Trans-disciplinary research approaches: integration of fluid mechanics with cell biology

At the level of the individual molecule, outstanding discoveries leading the the 2012 Nobel Prize in Chemistry clarified G-protein coupled receptors (GPCRs) crystallographic structures and functions. Nevertheless, a large number of such receptors cooperate to ultimately determine the cell response. Henceforth, understanding their group effects becomes crucial to predict the activity of the GPCRs populating the various cell aggregates

Minimum Technical Data Elements for Liquid Biopsy Data Submitted to Public Databases

Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/154656/1/cpt1747.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/154656/2/cpt1747-sup-0001-FigS1.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/154656/3/cpt1747_am.pd

Cdc42 Mediates Nucleus Movement and MTOC Polarization in Swiss 3T3 Fibroblasts under Mechanical Shear Stress

Nucleus movement is essential during nucleus positioning for tissue growth and development in eukaryotic cells. However, molecular regulators of nucleus movement in interphase fibroblasts have yet to be identified. Here, we report that nuclei of Swiss 3T3 fibroblasts undergo enhanced movement when subjected to shear flows. Such movement includes both rotation and translocation and is dependent on microtubule, not F-actin, structure. Through inactivation of Rho GTPases, well-known mediators of cytoskeleton reorganization, we demonstrate that Cdc42, not RhoA or Rac1, controls the extent of nucleus translocation, and more importantly, of nucleus rotation in the cytoplasm. In addition to generating nuclei movement, we find that shear flows also causes repositioning of the MTOC in the direction of flow. This behavior is also controlled by Cdc42 via the Par6/protein kinase Cζ pathway. These results are the first to establish Cdc42 as a molecular regulator of not only shear-induced MTOC polarization in Swiss 3T3 fibroblasts, but also of shear-induced microtubule-dependent nucleus movement. We propose that the movements of MTOC and nucleus are coupled chemically, because they are both regulated by Cdc42 and dependent on microtubule structure, and physically, possibly via Hook/SUN family homologues similar to those found in Caenorhabditis elegans

Nuclear Lamin A/C Deficiency Induces Defects in Cell Mechanics, Polarization, and Migration

Lamin A/C is a major constituent of the nuclear lamina, a thin filamentous protein layer that lies beneath the nuclear envelope. Here we show that lamin A/C deficiency in mouse embryonic fibroblasts (Lmna−/− MEFs) diminishes the ability of these cells to polarize at the edge of a wound and significantly reduces cell migration speed into the wound. Moreover, lamin A/C deficiency induces significant separation of the microtubule organizing center (MTOC) from the nuclear envelope. Investigations using ballistic intracellular nanorheology reveal that lamin A/C deficiency also dramatically affects the micromechanical properties of the cytoplasm. Both the elasticity (stretchiness) and the viscosity (propensity of a material to flow) of the cytoplasm in Lmna−/− MEFs are significantly reduced. Disassembly of either the actin filament or microtubule networks in Lmna+/+ MEFs results in decrease of cytoplasmic elasticity and viscosity down to levels found in Lmna−/− MEFs. Together these results show that both the mechanical properties of the cytoskeleton and cytoskeleton-based processes, including cell motility, coupled MTOC and nucleus dynamics, and cell polarization, depend critically on the integrity of the nuclear lamina, which suggest the existence of a functional mechanical connection between the nucleus and the cytoskeleton. These results also suggest that cell polarization during cell migration requires tight mechanical coupling between MTOC and nucleus, which is mediated by lamin A/C

Microrheology and ROCK Signaling of Human Endothelial Cells Embedded in a 3D Matrix

Cell function is profoundly affected by the geometry of the extracellular environment confining the cell. Whether and how cells plated on a two-dimensional matrix or embedded in a three-dimensional (3D) matrix mechanically sense the dimensionality of their environment is mostly unknown, partly because individual cells in an extended matrix are inaccessible to conventional cell-mechanics probes. Here we develop a functional assay based on multiple particle tracking microrheology coupled with ballistic injection of nanoparticles to measure the local intracellular micromechanical properties of individual cells embedded inside a matrix. With our novel assay, we probe the mechanical properties of the cytoplasm of individual human umbilical vein endothelial cells (HUVECs) embedded in a 3D peptide hydrogel in the presence or absence of vascular endothelial growth factor (VEGF). We found that VEGF treatment, which enhances endothelial migration, increases the compliance and reduces the elasticity of the cytoplasm of HUVECs in a matrix. This VEGF-induced softening response of the cytoplasm is abrogated by specific Rho-kinase (ROCK) inhibition. These results establish combined particle-tracking microrheology and ballistic injection as the first method able to probe the micromechanical properties and mechanical response to agonists and/or drug treatments of individual cells inside a matrix. These results suggest that ROCK plays an essential role in the regulation of the intracellular mechanical response to VEGF of endothelial cells in a 3D matrix

Cohort profile: Growing up in Singapore towards healthy outcomes (GUSTO) birth cohort study

10.1093/ije/dyt125International Journal of Epidemiology4351401 - 1409IJEPBGUSTO (Growing up towards Healthy Outcomes