial membrane, articular adipose tissue may take part in the local inflammatory response and affect the metabolism of the cartilage and subchondral osseous tissue. In in vitro conditions, the explants of this tissue obtained from patients suffering from osteoarthritis and rheumatoid arthritis produce similar pro- and anti-inflammatory cytokines to the explants of the synovial membrane. At this stage already, knowledge translates into imaging diagnostics. In radiological images, the shadowing of the periarticular soft tissues may not only reflect synovial membrane pathologies or joint effusion, but may also suggest inflammatory edema of the adipose tissue. On ultrasound examinations, abnormal presentation of the adipose tissue, i.e. increased echogenicity and hyperemia, may indicate its inflammation. Such images have frequently been obtained during ultrasound scanning and have been interpreted as inflammation, edema, hypertrophy or fibrosis of the adipose tissue. At present, when the knowledge concerning pathogenic mechanisms is taken into account, abnormal echogenicity and hyperemia of the adipose tissue may be considered as a proof of its inflammation. In the authors’ own practice, the inflammation of the adipose tissue usually accompanies synovitis. However, we also diagnose cases in which the inflammatory process in the joint is no longer active, but abnormal vascularity still persists in the adipose tissue. There are also cases where abnormal adipose tissue is the only sign of inflammation. Therefore, ultrasound examination confirms the existence of the additional site of inflammation, i.e. the adipose tissue which should be evaluated at the stage of initial diagnosis and during follow-up, also in terms of remission.

The pathogenesis of rheumatoid arthritis in radiological studies. Part I: Formation of inflammatory infiltrates within the synovial membrane.

Rheumatoid arthritis is a chronic inflammatory disease with a multifactorial etiology and varied course, which in the majority of patients leads to partial disability or to permanent handicap. Its characteristic trait is a persistent inflammation of the synovial membrane and the formation of an invasive synovial tissue, called the pannus, which in time leads to destruction of the cartilage, subchondral bone tissue, and the soft tissue of the affected joint(s). The pathogenesis of rheumatoid arthritis is complex and involves cells of both innate and adaptive immunity, a network of various cytokines and an immunoregulatory dysfunction. An important role in the discovery of rheumatoid arthritis pathogenesis was played by magnetic resonance imaging, which showed the disease process to extend beyond the synovium into the bone marrow. Many studies have shown a strict correlation between the vascularity of the synovium (assessed through the power Doppler ultrasound and magnetic resonance examinations), bone marrow edema and the clinical, laboratory and histopathological parameters of rheumatoid arthritis. From the current understanding of rheumatoid arthritis, bone erosions could occur from two directions: from the joint cavity and from the bone marrow. With power Doppler ultrasound, as well as in magnetic resonance imaging, it is possible to visualize the well-vascularized pannus and its destructive effects on joint structures and ligaments. In addition, the magnetic resonance study shows inflammatory and destructive changes within the bone marrow (bone marrow edema, inflammatory cysts, and erosions). Bone marrow edema occurs in 68–75% of patients with early rheumatoid arthritis and is considered to be a predictor of rapid disease progression.

Stress: Where are we now? Does immunity play an intrinsic role?

The whole world experiences progress and development, however it is the human being who pays the price in stress--an inevitable part of modern life. When encountering stress, an individual reacts at the level of both the micro- and macroenvironment. Nowadays, stress is defined as a real or interpreted threat to the physiological or psychological integrity of an individual, which results in a physiological and/or behavioral response. In the article a review of the stress conceptualization, health consequences of stress (its neuro- physiology and relation to autoimmune disease) as well as ways of management (exercises and psychotherapeutic intervention) is given.