MMP-2 mediates local degradation and remodeling of collagen by annulus fibrosus cells of the intervertebral disc

Abstract

IntroductionDegeneration of the intervertebral disc (IVD) is characterized by marked degradation and restructuring of the annulus fibrosus (AF). Although several matrix metalloproteinases (MMPs) have been found to be more prevalent in degenerate discs, their coordination and function within the context of the disease process are still not well understood. In this study, we sought to determine whether MMP-2 is associated with degenerative changes in the AF and to identify the manner by which AF cells use MMP-2.MethodsTwo established animal models of disc degeneration, static compression and transannular needle puncture of rodent caudal discs, were examined for MMP-2 immunopositivity. With lentiviral transduction of an shRNA expression cassette, we screened and identified an effective shRNA sequence for generating stable RNA interference to silence MMP-2 expression in primary rat AF cells. Gelatin films were used to compare gelatinase activity and spatial patterns of degradation between transduced cells, and both noninfected and nonsense shRNA controls. The functional significance of MMP-2 was determined by assessing the ability for cells to remodel collagen gels.ResultsBoth static compression and 18-g annular puncture of rodent caudal discs stimulated an increase in MMP-2 activity with concurrent lamellar disorganization in the AF, whereas 22-g and 26-g needle injuries did not. To investigate the functional role of MMP-2, we established lentivirus-mediated RNAi to induce stable knockdown of transcript levels by as much as 88%, and protein levels by as much as 95% over a 10-day period. Culturing transduced cells on gelatin films confirmed that MMP-2 is the primary functional gelatinase in AF cells, and that MMP-2 is used locally in regions immediately around AF cells. In collagen gels, transduced cells demonstrated an inability to remodel collagen matrices.ConclusionsOur study indicates that increases in MMP-2 observed in human degenerate discs are mirrored in experimentally induced degenerative changes in rodent animal models. AF cells appear to use MMP-2 in a very directed fashion for local matrix degradation and collagen remodeling. This suggests that MMP-2 may have a functionally significant role in the etiology of degenerative disc disease and could be a potential therapeutic target.

Tumor Cell Development: A Role for Viruses and Telomerase Activity?

“…This miRNA was observed to be overexpressed in HBx-transfected HCC cells and HepG2 cells that constitutively replicate HBV, as well as in p21-HBx transgenic mice, and such up-regulation favored migration of HepG2 cells. In the same study, miR-29a was shown to target PTEN mRNA, thus reducing PTEN levels and favoring the activation AKT and MMP2 proteins, 110 which are involved in cell proliferation 111 and degradation of extracellular matrix, 112 respectively. Regarding telomerase modulation, there is evidence supporting that PTEN inhibits telomerase activity by reducing TERT mRNA levels, 113 while AKT up-regulates telomerase by phosphorylation of the TERT subunit.…”

Altered expression of metalloproteinase-2 and tissue inhibitor of metalloproteinase-2 in cervical disc herniation patients

“…In the AF, matrix changes are accelerated with increase in age, which is thought to be a result of enhanced catabolic processing of the ECM (Akeda et al, 2007;Salo et al, 2008). While MMP-2 generally exists in an inactive pro-enzymatic form, it was observed that mechanical stress could induce MMP-2 activation (MacLean et al, 2008;Rastogi et al, 2013). Since mechanical stress is a major risk factor for the development of CDH, we hypothesized that MMP expression may be associated with the occurrence of CDH.…”

“…MMP-2 was demonstrated to play an important part in degenerative disc disease as it interferes with the activity of AF cells (Rastogi et al, 2013). Particularly, the knockdown of MMP-2 in AF cells leads to a significantly decreased ability to reorganize and contract collagen gels (Rastogi et al, 2013). IVD tissue structure is thought to be mainly composed of collagens and proteoglycans (Kozaci et al, 2006).…”

“…It is widely accepted that IVD degeneration may be linked to proteolytic degradation of the ECM, resulting in several structural changes, such as AF disorganization, calcification of the cartilaginous end plates, and dehydration and fibrosis of the NP (Boos et al, 2002). MMP-2 was demonstrated to play an important part in degenerative disc disease as it interferes with the activity of AF cells (Rastogi et al, 2013). Particularly, the knockdown of MMP-2 in AF cells leads to a significantly decreased ability to reorganize and contract collagen gels (Rastogi et al, 2013).…”

“…As one important member of the MMP family, MMP-2 actively functions in the secondary breakdown of collagen during the tissue remodeling process (Kozaci et al, 2006;Séguin et al, 2008). Additionally, a large amount of evidence in animal models suggest that MMP-2 is significantly correlated with degenerative changes (Hsieh et al, 2009;Rastogi et al, 2013). It is widely accepted that IVD degeneration may be linked to proteolytic degradation of the ECM, resulting in several structural changes, such as AF disorganization, calcification of the cartilaginous end plates, and dehydration and fibrosis of the NP (Boos et al, 2002).…”