Aggrecanases and aggrecanase-generated fragments in the human intervertebral disc at early and advanced stages of disc degeneration

Study Design. A comparative study of aggrecanases and aggrecan fragmentation profile in the human intervertebral disc at early and advanced stages of disc degeneration. Objective. To determine differences in the content of the aggrecanases and the profile of aggrecan fragmentation in early and advanced stages of disc degeneration using cadaveric human intervertebral discs. Summary of Background Data. Aggrecanases and aggrecanase-generated aggrecan fragments have been found in human degenerated discs. However, the association between the grade of disc degeneration and the content of the aggrecanases and the profile of aggrecan fragments has not been well studied. Methods. A total of 108 cadaveric donor spines were assessed by MRI T2 imaging and graded based on the Thompson scale. Twelve donor spines ( average age, 63 years), each specifically exhibiting 2 different stages ( Grade 2 and Grade 4) of disc degeneration at different disc levels, were included in this study. After harvesting the preselected discs, tissue samples were obtained from the center of the nucleus pulposus (NP) and the middle zone of the anulus fibrosus (AF). The amount of the aggrecanases, specifically ADAMTS-4 and ADAMTS-5, and the pattern of aggrecan fragmentation in the isolated tissues were assessed by western blot using specific antibodies. Results. In both NP and the AF tissues, the amount of ADAMTS-4 detected was higher in disc tissues with a higher level of degeneration (Grade 4) than in Grade 2 disc tissues with a lower level of degeneration. However, the amount of ADAMTS-5 detected did not differ between the 2 disc tissue grades. The aggrecan fragmentation analysis of these samples demonstrated the presence of aggrecanase-mediated fragmentation in both groups; however, there was no apparent difference in the aggrecan fragmentation profile between discs at early and advanced stages of disc degeneration. Conclusion. Aggrecanases are involved in aggrecanolysis at both the early and advanced stages of disc degeneration. The aggrecan fragmentation profile analysis demonstrates the involvement of aggrecanases, as well as that of matrix metalloproteinases and/or cathepsins, during disc degeneration.