Replicating the hemodynamic signal of endothelial cells (ECs) induced by an external counterpulsation (ECP) using a microfluidic system is crucial for further studying EC mechanobiology associated with ECP theraphy. However, there is currently no suitable method to construct a microfluidic system that can accurately reproduce the hemodynamic signals including blood pressure (BP) and wall shear stress (WSS) on the inner surface of the common carotid artery following ECP intervention. In this study, we propose a model-based approach to build a microfluidic feedback control system (MFCS), which consists of a microfluidic chip and a pump system (PS) to resolve the above issue. An integrated transfer function (TF) model formed by the expression of the input impedance and the proportion integration differentiation (PID) controller is used to design the microfluidic chip and PS. The performance of the proposed MFCS for replicating the hemodynamic signals is evaluated by flow-field analysis, and calcium ion (Ca2+) response experiments are conducted to validate the functionality of the proposed MFCS for investigating mechanobiological mechanisms.