Nano-Stenciled RGD-Gold Patterns That Inhibit Focal Contact Maturation Induce Lamellipodia Formation in Fibroblasts

Abstract

Cultured fibroblasts adhere to extracellular substrates by means of cell-matrix adhesions that are assembled in a hierarchical way, thereby gaining in protein complexity and size. Here we asked how restricting the size of cell-matrix adhesions affects cell morphology and behavior. Using a nanostencil technique, culture substrates were patterned with gold squares of a width and spacing between 250 nm and 2 µm. The gold was functionalized with RGD peptide as ligand for cellular integrins, and mouse embryo fibroblasts were plated. Limiting the length of cell-matrix adhesions to 500 nm or less disturbed the maturation of vinculin-positive focal complexes into focal contacts and fibrillar adhesions, as indicated by poor recruitment of α5-integrin. We found that on sub-micrometer patterns, fibroblasts spread extensively, but did not polarize. Instead, they formed excessive numbers of lamellipodia and a fine actin meshwork without stress fibers. Moreover, these cells showed aberrant fibronectin fibrillogenesis, and their speed of directed migration was reduced significantly compared to fibroblasts on 2 µm square patterns. Interference with RhoA/ROCK signaling eliminated the pattern-dependent differences in cell morphology. Our results indicate that manipulating the maturation of cell-matrix adhesions by nanopatterned surfaces allows to influence morphology, actin dynamics, migration and ECM assembly of adhering fibroblasts.

Reversible Holographic Patterns on Azopolymers for Guiding Cell Adhesion and Orientation

“…33 Topographic patterns imprinted on pDR1m proved to be effective in controlling different aspects of the cell-material interactions and macroscopic cell behavior. More interestingly, though, surface modifications induced on azopolymers are, in principle, reversible, i.e.…”

Micrometer scale spacings between fibronectin nanodots regulate cell morphology and focal adhesions

“…There have been important cell adhesion studies using both micro and nanopatterned surfaces [10][11][12][13][14][15][16][17][18][19][20][21][22][23]. The pioneering contributions of Mirkin and Spatz are particularly noteworthy [24,25].…”

Nano‐spaced Gold on Glassy Carbon Substrate for Controlling Cell Behavior

“…Phalloidin was used to stain the F‐actin filaments of the cytoskeleton which are responsible for the interaction between the focal points for cell attachment, [ 89 ] whereas Hoechst is used for specifically staining the cell nuclei ( Figure ). In the case of GC (control and FN) the cells presented a round morphology which is typical of non‐attached cells.…”