The Mechanism Of Curcumin Regulates Chondrocytes Autophagy Through ERK1/2 Signaling Pathways

ObjectiveOsteoarthritis(OA)has become an increasingly prominent chronic degeneration disease affecting the elderly life quality and increasing the income of medical expenses.The nonsteroidal anti-inflammatory drugs are the currently main available treatments for OAand related arthritic diseases,while these drugs reveal numerous side effects and only temporarily effective.Therefore safe and more efficacious anti-inflamma-tory drugs are needed to treat OA.Curcumin,which is a naturally occurring polyphenolic compound,is present in turmeric,possesses both anti-inflammatory and antioxidant properties.Autophagy is generally thought a protective mechanism which is activated during nutritionally depleted or hypoxic conditions in order to facilitate cell survival.The aim of this study was to investigate if there is a link between curcumin and apoptosis on IL-1β-stimulated primary chondrocytes in vitro and whether the ERK signaling pathway is involved in autophagy.Methods1.Analysis of the effects of curcumin on chondrocytes apoptosis:chondrocytes were isolated from Sprague Dawley rat articular cartilage.Chondrocytes were stimulated with IL-1β or/with curcumin.CCK-8 method,TUNEL staining and flow cytometry(FCM)were used to detect the cell viability and cell apoptosis;The expression of apoptosis related proteins in chondrocytes was detected by Western blot.2.Analysis of the effects of curcumin on chondrocytes autophagy:chondrocytes were isolated from Sprague Dawley rat articular cartilage.Chondrocytes were stimulated with IL-1β to mimic osteoarthritis model.The optimal concentration of curcumin was used to treat the chondrocytes MDC staining and transmission electron microscope(TEM)were used to detect the intracellular autophagy level;The expression of autophagy related proteins in chondrocytes was detected by Western blot.3.The role of ERK1/2 signal pathway on OA chondrocytes treated with curcumin:Chondrocytes were incubated with IL-1β alone or were preincubated with curcumin and then co-treated with IL-1β,the expression of phosphorylated ERK1/2 was detected by Western blot.The expression of autophagy molecular markers was also detected with Western blot in chondrocytes treated with ERK1/2 signaling pathway inhibitor U0126 or pretreated with curcumin.Results1.Curcumin can significantly inhibit the apoptosis of chondrocytes incubated with IL-1β:IL-1β decreased cell viability significantly in a dose-and time-dependent manner.Low dose of curcumin significantly inhibited the apoptosis induced by IL-1β.Western blot results showed that curcumin inhibited caspase-3 activation and increase the expression of Bcl-2 in chondrocytes incubated with IL-1β.2.Curcumin can obviously activate autophagy in chondrocytes treated with IL-1β:the results of MDC staining and the transmission electron microscope(TEM)indicated that the autophagosome were increased in the chondrocytes cotreated with curcumin and IL-1βcompared with that in chondrocytes only treated with IL-1β.Western blot results demonstrated that the expression of LC-3Ⅱ and Beclinl were increased in chondrocytes treated with curcumin.All these results showed that the level of autophagy was significantly increased.3.ERK1/2 signaling pathway participates in the curcumin regulation of chondrocytes autophagy:curcumin can regulate the expression of molecular markers ERK1/2 in chondrocytes in a time-dependent manner.However,the expression of autophagy molecular markers LC-3Ⅱ and Beclinl induced by curcumin were significantly decreased after treated with ERK inhibitor U0126.ConclusionOur results sugest that curcumin suppresses apoptosis and inflammatory signaling through its actions on the ERK1/2-induced autophagy in chondrocytes.