Wogonin, a plant derived small molecule, exerts potent anti-inflammatory and chondroprotective effects through the activation of ROS/ERK/Nrf2 signaling pathways in human Osteoarthritis chondrocytes

Abstract

Osteoarthritis (OA), characterized by progressive destruction of articular cartilage, is the most common form of human arthritis. Here, we evaluated the potential chondroprotective and anti-inflammatory effects of Wogonin, a naturally occurring flavonoid, in IL-1β-stimulated human OA chondrocytes and cartilage explants. Wogonin completely suppressed the expression and production of inflammatory mediators including IL-6, COX-2, PGE2, iNOS and NO in IL-1β-stimulated OA chondrocytes. Further, Wogonin exhibits potent chondroprotective potential by switching the signaling axis of matrix degradation from catabolic towards anabolic ends and inhibited the expression, production and activities of matrix degrading proteases including MMP-13, MMP-3, MMP-9, and ADAMTS-4 in OA chondrocytes, and blocked the release of s-GAG and COL2A1 in IL-1β-stimulated OA cartilage explants. Wogonin also elevated the expression of cartilage anabolic factors COL2A1 and ACAN in chondrocytes and inhibited the IL-1β-mediated depletion of COL2A1 and proteoglycan content in the matrix of cartilage explants. The suppressive effect of Wogonin was not mediated through the inhibition of MAPKs or NF-κB activation. Instead, Wogonin induced mild oxidative stress through the generation of ROS and depletion of cellular GSH, thereby modulating the cellular redox leading to the induction of Nrf2/ARE pathways through activation of ROS/ERK/Nrf2/HO-1-SOD2-NQO1-GCLC signaling axis in OA chondrocytes. Molecular docking studies revealed that Wogonin can disrupt KEAP-1/Nrf-2 interaction by directly blocking the binding site of Nrf-2 in the KEAP-1 protein. Genetic ablation of Nrf2 using specific siRNA, significantly abrogated the anti-inflammatory and chondroprotective potential of Wogonin in IL-1β-stimulated OA chondrocytes. Our data indicates that Wogonin exerts chondroprotective effects through the suppression of molecular events involved in oxidative stress, inflammation and matrix degradation in OA chondrocytes and cartilage explants. The study provides novel insights into the development of Nrf2 as a promising candidate and Wogonin as a therapeutic agent for the management of OA.

Wogonin, a natural flavonoid, intercalates with genomic DNA and exhibits protective effects in IL-1β stimulated osteoarthritis chondrocytes

“…Wogonin possess diverse biological properties including anti-cancer and anti-inflammatory activities [1–3]. Recently, we have shown the chondroprotective effect of Wogonin through effective suppression of MMPs and other catabolic genes in human osteoarthritis (OA) chondrocytes [4, 5]. Our studies demonstrate that Wogonin exerts protective effects through modulation of redox sensitive transcription factors AP-1 and Nrf2 [4, 5].…”

“…Recently, we have shown the chondroprotective effect of Wogonin through effective suppression of MMPs and other catabolic genes in human osteoarthritis (OA) chondrocytes [4, 5]. Our studies demonstrate that Wogonin exerts protective effects through modulation of redox sensitive transcription factors AP-1 and Nrf2 [4, 5]. However, molecular targets of these flavonoids pertinent to wide range of biological activities are subject to intensive research.…”

“…Cellular uptake of Wogonin in human OA chondrocytes was monitored using LC-MS/MS analysis [5]. The cell lysate for mass spectrometric analysis for Wogonin was prepared as described earlier [5].…”

“…The cell lysate for mass spectrometric analysis for Wogonin was prepared as described earlier [5]. Briefly, after the treatment with Wogonin (50–100 μM) for 4 or 24h, OA chondrocytes were washed three times with phosphate-buffered saline, lysed with lysis buffer (10 mM Tris-HCl (pH 9.6), 1% Triton X-100,150 mM NaCl, 5 mM EDTA) and deproteinized with an equivalent amount of acetonitrile.…”

“…Chondrocytes treated with Wogonin (50 μM) under stimulation with IL-1β (10 ng/ml, 48 h) were harvested, washed with cold PBS and lysed in ice-cold RIPA buffer and lysate were prepared as described previously [5]. Equivalent amounts of lysate protein (35 μg) were resolved by 10% SDS-PAGE and transferred to a PVDF membrane (Bio-Rad, USA) and the blots were incubated with primary antibodies diluted in 2% blocking buffer for overnight at 4°C.…”