Observations on the sorting-out of embryonic cells in monolayer culture

1975 ◽  
Vol 18 (3) ◽  
pp. 385-403
Author(s):  
M.S. Steinberg ◽  
D.R. Garrod
Keyword(s):  
Monolayer Culture ◽  
Cell Movement ◽  
Time Lapse ◽  
Cell Types ◽  
Liver Cells ◽  
Contact Inhibition ◽  
Limb Bud ◽  
Embryonic Cells ◽  
Embryo Cells ◽  
Embryonic Limb

Two problems are raised concerning the movement of cells during tissue-specific sorting-out of chick embryo cells in mixed aggregates. (i) A possible expectation from the hypothesis of ‘contact inhibition’ is that cells which are entirely surrounded by other cells in monolayer should be held stationary. Cells within solid aggregates, being totally surrounded by others, might also not be expected to move. How is it then that cell movement takes place within solid aggregates during sorting-out? (ii) Are the movements of cells within sorting aggregates ‘passive’, being driven by adhesive differentials or ‘active’, being merely guided by such differentials? In order to study these questions, sorting out experiments with chick embryonic limb bud mesenchyme and liver cells were carried out in monolayer culture, permitting direct observation of cell movements. Cell behavior was observed by time-lapse cinematography. Sorting-out of these cells in monolayer began before and continued after the cells had spread to confluency. During sorting, liver cells showed ruffing activity even when they appeared to be totally surrounded by other cells. Both cell types showed contact inhibition as judged by the criterion of monolayering, for they did not move over each other but remained attached to the substratum. Yet the cells in the confluent monolayer were not immobilized. Because of this, we suggest that the observed restraint against overlapping did not result from an inhibition of movement. Several considerations, detailed in the text, suggest that cell movement during sorting-out involve active locomotion. Previous work suggest that sorting-out configurations are determined by the relative intensities of intercellular adhesive strengths, the more cohesive of 2 cell populations tending to adopt the internal position. While limb bud cells form internal islands surrounded by liver cells in solid aggregates, the reverse was found to be the case in these monolayers. This suggests that, in the monolayer, limb bud cohesiveness is depressed relative to liver cell cohesiveness. This is consistent with the observation that the limb bud cells flattened themselves markedly against the substratum, significantly decreasing their area of mutual apposition.

Polygonal networks in living chick embryonic cells

1981 ◽  
Vol 52 (1) ◽  
pp. 55-69
Author(s):  
G.W. Ireland ◽  
F.C. Voon
Keyword(s):  
Dorsal Surface ◽  
Time Lapse ◽  
Cell Types ◽  
Outer Edge ◽  
Live Cells ◽  
Embryonic Cells ◽  
Embryo Cells ◽  
Dissociated Cells ◽  
The Relationship ◽  
Free Edges

Regular polygonal networks have been found in explants and dissociated cells of early chick embryos. These networks are readily observable in live cells with phase-contrast optics thus allowing time-lapse cinemicroscopy. They consisted of a regular pattern of nodes and radiating struts found predominantly in the lamelliplasm of the free edges of the cells bordering explants. At the outer edge, the network was terminated by radial struts associated with substrate-attached retraction processes whilst toward the centre of the cells it faded out. The network was also associated with stress fibres running across the cell and with microextensions on the dorsal surface. Even within one cell the network varied in size. Time-lapse films showed that microvilli were protruded from the dorsal surface over the nodes. Although the cells containing the networks were poorly motile the network itself was a mobile structure. Many explants from regions differing in prospective fates developed these networks after 2–4 days in culture. They appeared earlier in the smaller less yolky cells of definitive endoblast and epiblast. Experiments with dissociated and reaggregated cells confirmed their occurrence mainly in free edges of cells. The relationship between these networks seen in living chick embryo cells and those seen in other cell types using immunofluorescent techniques is discussed and a mechanism is proposed for their formation.

scholarly journals A Spindle Checkpoint Functions during Mitosis in the Early Caenorhabditis elegans Embryo

2005 ◽  
Vol 16 (3) ◽  
pp. 1056-1070 ◽  
Author(s):  
Sandra E. Encalada ◽  
John Willis ◽  
Rebecca Lyczak ◽  
Bruce Bowerman
Keyword(s):  
Caenorhabditis Elegans ◽  
Chromosome Segregation ◽  
Spindle Checkpoint ◽  
Metaphase Plate ◽  
Time Lapse ◽  
Cell Types ◽  
Embryonic Cells ◽  
Mitotic Spindles ◽  
Animal Cells ◽  
Dividing Cells

During mitosis, chromosome segregation is regulated by a spindle checkpoint mechanism. This checkpoint delays anaphase until all kinetochores are captured by microtubules from both spindle poles, chromosomes congress to the metaphase plate, and the tension between kinetochores and their attached microtubules is properly sensed. Although the spindle checkpoint can be activated in many different cell types, the role of this regulatory mechanism in rapidly dividing embryonic animal cells has remained controversial. Here, using time-lapse imaging of live embryonic cells, we show that chemical or mutational disruption of the mitotic spindle in early Caenorhabditis elegans embryos delays progression through mitosis. By reducing the function of conserved checkpoint genes in mutant embryos with defective mitotic spindles, we show that these delays require the spindle checkpoint. In the absence of a functional checkpoint, more severe defects in chromosome segregation are observed in mutants with abnormal mitotic spindles. We also show that the conserved kinesin CeMCAK, the CENP-F-related proteins HCP-1 and HCP-2, and the core kinetochore protein CeCENP-C all are required for this checkpoint. Our analysis indicates that spindle checkpoint mechanisms are functional in the rapidly dividing cells of an early animal embryo and that this checkpoint can prevent chromosome segregation defects during mitosis.

Two distinct classes of prechondrogenic cell types in the embryonic limb bud

1983 ◽  
Vol 97 (1) ◽  
pp. 59-69 ◽  
Author(s):  
B.J. Swalla ◽  
E.M. Owens ◽  
T.F. Linsenmayer ◽  
M. Solursh
Keyword(s):  
Cell Types ◽  
Limb Bud ◽  
Embryonic Limb

Cell-sorting in aggregates of Dictyostelium discoideum

1999 ◽  
Vol 112 (22) ◽  
pp. 3923-3929 ◽  
Author(s):  
A. Nicol ◽  
W. Rappel ◽  
H. Levine ◽  
W.F. Loomis
Keyword(s):  
Cell Sorting ◽  
Fluorescent Protein ◽  
Cell Mass ◽  
Three Dimensional ◽  
Cell Movement ◽  
Time Lapse ◽  
Cell Types ◽  
Cell Type ◽  
Prespore Cell ◽  
Green Fluorescent

When Dictyostelium cells are induced to develop between a coverslip and a layer of agarose, they aggregate normally into groups containing up to a thousand cells but are then constrained to form disks only a few cells thick that appear to be equivalent to the three-dimensional mounds formed on top of agarose. Such vertically restricted aggregates frequently develop into elongated motile structures, the flattened equivalent of three-dimensional slugs. The advantage of using this system is that the restricted z-dimension enables direct microscopic visualization of most of the cells in the developing structure. We have used time lapse digital fluorescence microscopy of Dictyostelium strains expressing green fluorescent protein (GFP) under the control of either prestalk or prespore specific promoters to follow cell sorting in these flattened mounds. We find that prestalk and prespore cells expressing GFP arise randomly in early aggregates and then rotate rapidly around the disk mixed with the other cell type. After a few hours, the cell types sort out by a process which involves striking changes in relative cell movement. Once sorted, the cell types move independently of each other showing very little heterotypic adhesion. When a group of prestalk cells reaches the edge of the disk, it moves out and is followed by the prespore cell mass. We suggest that sorting may result from cell type specific changes in adhesion and the consequent disruption of movement in the files of cells that are held together by end-to-end adhesion.

scholarly journals Distribution of microtubule organizing centers in migrating sheets of endothelial cells.

1981 ◽  
Vol 91 (2) ◽  
pp. 589-594 ◽  
Author(s):  
A I Gotlieb ◽  
L M May ◽  
L Subrahmanyan ◽  
V I Kalnins
Keyword(s):  
Endothelial Cells ◽  
Monolayer Culture ◽  
Cell Movement ◽  
Time Lapse ◽  
Microtubule Organizing Center ◽  
Organizing Center ◽  
Direction Of Movement ◽  
The Relationship ◽  
Wounded Area

This study was designed to investigate the relationship between the position of the microtubule organizing center (MTOC) and the direction of migration of a sheet of endothelial cells (EC). Using immunofluorescence and phase microscopy the MTOC's of migrating EC were visualized as the cells moved into an in vitro experimental wound produced by mechanical denudation of part of a confluent monolayer culture. Although the MTOC's in nonmigrating EC were randomly positioned in relation to the nucleus, in migrating cells the position of the MTOC's changed so that 80% of the cells had the MTOC positioned in front of the nucleus toward the direction of movement of the endothelial sheet. This repositioning of the MTOC occurred within the first 4 h after wounding and was associated with the beginning of migration of EC's into the wounded area as seen by time-lapse cinemicrophotography. These studies focus attention on the MTOC as a cytoskeletal structure that may play a role in determining the direction of cell movement.

scholarly journals Locomotory behavior, contact inhibition, and pattern formation of 3T3 and polyoma virus-transformed 3T3 cells in culture

1977 ◽  
Vol 74 (3) ◽  
pp. 963-982 ◽  
Author(s):  
PB Bell
Keyword(s):  
Social Behavior ◽  
Adhesion Formation ◽  
Time Lapse ◽  
Cell Types ◽  
Contact Inhibition ◽  
Locomotory Activity ◽  
3T3 Cells ◽  
Reduced Frequency ◽  
The Social ◽  
The Individual

The social behavior of 3T3 cells and their polynoma virus-transformed derivative (Py3T3 cells) was examined by time-lapse cinemicrography in order to determine what factors are responsible for the marked differences in the patterns formed by the two cell lines in culture. Contrary to expectations, both cell types have been found to exhibit contact inhibition of cell locomotion. Therefore, the tendency of 3T3 cells to form monolayers and of Py3T3 cells to form crisscrossed multilayers cannot be explained on the basis of the presence versus the absence of contact inhibition. Morevover, with the exception of cell division control, the social behavior of the two cell types is qualitively similar. Both exhibit cell underlapping and, after contact between lamelliopodia, both show inhibition of locomotory activity and adhesion formation. Neither cell type was observed to migrate over the surface of another cell. The two cell types do show quantitative differences in the frequency of underlapping, the frequency with which contact results in inhibition of locomotion, and the proportion of the cell margin that adheres to the substratum. The increased frequency pf Py3T3 underlapping is correlated with the reduced frequency of substratum adhesions, which in turn favors underlapping. On the basis of these observations, it is concluded that the differences in culture patterns are the result of differences in the shapes of the individual cells, such that underlapping, and hence crisscrossing, is favored in Py3T3 cell interactions and discouraged in 3T3 cells.

scholarly journals CELL SORTING IN THE PRESENCE OF CYTOCHALASIN B

1972 ◽  
Vol 55 (3) ◽  
pp. 542-553 ◽  
Author(s):  
Peter B. Armstrong ◽  
David Parenti
Keyword(s):  
Chick Embryo ◽  
Cell Sorting ◽  
Cytochalasin B ◽  
Monolayer Culture ◽  
Cell Types ◽  
Neural Retina ◽  
Limb Bud ◽  
Cell Contact ◽  
Adhesive Interaction ◽  
Cellular Locomotion

The ability of cytochalasin B to inhibit ruffled membrane activity and cellular locomotion of vertebrate cells in monolayer culture prompted its use to study the necessity for this kind of active cellular locomotion in cell sorting in heterotypic cell aggregates. Cell sorting was inhibited in chick embryo heart-pigmented retina aggregates but a remarkable degree of sorting did occur in neural retina-pigmented retina aggregates. In these experiments, the levels of cytochalasin B employed (5 or 10 µg/ml) are sufficient to inhibit completely locomotion of these cell types in monolayer culture. It is proposed that the degree of cell movement achieved during sorting in neural retina-pigmented retina aggregates in the presence of cytochalasin B is the result of changes in cell contact resulting from adhesive interaction of cells. The effect of cytochalasin B on the initial aggregation of dissociated cells was also tested. With the cell types used in this study (chick embryo neural retina and limb bud), aggregation was not affected for a period of several hours.

scholarly journals Intercellular adhesive selectivity. III. Species selectivity of embryonic liver intercellular adhesion.

1976 ◽  
Vol 71 (1) ◽  
pp. 96-106 ◽  
Author(s):  
S R Grady ◽  
E J McGuire
Keyword(s):  
Species Difference ◽  
Cell Types ◽  
Liver Cells ◽  
Evolutionary Divergence ◽  
Embryonic Liver ◽  
Embryonic Cells ◽  
Adhesive Properties ◽  
Assay Procedure ◽  
Double Label ◽  
Different Cell Types

A species difference in the intercellular adhesive selectivity of mixtures of embryonic liver cells is reported. This is first quantitative assessment of species differences in the intercellular adhesive properties of embryonic cells. A collecting aggregate assay, a new double-label assay procedure, and histological and autoradiographic procedures were used to elucidate the intercellular adhesive selectivity of developing mammalian and avian liver cells. Evidence is presented that the reported adhesive differences are not due to the different cell types composing the respective embryonic mammalian and avian livers. Finally, such heterolgous-homotypic selectivity of adhesion is not a property of all tissues, since it is shown that developing brain cells (mesencephalon) do not exhibit the avove intercellular adhesive selectivity (mammalian vs. avian). These findings provide further support for the hypothesis that generic identity as well as cell type may play an important part in determining the intercellular adhesive behavior of heterologous-homotypic mixtures of embryonic cells. A possible evolutionary divergence of morphogenetic mechanisms is discussed.

scholarly journals Robust Optical Flow Algorithm for General, Label-free Cell Segmentation

2020 ◽  
Author(s):  
Michael C. Robitaille ◽  
Jeff M. Byers ◽  
Joseph A. Christodoulides ◽  
Marc P. Raphael
Keyword(s):  
Optical Flow ◽  
Cell Biology ◽  
Cell Movement ◽  
Time Lapse ◽  
Cell Types ◽  
Free Cell ◽  
Cell Segmentation ◽  
Label Free ◽  
Segmentation Approach

ABSTRACTCell segmentation is crucial to the field of cell biology, as the accurate extraction of cell morphology, migration, and ultimately behavior from time-lapse live cell imagery are of paramount importance to elucidate and understand basic cellular processes. Here, we introduce a novel segmentation approach centered around optical flow and show that it achieves robust segmentation by validating it on multiple cell types, phenotypes, optical modalities, and in-vitro environments without the need of labels. By leveraging cell movement in time-lapse imagery as a means to distinguish cells from their background and augmenting the output with machine vision operations, our algorithm reduces the number of adjustable parameters needed for optimization to two. The code is packaged within a MATLAB executable file, offering an accessible means for general cell segmentation typically unavailable in most cell biology laboratories.

scholarly journals Studies of intercellular invasion in vitro using rabbit peritoneal neutrophil granulocytes (PMNS). I. Role of contact inhibition of locomotion.

1975 ◽  
Vol 65 (2) ◽  
pp. 439-462 ◽  
Author(s):  
P B Armstrong ◽  
J M Lackie
Keyword(s):  
Monolayer Culture ◽  
Time Lapse ◽  
Cell Types ◽  
Host Tissue ◽  
Necessary Condition ◽  
Neutrophil Granulocytes ◽  
Cell Type ◽  
A Cell

Intercellular invasion is the active migration of cells on one type into the interiors of tissues composed of cells of dissimilar cell types. Contact paralysis of locomotion is the cessation of forward extension of the pseudopods of a cell as a result of its collision with another cell. One hypothesis to account for intercellular invasion proposes that a necessary condition for a cell type to be invasive to a given host tissue is that it lack contact paralysis of locomotion during collision with cells of that host tissue. The hypothesis has been tested using rabbit peritoneal neutrophil granulocytes (PMNs) as the invasive cell type and chick embryo fibroblasts as the host tissue. In organ culture, PMNs rapidly invade aggregates of fibroblasts. The behavior of the pseudopods of PMNs during collision with fibroblasts was analyzed for contact paralysis by a study of time-lapse films of cells in mixed monolayer culture. In monolayer culture, PMNs show little sign of paralysis of the pseudopods upon collision with fibroblasts and thus conform in their behavior to that predicted by the hypothesis.

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