Toll-like receptors and chondrocytes: the lipopolysaccharide-induced decrease in cartilage matrix synthesis is dependent on the presence of toll-like receptor 4 and antagonized by bone morphogenetic protein 7.

OBJECTIVE To assess the presence of Toll-like receptors (TLRs) 1-9 in human articular cartilage, and to investigate the effects of lipopolysaccharide (LPS)-induced activation of TLR-4 on biosynthetic activity and matrix production by human articular chondrocytes. METHODS TLRs 1-9 were assessed in human articular cartilage by reverse transcription-polymerase chain reaction (RT-PCR); TLR-4 was also analyzed by Western blotting and immunohistochemistry. Articular chondrocytes were isolated from human donors and from wild-type or TLR-4(-/-) mice. Chondrocyte monolayer cultures were incubated with interleukin-1beta (IL-1beta) and LPS in the absence or presence of bone morphogenetic protein 7 (BMP-7) and IL-1 receptor antagonist (IL-1Ra). Neosynthesis of sulfated glycosaminoglycans (sGAG) was measured by (35)S-sulfate incorporation. Endogenous gene expression of cartilage markers as well as IL-1beta was examined using RT-PCR. The involvement of p38 kinase or p44/42 kinase (ERK-1/2) in LPS-mediated TLR-4 signaling was investigated by immunoblotting, RT-PCR, and sGAG synthesis. RESULTS TLRs 1-9 were found on the messenger RNA (mRNA) level in human articular chondrocytes. The presence of TLR-4 was also observed on the protein level. In murine and human articular chondrocytes, but not in chondrocytes derived from TLR-4(-/-) mice, stimulation with LPS resulted in a decrease in total proteoglycan synthesis. IL-1beta mRNA expression was increased by TLR-4 activation, whereas expression of aggrecan and type II collagen was significantly decreased. The presence of BMP-7 and IL-1Ra antagonized the anti-anabolic effects of LPS. Blocking of p38, but not ERK-1/2, resulted in inhibition of both LPS-mediated IL-1beta gene expression and the negative effects of LPS on matrix biosynthesis. CONCLUSION These data demonstrate the presence of TLRs in human articular cartilage. The suppressive effects of LPS on cartilage biosynthetic activity are dependent on the presence of TLR-4, are governed, at least in part, by an up-regulation of IL-1beta, and are mediated by p38 kinase. These in vitro data indicate an anti-anabolic effect of TLR-4 in articular chondrocytes that may hamper cartilage repair in various joint diseases.

[1]  A. Blom,et al.  Toll-like receptor 4 induced FcγR expression potentiates early onset of joint inflammation and cartilage destruction during immune complex arthritis: Toll-like receptor 4 largely regulates FcγR expression by interleukin 10 , 2006, Annals of the rheumatic diseases.

[2]  R. Zimmer,et al.  IL-1beta, but not BMP-7 leads to a dramatic change in the gene expression pattern of human adult articular chondrocytes--portraying the gene expression pattern in two donors. , 2006, Cytokine.

[3]  M. Ojaniemi,et al.  TLR-2 is upregulated and mobilized to the hepatocyte plasma membrane in the space of Disse and to the Kupffer cells TLR-4 dependently during acute endotoxemia in mice. , 2006, Immunology letters.

[4]  F. C. Gibson,et al.  Sensitization of Human Aortic Endothelial Cells to Lipopolysaccharide via Regulation of Toll-Like Receptor 4 by Bacterial Fimbria-Dependent Invasion , 2005, Infection and Immunity.

[5]  D. Mennerich,et al.  Characterization of Toll-like receptors in primary lung epithelial cells: strong impact of the TLR3 ligand poly(I:C) on the regulation of Toll-like receptors, adaptor proteins and inflammatory response , 2005, Journal of inflammation.

[6]  D. Salter,et al.  Expression and regulation of Toll-like receptor 2 by IL-1β and fibronectin fragments in human articular chondrocytes , 2005 .

[7]  Jordi Bolós,et al.  Structure-activity relationships of p38 mitogen-activated protein kinase inhibitors. , 2005, Mini reviews in medicinal chemistry.

[8]  R. Terkeltaub,et al.  TLR2 Signaling in Chondrocytes Drives Calcium Pyrophosphate Dihydrate and Monosodium Urate Crystal-Induced Nitric Oxide Generation1 , 2005, The Journal of Immunology.

[9]  Shizuo Akira,et al.  Pleiotropic function of Toll-like receptors. , 2004, Microbes and infection.

[10]  L. Joosten,et al.  Expression of toll-like receptors 2 and 4 in rheumatoid synovial tissue and regulation by proinflammatory cytokines interleukin-12 and interleukin-18 via interferon-gamma. , 2004, Arthritis and rheumatism.

[11]  R. Wait,et al.  Induction of interleukin-1 in articular cartilage by explantation and cutting. , 2004, Arthritis and rheumatism.

[12]  M. Pierer,et al.  Chemokine Secretion of Rheumatoid Arthritis Synovial Fibroblasts Stimulated by Toll-Like Receptor 2 Ligands1 , 2004, The Journal of Immunology.

[13]  R. Bergman,et al.  Chondrocyte response to growth factors is modulated by p38 mitogen-activated protein kinase inhibition , 2003, Arthritis research & therapy.

[14]  J. Smolen,et al.  Chondrocyte number and proteoglycan synthesis in the ageing and osteoarthritic human articular cartilage , 2003, Annals of the rheumatic diseases.

[15]  L. Joosten,et al.  Toll-Like Receptor 2 Pathway Drives Streptococcal Cell Wall-Induced Joint Inflammation: Critical Role of Myeloid Differentiation Factor 88 , 2003, The Journal of Immunology.

[16]  K. Isse,et al.  Lipopolysaccharide Activates Nuclear Factor-KappaB through Toll-Like Receptors and Related Molecules in Cultured Biliary Epithelial Cells , 2003, Laboratory Investigation.

[17]  J. Smolen,et al.  Therapeutic strategies for rheumatoid arthritis , 2003, Nature Reviews Drug Discovery.

[18]  R. Gay,et al.  Bacterial peptidoglycans but not CpG oligodeoxynucleotides activate synovial fibroblasts by toll-like receptor signaling. , 2003, Arthritis and rheumatism.

[19]  B. Beutler,et al.  Innate immune sensing and its roots: the story of endotoxin , 2003, Nature Reviews Immunology.

[20]  J. Gaston,et al.  Investigation of infectious agents associated with arthritis by reverse transcription PCR of bacterial rRNA , 2002, Arthritis research & therapy.

[21]  S. Akira,et al.  Toll-like receptors. , 2003, Annual review of immunology.

[22]  R. Gay,et al.  Detection of interleukin 1beta (IL-1beta), IL-6, and tumor necrosis factor-alpha in skin of patients with fibromyalgia. , 2003, The Journal of rheumatology.

[23]  Marina A Dobrovolskaia,et al.  Toll receptors, CD14, and macrophage activation and deactivation by LPS. , 2002, Microbes and infection.

[24]  Elisabeth L. Humphris,et al.  Role of Toll-Like Receptors in Changes in Gene Expression and NF-κB Activation in Mouse Hepatocytes Stimulated with Lipopolysaccharide , 2002, Infection and Immunity.

[25]  A. Silman,et al.  Epidemiology and genetics of rheumatoid arthritis , 2002, Arthritis research.

[26]  W. Graninger,et al.  Cartilage-derived morphogenetic protein-1 and -2 are endogenously expressed in healthy and osteoarthritic human articular chondrocytes and stimulate matrix synthesis. , 2002, Osteoarthritis and cartilage.

[27]  Shizuo Akira,et al.  Toll-like receptors as adjuvant receptors. , 2002, Biochimica et biophysica acta.

[28]  J. Pelletier,et al.  The role of cytokines in osteoarthritis pathophysiology. , 2002, Biorheology.

[29]  C. Janeway,et al.  Innate immune recognition. , 2002, Annual review of immunology.

[30]  Ruslan Medzhitov,et al.  Toll-like receptors and innate immunity , 2001, Nature Reviews Immunology.

[31]  A. Tarkowski,et al.  Staphylococcal peptidoglycans induce arthritis , 2001, Arthritis research.

[32]  J. Bertin,et al.  CARD4/Nod1 mediates NF‐κB and JNK activation by invasive Shigella flexneri , 2001 .

[33]  A. Steere,et al.  Differences in synovial fluid levels of matrix metalloproteinases suggest separate mechanisms of pathogenesis in Lyme arthritis before and after antibiotic treatment. , 2001, The Journal of infectious diseases.

[34]  W. Arend.  Cytokine imbalance in the pathogenesis of rheumatoid arthritis: the role of interleukin-1 receptor antagonist. , 2001, Seminars in arthritis and rheumatism.

[35]  S. Akira,et al.  Toll-like receptors and their signaling mechanism in innate immunity , 2001, Acta odontologica Scandinavica.

[36]  K. Huch.  Knee and ankle: human joints with different susceptibility to osteoarthritis reveal different cartilage cellularity and matrix synthesis in vitro , 2001 .

[37]  F. Luyten,et al.  Human periosteum-derived cells maintain phenotypic stability and chondrogenic potential throughout expansion regardless of donor age. , 2001, Arthritis and rheumatism.

[38]  S. Ghosh,et al.  Toll-like receptor-mediated NF-kappaB activation: a phylogenetically conserved paradigm in innate immunity. , 2001, The Journal of clinical investigation.

[39]  W. B. Berg.  Arguments for interleukin 1 as a target in chronic arthritis. , 2000 .

[40]  A. Aderem,et al.  Toll-like receptors in the induction of the innate immune response , 2000, Nature.

[41]  E. Bier,et al.  sog and dpp exert opposing maternal functions to modify toll signaling and pattern the dorsoventral axis of the Drosophila embryo. , 2000, Development.

[42]  Hiroshi Shibuya,et al.  BMP2-induced Apoptosis Is Mediated by Activation of the TAK1-p38 Kinase Pathway That Is Negatively Regulated by Smad6* , 2000, The Journal of Biological Chemistry.

[43]  P. D. Kraan,et al.  Anabolic and destructive mediators in osteoarthritis. , 2000 .

[44]  C. Dinarello,et al.  The IL-1 receptor/toll-like receptor superfamily: crucial receptors for inflammation and host defense. , 2000, Immunology today.

[45]  J. Swantek,et al.  IL-1 Receptor-Associated Kinase Modulates Host Responsiveness to Endotoxin1 , 2000, The Journal of Immunology.

[46]  F. Breedveld,et al.  Presence of bacterial DNA and bacterial peptidoglycans in joints of patients with rheumatoid arthritis and other arthritides. , 2000, Arthritis and rheumatism.

[47]  L. Joosten,et al.  Role of tumour necrosis factor α in experimental arthritis: separate activity of interleukin 1β in chronicity and cartilage destruction , 1999 .

[48]  S. Wright,et al.  Transport of Bacterial Lipopolysaccharide to the Golgi Apparatus , 1999, The Journal of experimental medicine.

[49]  M. Bayliss,et al.  Sulfation of Chondroitin Sulfate in Human Articular Cartilage , 1999, The Journal of Biological Chemistry.

[50]  S. Tashiro,et al.  ATF-2 Is a Common Nuclear Target of Smad and TAK1 Pathways in Transforming Growth Factor-β Signaling* , 1999, The Journal of Biological Chemistry.

[51]  F. Luyten,et al.  Cartilage‐Derived Morphogenetic Proteins and Osteogenic Protein‐1 Differentially Regulate Osteogenesis , 1998, Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research.

[52]  J. Ralphs,et al.  Characterization of collagens and proteoglycans at the insertion of the human Achilles tendon. , 1998, Matrix biology : journal of the International Society for Matrix Biology.

[53]  F. Luyten,et al.  Presence of cartilage-derived morphogenetic proteins in articular cartilage and enhancement of matrix replacement in vitro. , 1998, Arthritis and rheumatism.

[54]  E. Thonar,et al.  Effects of recombinant human osteogenic protein 1 on the production of proteoglycan, prostaglandin E2, and interleukin‐1 receptor antagonist by human articular chondrocytes cultured in the presence of interleukin‐1β , 1997 .

[55]  W. Feeser,et al.  The effects of IL-1 on mitogen-activated protein kinases in rabbit articular chondrocytes. , 1997, Biochemical and biophysical research communications.

[56]  R. Tapping,et al.  Lipopolysaccharide dependent cellular activation. , 1997, Journal of periodontal research.

[57]  E. Thonar,et al.  Recombinant human osteogenic protein 1 is a potent stimulator of the synthesis of cartilage proteoglycans and collagens by human articular chondrocytes. , 1996, Arthritis and rheumatism.

[58]  J. Pelletier,et al.  Chondroprotective effect of intraarticular injections of interleukin-1 receptor antagonist in experimental osteoarthritis. Suppression of collagenase-1 expression. , 1996, Arthritis and rheumatism.

[59]  L. Joosten,et al.  Role of interleukin-1, tumor necrosis factor alpha, and interleukin-6 in cartilage proteoglycan metabolism and destruction. Effect of in situ blocking in murine antigen- and zymosan-induced arthritis. , 1995, Arthritis and rheumatism.

[60]  M. McMahon,et al.  Examination of the role of MAP kinase in the response of macrophages to lipopolysaccharide. , 1995, Progress in clinical and biological research.

[61]  Jiahuai Han,et al.  Endotoxin induces rapid protein tyrosine phosphorylation in 70Z/3 cells expressing CD14. , 1993, The Journal of biological chemistry.

[62]  P. Baeuerle,et al.  Nuclear factor kappa B, a mediator of lipopolysaccharide effects. , 1993, Immunobiology.

[63]  A. Recklies,et al.  Induction of synthesis and release of interleukin-8 from human articular chondrocytes and cartilage explants. , 1992, Arthritis and rheumatism.

[64]  T. Luger,et al.  Production of natural killer cell activity-augmenting factor (interleukin-6) by human epiphyseal chondrocytes. , 1992, Arthritis and rheumatism.

[65]  M. Tiku,et al.  Articular chondrocytes secrete IL-1, express membrane IL-1, and have IL-1 inhibitory activity. , 1992, Cellular immunology.

[66]  T. Suzuki,et al.  Lipopolysaccharide receptors and signal transduction pathways in mononuclear phagocytes. , 1992, Current topics in microbiology and immunology.

[67]  P. D. Kittlick,et al.  Effect of the microbial constituents, LPS and BCG, on the glycosaminoglycans of chondrocyte cultures. , 1991, Experimental pathology.

[68]  J. B. Liesch,et al.  Production of hydrogen peroxide by rabbit articular chondrocytes. Enhancement by cytokines. , 1990, Journal of immunology.

[69]  A. J. Severijnen,et al.  Chronic arthritis induced in rats by cell wall fragments of Eubacterium species from the human intestinal flora , 1990, Infection and immunity.

[70]  S. Bulstra,et al.  Metabolic characteristics of in vitro cultured human chondrocytes in relation to the histopathologic grade of osteoarthritis. , 1989, Clinical orthopaedics and related research.

[71]  D. Heinegård,et al.  Cartilage proteoglycans in synovial fluid and serum in patients with inflammatory joint disease. Relation to systemic treatment. , 1987, Arthritis and rheumatism.

[72]  C. Sledge,et al.  Chondrocyte-mediated breakdown of cartilage. , 1987, The Journal of rheumatology.

[73]  P. A. Leonard,et al.  Hematuria in patients with rheumatoid arthritis receiving gold and D-penicillamine. , 1987, The Journal of rheumatology.

[74]  C. Laurencin,et al.  Expression of IL-1 genes in human and bovine chondrocytes: a mechanism for autocrine control of cartilage matrix degradation. , 1986, Biochemical and biophysical research communications.

[75]  M. Tiku,et al.  Class II histocompatibility antigen-mediated immunologic function of normal articular chondrocytes. , 1985, Journal of immunology.

[76]  D. Heinegård,et al.  DIFFERENCE IN CARTILAGE PROTEOGLYCAN LEVEL IN SYNOVIAL FLUID IN EARLY RHEUMATOID ARTHRITIS AND REACTIVE ARTHRITIS , 1985, The Lancet.

[77]  H. Jasin.  Bacterial lipopolysaccharides induce in vitro degradation of cartilage matrix through chondrocyte activation. , 1983, The Journal of clinical investigation.

[78]  K. Paigen,et al.  A simple, rapid, and sensitive DNA assay procedure. , 1980, Analytical biochemistry.

[79]  Christine A. Lee,et al.  COMBINED DIETARY VITAMIN-B12 DEFICIENCY AND BETA-THALASSÆMIA TRAIT , 1975, The Lancet.

[80]  R. Stockwell.  The cell density of human articular and costal cartilage. , 1967, Journal of anatomy.

[81]  J. S. Miles,et al.  BIOCHEMICAL STUDIES OF HUMAN CARTILAGE DURING THE AGING PROCESS * , 1964, Journal of the American Geriatrics Society.

[82]  D. Collins,et al.  Cell Counts of Normal and Osteo-Arthritic Articular Cartilage in Relation to the Uptake of Sulphate (35SO4) in vitro , 1962, Annals of the rheumatic diseases.