Cultured Rabbit Articular Chondrocytes Research Articles

Chloride conductance determining membrane potential of rabbit articular chondrocytes.

Membrane conductance of cultured rabbit articular chondrocytes was characterized by means of the patch-clamp technique. The resting membrane potential of the articular chondrocytes was about -42 mV. The membrane potential shifted in accordance with the prediction by the Nernst equation for Cl- when intracellular and extracellular concentrations of Cl- were changed. On the other hand, change in extracellular concentration of K+ produced no shift in the membrane potential of chondrocytes. The Cl- channel blocker 4-acetamido-4'-isothiocyanatostilbene-2'2-disulfonic acid (SITS) depolarized the membrane potential. These findings suggest that the membrane potential of the chondrocytes is determined mainly by Cl- conductance. Using the cell-attached patch-clamp method, a large unitary conductance of 217 pS was observed in the articular chondrocytes. The unitary current was reversibly blocked by SITS. Therefore, the unitary current was carried by Cl-. The Cl- channel showed voltage-dependent activation and the channels exhibited long-lasting openings. Therefore, the membrane potential of rabbit cultured articular chondrocytes was mainly determined by the activities of the large-conductance and voltage-dependent Cl- channels.

Rhein, an active metabolite of diacerein, down-regulates the production of pro-matrix metalloproteinases-1, -3, -9 and -13 and up-regulates the production of tissue inhibitor of metalloproteinase-1 in cultured rabbit articular chondrocytes

Objective Diacerein has proved to be effective in the treatment of osteoarthritis (OA). However, the precise action mechanism of diacerein on OA is not yet fully understood. Therefore, we investigated the effects of rhein, an active metabolite of diacerein, on the production of promatrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinase-1 (TIMP-1) in rabbit articular chondrocytes.Design Confluent rabbit chondrocytes were treated for 24 or 48h with rhein, naproxen or dexamethasone in the presence of recombinant human IL-1α (rhIL-1α). ProMMP-9/progelatinase B in the culture medium was monitored by gelatin zymography, and proMMP-1/procollagenase 1, proMMP-3/prostromelysin 1, proMMP-13/procollagenase 3 and TIMP-1 were analysed by Western blot analysis. The steady-state levels of proMMP mRNAs were examined by Northern blot analysis. Total MMPs activity was also determined using FITC-labeled casein.Results Rhein suppressed the rhIL-1α-induced production of proMMPs-1, -3, -9 and -13 in a dose-dependent manner (0.1–30μM). The suppressed production of proMMPs-1 and -3 was accompanied by a decrease in the steady-state levels of their mRNAs. Interestingly, rhein increased the production of TIMP-1. These observations were further supported by the fact that rhein decreased the apparent total activity of MMPs in the culture medium.Conclusion We have demonstrated that rhein, an active metabolite of diacerein down-regulates the gene-expression and production of proMMPs and up-regulates the TIMP-1 production. The therapeutic effects of diacerein on OA may be due, at least in part, to the chondroprotective effect of rhein, its active metabolite.

Effects of mechanical vibration on DNA and proteoglycan syntheses in cultured articular chondrocytes

The objective of this study was to determine the effects of mechanical vibration loading on DNA and proteoglycan syntheses in cultured rabbit articular chondrocytes. Chondrocyte culture plates were placed in a vibratory apparatus and subjected to a mechanical vibratory load at various frequencies and periods during culture. Mechanical vibration was applied at a sinusoidal waveform of 1.4 G-acceleration with frequencies of 200, 300, 400, 800, and 1600 Hz. 3H-thymidine and H235SO4 incorporation were used to detect radiolabeled DNA and proteoglycan syntheses, respectively. A frequency of 300 Hz showed a time-dependent augmentation of DNA synthesis and gave a maximal increase on day 3 with periodic vibration (8 h per day), and at 72 h or longer with continuous vibration. It also promoted proteoglycan synthesis in long-term cul-ture (from 3 to 15 days) by periodic vibration. However, frequencies above 400 Hz suppressed biosynthesis. These results suggest that mechanical vibration at certain frequencies may modulate the biosynthetic response of articular chondrocytes.

Effects of mechanical vibration on DNA and proteoglycan syntheses in cultured articular chondrocytes

The objective of this study was to determine the effects of mechanical vibration loading on DNA and proteoglycan syntheses in cultured rabbit articular chondrocytes. Chondrocyte culture plates were placed in a vibratory apparatus and subjected to a mechanical vibratory load at various frequencies and periods during culture. Mechanical vibration was applied at a sinusoidal waveform of 1.4 G-acceleration with frequencies of 200, 300, 400, 800, and 1600 Hz. 3H-thymidine and H235SO4 incorporation were used to detect radiolabeled DNA and proteoglycan syntheses, respectively. A frequency of 300 Hz showed a time-dependent augmentation of DNA synthesis and gave a maximal increase on day 3 with periodic vibration (8 h per day), and at 72 h or longer with continuous vibration. It also promoted proteoglycan synthesis in long-term cul-ture (from 3 to 15 days) by periodic vibration. However, frequencies above 400 Hz suppressed biosynthesis. These results suggest that mechanical vibration at certain frequencies may modulate the biosynthetic response of articular chondrocytes.

Rhein, an Active Metabolite of Diacerein, Suppresses the Interleukin-1 α-Induced Proteoglycan Degradation in Cultured Rabbit Articular Chondrocytes

Diacerein has proved to be effective in the treatment of osteoarthritis. We investigated the effects of rhein, an active metabolite of diacerein, on the degradation of recombinant human interleukin-1a (rhIL-1α)-induced proteoglycan and matrix metalloproteinases (MMPs) release from rabbit articular chondrocytes. Chondrocytes were treated for 24 h with rhein in the presence of rhIL-1α. Rhein suppressed the rhIL-1α-induced proteoglycan degradation. In addition, rhein decreased the rhlL-1α-induced proMMP-3 production and reduced the MMPs activity. These results suggested that diacerein might have a protective effect on proteoglycan degradation via the suppressive effect of rhein on the production of proMMPs.

Arterial remodeling. Mechanisms and clinical implications.

The presentation of coronary atherosclerosis can be gradual, because of progressive flow-limiting stenosis and exertional angina, or dramatic, with plaque rupture and thrombosis causing unstable angina, myocardial infarction, or sudden death. The importance of arterial remodeling, or persistent change in vessel size, has recently become apparent in both situations. Arterial remodeling, not plaque size, has been identified as the primary determinant of lumen size in the presence of stable lesions. Similarly, luminal stenosis in transplant vasculopathy and with restenosis after angioplasty occur mainly because of inward remodeling rather than plaque growth. However, recent evidence also suggests that adequate outward remodeling may be associated with an increased risk of plaque rupture, the underlying cause of acute coronary syndromes and sudden cardiac death. The term “arterial remodeling” has previously been used to describe any change in vessel wall structure. More recently, however, it has been used specifically to refer to a change in vessel size (or cross-sectional area within the external elastic lamina), and it is on this entity that this review is focused. Inward remodeling denotes a reduction in vessel size. Outward remodeling denotes an increase in vessel size. Various other terms are used in the literature (the Table⇓). When outward remodeling is present but insufficient to prevent luminal stenosis, it is referred to as inadequate outward remodeling. View this table: Table 1. Terminology of Arterial Remodeling It has been known for more than a century1 that blood vessels enlarge to accommodate increasing flow to the organ downstream (eg, during natural growth or in left ventricular hypertrophy). Widespread interest in this phenomenon was stimulated by observations that radial enlargement of vessels (outward remodeling) can compensate for progressive growth of atherosclerotic plaques, thus postponing the development of flow-limiting stenosis.2 3 These pathological findings were subsequently supported by in vivo intravascular ultrasound (IVUS) studies that …

Interleukin-1beta increases the functional expression of connexin 43 in articular chondrocytes: evidence for a Ca2+-dependent mechanism.

Cell-to-cell interactions and gap junctions-dependent communication are crucially involved in chondrogenic differentiation, whereas in adult articular cartilage direct intercellular communication occurs mainly among chondrocytes facing the outer cartilage layer. Chondrocytes extracted from adult articular cartilage and grown in primary culture express connexin 43 (Cx43) and form functional gap junctions capable of sustaining the propagation of intercellular Ca2+ waves. Degradation of articular cartilage is a characteristic feature of arthritic diseases and is associated to increased levels of Interleukin-1 (IL-1) in the synovial fluid. We have examined the effects of IL-1 on gap junctional communication in cultured rabbit articular chondrocytes. Incubation with IL-1 potentiated the transmission of intercellular Ca2+ waves and the intercellular transfer of Lucifer yellow. The stimulatory effect was accompanied by a dose-dependent increase in the expression of Cx43 and by an enhanced Cx43 immunostaining at sites of cell-to-cell contact. IL-1 stimulation induced a dose-dependent increase of cytosolic Ca2+ and activates protein tyrosine phosphorylation. IL-1-dependent up-regulation of Cx43 could be prevented by intracellular Ca2+ chelation but not by inhibitors of protein tyrosine kinases, suggesting a crucial role of cytosolic Ca2+ in regulating the expression of Cx43. IL-1 is one of the most potent cytokines that promotes cartilage catabolism; its modulation of intercellular communication represents a novel mechanism by which proinflammatory mediators regulate the activity of cartilage cells.

Oxidative injury induced by synthetic hum1c acid polymer and monomer in cultured rabbit articular chondrocytes

Humic substance has been proposed as one of the causative factors of Kashin-Beck disease (KBD), an endemic osteoarthritic disorder with necrosis of chondrocytes widely prevalent in some regions of China In order to exclude the complications of natural humic substance, here we prepared phenolic polymers of synthetic humic acid (SHA) by oxidation of phenolic monomer, the protocatechuic acid (PCA). The biological effects of SHA and PCA on primary culture of rabbit articular chondrocytes were investigated. We found that not only SHA but also PCA caused chondrocyte injury, as evidenced by the loss of cell viability measured with methylthiazol tetrazolium (MTT) assay and the increased release of intracellular lactate dehydrogenase (LDH) Both SHA and PCA could result in lipid peroxidation and glutathione (GSH) depletion in chondrocytes, indicating that oxidative stress may be involved in chondrocyte injury. Furthermore, a marked increase in intracellular calcium level ([Ca 2+]i) occurred after chondrocytes treated with SHA or PCA. These results suggest that chondrocyte injury elicited by SHA or PCA may be mediated through the occurrence of oxidative stress and the disruption of intracellular Ca 2+ homeostasis. Data also suggest that the monomeric phenolic acid may be considered one of the causative factors of KBD in addition to humic substance.

Esculetin suppresses proteoglycan metabolism by inhibiting the production of matrix metalloproteinases in rabbit chondrocytes

The possible mechanism of the chondroprotective effect of 6,7-dihydroxycoumarin (esculetin) was investigated using primary cultures of rabbit articular chondrocytes. Esculetin (EST) significantly suppressed the proteoglycan depletion and the release of pulse-labeled [ 35 S] proteoglycan from the matrix layer of rabbit chondrocytes treated with recombinant human interleukin-1α. The matrix metalloproteinase inhibitor, 1,10-phenanthroline, also blocked the proteoglycan depletion and [ 35 S] proteoglycan release. From these results, it is likely that recombinant human interleukin-1α-induced proteoglycan depletion is mediated by matrix metalloproteinases. Although esculetin did not directly inhibit collagenolytic activity in the culture media, it significantly suppressed the production of pro-matrix metalloproteinase-1/interstitial procollagenase and pro-matrix metalloproteinase-3/prostromelysin 1, accompanied by a decrease in the steady-state levels of their mRNAs. These results suggest that esculetin is a therapeutically effective candidate for inhibition of cartilage destruction in osteoarthritis and rheumatoid arthritis.

Hyaluronic acid suppresses the reduction of alpha2(VI) collagen gene expression caused by interleukin-1beta in cultured rabbit articular chondrocytes.

In order to investigate how alpha2(VI) collagen gene is regulated by inflammatory cytokines in cultured rabbit articular chondrocytes, we examined the effect of interleukin-1beta (IL-1beta) on this collagen mRNA expression. Polylayer cultures of chondrocytes were exposed to IL-1beta (0.1, 1, 10 ng/ml). Quantitative detection of specific mRNA for this collagen was carried out by reverse transcription-polymerase chain reaction (RT-PCR). Furthermore, to investigate the effect of hyaluronic acid (HA) on alpha2(VI) collagen mRNA expression by IL-1beta chondrocytes were exposed to IL-1beta (10 ng/ml) in the presence of HA (0.01, 0.1, 1 mg/ml) with molecular weight of 900 kDa. Chondrocytes were also exposed to IL-1beta (10 ng/ml) in the presence of HA (1 mg/ml) with molecular weights of 200, 900 and 2000 kDa. Alpha2(VI) collagen mRNA expression was decreased significantly in chondrocytes cultured with 1 and 10 ng/ml of IL-1beta. However, the addition of both IL-1beta and HA (0.1, 1 mg/ml) or both IL-1beta and HA (1 mg/ml) with all the molecular weight significantly suppressed these reduced mRNA levels. No tendency for this suppression to depend on the molecular weight was observed. These results suggest that suppression of transcriptional activity for type VI collagen will be associated with the reduction of cartilage matrix tissue and that HA will be associated with the suppression of the effect of IL-1beta.

Effects of the existence of nitric oxide and the degradation of extracellular matrix on CD44 mRNA expression in cultured chondrocytes

We previously reported that interleukin-1β (IL-1β) promoted CD44 mRNA expression in cultured rabbit articular chondrocytes, together with the reduction of extracellular matrix. It was assumed from this result that the existence of nitric oxide (NO) induced by IL-1β and degradation of the extracellular matrix affects CD44 mRNA expression in the chondrocytes. To confirm this assumption, we first carried out production of NO by IL-1β through quantitative detection of inducible nitric oxide synthase (iNOS) mRNA expression in the chondrocytes and measurement of nitrite plus nitrate in the culture medium. IL-1β evidently promoted NO production in the chondrocytes. A tendency for increased release of chondroitin sulfates (CS), as extracellular matrix, to the culture medium was observed by addition of IL-1β, but release of CS to the culture medium did not increase as the result of addition of S-nitroso-N-acetyl penicillamine (SNAP) as NO donor. We then investigated the effects of SNAP, chondroitinase, and hyaluronidase on CD44 mRNA expression in the cultured chondrocytes. CD44 mRNA expression was increased significantly in chondrocytes cultured with SNAP and chondroitinase but not in chondrocytes cultured with hyaluronidase. These results suggest that the existence of NO and the degradation of CS may be associated with upregulation of CD44 mRNA expression in chondrocytes..

OXIDATIVE STRESS INDUCED BY HUMIC ACID SOLVENT EXTRACTION FRACTION IN CULTURED RABBIT ARTICULAR CHONDROCYTES

Kashin-Beck disease (KBD) , an endemic, chronic osteoarthritic disorder with necrosis of chondrocytes, commonly occurs in China. The humic substance present in the drinking water of endemic areas has been proposed as one of the causative factors. In this study an in vitro cell culture system was used to investigate the damaging effects of humic acid (HA), a constituent of humic substance, on cultured rabbit articular chondrocytes. The commercial Aldrich humic acid (AHA) was fractionated with a series of organic solvents including n -hexane, benzene, ethyl acetate, and methanol. Among the several fractions of AHA, the ethyl acetate fraction (AHA-[EA]) displayed the most potent inhibitory effect on the survival of chondrocytes in clonogenic assays. Cellular injury induced by AHA-[EA] was evaluated by measuring cell viability with methylthiazol tetrazolium (MTT) and by determining the release of intracellular lactate dehydrogenase (LDH). Incubation of chondrocytes with AHA-[EA] (100-500 mug/ml) for 12 h produced a concentration-dependent decrease in cell viability and increase in LDH release. In addition, AHA-[EA] triggered lipid peroxidation manifested by elevated malondialdehyde (MDA) formation. In chemiluminescence assay, AHA-[EA] at the concentrations of 150-600 mug/ ml caused 6- to 15-fold increases of luminol-amplified chemiluminescence responses, which are considered to reflect the production of hydrogen peroxide (H2O2). Moreover, pretreating the cells with 500-750 U/ml of catalase significantly prevented the loss of cell viability, while superoxide dismutase (SOD) enhanced the adverse effect of 300 mug/ml AHA-[EA]. Data suggest that the injury to chondrocytes induced by AHA[EA] may be first through O2 production, which is then converted into H2O2, thus initiating lipid peroxidation and leading to chondronecrosis observed in KBD.

Nitric oxide mediates interleukin-1-induced gene expression of matrix metalloproteinases and basic fibroblast growth factor in cultured rabbit articular chondrocytes.

We recently reported that nitric oxide (NO), which is produced by chondrocytes treated with interleukin-1beta (IL-1), releases basic fibroblast growth factor (bFGF) stored in the matrix of articular chondrocytes. To clarify the mechanism of the IL-1-induced bFGF release, we investigated the production and gene expression of bFGF, matrix metalloproteinases (MMPs), syndecan 3, and inducible NO synthase (iNOS) by IL-1-treated rabbit articular chondrocytes. IL-1 stimulated not only the release of bFGF but also the production of it. Gelatin and casein zymography revealed that IL-1 stimulated the production of not only MMP-9 but also MMP-3. The increase in the production of these MMPs preceded the IL-1-stimulated bFGF release. An MMP inhibitor partially suppressed the release of bFGF, indicating that matrix degradation is at least partially involved in the IL-1-stimulated bFGF release even if increased production of bFGF is related to the release. IL-1 sequentially stimulated mRNA expression of iNOS, membrane type 1-MMP, MMP-9 and -3, and bFGF, in that order. NG-Monomethyl-L-arginine, an inhibitor of NO production, inhibited gene expression of MMP-9 and bFGF. These findings suggest that elevation of the NO level via iNOS mRNA expression stimulated by IL-1 mediates gene expression and production of MMPs and bFGF, resulting in the release of bFGF, and also reveal molecular mechanisms implicating the degradation of articular cartilage followed by angiogenesis in the synovium in arthritic joints.

Upregulation of CD44 mRNA expression by interleukin-1β in cultured rabbit articular chondrocytes

We investigated the effects of interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) on CD44 mRNA expression in cultured rabbit articular chondrocytes. High-density-suspension-cultured chondrocytes were exposed to IL-1β (0.1, 1, or 10 ng/ml) or TNF-α (0.1, 1, or 10 ng/ml). Quantitative detection of specific mRNA for CD44 was carried out by reverse transcription, polymerase chain reaction (RT-PCR). Furthermore, to determine the degradation of cartilage matrix by IL-1β and TNF-α, the concentrations of chondroitin 4-sulfate (C4S) and chondroitin 6-sulfate (C6S) released to the culture medium were measured with high-performance liquid chromatography (HPLC). CD44 mRNA expression was increased significantly in chondrocytes cultured with IL-1β, but not in the chondrocytes cultured with TNF-α. The release of chondroitin sulfates (C4S+C6S) to the culture medium was also accelerated by IL-1β but was not affected by TNF-α. These results suggest that IL-1β can promote CD44 mRNA expression, together with the degradation of the cartilage matrix, and may assist in binding hyaluronic acid (HA) and CD44 in the chondrocytes.

Voltage-gated ionic channels in cultured rabbit articular chondrocytes

The membrane properties of cultured cells of rabbit articular chondrocytes were studied using the whole-cell patch clamp technique. The average cell capacitance was 37.9 ± 9.0 pF ( n = 13), and the cell resting potential was −41.0 ± 7.0 mV ( n = 11). We were unable to induce an action potential by applying a depolarizing current. Upon step depolarization, under voltage clamp conditions, one kind of inward and two kinds of outward currents were elicited. The inward current was initially observed at around −30 mV, peaked at 0 mV, and reversed at around +90 mV. Tetrodotoxin (TTX; 1 μM) was shown to completely block this inward current. At steady state, the inward current was half-inactivated at −51 mV, with a slope factor of 6.3 mV. Two outward currents were determined from measurements of activation threshold, reversal potential, and pharmacological responses. One was observed at around −30 mV, and its amplitude increased with membrane depolarization. Extracellularly applied 4-aminopyridine (4-AP) (1 mM) and tetraethyl ammonium chloride (TEA) (5 mM) blocked this current. The other outward current was observed at around +10 mV, and its direction reversed at a potential close to that predicted by the Nernst equation for a Cl − selective channel. This current fluctuated markedly, and the fluctuation did not decline throughout the 100 ms of the step pulse. Extracellularly applied 4-acetamido-4′-isothiocyanostilbenezene-2,2-disulfonic acid (SITS) (0.25 mM) blocked this current, but the same dose of 4,4′-diisothiocyanatostilbene-2,2′-disulfonic acid (DIDS) had little effect. These results suggest the presence of TTX-sensitive Na +, 4-AP- and TEA-sensitive K +, and SITS-sensitive Cl − channels in rabbit articular chondrocyte membrane. The functional significance of these channels is discussed.

Lack of correlation between hydrogen peroxide production and nitric oxide production by cultured rabbit articular chondrocytes treated with fluoroquinolone antimicrobial agents

The arthrotoxicity of fluoroquinolone antibacterial agents so far remains unexplained. Recent experimental data have indicated an early stimulation of the oxidative metabolism within articular chondrocytes. An in vitro model was designed to analyse the production of oxygen-derived reactive species and glutathione by immature rabbit articular chondrocytes, and the influence of different fluoroquinolones on this model was examined. Primary cultures of chondrocytes were exposed to pefloxacin, ofloxacin or ciprofloxacin at 10 μg/ml, for 24 or 48 hr. Flow cytometric analysis used the vital tracer 2′,7′-dichlorofluorescein diacetate (DCFH-DA) and evaluated the production of H 2O 2 and NO by chondrocytes. Separately, NO production and intracellular glutathione levels were evaluated, with the Greiss colorimetric technique and the Tietze method, respectively. With each fluoroquinolone tested, intracellular levels of the fluorescent compound dichlorofluorescein (oxidized form of DCFH-DA) were significantly higher in treated chondrocytes than in control cells. No significant modification of NO or of glutathione cellular levels was noted. Fluoroquinolones stimulate H 2O 2 production in immature articular chondrocytes, but have no apparent effect on either NO or glutathione production, at least in the early stages of the chondrotoxicity.

Nitric oxide mediates interleukin-1-induced matrix degradation and basic fibroblast growth factor release in cultured rabbit articular chondrocytes: a possible mechanism of pathological neovascularization in arthritis.

Prolonged incubation with interleukin-1 beta (IL-1) induced the release of large amounts of NO and subsequently inhibited DNA synthesis and the biosynthesis and accumulation of proteoglycans in cultured rabbit articular chondrocytes (RAC). IL-1 also inhibited DNA synthesis in bovine aortic endothelial cells (BAE). On the other hand, DNA synthesis in BAE cocultured with RAC was not inhibited by prolonged incubation with IL-1. Moreover, conditioned media from RAC incubated for a long period with IL-1 stimulated DNA synthesis in BAE alone. This growth stimulatory activity was mainly due to the release of basic fibroblast growth factor, a heparin-binding growth factor, into RAC culture. Gelatin zymography of the RAC culture medium revealed that IL-1 increased the production of matrix metalloproteinase-2 (MMP-2) and MMP-9. NG-Monomethyl-L-arginine, an inhibitor of NO synthesis, inhibited all of these actions of IL-1. These results indicate that NO from RAC treated with IL-1 stimulates MMPs, which, in turn, degrade the extracellular matrix produced by RAC, resulting in the release of large amounts of basic fibroblast growth factor stored in the matrix, which then stimulates adjacent BAE proliferation. Thus, NO produced from RAC treated with IL-1 may modulate angiogenesis in the synovium of arthritic patients.

Nitric oxide mediates interleukin-1-induced matrix degradation and basic fibroblast growth factor release in cultured rabbit articular chondrocytes: a possible mechanism of pathological neovascularization in arthritis

Nitric oxide mediates interleukin-1-induced matrix degradation and basic fibroblast growth factor release in cultured rabbit articular chondrocytes: a possible mechanism of pathological neovascularization in arthritis

Type II transglutaminase expression in rabbit articular chondrocytes in culture: relation with cell differentiation, cell growth, cell adhesion and cell apoptosis.

Depending on the cell type studied, the involvement of type II transglutaminase (TGase) has been proposed in almost any event of the cell life such as differentiation, apoptosis, growth, aging, cell morphology and adhesion, metastatic capacity or extracellular matrix stabilization. In order to define the field(s) where this enzyme may be implicated in chondrocytes, type II TGase expression was studied in chondrocytes at different passages which differentiated state was modulated by retinoic acid, dihydrocytochalasin B or staurosporin. Results showed that (i) type II TGase expression is not incompatible with type II collagen expression, a main marker of chondrocyte differentiation (ii) type II TGase expression is higher when cells are in the exponential phase of growth than when growth arrested (iii) a high type II TGase expression does not imply that cells are apoptotic although cell apoptosis correlates with increased type II TGase expression (iv) non-adherent cells do not express type II TGase whereas adherent cells do whatever their differentiation state as assessed by type II collagen synthesis. These results suggest that, in articular chondrocytes, type II TGase is specifically implicated in the cell adhesion capacity.

Synergistic Effect of Interleukin-1β and Tumor Necrosis Factor α on PGE2Production by Articular Chondrocytes Does Not Involve PLA2Stimulation

This study investigates the ways in which two proinflammatory cytokines, tumor necrosis factor α (TNF) and interleukin-1β (IL1), cause increased production of prostaglandin E2(PGE2) in rabbit articular chondrocytes (RAC). Rabbit articular chondrocytes in primary culture were incubated with IL1, TNF, or both. Arachidonic acid (AA) release, PGE2production, and the activities of cytosolic phospholipase A2(cPLA2), secreted phospholipase A2(sPLA2), and cyclooxygenase (COX) were measured. The mRNA levels of cPLA2, sPLA2, and COX-2 were also measured by Northern blotting, using specific complementary DNA probes. Incubation of IL1-stimulated RAC with TNF further increased PGE2production. This synergy did not involve PLA2stimulation, as there were no increases in AA release, cPLA2and sPLA2activities, or mRNA. In contrast, TNF increased the effect of IL1 on COX-2 activity and mRNA level. These results show that TNF and IL1 act in synergy in PGE2production in articular chondrocytes. As sPLA2and cPLA2do not seem to be involved, COX-2 appears to be the best target for a specific anti-inflammatory strategy against cartilage degradation.