To compare the response of chondrocytes and cartilage matrix to injurious mechanical compression and treatment with interleukin-1beta (IL-1beta) and tumor necrosis factor alpha (TNFalpha), by ...characterizing proteins lost to the medium from cartilage explant culture.
Cartilage explants from young bovine stifle joints were treated with 10 ng/ml of IL-1beta or 100 ng/ml of TNFalpha or were subjected to uniaxial, radially-unconfined injurious compression (50% strain; 100%/second strain rate) and were then cultured for 5 days. Pooled media were subjected to gel-based separation (sodium dodecyl sulfate-polyacrylamide gel electrophoresis) and analysis by liquid chromatography tandem mass spectrometry, and the data were analyzed by Spectrum Mill proteomics software, focusing on protein identification, expression levels, and matrix protein proteolysis.
More than 250 proteins were detected, including extracellular matrix (ECM) structural proteins, pericellular matrix proteins important in cell-cell interactions, and novel cartilage proteins CD109, platelet-derived growth factor receptor-like, angiopoietin-like 7, and adipocyte enhancer binding protein 1. IL-1beta and TNFalpha caused increased release of chitinase 3-like protein 1 (CHI3L1), CHI3L2, complement factor B, matrix metalloproteinase 3, ECM-1, haptoglobin, serum amyloid A3, and clusterin. Injurious compression caused the release of intracellular proteins, including Grp58, Grp78, alpha4-actinin, pyruvate kinase, and vimentin. Injurious compression also caused increased release and evidence of proteolysis of type VI collagen subunits, cartilage oligomeric matrix protein, and fibronectin.
Overload compression injury caused a loss of cartilage integrity, including matrix damage and cell membrane disruption, which likely occurred through strain-induced mechanical disruption of cells and matrix. IL-1beta and TNFalpha caused the release of proteins associated with an innate immune and stress response by the chondrocytes, which may play a role in host defense against pathogens or may protect cells against stress-induced damage.
Collagen and other polyelectrolyte materials in a homogeneous, aqueous electrolyte are studied and modeled as a seat of electromechanical transduction. The frequency response of collagen membranes in ...a range of chemical environments shows how transduction with a deformable membrane coupled hydrodynamically to an external mechanical system can lead to a mechanical response resulting from an electrical stimulus and an electrical response to a mechanical excitation. Dynamic measurements, which can more easily distinguish between membrane and electrode phenomena, result in electromechanical coupling coefficients that show reciprocity and agree with values determined by stationary techniques. A model representing the membrane at the interfibrillar level by a system of cylindrical pores is developed relating externally measured potentials, membrane deformations, currents, and mass fluxes to equivalent pore radius, fibril diameter, and polyelectrolyte charge. Measured values of the membrane coupling coefficients are then used to infer average microstructural parameters of the membrane that compare favorably to available data based on electron microscopy.
Bone marrow stromal cells (BMSCs) are an established cell choice for cartilage repair because they are easily harvested, expanded, and differentiated into a cartilage phenotype characterized by ...aggrecan and type II collagen production. Transforming growth factor β (TGF-β), insulin-like growth factor 1 (IGF-1), and Dexamethasone (Dex) all influence the process of chondrogenesis. Although difficult challenges remain for optimizing the use of BMSCs for cartilage tissue engineering, in vitro culture systems present an excellent opportunity for studying chondrogenesis and understanding how progenitor cells respond to their biological, chemical, and mechanical microenvironment.
Objective Traumatic joint injury can damage cartilage and release inflammatory cytokines from adjacent joint tissue. The present study was undertaken to study the combined effects of compression ...injury, tumor necrosis factor (TNF), and interleukin-6 (IL-6) and its soluble receptor (sIL-6R) on immature bovine and adult human knee and ankle cartilage, using an in vitro model, and to test the hypothesis that endogenous IL-6 plays a role in proteoglycan loss caused by a combination of injury and TNF. Methods Injured or uninjured cartilage disks were incubated with or without TNF and/or IL-6/sIL-6R. Additional samples were preincubated with an IL-6-blocking antibody Fab fragment and subjected to injury and TNF treatment. Treatment effects were assessed by histologic analysis, measurement of glycosaminoglycan (GAG) loss, Western blot to determine proteoglycan degradation, zymography, radiolabeling to determine chondrocyte biosynthesis, and Western blot and enzyme-linked immunosorbent assay to determine chondrocyte production of IL-6. Results In bovine cartilage samples, injury combined with TNF and IL-6/sIL-6R exposure caused the most severe GAG loss. Findings in human knee and ankle cartilage were strikingly similar to those in bovine samples, although in human ankle tissue, the GAG loss was less severe than that observed in human knee tissue. Without exogenous IL-6/sIL-6R, injury plus TNF exposure up-regulated chondrocyte production of IL-6, but incubation with the IL-6-blocking Fab significantly reduced proteoglycan degradation. Conclusion Our findings indicate that mechanical injury potentiates the catabolic effects of TNF and IL-6/sIL-6R in causing proteoglycan degradation in human and bovine cartilage. The temporal and spatial evolution of degradation suggests the importance of transport of biomolecules, which may be altered by overload injury. The catabolic effects of injury plus TNF appeared partly due to endogenous IL-6, since GAG loss was partially abrogated by an IL-6-blocking Fab.
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