Nitration products of unsaturated fatty acids are formed via NO-dependent oxidative reactions and appear to be a new class of endogenous anti-inflammatory mediators. Nitroalkene derivatives of LNO₂ and OA-NO₂ alleviate inflammatory responses in macrophages, but the underlying mechanisms remain to be fully defined. Herein we report that LNO₂ and OA-NO₂ suppress pro-inflammatory STAT signaling in macrophages. In RAW264.7 cells, a murine macrophage cell line, LNO₂ and OA-NO₂ inhibited the LPS-induced STAT1 phosphorylation and the STAT1-dependent transcriptional activity, thereby suppressing expression of its target gene such as iNOS and MCP-1. The nitroalkene-mediated inhibition of STAT1 activity was not affected by cPITO (a NO scavenger), GW9662 (a PPAR specific antagonist) or glutathione (an antioxidant), suggesting an underlying mechanism independent of NO, PPAR or thio-nitralkylation. In contrast, LNO₂ or OA-NO₂ alone up-regulated both mRNA and protein levels of MKP-1, and strongly augmented the LPS-induced MKP-1 protein expression. Knockdown of MKP-1 by MKP-1 siRNA enhanced the LPS-induced STAT1 phosphorylation, suggesting that MKP-1 acts as a negative regulator for LPS-induced STAT signaling. In addition, the nitroalkene-mediated inhibitory effects on STAT1 phosphorylation, iNOS expression and MCP-1 secretion were also largely attenuated by the MKP-1 siRNA approach. Taken together, our data demonstrate that nitroalkenes inhibit pro-inflammatory STAT signaling through inducting MKP-1 in macrophages.