This study proposed that attenuated expression of inflammatory factors is an underlying mechanism driving the repeated-bout effect (rapid adaptation to eccentric exercise). We investigated changes in mRNA levels and protein localization of inflammatory genes after two bouts of muscle-lengthening exercise. Seven male subjects performed two bouts of lower body exercise (separated by 4 wk) in which one leg performed 300 eccentric-concentric actions, and the contralateral leg performed 300 concentric actions only. Vastus lateralis biopsies were collected at 6 h, and strength was assessed at baseline and at 0, 3, and 5 days after exercise. mRNA levels were measured via semiquantitative RT-PCR for the following genes: CYR61, HSP40, HSP70, IL1R1, TCF8, ZFP36, CEBPD, and MCP1. Muscle functional adaptation was demonstrated via attenuated strength loss (16% less, P = 0.04) at 5 days after bout 2 compared with bout 1 in the eccentrically exercised leg. mRNA expression of three of the eight genes tested was significantly elevated in the eccentrically exercised leg from bout 1 to bout 2 (+3.9-fold for ZFP36, +2.3-fold for CEBPD, and +2.6-fold for MCP1), while all eight mRNA levels were unaffected by bout in the concentrically exercised leg. Immunohistochemistry further localized the protein of one of the elevated factors [monocyte chemoattractant protein-1 (MCP1)] within the tissue. MCP1 colocalized with resident macrophage and satellite cell populations, suggesting that alterations in cytokine signaling between these cell populations may play a role in muscle adaptation to exercise. Contrary to our hypothesis, several inflammatory genes were transcriptionally upregulated (rather than attenuated) after a repeated exercise bout, potentially indicating a role for these genes in the adaptation process.