Vivian C. Hecht

Characterizing deformability and surface friction of cancer cells

Metastasis requires the penetration of cancer cells through tight spaces, which is mediated by the physical properties of the cells as well as their interactions with the confined environment. Various microfluidic approaches have been devised to mimic traversal in vitro by measuring the time required for cells to pass through a constriction. Although a cell’s passage time is expected to depend on its deformability, measurements from existing approaches are confounded by a cells size and its frictional properties with the channel wall. Here, we introduce a device that enables the precise measurement of (i) the size of a single cell, given by its buoyant mass, (ii) the velocity of the cell entering a constricted microchannel (entry velocity), and (iii) the velocity of the cell as it transits through the constriction (transit velocity). Changing the deformability of the cell by perturbing its cytoskeleton primarily alters the entry velocity, whereas changing the surface friction by immobilizing positive charges on the constrictions walls primarily alters the transit velocity, indicating that these parameters can give insight into the factors affecting the passage of each cell. When accounting for cell buoyant mass, we find that cells possessing higher metastatic potential exhibit faster entry velocities than cells with lower metastatic potential. We additionally find that some cell types with higher metastatic potential exhibit greater than expected changes in transit velocities, suggesting that not only the increased deformability but reduced friction may be a factor in enabling invasive cancer cells to efficiently squeeze through tight spaces.

Intracellular Water Exchange for Measuring the Dry Mass, Water Mass and Changes in Chemical Composition of Living Cells

We present a method for direct non-optical quantification of dry mass, dry density and water mass of single living cells in suspension. Dry mass and dry density are obtained simultaneously by measuring a cell’s buoyant mass sequentially in an H2O-based fluid and a D2O-based fluid. Rapid exchange of intracellular H2O for D2O renders the cell’s water content neutrally buoyant in both measurements, and thus the paired measurements yield the mass and density of the cell’s dry material alone. Utilizing this same property of rapid water exchange, we also demonstrate the quantification of intracellular water mass. In a population of E. coli, we paired these measurements to estimate the percent dry weight by mass and volume. We then focused on cellular dry density – the average density of all cellular biomolecules, weighted by their relative abundances. Given that densities vary across biomolecule types (RNA, DNA, protein), we investigated whether we could detect changes in biomolecular composition in bacteria, fungi, and mammalian cells. In E. coli, and S. cerevisiae, dry density increases from stationary to exponential phase, consistent with previously known increases in the RNA/protein ratio from up-regulated ribosome production. For mammalian cells, changes in growth conditions cause substantial shifts in dry density, suggesting concurrent changes in the protein, nucleic acid and lipid content of the cell.

Deformability of Tumor Cells versus Blood Cells

The potential for circulating tumor cells (CTCs) to elucidate the process of cancer metastasis and inform clinical decision-making has made their isolation of great importance. However, CTCs are rare in the blood, and universal properties with which to identify them remain elusive. As technological advancements have made single-cell deformability measurements increasingly routine, the assessment of physical distinctions between tumor cells and blood cells may provide insight into the feasibility of deformability-based methods for identifying CTCs in patient blood. To this end, we present an initial study assessing deformability differences between tumor cells and blood cells, indicated by the length of time required for them to pass through a microfluidic constriction. Here, we demonstrate that deformability changes in tumor cells that have undergone phenotypic shifts are small compared to differences between tumor cell lines and blood cells. Additionally, in a syngeneic mouse tumor model, cells that are able to exit a tumor and enter circulation are not required to be more deformable than the cells that were first injected into the mouse. However, a limited study of metastatic prostate cancer patients provides evidence that some CTCs may be more mechanically similar to blood cells than to typical tumor cell lines.

Biophysical changes reduce energetic demand in growth factor–deprived lymphocytes

Changes to the biophysical properties of lymphocytes are identified as an adaptive response to acute nutrient stress that occurs before the induction of autophagy.

Characterizing Cellular Biophysical Responses to Stress by Relating Density, Deformability, and Size

Cellular physical properties are important indicators of specific cell states. Although changes in individual biophysical parameters, such as cell size, density, and deformability, during cellular processes have been investigated in great detail, relatively little is known about how they are related. Here, we use a suspended microchannel resonator (SMR) to measure single-cell density, volume, and passage time through a narrow constriction of populations of cells subjected to a variety of environmental stresses. Osmotic stress significantly affects density and volume, as previously shown. In contrast to density and volume, the effect of an osmotic challenge on passage time is relatively small. Deformability, as determined by comparing passage times for cells with similar volume, exhibits a strong dependence on osmolarity, indicating that passage time alone does not always provide a meaningful proxy for deformability. Finally, we find that protein synthesis inhibition, cell-cycle arrest, protein kinase inhibition, and cytoskeletal disruption result in unexpected relationships among deformability, density, and volume. Taken together, our results suggest that by measuring multiple biophysical parameters, one can detect unique characteristics that more specifically reflect cellular behaviors.

Abstract A36: Amino acids rather than glucose accounts for the majority of cell mass in rapidly proliferating mammalian cells

To facilitate growth and division, proliferating cells must duplicate their mass over the cell cycle. Glucose and glutamine are the most consumed nutrients by proliferating mammalian cells, but the extent to which these and other nutrients contribute directly to cell mass has not been investigated. Using isotope labeled nutrients, we quantified the fraction of cell mass derived from different fuels and find that surprisingly neither glucose nor glutamine provide the majority of carbon present in cells. Instead, amino acids, which are consumed at substantially lower rates, together account for the majority of cell mass. While glucose carbon has diverse fates, glutamine is overwhelmingly diverted into protein and its incorporation into cell mass greatly exceeds that of other amino acids, suggesting that glutamine-driven anapleurosis is primarily used for amino acid biosynthesis. Our findings demonstrate that the most heavily consumed nutrients do not make a proportionately large contribution of carbon to macromolecule biosynthesis. Citation Format: Aaron M. Hosios, Vivian C. Hecht, Marc Johnson, Jeffrey C. Rathmell, Scott R. Manalis, Matthew G. Vander Heiden. Amino acids rather than glucose accounts for the majority of cell mass in rapidly proliferating mammalian cells. [abstract]. In: Proceedings of the AACR Special Conference: Metabolism and Cancer; Jun 7-10, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(1_Suppl):Abstract nr A36.