Directing Left-Right Asymmetry in Tissue Formation via Micropatterned Substrate with Varying Stiffness

Description

Tissue engineering aims at using cell-based therapy to rebuild missing or damagedtissue. Many physiological functions, such as heart contraction, requires tissue alignmentwith specific left-right (LR) asymmetry. Recent evidences indicated that LR asymmetryshould be originated from single cell’s LR biased motion, e.g. rotation or migration.However, how to guide LR asymmetry from single cells to tissue formation in apredictive manner is still a major challenge. If it is uncontrolled, such intrinsic cellbehavior may lead to disorganized patterns and eventually malfunction the tissueconstruct.In this proposal, we aim at exploring an engineering strategy to guide the LR asymmetryin a type of tissue formation, skeletal muscle myogenesis. Recently we found thatmicropatterned substrate with decreased stiffness could reverse the LR polarity of cellslocated at the micropatterned interface. Importantly, based on our previous findings, theLR polarity may propagate from cells at the interface to cells remote from the interface,eventually leading to a coherent LR biased cell-cell alignment throughout the entiremulticellular structure. Taking together, it suggests that micropatterned substrate withvarying stiffness may be an approach, for the first time, to guide the LR asymmetry notonly in single cells but also the associated tissue formation. To explore this possibility,we will hierarchically investigate how substrate stiffness can mediate the LR bias from1) single cell’s torque to 2) its corresponding cell-cell alignment for 3) applications ofguiding LR development in skeletal muscle myogenesis. First, we will apply the nanowiremagnetoscope, our newly developed platform, to measure cellular torque representingsingle cell’s LR bias in response to substrate stiffness. Second, we will investigate howsubstrate stiffness can influence the propagation of LR bias using time-lapse nucleusrotation that automatically measures the long axis of stained nucleus in live cells toreveal the dynamics of cell-cell alignment. At last, we will apply the knowledge gainedto a model of tissue formation, skeletal muscle myogenesis. Using C2C12 myoblasts, wewill explore whether the changed substrate stiffness would switch the LR orientation ofdifferentiated myotube, eventually controlling the LR direction of muscle orientation andcontraction. Together, this integrated analysis on substrate stiffness will provide avaluable understanding of how LR asymmetry is formed from single cell to multicellularstructure, opening up a new direction for controlling LR asymmetry in tissue formationin the future.