Alignment of corneal and lens epithelial cells by co-operative effects of substratum topography and DC electric fields.

Corneal and lens epithelial cells (CECs and LECs) in the eye encounter precisely ordered fibre arrays on the nanoscale in tandem with an endogenous electric field (EF). Prosthetic biomaterials often incorporate topographical features intended to mimic those in situ. However, the cellular basis for control of cell morphology by nanotopography or by an EF is not clear. We examined cell axis alignment in response to substratum nanotopography and a physiological EF separately and in combination. Bovine CECs aligned parallel to substratum nanogrooves (NGs) as shallow as 14 nm but LECs were less sensitive. Actin filaments of both cell types concentrated at substratum ridges so we tested the mechanistic roles of rho, rac and cdc42, molecules that control cytoskeletal organization. CEC alignment to 130 nm deep NGs was prevented by the inhibition of rho, but not by the inhibition of cdc42, rac, or the rho effectors myosin light chain kinase or rho kinase. Conversely, CEC alignment was enhanced by the activation of rho. CECs on planar quartz substrata aligned orthogonal to an EF of 150 mV/mm. Alignment required signalling by cdc42 and rho but not rac, and was accompanied by lamellipodial reorganisation and cell migration toward the cathode. When CECs on vertically oriented NGs were exposed simultaneously to a horizontal EF, they aligned more robustly than to either cue alone and the enhanced alignment required rho signalling. Therefore, nanoscale substratum features and EFs co-operate to control cell axis alignment via rho, and cdc42-mediated intracellular signals, which can be exploited in tissue engineering.