Modulation of IL‐1‐induced cartilage injury by NO synthase inhibitors: a comparative study with rat chondrocytes and cartilage entities

Abstract

1 Nitric oxide (NO) is produced in diseased joints and may be a key mediator of IL-1 eects on cartilage. Therefore, we compared the potency of new [aminoguanidine (AG), S-methylisothiourea (SMT), S-aminoethylisothiourea (AETU)] and classical [N o -monomethyl-L-arginine (L-NMMA), N onitro-L-arginine methyl ester (L-NAME)] NO synthase (NOS) inhibitors on the inhibitory eect of recombinant human interleukin-1b (rhIL-1b) on rat cartilage anabolism. Three dierent culture systems were used: (1) isolated chondrocytes encapsulated in alginate beads; (2) patellae and (3) femoral head caps. 2 Chondrocyte beads and cartilage entities were incubated in vitro for 48 h in the presence of rhIL-1b with a daily change of incubation medium to obtain optimal responses on proteoglycan synthesis and NO production. Proteoglycan synthesis was assessed by incorporation of radiolabelled sodium sulphate [Na 2 35 SO 4 ] and NO production by cumulated nitrite release during the period of study. 3 Chondrocytes and patellae, as well as femoral head caps, responded concentration-dependently to IL1b challenge (0 to 250 U ml 71 and 0 to 15 U ml 71 respectively) by a large increase in nitrite level and a marked suppression of proteoglycan synthesis. Above these concentrations of IL-1b (2500 U ml 71 and 30 U ml 71 respectively), proteoglycan synthesis plateaued whereas nitrite release still increased thus suggesting dierent concentration-response curves. 4 When studying the eect of NOS inhibitors (1 to 1000 mM) on NO production by cartilage cells stimulated with IL-1b (25 U ml 71 or 5 U ml 71 ), we observed that: (i) their ability to reduce nitrite level decreased from chondrocytes to cartilage samples, except for L-NMMA and AETU; (ii) they could be roughly classi®ed in the following rank order of potency: AETU4L-NMMA5SMT4AG5L-NAME and (iii) AETU was cytotoxic when used in the millimolar range. 5 When studying the eect of NOS inhibitors on proteoglycan synthesis by cartilage cells treated with IL-1b, we observed that: (i) they had more marked eects on proteoglycan synthesis in chondrocytes than in cartilage samples; (ii) they could be roughly classi®ed in the following rank order of potency: L-NAME5L-NMMA44AG4SMT44AETU and (iii) potentiation of the IL-1 eect by AETU was consistent with cytotoxicity in the millimolar range. 6 D-isomers of L-arginine analog inhibitors (1000 mM) were unable to correct nitrite levels or proteoglycan synthesis in IL-1b treated cells. L-arginine (5000 mM) tended to reverse the correcting eect of L-NMMA (1000 mM) on proteoglycan synthesis, thus suggesting a NO-related chondroprotective eect. However, data with L-NAME and SMT argued against a general inverse relationship between nitrite level and proteoglycan synthesis. 7 Dexamethasone (0.1 to 100 mM) (i) failed to inhibit NO production in femoral head caps and chondrocytes beads whilst reducing it in patellae (50%) and (ii) did not aect or worsened the inhibitory eect of IL-1b on proteoglycan synthesis. Such results suggested a corticosteroid-resistance of rat chondrocy...

Protective effect of Capparis spinosa on chondrocytes

“…It appeared to be the result of an imbalance between the destructive and reparative/synthetic processes of the articular cartilage due to the destructive effects of free radicals (ROS and NO), inflammatory cytokines (e.g. IL and TNF-alpha) and metalloproteinases (MMP) (Moulton, 1996;Cipoletta et al, 1998). Cytokines, such as interleukin-1h interfere in extracellular matrix turnover, accelerate the degradation of cartilage matrix, and induce chondrocytes apoptosis.…”

Identification and cloning of a human urea transporter HUT11, which is downregulated during adipogenesis of explant cultures of human bone

“…NO is an important mediator of osteoblast activity and a stimulator of bone formation [Macpherson et al, 1999]. NO is also known to play distinct roles in the biochemistry of chondrocytes, osteoblasts, and adipocytes [Cipolletta et al, 1998;Gaudiot et al, 1998;Macpherson et al, 1999]. The formation of arginine in the urea cycle leads metabolically to the synthesis of polyamines via the rapidly inducible ornithine decarboxylase [Miyanka et al, 1998;Promeneur et al, 1996;Xu et al, 1997], which, interestingly, is co-localized with HUT11 in the renal outer medulla.…”

Evidence for the Presence of Peroxisome Proliferator-activated Receptor (PPAR) α and γ and Retinoid Z Receptor in Cartilage

“…This concentration was chosen because we previously found that it corresponded to the middle range of the concentration-response curve for proteoglycan synthesis inhibition and NO production (30). The ligands were added to cells simultaneously or 1 h before the addition of rhIL-1␤ (31), as Me 2 SO solutions with the final concentration of Me 2 SO never exceeding 0.1%.…”