1,720,967 research outputs found
Quantitative in vitro characterization of transformed cell lines as a model for metastatic dissemination
No full textAnalysis and modelling of motility of cell populations with MotoCell
Full text linkBackground
Cell motility plays a central role in development, wound-healing and tumour invasion. Cultures of eucariotic cells are a complex system where most cells move according to 'random' patterns, but may also be induced to a more coordinate migration by means of specific stimuli, such as the presence of chemical attractants or the introduction of a mechanical stimulus. Various tools have been developed that work by keeping track of the paths followed by specific objects and by performing statistical analysis on the recorded path data. The available tools include desktop applications or macros running within a commercial package, which address specific aspects of the process.
Results
An online application, MotoCell, was developed to evaluate the motility of cell populations maintained in various experimental conditions. Statistical analysis of cell behaviour consists of the evaluation of descriptive parameters such as average speed and angle, directional persistence, path vector length, calculated for the whole population as well as for each cell and for each step of the migration; in this way the behaviour of a whole cell population may be assessed as a whole or as a sum of individual entities. The directional movement of objects may be studied by eliminating the modulo effect in circular statistics analysis, able to evaluate linear dispersion coefficient (R) and angular dispersion (S) values together with average angles. A case study is provided where the system is used to characterize motility of RasV12 transformed NIH3T3 fibroblasts.
Conclusion
Here we describe a comprehensive tool which takes care of all steps in cell motility analysis, including interactive cell tracking, path editing and statistical analysis of cell movement, all within a freely available online service. Although based on a standard web interface, the program is very fast and interactive and is immediately available to a large number of users, while exploiting the web approach in a very effective way. The ability to evaluate the behaviour of single cells allows to draw the attention on specific correlations, such as linearity of movement and deviation from the expected direction. In addition to population statistics, the analysis of single cells allows to group the cells into subpopulations, or even to evaluate the behaviour of each cell with respect to a variable reference, such as the direction of a wound or the position of the closest cell
Time-Lapse Phase-Contrast Microscopy Fibroblast Automated Tracking
No full textHigh-throughput applications on time-lapse microscopy allows us to follow the in vitro temporal and spatial evolution of cell populations; analysis on those kind of data will reveal cell motion parameters such as average speed, persistence and directionality, important informations for many research and therapeutic applications such as drug development or wound healing. The large quantity of frames usually acquired containing multiple cells require automated analysis methods dealing with a cell tracking process. In this work an either semi- or fully automated cell tracking system is proposed, developed to deal especially with time-lapse phase-contrast microscopy of fibroblasts, which are difficult to follow cells in some cases also for a human expert
Ras activated ERK and PI3K pathways differentially affect directional movement of cultured fibroblasts.
No full textBackground: Cell migration is essential in physiological and pathological processes, such as wound healing and metastasis formation. Ras involvement in these processes has been extensively demonstrated. This work attempts to characterize Ras regulation of the phenomena determining directional cell migration by separately analyzing the role of its principal effector pathways, MAPK and PI3K. Methods: NIH3T3 and NIHRasV12 fibroblasts were followed in wound healing assays to study, in time and under a directional stimulus, cell migration both under standard conditions and in presence of MAPK and PI3K inhibitors. Several parameters, descriptive of specific aspects of cell motion, were evaluated by coupling dynamic microscopy with quantitative and statistical methods. Quantitative Western Blots coupled with immunofluorescence stainings, were used to evaluate ERK activation. Results: Constitutive RasV12 activation confers to NIH3T3 the ability to close the wound faster. Neither increased cell proliferation nor higher speed explains the accelerated healing, but the increased directional migration drives the wound closure. Inhibition of ERK activation, which occurs immediately after wound, greatly blocks the directional migration, while inhibition of PI3K pathway reduces cell speed but does not prevent wound closure. Conclusion: Ras is greatly involved in determining and regulating directionality, ERK is its key effector for starting, driving and regulating directional movemen
A three component model for superdiffusive motion effectively describes migration of eukaryotic cells moving freely or under a directional stimulus
No full textAlthough the simple diffusion model can effectively describe the movement of eukaryotic cells on a culture surface observed at relatively low sampling frequency, at higher sampling rates more complex models are often necessary to better fit the experimental data. Currently available models can describe motion paths by involving additional parameters, such as linearity or directional persistence in time. However sometimes difficulties arise as it is not easy to effectively evaluate persistence in presence of a directional bias. Here we present a procedure which helps solve this problem, based on a model which describes displacement as the vectorial sum of three components: diffusion, persistence and directional bias. The described model has been tested by analysing the migratory behaviour of simulated cell populations and used to analyse a collection of experimental datasets, obtained by observing cell cultures in time lapse microscopy. Overall, the method produces a good description of migration behaviour as it appears to capture the expected increase in the directional bias in presence of wound without a large concomitant increase in the persistence module, allowing it to remain as a physically meaningful quantity in the presence of a directional stimulus
Methods and computational tools to study eukaryotic cell migration in vitro
Full text linkISSN:2296-634
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
SARS-CoV-2 pandemic tracing in Italy highlights lineages with mutational burden in growing subsets
Full text linkTracing the appearance and evolution of virus variants is essential in the management
of the COVID-19 pandemic. Here, we focus on SARS-CoV-2 spread in Italian patients by using
viral sequences deposited in public databases and a tracing procedure which is used to monitor the
evolution of the pandemic and detect the spreading, within the infected population of emergent
sub-clades with a potential positive selection. Analyses of a collection of monthly samples focused
on Italy highlighted the appearance and evolution of all the main viral sub-trees emerging at the
end of the first year of the pandemic. It also identified additional expanding subpopulations which
spread during the second year (i.e., 2021). Three-dimensional (3D) modelling of the main amino acid
changes in mutated viral proteins, including ORF1ab (nsp3, nsp4, 2’-o-ribose methyltransferase, nsp6,
helicase, nsp12 [RdRp]), N, ORF3a, ORF8, and spike proteins, shows the potential of the analysed
structural variations to result in epistatic modulation and positive/negative selection pressure. These
analyzes will be of importance to the early identification of emerging clades, which can develop into
new “variants of concern” (i.e., VOC). These analyses and settings will also help SARS-CoV-2 coronet
genomic centers in other countries to trace emerging worldwide varian
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