Control of extracellular matrix homeostasis of normal cartilage by a TGFβ autocrine pathway. Validation of flow cytometry as a tool to study chondrocyte metabolism in vitro

Wang, Lai; Almqvist, Karl; Veys, Eric; Verbruggen, G.

doi:10.1053/joca.2001.0492

Cited by 41 publications

(23 citation statements)

References 46 publications

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“…This finding is consistent with the aforementioned studies. Moreover, MMP-1 levels were much lower than those of MMP-3, confirming the results of the previous report [44]. This work shows the important role of MMPs in loading response in vivo .…”

Section: Discussionsupporting

confidence: 91%

Decreased metalloproteinase production as a response to mechanical pressure in human cartilage: a mechanism for homeostatic regulation

et al. 2006

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Articular cartilage is optimised for bearing mechanical loads. Chondrocytes are the only cells present in mature cartilage and are responsible for the synthesis and integrity of the extracellular matrix. Appropriate joint loads stimulate chondrocytes to maintain healthy cartilage with a concrete protein composition according to loading demands. In contrast, inappropriate loads alter the composition of cartilage, leading to osteoarthritis (OA). Matrix metalloproteinases (MMPs) are involved in degradation of cartilage matrix components and have been implicated in OA, but their role in loading response is unclear. With this study, we aimed to elucidate the role of MMP-1 and MMP-3 in cartilage composition in response to mechanical load and to analyse the differences in aggrecan and type II collagen content in articular cartilage from maximum-and minimum-weight-bearing regions of human healthy and OA hips. In parallel, we analyse the apoptosis of chondrocytes in maximal and minimal load areas. Because human femoral heads are subjected to different loads at defined sites, both areas were obtained from the same hip and subsequently evaluated for differences in aggrecan, type II collagen, MMP-1, and MMP-3 content (enzyme-linked immunosorbent assay) and gene expression (real-time polymerase chain reaction) and for chondrocyte apoptosis (flow cytometry, bcl-2 Western blot, and mitochondrial membrane potential analysis). The results showed that the load reduced the MMP-1 and MMP-3 synthesis (p < 0.05) in healthy but not in OA cartilage. No significant differences between pressure areas were found for aggrecan and type II collagen gene expression levels. However, a trend toward significance, in the aggrecan/ collagen II ratio, was found for healthy hips (p = 0.057) upon comparison of pressure areas (loaded areas > non-loaded areas). Moreover, compared with normal cartilage, OA cartilage showed a 10-to 20-fold lower ratio of aggrecan to type II collagen, suggesting that the balance between the major structural proteins is crucial to the integrity and function of the tissue. Alternatively, no differences in apoptosis levels between loading areas were found -evidence that mechanical load regulates cartilage matrix composition but does not affect chondrocyte viability. The results suggest that MMPs play a key role in regulating the balance of structural proteins of the articular cartilage matrix according to local mechanical demands.

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Section: Discussionsupporting

confidence: 91%

Decreased metalloproteinase production as a response to mechanical pressure in human cartilage: a mechanism for homeostatic regulation

et al. 2006

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“…MMP-1 is an important component in cartilage degradation and tissue remodeling, whereas TIMP-1 acts to keep the effects of MMP-1 in check (Wang et al 2002). Together, the balance of MMP-1 and TIMP-1 mediates matrix degradation in cartilage.…”

Section: Discussionmentioning

confidence: 99%

Gene expression of single articular chondrocytes

2006

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Although previous studies in the field of tissue engineering have provided important information about articular cartilage, their conclusions are based on population averages and do not account for variations in cell subpopulations. To obtain a precise understanding of chondrocytes, we investigated the effects of cartilage zone and seeding duration on single chondrocyte gene expression to select an optimal zone for tissue engineering (Phase I), followed by an evaluation of growth factor exposure on the zone selected in Phase I (Phase II). In Phase I, superficial and middle/deep bovine articular chondrocytes were seeded in monolayers for 3 or 18 h. In Phase II, middle/deep chondrocytes (selected in Phase I) received 100 ng/ml insulin-like growth factor-I (IGF-I) for 3 h. Real-time reverse transcription/polymerase chain reaction was used to quantify the abundance of D-glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and the relative abundances of aggrecan, collagens I and II, cartilage oligomeric matrix protein (COMP), matrix metalloproteinase-1 (MMP-1), and tissue inhibitor of metalloproteinase-1 (TIMP-1). GAPDH varied zonally, but neither time nor IGF-I had an effect on it, suggesting that GAPDH is a suitable housekeeping gene for comparisons within each zone, but not across zones. IGF-I increased the expression of aggrecan and collagen II in middle/deep chondrocytes seeded for 18 h. TIMP-1 expression increased with time in control cells, suggesting that chondrocytes enter a matrix protective state after seeding. IGF-I diminished this effect, suggesting that treatment with IGF-I refocuses chondrocytes on matrix production rather than on protection from metalloproteinases. Concomitant to increasing TIMP-1, MMP-1 was detectable by 18 h in superficial cells, providing further evidence of a trend toward matrix degradation with time. Collagen I was undetected in all cells, and no differences were observed for COMP, confirming that no dedifferentiation or osteoarthritic changes occurred. Taken together, these results establish a unique understanding of individual chondrocyte behavior.

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“…40,41 MMP-1 is also reported to be downregulated in PC-3 cells following TGFb1 treatment; 20 however, we found no detectable levels of MMP-1 at the concentration of media we used. In the stromal-epithelia context we have analyzed, it appears that the relative contribution of MMP-1 from PC-3 cells alone is minor, although MMP-1 secretion by prostate cancer cells could certainly play important roles in other stages of metastasis.…”

Section: Discussionmentioning

confidence: 53%

Growth factor regulation of secreted matrix metalloproteinase and plasminogen activators in prostate cancer cells, normal prostate fibroblasts and normal osteoblasts

Forbes

Webb

Sehgal

2003

Prostate Cancer Prostatic Dis

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We assessed the relative levels of secreted matrix metalloproteinases (MMPs) and plasminogen activators (PAs) in PC-3 cells, prostate fibroblasts and osteoblasts in the presence and absence of VEGF, TGFb1 and bFGF. Fibroblasts and osteoblasts secreted more MMPs -1 and -2 than did PC-3 cells, while PC-3 s contributed the majority of PAs. MMP-1 expression was downregulated by transforming growth factor b-1 (TGFb1) treatment in prostate fibroblasts and upregulated by basic fibroblast growth factor (bFGF) in both stromal lines. In PC-3 cells, TGFb1 and bFGF increased urokinase plasminogen activator secretion. TGFb1 decreased tissue plasminogen activator secretion in all cell lines. Prostate cancer cells associated with fibroblasts or osteoblasts have a variety of MMPs and PAs to facilitate matrix degradation.

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Control of extracellular matrix homeostasis of normal cartilage by a TGFβ autocrine pathway. Validation of flow cytometry as a tool to study chondrocyte metabolism in vitro

Cited by 41 publications

References 46 publications

Decreased metalloproteinase production as a response to mechanical pressure in human cartilage: a mechanism for homeostatic regulation

Decreased metalloproteinase production as a response to mechanical pressure in human cartilage: a mechanism for homeostatic regulation

Gene expression of single articular chondrocytes

Growth factor regulation of secreted matrix metalloproteinase and plasminogen activators in prostate cancer cells, normal prostate fibroblasts and normal osteoblasts

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