Collective migration during a gap closure in a two-dimensional haptotactic model

The ability of cells to respond to substrate-bound protein gradients is crucial for many physiological processes, such as immune response, neurogenesis and cancer cell migration. However, the difficulty to produce well-controlled protein gradients has long been a limitation to our understanding of collective cell migration in response to haptotaxis. Here we use a photopatterning technique to create circular, square and linear fibronectin (FN) gradients on two-dimensional (2D) culture substrates. We observed that epithelial cells spread preferentially on zones of higher FN density, creating rounded or elongated gaps within epithelial tissues over circular or linear FN gradients, respectively. Using time-lapse experiments, we demonstrated that the gap closure mechanism in a 2D haptotaxis model requires a significant increase of the leader cell area. In addition, we found that gap closures are slower on decreasing FN densities than on homogenous FN-coated substrate and that fresh closed gaps are characterized by a lower cell density. Interestingly, our results showed that cell proliferation increases in the closed gap region after maturation to restore the cell density, but that cell–cell adhesive junctions remain weaker in scarred epithelial zones. Taken together, our findings provide a better understanding of the wound healing process over protein gradients, which are reminiscent of haptotaxis.

Critical role of three-dimensional tumorsphere size on experimental outcome

Three-dimensional in vitro spheroids are a reliable model to study tumor biology and drug toxicity. However, inconsistencies exist in terms of seeding cell density that governs spheroid size and shape, influencing the experimental outcome. We investigated the effect of varying cell densities using glioblastoma cells on tumorsphere formation and their responsiveness to drug treatment. Our results demonstrated that in comparison with spheroids formed with lower cell density, spheroids formed with higher cell density were not only larger in size but also had a larger necrotic core surrounded by a higher number of quiescent cells and were irresponsive to drug treatment. Our study highlights the importance of predetermination of cell density to obtain desired/appropriate spheroid size to produce consistent and reliable data on drug toxicity studies in tumor cells.

Comparative Analysis of Rhino-Cytological Specimens with Image Analysis and Deep Learning Techniques

Cytological study of the nasal mucosa (also known as rhino-cytology) represents an important diagnostic aid that allows highlighting of the presence of some types of rhinitis through the analysis of cellular features visible under a microscope. Nowadays, the automated detection and classification of cells benefit from the capacity of deep learning techniques in processing digital images of the cytological preparation. Even though the results of such automatic systems need to be validated by a specialized rhino-cytologist, this technology represents a valid support that aims at increasing the accuracy of the analysis while reducing the required time and effort. The quality of the rhino-cytological preparation, which is clearly important for the microscope observation phase, is also fundamental for the automatic classification process. In fact, the slide-preparing technique turns out to be a crucial factor among the multiple ones that may modify the morphological and chromatic characteristics of the cells. This paper aims to investigate the possible differences between direct smear (SM) and cytological centrifugation (CYT) slide-preparation techniques, in order to preserve image quality during the observation and cell classification phases in rhino-cytology. Firstly, a comparative study based on image analysis techniques has been put forward. The extraction of densitometric and morphometric features has made it possible to quantify and describe the spatial distribution of the cells in the field images observed under the microscope. Statistical analysis of the distribution of these features has been used to evaluate the degree of similarity between images acquired from SM and CYT slides. The results prove an important difference in the observation process of the cells prepared with the above-mentioned techniques, with reference to cell density and spatial distribution: the analysis of CYT slides has been more difficult than of the SM ones due to the spatial distribution of the cells, which results in a lower cell density than the SM slides. As a marginal part of this study, a performance assessment of the computer-aided diagnosis (CAD) system called Rhino-cyt has also been carried out on both groups of image slide types.

An experimental study on one-step and two-step foaming of natural rubber/silica nanocomposites

The curing and cellular structure of natural rubber (NR)/silica composite foams were investigated. The presence of an activator in the rubber formulation significantly lowered the decomposition temperature of the azodicarbonamide foaming agent, which allowed foaming before NR curing. Therefore, two foam methods were designed: foaming initially at 90°C and then curing at 140°C, and foaming and curing simultaneously at 140°C. Two-step foaming generated a lower cell density and higher cell size. Incorporation of nano silica into NR increased the foam density, but decreased the cell size. The higher foaming temperature restricted the bubble growth because of a higher curing rate and inhibited cell coalescence.

Effect of polyploidization on morphology in two apple (Malus × domestica) genotypes

Because polyploidy often results in enhancement of desirable properties, artificial genome doubling is commonly used in agri- and horticultural crop breeding programs. In this study genome doubling was induced in two apple genotypes. The effect on vegetative morphological and physiological traits of the plants was then comprehensively determined by comparing the obtained tetraploid apple plants with their diploid counterparts. Out of 17 different physio- and morphological characteristics, 15 were significantly affected in one or both genotypes. The response of these 15 characteristics also appeared to have been caused by two effects; 10 of the 15 characteristics exhibited a common response to ploidy change over both genotypes while five traits showed a genotype-specific response to polyploidization. Tetraploid leaves also exhibited a darker leaf colour, which could be correlated to a higher pigment concentration. Furthermore, the results also show a decreased elongation rate and leaf size in tetraploids, which is suggested to be due to the observed lower cell density in the polyploid apple plants.

The Iron-Dependent Regulator Fur Controls Pheromone Signaling Systems and Luminescence in the Squid Symbiont Vibrio fischeri ES114

ABSTRACTBacteria often use pheromones to coordinate group behaviors in specific environments. While high cell density is required for pheromones to achieve stimulatory levels, environmental cues can also influence pheromone accumulation and signaling. For the squid symbiontVibrio fischeriES114, bioluminescence requires pheromone-mediated regulation, and this signaling is induced in the host to a greater extent than in culture, even at an equivalent cell density. Our goal is to better understand this environment-specific control over pheromone signaling and bioluminescence. Previous work withV. fischeriMJ1 showed that iron limitation induces luminescence, and we recently found that ES114 encounters a low-iron environment in its host. Here we show that ES114 induces luminescence at lower cell density and achieves brighter luminescence in low-iron media. This iron-dependent effect on luminescence required ferric uptake regulator (Fur), which we propose influences two pheromone signaling master regulators, LitR and LuxR. Genetic and bioinformatic analyses suggested that under low-iron conditions, Fur-mediated repression oflitRis relieved, enabling more LitR to perform its established role as an activator ofluxR. Interestingly, Fur may similarly control the LitR homolog SmcR ofVibrio vulnificus. These results reveal an intriguing regulatory link between low-iron conditions, which are often encountered in host tissues, and pheromone-dependent master regulators.

Crumb evaluation of bread with hemp products addition by means of image analysis

Hemp flour composition (20–30% proteins, 7–13% fat and more than 40% saccharides) is a precondition for its usage into non-traditional cereal products. Corresponding to the fact, that hemp proteins are represented mostly by edestin, a low-molecular globulin, technological behaviour of composites containing 5–20% of hemp flour is basically different. The effect was clearly reflected in specific bread volume decrease, comparing standard wheat bread vs. wheat-hemp one. Sensorial profile of such fortified product depends on hemp sample origin, the better one was observed for dehulled wholemeal hemp flour addition. Image analysis of black-white bread cut prints revealed increasing pore densities (up to about 74%) at reversely diminishing mean cell areas (up to about 31%) for bread altered by hulled wholemeal hemp flour. Comparing to wheat standard W2, crumb appearance of bread enhanced by 5% and 20% of dehulled hemp wholemeal was described by conversely lower cell density (11 and 9 vs. 13 pores per cm2) with verifiably larger cells (3.13 a 4.25 mm2 against 2.35 mm2). Specific bread volume and crumb penetration were significantly correlated to both cell density (r −0.69 and −0.65, respectively; P = 99.9 %) and to cell mean area (0.79 and 0.69, respectively; P = 99.9 %).

Biocontrol of Ralstonia solanacearum by Treatment with Lytic Bacteriophages

ABSTRACTRalstonia solanacearumis a Gram-negative bacterium and the causative agent of bacterial wilt in many important crops. We treatedR. solanacearumwith three lytic phages: φRSA1, φRSB1, and φRSL1. Infection with φRSA1 and φRSB1, either alone or in combination with the other phages, resulted in a rapid decrease in the host bacterial cell density. Cells that were resistant to infection by these phages became evident approximately 30 h after phage addition to the culture. On the other hand, cells infected solely with φRSL1 in a batch culture were maintained at a lower cell density (1/3 of control) over a long period. Pretreatment of tomato seedlings with φRSL1 drastically limited penetration, growth, and movement of root-inoculated bacterial cells. All φRSL1-treated tomato plants showed no symptoms of wilting during the experimental period, whereas all untreated plants had wilted by 18 days postinfection. φRSL1 was shown to be relatively stable in soil, especially at higher temperatures (37 to 50°C). Active φRSL1 particles were recovered from the roots of treated plants and from soil 4 months postinfection. Based on these observations, we propose an alternative biocontrol method using a unique phage, such as φRSL1, instead of a phage cocktail with highly virulent phages. Using this method, φRSL1 killed some but not all bacterial cells. The coexistence of bacterial cells and the phage resulted in effective prevention of wilting.

Effects of Growth in the Presence of Subinhibitory Concentrations of Dicloxacillin on Staphylococcus epidermidis and Staphylococcus haemolyticus Biofilms

ABSTRACT Low concentrations of antibiotics can inhibit microbial adherence to medical device surfaces. However, little is known about the changes that occur in the physiology of bacteria within biofilms formed in the presence of subinhibitory (sub-MIC) concentrations of antibiotics. In this study, the densities and matrix compositions ofbiofilms formed by two coagulase-negative Staphylococcus species in the absence and in the presence of sub-MIC concentrations of dicloxacillin were evaluated. Biofilms formed in the presence of sub-MIC concentrations of dicloxacillin contained less biomass, and there were notable changes in the composition of the biofilm matrix. Changes in the spatial structure were also verified by confocal scanning laser microscopy, indicating that biofilms grown in the presence of sub-MIC concentrations of dicloxicilln had a lower cell density. Physiological alterations in the bacteria within biofilms grown in the presence of subinhibitory concentrations of the antibiotic were also evaluated. The results showed that there were differences in bacterial surface characteristics when cultures were grown in the presence of sub-MIC concentrations of dicloxacillin, including decreased hydrophobicity and decreased expression of the exopolysaccharide poly-N-acetylglucosamine. The elemental composition of the cell surface was also analyzed, and whereas in Staphylococcus epidermidis there were decreases in the oxygen and nitrogen contents, in Staphylococcus haemolyticus there were increases in these two parameters. Additionally, increases in resistance to several antibiotics were observed for the cells within biofilms formed in the presence of dicloxacillin.