Characterization of 3D Printed Stretching Devices for Imaging Force Transmission in Live-Cells
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Cell stretch is a method which can rapidly apply mechanical force through cell-matrix and cell-cell adhesions and can be utilized to better understand underlying biophysical questions related to intracellular force transmission and mechanotransduction.
3D printable stretching devices suitable for live-cell fluorescent imaging were designed using finite element modeling and validated experimentally. These devices were then used along with FRET based nesprin-2G force sensitive biosensors as well as live cell fluorescent staining to understand how the nucleus responds to externally applied mechanical force in cells with both intact LINC (linker of nucleoskeleton and cytoskeleton) complex and cells with the LINC complex disrupted using expression of dominant negative KASH protein.
The devices were shown to provide a larger strain ranges (300% uniaxial and 60% biaxial) than currently available commercial or academic designs we are aware of. Under uniaxial deformation, the deformation of the nucleus of NIH 3T3 cells per unit of imposed cell strain was shown to be approximately 50% higher in control cells compared to cells with a disrupted LINC complex. Under biaxial deformation, MDCK II cells showed permanent changes in the nuclear morphology as well as actin organization upon unloading, indicating that failure, plastic deformation, or remodeling of the cytoskeleton is occurring in response to the applied stretch.
Development and open distribution of low-cost, 3D-printable uniaxial and biaxial cell stretching devices compatible with live-cell fluorescent imaging allows a wider range of researchers to investigate mechanical influences on biological questions with only a minimal investment of resources.
KeywordsCell Stretch Cell mechanics Nuclear mechanics Nuclear LINC complex
We wish to thank Teemu Ihalainen for providing reagents. We acknowledge support from NIH R35GM119617 and R03AR068096, NSF CMMI-1653299, and the Thomas F. and Kate Miller Jeffress Memorial Trust. Microscopy was performed at the VCU Nano Characterization Core Facility.
CM and PA wrote the manuscript. CM, PA, KB and KD performed the experiments. CM designed the biaxial stretcher, PA and KB designed the uniaxial stretcher. DC led the project and provided input to the manuscript. All authors helped edit and revise the manuscript.
Any datasets generated during and/or analyzed during the study are available from the corresponding author on reasonable request.
Conflict of interest
Carl R. Mayer, Paul T. Arsenovic, Kranthidhar Bathula, Kevin B. Denis, and Daniel E. Conway declare that they have no conflicts of interest.
No human studies were carried out by the authors for this article. No animal studies were carried out by the authors for this article.
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