Objective: To investigate the effect of parthenolide (PAR) on the tumor necrosis factor (TNF)-alpha induced aggrecan catabolism of chondrocytes in osteoarthritis (OA).
Methods: Human chondrocytes were obtained from the condyles of femur of OA patients undergoing knee joint replacement during operation, cultured, and randomly divided into 4 groups: control group, TNF group (cultured in medium containing 10 ng/ml TNF-alpha) , PAR group (cultured in medium containing 10 micromol/L PAR), and PAR + TNF group (cultured in medium containing 10 micromol/L PAR and 10 ng/ml TNF-alpha). Eight days later 1, 9-dimethylmethylene blue spectrophotometric assay was used to measure the content of glycosaminoglycan (GAG) , marker of aggrecan catabolism, in the culture fluid and the cells. Using anti-aggrecan monoclonal antibodies Mab 5D4 and 3B3, ELISA was employed to detect the contents of 5D4 and 3B3, aggrecan catabolic fragments. RT-PCR was used to detect the mRNA expression of the aggrecan, ADAMTS-4 and ADAMTS-5, both aggrecanases, tissue inhibitor of metalloprotease (TIMP)-1, mitogen-activated protein kinase (MAPK)-1 and 18S, a marker.
Results: The GAG percentage in the culture fluid of the TNF-alpha was 57.1% +/- 2.0%, significantly higher than those of the control and TNF-alpha + PAR groups (P = 0.001 and 0.02). The 5D4 fragment level of the TNF-alpha group was 509 ng/ml +/- 32 ng/ml, significantly higher than that of the control group (166 ng/ml +/- 15 ng/ml, t = 11.60, P =0.007), and the level of 5D4 fragment of the PAR + TNF-alpha group was 333 ng/ml +/- 15 ng/ml, significantly lower than that of the TNF-alpha group (t = 7.93, P = 0.016). There was not significant difference in the 3B3 fragment level among the 4 groups (F = 1.316, P = 0.335). The aggrecan mRNA expression level of the TNF-alpha group was significantly lower than those of the other 3 groups (F = 133.7, P = 0.000), the mRNA expression levels of ADAMTS-5 and MAPK-1 of the TNF-alpha group were significantly higher than those of the other 3 groups (F = 209. 7, 117.1; P =0. 000), the ADAMTS-5 mRNA expression level of the PAR group was significantly lower than that of the control group (t = 11.1, P= 0.008) , and there was not significant differences in the mRNA expression levels of ADAMTS-4 and TIMP-1 among the 4 groups (F = 1.87, 0.73; P > 0.05) .
Conclusion: PAR inhibits TNF-alpha induced catabolism of aggrecan in the chondrocytes of OA and reduces the mRNA expression of ADAMTS-5 and MAPK-1. PAR may be useful in the treatment of OA and other inflammatory joint diseases.