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Stiffness-controlled three-dimensional extracellular matrices for high-resolution imaging of cell behavior
Robert S Fischer1, Kenneth A Myers, Margaret L Gardel
1Cell Biology and Physiology Center, National Heart, Lung, and Blood Institute (NHBLI), US National Institutes of Health (NIH), Bethesda, Maryland, USA. fischerr2@nhlbi.nih.gov
Nature Protocols
|October 27, 2012
Summary
Researchers developed a new method to study how extracellular matrix (ECM) stiffness and dimensionality affect cell behavior. This technique allows for precise control over physical properties in three-dimensional (3D) environments.
Area of Science:
- Biomedical Engineering
- Cell Biology
- Biophysics
Background:
- Cell functions are regulated by the physical properties of the extracellular matrix (ECM).
- ECM stiffness and dimensionality critically influence cell signaling and behavior.
- Disentangling the specific roles of ECM stiffness and dimensionality has been challenging due to their interconnectedness with other factors like ligand density.
Purpose of the Study:
- To develop a straightforward protocol for investigating the independent effects of ECM stiffness and dimensionality on cell function.
- To create a system that allows for the study of cell behavior in physiologically relevant three-dimensional (3D) ECM environments with controlled mechanical properties.
Main Methods:
- A novel 'sandwich gel' system was designed, integrating cells within a 3D fibrillar ECM.
- This 3D ECM was coupled to an underlying polyacrylamide gel with precisely controlled stiffness (compliance).
- The protocol is rapid, completable within 1-2 days, and facilitates high-resolution time-lapse imaging.
Main Results:
- The 'sandwich gel' system successfully isolates and controls ECM stiffness in a 3D context.
- This method enables the study of how defined ECM compliance specifically impacts cell signaling and function within 3D matrices.
- The system is adaptable for various cell types and molecular perturbations.
Conclusions:
- The developed 'sandwich gel' protocol provides a powerful tool for dissecting the specific contributions of ECM stiffness to cell behavior in 3D.
- This technique overcomes previous limitations in isolating ECM physical properties.
- It offers a versatile platform for advancing our understanding of mechanobiology in development and disease.
