Interstitial flow (IF) modulates both the biochemical and biophysical cues surrounding cells. It represents a very important regulating mechanism for cell/tissue function and has been commonly utilized in tissue engineering (TE). This article discusses the possible regulating mechanisms of IF on fibroblasts, the various fibroblast responses to IF, the current challenges in understanding the IF-fibroblast relationship and the application of IF for fibroblast involved TE. In particular, IF can affect fibroblast growth at both intracellular (e. g., calcium signaling, protein/proteinase secretion) and cellular (e. g., autocrine/paracrine signaling, proliferation, differentiation, alignment, adhesion, migration) levels. One major challenge for understanding IF-fibroblast interaction has been the determination of the flow and cell growth condition at microlevel especially in a three-dimensional environment. To utilize IF and optimize the fluidic environment for TE, several influencing factors in the system including perfusate composition, flow profile, nutrient supply, signaling molecule effect, scaffold property, and fibroblast type should be considered. Biotechnol. Bioeng. 2010; 107: 1-10. (C) 2010 Wiley Periodicals, Inc.