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Cartilage Chondrocytes (cartilage + chondrocyte)
Kinds of Cartilage Chondrocytes Selected AbstractsInvolvement of the cytoskeletal elements in articular cartilage homeostasis and pathologyINTERNATIONAL JOURNAL OF EXPERIMENTAL PATHOLOGY, Issue 1 2009Emma J. Blain Summary The cytoskeleton of all cells is a three-dimensional network comprising actin microfilaments, tubulin microtubules and intermediate filaments. Studies in many cell types have indicated roles for these cytoskeletal proteins in many diverse cellular processes including alteration of cell shape, movement of organelles, migration, endocytosis, secretion, cell division and extracellular matrix assembly. The cytoskeletal networks are highly organized in structure enabling them to fulfil their biological functions. This review will primarily focus on the organization and function of the three major cytoskeletal networks in articular cartilage chondrocytes. Articular cartilage is a major load-bearing tissue of the synovial joint; it is well known that the cytoskeleton acts as a physical interface between the chondrocytes and the extracellular matrix in ,sensing' mechanical stimuli. The effect of mechanical load on cytoskeletal element expression and organization will also be reviewed. Abnormal mechanical load is widely believed to be a risk factor for the development of osteoarthritis. Several studies have intimated that the major cytoskeletal networks are disorganized or often absent in osteoarthritic cartilage chondrocytes. The implications and possible reasoning for this are more widely discussed and placed into context with their potential relevance to disease and therapeutic strategies. [source] ERK-1/2 and p38 in the regulation of hypertrophic changes of normal articular cartilage chondrocytes induced by osteoarthritic subchondral osteoblastsARTHRITIS & RHEUMATISM, Issue 5 2010Indira Prasadam Objective Previous studies have shown the influence of subchondral bone osteoblasts (SBOs) on phenotypical changes of articular cartilage chondrocytes (ACCs) during the development of osteoarthritis (OA). The molecular mechanisms involved during this process remain elusive, in particular, the signal transduction pathways. The aim of this study was to investigate the in vitro effects of OA SBOs on the phenotypical changes in normal ACCs and to unveil the potential involvement of MAPK signaling pathways during this process. Methods Normal and arthritic cartilage and bone samples were collected for isolation of ACCs and SBOs. Direct and indirect coculture models were applied to study chondrocyte hypertrophy under the influence of OA SBOs. MAPKs in the regulation of the cell,cell interactions were monitored by phosphorylated antibodies and relevant inhibitors. Results OA SBOs led to increased hypertrophic gene expression and matrix calcification in ACCs by means of both direct and indirect cell,cell interactions. In this study, we demonstrated for the first time that OA SBOs suppressed p38 phosphorylation and induced ERK-1/2 signal phosphorylation in cocultured ACCs. The ERK-1/2 pathway inhibitor PD98059 significantly attenuated the hypertrophic changes induced by conditioned medium from OA SBOs, and the p38 inhibitor SB203580 resulted in the up-regulation of hypertrophic genes in ACCs. Conclusion The findings of this study suggest that the pathologic interaction of OA SBOs and ACCs is mediated via the activation of ERK-1/2 phosphorylation and deactivation of p38 phosphorylation, resulting in hypertrophic differentiation of ACCs. [source] Proteomic analysis by two-dimensional electrophoresis to identify the normal human chondrocyte proteome stimulated by tumor necrosis factor , and interleukin-1,ARTHRITIS & RHEUMATISM, Issue 3 2010Berta Cillero-Pastor Objective To determine the intracellular proteome of normal human chondrocytes stimulated with interleukin-1, (IL-1,) and tumor necrosis factor , (TNF,) and to ascertain differences in the protein expression patterns of these 2 cytokines. Methods Normal human knee cartilage chondrocytes were incubated for 48 hours without stimulation or stimulated with IL-1, (5 ng/ml) or with TNF, (10 ng/ml). For each culture condition, protein extracts from 4 normal subjects were pooled and resolved using 2-dimensional electrophoresis. Protein spots were visualized with Sypro stain, and qualitative and quantitative analyses were performed using PDQuest software. Protein spots were then identified by mass spectrometry, using matrix-assisted laser desorption ionization,time-of-flight/time-of-flight technology. Results We identified 37 spots by mass spectrometry (MS) or by MS/MS, corresponding to 35 different proteins. In IL-1,,stimulated chondrocytes, IL-1, was found to modulate 22 proteins, as compared with unstimulated chondrocytes. All of these proteins except connective tissue growth factor (CCND2) were up-regulated. Proteins involved in cellular metabolism and energy (23%) that were up-regulated or induced by IL-1, included nicotinamide phosphoribosyltransferase, long-chain fatty acid,coenzyme A ligase 4, ,-aminolevulinic acid dehydratase, triosephosphate isomerase, and an isoform of glyceraldehyde-3-phosphate dehydrogenase. In TNF,-stimulated chondrocytes, TNF, was found to modulate 20 proteins, as compared with unstimulated chondrocytes. All of these except chitinase 3,like 1 (cartilage glycoprotein 39), proteasome activator complex subunit 2, and G3PDH, were up-regulated. Eighteen proteins were differently modulated by IL-1, and TNF,. Of these, 45% were related to metabolism. Conclusion IL-1, and TNF, induce different profiles of intracellular protein expression in healthy human chondrocytes. Most of the proteins that are differently regulated are proteins that are implicated in the generation of cellular energy and in glycolysis. [source] Trefoil factor 3 is induced during degenerative and inflammatory joint disease, activates matrix metalloproteinases, and enhances apoptosis of articular cartilage chondrocytesARTHRITIS & RHEUMATISM, Issue 3 2010Sophie Rösler Objective Trefoil factor 3 (TFF3, also known as intestinal trefoil factor) is a member of a family of protease-resistant peptides containing a highly conserved motif with 6 cysteine residues. Recent studies have shown that TFF3 is expressed in injured cornea, where it plays a role in corneal wound healing, but not in healthy cornea. Since cartilage and cornea have similar matrix properties, we undertook the present study to investigate whether TFF3 could induce anabolic functions in diseased articular cartilage. Methods We used reverse transcriptase,polymerase chain reaction, Western blot analysis, and immunohistochemistry to measure the expression of TFF3 in healthy articular cartilage, osteoarthritis (OA),affected articular cartilage, and septic arthritis,affected articular cartilage and to assess the effects of cytokines, bacterial products, and bacterial supernatants on TFF3 production. The effects of TFF3 on matrix metalloproteinase (MMP) production were measured by enzyme-linked immunosorbent assay, and effects on chondrocyte apoptosis were studied by caspase assay and annexin V assay. Results Trefoil factors were not expressed in healthy human articular cartilage, but expression of TFF3 was highly up-regulated in the cartilage of patients with OA. These findings were confirmed in animal models of OA and septic arthritis, as well as in tumor necrosis factor ,, and interleukin-1,,treated primary human articular chondrocytes, revealing induction of Tff3/TFF3 under inflammatory conditions. Application of the recombinant TFF3 protein to cultured chondrocytes resulted in increased production of cartilage-degrading MMPs and increased chondrocyte apoptosis. Conclusion In this study using articular cartilage as a model, we demonstrated that TFF3 supports catabolic functions in diseased articular cartilage. These findings widen our knowledge of the functional spectrum of TFF peptides and demonstrate that TFF3 is a multifunctional trefoil factor with the ability to link inflammation with tissue remodeling processes in articular cartilage. Moreover, our data suggest that TFF3 is a factor in the pathogenesis of OA and septic arthritis. [source] HLA,B27,restricted antigen presentation by human chondrocytes to CD8+ T cells: Potential contribution to local immunopathologic processes in ankylosing spondylitisARTHRITIS & RHEUMATISM, Issue 6 2009Maren Kuhne Objective Analysis of the histopathologic features of hip arthritis in patients with ankylosing spondylitis (AS) has revealed accumulation of infiltrating mononuclear cells in the bone end plate and presence of hyaline articular cartilage that is not found in areas of total cartilage destruction. This study was undertaken to assess whether chondrocytes attract lymphocytes and whether cartilage chondrocytes from patients with AS have the potential to directly stimulate T cells in an HLA-restricted manner. Methods Human HLA,B27+ T cell lines, specific for the Epstein-Barr virus,derived peptide EBNA258,266, and autologous chondrocytes, serving as nonprofessional antigen-presenting cells (APCs), were available for use in a model system to study chondrocyte functions in femoral head joint cartilage of patients with AS. Peptide functionality of cytotoxic T cells was assessed by flow cytometry, and cellular interactions were detected by fluorescence confocal microscopy. Results When maintained in an alginate matrix, chondrocytes isolated from the femoral heads of patients with AS constitutively expressed type II collagen and CD80. When pulsed with the EBNA258,266 peptide, autologous chondrocytes functioned as APCs and, specifically, induced interferon-, production in CD8+ T cells. In mixed chondrocyte,T cell cultures, cell,cell contacts were dependent on the presence of the EBNA258,266 peptide. T cells adjacent to chondrocytes produced perforin and granzyme B; both molecules were found in focal aggregates, a prerequisite for antigen-specific lysis of target cells. Conclusion Antigen presentation through human chondrocytes allows the stimulation of peptide-specific CD8+ T cells. These results indicate that human chondrocytes can act as nonprofessional APCs, and suggest that there is an interferon-,,triggered autocrine loop of immune cell,mediated chondrocyte activation in the already inflamed environment. Thus, local HLA-dependent activation of peptide-specific cytotoxic CD8+ T cells by chondrocytes might contribute to inflammatory processes in the spondylarthritides. [source] The accumulation of intracellular ITEGE and DIPEN neoepitopes in bovine articular chondrocytes is mediated by CD44 internalization of hyaluronanARTHRITIS & RHEUMATISM, Issue 2 2006Jennifer J. Embry Flory Objective A dramatic loss of aggrecan proteoglycan from cartilage is associated with osteoarthritis. The fate of residual G1 domains of aggrecan is unknown, but inefficient turnover of these domains may impede subsequent repair and retention of newly synthesized aggrecan. Thus, the objective of this study was to determine whether ITEGE- and DIPEN-containing G1 domains, generated in situ, are internalized by articular chondrocytes, and whether these events are dependent on hyaluronan (HA) and its receptor, CD44. Methods ITEGE and DIPEN neoepitopes were detected by immunofluorescence staining of bovine articular cartilage chondrocytes treated with or without interleukin-1, (IL-1,). Additionally, purified ITEGE- or DIPEN-containing G1 domains were aggregated with HA and then added to articular chondrocytes, articular chondrocytes transfected with CD44,67, or COS-7 cells transfected with or without full-length CD44. Internalized epitopes were distinguished by their resistance to extensive trypsinization of the cell surface. Results Both ITEGE and DIPEN were visualized within the extracellular cell-associated matrix of chondrocytes as well as within intracellular vesicles. Following trypsinization, the intracellular accumulation of both epitopes was clearly visible. IL-1 treatment increased extracellular as well as intracellular ITEGE epitope accumulation. Once internalized, the ITEGE neoepitope became localized within the nucleus and displayed little colocalization with HA, DIPEN, or other G1 domain epitopes. The internalization of both ITEGE and DIPEN G1 domains was dependent on the presence of HA and CD44. Conclusion One important mechanism for the elimination of residual G1 domains following extracellular degradation of aggrecan is CD44-mediated co-internalization with HA. [source] In vivo selective inhibition of mitogen-activated protein kinase kinase 1/2 in rabbit experimental osteoarthritis is associated with a reduction in the development of structural changesARTHRITIS & RHEUMATISM, Issue 6 2003Jean-Pierre Pelletier Objective The primary aim of this study was to investigate, using an experimental rabbit model of osteoarthritis (OA), the effect of a selective mitogen-activated protein kinase kinase 1/2 (MEK-1/2) inhibitor, PD 198306, on the development of structural changes. Additional aims were to assess the effects of the inhibitor on levels of phosphorylated extracellular signal,regulated kinase 1/2 (phospho,ERK-1/2) and matrix metalloproteinase 1 (MMP-1; collagenase 1) in OA chondrocytes. Methods After surgical sectioning of the anterior cruciate ligament of the right knee joint, rabbits with OA were separated into 3 experimental groups: oral treatment with placebo or with PD 198306 at a therapeutic concentration of 10 mg/kg/day or 30 mg/kg/day. Each treatment started immediately after surgery. The animals were killed 8 weeks after surgery. Macroscopic and histologic studies were performed on the cartilage and synovial membrane. The levels of phospho,ERK-1/2 and MMP-1 in OA cartilage chondrocytes were evaluated by immunohistochemistry. Normal, untreated rabbits were used as controls. Results OA rabbits treated with the highest dosage of MEK-1/2 inhibitor showed decreases in the surface area (size) of cartilage macroscopic lesions (P < 0.002) and in osteophyte width on the lateral condyles (P = 0.05). Histologically, the severity of synovial inflammation (villous hyperplasia) was also reduced (P < 0.02). In cartilage from placebo-treated OA rabbits, a significantly higher percentage of chondrocytes in the superficial layer stained positive for phospho,ERK-1/2 and MMP-1 compared with normal controls. Rabbits treated with the highest dosage of PD 198306 demonstrated a significant and dose-dependent reduction in the level of phospho,ERK-1/2 and a lower level of MMP-1. Conclusion This study demonstrates that, in vivo, PD 198306, a selective inhibitor of MEK-1/2, can partially decrease the development of some of the structural changes in experimental OA. This effect was associated with a reduction in the level of phospho,ERK-1/2 in OA chondrocytes, which probably explains the action of the drug. [source] Inhibition of cartilage degradation: A combined tissue engineering and gene therapy approachARTHRITIS & RHEUMATISM, Issue 3 2003Wael Kafienah Objective To determine if tissue-engineered cartilage can be protected from cytokine-induced degradation using a gene therapy approach. Methods Chemical and pantropic retroviral gene transfer methodologies were compared for their ability to introduce a luciferase reporter gene into adult bovine cartilage chondrocytes grown in monolayer. Pantropic retrovirus was then used to transduce these cells with human tissue inhibitor of metalloproteinases 1 (TIMP-1), and the stability of expression in monolayer or pellet culture was monitored for 6 weeks. Untransduced and TIMP-1,transduced cells were also used to tissue engineer 3-dimensional cartilage constructs that were then challenged with interleukin-1 (IL-1) for 4 weeks. Conditioned media and residual cartilage were collected for analysis of matrix components, including type II collagen and proteoglycans, and for TIMP-1 production and matrix metalloproteinase (MMP) activity. Results Chemical transfection of adult bovine chondrocytes gave rise to short-lived reporter expression that was virtually undetectable after 4 weeks of culture. In contrast, pantropic retroviral transduction gave rise to stable expression that persisted at a high level for at least 6 weeks. Pantropic transduction of the cells with TIMP-1 gave rise to similar long-term expression, both in monolayer and pellet cultures. TIMP-1,transduced tissue-engineered cartilage also retained TIMP-1 expression for an additional 4 weeks of culture in the presence of IL-1. Compared with control samples, TIMP-1,transgenic cartilage resisted the catabolic effects of IL-1, with MMP activity reduced to basal levels and a decreased loss of type II collagen. Conclusion Pantropic retroviral transduction permits long-term expression of potentially therapeutic transgenes in adult tissue-engineered cartilage. While TIMP-1 transduction could be used to prevent collagen breakdown, alternative transgenes may be necessary to protect cartilage proteoglycans. [source] | ||||||||||