Histotypic Cell Aggregation in the Presence of Cytochalasin B

1974 ◽  
Vol 16 (3) ◽  
pp. 651-663
Author(s):  
D. E. MASLOW ◽  
E. MAYHEW
Keyword(s):  
Cell Suspension ◽  
Cytochalasin B ◽  
Cell Aggregation ◽  
Cell Types ◽  
Neural Retina ◽  
Heart Cell ◽  
Heart Cells ◽  
Cell Type ◽  
Mixed Aggregates ◽  
Gyratory Shaker

A study was made of the effects of cytochalasin B on (a) specific sorting of reaggregating cells; (b) redistribution of cell types after treatment of preformed aggregates; and (c) the ability of aggregates of one cell type to incorporate and sort cells of another type. Freshly disaggregated neural retina and heart cells were cultured on a gyratory shaker at 70 rev/min and the aggregates formed analysed for sorting of cell types. Cytochalasin B disrupted the sorting of forming aggregates at concentrations of 1 µg/ml and greater. The distribution of cell types in aggregates that were treated with cytochalasin after 24 h of culture was more random than the control. Untreated cultures of retinal aggregates and heart cell suspension resulted in pure retinal and pure heart aggregates, but with more than 50 % mixed and sorted aggregates. Cytochalasin B treatment resulted in fewer mixed aggregates and a higher proportion of pure retina and pure heart aggregates.

The effects of tumour cells on embryonic cell aggregation, adhesion and detachment

1978 ◽  
Vol 29 (1) ◽  
pp. 271-275
Author(s):  
D.E. Maslow ◽  
L. Weiss
Keyword(s):  
Aggregate Size ◽  
Cell Aggregation ◽  
Inhibitory Effect ◽  
Neural Retina ◽  
Embryonic Cell ◽  
Tumour Cells ◽  
Ehrlich Ascites ◽  
Initial Adhesion ◽  
Ehrlich Ascites Cells ◽  
Gyratory Shaker

The presence of small numbers of tumour cells inhibits the aggregation of embryonic chicken neural retina cells grown in gyratory shaker culture. The aggregation of neural retina cells was also inhibited by ascites cell medium. We investigated whether the inhibitory effect of the tumour cells on aggregate size is effected by inhibition of the initial adhesion or by enhancement of their separation. The number of neural retina cells adherent to microtest plate surfaces was significantly reduced after incubation with either Ehrlich ascites cells or cell-free, conditioned medium, while the percentage of cells removed from glass by shearing was unchanged under those conditions. These results suggest that the reduction in neural retina cell aggregate size produced by Ehrlich ascites cells and their products is due to partial inhibition of neural retina cell adhesion processes, as distinct from enhancement of separation.

The analysis of spatial distributions in mixed cell populations: a statistical method for detecting sorting out

Development ◽  
1971 ◽  
Vol 26 (1) ◽  
pp. 135-156
Author(s):  
R. A. Elton ◽  
C. A. Tickle
Keyword(s):  
Plant Communities ◽  
Quantitative Measure ◽  
Cell Types ◽  
Limb Bud ◽  
Cell Populations ◽  
Spatial Distributions ◽  
Mixed Cell ◽  
Mixed Aggregates ◽  
Chick Heart ◽  
Gyratory Shaker

1. This work presents a quantitative measure, α, of the degree of segregation of two cell types in sections of aggregates, and some results obtained with the measure relating to ‘sorting out’. The method is designed particularly for the case where labelling of one type of cell is incomplete, and the importance of this effect is assessed. Possible problems in formulating such a model are discussed. The measure α is compared with methods used in investigations of segregation in plant communities. 2. Segregation of chick heart and limb-bud cells in mixed aggregates has been analysed using α. In control aggregates of mixtures of labelled and unlabelled cells of one type, α is near to its random value of 1, and we suggest that the departure from random can be adequately accounted for by cell division. In mixed aggregates, significant segregation is consistently found, even in aggregates formed after 2 and 4 h. Both disaggregation procedures (EDTA, trypsin or trypsin + EDTA) and reaggregation methods (reciprocating or gyratory shaker) are found to have an effect on the degree of segregation. Possible reasons for these findings are discussed. 3. Positioning of the cells relative to the outside of aggregates is also investigated for some of the aggregates.

Cell contacts and sorting out in vivo: the behaviour of some embryonic tissues implanted into the developing chick wing

Development ◽  
1978 ◽  
Vol 48 (1) ◽  
pp. 225-237
Author(s):  
C. Tickle ◽  
M. Goodman ◽  
L. Wolpert
Keyword(s):  
Neural Tube ◽  
Cell Types ◽  
Neural Retina ◽  
Limb Bud ◽  
Embryonic Liver ◽  
Malignant Cells ◽  
Embryonic Tissues ◽  
Mixed Aggregates ◽  
Wing Bud

The interaction of cells from embryonic liver, neural retina and mesonephros with cells from limb-bud mesenchyme has been investigated in vivo by grafting these tissues into the developing chick wing-bud. The implanted cells were in all cases from quail tissue which can be recognized histologically. As embryonic liver and neural tube are tissues that sort externally to limb-bud mesenchyme in mixed aggregates, it would be expected, from a differential adhesiveness hypothesis, that heterotypic adhesions along the borders of graft and host would be favoured over cell-cell adhesions in the graft. No morphological signs of this were evident: rather the grafted cells maximized like-like contacts. The cells of the grafts, including those from control mesenchyme, did not invade into the wing. The results were the same irrespective of whether the graft was a fragment of tissue or a pellet of reaggregated cells. This supports the idea that cells within tissues are not actively moving around and also provides controls for assaying the invasiveness of other cell types, such as malignant cells into the wing.

scholarly journals Single-Nucleus Sequencing of an Entire Mammalian Heart: Cell Type Composition and Velocity

Cells ◽  
2020 ◽  
Vol 9 (2) ◽  
pp. 318 ◽  
Author(s):  
Markus Wolfien ◽  
Anne-Marie Galow ◽  
Paula Müller ◽  
Madeleine Bartsch ◽  
Ronald M. Brunner ◽  
...  
Keyword(s):  
Cell Types ◽  
Cellular Level ◽  
Heart Cell ◽  
Cell Type ◽  
Mammalian Heart ◽  
Cell Type Composition ◽  
Type Composition ◽  
A Cell ◽  
Single Nucleus ◽  
First Time

Analyses on the cellular level are indispensable to expand our understanding of complex tissues like the mammalian heart. Single-nucleus sequencing (snRNA-seq) allows for the exploration of cellular composition and cell features without major hurdles of single-cell sequencing. We used snRNA-seq to investigate for the first time an entire adult mammalian heart. Single-nucleus quantification and clustering led to an accurate representation of cell types, revealing 24 distinct clusters with endothelial cells (28.8%), fibroblasts (25.3%), and cardiomyocytes (22.8%) constituting the major cell populations. An additional RNA velocity analysis allowed us to study transcription kinetics and was utilized to visualize the transitions between mature and nascent cellular states of the cell types. We identified subgroups of cardiomyocytes with distinct marker profiles. For example, the expression of Hand2os1 distinguished immature cardiomyocytes from differentiated cardiomyocyte populations. Moreover, we found a cell population that comprises endothelial markers as well as markers clearly related to cardiomyocyte function. Our velocity data support the idea that this population is in a trans-differentiation process from an endothelial cell-like phenotype towards a cardiomyocyte-like phenotype. In summary, we present the first report of sequencing an entire adult mammalian heart, providing realistic cell-type distributions combined with RNA velocity kinetics hinting at interrelations.

Reconstruction of Mammalian Heart Tissue from Isolated Embryonic Heart Cells

1977 ◽  
Vol 35 ◽  
pp. 580-581
Author(s):  
Asish C. Nag ◽  
Debra S. Buszke
Keyword(s):  
Basal Medium ◽  
Cell Aggregation ◽  
Heart Tissue ◽  
Cell Suspensions ◽  
Heart Cells ◽  
Mammalian Heart ◽  
Dissociated Cells ◽  
Gyratory Shaker ◽  
And Function

Although monolayer cultures are useful in various cell studies, they are not always adequate for examining the nature of cellular interrelationships and interactions in the formation, differentiation, and function of tissues. The procedures of aggregation in vitro of dissociated cells (Moscona & Moscona, 1966) enable one to study in detail the formation of cell contacts, the assembly of cells into multicellular systems, and cell cooperation in forming organized and differentiating tissues. We adapted the techniques of cell aggregation by rotation to studies on embryonic mammalian heart cells. Cell suspensions from the 18-day-old embryonic rat were prepared by dissociation with 0. 5% trypsin. The cells were dispersed in a culture medium which consisted of Eagle's basal medium with 10% fetal bovine serum, 1% glutamine solution, and 1% penicillin-streptomycin mixture. Cultured in 25ml Erlenmeyer flasks on a gyratory shaker at 70 rpm at 37°C were 3ml aliquots of the cell suspensions.

scholarly journals DO MORPHOGENETIC TISSUE REARRANGEMENTS REQUIRE ACTIVE CELL MOVEMENTS?

1972 ◽  
Vol 55 (3) ◽  
pp. 606-615 ◽  
Author(s):  
Malcolm S. Steinberg ◽  
Lawrence L. Wiseman
Keyword(s):  
Solid Surface ◽  
Cell Sorting ◽  
Cytochalasin B ◽  
Cell Aggregation ◽  
Heart Cell ◽  
Embryonic Cells ◽  
Morphogenetic Movements ◽  
Cell Locomotion ◽  
Mutual Contact ◽  
Motile Cells

Previous studies have indicated that cell sorting and tissue spreading are caused by cell combination-specific differences in intercellular adhesive energies, acting in a system of motile cells. We wished to determine whether these adhesive energies could drive cell rearrangements as well as guide them. Accordingly, aggregates of intermixed embryonic cells were cultured in solutions of the drug cytochalasin B (CCB) at a concentration shown to inhibit the locomotion of cells on a solid surface. In addition, spherical aggregates of several kinds were cultured in mutual contact under similar conditions. Both cell sorting and tissue spreading were found to be inhibited. The prompt release of this inhibition upon removal of the CCB showed that the inhibited cells were not merely injured. Moreover, aggregation experiments showed that CCB did not prevent cells of several kinds from initiating mutual adhesions. In fact, heart cell aggregation was enhanced by CCB. We conclude that interfacial forces, originating outside the cell, act together with forces originating inside it in bringing about the morphogenetic movements of cell sorting and tissue spreading. We propose the term "cooperative cell locomotion" to describe translational movements of cells arising from such a combination of intrinsic and extrinsic forces.

STEROID RELEASE IN VITRO BY TWO LUTEAL CELL TYPES IN THE CORPUS LUTEUM OF THE PREGNANT SOW

1977 ◽  
Vol 72 (3) ◽  
pp. 351-359 ◽  
Author(s):  
MEREDITH LEMON ◽  
M. LOIR
Keyword(s):  
Cell Suspension ◽  
Corpus Luteum ◽  
Initial Rate ◽  
Cell Types ◽  
Luteal Cell ◽  
Cell Populations ◽  
Corpora Lutea ◽  
Cell Type ◽  
Three Stages

SUMMARY Corpora lutea from sows at 30, 60 and 90 days of gestation were dissociated enzymically, and the components of the resulting cell suspension were separated by sedimentation at unit gravity. Two luteal cell populations of 30–50 μm diameter and 15–20 μm diameter were obtained and superfused for up to 18 h with Dulbecco's modified Eagle medium, the cells being supported in a column in a matrix of Biogel. Fractions were collected every 30 min and assayed for progesterone and oestradiol-17β. At 30 and 60 days of gestation the large luteal cells produced progesterone at an initial rate of approximately 100 ng/h/105 cells, which decreased to half this rate at 90 days. The smaller cells also released progesterone into the medium at approximately 15–20 ng/h/105 cells at all stages of gestation. At 30 days of gestation, neither cell type released significant amounts of oestradiol-17β, but from 60 days onwards, significant and increasing quantities were measured in the superfusates from the larger cells. Both cell types were perfused with porcine LH at the three stages of gestation, and both showed an immediate response in terms of progesterone release which decreased in magnitude with increasing age of gestation. The response of the smaller cells was greater than that of the larger 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.

Abstract 16742: Single-Cell Transcriptional and Epigenomic Dissection of Human Heart in Health and Coronary Artery Disease Reveals Cell-type-Specific Driver Genes and Pathways

Circulation ◽  
2020 ◽  
Vol 142 (Suppl_3) ◽  
Author(s):  
Suvi Linna Kuosmanen ◽  
Eloi Schmauch ◽  
Kyriakitsa Galani ◽  
Carles Boix ◽  
Yongjin P Park ◽  
...  
Keyword(s):  
Coronary Artery Disease ◽  
Single Cell ◽  
Human Heart ◽  
Target Genes ◽  
Expression Patterns ◽  
Cell Types ◽  
Heart Cell ◽  
Cell Type ◽  
Cell Type Specific ◽  
Artery Disease

Genome-wide association studies have uncovered over 200 genetic loci underlying coronary artery disease (CAD), providing great hope for a deeper understanding of the causal mechanisms leading to this disease. However, in order to understand CAD at the molecular level, it is necessary to uncover cell-type-specific circuits and to use these circuits to dissect driver variants, genes, pathways, and cell types, in normal and diseased tissues. Here, we provide the most detailed single-cell dissection of human heart cell types, using cardiac biopsies collected during open-heart surgery from healthy, CAD, and CAD-related heart failure donors, and profiling both transcriptional (scRNA-seq) and epigenomic (scATAC-seq) changes. Using this approach, we identify 12 major heart cell types, including typical cardiovascular cells (cardiomyocytes, endothelial cells, fibroblasts), rarer cell types (B cells, neurons, Schwann cells), and previously-unrecognized layer-specific epithelial and endothelial cell types. We define markers for each cell type, providing the first extensive reference set for the living human heart. In addition, we define differential gene expression patterns in CAD relative to control samples, revealing substantial differences in cell-type-specific expression of disease-related genes, emphasizing, for example, the importance of the vascular endothelium in the pathogenesis of CAD. Strikingly, further clustering of the cell types based on specific subtypes revealed important differences in their expression patterns of disease-associated genes. These changes enrich in known CAD genetic loci, enabling us to recognize their likely target genes from scRNA-seq expression changes, candidate driver variants based on scATAC-seq localization and differential DNA accessibility, and candidate upstream regulators based on their enriched motif occurrences in scATAC loci. Overall, our results highlight the relevance and potential of single-cell transcriptional and epigenomic analyses to gain new biological insights into cardiovascular disease, and to recognize novel therapeutic target genes, pathways, and the cell types where they act.

69 EFFECTS OF CELL TYPE, PRE-ACTIVATION PROTOCOL, AND CULTURE CONDITIONS ON DEVELOPMENT OF PORCINE HANDMADE CLONED EMBRYOS

2010 ◽  
Vol 22 (1) ◽  
pp. 193
Author(s):  
J. C. Mezzalira ◽  
L. U. Ohlweiler ◽  
A. Massie ◽  
E. Monaco ◽  
E. P. Silva ◽  
...  
Keyword(s):  
In Vitro Culture ◽  
Cytochalasin B ◽  
Cell Types ◽  
Blastocyst Stage ◽  
Culture Conditions ◽  
Dependent Manner ◽  
Cell Type ◽  
Embryo Production ◽  
Distinct Cell

Despite the rather successful and widespread use of cloning in various species, distinct cell types from the same species and even the same genotype display differences in blastocyst yield. Moreover, variations in the protocol for embryo production can influence development to the blastocyst stage and subsequent fetal development. The aim of this study was to evaluate the effect of 2 cell types and 2 embryo pre-activation protocols with or without the presence of FCS in the in vitro culture medium on development of handmade pig cloned embryos to the blastocyst stage. Cumulus-oocyte complexes recovered from sow ovaries were in vitro-matured for 38 to 40 h. Denuded matured oocytes selected by the presence of a polar body had the zona pellucida removed in a 0.2% protease HEPES-buffered solution +25% FCS, followed by manual bisection and UV screening of enucleated halves using Hoechst stain. Clone embryo reconstruction was performed using a phytohemoagglutinin solution to adhere 2 cytoplasts and a somatic cell. Adipocyte-derived mesenchymal stem cells (ADMSC) from a Yorkshire pig or granulosa cells (GC) from an Ossabaw pig were used as nuclear donors. Following electrical fusion, couplets were pretreated with a brief exposure to cytochalasin B (CB) or cytochalasin B + cycloheximide (CB+CX) in the presence of serum before the electrical activation (Naruse et al. 2007 Theriogenology 68, 709-716; Du et al. 2009 Reprod. Fertil. Dev. 21, 114). Activated embryos were in vitro-cultured in the well of the well (WOW) system, with 2 embryos per microwell, for 7 days in PZM-3 medium +0.3% BSA in the presence (FBS+) or absence (FBS-) of 10% FCS. Cleavage (Day 2, chi-square test) and blastocyst (Day 7, Fisher test) rates, on a per WOW basis, were compared for a level of significance of 5%. Our preliminary data indicate that the presence of serum in the IVC affected cleavage and blastocyst yield in a cell-type-dependent manner. The presence of serum enhanced the blastocyst yield for ADMSC, whereas for GC, only the absence of serum allowed any blastocyst development. The cell type and the pre-activation protocol did not appear to affect cleavage and embryo development to the blastocyst stage. Despite the low number of replications, our results reinforce the importance of optimizing the embryo production system taking into consideration the individual requirements for distinct cell types, procedures, and culture conditions. Table 1.Effects of cell type, pre-activation process and in vitro culture (IVC) medium on development of handmade pig cloned embryos

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