Osteopontin Isoforms Differentially Promote Arteriogenesis in Response to Ischemia via Macrophage Accumulation and Survival

OBJECTIVE: Osteopontin (OPN) is critical for ischemia-induced neovascularization. Unlike rodents, humans express 3 OPN isoforms (a, b, and c); however, the roles of these isoforms in post-ischemic neovascularization and cell migration remain undefined. Our objective was to determine if OPN isoforms differentially affect post-ischemic neovascularization and to elucidate the mechanisms underlying these differences. APPROACH AND RESULTS: To investigate if human OPN isoforms exert divergent effects on post-ischemic neovascularization, we utilized OPN(−/−) mice and a loss-of-function/gain-of-function approach in vivo and in vitro. In this study OPN(−/−) mice underwent hindlimb ischemia surgery and 1.5×10(6) lentivirus particles were administered intramuscularly to overexpress OPNa, OPNb or OPNc. OPNa and OPNc significantly improved limb perfusion 30.4%±0.8 and 70.9%±6.3, respectively, and this translated to improved functional limb use, as measured by voluntary running wheel utilization. OPNa and OPNc treated animals exhibited significant increases in arteriogenesis, defined here as the remodelling of existing arterioles into larger conductance arteries. Macrophages play a prominent role in the arteriogenesis process and OPNa and OPNc treated animals showed significant increases in macrophage accumulation in vivo. In vitro, OPN isoforms did not affect macrophage polarization, whereas all 3 isoforms increased macrophage survival and decreased macrophage apoptosis. However, OPN isoforms exert differential effects on macrophage migration, where OPNa and OPNc significantly increased macrophage migration with OPNc serving as the most potent isoform for macrophage migration. CONCLUSIONS: Human OPN isoforms exert divergent effects on neovascularization through differential effects on arteriogenesis and macrophage accumulation in vivo and on macrophage migration and survival, but not polarization, in vitro. Altogether, these data support that human OPN isoforms may represent novel therapeutic targets to improve neovascualrization and preserve tissue function in patients with obstructive artery diseases.