A Microfluidic Hydrogel Chip with Orthogonal Dual Gradients of Matrix Stiffness and Oxygen for Cytotoxicity Test

The oxygen tensions and matrix stiffness play important roles in regulating cell response to cytotoxic drugs. In recent years, single spatiotemporal oxygen or matrix stiffness gradient has been established by various technologies for cell studies. However, the synergistic effects of the two factors on tumor cells remained elusive. In this study, we created a highly integrated and easy-to-operate microfluidic device. It was capable of generating more continuous, linear, stable and diffusive hydrogel stiffness gradient over a well-defined oxygen gradient. Sodium hydroxide and pyrogallol were used to scavenge oxygen and generate oxygen gradient, skillfully avoiding the utilization of bulky pressurized gas cylinders and sophisticated flow control by conventional methods. Utilizing the newly developed microfluidic device, we successfully performed drug test with a hypoxia sensitive anti-cancer drug, triapazamine (TPZ), on A549 cells under perpendicular construction of oxygen and spatially hydrogel stiffness gradients. The cell experiment results demonstrated the matrix stiffness-dependent cell drug resistance and hypoxia-induced cytotoxicity of TPZ. In summary, the developed microfluidic device exhibits high potential in the study of matrix stiffness-dependent cell responses and oxygen-sensitive drug cytotoxicity. Furthermore, the design principle can be functionally extended to other dual factors system and will facilitate the study of cell response in complex physiological and pathological conditions.