Effects of Brazilin on human chondrocytes and synoviocytes

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Purpose: Osteoarthritis (OA) is the most prevalent disorder of the musculoskeletal system and has a very high socioeconomic impact. Current treatment options for OA are only symptomatic and do not act on the causes of OA or stop the progression of the disease. Sappan Lignum, the heartwood of Caesalpinia sappan L. (Leguminosae), has been widely used in oriental traditional medicine for improvement of blood circulation, sprains, and emmeniopathy. Different extracts and active compounds such as brazilin have been reported to possess antioxidative and anti-inflammatory effects. This study investigates the influence of brazilin on the expression of IL1ß, TNF-alpha, PGE-2, MMP3 and MMP13 in primary chondrocytes and synoviocytes of OA patients. Methods: Brazilin was isolated from Caesalpina sappan using preparative HPLC and identified using 1H-NMR. Chondrocytes and synoviocytes were isolated from OA patients undergoing joint replacement. The cytotoxicity of Brazilin was assessed using an MTT-based test. Cells were preincubated with Brazilin (5, 10, 20 μg/ml) for 1 h and then incubated for 6 h in the presence of 10 ng/ml IL-1ß prior to RNA isolation and qPCR analyses of target gene mRNA levels. Target protein concentrations in the cell culture supernatants were measured using ELISA. Results: There was no significant cytotoxicity of brazilin up to concentrations of 40 μg/ml in primary chondrocytes and synoviocytes. COX-2, TNF-alpha and MMP3 mRNA levels in primary chondrocytes were strongly increased up to 200 fold by 10 ng/ml IL-1ß, whereas pre-treatment with 5-20 μg/ml brazilin significantly reduced this effect dose dependently. In agreement, pretreatment with 10 μg/ml brazilin significantly suppressed the IL-1ß-induced TNF-alpha and MMP3 production in cell culture supernatants. In IL1ß stimulated primary synoviocytes, Brazilin markedly suppressed IL1ß, TNF-alpha and MMP13 mRNA and protein levels. Conclusions: In summary, we isolated brazilin, identified its structure, and determined its biological activities on OA chondrocytes and synoviocytes. Brazilin substantially suppressed the expression of proinflammatory mediators and major MMPs suggesting that brazilin may be beneficial for future management of OA.