Reprogramming cellular phenotype by soft collagen gels

Abstract

A variety of cell types exhibit phenotype changes in response to the mechanical stiffness of the substrate. Many cells excluding neurons display increase in spread area, actin stress fiber formation and larger focal adhesion complexes as substrate stiffness increases in sparsely populated culture. Cell proliferation is also known to directly correlate with these phenotype changes/change in substrate stiffness. Augmented spreading and proliferation on stiffer substrates require nuclear transcriptional regulator YAP (Yes associated protein) localization in cell nucleus and is tightly coupled with larger traction force generation. In this study, we show that different types of fibroblasts can exhibit spread morphology, well defined actin stress fibers, and larger focal adhesion seven on very soft collagen gels (modulus in hundreds of Pascals)as if they are on hard glass substrate (modulus in GPa, several orders of magnitude higher). Strikingly, we show, for the first time, that augmented spreading and other hard substrate cytoskeleton architecture on soft collagen gels are not correlated with cell proliferation pattern and do not require YAP localization in cell nucleus. Finally, we examine the response of human colon carcinoma (HCT-8) cells on soft collagen gels. Recent studies show that human colon carcinoma (HCT-8) cells form multi cellular clusters by 2–3 days when cultured on soft polyacrylamide (PA) gels with a wide range of stiffness (0.5 – 50 kPa) and coated with extracellular matrix, ECM (collagen monomer/ fibronectin). These clusters show limited spreading/wetting on PA gels, form 3D structures at the edges, and eventually display a remarkable, dissociative metastasis like phenotype (MLP), i.e., epithelial to rounded morphological transition after a week of culture on PA gels only, but not on collagen monomer coated stiff polystyrene/glass where they exhibit enhanced wetting and form confluent monolayer. Here, we show that HCT-8 cell clusters also show augmented spreading/wetting on soft collagen gels and eventually form confluent monolayer as on rigid glass substrates and MLP is completely inhibited on soft collagen gels. Overall, these results suggest that cell-material interaction (soft collagen gels in this case) can induce cellular phenotype and cytoskeleton organization in a remarkably distinct manner compared to a classical synthetic polyacrylamide (PA) hydrogel cell culture model and may contribute in designing new functional biomaterials.

Fiber Network Models Predict Enhanced Cell Mechanosensing on Fibrous Gels

“…The results from this study support the growing body of experimental and theoretical work that indicates that the fibrous nature of biological gels is responsible for enhanced long-distance cell mechanosensing compared to nonfibrous gels [14][15][16][17]30,[46][47][48]. Several of these theoretical studies have found that the mechanosensing length scale of a cell increases when fibers are accounted for in the model [14,30,46,48].…”

“…Much of this character derives from the fibrous nature of the ECM and the fact that these fibers are often connected into networks that organize into hierarchical structures at each scale of the tissue. Experiments on fibrous substrates, such as fibrin and collagen, have demonstrated that the cellular response is quite different than for a nonfibrous substrate of comparable bulk stiffness [14][15][16][17].…”

Extraction and Characterization of Collagen from Buffalo Skin for Biomedical Applications

“…It is most frequent used as hard and soft capsules, microspheres, sealants for vascular prostheses, wound dressing, drug delivery agent and three-dimensional tissue scaffold in tissue engineering 11,12,14,15 Collagen is used in tissue engineering as scaffolds in three forms. The first form is the collagen gel system 16,17 . The second form of collagen scaffold is the nanofiber sheets 18,19 .…”

A Multi-well Format Polyacrylamide-based Assay for Studying the Effect of Extracellular Matrix Stiffness on the Bacterial Infection of Adherent Cells

“…The stiffness of the ECM of cells is increasingly appreciated as an important modulator of cellular motility, differentiation, proliferation, and ultimately cell fate474849. Although there have been many recent advances in understanding the complex biomechanical interaction between cells and their ECM, little is known about how environmental stiffness affects the susceptibility of cells to bacterial infection.…”