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Pathogenic role of basic calcium phosphate crystals in destructive arthropathies.

Ea HK, Chobaz V, Nguyen C, Nasi S, van Lent P, Daudon M, Dessombz A, Bazin D, McCarthy G, Jolles-Haeberli B, Ives A, Van Linthoudt D, So A, Lioté F, Busso N - PLoS ONE (2013)

Bottom Line: BCP crystal-induced synovitis was totally independent of IL-1α and IL-1β signalling and no alterations of inflammation were observed in mice deficient for components of the NLRP3-inflammasome, IL-1α or IL-1β.Similarly, treatment with anakinra did not prevent BCP crystal effects.The effects are independent of IL-1 and NLRP3 inflammasome.

View Article: PubMed Central - PubMed

Affiliation: INSERM, UMR-S 606, Hospital Lariboisière, Paris, France.

ABSTRACT

Background: basic calcium phosphate (BCP) crystals are commonly found in osteoarthritis (OA) and are associated with cartilage destruction. BCP crystals induce in vitro catabolic responses with the production of metalloproteases and inflammatory cytokines such as interleukin-1 (IL-1). In vivo, IL-1 production induced by BCP crystals is both dependant and independent of NLRP3 inflammasome. We aimed to clarify 1/ the role of BCP crystals in cartilage destruction and 2/ the role of IL-1 and NLRP3 inflammasome in cartilage degradation related to BCP crystals.

Methodology principal findings: synovial membranes isolated from OA knees were analysed by alizarin Red and FTIR. Pyrogen free BCP crystals were injected into right knees of WT, NLRP3 -/-, ASC -/-, IL-1α -/- and IL-1β-/- mice and PBS was injected into left knees. To assess the role of IL-1, WT mice were treated by intra-peritoneal injections of anakinra, the IL-1Ra recombinant protein, or PBS. Articular destruction was studied at d4, d17 and d30 assessing synovial inflammation, proteoglycan loss and chondrocyte apoptosis. BCP crystals were frequently found in OA synovial membranes including low grade OA. BCP crystals injected into murine knee joints provoked synovial inflammation characterized by synovial macrophage infiltration that persisted at day 30, cartilage degradation as evidenced by loss of proteoglycan staining by Safranin-O and concomitant expression of VDIPEN epitopes, and increased chondrocyte apoptosis. BCP crystal-induced synovitis was totally independent of IL-1α and IL-1β signalling and no alterations of inflammation were observed in mice deficient for components of the NLRP3-inflammasome, IL-1α or IL-1β. Similarly, treatment with anakinra did not prevent BCP crystal effects. In vitro, BCP crystals elicited enhanced transcription of matrix degrading and pro-inflammatory genes in macrophages.

Conclusions significance: intra-articular BCP crystals can elicit synovial inflammation and cartilage degradation suggesting that BCP crystals have a direct pathogenic role in OA. The effects are independent of IL-1 and NLRP3 inflammasome.

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OCP crystal-induced inflammation and cartilage degradation is NLRP3 inflammasome- and IL-1 independent.WT (n = 6), ASC-/- (n = 4), NLRP3-/- (n = 6), IL-1α-/- (n = 5) and IL-1β-/- (n = 6) mice were injected i.a. with OCP crystals (200 µg in 20 µl) or PBS. In a second set of experiment, anakinra, the recombinant form of IL-1Ra, or PBS were injected for 4 days (7 mice per group), the first injection being 30 min prior to OCP injection into the knee of WT (F). Ratio of isotope uptake into OCP injected knee versus PBS-injected ones was calculated at different time points (A). Synovial inflammation (B, E, F), cartilage PG loss (C, E, F) and VDIPEN immunohistochemistries (D, F) were assessed. Results are expressed as % of scores against WT (B,C,D) or in arbitrary units (E, F), and represent mean ± S.E.M. of at least n = 4 mice per group. For p values, *  =  p<0.05, **  =  p<0.01, ***  =  p<0.001.
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pone-0057352-g003: OCP crystal-induced inflammation and cartilage degradation is NLRP3 inflammasome- and IL-1 independent.WT (n = 6), ASC-/- (n = 4), NLRP3-/- (n = 6), IL-1α-/- (n = 5) and IL-1β-/- (n = 6) mice were injected i.a. with OCP crystals (200 µg in 20 µl) or PBS. In a second set of experiment, anakinra, the recombinant form of IL-1Ra, or PBS were injected for 4 days (7 mice per group), the first injection being 30 min prior to OCP injection into the knee of WT (F). Ratio of isotope uptake into OCP injected knee versus PBS-injected ones was calculated at different time points (A). Synovial inflammation (B, E, F), cartilage PG loss (C, E, F) and VDIPEN immunohistochemistries (D, F) were assessed. Results are expressed as % of scores against WT (B,C,D) or in arbitrary units (E, F), and represent mean ± S.E.M. of at least n = 4 mice per group. For p values, *  =  p<0.05, **  =  p<0.01, ***  =  p<0.001.

Mentions: Knee joints of NLRP3 and ASC deficient mice injected with BCP crystals had similar inflammation, PG loss and VDIPEN-staining scores compared to WT mice (Figure 3A, B, C, D). These results suggest that the NLRP3-inflammasome pathway of IL-1β production is not necessary in crystal-mediated cartilage destruction. To test if either IL-1α or-β were directly involved, we then injected crystals into knee joints of IL-1β or IL-1α deficient mice or WT mice. We observed no significant reduction of inflammation or cartilage damage in the deficient mice (Figure 3E). Finally, we investigated the effects of IL-1Ra, which blocks the binding of both IL-1α and -β to the IL-1 receptor (IL-1R). Mice were injected twice daily i.p. with recombinant anakinra at 200 µg per mouse for 4 days prior to sacrifice, at day 5. The treatment had no effect on crystal-induced synovial inflammation, PG depletion, or VDIPEN staining (Figure 3F).


Pathogenic role of basic calcium phosphate crystals in destructive arthropathies.

Ea HK, Chobaz V, Nguyen C, Nasi S, van Lent P, Daudon M, Dessombz A, Bazin D, McCarthy G, Jolles-Haeberli B, Ives A, Van Linthoudt D, So A, Lioté F, Busso N - PLoS ONE (2013)

OCP crystal-induced inflammation and cartilage degradation is NLRP3 inflammasome- and IL-1 independent.WT (n = 6), ASC-/- (n = 4), NLRP3-/- (n = 6), IL-1α-/- (n = 5) and IL-1β-/- (n = 6) mice were injected i.a. with OCP crystals (200 µg in 20 µl) or PBS. In a second set of experiment, anakinra, the recombinant form of IL-1Ra, or PBS were injected for 4 days (7 mice per group), the first injection being 30 min prior to OCP injection into the knee of WT (F). Ratio of isotope uptake into OCP injected knee versus PBS-injected ones was calculated at different time points (A). Synovial inflammation (B, E, F), cartilage PG loss (C, E, F) and VDIPEN immunohistochemistries (D, F) were assessed. Results are expressed as % of scores against WT (B,C,D) or in arbitrary units (E, F), and represent mean ± S.E.M. of at least n = 4 mice per group. For p values, *  =  p<0.05, **  =  p<0.01, ***  =  p<0.001.
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pone-0057352-g003: OCP crystal-induced inflammation and cartilage degradation is NLRP3 inflammasome- and IL-1 independent.WT (n = 6), ASC-/- (n = 4), NLRP3-/- (n = 6), IL-1α-/- (n = 5) and IL-1β-/- (n = 6) mice were injected i.a. with OCP crystals (200 µg in 20 µl) or PBS. In a second set of experiment, anakinra, the recombinant form of IL-1Ra, or PBS were injected for 4 days (7 mice per group), the first injection being 30 min prior to OCP injection into the knee of WT (F). Ratio of isotope uptake into OCP injected knee versus PBS-injected ones was calculated at different time points (A). Synovial inflammation (B, E, F), cartilage PG loss (C, E, F) and VDIPEN immunohistochemistries (D, F) were assessed. Results are expressed as % of scores against WT (B,C,D) or in arbitrary units (E, F), and represent mean ± S.E.M. of at least n = 4 mice per group. For p values, *  =  p<0.05, **  =  p<0.01, ***  =  p<0.001.
Mentions: Knee joints of NLRP3 and ASC deficient mice injected with BCP crystals had similar inflammation, PG loss and VDIPEN-staining scores compared to WT mice (Figure 3A, B, C, D). These results suggest that the NLRP3-inflammasome pathway of IL-1β production is not necessary in crystal-mediated cartilage destruction. To test if either IL-1α or-β were directly involved, we then injected crystals into knee joints of IL-1β or IL-1α deficient mice or WT mice. We observed no significant reduction of inflammation or cartilage damage in the deficient mice (Figure 3E). Finally, we investigated the effects of IL-1Ra, which blocks the binding of both IL-1α and -β to the IL-1 receptor (IL-1R). Mice were injected twice daily i.p. with recombinant anakinra at 200 µg per mouse for 4 days prior to sacrifice, at day 5. The treatment had no effect on crystal-induced synovial inflammation, PG depletion, or VDIPEN staining (Figure 3F).

Bottom Line: BCP crystal-induced synovitis was totally independent of IL-1α and IL-1β signalling and no alterations of inflammation were observed in mice deficient for components of the NLRP3-inflammasome, IL-1α or IL-1β.Similarly, treatment with anakinra did not prevent BCP crystal effects.The effects are independent of IL-1 and NLRP3 inflammasome.

View Article: PubMed Central - PubMed

Affiliation: INSERM, UMR-S 606, Hospital Lariboisière, Paris, France.

ABSTRACT

Background: basic calcium phosphate (BCP) crystals are commonly found in osteoarthritis (OA) and are associated with cartilage destruction. BCP crystals induce in vitro catabolic responses with the production of metalloproteases and inflammatory cytokines such as interleukin-1 (IL-1). In vivo, IL-1 production induced by BCP crystals is both dependant and independent of NLRP3 inflammasome. We aimed to clarify 1/ the role of BCP crystals in cartilage destruction and 2/ the role of IL-1 and NLRP3 inflammasome in cartilage degradation related to BCP crystals.

Methodology principal findings: synovial membranes isolated from OA knees were analysed by alizarin Red and FTIR. Pyrogen free BCP crystals were injected into right knees of WT, NLRP3 -/-, ASC -/-, IL-1α -/- and IL-1β-/- mice and PBS was injected into left knees. To assess the role of IL-1, WT mice were treated by intra-peritoneal injections of anakinra, the IL-1Ra recombinant protein, or PBS. Articular destruction was studied at d4, d17 and d30 assessing synovial inflammation, proteoglycan loss and chondrocyte apoptosis. BCP crystals were frequently found in OA synovial membranes including low grade OA. BCP crystals injected into murine knee joints provoked synovial inflammation characterized by synovial macrophage infiltration that persisted at day 30, cartilage degradation as evidenced by loss of proteoglycan staining by Safranin-O and concomitant expression of VDIPEN epitopes, and increased chondrocyte apoptosis. BCP crystal-induced synovitis was totally independent of IL-1α and IL-1β signalling and no alterations of inflammation were observed in mice deficient for components of the NLRP3-inflammasome, IL-1α or IL-1β. Similarly, treatment with anakinra did not prevent BCP crystal effects. In vitro, BCP crystals elicited enhanced transcription of matrix degrading and pro-inflammatory genes in macrophages.

Conclusions significance: intra-articular BCP crystals can elicit synovial inflammation and cartilage degradation suggesting that BCP crystals have a direct pathogenic role in OA. The effects are independent of IL-1 and NLRP3 inflammasome.

Show MeSH
Related in: MedlinePlus