Age-associated differences in estrogen levels critically modify the cutaneous wound healing response. Using a microarray-based approach, we profiled changes in gene expression within the wounds of mice that were wild type or null for the pro-inflammatory cytokine macrophage migration inhibitory factor (MIF) in the presence or absence of estrogen. This experimental design identified more than 600 differentially expressed genes and established MIF as a key player in the wound healing process, regulating many novel repair/inflammation-associated gene targets. Moreover, MIF affected virtually all of the effects of reduced estrogen on wound repair. In humans, serum and wound levels of MIF increased with age and were strongly down-regulated by estrogen in vivo. Estrogen-regulated MIF transcription in vitro via a nuclear factor kappaB-dependent mechanism. These findings have wide-ranging implications for the many pathophysiological states in which MIF plays an important regulatory role and suggest a potential therapeutic role for MIF in modulating clinical conditions associated with age-related decline in estrogen levels.