Epithelial ovarian cancer (EOC) is usually discovered after extensive metastasis have developed in the peritoneal cavity. The ovarian surface is exposed to peritoneal fluid pressures and shear forces due to the continuous peristaltic motions of the gastro-intestinal system, creating a mechanical micro-environment for the cells. An in vitro experimental model was developed to expose EOC cells to steady fluid flow induced wall shear stresses (WSS). The EOC cells were cultured from OVCAR-3 cell line on denuded amniotic membranes in special wells. Wall shear stresses of 0.5, 1.0 and 1.5 dyne/cm2 were applied on the surface of the cells under conditions that mimic the physiological environment, followed by fluorescent stains of actin and β-tubulin fibers. The cytoskeleton response to WSS included cell elongation, stress fibers formation and generation of microtubules. More cytoskeletal components were produced by the cells and arranged in a denser and more organized structure within the cytoplasm. This suggests that WSS may have a significant role in the mechanical regulation of EOC peritoneal spreading.