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Pro-inflammatory cytokines such as interleukin-1 (IL-1may participate in the pathogenesis of cartilage damage in osteoarthritis (OA) through the production of catabolic enzymes and inflammatory mediators. Induction of heme oxygenase-1 (HO-1) has previously been shown to exert anti-inflammatory effects in different cel types. We

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These data have opened new mechanisms of action for CORM-2, raising the prospect that CO-releasing molecules are an interesting strategy for the development of new treatments in articular conditions.

IL (interleukin)-1β plays an important role in cartilage extracellular matrix degradation and bone resorption in OA (osteoarthritis) through the induction of degradative enzymes and pro-inflammatory mediators. In the present study, we have determined the consequences of HO-1 (haem oxygenase-1) induction on markers of inflammation and senescence in the functional unit cartilage-subchondral bone stimulated with IL-1β. Cartilage-subchondral bone specimens were obtained from the knees of osteoarthritic patients. Treatment with the HO-1 inducer CoPP (cobalt protoporphyrin IX) counteracted the stimulatory effects of IL-1β on IL-6, nitrite, PGE2 (prostaglandin E2), TGF (transforming growth factor) β2, TGFβ3 and osteocalcin. Immunohistochemical analyses indicated that CoPP treatment of explants down-regulated iNOS (inducible nitric oxide synthase), COX-2 (cyclooxygenase-2) and mPGES-1 (microsomal prostaglandin E synthase-1) induced by IL-1β. In contrast, the expression of HMGB1 (high-mobility group box 1) was not significantly modified. In addition, CoPP decreased the expression of iNOS and mPGES-1 in cells isolated from the explants and stimulated with IL-1β, which was counteracted by an siRNA (small interfering RNA) specific for human HO-1. In isolated primary chondrocytes, we determined senescence-associated β-galactosidase activity and the expression of senescence markers by real-time PCR. We have found that HO-1 induction could regulate senescence markers in the presence of IL-1β and significantly affected telomerase expression, as well as β-galactosidase activity and hTERT (human telomerase reverse transcriptase) and p21 expression in chondrocytes. The findings of the present study support the view that HO-1 induction results in the down-regulation of inflammatory and senescence responses in OA articular tissues.

HO-1 (haem oxygenase-1) catalyses the degradation of haem and possesses anti-inflammatory and cytoprotective properties. The role of inflammatory mediators in the pathogenesis of OA (osteoarthritis) is becoming increasingly appreciated. In the present study, we investigated the effects of HO-1 induction in OA and healthy HACs (human articular chondrocytes) in response to inflammatory cytokine IL-1 β (interleukin-1β) under hypoxic conditions. Hypoxia was investigated as it is a more physiological condition of the avascular cartilage. Hypoxic signalling is mediated by HIFs (hypoxia-inducible factors), of which there are two main isoforms, HIF-1α and HIF-2α. Normal and OA chondrocytes were stimulated with IL-1β. This cytokine suppresses HO-1 expression and exerts both catabolic and anti-anabolic effects, while increasing HIF-1α and suppressing HIF-2α protein levels in OA chondrocytes in hypoxia. Induction of HO-1 by CoPP (cobalt protoporphyrin IX) reversed these IL-1β actions. The hypoxia-induced anabolic pathway involving HIF-2α, SOX9 [SRY (sex determining region Y)-box 9] and COL2A1 (collagen type II α1) was suppressed by IL-1β, but importantly, levels were restored by HO-1 induction, which down-regulated TNFα (tumour necrosis factor α), MMP (matrix metalloproteinase) activity and MMP-13 protein levels. Depletion of HO-1 using siRNA (small interfering RNA) abolished the CoPP effects, further demonstrating that these were due to HO-1. The results of the present study reveal the different mechanisms by which HO-1 exerts protective effects on chondrocytes in physiological levels of hypoxia.

their isolation. The absence of cross-reaction with the IIA isoform was established by ELISA and WB. In addition, the Saos-2 cell line was chosen to test a possible labelling of other fibrillar procollagens, mainly type I, V and XI. In fact, this cell line is described to synthesize the (a1)I, (a2)I, (a1)V, (a2)V, (a1)XI and (a2)XI, but no (a3)XI chains. No signal was detected on WB of cellular extracts or conditioned media with anti-pNIIB52, whereas antibodies to the collagen I, V and XI triple-helical parts revealed indeed the presence of proforms of these collagens. Conclusions: Anti-pNIIB52 antibodies allow a very sensitive and specific detection of the procollagen IIB, the protein synthesized by mature chondrocytes. No cross-reactions with the IIA or with the major fibrillar procollagens synthesized by cells with an osteoblastic phenotype were observed. These antibodies offer a new tool in basic as well as in translational research concerning cartilage differentiation, homeostasis and repair.

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