Modulation of Extracellular Matrix Gene Expression in Bovine High-Density Chondrocyte Cultures by Ascorbic Acid and Enzymatic Resuspension

Hering, Thomas M.; Kollar, John; Huynh, Tru D.; Varelas, Judith B.; Sandell, Linda J.

doi:10.1006/abbi.1994.1415

Cited by 69 publications

(58 citation statements)

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“…Absence of an accelerated trans-differentiation into a fibroblastic phenotype was evidenced by the lack of increase in type I procollagen steady-state mRNA expression even as expression of type I1 procollagen decreased. In contrast, monolayer cultures of chondrocytes spontaneously increase the expression and synthesis of type I procollagen with time in culture and passage [25,30,36,49].…”

Section: Discussionmentioning

confidence: 95%

Corticosteroids alter the differentiated phenotype of articular chondrocytes

Fubini

Todhunter

Burton‐Wurster

et al. 2001

Journal Orthopaedic Research

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Experimental evidence suggests that recommended dosages of some corticosteroids used clinically as antiinflammatory agents for treating arthropathies damage articular cartilage, but low dosages may be chondroprotective. The purpose of this study was to evaluate how different concentrations of methylprednisolone affect chondrocyte function and viability. Articular cartilage and chondrocytes were obtained from young adult horses, 1 5 3 . 5 years of age. Corticosteroid-induced changes in collagen expression were studied at the transcriptional level by Northern blot analyses and at the translational level by measuring [3H]-proline incorporation into [3H]-hydroxyproline. Fibronectin mRNA splicing patterns were evaluated with ribonuclease protection assays. Cytotoxicity was studied using erythrosin B dye exclusion. Steady-state levels of type I1 procollagen mRNA decreased without concurrent changes in type I procollagen expression as the medium methylprednisolone concentrations were increased from 1 x lo1 to 1 x 10' pg/ml, dropping below 10% of control values by 1 Y lo5 pg/ml. Cytotoxicity occurred as methylprednisolone levels were increased further from 1 x loR to 1 A lo9 pg/ml. Changes in total collagen (protein) synthesis were less pronounced, but also demonstrated significant suppression between 1 x lo4 and 1 x lo8 pg/ml. Corticosteroid-induced changes in fibronectin isoform levels were evaluated in articular cartilage samples without in vitro culture. The cartilage-specific (V + C)-isoform was suppressed in both normal and inflamed joints by a single intraarticular injection (0.1 mg/kg) of methylprednisolone. Combined, these data indicate that methylprednisolone suppresses matrix protein markers of chondrocytic differentiation. Decreased and altered chondrocyte expression of matrix proteins likely contributes to the pathogenesis of corticosteroid-induced cartilage degeneration.

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Section: Discussionmentioning

confidence: 95%

Corticosteroids alter the differentiated phenotype of articular chondrocytes

Fubini

Todhunter

Burton‐Wurster

et al. 2001

Journal Orthopaedic Research

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“…After removal of articular chondrocytes from their matrix, normal chondrocytes regain the ability to proliferate and will divide freely in certain culture environments (18). However, release of chondrocytes from their matrix with subsequent growth in monolayer culture also results in modulation of the phenotype.…”

Section: Discussionmentioning

confidence: 99%

“…However, release of chondrocytes from their matrix with subsequent growth in monolayer culture also results in modulation of the phenotype. This "dedifferentiation" is manifest as loss of type-I1 collagen and aggrecan production and upregulation of type-I collagen (18). Attempts to study normal chondrocyte matrix component synthesis in vitro have resulted in development of culture techniques using agarose or alginate beads to entrap the chondrocytes (1,3,5,6,16).…”

Section: Discussionmentioning

confidence: 99%

“…In these three-dimensional culture systems, the chondrocytes appear to retain their phenotype. Culture of high-density chondrocytes also slows the loss of chondrocyte phenotype and matrix synthesis more consistently than monolayer cultures (18,23). The stimulus in the altered environment of the culture systems that slows or prevents dedifferentiation is not known.…”

Section: Discussionmentioning

confidence: 99%

“…Interestingly, both control and rhBMP-2-treated cultures expressed low levels of type-I collagen message after 7 days in culture. The expression of type-I collagen has recently been reported, even in freshly isolated chondrocytes (18,20), but whether this indicates protein secretion is as yet unclear. Reduction in the type-I collagen gene expression at 28 days may be a result of the added influence of increased cell density helping to maintain the cartilage phenotype.…”

Section: Discussionmentioning

confidence: 99%

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Recombinant human bone morphogenetic protein‐2 maintains the articular chondrocyte phenotype in long‐term culture

Sailor¹,

Hewick²,

Morris³

1996

Journal Orthopaedic Research

134

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Summary: Bone morphogenetic proteins have been shown to increase matrix synthesis by articular chondrocytes in short-term cultures. Members of this family of proteins have also been shown to induce endochondral ossification in vivo. The present study was performed to determine if the addition of human recombinant bone morphogenetic protein-2 to a long-term monolayer articular chondrocyte cell culture system affected the ability of the chondrocytes to divide in vitro, whether the cytokine altered expression of the articular chondrocyte phenotype and synthesis of matrix proteoglycans, and whether the cytokine was capable of inducing differentiation to a hypertrophic chondrocyte. Human recombinant bone morphogenetic protein-2 did not alter cell proliferation. It caused 3.5-6.2 times more proteoglycan synthesis by articular chondrocytes during each of the time points tested after 4 days in culture. Total proteoglycan accumulation in the extracellular matrix after 28 days in culture was 6.7 times as great in the treated cultures as in the control. Treatment with human recombinant bone morphogenetic protein-2 maintained the articular chondrocyte phenotype of cells in culture as demonstrated by Northern blot analysis: the expression of type-I collagen genes was increased and that of type-I1 collagen and aggrecan mRNA was lost in untreated chondrocyte cultures after 14-21 days in culture. In contrast, exposure to 100 ngiml human recombinant bone morphogenetic protein-2 maintained expression of type-I1 collagen and increased expression of aggrecan compared with controls during the 28-day culture period. Northern blot analysis of the expression of type-X collagen and osteocalcin by chondrocytes treated with human recombinant bone morphogenetic protein-2 showed a lack of expression of these genes, indicating no alteration in phenotype. These experiments demonstrated the ability of human recombinant bone morphogenetic protein-2 to promote the articular chondrocyte phenotype and matrix synthesis in long-term culture. Characteristics of cell growth were not affected, and the cytokine did not induce differentiation to a hypertrophic chondrocyte. ~Members of the bone morphogenetic protein (BMP) subgroup of the transforming growth factorp superfamily of proteins are believed to play important roles in embryonic development (24). They have also received attention because of their ability to affect adult tissues: BMPs induce new endochondral bone formation at ectopic (32,35) and skeletal (7,12,15) synthesized by the chondrocytes themselves. The specific biochemical and mechanical components of the matrix impart the characteristics necessary for frictionfree and painless movement of joints. The limited capacity of damaged adult articular cartilage to repair has stimulated many investigations into ways of modulating chondrocyte metabolism, some of which have involved exploration of the effect of BMPs. Osteogenin and BMP-4 have been shown to increase collagen and proteoglycan synthesis by cartilage explants and by chondrocytes in sh...

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