المؤلفون: Karim Boumediene, Jean-Marc Elissalde, Elise Duval, Philippe Galéra, Magali Demoor, Sylvain Leclercq

المصدر: Arthritis & Rheumatism. 60:3038-3048

مصطلحات موضوعية: Adult, medicine.medical_specialty, Immunology, Type II collagen, Collagen Type I, Chondrocyte, Chondrocytes, Rheumatology, Internal medicine, medicine, Humans, Immunology and Allergy, Pharmacology (medical), Aggrecans, Fibroblast, Collagen Type II, Cells, Cultured, Aggrecan, Aged, Aged, 80 and over, Chemistry, Cartilage homeostasis, Cell Differentiation, SOX9 Transcription Factor, Middle Aged, Hypoxia-Inducible Factor 1, alpha Subunit, Chondrogenesis, Cell Hypoxia, Cell biology, Collagen Type I, alpha 1 Chain, Collagen Type III, medicine.anatomical_structure, Endocrinology, Hypoxia-inducible factors, Ectopic expression, Collagen

الوصف: Objective Autologous chondrocyte implantation requires expansion of cells ex vivo, leading to dedifferentiation of chondrocytes (loss of aggrecan and type II collagen to the profit of type I and type III collagens). Several approaches have been described for redifferentiation of these cells. Among them, low oxygen tension has been exploited to restore the differentiated chondrocyte phenotype, but molecular mechanisms of this process remain unclear. However, under conditions of hypoxia, one of the major factors involved is hypoxia-inducible factor 1alpha (HIF-1alpha). The purpose of this study was to investigate the role of HIF-1alpha during human chondrocyte redifferentiation. Methods We used complementary approaches to achieving HIF-1alpha loss (inhibition by cadmium ions and dominant-negative expression) or gain (ectopic expression and cobalt ion treatment) of function. Expression of chondrocyte, as well as fibroblast-like, phenotype markers was determined using real-time reverse transcription-polymerase chain reaction and Western blot analyses. Binding activities of HIF-1alpha and SOX9, a pivotal transcription factor of chondrogenesis, were evaluated by electrophoretic mobility shift assays and by chromatin immunoprecipitation assay. Results We found that hypoxia and HIF-1alpha not only induced the expression of SOX9, COL2A1, and aggrecan, but they simultaneously inhibited the expression of COL1A1, COL1A2, and COL3A1. In addition, we identified the binding of HIF-1alpha to the aggrecan promoter, the first such reported demonstration of this binding. Conclusion This study is the first to show a bimodal role of HIF-1alpha in cartilage homeostasis, since HIF-1alpha was shown to favor specific markers and to impair dedifferentiation. This suggests that manipulation of HIF-1alpha could represent a promising approach to the treatment of osteoarthritis.

الوصول الحر: https://explore.openaire.eu/search/publication?articleId=doi_dedup___::beb2d979a87cfdb72433050e53a9fd45Test
https://doi.org/10.1002/art.24851Test

المؤلفون: Dominik R. Haudenschild, Jianfen Chen, Martin Lotz, Bao Nguyen, Darryl D. D'Lima

المصدر: Arthritis & Rheumatism. 58:2735-2742

مصطلحات موضوعية: Adult, Male, Time Factors, RHOA, Adolescent, Phalloidin, Interleukin-1beta, Immunology, Cell Culture Techniques, Gene Expression, Nitric Oxide Synthase Type II, Biology, Mechanotransduction, Cellular, Filamentous actin, Article, Chondrocyte, chemistry.chemical_compound, Chondrocytes, Rheumatology, 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine, Physical Stimulation, Osteoarthritis, medicine, Humans, Immunology and Allergy, Pharmacology (medical), RNA, Messenger, Cytoskeleton, Cell Shape, Rho-associated protein kinase, Cells, Cultured, rho-Associated Kinases, Chemokine CCL20, Microscopy, Confocal, Reverse Transcriptase Polymerase Chain Reaction, Middle Aged, Actin cytoskeleton, Actins, Cell biology, medicine.anatomical_structure, chemistry, biology.protein, Female, Stress, Mechanical, MDia1

الوصف: Objective Mechanical stimulation of cartilage affects tissue homeostasis and chondrocyte function. The chondrocyte phenotype is dependent on cell shape, which is largely determined by the actin cytoskeleton. Reorganization of the actin cytoskeleton results from Rho GTPase activation. The purpose of this study was to examine the roles of both actin and Rho in mechanotransduction in chondrocytes. Methods We embedded human articular chondrocytes in 2 × 6–mm agarose discs at 5 × 106 cells/ml and subjected the discs to unconfined dynamic compression at 0.5 Hz. By comparing samples with and without dynamic compression, we identified Rho activation according to the GTP-bound active RhoA measured in cell lysates. We identified rearrangements in filamentous actin structures using fluorescence-labeled phalloidin and confocal microscopy of fixed samples. We identified altered gene expression using TaqMan quantitative reverse transcription–polymerase chain reaction analysis. We tested for a requirement for Rho signaling by performing the dynamic compression in the presence of Rho kinase inhibitors. Results RhoA activation occurred within 5–10 minutes of dynamic compression. Rho kinase–dependent actin reorganization occurred within 20 minutes after application of dynamic compression and was apparent as “punctate” F-actin structures that were visible under confocal microscopy. We identified early-phase mechanoresponsive genes (CCL20 and inducible nitric oxide synthase) that were highly up-regulated within 1 hour of dynamic compression in a Rho kinase–dependent and actin-dependent manner. Conclusion Together, these results are the first demonstration that the Rho–Rho kinase pathway and actin cytoskeletal reorganization are required for changes in the expression of genes involved in human chondrocyte mechanotransduction.

الوصول الحر: https://explore.openaire.eu/search/publication?articleId=doi_dedup___::8bba39623c81f76c1491ba8084a1b5a0Test
https://doi.org/10.1002/art.23797Test

المؤلفون: Josef S Smolen, Ilse-Gerlinde Sunk, L. Amoyo, Lin Xu, Yefu Li, Jochen G. Hofstaetter, Afschin Soleiman, Klaus Bobacz

المصدر: Arthritis & Rheumatism. 56:3685-3692

مصطلحات موضوعية: Adult, Cartilage, Articular, Pathology, medicine.medical_specialty, Immunology, Type II collagen, Osteoarthritis, Matrix metalloproteinase, Severity of Illness Index, Chondrocyte, Chondrocytes, Rheumatology, Matrix Metalloproteinase 13, medicine, Humans, Immunology and Allergy, Pharmacology (medical), RNA, Messenger, Collagen Type II, Discoidin Domain Receptors, Cells, Cultured, Aggrecan, Aged, Aged, 80 and over, Chemistry, Cartilage, Receptor Protein-Tyrosine Kinases, Middle Aged, Osteoarthritis, Knee, medicine.disease, Cell biology, body regions, Collagen, type I, alpha 1, medicine.anatomical_structure, Receptors, Mitogen, Discoidin domain

الوصف: Objective To investigate the relationship between increased discoidin domain receptor 2 (DDR-2) expression and cartilage damage in osteoarthritis (OA). Methods Full-thickness cartilage tissue samples from 16 human knee joints were obtained and the grade of cartilage damage was evaluated according to the Mankin scale. Expression of DDR-2, matrix metalloproteinase 13 (MMP-13), and MMP-derived type II collagen fragments was visualized immunohistochemically. Moreover, upon stimulation with either type II collagen or gelatin, levels of DDR-2 and MMP-13 messenger RNA (mRNA) in primary human articular chondrocytes were assessed by real-time polymerase chain reaction. Results Immunohistochemical analysis showed an increase in DDR-2 expression in human articular cartilage, which was correlated with the degree of tissue damage. In parallel, the extent of MMP-13 and type II collagen breakdown products was elevated as a function of increased DDR-2 expression and cartilage damage. Furthermore, in vitro experiments revealed an up-regulation of both DDR-2 and MMP-13 mRNA in human articular chondrocytes after stimulation with type II collagen. Conclusion Our data indicate that 3 factors, DDR-2 expression, MMP-13 expression, and the degree of cartilage damage, are linked, such that DDR-2 promotes tissue catabolism, and tissue degradation promotes DDR-2 up-regulation and activation. Thus, the perpetuation of DDR-2 expression and activation can be seen as a vicious circle that ultimately leads to cartilage destruction in OA.

الوصول الحر: https://explore.openaire.eu/search/publication?articleId=doi_dedup___::312fa7ec664bd8ecbe1ea202d1db66b9Test
https://doi.org/10.1002/art.22970Test

المؤلفون: Jochen G. Hofstaetter, Josef S Smolen, C. D. Toma, Shizuo Akira, Thomas Weichhart, Marcus D. Säemann, Ilse-Gerlinde Sunk, L. Amoyo, K. Bobacz

المصدر: Arthritis & Rheumatism. 56:1880-1893

مصطلحات موضوعية: Adult, Lipopolysaccharides, Adolescent, Bone Morphogenetic Protein 7, Interleukin-1beta, Immunology, Type II collagen, Cartilage Oligomeric Matrix Protein, Biology, p38 Mitogen-Activated Protein Kinases, Chondrocyte, Mice, Chondrocytes, Rheumatology, Transforming Growth Factor beta, medicine, Animals, Humans, Matrilin Proteins, Immunology and Allergy, Pharmacology (medical), RNA, Messenger, Cells, Cultured, Aggrecan, Aged, Glycoproteins, Aged, 80 and over, Mice, Knockout, Cartilage oligomeric matrix protein, Extracellular Matrix Proteins, Toll-like receptor, Cartilage, Toll-Like Receptors, Middle Aged, Chondrogenesis, Cell biology, Mice, Inbred C57BL, Toll-Like Receptor 4, Bone morphogenetic protein 7, Interleukin 1 Receptor Antagonist Protein, medicine.anatomical_structure, Gene Expression Regulation, Bone Morphogenetic Proteins, biology.protein, Female

الوصف: 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-1β (IL-1β) 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 35S-sulfate incorporation. Endogenous gene expression of cartilage markers as well as IL-1β 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-1β 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-1β 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-1β, 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.

الوصول الحر: https://explore.openaire.eu/search/publication?articleId=doi_dedup___::5282316c147b3295338ad4e5779b42c4Test
https://doi.org/10.1002/art.22637Test

المؤلفون: M. B. Aydelotte, E. J-M. A. Thonar, K. E. Kuettner, Johannes Flechtenmacher, L. Michal, H. J. Häuselmann, M. Shinmei

المصدر: Arthritis & Rheumatism. 39:478-488

مصطلحات موضوعية: Adult, Cartilage, Articular, Adolescent, medicine.drug_class, Sialoglycoproteins, Immunology, Matrix metalloproteinase, Chondrocyte, Rheumatology, medicine, Humans, Immunology and Allergy, Pharmacology (medical), Catabolism, Chemistry, Cartilage, Receptors, Interleukin-1, Interleukin, Biological activity, Anatomy, Middle Aged, Receptor antagonist, Recombinant Proteins, Cell biology, Interleukin 1 Receptor Antagonist Protein, medicine.anatomical_structure, Cell culture, Proteoglycans, Interleukin-1

الوصف: Objective. To compare the responses of chondrocytes from superficial and deep layers of normal human articular cartilage to interleukin-1 (IL-1) and IL-1 receptor antagonist protein (IRAP), and to evaluate the binding sites for IL-1 on these cells. Methods. Cartilage and chondrocytes from superficial and deeper layers of human femoral condyles were cultured with and without IL-1 in the presence and absence of IRAP. The effect of these agents on 35S-proteoglycan synthesis and catabolism and production of stromelysin and tissue inhibitor of metalloproteinases 1 (TIMP-1) were measured by biochemical and immunologic assays. Receptor binding was evaluated using 125I-labeled IL-1. Results. IL-1 induced more severe inhibition of proteoglycan synthesis and a lower ratio of secreted TIMP-1:stromelysin in chondrocytes from superficial cartilage than those from deeper cartilage. IRAP blocked responses to IL-1 more effectively in chondrocytes from deep cartilage than those from superficial cartilage. Chondrocytes from the articular surface showed approximately twice the number of high-affinity binding sites for IL-1 as did cells from deep cartilage. Conclusion. Chondrocytes from the surface of articular cartilage show a greater vulnerability to the harmful effects of IL-1 and are less responsive to the potential therapeutic effects of IRAP than cells in the deeper layers of the tissue.

الوصول الحر: https://explore.openaire.eu/search/publication?articleId=doi_dedup___::615f8af66fb4322d3d6db9bbd2ff8677Test
https://doi.org/10.1002/art.1780390316Test