Influence of micro-scale substrate curvature on subcellular behaviors of vascular cells

Physiological characteristics of cells are affected by the extra-cellular matrix (ECM). Previous research have demonstrated that both biophysical and biochemical properties of ECM can determine cell behaviors such as cell spreading and morphology. Current studies on the biophysical factors have been largely related to the substrate rigidity and nano-roughness, yet how the three-dimensional (3D) micro-curvature affects cell behaviors remains elusive. In this work, we apply microfabrication and micro-contact printing technologies to develop arrays of ECM-coated micro-wells and micro-grooves with different substrate radii of curvature as a high-throughput, generic assay for characterizing different cell properties. The results show that the single third-dimension curvature factor could limit cell spreading and induce different cell morphologies compared with those on planar surface. Importantly, we discover for the first time that a part of vascular smooth muscle cells (VSMCs) stretch across the substrates (as the 'freestanding' behavior) while all endothelial cells conform to the substrate curvature. By immunofluorescence staining, we revealed that the freestanding behavior in curved substrates is related with actomyosin. It has been known that cell behavior is the reflection of cell response to ECM via various ways including mechanotrasduction. The further study the influence of micro-curvature on subcellular cytoskeletons, for instance the contractile system (actomyosin), will help explore their roles in cell communications with ECM, especially the freestanding behavior, and provide important insights for successful 3D culture and the 3D mechanotransductive responses.