Transforming growth factor alpha suppression of articular chondrocyte phenotype and Sox9 expression in a rat model of osteoarthritis.
ABSTRACT To define the roles of transforming growth factor alpha (TGFalpha) in cartilage degradation.
Primary rat articular chondrocytes and articular osteochondral explants were cultured with TGFalpha to assess the effects of TGFalpha on chondrocyte physiology and phenotype.
TGFalpha altered chondrocyte morphology through reorganization of the actin cytoskeleton and formation of stress fibers. Expression of anabolic genes, including aggrecan, type II collagen, and cartilage link protein, was reduced in response to TGFalpha. Proliferation of chondrocytes and formation of articular chondrocyte clusters was stimulated by TGFalpha. Expression of matrix metalloproteinase 13 and cathepsin C was increased by TGFalpha. We demonstrated the down-regulation of Sox9 messenger RNA and protein levels by TGFalpha. This was associated with reduced levels of phosphorylated and total SOX9 in cartilage explants upon TGFalpha treatment. In contrast, another growth factor identified in our microarrays, Kitl, had no effects on the chondrocyte parameters tested. To examine correlations between the increased levels of TGFalpha in experimental knee osteoarthritis (OA) with the levels of TGFalpha in humans with knee OA, a microarray analysis of mRNA from 13 normal and 12 late-stage OA cartilage samples was performed. Seven OA samples showed TGFA mRNA levels similar to those in the normal controls, but expression was markedly increased in the other 5 OA samples. These data confirm that TGFA transcript levels are increased in a subset of patients with OA.
This study adds TGFalpha to the list of dysregulated cytokines present in degrading cartilage in OA. Since TGFalpha inhibits articular chondrocyte anabolic capacity, increases catabolic factors, and contributes to the development of chondrocyte clusters, TGFalpha may be a potential target for therapeutic strategies in the treatment of OA.
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ABSTRACT: Objective: Mig-6 regulates epidermal growth factor receptor (EGFR) signalling in synovial joint tissues. Whole body knockout of the Mig-6 gene in mice has been shown to induce osteoarthritis and joint degeneration. To evaluate the role of chondrocytes in this process, Mig-6 was conditionally deleted from Col2a1 expressing cell types in the cartilage of mice.Methods: Bone and cartilage in the synovial joints of cartilage specific Mig-6 deleted (KO) mice and control littermates were compared. Histological stains and immunohistochemistry were used to evaluate joint pathology as well as expression of key extracellular matrix and regulatory proteins. Calcified tissue in synovial joints was assessed by microCT and whole skeletal staining.Results: Formation of long bones was found to be normal in KO animals. The cartilage thickness and proteoglycan staining of articular cartilage in the knee of 12 week old KO mice was increased compared to control, with higher cellularity throughout the tissue. Radio-opaque chondro-osseous nodules appear by 12 weeks of age in the knees of KO animals, and progressed to calcified bone-like tissue by 36 weeks of age. Nodules were also observed in the spine of 36 week old animals. Erosion of bone at ligament entheses was evident by 12 weeks of age in both histology and microCT.Conclusion: Mig-6 expression in chondrocytes is important for the maintenance of cartilage and joint homeostasis. Dysregulation of EGFR signalling in chondrocytes results in anabolic activity in cartilage, but erosion of ligament entheses and formation of ectopic chondro-osseous nodules severely disturb joint physiology. © 2014 American College of Rheumatology.Arthritis & Rheumatology. 06/2014;
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ABSTRACT: A deficiency of mitogen-inducible gene-6 (Mig-6) in mice leads to the development of an early-onset, osteoarthritis (OA)-like disorder in multiple synovial joints, underlying its importance in maintaining joint homeostasis. Here we determined what joint tissues Mig-6 is expressed in and what role chondrocytes play in the Mig-6-deficient OA-like disorder. A Mig-6/lacZ reporter mouse strain expressing β-galactosidase under the control of the Mig-6 gene promoter was generated to determine Mig-6 expression in joint tissues. By β-galactosidase staining, we demonstrated that Mig-6 was uniquely expressed in the cells across the entire surface of the synovial joint cavity, including chondrocytes in the superficial zone of articular cartilage and in the meniscus, as well as synovial lining cells. By crossing Mig-6-floxed mice to Col2a1-Cre transgenic mice, to generate cartilage-specific deletion of Mig-6, we demonstrated that deficiency of Mig-6 in the chondrocytes results in a joint phenotype that only partially recapitulates the OA-like disorder of the Mig-6-deficient mice: Ubiquitous deletion of Mig-6 led to the OA-like disorder in multiple joints, whereas cartilage-specific deletion affected the knees but rarely other joints. Furthermore, chondrocytes with Mig-6 deficiency showed excessive proliferative activities along with enhanced EGF receptor signaling in the articular cartilage and in the abnormally formed osteophytes. Our findings provide insight into the crucial requirement for Mig-6 in maintaining joint homeostasis and in regulating chondrocyte activities in the synovial joints. Our data also suggest that other cell types are required for fully developing the Mig-6-deficient OA-like disorder.Proceedings of the National Academy of Sciences 02/2014; 111(7):2590-5. · 9.81 Impact Factor
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ABSTRACT: Ligament and meniscus damage can cause joint disease. Arthritic joints contain increased amounts of EGFR protein, and polymorphisms in EGFR are associated with arthritis risk. The role of endogenous EGFR regulation during joint disease due to ligament and meniscus trauma is unknown. Mitogen-inducible gene 6 (MIG-6) can reduce EGFR phosphorylation and downstream signaling. We examined the effect of EGFR modulation by MIG-6 on joint disease development after ligament and meniscus injury. Knee ligament transection and meniscus removal was performed surgically on mice homozygous for a global inactivating mutation in MIG-6 (Mig-6-/-) and in WT animals. Two weeks after surgery, Mig-6-/- mice had bone erosion as well as greater fibrous tissue area and serum RANKL concentration. Four weeks after surgery, Mig-6-/- mice had less cartilage and increased proliferating cells relative to contralateral control and WT knees; increased apoptotic cells and growth outside of the articulating region occurred in Mig-6-/- mice. Tibia trabecular BMD and the number of trabeculae were lower in surgery knees relative to respective control knees for both groups. BMD, as well as trabecular thickness and number, were lower in surgery knees from Mig-6-/- mice relative to WT surgery knees. Phospho-EGFR staining in surgery knees decreased for WT mice and increased for Mig-6-/- mice. Fewer inflammatory cells were present in WT knees. Mig-6-/- mice have rapid and increased joint damage after ligament and meniscus trauma. Mig-6 modification could lessen degenerative disease development after this type of injury.Arthritis research & therapy 03/2014; 16(2):R81. · 4.12 Impact Factor