Cytoplasmic Flow and Mixing Due to Deformation of Motile Cells.

The cytoplasm of a living cell is a dynamic environment through which intracellular components must move and mix. In motile, rapidly deforming cells such as human neutrophils, bulk cytoplasmic flow couples cell deformation to the transport and dispersion of cytoplasmic particles. Using particle-tracking measurements in live neutrophil-like cells, we demonstrate that fluid flow associated with the cell deformation contributes to the motion of small acidic organelles, dominating over diffusion on timescales above a few seconds. We then use a general physical model of particle dispersion in a deforming fluid domain to show that transport of organelle-sized particles between the cell periphery and the bulk can be enhanced by dynamic deformation comparable to that observed in neutrophils. Our results implicate an important mechanism contributing to organelle transport in these motile cells: cytoplasmic flow driven by cell shape deformation

A First Exposure to Statistical Mechanics for Life Scientists

Statistical mechanics is one of the most powerful and elegant tools in the quantitative sciences. One key virtue of statistical mechanics is that it is designed to examine large systems with many interacting degrees of freedom, providing a clue that it might have some bearing on the analysis of the molecules of living matter. As a result of data on biological systems becoming increasingly quantitative, there is a concomitant demand that the models set forth to describe biological systems be themselves quantitative. We describe how statistical mechanics is part of the quantitative toolkit that is needed to respond to such data. The power of statistical mechanics is not limited to traditional physical and chemical problems and there are a host of interesting ways in which these ideas can be applied in biology. This article reports on our efforts to teach statistical mechanics to life science students and provides a framework for others interested in bringing these tools to a nontraditional audience in the life sciences.Comment: 27 pages, 16 figures. Submitted to American Journal of Physic

Listeria monocytogenes cell-to-cell spread in epithelia is heterogeneous and dominated by rare pioneer bacteria.

Listeria monocytogenes hijacks host actin to promote its intracellular motility and intercellular spread. While L. monocytogenes virulence hinges on cell-to-cell spread, little is known about the dynamics of bacterial spread in epithelia at a population level. Here, we use live microscopy and statistical modeling to demonstrate that L. monocytogenes cell-to-cell spread proceeds anisotropically in an epithelial monolayer in culture. We show that boundaries of infection foci are irregular and dominated by rare pioneer bacteria that spread farther than the rest. We extend our quantitative model for bacterial spread to show that heterogeneous spreading behavior can improve the chances of creating a persistent L. monocytogenes infection in an actively extruding epithelium. Thus, our results indicate that L. monocytogenes cell-to-cell spread is heterogeneous, and that rare pioneer bacteria determine the frontier of infection foci and may promote bacterial infection persistence in dynamic epithelia. Editorial note:This article has been through an editorial process in which the authors decide how to respond to the issues raised during peer review. The Reviewing Editor's assessment is that all the issues have been addressed (see decision letter)

Response and Discrimination Performance of Arrays of Organothiol-Capped Au Nanoparticle Chemiresistive Vapor Sensors

The response and discrimination performance of an array that consisted of 20 different organothiol-capped Au nanoparticle chemiresistive vapor sensors was evaluated during exposure to 13 different organic vapors. The passivating organothiol ligand library consisted of collections of straight-chain alkanethiols, branched alkanethiols, and aromatic thiols. A fourth collection of sensors was formed from composites of 2-phenylethanethiol-capped Au nanoparticles and nonpolymeric aromatic materials that were coembedded in a sensor film. The organic vapors consisted of six hydrocarbons (n-hexane, n-heptane, n-octane, isooctane, cyclohexane, and toluene), three polar aprotic vapors (chloroform, tetrahydrofuran, and ethyl acetate), and four alcohols (methanol, ethanol, isopropanol, and 1-butanol). Trends in the resistance response of the sensors were consistent with expected trends in sorption due to the properties of the test vapor and the molecular structure of the passivating ligands in the sensor films. Classification algorithms including principal components analysis and Fisher’s linear discriminant were used to evaluate the discrimination performance of an array of such sensors. Each collection of sensors produced accurate classification of most vapors, with misclassification occurring primarily for vapors that had mutually similar polarity. The classification performance for an array that contained all of the sensor collections produced nearly perfect discrimination for all vapors studied. The dependence of the array size (i.e., the number of sensors) and the array chemical diversity on the discrimination performance indicated that, for an array of 20 sensors, an array size of 13 sensors or more produced the maximum discrimination performance

Perylene as an Organic Photocatalyst for the Radical Polymerization of Functionalized Vinyl Monomers through Oxidative Quenching with Alkyl Bromides and Visible Light

The generation of carbon-centered radicals from alkyl bromides through an oxidative quenching pathway using perylene as an organic visible-light photocatalyst is described. This methodology is used to initiate the radical polymerization of methyl methacrylate and other functionalized vinyl monomers. The polymers possess bromide chain-end groups that can be used to reinitiate polymerization to produce block copolymers. Control over the polymerization propagation can be achieved through pulsed light sequences while the ability to use natural sunlight to promote carbon–carbon bond formation produces polymers with dispersity as low as 1.29

Coupling of actin hydrolysis and polymerization: Reduced description with two nucleotide states

The polymerization of actin filaments is coupled to the hydrolysis of adenosine triphosphate (ATP), which involves both the cleavage of ATP and the release of inorganic phosphate. We describe hydrolysis by a reduced two-state model with a cooperative cleavage mechanism, where the cleavage rate depends on the state of the neighboring actin protomer in a filament. We obtain theoretical predictions of experimentally accessible steady state quantities such as the size of the ATP-actin cap, the size distribution of ATP-actin islands, and the cleavage flux for cooperative cleavage mechanisms.Comment: 6 page

Hindlimb Suspension as a Model to Study Ophthalmic Complications in Microgravity Status Report: Optimization of Rat Retina Flat Mounts Staining to Study Vascular Remodeling

Preliminary data from a prior tissuesharing experiment has suggested that early growth response protein1 (Egr1), a transcription factor involved in various stress responses in the vasculature, is induced in the rat retina after 14 days of hindlimb suspension (HS) and may be evidence that mechanical stress is occurring secondary to the cephalad fluid shift. This mechanical stress could cause changes in oxygenation of the retina, and the subsequent ischemia or inflammationdriven hypoxia may lead to microvascular remodeling. This microvascular remodeling process can be studied using image analysis of retinal vessels and can be then be quantified by the VESsel GENeration Analysis (VESGEN) software, a computational tool that quantifies remodeling patterns of branching vascular trees and capillary or vasculogenic networks. Our project investigates whether rodent HS is a valid model to study the effects of simulatedweightlessness on ocular structures and their relationship with intracranial pressure (ICP). One of the hypotheses to be tested is that HSinduced cephalad fluid shift is accompanied by vascular engorgement that produces changes in retinal oxygenation, leading to oxidative stress, hypoxia, microvascular remodeling, and cellular degeneration. We have optimized the procedure to obtain flat mounts of rat retina, staining of the endothelial lining in vasculature and acquisition of high quality images suitable for VESGEN analysis. Briefly, eyes were fixed in 4% paraformaldehyde for 24 hours and retinas were detached and then mounted flat on microscope slides. The microvascular staining was done with endothelial cellspecific isolectin binding, coupled to Alexa488 fluorophore. Image acquisition at low magnification and high resolution was performed using a new Leica SP8 confocal microscope in a tile pattern across the X,Y plane and multiple sections along the Zaxis. This new confocal microscope has the added capability of dye separation using the Linear Unmixing method and allows us to remove the autofluorescence originating from the photoreceptor layer. In summary, we have an improved method for studying the retinal microvasculature that will provide an increase in the quality of images captured and will be applied throughout the various animal cohorts of the recentlyinitiated study that will evaluate rodent HS as a model to study ophthalmic complications in microgravity

Effects of Dietary Iron and Gamma Radiation on the Rat Retina

A health risk of concern for NASA relates to radiation exposure and its synergistic effects with other space environmental factors, includi ng nutritional status of the crew. Astronauts consume almost three times the recommended daily allowance of iron due to the use of fortifie d foods aboard the International Space Station, with iron intake occa sionally exceeding six times the recommended values. Recently, NASA has become concerned with visual changes associated with spaceflight, a nd research is being conducted to elucidate the etiology of eye structure alterations in the spaceflight environment. Terrestrially, iron o verload is also associated with certain optic neuropathies. In additi on, due to its role in Fenton reactions, iron can potentiate oxidative stress, which is a recognized cause of cataract formation. As part o f a study investigating the combined effects of radiation exposure an d iron overload on multiple physiological systems, we focused on defining the effects of both treatments on eye biology. In this study, 12- week-old Sprague-Dawley rats were assigned to one of four experimental groups: normal iron/no radiation (Control/Sham), high iron/no radiat ion (Fe/Sham), normal iron/gamma radiation (3 Gy cumulative dose, fra ctionated at 0.375 Gy/d every other day for 16 d) (Control/Rad), and high iron/gamma radiation (Fe/Rad). Oxidative stress-induced DNA damag e, measured as concentration of the marker 8-hydroxy-2'-deoxyguanosine (8OHdG) in eye retinal tissue by enzyme-immunoanalysis did not show significant changes among treatments. However, there was an overall i ncrease in 8OHdG immunostaining density in retina sections due to radiation exposure (P = 0.05). Increased dietary iron and radiation expos ure had an interactive effect (P = 0.02) on 8OHdG immunostaining of t he retinal ganglion cell layer with iron diet increasing the signal in the group not exposed to radiation (P = 0.05). qPCR gene expression profiling of relevant target genes indicated upregulation of ferritin light chain (P = 0.09) as a result of dietary iron but no change in e xpression of the gene for ferritin heavy chain. Immunolocalization of light chain and heavy chain of the iron storage protein ferritin showed the expected distribution in the choroid, photoreceptor layer, inn er nuclear layer and in the inner plexiform layer that corresponded t o the synaptic terminals of bipolar cells. Evidence of stress and damage in the retina was also suggested by a decrease in expression of th e survival marker Bcl2 (P = 0.01) and the protective proteins clusterin (P = 0.04) and heat shock factor 1 (Hsf1, P < 0.001), as a result o f increased dietary iron. The effect of increased iron on expression of the antioxidant enzyme heme oxygenase 1 (Hmox1) had a significant interaction with the effect of radiation (P < 0.001). In summary, the results of this study indicate that both gamma radiation exposure and a moderate increase in dietary iron can contribute to deleterious cha nges in retinal health and physiology