Proteoglycan Release Research Articles

The proteoglycan metabolism, morphology and viability of articular cartilage treated with a synthetic matrix metalloproteinase inhibitor.

Matrix metalloproteinases (MMP) are among the key enzymes responsible for the proteolytic destruction of articular cartilage during chronic rheumatic diseases. Articular cartilage is one potential target for drugs designed to inhibit the activity of MMPs in order to stop or to slow down the proteolytic destruction of the extracellular matrix of cartilage. The purpose of this study was to investigate the effect of the synthetic inhibitor of MMPs U-24522 for its ability (1) to inhibit in vitro the activity of MMP-proteoglycanases; (2) to modulate the morphology and viability of cartilage explants; and (3) to modify the biosynthesis and release of proteoglycans from articular cartilage explants. U-24522 dose-dependently inhibited the activity of MMP-proteoglycanases and significantly reduced the release of proteoglycans from interleukin-1 treated bovine articular cartilage explants when tested at concentrations ranging from 10(-4) to 10(-9) M. This hydroxamic acid derivative proved not to be harmful to chondrocyte viability and cartilage morphology. In addition, U-24522 had no effect on the rate of proteoglycan biosynthesis of interleukin-1 treated cartilage explants and increased the percentage of newly synthesized proteoglycans to form macromolecular aggregates. Thus U-24522 combines direct inhibitory potential on the activity of MMP-proteoglycanases with the inhibition of interleukin-1 stimulated proteoglycan loss from articular cartilage explants without affecting the morphology, viability and biosynthesis of proteoglycans of bovine articular cartilage explants.

Differential regulation of neuronal proteoglycans by activation of excitatory amino acid receptors.

Cell surface proteoglycans (PGs) have been implicated in neuronal growth, migration and differentiation and can play pivotal roles both in cell-cell and cell-substrate interactions during the development of the nervous system. We have previously shown that glutamate activation of excitatory amino acid receptors induces the synthesis and release of PGs with neurite-promoting activity from hippocampal neurones. In this study, we have investigated the activity-dependent regulation of mRNA expression of two PGs in fetal hippocampal neurones using a competitive reverse transcriptase-polymerase chain reaction and correlated this expression with neuronal growth. Both cerebroglycan (CBG), a glycosylphatidylinositol-anchored heparan sulphate PG, and neurocan, a developmentally regulated chondroitin sulphate PG, are expressed in hippocampal neurones. Exposure of hippocampal neurones to 100 microM glutamate for 5 min resulted in an increase in CBG mRNA levels and an increase in axonal and dendritic length. The increase in CBG mRNA levels following glutamate exposure was mediated via both N-methyl-D-aspartate and metabotropic receptor activation.

Degradation of type II collagen, but not proteoglycan, correlates with matrix metalloproteinase activity in cartilage explant cultures.

To determine the contribution of certain matrix metalloproteinases (MMPs) to the degradation of proteoglycan and type II collagen in cartilage. Bovine nasal and articular cartilage explants were cultured with recombinant human interleukin-1 alpha (IL-1 alpha) for up to 4 weeks. Release of proteoglycan and type II collagen into the medium was determined by colorimetric assay and immunoassay, respectively. The activity of MMPs in the medium was assayed using a quenched fluorescent substrate, as well as with a collagen fibril assay, by zymography, and in Western immunoblots. In some experiments, the effects of specific MMP inhibitors on type II collagen degradation were studied. In cultures of nasal cartilage with IL-1 alpha, almost all the proteoglycan was released within the first week, whereas there was no detectable release of type II collagen for the first 2 weeks of culture. A rapid period of almost complete dissolution of the collagen occurred in the third or fourth week. MMP activity measured using a quenched fluorescent substrate was negligible during the first 2 weeks of culture but was substantially increased in the third week of culture, at the time of collagen degradation. Similarly, there was a large increase in collagenolytic activity (by collagen fibril assay) and gelatinolytic activity (by zymography) during the third week of culture. Articular cartilage cultured with IL-1 alpha lost proteoglycan progressively during the 4-week period; however, there was no loss of type II collagen from the matrix in that time and no significant increase in MMP activity. The loss of type II collagen from nasal cartilage stimulated with IL-1 alpha was inhibited by BB87, an inhibitor of both collagenases and gelatinases, and by BB3003, a selective inhibitor of gelatinase A. In Western immunoblots, procollagenase and active interstitial collagenase could be readily detected in nasal cartilage cultures. Some procollagenase 3 and active collagenase 3 was also shown to be present. MMP activity correlates with degradation of type II collagen, but not proteoglycan, in cartilage cultures. Interstitial collagenase, collagenase 3, and gelatinases are all likely to contribute to cleavage and removal of collagen from the cartilage matrix. The proteinase(s) responsible for aggrecan breakdown remains unclear.

Effects of the hydroxamic acid derivate Ro 31-4724 on the metabolism and morphology of interleukin-1-treated cartilage explants.

Matrix metalloproteinases (MMPs) belong to the key enzymes of the proteolytic destruction of cartilage matrix during chronic rheumatic diseases. Our work focused on the inhibitory potential of the hydroxamate Ro 31-4724 on the activity of MMP-proteoglycanases as well as on the viability, morphology and proteoglycan metabolism of interleukin-1 (IL-1)-treated bovine articular cartilage explants. The in vitro activity of MMP-proteoglycanases as well as the release of proteoglycans from IL-1-treated cartilage explants were significantly and concentration-dependently inhibited by Ro 31-4724 tested at concentrations ranging from 1 nmol/l to 10 mumol/l. Histopathological evaluation of sections from cartilage explants treated with this drug revealed no microscopically discernible alterations, and did not show any cytotoxic effects of Ro 31-4724. In addition, Ro 31-4724 had no effect on the rate of proteoglycan biosynthesis by IL-1-treated cartilage explants and increased the percentage of newly synthesized proteoglycans to form macromolecular aggregates. In conclusion, Ro 31-4724 displayed MMP-proteoglycanase inhibitory activity both in vitro and ex vivo and proved to be not harmful to the morphology, viability and proteoglycan biosynthesis of bovine articular cartilage explants.

The effect of TGF-beta 1 on cell proliferation and proteoglycan production in human melanoma cells depends on the degree of cell differentiation.

The effect of transforming growth factor beta 1 (TGF-beta 1) on cell proliferation, colony formation in soft agar and synthesis and structure of proteoglycans was studied in three human melanoma cell lines at different stages of differentiation: SK-mel-1.36-1-5 (early), SK-mel-3.44 (intermediate) and SK-mel-23 (late). TGF-beta 1 potently inhibited cell growth in monolayer as well as in soft agar. TGF-beta 1 increased the release of sulfated proteoglycans into the medium, including the cell-specific melanoma proteoglycan, mel-PG, and induced changes in disaccharide composition and sulfation of the glycosaminoglycan chains. In all the cases, the effect of TGF-beta 1 was more pronounced in the most undifferentiated cell line SK-mel-1.36-1-5 than in the SK-mel-3.44, whereas it had no effect on the most differentiated SK-mel-23 cells.

Inhibitors of collagenase but not of gelatinase reduce cartilage explant proteoglycan breakdown despite only low levels of matrix metalloproteinase activity

Aims-To investigate the level of matrix metalloproteinase activity during the time-course of cartilage explant proteoglycan breakdown; to determine the effects of selective small-molecule inhibitors of matrix metalloproteinases on proteoglycan degradation.Methods-The levels of matrix metalloproteinase activity in cartilage explant cultures and conditioned media were monitored by use of a quenched fluorescent substrate. The constants for inhibition of certain matrix metalloproteinases by a series of synthetic inhibitors were determined. Bovine and human cartilage explant cultures were treated with interleukin-1, tumor necrosis factor or retinoic acid and the amount of proteoglycan released into the culture medium in the absence and presence of the inhibitors was quantified. Control experiments, examining the inhibition of other proteinases, and investigating possible toxic or non-specific effects of the inhibitors, were carried out.Results-The profile of inhibition of proteoglycan release suggested the involvement of interstitial collagenase-like, rather than gelatinase- or possibly stromelysin-like, proteinases. No evidence was found for toxic or non-specific mechanisms of inhibition. Very low levels of activity of the known matrix metalloproteinases were present during the time-course of aggrecan breakdown.Conclusions-A novel collagenase-like proteinase(s) may be involved in cartilage proteoglycan breakdown. Gelatinase-type matrix metalloproteinases do not seem to be involved in this process. Specific collagenase inhibitors may be therapeutically efficacious in the treatment of arthritis.

Excitatory amino acid regulation of astrocyte proteoglycans

Activity-dependent enduring change in cellular communication is essential for specific connectivity during development of the nervous system and for adaptive responses of the mature nervous system. Here we report that glutamate activation of excitatory amino acid receptors induces the synthesis and release of proteoglycans (PGs) from fetal hippocampal astrocytes in dissociated culture. PG synthesis and release are mediated via kainate and metabotropic receptor activation. Glutamate exposure did not regulate the release of a specific family of PG, but glutamate inhibited the synthesis of heparan sulfate (HS) PGs that appeared within the extracellular environment of the astrocyte. Particulate protein kinase C (PKC) activity was increased by glutamate and the PKC activator phorbol 10-myristate 13-acetate produced a dose-dependent increase in PG release. However, glutamate-induced PG release was not blocked by inhibition of PKC activity. These data suggest that PKC activation can lead to PG release, but is not necessary for it. Activity-dependent influences on a class of substrate-bound molecular species with growth-modulatory properties may be involved in spatial regulation of neuronal growth responses produced by excitatory amino acids.

Association of Proteoglycan Degradation with Catabolic Cytokine and Stromelysin Release from Cartilage Cultured with Fibronectin Fragments

Addition of fibronectin fragments to bovine articular cartilage explant cultures results in enhanced release of metalloproteinases and rapid cartilage proteoglycan (PG) degradation and loss. The chondrolysis begins with rapid PG degradation which markedly slows after 1 week. Preliminary observations suggest that catabolic cytokines mediate chondrolytic activities of the fibronectin fragments. The objectives of this work were to investigate the correlations between: (a) release of specific cytokines; (b) release of the metalloproteinase (MMP), stromelysin-1 (MMP-3); (c) release of the tissue inhibitor of MMPs, TIMP-1, and; (d) degradation and release of PG from cultured cartilage. We report that human articular cartilage cultured with an amino-terminal 29-kDa fragment (Fn-f), at 0.1 μM, released enhanced levels of TNF-α, IL-1β, and IL-1α with peaks at Days 2, 3, and 9, respectively. MMP-3 release was elevated with a peak at Day 6 and a profile similar to that for the Fn-f-induced cartilage PG depletion. IL-6 release was enhanced within 2 days and continued at the same level throughout the culture period but this did not lead to enhanced release of TIMP-1, a known activity of IL-6. These data suggest that in the early chondrolytic events induced in cultured cartilage by Fn-f, enhanced MMP-3 release and maximal degradation and release of PG from cultured cartilage are kinetically associated with elevated release of the catabolic cytokines, TNF-α, IL-1β, and IL-1α. Further, a later period of slowing PG loss and slowing MMP-3 release is associated with greatly slowed release of these cytokines, but prolonged release of IL-6. This model of cartilage damage may be useful for studies of the interplay between cytokines and the effects of combinations of cytokines on cartilage homeostasis.

Shedding of heparan sulfate proteoglycan by stimulated endothelial cells: Evidence for proteolysis of cell‐surface molecules

Activation of endothelial cells by cytokines and endotoxin causes procoagulant and pro-inflammatory changes over a period of hours. We postulated that the same functional state might be achieved more rapidly by changes in the metabolism of heparan sulfate, which supports many of the normal functions of endothelial cells. We previously found that binding of anti-endothelial cell antibodies and activation of complement on endothelial cells causes the rapid shedding of endothelial cell heparan sulfate. Here we report the biochemical mechanism responsible for the release of the heparan sulfate. Stimulation of endothelial cells by anti-endothelial cell antibodies and complement resulted in the release of 35S-heparan sulfate proteoglycan and partially degraded 35S-heparan sulfate chains. Degradation of the 35S-heparan sulfate chains was not necessary for release since heparin and suramin prevented cleavage of the heparan sulfate but did not inhibit release from stimulated endothelial cells. The 35S-heparan sulfate proteoglycan released from endothelial cells originated from the cell surface and had a core protein similar in size (70.5 kD) to syndecan-1. Release was due to proteolytic cleavage of the protein core by serine and/or cysteine proteinases since the release of heparan sulfate was inhibited 87% by antipain and 53% by leupeptin. Release of heparan sulfate coincided with a decrease of ∼︁7 kD in the mass of the protein core and with a loss of hydrophobicity of the proteoglycan, consistent with the loss of the hydrophobic transmembrane domain. The cleavage and release of cell-surface 35S-heparan sulfate proteoglycan might be a novel mechanism by which endothelial cells may rapidly acquire the functional properties of activated endothelial cells. © 1996 Wiley-Liss, Inc.

Shedding of heparan sulfate proteoglycan by stimulated endothelial cells: evidence for proteolysis of cell-surface molecules.

Activation of endothelial cells by cytokines and endotoxin causes procoagulant and pro-inflammatory changes over a period of hours. We postulated that the same functional state might be achieved more rapidly by changes in the metabolism of heparan sulfate, which supports many of the normal functions of endothelial cells. We previously found that binding of anti-endothelial cell antibodies and activation of complement on endothelial cells causes the rapid shedding of endothelial cell heparan sulfate. Here we report the biochemical mechanism responsible for the release of the heparan sulfate. Stimulation of endothelial cells by anti-endothelial cell antibodies and complement resulted in the release of 35S-heparan sulfate proteoglycan and partially degraded 35S-heparan sulfate chains. Degradation of the 35S-heparan sulfate chains was not necessary for release since heparin and suramin prevented cleavage of the heparan sulfate but did not inhibit release from stimulated endothelial cells. The 35S-heparan sulfate proteoglycan released from endothelial cells originated from the cell surface and had a core protein similar in size (70.5 kD) to syndecan-1. Release was due to proteolytic cleavage of the protein core by serine and/or cysteine proteinases since the release of heparan sulfate was inhibited 87% by antipain and 53% by leupeptin. Release of heparan sulfate coincided with a decrease of approximately 7 kD in the mass of the protein core and with a loss of hydrophobicity of the proteoglycan, consistent with the loss of the hydrophobic transmembrane domain. The cleavage and release of cell-surface 35S-heparan sulfate proteoglycan might be a novel mechanism by which endothelial cells may rapidly acquire the functional properties of activated endothelial cells.

Effects of X Irradiation on Metabolism of Proteoglycans

The effects of X irradiation on matrix formation by growth-plate and articular chondrocytes, as reflected by metabolism of proteoglycans and type II collagen, were examined in a rabbit chondrocyte culture system. Irradiation with 1 to 10 Gy selectively inhibited synthesis of proteoglycans (incorporation of [35S]sulfate) depending on the stage of differentiation of the irradiated cells; however, synthesis of type II collagen was not affected. Irradiation of an immature culture, in which chondrocytes had just reached confluence, suppressed incorporation of [35S]sulfate into the glycosaminoglycan, 10 Gy inducing approximately 45-50% inhibition. In contrast, the irradiation of mature cultures, in which chondrocytes had already secreted extensive cartilage matrix, did not affect the rate of synthesis of proteoglycans (incorporation of [35S]sulfate). We also found that here irradiation stimulated the degradation of proteoglycans, but with the effect differing in growth-plate chondrocytes and articular chondrocytes. In growth-plate chondrocytes, cleavage from a site close to the G1 globular domain induced by 10 Gy enhanced the release of 35S-labeled proteoglycans into the medium, whereas in articular chondrocytes, irradiation had only marginal effects on the release of 35S-labeled proteoglycans. Our results show that irradiation with 1-10 Gy impaired proteoglycan metabolism in cartilage, with differing effects according to the stage of cell differentiation and the type of chondrocyte.

ONCOSTATIN M (OSM) STIMULATES RESORPTION AND INHIBITS SYNTHESIS OF PROTEOGLYCAN IN PORCINE ARTICULAR CARTILAGE EXPLANTS

Oncostatin M (OSM) is structurally and functionally similar to leukaemia inhibitory factor (LIF), interleukin 6 (IL-6), interleukin 11 (IL-11) and ciliary neurotrophic factor (CNTF). We have previously shown that LIF stimulates proteoglycan release and suppresses proteoglycan synthesis in pig and goat cartilage explants. The aim of this study was to determine whether OSM and related cytokines influence proteoglycan metabolism in pig cartilage explants. Slices of pig articluar cartilage were incubated for 6 days in serum free DMEM with or without cytokines. The total proteoglycan content in papain digested cartilage explants and medium was determined by the 1,9 dimethylmethylene blue method. Cytokine activity was assessed by determining the percentage release of total proteoglycan. To evaluate proteoglycan synthesis, cartilage was cultured for 48 h under the same conditions and in the final 6 h the tissue was cultured in sulphate free DMEM containing 35SO 4. The radioactivity in the medium and tissue was determined in cetylpyridinium chloride precipitates. Biosynthetic activity was expressed as DPM per mg wet weight of cartilage. Dose dependent stimulation of proteoglycan release and suppression of proteoglycan synthesis were observed with rhOSM. IL-6, IL-11 and CNTF also inhibited proteoglycan synthesis in a dose dependent manner but the degree of inhibition was less than that for OSM and these cytokines had no significant effect on proteoglycan release. New biological effects have been identified for OSM and the related cytokines CNTF and IL-11. All three of these cytokines, like LIF and IL-6, suppress proteoglycan synthesis in pig cartilage explants. This common effect suggests that the gp130 subunit of the receptors for these cytokines may represent a common signalling pathway whereby proteoglycan synthesis is regulated. Whilst OSM and LIF stimulate proteoglycan catabolism; IL-6 IL-11 and OSM do not. Thus these effects are not always coupled and activation of gp130 alone may not be a suffcient signal for proteoglycan catabolism.

AUTOCRINE INFLUENCES OF PROTEOGLYCANS ON ACTIVITY-DEPENDENT GROWTH RESPONSES OF FETAL HIPPOCAMPAL NEURONS. • 2277

Excitatory amino acid (EAA) neurotransmitters have been shown to play important roles as molecular signals influencing the structure of neurons during development. However, the signal transduction and effector mechanisms responsible for these effects have yet to be fully elucidated. We have previously provided evidence that EAA agonists induce the synthesis and release of proteoglycans (PGs) with neurite-promoting activity from fetal hippocampal neurons (Sugiura M., Dow K.E. Dev. Biol. 164:102, 1994). In the present studies, we examined expression of the neuron-specific growth associated gene Tα1 α-tubulin (Tα1) and genes that encode for proteins targetted to axons (growth-associated protein-43 (GAP-43)) and dendrites (microtubule-associated protein-2 (MAP-2)) following exposure to EAAs.

Effects of ipriflavone and its metabolites on human articular chondrocytes cultivated in clusters

Ipriflavone (IP) is an isoflavone derivative that was suggested to have bone-sparing effects in post-menopausal and senile osteoporosis. A moderate stimulatory effect of IP and its metabolites on proliferation of osteoblastic cells was reported in rat osteoblastic osteosarcoma cell line. We investigated the effects of different concentrations (0, 1, 10 and 100 micrograms/ml) of IP and its metabolites (MET I, II, III and V) on the incorporation of [3H] thymidine and production of proteoglycans (PG) and type II collagen (COL II) by human articular chondrocytes during a 12-day period, in a three-dimensional chondrocyte culture model. [3H]thymidine uptake was measured in chondrocyte clusters, and specific PG and COL II radioimmunoassays were performed every 4 days on the culture medium and cell clusters. Incubation with IP or its metabolites did not affect [3H]thymidine uptake regardless of the dose. PG released into the culture medium and PG cluster content rose significantly (P < 0.025) in presence of IP (1, 10 and 100 micrograms/ml). MET I increased PG release in culture medium (10 and 100 micrograms/ml) and PG cluster content (100 micrograms/ml). MET II has no effect on PG production. MET III increased PG in culture medium (100 microgram/ml) but did not influence PG cluster content while MET V (100 micrograms/ml) increased both PG release in culture medium and PG cluster content. COL II release in culture medium and COL II cluster content were significantly (P < 0.025) increased in presence of IP (10 and 100 micrograms/ml), MET III (1, 10 and 100 micrograms/ml) or MET V (100 micrograms/ml). MET I and II did not significantly affect COL II production.

The role of nitric oxide in proteoglycan turnover by bovine articular cartilage organ cultures.

Monolayer cultures of articular chondrocytes synthesize large amounts of nitric oxide (NO) following exposure to IL-1. The latter has antianabolic and procatabolic activities on these cells, but little is known about the role, if any, of NO in the integrated metabolic pathways of the chondrocyte. In the present study, the role of endogenously produced NO in both the synthesis and degradation of proteoglycans was investigated for the first time. Bovine articular cartilage slices exposed to 20 U/ml human rIL-1 beta (hrIL-1 beta) synthesized large amounts of NO for 1 to 2 days, after which production fell to a steady state level approximately 20% of the peak value for the remainder of the 14-day incubation. The NO synthase inhibitor, N-monomethyl L-arginine (L-NMA, 1 mM), blocked NO production and enhanced the acute catabolic effects of hrIL-1 beta in cartilage derived from both cartilage derived from both calves and adult animals. However, in late cultures, release of proteoglycans was reduced in the presence of L-NMA. The proteolytic activity in conditioned medium of these cultures (measured as caseinolytic activity) was enhanced by L-NMA; however, this inhibitor did not affect the rates of synthesis of proteoglycans. Although NO is widely assumed to be a mediator of cartilage catabolism, our data suggest that it may instead have an acute protective effect. Whether this effect is maintained chronically is less clear.

Proteoglycan-degrading activity associated with the 40 kDa collagen-binding fragment of fibronectin.

The osteoarthritis (OA) process is characterized by the progressive destruction of articular cartilage. There is a loss of cartilage proteoglycan content and disorganization of the collagen network, as well as an increase in other non-collagenous protein such as fibronectin (Fn). Increased proteolytic activity may lead to the degradation of native Fn and generation of Fn proteolytic fragments. Among them, the 45 kDa collagen-binding Fn fragment can be autoactivated in vitro into a 40 kDa fragment. This 40 kDa fragment induces an average of 30% of proteoglycan release per day from human OA cartilage explants and can degrade proteoglycan using dead cartilage sections. Proteoglycan-degrading activity related to the 40 kDa Fn fragment was decreased up to 66% by fetal calf serum (10%), but was not prevented by protein synthesis inhibitors (cycloheximide or actinomycin D). The action of this 40 kDa Fn fragment was greater on OA than on normal cartilage. This study suggests that enzymatic activity induced by the 40 kDa collagen-binding fragment of Fn might be involved in cartilage matrix turnover.

Signal transduction mechanisms subserving activity-dependent release of neuronal proteoglycans

We demonstrate here using dissociated hippocampal neurons that glutamate-induced release of proteoglycans which have been shown to have neurite growth-promoting activity is regulated by serine/threonine kinases of the protein kinase C and calcium/calmodulin type II kinase families, and that the state of phosphorylation of hippocampal neurons is a determinant of the magnitude and duration of the release response. Nitric oxide is also involved in mediating glutamate-induced PG release.

Preferential requirement for protein tyrosine phosphatase activity in the 12- O-tetradecanoylphorbol-13-acetate-induced differentiation of human colon cancer cells

Some lines of colon cancer cells are forced to undergo differentiation by 12- O-tetradecanoylphorbol-13-acetate (TPA). The increases in activities of both protein tyrosine phosphatase (PTP) and protein tyrosine kinase (PTK) have been reported to be associated with the TPA-induced differentiation of HL-60 leukemia cells. In the present study, a 2-fold increase in PTP activity was observed in SW620 human colon cancer cells after 30 min of TPA treatment; a maximal level (4- to 5-fold) was reached at 60 min and continued for more than 6 hr. In addition, two TPA-induced differentiated characteristics, morphological alteration and release of cellular surface proteoglycan, were effectively blocked by PTP inhibitors, such as sodium orthovanadate (50 μM), zinc chloride (100 μM), and iodoacetate (250 μM), but not by the protein serine/threonine phosphatase inhibitor okadaic acid (20 nM). On the other hand, although TPA induced a transient slight increase in PTK activity (1.4-fold) at 60 min, four PTK inhibitors (genistein, herbimycin A, tyrphostin-23 and quercetin) had different effects on the TPA-induced release of cell surface proteoglycan. Genistein (60 μM) potentiated this process, but in contrast, quercetin (45 μM) could partially inhibit the TPA effect. Taken together, these observations suggest that both PTP and PTK activities were increased in SW620 cells in response to TPA; however, the activation of PTP seems to be preferentially required for the TPA-induced differentiation of SW620 human colon cancer cells.

Interleukin-1 and Oncostatin M in Combination Promote the Release of Collagen Fragments from Bovine Nasal Cartilage in Culture

Interleukin-1 (IL-1) and Oncostatin M (OM) induce a rapid and reproducible release of proteoglycan and collagen fragments from bovine nasal cartilage in culture. Over 90% of the total collagen was released by day 14 compared to a variable release with IL-1 alone. This release was accompanied by the appearance of collagenolytic activity in the medium that cleaved collagen specifically at the one quarter/three quarter position. Tissue inhibitor of metalloproteinases (TIMP-1) activity was low or absent in media from resorbing tissue. The breakdown of cartilage collagen could be prevented by the addition of BB94, a specific matrix metalloproteinase (MMP) inhibitor. These results suggest that T-cell/macrophage products within inflammed joints can interact with pro-inflammatory cytokines and lead to the rapid destruction of connective tissue collagen.

Oral administration of a matrix metalloproteinase inhibitor, CGS 27023A, protects the cartilage proteoglycan matrix in a partial meniscectomy model of osteoarthritis in rabbits

Matrix metalloproteinases (MMP) are elevated in human osteoarthritic cartilage [1] and in cartilage from rabbits with experimental osteoarthritis (OA) following partial meniscectomy [2]. CGS 27023A is a M M P inhibitor that inhibits stromelysin, collagenase and gelatinase. CGS 27023A is an orally active inhibitor of stromelysin. CGS 27023A at 75 gmoles/kg p.o. inhibits release of proteoglycan into synovial fluid following intra-articular injection of stromelysin into rabbit knees. The purpose of these experiments was to determine whether CGS 27023A would inhibit cartilage proteoglycan loss in a partial meniscectomy model of osteoarthritis in rabbits [3]. Doxycycline was used as a reference drug [4].