Methylation of the OP-1 promoter: potential role in the age-related decline in OP-1 expression in cartilage R.F. Loeser, M.D., H.-J. Im, Ph.D., B. Richardson,

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General Information Osteoarthritis and Cartilage

Presentation transcript:

Methylation of the OP-1 promoter: potential role in the age-related decline in OP-1 expression in cartilage R.F. Loeser, M.D., H.-J. Im, Ph.D., B. Richardson, M.D., Ph.D., Q. Lu, M.D., Ph.D., S. Chubinskaya, Ph.D. Osteoarthritis and Cartilage Volume 17, Issue 4, Pages 513-517 (April 2009) DOI: 10.1016/j.joca.2008.08.003 Copyright © 2008 Osteoarthritis Research Society International Terms and Conditions

Fig. 1 Age-related increase in methylation of OP-1 promoter DNA. (A) Methylation specific PCR for OP-1 using DNA isolated from cultures of young (20 years old) and old (74 years old) donor chondrocytes. DNA was amplified using primers recognizing sequences corresponding to methylated and unmethylated regions of bisulfite treated DNA. Samples with H2O substituted for DNA were used as a control to exclude primer–dimer formation. (B) Relationship of age to OP-1 promoter methylation status. The graph shows the ratio of PCR products obtained using the methylation specific primers normalized to results with primers from the unmethylated region (n=22, R2=0.277, P=0.014). Osteoarthritis and Cartilage 2009 17, 513-517DOI: (10.1016/j.joca.2008.08.003) Copyright © 2008 Osteoarthritis Research Society International Terms and Conditions

Fig. 2 Treatment of chondrocytes with 5-azacytidine (0–10μM) increases the production of OP-1. An ELISA was used to measure OP-1 levels in the media from cells of two different donors aged 72 and 80 years. OP-1 results were normalized to the DNA content measured in the cell-layer. *P<0.001 vs control. Osteoarthritis and Cartilage 2009 17, 513-517DOI: (10.1016/j.joca.2008.08.003) Copyright © 2008 Osteoarthritis Research Society International Terms and Conditions

Fig. 3 Effects of 5-azacytidine on chondrocyte gene expression. Primary chondrocytes isolated from donors with ages of 71–82 years were treated in culture with the indicated doses of 5-azacytidine for 120h. RNA was isolated and used for real-time PCR with primers specific for IGF-1, the IGF-1R, OP-1, aggrecan, or GAPDH as a control. (A–D) Results using cells cultured from the ankle joint (n=6) and (E) a comparison of cells cultured from ankle (n=7) and knee cartilage (n=2). *P<0.05. Osteoarthritis and Cartilage 2009 17, 513-517DOI: (10.1016/j.joca.2008.08.003) Copyright © 2008 Osteoarthritis Research Society International Terms and Conditions