Matrix metalloproteinase (MMP)-8 and MMP-9 play several roles in inflammation, including degradation of extracellular matrix (ECM) components and regulation of cytokine activity. To determine the roles of MMP-8 and MMP-9 in a neutrophil-dependent inflammatory response, we used a murine model of corneal inflammation in which LPS is injected into the corneal stroma. In contrast to wild-type mice, we found that i) lipopolysaccharide (LPS)-injected CXCR2 -/- corneas had impaired neutrophil infiltration and did not express either MMP-8 or MMP-9; ii) neutrophil migration through the central cornea was impaired in Mmp8-/-, but not Mmp9-/-, mice; iii) neutrophil migration was inhibited in collagenase-resistant mice; iv) the chemotactic Pro-Gly-Pro (PGP) tripeptide that binds CXCR2 was decreased in CXCR2-/- mice; v) PGP production was impaired in Mmp8-/- corneas; and vi) neutralizing anti-PGP antibody did not inhibit neutrophil infiltration in Mmp8-/- mice. We found no effects of MMP-8 on LPS-induced CXC chemokine (LIX, or CXCL5)-induced neutrophil recruitment or on LPS-induced CXC chemokine production. Together, these studies indicate that neutrophils contribute to the production of both MMP-8 and MMP-9 in LPS-injected corneas and that MMP-8 regulates neutrophil migration through the dense collagenous ECM of the corneal stroma by generating chemotactic PGP during inflammation. Copyright © American Society for Investigative Pathology.