Matrix metalloproteinase-3–dependent generation of a macrophage chemoattractant in a model of herniated disc resorption
Hirotaka Haro, Howard C. Crawford, Barbara Fingleton, John R. MacDougall, Kenichi Shinomiya, Dan M. Spengler, Lynn M. Matrisian
Hirotaka Haro, Howard C. Crawford, Barbara Fingleton, John R. MacDougall, Kenichi Shinomiya, Dan M. Spengler, Lynn M. Matrisian
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Matrix metalloproteinase-3–dependent generation of a macrophage chemoattractant in a model of herniated disc resorption

Abstract

Herniated disc (HD) is a common health problem that is resolved by surgery unless spontaneous resorption occurs. HD tissue contains abundant macrophage infiltration and high levels of matrix metalloproteinases (MMPs) MMP-3 and MMP-7. We developed a model system in which disc tissue or isolated chondrocytes from wild-type or MMP-null mice were cocultured with peritoneal macrophages and used this system to investigate the role of MMPs and chondrocyte/macrophage interactions in disc resorption. We observed a marked enhancement of MMP-3 protein and mRNA in chondrocytes after exposure to macrophages. Chondrocytic MMP-3, but not MMP-7, was required for disc resorption, as determined by assaying for a reduction in wet weight and proteoglycan content after 3 days of coculture. Surprisingly, chondrocyte MMP-3 was required for the generation of a macrophage chemoattractant and the subsequent infiltration of the disc tissue by proteolytically active macrophages. We conclude that macrophage induction of chondrocyte MMP-3 plays a major role in disc resorption by mechanisms that include the generation of a bioactive macrophage chemoattractant.

Authors

Hirotaka Haro, Howard C. Crawford, Barbara Fingleton, John R. MacDougall, Kenichi Shinomiya, Dan M. Spengler, Lynn M. Matrisian

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Chemotactic activity of macrophage/disc coculture–conditioned medium. Pe...
Chemotactic activity of macrophage/disc coculture–conditioned medium. Peritoneal macrophages were placed in the upper level of a Boyden chamber, and migration was quantified by counting the number of cells migrating to the lower side of the filter in 6 high-power fields (×400) per well. (a) Medium in lower chamber was from wild-type (wt) or MMP-3–null (3–) discs cocultured with or without macrophages (mφ). *Significant migration (P < 0.05) compared with conditioned medium without macrophages. (b) Migration of wild-type macrophages toward macrophage/disc coculture media was analyzed using a modified Boyden chamber assay and diluting (< 100%) or concentrating (> 100%) conditioned medium (CM) from either wt macrophage/wt disc or wt macrophage/MMP-3–null disc cocultures. A checkerboard analysis of migration toward a concentration gradient was performed by placing increasing concentrations of coculture medium in the lower chamber with 0% (squares), 50% (diamonds), or 100% (circles) media in the upper chamber. Plotted are the average number of cells induced to migrate by wt/wt coculture medium with the MMP-3–independent background (number of cells induced to migrate by wt/3– medium) subtracted. Note that enhanced migration does not occur until the concentration of the medium in the lower chamber exceeds the concentration of the medium in the upper chamber; i.e., a concentration gradient is established.