Skeletal muscle has recently been described as an endocrine organ, capable of releasing cytokines and regulators of metabolism. Microdialysis of the interstitial space of skeletal muscle enables analysis of the release of such cytokines. The purpose of this study was to determine the transient changes in concentration of metabolites and cytokines in human skeletal muscle in a 7h period following the insertion of a microdialysis probe. In total, sixteen microdialysis catheters were inserted into the vastus lateralis of male participants (age 26.2±1.35y, height 180.8±3.89cm, mass 83.9±3.86kg, BMI 25.7±0.87kgm(-2), body fat 26.1±3.0%). Serial samples were analyzed by micro-enzymatic and multiplexed immunoassay. Muscle interstitial glucose and lactate levels remained stable throughout, amino acid concentrations stabilized after 2.5h, however, insertion of a microdialysis catheter induced a 29-fold increase in peak IL-6 (p<0.001) and 35-fold increase in peak IL-8 concentrations (p<0.001) above basal levels 6h post insertion. In contrast to stable amino acid, glucose and lactate concentrations after 2h, commonly reported markers of tissue homeostasis in in vivo microdialysis, the multi-fold increase in IL-6 and IL-8 following insertion of a microdialysis catheter is indicative of a sustained disturbance of tissue homeostasis.