Immunoblotting study to evaluate aggrecan
degradation patterns in rat intervertebral discs (IVDs) subjected to mechanical overload.
To evaluate the effects of in vivo
dynamic compression overloading on aggrecan
degradation products associated with matrix metalloproteinase
(MMP) and aggrecanase
activity in different regions of the IVD.
Summary of Background Data. Aggrecan
cleavage at the MMP and aggrecanase
sites is an important event in human IVD aging, with distinct cleavage patterns in the anulus and nucleus regions. No such information is available on regional variations in rat IVDs, nor on how such cleavage is affected by mechanical loading.
Sprague-Dawley rats were instrumented with an Ilizarov-type device and subjected to dynamic compression (1 MPa and 1 Hz for 8 hours per day for 8 weeks). Control, sham, and overloaded IVDs were separated by disc region and analyzed for aggrecan
degradation products using immunoblotting techniques, with antibodies specific for the aggrecanase
and MMP cleavage sites in the interglobular domain of aggrecan
Control IVDs exhibited strong regional variation in aggrecan
degradation patterns with minimal degradation products being present in the nucleus pulposus, degradation products associated with aggrecanase
cleavage predominating in the inner anulus fibrosus (AF), and degradation products associated with MMP cleavage predominating in the outer AF. Dynamic compression overloading increased the amount of aggrecan
degradation products associated with MMP cleavage not only in the AF but also in the nucleus pulposus. Degradation profiles of sham IVDs were similar to control.
G1 regions resulting from proteolysis were found to have a strong regionally specific pattern in the rat IVD, which was altered under excessive loading. The shift from aggrecanase
to MMP-induced degradation products with dynamic compression overloading suggests that protein degradation and loss can precede major structural disruption in the IVD, and that MMP-induced aggrecan
degradation may be a marker of mechanically induced disc degeneration