This study investigated the effects of a fish oil- (FO-) based lipid emulsion on muscle leukocyte chemotaxis and inflammatory responses in a murine model of limb ischemia-reperfusion (IR) injury. Mice were assigned randomly to 1 sham (sham) group, 2 ischemic groups, and 2 IR groups. The sham group did not undergo the ischemic procedure. The mice assigned to the ischemic or IR groups were pretreated intraperitoneally with either saline or FO-based lipid emulsion for 3 consecutive days. The IR procedure was induced by applying a 4.5 oz orthodontic rubber band to the left thigh above the greater trochanter for 120 min and then cutting the band to allow reperfusion. The ischemic groups were sacrificed immediately while the IR groups were sacrificed 24 h after reperfusion. Blood, IR-injured gastrocnemius, and lung tissues were collected for analysis. The results showed that FO pretreatment suppressed the local and systemic expression of several IR-induced proinflammatory mediators. Also, the FO-pretreated group had lower blood Ly6ChiCCR2hi monocyte percentage and muscle M1/M2 ratio than the saline group at 24 h after reperfusion. These findings suggest that FO pretreatment may have a protective role in limb IR injury by modulating the expression of proinflammatory mediators and regulating the polarization of macrophage.