Cytokines act as chemical mediators during the inflammatory process. Measurements of cytokine level in tissue have previously been performed in homogenized tissue, and the true concentrations in native interstitial fluid (ISF), i.e. the compartment where cytokines exert their biologically active role, have remained unknown. The role of skeletal muscle myocytes as a source for cytokines during endotoxemia was explored by collecting muscle ISF using a wick method, and the level of 14 cytokines in ISF and plasma was related to corresponding changes in mRNA level to reveal any potential discrepancies between gene expression and protein release of cytokines to ISF. The majority of investigated cytokines were elevated in muscle ISF during endotoxemia, and an analysis of cytokine mRNA level revealed consistency between gene expression and protein release. Elevated cytokine level in ISF, in addition to elevated gene expression in muscle, indicated a significant local production and release of several proinflammatory cytokines and chemokines within skeletal muscle tissue during endotoxemia. Immunohistochemistry revealed that myocytes constituted a significant source of IL-1 and TNF- production during endotoxemia, while the contribution from inflammatory cells i.e. leukocytes, were found to be less significant. Muscle cells apparently constitutes an important source of several different cytokines during endotoxemia, governing the level in the microenvironment surrounding muscle cells, and are likely to contribute significantly to the cytokine level in plasma.