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Peter Ghosh - One of the best experts on this subject based on the ideXlab platform.
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effects of transforming growth factor beta on proteoglycan synthesis by cell and explant cultures derived from the knee joint meniscus
Osteoarthritis and Cartilage, 1995Co-Authors: Simon Collier, Peter GhoshAbstract:Repair of meniscal tears depends in part upon the ability of the resident fibrochondrocytes to produce new extracellular matrix molecules including Proteoglycans. Three culture systems have been used to investigate proteoglycan production by meniscal fibrochondrocytes from the inner, middle and outer zones of medial and lateral menisci of the sheep stifle joint. Cultures of meniscal explants, monolayered cells, and cells encapsulated in alginate beads were labeled with 35SO4H2 for 48 h in the absence and presence of transforming growth factor beta (TGF beta) and the Proteoglycans were analysed by Sephacryl S-1000 chromatography. In general, the lateral meniscus produced more proteoglycan than the medial. Explants from the inner and middle zones produced predominantly aggrecan-like proteoglycan, together with a smaller proteoglycan population eluting with an average distribution coefficient of around 0.65. The outer meniscal zones synthesized less proteoglycan overall, the majority of which consisted of the smaller Proteoglycans. These characteristic proteoglycan size profiles obtained with explant cultures also were preserved when cells isolated from the respective zones were cultured in alginate beads. Monolayer cell cultures, however, produced almost entirely small Proteoglycans, regardless of their zone of origin. Chromatography of chondroitinase AC and ABC digested samples indicated that the small proteoglycan population comprised mostly dermatan sulphate-containing Proteoglycans. In all meniscal zones and in all culture systems, TGF beta stimulated proteoglycan production by up to 100% and the Proteoglycans were slightly larger. TGF beta also stimulated cell division in fibrochondrocyte monolayer cultures. Long term intermittent stimulation of alginate bead cultures with TGF beta resulted in large increases in proteoglycan synthesis, increased aggregation of large proteoglycan monomers, and an increase in the production of the larger of two small Proteoglycans, putatively, biglycan.
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osteoarthritis and cartilage effects of transforming growth factor beta on proteoglycan synthesis by cell and explant cultures derived from the knee joint meniscus
1995Co-Authors: Simon Collier, Peter Ghosh, Raymond PurvesAbstract:Summary Repair of meniscal tears depends in part upon the ability of the resident fibrochondrocytes to produce new extracellular matrix molecules including Proteoglycans. Three culture systems have been used to investigate proteoglycan production by meniscal fibrochondrocytes from the inner, middle and outer zones of medial and lateral menisci of the sheep stifle joint. Cultures of meniscal explants, monolayered cells, and cells encapsulated in alginate beads were labeled with 3~SO4H~ for 48h in the absence and presence of tranforming growth factor beta (TGFfl) and the Proteoglycans were analysed by Sephacryl S-1000 chromatography. In general, the lateral meniscus produced more proteoglycan than the medial. Explants from the inner and middle zones produced predominantly aggrecan-like proteoglycan, together with a smaller proteoglycan population eluting with an average distribution coefficient of around 0.65. The outer meniscal zones synthesized less proteoglycan overall, the majority of which consisted of the smaller Proteoglycans. These characteristic proteoglycan size profiles obtained with explant cultures also were preserved when cells isolated from the respective zones were cultured in alginate beads. Monolayer cell cultures, however, produced almost entirely small Proteoglycans, regardless of their zone of origin. Chromatography of chondroitinase AC and ABC digested samples indicated that the small proteoglycan population comprised mostly dermatan sulphate-containing Proteoglycans. In all meniscal zones and in all culture systems, TGFfl stimulated proteoglycan production by up to 100% and the Proteoglycans were slightly larger. TGFfl also stimulated cell division in fibrochondrocyte monolayer cultures. Long term intermittent stimulation of alginate bead cultures with TGFfl resulted in large increases in proteoglycan synthesis, increased aggregation of large proteoglycan monomers, and an increase in the production of the larger of two small Proteoglycans, putatively, biglycan.
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comparison of the effects of non steroidal anti inflammatory drugs nsaids on proteoglycan synthesis by articular cartilage explant and chondrocyte monolayer cultures
Biochemical Pharmacology, 1991Co-Authors: Simon Collier, Peter GhoshAbstract:Abstract Experiments were conducted to study proteoglycan biosynthesis by rabbit articular chondrocytes cultured in the presence of NSAIDs and 35 SO 2− 4 for up to 8 days. Both articular cartilage explants and confluent chondrocyte monolayer culture models were used. Medium was changed every 2 days and the [ 35 SO 4 ]Proteoglycans which had accumulated in the medium and the extracellular matrix during the culture intervals were assayed separately. In long-term experiments, drugs were removed on day 8, and proteoglycan production during a 10–12 day culture interval also was assayed. The drugs studied were diclofenac, indomethacin, ketoprofen, piroxicam and tiaprofenic acid, at concentrations of 0, 0.1, 1, 10, 50 and 100μg/mL. Whereas proteoglycan production by cell cultures was maximal early in the culture period, explants produced more Proteoglycans as time progressed. The highest concentrations of all of the drugs, especially diclofenac and indomethacin, inhibited proteoglycan secretion by both cell and explant cultures. However, after removal of the drugs from the cultures, suppressed proteoglycan production reversed to levels equivalent to, or higher than controls in the cell cultures, but largely persisted in explant cultures. About 70–80% of Proteoglycans produced by explants were retained in the matrix, whereas about 80–90% of Proteoglycans produced by cell cultures were secreted into the medium. Where drugs inhibited proteoglycan production, the levels were reduced by approximately the same proportions in both extracellular matrix and culture medium fractions. Of the NSAIDs examined only ketoprofen demonstrated a stimulatory effect on PG synthesis in explant cultures at a physiological concentration (0.1μg/mL).
Hans J Hausselmann - One of the best experts on this subject based on the ideXlab platform.
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differential effects of igf 1 ans tgfβ 2 on the assembly of Proteoglycans in pericellular and territorial matrix by cultured bovine articular chondrocytes
Osteoarthritis and Cartilage, 1998Co-Authors: Gerjo J V M Van Osch, Wim B Van Den Berg, Ernst B Hunziker, Hans J HausselmannAbstract:Objectives: Knowledge of matrix assembly is necessary to understand the pathogenesis of disease processes and to find solutions for repair of articular cartilage lesions. The influence of growth factors on matrix assembly is largely unknown. We investigated whether, and to what degree, insulin-like growth factor (IGF-1) and transforming growth factor β-2 (TGFβ-2) influence proteoglycan synthesis and accumulation in the cell-associated matrix compartment (consisting of pericellular and territorial matrix) compared to the further-removed matrix compartment (consisting of the interterritorial matrix). Design: Bovine articular chondrocytes were cultured in alginate beads for day 13. The effect of addition of 25 ng/ml IGF-1 or 25 ng/ml TGFβ-2 during the last 7 days in culture was determined. Cell-associated and further-removed matrix compartments were separated by centrifugation after sodium citrate/EDTA treatment. The amount of DNA, the total amount of Proteoglycans and the amount of newly synthesized Proteoglycans were analyzed biochemically. Morphometric analysis on electron micrographs was used to calculate the volumes of the cell-associated and further-removed matrix components. Results: It was demonstrated in control beads that 25±8% of the Proteoglycans were laid down in the cell-associated matrix compartment compared with 75±8% in the further-removed matrix compartment. The cell-associated matrix compartment in intact beads could be recognized in electron microscopy by a delineation of dense amorph material. Morphometric evaluation showed a relative volume of the cell-associated matrix compartment of 5.2±0.6% compared with 91.3±0.8% of the further-removed matrix compartment and 3.5±0.3% of the area occupied by cells. Combination of biochemical and morphometric results showed that the concentration of Proteoglycans in the cell-associated matrix compartment was 3.63±0.32 mg/ml. By adding IGF-1 or TGFβ-2, the amount of both total accumulated Proteoglycans and newly synthesized [35S]Proteoglycans at day 13 in culture increased. In addition to an overall rise in proteoglycan content, IGF-1 significantly increased (24%) the percentage of Proteoglycans laid down in the cell-associated matrix compartment while not changing the relative volume of this compartment (5.2±0.8%). This leads to a 82% (P<0.05) increase in the proteoglycan concentration in the cell-associated matrix compartment compared to control beads. In contrast, TGFβ-2 significantly decreased (24%) the relative amount of Proteoglycans in the cell-associated matrix compartment which was paralleled by a reduction of the relative volume from 5.2±0.6 to 3.6±1.4%. This leads to a significant increase of 87% of the proteoglycan concentration in the cell-association matrix compartment. Conclusions: This study demonstrates that both IGF-1 and TGFβ-2 significantly but differently influence proteoglycan synthesis and accumulation in the cell-associated matrix compartment of cultured bovine chondrocytes in alginate. Both growth factors increase the concentration of Proteoglycans in the cell-associated matrix compartment. However, addition of TGFβ-2 to bovine articular chondrocytes cultured in alginate beads for 13 days results in a significant reduction of the relative volume of the pericellular matrix compartment compared to controls, indicating differences in assembly of the matrix.
Simon Collier - One of the best experts on this subject based on the ideXlab platform.
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effects of transforming growth factor beta on proteoglycan synthesis by cell and explant cultures derived from the knee joint meniscus
Osteoarthritis and Cartilage, 1995Co-Authors: Simon Collier, Peter GhoshAbstract:Repair of meniscal tears depends in part upon the ability of the resident fibrochondrocytes to produce new extracellular matrix molecules including Proteoglycans. Three culture systems have been used to investigate proteoglycan production by meniscal fibrochondrocytes from the inner, middle and outer zones of medial and lateral menisci of the sheep stifle joint. Cultures of meniscal explants, monolayered cells, and cells encapsulated in alginate beads were labeled with 35SO4H2 for 48 h in the absence and presence of transforming growth factor beta (TGF beta) and the Proteoglycans were analysed by Sephacryl S-1000 chromatography. In general, the lateral meniscus produced more proteoglycan than the medial. Explants from the inner and middle zones produced predominantly aggrecan-like proteoglycan, together with a smaller proteoglycan population eluting with an average distribution coefficient of around 0.65. The outer meniscal zones synthesized less proteoglycan overall, the majority of which consisted of the smaller Proteoglycans. These characteristic proteoglycan size profiles obtained with explant cultures also were preserved when cells isolated from the respective zones were cultured in alginate beads. Monolayer cell cultures, however, produced almost entirely small Proteoglycans, regardless of their zone of origin. Chromatography of chondroitinase AC and ABC digested samples indicated that the small proteoglycan population comprised mostly dermatan sulphate-containing Proteoglycans. In all meniscal zones and in all culture systems, TGF beta stimulated proteoglycan production by up to 100% and the Proteoglycans were slightly larger. TGF beta also stimulated cell division in fibrochondrocyte monolayer cultures. Long term intermittent stimulation of alginate bead cultures with TGF beta resulted in large increases in proteoglycan synthesis, increased aggregation of large proteoglycan monomers, and an increase in the production of the larger of two small Proteoglycans, putatively, biglycan.
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osteoarthritis and cartilage effects of transforming growth factor beta on proteoglycan synthesis by cell and explant cultures derived from the knee joint meniscus
1995Co-Authors: Simon Collier, Peter Ghosh, Raymond PurvesAbstract:Summary Repair of meniscal tears depends in part upon the ability of the resident fibrochondrocytes to produce new extracellular matrix molecules including Proteoglycans. Three culture systems have been used to investigate proteoglycan production by meniscal fibrochondrocytes from the inner, middle and outer zones of medial and lateral menisci of the sheep stifle joint. Cultures of meniscal explants, monolayered cells, and cells encapsulated in alginate beads were labeled with 3~SO4H~ for 48h in the absence and presence of tranforming growth factor beta (TGFfl) and the Proteoglycans were analysed by Sephacryl S-1000 chromatography. In general, the lateral meniscus produced more proteoglycan than the medial. Explants from the inner and middle zones produced predominantly aggrecan-like proteoglycan, together with a smaller proteoglycan population eluting with an average distribution coefficient of around 0.65. The outer meniscal zones synthesized less proteoglycan overall, the majority of which consisted of the smaller Proteoglycans. These characteristic proteoglycan size profiles obtained with explant cultures also were preserved when cells isolated from the respective zones were cultured in alginate beads. Monolayer cell cultures, however, produced almost entirely small Proteoglycans, regardless of their zone of origin. Chromatography of chondroitinase AC and ABC digested samples indicated that the small proteoglycan population comprised mostly dermatan sulphate-containing Proteoglycans. In all meniscal zones and in all culture systems, TGFfl stimulated proteoglycan production by up to 100% and the Proteoglycans were slightly larger. TGFfl also stimulated cell division in fibrochondrocyte monolayer cultures. Long term intermittent stimulation of alginate bead cultures with TGFfl resulted in large increases in proteoglycan synthesis, increased aggregation of large proteoglycan monomers, and an increase in the production of the larger of two small Proteoglycans, putatively, biglycan.
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comparison of the effects of non steroidal anti inflammatory drugs nsaids on proteoglycan synthesis by articular cartilage explant and chondrocyte monolayer cultures
Biochemical Pharmacology, 1991Co-Authors: Simon Collier, Peter GhoshAbstract:Abstract Experiments were conducted to study proteoglycan biosynthesis by rabbit articular chondrocytes cultured in the presence of NSAIDs and 35 SO 2− 4 for up to 8 days. Both articular cartilage explants and confluent chondrocyte monolayer culture models were used. Medium was changed every 2 days and the [ 35 SO 4 ]Proteoglycans which had accumulated in the medium and the extracellular matrix during the culture intervals were assayed separately. In long-term experiments, drugs were removed on day 8, and proteoglycan production during a 10–12 day culture interval also was assayed. The drugs studied were diclofenac, indomethacin, ketoprofen, piroxicam and tiaprofenic acid, at concentrations of 0, 0.1, 1, 10, 50 and 100μg/mL. Whereas proteoglycan production by cell cultures was maximal early in the culture period, explants produced more Proteoglycans as time progressed. The highest concentrations of all of the drugs, especially diclofenac and indomethacin, inhibited proteoglycan secretion by both cell and explant cultures. However, after removal of the drugs from the cultures, suppressed proteoglycan production reversed to levels equivalent to, or higher than controls in the cell cultures, but largely persisted in explant cultures. About 70–80% of Proteoglycans produced by explants were retained in the matrix, whereas about 80–90% of Proteoglycans produced by cell cultures were secreted into the medium. Where drugs inhibited proteoglycan production, the levels were reduced by approximately the same proportions in both extracellular matrix and culture medium fractions. Of the NSAIDs examined only ketoprofen demonstrated a stimulatory effect on PG synthesis in explant cultures at a physiological concentration (0.1μg/mL).
Lisa R Tannock - One of the best experts on this subject based on the ideXlab platform.
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serum amyloid a but not c reactive protein stimulates vascular proteoglycan synthesis in a pro atherogenic manner
American Journal of Pathology, 2008Co-Authors: Patricia G Wilson, Lisa R Tannock, Joel C Thompson, Nancy R Webb, Frederick C De Beer, Victoria L KingAbstract:Inflammatory markers serum amyloid A (SAA) and C-reactive protein (CRP) are predictive of cardiac disease and are proposed to play causal roles in the development of atherosclerosis, in which the retention of lipoproteins by vascular wall Proteoglycans is critical. The purpose of this study was to determine whether SAA and/or CRP alters vascular proteoglycan synthesis and lipoprotein retention in a pro-atherogenic manner. Vascular smooth muscle cells were stimulated with either SAA or CRP (1 to 100 mg/L) and Proteoglycans were then isolated and characterized. SAA, but not CRP, increased proteoglycan sulfate incorporation by 50 to 100% in a dose-dependent manner ( P d , 29 μg/ml LDL versus 90 μg/ml LDL for SAA versus control Proteoglycans; P in vivo , ApoE −/− mice were injected with an adenovirus expressing human SAA-1, a null virus, or saline. Mice that received adenovirus expressing SAA had increased TGF-β concentrations in plasma and increased aortic biglycan content compared with mice that received either null virus or saline. Thus, SAA alters vascular Proteoglycans in a pro-atherogenic manner via the stimulation of TGF-β and may play a causal role in the development of atherosclerosis.
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Proteoglycans synthesized by arterial smooth muscle cells in the presence of transforming growth factor β1 exhibit increased binding to ldls
Arteriosclerosis Thrombosis and Vascular Biology, 2002Co-Authors: Peter J Little, Alan Chait, Lisa R Tannock, Katherine L Olin, Thomas N WightAbstract:The "response-to-retention" hypothesis of atherogenesis states that atherogenic lipoproteins, such as low density lipoprotein (LDL), are retained in vessels by Proteoglycans and undergo proatherosclerotic modifications. Transforming growth factor (TGF)-1 has been identified in atherosclerotic vessels and has been shown to stimulate the synthesis of chondroitin sulfate- and dermatan sulfate- containing Proteoglycans by arterial smooth muscle cells (ASMCs), but whether it promotes lipid retention has not been addressed. We investigated whether TGF-1 modulates the biosynthesis of Proteoglycans by ASMCs in a manner that promotes binding to LDL. Proteoglycans isolated from TGF-1-treated ASMCs exhibited enhanced binding to native LDL compared with the binding of Proteoglycans isolated from control cultures (Kd 18 g/mL LDL versus 81 g/mL LDL, respectively). The increase in proteoglycan-LDL binding caused by TGF-1 could be attributed primarily to the glycosaminoglycan portion of the Proteoglycans, since the glycosaminoglycan chains liberated from the core proteins of these Proteoglycans synthesized in the presence of TGF-1 exhibited increased LDL binding as well. Furthermore, glycosaminoglycan chains initiated on xyloside (an initiator of glycosaminoglycan synthesis) in the presence of TGF-1 were longer and displayed enhanced binding to LDL compared with the LDL binding of xyloside-initiated glycosaminoglycan chains from control cultures. These results indicate that TGF-1 promotes LDL-proteoglycan interaction primarily by its effects on the glycosaminoglycan synthetic machinery of the ASMCs. Therefore, this study supports a proatherogenic role for TGF-1. (Arterioscler Thromb Vasc Biol. 2002;22:55-60.)
Ernst B Hunziker - One of the best experts on this subject based on the ideXlab platform.
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differential effects of igf 1 ans tgfβ 2 on the assembly of Proteoglycans in pericellular and territorial matrix by cultured bovine articular chondrocytes
Osteoarthritis and Cartilage, 1998Co-Authors: Gerjo J V M Van Osch, Wim B Van Den Berg, Ernst B Hunziker, Hans J HausselmannAbstract:Objectives: Knowledge of matrix assembly is necessary to understand the pathogenesis of disease processes and to find solutions for repair of articular cartilage lesions. The influence of growth factors on matrix assembly is largely unknown. We investigated whether, and to what degree, insulin-like growth factor (IGF-1) and transforming growth factor β-2 (TGFβ-2) influence proteoglycan synthesis and accumulation in the cell-associated matrix compartment (consisting of pericellular and territorial matrix) compared to the further-removed matrix compartment (consisting of the interterritorial matrix). Design: Bovine articular chondrocytes were cultured in alginate beads for day 13. The effect of addition of 25 ng/ml IGF-1 or 25 ng/ml TGFβ-2 during the last 7 days in culture was determined. Cell-associated and further-removed matrix compartments were separated by centrifugation after sodium citrate/EDTA treatment. The amount of DNA, the total amount of Proteoglycans and the amount of newly synthesized Proteoglycans were analyzed biochemically. Morphometric analysis on electron micrographs was used to calculate the volumes of the cell-associated and further-removed matrix components. Results: It was demonstrated in control beads that 25±8% of the Proteoglycans were laid down in the cell-associated matrix compartment compared with 75±8% in the further-removed matrix compartment. The cell-associated matrix compartment in intact beads could be recognized in electron microscopy by a delineation of dense amorph material. Morphometric evaluation showed a relative volume of the cell-associated matrix compartment of 5.2±0.6% compared with 91.3±0.8% of the further-removed matrix compartment and 3.5±0.3% of the area occupied by cells. Combination of biochemical and morphometric results showed that the concentration of Proteoglycans in the cell-associated matrix compartment was 3.63±0.32 mg/ml. By adding IGF-1 or TGFβ-2, the amount of both total accumulated Proteoglycans and newly synthesized [35S]Proteoglycans at day 13 in culture increased. In addition to an overall rise in proteoglycan content, IGF-1 significantly increased (24%) the percentage of Proteoglycans laid down in the cell-associated matrix compartment while not changing the relative volume of this compartment (5.2±0.8%). This leads to a 82% (P<0.05) increase in the proteoglycan concentration in the cell-associated matrix compartment compared to control beads. In contrast, TGFβ-2 significantly decreased (24%) the relative amount of Proteoglycans in the cell-associated matrix compartment which was paralleled by a reduction of the relative volume from 5.2±0.6 to 3.6±1.4%. This leads to a significant increase of 87% of the proteoglycan concentration in the cell-association matrix compartment. Conclusions: This study demonstrates that both IGF-1 and TGFβ-2 significantly but differently influence proteoglycan synthesis and accumulation in the cell-associated matrix compartment of cultured bovine chondrocytes in alginate. Both growth factors increase the concentration of Proteoglycans in the cell-associated matrix compartment. However, addition of TGFβ-2 to bovine articular chondrocytes cultured in alginate beads for 13 days results in a significant reduction of the relative volume of the pericellular matrix compartment compared to controls, indicating differences in assembly of the matrix.
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mechanical compression alters proteoglycan deposition and matrix deformation around individual cells in cartilage explants
Journal of Cell Science, 1998Co-Authors: Thomas M Quinn, Alan J Grodzinsky, Michael D Buschmann, Ernst B HunzikerAbstract:JCS4443 We have used new techniques of cell-length scale quantitative autoradiography to assess matrix synthesis, deposition, and deformation around individual chondrocytes in mechanically compressed cartilage explants. Our objectives were to: (1) quantify the effects of static and dynamic compression on the deposition of newly synthesized Proteoglycans into cell-associated and furtherremoved matrices; (2) measure cell-length scale matrix strains and morphological changes of the cell and matrix associated with tissue compression; and (3) relate microscopic physical stimuli to changes in proteoglycan synthesis as functions of compression level and position within mechanically compressed explants. Results indicate a high degree of structural organization in the extracellular matrix, with the pericellular matrix associated with the most rapid rates of proteoglycan deposition, and greatest sensitivity to mechanical compression. Static compression could stimulate directional deposition of secreted Proteoglycans around chondrocytes, superimposed on an inhibition of proteoglycan synthesis; these events followed trends for compressive strain in the cell-associated matrix. Conversely, proteoglycan synthesis and pericellular deposition was stimulated by dynamic compression. Results suggest that cell-matrix interactions in the cellassociated matrix may be a particularly important aspect of the chondrocyte response to mechanical compression, possibly involving macromolecular transport limitations and morphological changes associated with fluid flow and local compaction of the matrix around cells. SUMMARY
