control-cell-geometry-through-infrared-laser-assisted

Control of Cell Geometry through Infrared Laser Assisted Micropatterning

Micropatterning is an established technique in the cell biology community used to study connections between the morphology and function of cellular ...

University of Toronto

Cells in the human body experience and integrate a wide variety of environmental cues. A growing interest in tissue mechanics in the past four decades has shown that the mechanical properties of tissue drive key biological processes and facilitate disease development. However, tissue stiffness is not only a potent behavioral cue, but also a product of cellular signaling activity. This review explores both roles of tissue stiffness in the context of inflammation and fibrosis, and the important molecular players driving such processes. During inflammation, proinflammatory cytokines upregulate tissue stiffness by increasing hydrostatic pressure, ECM deposition, and ECM remodeling. As the ECM stiffens, cells involved in the immune response employ intricate molecular sensors to probe and alter their mechanical environment, thereby facilitating immune cell recruitment and potentiating the fibrotic phenotype. This powerful feedforward loop raises numerous possibilities for drug development and warrants further investigation into the mechanisms specific to different fibrotic diseases.

Mechanosensitive Regulation of Fibrosis.

Myofibroblasts play important roles in physiological processes such as wound healing and tissue repair. While high contractile forces generated by the actomyosin network enable myofibroblasts to physically contract the wound and bring together injured tissue, prolonged and elevated levels of contraction also drive the progression of fibrosis and cancer. However, quantitative mapping of these forces has been difficult due to their extremely low magnitude ranging from 100 pN/μm to 2 nN/μm. Here, we provide a protocol to measure cellular forces exerted on two-dimensional compliant elastic hydrogels. We describe the fabrication of polyacrylamide hydrogels labeled with fluorescent fiducial markers, functionalization of substrates with ECM proteins, setting up the experiment, and imaging procedures. We demonstrate the application of this technique for quantitative analysis of traction forces exerted by myofibroblasts.

Shuying Yang

Department of Cell and Systems Biology

University of Toronto

Control of Cell Geometry through Infrared Laser Assisted Micropatterning