Modulation of TGF-beta signaling by proinflammatory cytokines in articular chondrocytes.

OBJECTIVE: The normal structure and function of articular cartilage are the result of a precisely balanced interaction between anabolic and catabolic processes. The transforming growth factor-beta (TGF-beta) family of growth factors generally exerts an anabolic or repair response; in contrast, proinflammatory cytokines such as interleukin 1 beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) exert a strong catabolic effect. Recent evidence has shown that IL-1beta, and TNF-alpha, and the TGF-beta signaling pathways share an antagonistic relationship. The aim of this study was to determine whether the modulation of the response of articular chondrocytes to TGF-beta by IL-1beta or TNF-alpha signaling pathways occurs through regulation of activity and availability of mothers against DPP (Drosophila) human homologue (Smad) proteins. METHODS: Human articular chondrocytes isolated from knee joints from patients with osteoarthritis (OA) or normal bovine chondrocytes were cultured in suspension in poly-(2-hydroxyethyl methacrylate)-coated dishes with either 10% fetal bovine serum media or serum-deprived media 6h before treatment with IL-1beta alone, TNF-alpha alone or IL-1beta followed by TGF-beta. Nuclear extracts were examined by electrophoretic mobility-shift assays (EMSA) for nuclear factor-kappa B (NF-kappaB) and Smad3/4 deoxyribonucleic acid (DNA) binding. Nuclear extracts were also subjected to the TranSignal Protein/DNA array (Panomics, Redwood City, CA) enabling the simultaneous semiquantitative assessment of DNA-binding activity of 54 different transcription factors. Nuclear phospho-Smad2/3 and total Smad7 protein expression in whole cell lysates were studied by Western blot. Cytoplasmic Smad7, type II collagen alpha 1 (COL2A1), aggrecan and SRY-related high mobility group-Box gene 9 (SOX-9) mRNA expression were measured by real-time polymerase chain reaction (PCR). RESULTS: The DNA-binding activity of Smad3/4 in the TranSignal Protein/DNA array was downregulated by TNF-alpha (46%) or IL-1beta treatment (42%). EMSA analysis showed a consistent reduction in Smad3/4 DNA-binding activity in human articular chondrocytes treated with IL-1beta or TNF-alpha. TGF-beta-induced Smad3/4 DNA-binding activity and Smad2/3 phosphorylation were also reduced following pretreatment with IL-1beta in human OA and bovine chondrocytes. Real-time PCR and Western blot analysis showed that IL-1beta partially reversed the TGF-beta stimulation of Smad7 mRNA and protein levels in TGF-beta-treated human OA cells. In contrast, TGF-beta-stimulated COL2A1, aggrecan, and SOX-9 mRNA levels were abrogated by IL-1beta. CONCLUSIONS: IL-1beta or TNF-alpha exerted a suppressive effect on Smad3/4 DNA-binding activity in human articular chondrocytes, as well as on TGF-beta-induced stimulation of Smad3/4 DNA-binding activity and Smad2/3 phosphorylation in human OA and bovine articular chondrocytes. IL-1beta partially reversed the increase in TGF-beta-stimulated Smad7 mRNA or protein levels suggesting that Smad7 may not be involved in the suppression of TGF-beta signaling induced by IL-1beta or TNF-alpha in articular chondrocytes. The balance between the IL-1beta or TNF-alpha and the TGF-beta signaling pathways is crucial for maintenance of articular cartilage homeostasis and its disruption likely plays a substantial role in the pathogenesis of OA

Targeting NF-kappaB: a promising molecular therapy in inflammatory arthritis.

The nuclear factor-kappa B family of transcription factors is intimately involved in the regulation of the inflammatory responses that play a fundamental role in the damage of articular tissues. Thus, many studies have examined the important contributions of components of the NF-kappaB signaling pathways to the pathogenesis of various rheumatic diseases and their pharmacologic modulation. Currently available therapeutic agents including nonsteroidal anti-inflammatory drugs, corticosteroids, nutraceuticals, and disease-modifying antirheumatic drugs, as well as novel specific small-molecule inhibitors have been employed. In addition, promising nucleic acid-based strategies have shown encouraging results. However, further research will be needed before NF-kappaB-aimed strategies become an effective therapy for inflammatory arthritis

Osteoarthritis associated with estrogen deficiency

Osteoarthritis (OA) affects all articular tissues and finally leads to joint failure. Although articular tissues have long been considered unresponsive to estrogens or their deficiency, there is now increasing evidence that estrogens influence the activity of joint tissues through complex molecular pathways that act at multiple levels. Indeed, we are only just beginning to understand the effects of estrogen deficiency on articular tissues during OA development and progression, as well as on the association between OA and osteoporosis. Estrogen replacement therapy and current selective estrogen receptor modulators have mixed effectiveness in preserving and/or restoring joint tissue in OA. Thus, a better understanding of how estrogen acts on joints and other tissues in OA will aid the development of specific and safe estrogen ligands as novel therapeutic agents targeting the OA joint as a whole organ

Hypercholesterolemia boosts joint destruction in chronic arthritis. An experimental model aggravated by foam macrophage infiltration

Objective: The aim of this study was to determine whether hypercholesterolemia increases articular damage in a rabbit model of chronic arthritis. Methods: Hypercholesterolemia was induced in 18 rabbits by administrating a high-fat diet (HFD). Fifteen rabbits were fed normal chow as controls. Chronic antigen-induced arthritis (AIA) was induced in half of the HFD and control rabbits, previously immunized, by intra-articular injections of ovalbumin. After sacrifice, lipid and systemic inflammation markers were analyzed in blood serum. Synovium was analyzed by Krenn score, multinucleated cell counting, immunohistochemistry of RAM11 and CD31, and TNF-a and macrophage chemoattractant protein-1 (MCP-1) gene expression. Active bone resorption was assessed by protein expression of receptor activator of nuclear factor kappa-B ligand (RANKL), osteoprotegerin (OPG) and quantification of cathepsin K, contact surface and the invasive area of pannus into bone. Results: Rabbits receiving the HFD showed higher total serum cholesterol, HDL, triglycerides and CRP levels than rabbits fed a normal diet. Synovitis score was increased in HFD, and particularly in AIA and AIA + HFD groups. AIA + HFD synovium was characterized by a massive infiltration of RAM11+ cells, higher presence of multinucleated foam cells and bigger vascularization than AIA. Cathepsin K+ osteoclasts and the contact surface of bone resorbing pannus were also increased in rabbits with AIA + HFD compared with AIA alone. Synovial TNF-a and MCP-1 gene expression was increased in AIA and HFD rabbits compared with healthy animals. RANKL protein expression in AIA and AIA + HFD groups was higher compared with either HFD or normal groups. Conclusions: This experimental model demonstrates that hypercholesterolemia increments joint tissue damage in chronic arthritis, with foam macrophages being key players in this process.This work was supported by research grants from Instituto de Salud Carlos III (PS09/01625, GH-B and PS09/00034, RL). IP-P is the recipient of a fellowship from Fundacion Conchita Rábago. MJM-C and RG are both recipients of a Sara Borrell contract from Instituto de Salud Carlos III. RL was funded by Instituto de Salud Carlos III through a research staff stabilization progra

Characterization of multinucleated giant cells in synovium and subchondral bone in knee osteoarthritis and rheumatoid arthritis

Background: Multinucleated giant cells have been noticed in diverse arthritic conditions since their first description in rheumatoid synovium. However, their role in the pathogenesis of osteoarthritis (OA) or rheumatoid arthritis (RA) still remains broadly unknown. We aimed to study the presence and characteristics of multinucleated giant cells (MGC) both in synovium and in subchondral bone tissues of patients with OA or RA. Methods: Knee synovial and subchondral bone samples were from age-matched patients undergoing total joint replacement for OA or RA, or non-arthritic post mortem (PM) controls. OA synovium was stratified by histological inflammation grade using index tissue sections. Synovitis was assessed by Krenn score. Histological studies employed specific antibodies against macrophage markers or cathepsin K, or TRAP enzymatic assay. Results: Inflamed OA and RA synovia displayed more multinucleated giant cells than did non-inflamed OA and PM synovia. There was a significant association between MGC numbers and synovitis severity. A TRAP negative/cathepsin K negative Langhans-like subtype was predominant in OA, whereas both Langhans-like and TRAP-positive/ cathepsin K negative foreign-body-like subtypes were most commonly detected in RA. Plasma-like and foam-like subtypes also were observed in OA and RA synovia, and the latter was found surrounding adipocytes. TRAP positive/ cathepsin K positive osteoclasts were only identified adjacent to subchondral bone surfaces. TRAP positive osteoclasts were significantly increased in subchondral bone in OA and RA compared to PM controls. Conclusions: Multinucleated giant cells are associated with synovitis severity, and subchondral osteoclast numbers are increased in OA, as well as in RA. Further research targeting multinucleated giant cells is warranted to elucidate their contributions to the symptoms and joint damage associated with arthritis

Clinical settings in knee osteoarthritis: Pathophysiology guides treatment

Osteoarthritis (OA) is the most common chronic joint disorder and its prevalence increases rapidly during midlife. Complex interactions of genetic alterations, sex hormone deficit, and aging with mechanical factors and systemic inflammation-associated metabolic syndrome lead to joint damage. Thus, the expression of a clinical phenotype in the early stages of OA relies on the main underlying pathway and predominant joint tissue involved at a given time. Moreover, OA often coexists with other morbidities in the same patient, which in turn condition the OA process. In this scenario, an appropriate identification of clinical phenotypes, especially in the early stages of the disease, may optimize the design of individualized treatments in OA. An ESCEO-EUGMS (European Union Geriatric Medicine Society) working group has recently suggested possible patient profiles in OA. Hereby, we propose the existence of 4 clinical phenotypes – biomechanical, osteoporotic, metabolic and inflammatory – whose characterization would help to properly stratify patients with OA in clinical trials or studies. Further research in this field is warranted