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TNF/TNFR signal transduction pathway-mediated anti-apoptosis and anti-inflammatory effects of sodium ferulate on IL-1β-induced rat osteoarthritis chondrocytes in vitro.

Qin J, Shang L, Ping AS, Li J, Li XJ, Yu H, Magdalou J, Chen LB, Wang H - Arthritis Res. Ther. (2012)

Bottom Line: SF also inhibited activities of caspase-8 and caspase-3 compared with the OA model control (P < 0.01).The results of EMSA showed that SF inhibited the activity of NF-κB.SF has anti-apoptosis and anti-inflammatory effects on an OA model induced by IL-1β in vitro, which were due to inhibitory actions on the caspase-dependent apoptosis pathway and the IKK/NF-κB signal transduction pathway of the TNF/TNFR pathway.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Introduction: Sodium ferulate (SF) is a natural component of traditional Chinese herbs. Our previous study shows that SF has a protective effect on osteoarthritis (OA). The objective of this study was to investigate the effect of SF on the TNF/TNF receptor (TNFR) signal transduction pathway of rat OA chondrocytes.

Methods: Primary rat articular chondrocytes were co-treated with IL-1β and SF. Chondrocyte apoptosis was assessed by fluorescein isothiocyanate-annexin V/propidium iodide assay. The PCR array was used to screen the expression of 84 key genes involved in apoptosis. The release of TNFα and prostaglandin E2 were analyzed by ELISA. Expressions of proteins were assessed by western blotting. The activity of NF-κB was determined by electrophoretic mobility shift assay (EMSA). Gene expression of inducible nitric oxide synthase (iNOS) was evaluated by real-time quantitative PCR. The nitric oxide content was measured with the Griess method.

Results: After treatment with SF, the apoptosis rate of chondrocytes significantly attenuated (P < 0.01). Results of the apoptosis PCR array suggested that mRNA expression of some core proteins in the TNF/TNFR pathway showed valuable regulation. The protein expressions of TNFα, TNFR-1, TNF receptor-associated death domain, caspase-8 and caspase-3 were prevented by SF in a concentration-dependent manner. SF also inhibited activities of caspase-8 and caspase-3 compared with the OA model control (P < 0.01). TNF receptor-associated factor-2 expression, phosphorylations of inhibitor of NF-κB kinase (IKK) subunits alpha and beta, and NF-κB inhibitor, alpha (IκBα) were all concentration-dependently suppressed by SF treatment. The results of EMSA showed that SF inhibited the activity of NF-κB. In addition, the expressions of cycloxygenase-2 and iNOS and the contents of prostaglandin E2 and NO were attenuated with the treatment of SF (P < 0.01).

Conclusion: SF has anti-apoptosis and anti-inflammatory effects on an OA model induced by IL-1β in vitro, which were due to inhibitory actions on the caspase-dependent apoptosis pathway and the IKK/NF-κB signal transduction pathway of the TNF/TNFR pathway.

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Effect of sodium ferulate on the apoptosis rate of IL-1β-induced rat osteoarthritis chondrocytes. Chondrocytes were incubated with sodium ferulate (SF) alone for 24 hours, and then co-treated with IL-1β and SF for 48 hours. After staining with annexin-V-fluorescein isothiocyanate and propidium iodide, chondrocytes were measured on flow cytometry and analyzed with Multi-cycle software (Phoenix Flow Systems, San Diego, CA, USA). Results expressed as percentage of apoptotic cells. Values represent mean ± standard error of the mean of four different simples. **P < 0.01 versus normal control. ##P < 0.01 versus osteoarthritis model control.
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Figure 2: Effect of sodium ferulate on the apoptosis rate of IL-1β-induced rat osteoarthritis chondrocytes. Chondrocytes were incubated with sodium ferulate (SF) alone for 24 hours, and then co-treated with IL-1β and SF for 48 hours. After staining with annexin-V-fluorescein isothiocyanate and propidium iodide, chondrocytes were measured on flow cytometry and analyzed with Multi-cycle software (Phoenix Flow Systems, San Diego, CA, USA). Results expressed as percentage of apoptotic cells. Values represent mean ± standard error of the mean of four different simples. **P < 0.01 versus normal control. ##P < 0.01 versus osteoarthritis model control.

Mentions: Chondrocyte apoptosis was identified by FITC-annexin V/PI double-labeled assay. IL-1β significantly increased the percentage of apoptotic chondrocytes in model control compared with the normal control (P < 0.01; n = 4). With treatments of 250, 500 and 1,000 μmol/l SF, the apoptotic percentage of OA chondrocytes was attenuated in a concentration-dependent manner (all P < 0.01; n = 4). However, 125 μmol/l SF failed to prevent OA chondrocyte apoptosis (Figure 2).


TNF/TNFR signal transduction pathway-mediated anti-apoptosis and anti-inflammatory effects of sodium ferulate on IL-1β-induced rat osteoarthritis chondrocytes in vitro.

Qin J, Shang L, Ping AS, Li J, Li XJ, Yu H, Magdalou J, Chen LB, Wang H - Arthritis Res. Ther. (2012)

Effect of sodium ferulate on the apoptosis rate of IL-1β-induced rat osteoarthritis chondrocytes. Chondrocytes were incubated with sodium ferulate (SF) alone for 24 hours, and then co-treated with IL-1β and SF for 48 hours. After staining with annexin-V-fluorescein isothiocyanate and propidium iodide, chondrocytes were measured on flow cytometry and analyzed with Multi-cycle software (Phoenix Flow Systems, San Diego, CA, USA). Results expressed as percentage of apoptotic cells. Values represent mean ± standard error of the mean of four different simples. **P < 0.01 versus normal control. ##P < 0.01 versus osteoarthritis model control.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3674623&req=5

Figure 2: Effect of sodium ferulate on the apoptosis rate of IL-1β-induced rat osteoarthritis chondrocytes. Chondrocytes were incubated with sodium ferulate (SF) alone for 24 hours, and then co-treated with IL-1β and SF for 48 hours. After staining with annexin-V-fluorescein isothiocyanate and propidium iodide, chondrocytes were measured on flow cytometry and analyzed with Multi-cycle software (Phoenix Flow Systems, San Diego, CA, USA). Results expressed as percentage of apoptotic cells. Values represent mean ± standard error of the mean of four different simples. **P < 0.01 versus normal control. ##P < 0.01 versus osteoarthritis model control.
Mentions: Chondrocyte apoptosis was identified by FITC-annexin V/PI double-labeled assay. IL-1β significantly increased the percentage of apoptotic chondrocytes in model control compared with the normal control (P < 0.01; n = 4). With treatments of 250, 500 and 1,000 μmol/l SF, the apoptotic percentage of OA chondrocytes was attenuated in a concentration-dependent manner (all P < 0.01; n = 4). However, 125 μmol/l SF failed to prevent OA chondrocyte apoptosis (Figure 2).

Bottom Line: SF also inhibited activities of caspase-8 and caspase-3 compared with the OA model control (P < 0.01).The results of EMSA showed that SF inhibited the activity of NF-κB.SF has anti-apoptosis and anti-inflammatory effects on an OA model induced by IL-1β in vitro, which were due to inhibitory actions on the caspase-dependent apoptosis pathway and the IKK/NF-κB signal transduction pathway of the TNF/TNFR pathway.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Introduction: Sodium ferulate (SF) is a natural component of traditional Chinese herbs. Our previous study shows that SF has a protective effect on osteoarthritis (OA). The objective of this study was to investigate the effect of SF on the TNF/TNF receptor (TNFR) signal transduction pathway of rat OA chondrocytes.

Methods: Primary rat articular chondrocytes were co-treated with IL-1β and SF. Chondrocyte apoptosis was assessed by fluorescein isothiocyanate-annexin V/propidium iodide assay. The PCR array was used to screen the expression of 84 key genes involved in apoptosis. The release of TNFα and prostaglandin E2 were analyzed by ELISA. Expressions of proteins were assessed by western blotting. The activity of NF-κB was determined by electrophoretic mobility shift assay (EMSA). Gene expression of inducible nitric oxide synthase (iNOS) was evaluated by real-time quantitative PCR. The nitric oxide content was measured with the Griess method.

Results: After treatment with SF, the apoptosis rate of chondrocytes significantly attenuated (P < 0.01). Results of the apoptosis PCR array suggested that mRNA expression of some core proteins in the TNF/TNFR pathway showed valuable regulation. The protein expressions of TNFα, TNFR-1, TNF receptor-associated death domain, caspase-8 and caspase-3 were prevented by SF in a concentration-dependent manner. SF also inhibited activities of caspase-8 and caspase-3 compared with the OA model control (P < 0.01). TNF receptor-associated factor-2 expression, phosphorylations of inhibitor of NF-κB kinase (IKK) subunits alpha and beta, and NF-κB inhibitor, alpha (IκBα) were all concentration-dependently suppressed by SF treatment. The results of EMSA showed that SF inhibited the activity of NF-κB. In addition, the expressions of cycloxygenase-2 and iNOS and the contents of prostaglandin E2 and NO were attenuated with the treatment of SF (P < 0.01).

Conclusion: SF has anti-apoptosis and anti-inflammatory effects on an OA model induced by IL-1β in vitro, which were due to inhibitory actions on the caspase-dependent apoptosis pathway and the IKK/NF-κB signal transduction pathway of the TNF/TNFR pathway.

Show MeSH
Related in: MedlinePlus