Background Cartilage degradation is a hallmark of arthritic disease. The main constituents of cartilage is aggrecan and type II collagen. Previous studies have shown that aggrecan degradation is prior to type II collagen degradation and that only aggrecanase degradation but not matrix metalloproteinases (MMP) degradation of aggrecan was reversible1. It is therefore essential to have biomarkers that can detect reversible and irreversible aggrecan degradation to identify the condition of the cartilage. Of the well-known aggrecanase degradation site at NITEGE373_374ARGS, ARGS is released by only aggrecanse, but NITEGE is retained in the aggrecan molecule and in theory needs additional aggrecan degradation to be released. The numbers indicate the aggrecanase cleavage site.
Objectives Investigate the profile of two neo-epitope biomarkers of aggrecanase degraded aggrecan at NITEGE373–374ARGS in a bovine cartilage explants model.
Methods Catabolic stimulated (oncostatin M and TNF-a; O+T) bovine cartilage explants were treated with or without the generic MMP inhibitor, GM6001. Explants without catabolic stimulation was used as negative control. In the culture supernatant two biomarkers investigating aggrecanase degraded aggrecan was measured. The AGNxI competitive ELISA detects the NITEGE373 neo-epitope and the ARGS sandwich ELISA detects the 374ARGSVI neo-epitope. Statistical differences between groups were tested by two-way ANOVA.
Results The AGNxI level was significantly increased compared to w/o from day 5 to day 17. GM6001 significantly lowered the release of AGNxI compared to O+T alone at day 7 (p<0.0001), day 10 (p=0.0007) and 12 (p=0.01). In addition, GM6001 shifted the release to 2days later in the culture period, as the AGNxI level was first significantly increased compared to w/o at day 7 to day 17, where the level was higher than O+T, but non-significant. The ARGS level was significantly increased at day 5, 10 and 12 for both O+T and O+T + GM6001 compared to w/o. At day 10 the ARGS level was higher in O+T + GM6001 than O+T (p<0.0001).
Conclusions The prolonged release of AGNxI and the significantly lowered level of AGNxI with the MMP-inhibitor, suggest that the AGNxI release is somewhat dependent of MMP activity. In contrast, the ARGS release seems not to be MMP-dependent as there was a significant increase in O+T with the MMP-inhibitor compared to w/o and the peak was pronounced. In summary, aggrecan degradation is not just aggrecan degradation and different neo-epitopes have different importance in cartilage degradation.
Arthritis Res Ther. 2008;10(3):R63
Disclosure of Interest A. S. Siebuhr Employee of: NORDIC BIOSCIENCE A/S, Y. He Employee of: NORDIC BIOSCIENCE A/S, S. Hoielt: None declared, Y. Luo: None declared, M. Karsdal Shareholder of: NORDIC BIOSCIENCE A/S, Employee of: NORDIC BIOSCIENCE A/S, A.-C. Bay-Jensen Employee of: NORDIC BIOSCIENCE A/S