RESUMO
Cartilage loss in osteoarthritis (OA) results from altered local production of growth factors and metalloproteases (MMPs). Furin, an enzyme involved in the protein maturation of MMPs, might regulate chondrocyte function. Here, we tested the effect of furin on chondrocyte catabolism and the development of OA. In primary chondrocytes, furin reduced the expression of MMP-13, which was reversed by treatment with the furin inhibitor α1-PDX. Furin also promoted the activation of Smad3 signaling, whereas activin receptor-like kinase 5 (ALK5) knockdown mitigated the effects of furin on MMP-13 expression. Mice underwent destabilization of the medial meniscus (DMM) to induce OA, then received furin (1 U/mice), α1-PDX (14 µg/mice) or vehicle. In mice with DMM, the OA score was lower with furin than vehicle treatment (6.42 ± 0.75 vs 9.16 ± 0.6, p < 0.01), and the number of MMP-13(+) chondrocytes was lower (4.96 ± 0.60% vs 20.96 ± 8.49%, p < 0.05). Moreover, furin prevented the increase in ALK1/ALK5 ratio in cartilage induced by OA. Conversely, α1-PDX had no effect on OA cartilage structure. These results support a protective role for furin in OA by maintaining ALK5 receptor levels and reducing MMP-13 expression. Therefore, furin might be a potential target mediating the development of OA.
Assuntos
Furina/farmacologia , Metaloproteinase 13 da Matriz/efeitos dos fármacos , Osteoartrite/prevenção & controle , Fator de Crescimento Transformador beta/farmacologia , Receptores de Ativinas Tipo I/análise , Receptores de Ativinas Tipo I/efeitos dos fármacos , Receptores de Activinas Tipo II , Animais , Condrócitos/efeitos dos fármacos , Condrócitos/metabolismo , Camundongos , Osteoartrite/tratamento farmacológico , Pró-Proteína Convertases/farmacologia , Receptor do Fator de Crescimento Transformador beta Tipo I/efeitos dos fármacosRESUMO
OBJECTIVE: To investigate the impact of systemic inhibition of interleukin 6 (IL-6) or signal transducer and activator of transcription (Stat3) in an experimental model of osteoarthritis (OA). METHODS: Expression of major catabolic and anabolic factors of cartilage was determined in IL-6-treated mouse chondrocytes and cartilage explants. The anti-IL-6-receptor neutralising antibody MR16-1 was used in the destabilisation of the medial meniscus (DMM) mouse model of OA. Stat3 blockade was investigated by the small molecule Stattic ex vivo and in the DMM model. RESULTS: In chondrocytes and cartilage explants, IL-6 treatment reduced proteoglycan content with increased production of matrix metalloproteinase (MMP-3 and MMP-13) and a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS-4 and ADAMTS-5). IL-6 induced Stat3 and extracellular signal-regulated kinase (ERK) 1/2 signalling but not p38, c-Jun N-terminal kinase or Akt. In the DMM model, Stat3 was activated in cartilage, but neither in the synovium nor in the subchondral bone. Systemic blockade of IL-6 by MR16-1 alleviated DMM-induced OA cartilage lesions, impaired the osteophyte formation and the extent of synovitis. In the same model, Stattic had similar beneficial effects on cartilage and osteophyte formation. Stattic, but not an ERK1/2 inhibitor, significantly counteracted the catabolic effects of IL-6 on cartilage explants and suppressed the IL-6-induced chondrocytes apoptosis. CONCLUSION: IL-6 induces chondrocyte catabolism mainly via Stat3 signalling, a pathway activated in cartilage from joint subjected to DMM. Systemic blockade of IL-6 or STAT-3 can alleviate DMM-induced OA in mice.
Assuntos
Cartilagem/metabolismo , Interleucina-6/antagonistas & inibidores , Interleucina-6/metabolismo , Osteoartrite/metabolismo , Fator de Transcrição STAT3/antagonistas & inibidores , Fator de Transcrição STAT3/metabolismo , Proteína ADAMTS4/metabolismo , Proteína ADAMTS5/metabolismo , Animais , Anticorpos/farmacologia , Apoptose/efeitos dos fármacos , Células Cultivadas , Condrócitos , Óxidos S-Cíclicos/farmacologia , Modelos Animais de Doenças , Interleucina-6/farmacologia , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Metaloproteinase 13 da Matriz/metabolismo , Metaloproteinase 3 da Matriz/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Osteoartrite/prevenção & controle , Osteófito/prevenção & controle , Proteoglicanas/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores de Interleucina-6/imunologia , Sinovite/prevenção & controle , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
Sclerostin, mainly produced by osteocytes, is now considered a major regulator of bone formation. Identified from patients with a low bone mass, sclerostin inhibits the Wnt pathway by binding to LRP5/6 and subsequently increases bone formation. Sclerostin may also play a role in the mediation of systemic and local factors such as calcitriol, PTH, glucocorticoids and tumor necrosis factor-alpha. Circulating sclerostin levels increase with age and with the decline of kidney function. However, they are surprisingly higher in patients with a high bone mineral density, suggesting that sclerostin may be a relevant marker of the pool of mature osteocytes. The anti-anabolic properties lead to the development of anti-sclerostin biotherapies that are under current evaluation. The results of these clinical trials will open new promising opportunities for the treatment of osteoporosis and bone fragility fractures.
Assuntos
Envelhecimento/genética , Proteínas Morfogenéticas Ósseas/genética , Marcadores Genéticos/genética , Osteogênese/genética , Osteoporose/genética , Proteínas Adaptadoras de Transdução de Sinal , Proteínas Morfogenéticas Ósseas/fisiologia , Osso e Ossos/metabolismo , Fraturas Espontâneas/genética , Fraturas Espontâneas/fisiopatologia , Marcadores Genéticos/fisiologia , Humanos , Osteoporose/tratamento farmacológico , Via de Sinalização Wnt/genética , Via de Sinalização Wnt/fisiologiaRESUMO
Low oxygen tension (hypoxia) regulates chondrocyte differentiation and metabolism. Hypoxia-inducible factor 1α (HIF1α) is a crucial hypoxic factor for chondrocyte growth and survival during development. The major metalloproteinase matrix metalloproteinase 13 (MMP13) is also associated with chondrocyte hypertrophy in adult articular cartilage, the lack of which protects from cartilage degradation and osteoarthritis (OA) in mice. MMP13 is up-regulated by the Wnt/ß-catenin signaling, a pathway involved in chondrocyte catabolism and OA. We studied the role of HIF1α in regulating Wnt signaling in cartilage and OA. We used mice with conditional knockout of Hif1α (∆Hif1α(chon)) with joint instability. Specific loss of HIF1α exacerbated MMP13 expression and cartilage destruction. Analysis of Wnt signaling in hypoxic chondrocytes showed that HIF1α lowered transcription factor 4 (TCF4)-ß-catenin transcriptional activity and inhibited MMP13 expression. Indeed, HIF1α interacting with ß-catenin displaced TCF4 from MMP13 regulatory sequences. Finally, ΔHif1α(chon) mice with OA that were injected intraarticularly with PKF118-310, an inhibitor of TCF4-ß-catenin interaction, showed less cartilage degradation and reduced MMP13 expression in cartilage. Therefore, HIF1α-ß-catenin interaction is a negative regulator of Wnt signaling and MMP13 transcription, thus reducing catabolism in OA. Our study contributes to the understanding of the role of HIF1α in OA and highlights the HIF1α-ß-catenin interaction, thus providing new insights into the impact of hypoxia in articular cartilage.
Assuntos
Cartilagem Articular/metabolismo , Cartilagem Articular/patologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Metaloproteinase 13 da Matriz/metabolismo , Osteoartrite/metabolismo , beta Catenina/metabolismo , Animais , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Osteoartrite/patologia , Ligação Proteica , Transdução de SinaisRESUMO
INTRODUCTION: Sclerostin is a Wnt inhibitor produced by osteocytes that regulates bone formation. Because bone tissue contributes to the development of osteoarthritis (OA), we investigated the role of sclerostin in bone and cartilage in a joint instability model in mice. METHODS: Ten-week-old SOST-knockout (SOST-KO) and wild-type (WT) mice underwent destabilization of the medial meniscus (DMM). We measured bone volume at the medial femoral condyle and osteophyte volume and determined the OA score and expression of matrix proteins. Primary murine chondrocytes were cultured with Wnt3a and sclerostin to assess the expression of matrix proteins, proteoglycan release and glycosaminoglycan accumulation. RESULTS: Sclerostin was expressed in calcified cartilage of WT mice with OA. In SOST-KO mice, cartilage was preserved despite high bone volume. However, SOST-KO mice with DMM had a high OA score, with increased expression of aggrecanases and type X collagen. Moreover, SOST-KO mice with OA showed disrupted anabolic-catabolic balance and cartilage damage. In primary chondrocytes, sclerostin addition abolished Wnt3a-increased expression of a disintegrin and metalloproteinase with thrombospondin motifs, matrix metalloproteinases and type X collagen by inhibiting the canonical Wnt pathway. Moreover, sclerostin inhibited Wnt-phosphorylated c-Jun N-terminal kinase (JNK) and rescued the expression of anabolic genes. Furthermore, sclerostin treatment inhibited both Wnt canonical and non-canonical JNK pathways in chondrocytes, thus preserving metabolism. CONCLUSION: Sclerostin may play an important role in maintaining cartilage integrity in OA.
Assuntos
Glicoproteínas/fisiologia , Osteoartrite do Joelho/metabolismo , Via de Sinalização Wnt/fisiologia , beta Catenina/fisiologia , Proteínas Adaptadoras de Transdução de Sinal , Animais , Western Blotting , Cartilagem Articular/metabolismo , Condrócitos/citologia , Condrócitos/efeitos dos fármacos , Colágeno Tipo X/metabolismo , Desintegrinas/metabolismo , Glicoproteínas/farmacologia , Peptídeos e Proteínas de Sinalização Intercelular , Instabilidade Articular/metabolismo , Masculino , Metaloproteases/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Osteoartrite do Joelho/patologia , Reação em Cadeia da Polimerase , Proteína Wnt3A/farmacologiaRESUMO
OBJECTIVE: Wnt signaling is a master regulator of joint homeostasis, but its role in osteoarthritis (OA) remains unclear. This study was undertaken to characterize the activation of Wnt/ß-catenin in knee joints of mice with OA and to assess how inhibiting this pathway in bone could affect cartilage. METHODS: OA was induced by partial meniscectomy in Topgal mice and in transgenic mice overexpressing Dkk-1 under the control of the 2.3-kb Col1a1 promoter (Col1a1-Dkk-1-Tg mice). Wnt/ß-catenin activation was assessed by X-Gal staining at baseline and at weeks 4, 6, and 9. Cartilage and bone damage was analyzed in Col1a1-Dkk-1-Tg mice with OA at week 6. Primary chondrocytes and cartilage explants were used to assess the effect of Dkk-1 on cartilage catabolism. RESULTS: In meniscectomized Topgal mice, Wnt was mainly activated in osteocytes from the subchondral bone at week 6 after OA induction, as well as in osteophytes and synovium at week 4. Chondrocytes from damaged zones expressed X-Gal from week 4. Dkk-1 expression was high in chondrocytes in control mouse knees (mean ± SEM 84.2 ± 3.1%) but decreased greatly in knees of meniscectomized mice from week 4 (mean ± SEM 14.4 ± 3.8%). The OA score was lower in meniscectomized Col1a1-Dkk-1-Tg mice at week 6 compared with wild-type mice (5.1 ± 0.6 versus 8.4 ± 0.6; P = 0.002). Subchondral bone fraction and osteophyte volume were decreased. However, cartilage explants from Col1a1-Dkk-1-Tg mice showed proteoglycan loss and increased NITEGE expression. Expression of vascular endothelial growth factor (VEGF) was reduced in osteoblasts from Col1a1-Dkk-1-Tg mice, thereby decreasing expression of messenger RNA for matrix metalloproteinases in chondrocytes. CONCLUSION: Wnt activation in OA affects the whole joint, particularly bone. Selective inhibition of this pathway in bone by Dkk-1 decreased OA severity through VEGF inhibition.