Osteopontin (OPN) is a pro-inflammatory protein that paradoxically protects against inflammation and bone destruction in a mouse model of endodontic infection. Here we have tested the hypothesis that this effect of OPN is mediated by effects on migration of innate immune cells to the site of infection. Using the air pouch as a model of endodontic infection in mice, we showed that neutrophil accumulation at the site of infection with a mixture of endodontic pathogens is significantly reduced in OPN-deficient mice. Reduced neutrophil accumulation in the absence of OPN was accompanied by an increase in bacterial load. OPN-deficiency did not affect neutrophil survival, CXCR2 ligand expression, or the production of inflammatory cytokines in the air pouch. In vitro, OPN enhanced neutrophil migration to CXCL1, whereas in vivo, inhibition of CXCR2 suppressed cellular infiltration in air pouches of infected wild-type mice by > 50%, but had no effect in OPN-deficient mice. OPN increased cell surface expression of CXCR2 on bone marrow neutrophils in an integrin-α -dependent manner, and suppressed the internalization of CXCR2 in the absence of ligand. Together, these results support a model where the protective effect of OPN results from enhanced initial neutrophil accumulation at sites of infection resulting in optimal bacterial killing. We describe a novel mechanism for this effect of OPN: integrin-α -dependent suppression of CXCR2 internalization in neutrophils, which increases the ability of these cells to migrate to sites of infection in response to CXCR2 ligands.