Tissue regeneration at an injured site depends on proliferation, migration, and differentiation of resident stem or progenitor cells, but solid tissues are often sufficiently dense and constricting that nuclei are highly stressed by migration. In this study, constricted migration of myoblastic cell types and mesenchymal stem cells (MSCs) increases nuclear rupture, increases DNA damage, and modulates differentiation. Fewer myoblasts fuse into regenerating muscle in vivo after constricted migration in vitro, and myodifferentiation in vitro is likewise suppressed. Myosin II inhibition rescues rupture and DNA damage, implicating nuclear forces, while mitosis and the cell cycle are suppressed by constricted migration, consistent with a checkpoint. Although perturbed proliferation fails to explain defective differentiation, nuclear rupture mislocalizes differentiation-relevant MyoD and KU80 (a DNA repair factor), with nuclear entry of the DNA-binding factor cGAS. Human MSCs exhibit similar damage, but osteogenesis increases-which is relevant to bone and to calcified fibrotic tissues, including diseased muscle. Tissue repair can thus be modulated up or down by the curvature of pores through which stem cells squeeze.

Motor neuron (MN) diseases are progressive disorders resulting from degeneration of neuromuscular junctions (NMJs), which form the connection between MNs and muscle fibers. NMJ-in-a-dish models have been developed to examine human MN-associated dysfunction with disease; however such coculture models have randomly oriented myotubes with immature synapses that contract asynchronously. Mechanically patterned (MP) extracellular matrix with alternating soft and stiff stripes improves current NMJ-in-a-dish models by inducing both mouse and human myoblast durotaxis to stripes where they aligned, differentiated, and fused into patterned myotubes. Compared to conventional culture on rigid substrates or unpatterned hydrogels, MP substrates supported increased differentiation and fusion, significantly larger acetylcholine (ACh) receptor clusters, and increased expression of MuSK and Lrp4, two cell surface receptors required for NMJ formation. Robust contractions were observed when mouse myotubes were stimulated by ACh, with twitch duration and frequency most closely resembling those for mature muscle on MP substrates. Fused myotubes, when cocultured with MNs, were able to form even larger NMJs. Thus MP matrices produce more functionally active NMJs-in-a-dish, which could be used to elucidate disease pathology and facilitate drug discovery.

Matrix stiffness and dimensionality have been shown to be major determinants of cell behavior. However, a workflow for examining nanometer-scale responses of the associated molecular machinery is not available. Here, we describe a comprehensive, quantitative workflow that permits the analysis of cells responding to mechanical and dimensionality cues in their native state at nanometer scale by cryogenic electron tomography. Using this approach, we quantified distinct cytoskeletal nanoarchitectures and vesicle phenotypes induced in human mammary epithelial cells in response to stiffness and dimensionality of reconstituted basement membrane. Our workflow closely recapitulates the microenvironment associated with acinar morphogenesis and identified distinct differences in situ at nanometer scale. Using drug treatment, we showed that molecular events and nanometer-scale rearrangements triggered by engagement of apical cell receptors with reconstituted basement membrane correspond to changes induced by reduction of cortical tension. Our approach is fully adaptable to any kind of stiffness regime, extracellular matrix composition, and drug treatment.

Mechanical linkage between cell-cell and cell-extracellular matrix (ECM) adhesions regulates cell shape changes during embryonic development and tissue homoeostasis. We examined how the force balance between cell-cell and cell-ECM adhesions changes with cell spread area and aspect ratio in pairs of MDCK cells. We used ECM micropatterning to drive different cytoskeleton strain energy states and cell-generated traction forces and used a Förster resonance energy transfer tension biosensor to ask whether changes in forces across cell-cell junctions correlated with E-cadherin molecular tension. We found that continuous peripheral ECM adhesions resulted in increased cell-cell and cell-ECM forces with increasing spread area. In contrast, confining ECM adhesions to the distal ends of cell-cell pairs resulted in shorter junction lengths and constant cell-cell forces. Of interest, each cell within a cell pair generated higher strain energies than isolated single cells of the same spread area. Surprisingly, E-cadherin molecular tension remained constant regardless of changes in cell-cell forces and was evenly distributed along cell-cell junctions independent of cell spread area and total traction forces. Taken together, our results showed that cell pairs maintained constant E-cadherin molecular tension and regulated total forces relative to cell spread area and shape but independently of total focal adhesion area.

Health care personnel (HCP) caring for patients with coronavirus disease 2019 (COVID-19) might be at high risk for contracting SARS-CoV-2, the virus that causes COVID-19. Understanding the prevalence of and factors associated with SARS-CoV-2 infection among frontline HCP who care for COVID-19 patients are important for protecting both HCP and their patients. During April 3-June 19, 2020, serum specimens were collected from a convenience sample of frontline HCP who worked with COVID-19 patients at 13 geographically diverse academic medical centers in the United States, and specimens were tested for antibodies to SARS-CoV-2. Participants were asked about potential symptoms of COVID-19 experienced since February 1, 2020, previous testing for acute SARS-CoV-2 infection, and their use of personal protective equipment (PPE) in the past week. Among 3,248 participants, 194 (6.0%) had positive test results for SARS-CoV-2 antibodies. Seroprevalence by hospital ranged from 0.8% to 31.2% (median = 3.6%). Among the 194 seropositive participants, 56 (29%) reported no symptoms since February 1, 2020, 86 (44%) did not believe that they previously had COVID-19, and 133 (69%) did not report a previous COVID-19 diagnosis. Seroprevalence was lower among personnel who reported always wearing a face covering (defined in this study as a surgical mask, N95 respirator, or powered air purifying respirator [PAPR]) while caring for patients (5.6%), compared with that among those who did not (9.0%) (p = 0.012). Consistent with persons in the general population with SARS-CoV-2 infection, many frontline HCP with SARS-CoV-2 infection might be asymptomatic or minimally symptomatic during infection, and infection might be unrecognized. Enhanced screening, including frequent testing of frontline HCP, and universal use of face coverings in hospitals are two strategies that could reduce SARS-CoV-2 transmission.