We herein introduce a novel multi-well stretching device that is made of three polydimethylsiloxane layers, consisting of a top hole-punched layer, middle thin membrane, and bottom patterned layer. It is the first time that such a simple device has been used to supply axisymmetric and nonuniform strains to cells cultured on well bottoms that are stretchable. These mechanical stimuli can somewhat mimic the stretching at the bending sites of blood vessels, where the strains are complicated. In this device, nonuniform strain is given to cells through the deformation of a membrane from a flat surface to a spherical cap during the injection of a certain volume of water into the chamber between the middle membrane and bottom layer. EA.hy926 cells (a human umbilical vein endothelial cell line) were seeded on the well bottoms and exposed to axisymmetric strain under a 5, 10, 15, and 20% degree of deformation of the membrane. The cellular responses were characterized in terms of cell morphology, cell viability, and expression of inflammatory mRNAs and proteins. With increasing the degree of deformation, the cells exhibited an inclination toward detachment and apoptosis; meanwhile the expression of inflammatory mRNAs and proteins, such as MCP-1, IL-8, IL-6 and ICAM-1, showed a significant increment. The obtained results demonstrate that the inflammatory responses of EA.hy926 cells can be induced by increasing the magnitude of the strain. This simple device provides a useful tool for in vitro investigation of the inflammatory mechanisms related to vascular diseases.