Ab0048 the Suppression of Inflammatory Processes and Modulation of Mapk Signaling by Shikonin Derivatives in Primary Oa Chondrocytes

Abstract

BackgroundOsteoarthritis (OA) is one of the most common joint disorders and is characterized by the degeneration and loss of articular cartilage with chronic arthritis of the joint edge and subchondral bone. OA is causally influenced by several factors, including age, gender, familial susceptibility, as well as local biomechanics, cartilage cell apoptosis, and the action of degenerative enzymes. Despite intensive research, there are still few effective therapeutic approaches.ObjectivesIn the rat OA model, shikonin was shown to inhibit inflammatory processes and chondrocyte apoptosis by regulating the PI3K/AKT pathway. Therefore, we investigated the effect of shikonin and its derivatives acetylshikonin and cyclopropylshikonin on inflammation, MMP expression, and regulation of MAPK signaling in human OA chondrocytes.MethodsViability was analyzed using the CellTiter 96 AQueous Luminescence Assay on human healthy chondrocytes (HC) and primary OA chondrocytes (pCH-OA). For the study of inflammatory processes, we performed a proteome profile screening assay. As MAPK signaling pathways play a key role in cartilage destruction in OA, we analyzed the effects of shikonin and its derivatives using protein expression analysis of the phosphorylation pattern and the corresponding downstream gene regulation using RT-qPCR.ResultsBoth HC and pCH-OA showed a dose-dependent inhibition of cell viability after treatment with shikonin derivatives, whereby the strongest effects were found for shikonin with IC50 values of 1.2 µM and 1.3 µM, respectively. Shikonin counteracts inflammatory response caused by IL-1β by massively reducing the expression of pro-inflammatory mediators.Phosphorylation level of ERK changed slightly, pJNK and pp38 showed a significant increase after treatment with the shikonin derivatives both in HC and pCH-OA cells. The downstream targets c/EBPs and MEF2c may play a role in the homeostasis of joint cartilage under physiological and pathological conditions. The phosphorylation level of STAT3 significantly decreased in a dose-dependent manner after treatment. STAT3 blockade has a chondroprotective function through a regulation of cyclin D1 or Sox9.ConclusionOur results demonstrate for the first time that shikonin and its derivatives acetylshikonin and cyclopropylshikonin have extensive effects on inflammatory processes, MAPKs, and IL6/STAT3 downstream regulation in human healthy chondrocytes and primary OA chondrocytes.ReferencesnoneDisclosure of InterestsNone declared