Probing the Nanomechanical Behavior of Cells and Cell Nuclei

Atomic force microscopy (AFM) is a nanoscale characterization technique that at its most basic level employs a nanometer-scale probe tip to physically trace a surface, generating a topographical map of the sample. However AFM has many applications beyond topography, including nanomechanical property analysis via cantilevered nanoindentation. In this project, tipless AFM probes functionalized with a 10 µm diameter glass bead have been used to measure the elastic modulus of live multipotent stromal stem cell nuclei before and after vibration treatments and/or structural component knockouts. The goal of these nanoindentation measurements of nuclear stiffness is to gain a better understanding of how mesenchymal stem cells respond to mechanical (in addition to chemical) signals in their environment to guide differentiation into osteoblasts, chondrocytes, or other cell types

Nuclear Response to Low Intensity Vibrations

The nucleus, central to all cellular activity, relies on both direct mechanical input as well as its molecular transducers to sense external stimuli and respond by regulating intra-nuclear organization that ultimately determines gene expression to control cell function and fate. It is long studied that signals propagate from extracellular environment to cytoskeleton and into the nucleus (outside-in signaling) to regulate cell behavior. Emerging evidence, however, has shown that both the cytoskeleton and nucleus have an inherent ability to sense and adapt to mechanical force independent of each other. This suggests mechano-signaling and cytoskeleton remodeling events in response to exercise mimetics, like low intensity vibration (LIV), may directly be sensed at the nucleus (inside-out signaling). Here we hypothesize that cell nuclei will directly adapt to dynamic accelerations in response to LIV. To answer this question directly, we isolated live nuclei from cells to test their mechanical responses to LIV. Isolated nuclei are introduced to a low intensity vibration (LIV, 0.7g, 90Hz) and their stiffness’ measured via AFM (Atomic force microscope) protocol. Findings from this study will allow evaluation of the mechanical role that the nucleus plays in the cell as an individual organelle

Developing Nucleus Specific Finite Element Models Using Confocal Microscopy Scans

Emerging evidence suggests that nucleus have an inherent ability to adapt to mechanical force. Current approaches, however, are unable to quantify native forces generated in on cell nuclei without inserting sensors that affect cell function. Our goal in this research therefore is to use Finite Element Modeling in combination with confocal microscopy to generate mechanical models of nuclei. To build nuclear Finite element models chromatin of mesenchymal stem cell nuclei were imaged with a Zeiss 810 confocal microscope at 120nm planar resolution at every 360nm. These images were developed into hexahedral based models. To calibrate the model, isolated live cell nuclei stiffness were deduced using an Atomic force microscope (AFM). Establishing this process will enable the creation of nucleus specific models that allow further research into how the mechanical stiffness of a nucleus is regulated