Nano-patterned oxidized porous silicon (PS) substrates are topographically biocompatible to provide in vitro environment for improved understanding of in vivo wound healing behavior. But the wound healing studies reported on various featured substrates are mainly based on time lapse microscopy coupled with fluorescent labeling. This study is the first attempt to perform continuous monitoring of wound healing phenomena using PS substrates with two pore morphologies by means of impedance measurement. A distributed electrical model has been developed for the PS substrates to extract the cell migration rate, proliferation rate, and time-dependent coverage of cell free area. It has been observed from impedance measurements and from model that the wound closure is faster on 50 nm pores followed by 500 nm pores and planar ECIS substrates. This has been qualitatively explained from the extracted time-dependent cell–cell and cell-substrate interaction parameters using a well-known mechanical model of wound healing process. The electrical readings have been closely correlated with scanning electron microscopy (SEM) and optical microscopy images. Thus, the impedance spectroscopy method coupled with the proposed theoretical framework can provide information about the underlying mechanisms of wound regeneration on nano-patterned substrates in a non-invasive manner.