Fibronectin, intercellular junctions and the sorting-out of chick embryonic tissue cells in monolayer

1982 ◽  
Vol 54 (1) ◽  
pp. 357-372
Author(s):  
A. Nicol ◽  
D.R. Garrod
Keyword(s):  
Corneal Epithelium ◽  
Fluorescent Antibody ◽  
Intercellular Junctions ◽  
Liver Parenchyma ◽  
Cell Types ◽  
Limb Bud ◽  
Antibody Staining ◽  
Intercellular Contacts ◽  
Mesenchyme Cells ◽  
Chick Embryonic Tissue

A hierarchy of relative cohesiveness in monolayer of four different embryonic chick tissues was determined in a previous study. The hierarchy is: corneal epithelium congruent to liver parenchyma greater than pigmented epithelium greater than limb bud mesenchyme. The purpose of this paper is to describe the correlation between these adhesive relationships and, firstly, the amount of the adhesive glycoprotein, fibronectin, associated with the cells and, secondly, the morphology of their intercellular contacts. Fluorescent antibody staining of the cells with anti-fibronectin antibody showed that limb bud mesenchyme cells, the most weakly cohesive, had much more fibronectin than the other cell types. Thus there was a negative correlation between the amount of fibronectin and cellular cohesiveness. Analysis of intercellular contacts by electron microscopy showed that the most strongly cohesive cell types, corneal epithelium and liver parenchyma, were also those that possessed desmosomes.

Mutual desmosome formation between all binary combinations of human, bovine, canine, avian and amphibian cells: desmosome formation is not tissue- or species-specific

1985 ◽  
Vol 75 (1) ◽  
pp. 377-399 ◽  
Author(s):  
D.L. Mattey ◽  
D.R. Garrod
Keyword(s):  
Epithelial Cell ◽  
Corneal Epithelium ◽  
Fluorescent Antibody ◽  
Cell Types ◽  
Recognition Mechanism ◽  
Antibody Staining ◽  
Species Specific ◽  
Molecular Components ◽  
Different Cell Types ◽  
Darby Canine Kidney

Our previous work has suggested that the molecular components of desmosomes are highly conserved between different tissues and different vertebrate species. In order to determine whether the adhesion recognition mechanism of desmosomes is also conserved we have examined the specificity of desmosome formation between different epithelial cell types by co-culturing binary combinations of cells from different species and from epidermal and non-epidermal origin. The following cell types were used: human (HeLa, cervical carcinoma), bovine (Madin Darby bovine kidney, MDBK), canine (Madin Darby canine kidney, MDCK), avian (chick embryonic corneal epithelium) and amphibian (Rana pipiens, adult corneal epithelium). Different cells in co-culture were identified on the basis of at least one of the following criteria: (1) morphology by phase-contrast microscopy; (2) presence or absence of staining of cytokeratin with monoclonal antibody LE61; (3) morphology at the electron microscope level. Mutual desmosome formation between different cell types was assessed using fluorescent antibody staining with anti-desmoplakin antibodies and confirmed using electron microscopy. We have found that mutual desmosome formation occurred between all binary combinations of human, bovine, canine, avian and amphibian cells. Thus there is complete non-selectivity of desmosome formation between five different epithelial cell types from three vertebrate classes. Our results suggest that desmosome formation is not tissue- or species-specific and that the mechanism for intercellular binding involved in desmosomal adhesion is highly conserved.

The sorting out of embryonic cells in monolayer, the differential adhesion hypothesis and the non-specificity of cell adhesion

1979 ◽  
Vol 38 (1) ◽  
pp. 249-266
Author(s):  
A. Nicol ◽  
D.R. Garrod
Keyword(s):  
Corneal Epithelium ◽  
Liver Parenchyma ◽  
Cell Types ◽  
Limb Bud ◽  
The Other ◽  
Embryonic Cells ◽  
Differential Adhesion ◽  
Discontinuous Phase ◽  
Different Cell Types ◽  
Differential Adhesion Hypothesis

It has been reported previously that sorting out of chick embryonic liver parenchyma and limb bud mesenchymal cells would take place in monolayer culture. The distribution of cell types obtained (liver formed the internal, discontinuous phase) was interpreted in terms of the differential adhesion hypothesis. It was suggested that, in monolayer, liver cells were more cohesive than limb bud cells. In this paper we set out to extend the previous observations with 2 particular questions in mind: (i) Is sorting out in monolayer a general phenomenon occurring between a wider range of cell types? (ii) Can evidence be provided for or against the interpretation of results in terms of the differential adhesion hypothesis? Sorting-out experiments were conducted on circular hydrophilic islands, on an otherwise hydrophobic substratum. Under these conditions, sorting-out in monolayer was obtained with binary combinations of 4 chick embryonic tissue types: liver parenchyma, limb bud mesenchyme, pigmented epithelium of the eye and corneal epithelium. With every combination but one, the cells of one type surrounded the cells of the other type, generating what we have called a ‘circle-within-a-circle’ configuration. With the remaining combination, liver parenchyma and corneal epithelium, only localized sorting was obtained. The ‘circle-within-a-circle’ configuration is consistent with an interpretation in terms of the differential adhesion hypothesis, according to which the distribution of cells is determined by the relative strengths of cohesions between their lateral surfaces. In direct support of this is the finding from plating the different cell types at sub-confluent density on hydrophilic substrata that limb bud is the cell tye having the weakest lateral cohesion in monolayer. Limb bud surrounded the other 3 tissues on hydrophilic island. A hierachy of lateral cohesiveness between the 4 cell types has been constructed. It is unlikely that the results can be explained in terms of specific cohesion. When plated together at subconfluent density, the 3 epithelial cell types aggregate together to form mixed monolayered islands, suggesting that they share common adhesive mechanisms.

Cell types in regenerating claws of the snapping shrimp, Alpheus heterochelis

1998 ◽  
Vol 76 (6) ◽  
pp. 1080-1090 ◽  
Author(s):  
A T Read ◽  
C K Govind
Keyword(s):  
Epithelial Cells ◽  
Blood Vessels ◽  
Cell Types ◽  
Limb Bud ◽  
Efferent Innervation ◽  
Snapping Shrimp ◽  
Intercellular Contacts ◽  
Snapping Shrimps ◽  
Cytoplasmic Processes

Cell types in the regenerating claws of adult snapping shrimps, Alpheus heterochelis, are described, based onelectron microscopy. Following autotomy of a limb, the coxal stump is secured by a membrane lined by a layer of proliferatingepithelial cells. Numerous fibroblasts with long cytoplasmic processes form small fluid-filled compartments that provide astructural framework and are inundated with mostly hemocytes and blood vessels. Agranular hemocytes are uncommoncompared with granular ones, which have prominent pseudopodia, vacuoles, and lysosomes, features that suggest a phagocyticfunction. The cytoplasmic network formed by fibroblasts persists in the regenerating blastema and papilla, together withgranular hemocytes and blastemal cells. Close structural associations were observed amongst all four cell types. Regionalproliferation of epithelial cells subdivides the distal tip of the papilla into the presumptive propus and dactyl and marks thebeginning of segmentation, which proceeds in a distal to proximal direction. This is accompanied by the appearance of firstafferent innervation, also proceeding in a distal to proximal direction, and multinucleate myoblasts identified by fragments ofmyofibrils, then efferent innervation and well-organized muscle. Prominent intercellular contacts between hemocytes and othercell types within the papilla may serve for adhesion as well as for communication. The early and prevalent appearance ofhemocytes in the regenerating limb bud, as well as their pluripotent nature in other regenerating tissues, implicates them as theorigin of blastemal cells.

scholarly journals Integrin alpha 6/beta 4 complex is located in hemidesmosomes, suggesting a major role in epidermal cell-basement membrane adhesion.

1991 ◽  
Vol 113 (4) ◽  
pp. 907-917 ◽  
Author(s):  
A Sonnenberg ◽  
J Calafat ◽  
H Janssen ◽  
H Daams ◽  
L M van der Raaij-Helmer ◽  
...  
Keyword(s):  
Basement Membrane ◽  
Epidermal Cell ◽  
Fluorescent Antibody ◽  
Cell Types ◽  
Similar Distribution ◽  
Basal Region ◽  
Basal Cells ◽  
Antibody Staining ◽  
Membrane Adhesion ◽  
Integrin Alpha 6

The alpha 6/beta 4 complex is a member of the integrin family of adhesion receptors. It is found on a variety of epithelial cell types, but is most strongly expressed on stratified squamous epithelia. Fluorescent antibody staining of human epidermis suggests that the beta 4 subunit is strongly localized to the basal region showing a similar distribution to that of the 230-kD bullous pemphigoid antigen. The alpha 6 subunit is also strongly localized to the basal region but in addition is present over the entire surfaces of basal cells and some cells in the immediate suprabasal region. By contrast staining for beta 1, alpha 2, and alpha 3 subunits was very weak basally, but strong on all other surfaces of basal epidermal cells. These results suggest that different integrin complexes play differing roles in cell-cell and cell-matrix adhesion in the epidermis. Immunoelectron microscopy showed that the alpha 6/beta 4 complex at the basal epidermal surface is strongly localized to hemidesmosomes. This result provides the first well-characterized monoclonal antibody markers for hemidesmosomes and suggests that the alpha 6/beta 4 complex plays a major role in epidermal cell-basement membrane adhesion. We suggest that the cytoplasmic domains of these transmembrane glycoproteins may contribute to the structure of hemidesmosomal plaques. Immunoultrastructural localization of the BP antigen suggests that it may be involved in bridging between hemidesmosomal plaques and keratin intermediate filaments of the cytoskeleton.

Calcium-induced desmosome formation in cultured kidney epithelial cells

1986 ◽  
Vol 85 (1) ◽  
pp. 95-111
Author(s):  
D.L. Mattey ◽  
D.R. Garrod
Keyword(s):  
Epithelial Cells ◽  
Mdck Cells ◽  
Fluorescent Antibody ◽  
Cell Types ◽  
Cell Periphery ◽  
Cell Type ◽  
Antibody Staining ◽  
Mdbk Cells ◽  
Triton X ◽  
Do So

Previous work has shown that cultured keratinocytes do not form desmosomes at low [Ca2+] (less than 0.1 mM) but may be induced to do so by raising [Ca2+] to physiological levels (1.8-2 mM). Here, fluorescent antibody staining with specific anti-desmosomal antibodies and electron microscopy have been used to determine whether Ca2+-induced desmosome formation also occurs in simple epithelial cells. Both Madin-Darby canine and bovine kidney cells (MDCK and MDBK) exhibit Ca2+-induced desmosome formation, but there are significant differences between them. MDCK cells resemble keratinocytes in showing rapid desmosome formation characterized by the simultaneous appearance of four desmosomal antigens at the cell periphery within 15–20 min of raising the [Ca2+]. In contrast MDBK cells take between 7 and 8 h to form desmosomes after Ca2+ switching, and this is characterized by slow appearance of two desmosomal antigens, the 175–164(X 10(3)) Mr glycoprotein and desmoplakin, at the cell periphery. Differences in the pattern of staining for desmosomal antigens between the two cell types in low and high [Ca2+] are described and discussed in relation to desmosome formation and internalization. Triton X-100 extractability of desmosomal antigen staining is also considered. While most is non-extractable, staining for the glycoproteins known as desmocollins is completely extractable from MDCK cells in low [Ca2+], but that which reaches the cell periphery after Ca2+ switching becomes non-extractable. Although neither cell type forms desmosomes in low [Ca2+], both possess zonulae adhaerentes, suggesting a difference in Ca2+ requirement for formation of these two junctions.

A Most Probable Number Method for the Enumeration of Legionella Bacteria in Water

1991 ◽  
Vol 24 (2) ◽  
pp. 143-147 ◽  
Author(s):  
N. A. Grabow ◽  
R. Kfir ◽  
W. O. K. Grabow
Keyword(s):  
Water Samples ◽  
Membrane Filtration ◽  
Quantitative Method ◽  
Fluorescent Antibody ◽  
Most Probable Number ◽  
Probable Number ◽  
Comparative Tests ◽  
Antibody Staining ◽  
Direct Fluorescent Antibody ◽  
Yeast Extract Agar

A new quantitative method for the enumeration of Legionella bacteria in water is described. Appropriate tenfold serial dilutions of water samples concentrated by membrane filtration are plated in triplicate on buffered charcoal yeast extract agar. After incubation for 3 days representative smears from individual plates are tested for the presence of Legionella by direct fluorescent antibody staining. The number of positive plates in each dilution is used to calculate the Legionella count by means of conventional most probable number statistics. In comparative tests on a variety of water samples this method yielded significantly higher counts than previously used procedures.

Morphilogy and Ultrastructure of in Vitro Cultures of Aortic Endothelial Cells Interacting with Antibodies

1979 ◽  
Author(s):  
S. Korach ◽  
D. Ngo
Keyword(s):  
Endothelial Cells ◽  
Cell Surface ◽  
Vascular Endothelial Cells ◽  
Intercellular Junctions ◽  
Cell Types ◽  
Endothelial Damage ◽  
Antibody Concentration ◽  
High Yields ◽  
Scanning Em

Adult pig aortas, sectioned longitudinally, were incubated in 0.1% collagenase-PBS (15 mn, 37°C). Gentle scraping of the lumenal surface resulted in high yields (3-4 x 106 cell/aorta) of viable endothelial cells, essentially devoid of other cell types by morphological and immunochemical (F VIII-antigen) criteria. Confluent monolayers were incubated for various times (5 mn to 1 wk) with decomplemented rabbit antisera raised against pig endothelial cells. Changes in cell morphology appeared to depend on antibody concentration rather than on duration of contact with antiserum. High concentrations of antiserum (5 to 20%) led to cytoplasmic shredding, bulging of cells and extensive vacuolization, whereas at lower concentrations, cells appeared almost normal. Transmission EM studies by the indirect immunoperoxydase method showed antibodies reacting with unfixed cells to be distributed all over the upper cell surface, in the outer parts of intercellular junctions, and within numerous pinocytotic vesicles. Much weaker reactions could also be seen at the lower cell surface. When viewed under the Scanning EM, antiserum-treated endothelial cells also disclosed antibody concentration-dependent bulging and release of cells from their substrate. In vitro studies of gradual modifications of vascular endothelial cells acted upon by antibodies should provide a better understanding of the structural and biochemical processes underlying endothelial damage and detachment.

Contact behaviour during the reassociation of dissociated epithelial cells in primary culture

1987 ◽  
Vol 88 (4) ◽  
pp. 521-526
Author(s):  
R.M. Brown ◽  
C.A. Middleton
Keyword(s):  
Epithelial Cells ◽  
Chick Embryo ◽  
Primary Culture ◽  
Cell Cultures ◽  
Corneal Epithelium ◽  
Time Lapse ◽  
Cell Types ◽  
Epidermal Cells ◽  
Isolated Cells ◽  
Contact Behaviour

The behaviour in culture of dissociated epithelial cells from chick embryo pigmented retina epithelium (PRE), corneal epithelium (CE) and epidermis has been studied using time-lapse cinematography. The analysis concentrated on the contact behaviour of 60 previously isolated cells of each type during a 24 h period starting 3.5 h after the cells were plated out. During the period analysed the number of isolated cells in cultures of all three types gradually decreased as they became incorporated into islands and sheets of cells. However, there were significant differences in behaviour between the cell types during the establishment of these sheets and islands. In PRE cell cultures, islands of cells developed because, throughout the period of analysis, collisions involving previously isolated cells almost invariably resulted in the development of a stable contact. Once having established contact with another cell these cells rarely broke away again to become reisolated. In contrast the contacts formed between colliding CE and epidermal cells were, at least initially, much less stable and cells of both these types were frequently seen to break away and become reisolated after colliding with other cells. Sheets and islands of cells eventually developed in these cultures because the frequency with which isolated cells become reisolated decreased with increasing time in culture. The possible reasons underlying the different behaviour of PRE cells, when compared with that of CE and epidermal cells, are discussed. It is suggested that the decreasing tendency of isolated CE and epidermal cells to become reisolated may be related to the formation of desmosomes.

Retinoic acid and chick limb bud development

Development ◽  
1991 ◽  
Vol 113 (Supplement_1) ◽  
pp. 113-121 ◽  
Author(s):  
C. Tickle
Keyword(s):  
Retinoic Acid ◽  
Target Genes ◽  
Homeobox Gene ◽  
Experimental System ◽  
Shoulder Girdle ◽  
Limb Bud ◽  
Local Concentration ◽  
Anterior Position ◽  
Mesenchyme Cells ◽  
Vitamin A Derivative

The chick limb bud is a powerful experimental system in which to study pattern formation in vertebrate embryos. Exogenously applied retinoic acid, a vitamin A derivative, can bring about changes in pattern and, on several grounds, is a good candidate for an endogenous morphogen. As such, the local concentration of retinoic acid might provide cells with information about their position in relation to one axis of the limb. Alternatively, retinoic acid may be part of a more complex signalling system. Homeobox genes are possible target genes for regulation by retinoic acid in the limb. In particular, one homeobox gene, XlHbox 1 is expressed locally in the mesenchyme of vertebrate forelimbs and might code for an anterior position. When the pattern of the chick wing is changed by retinoic acid or by grafts of signalling tissue such that anterior cells now form posterior structures, the domain of XlHbox 1 expression expands rather than contracts. The expansion of XlHbox 1 expression correlates with shoulder girdle abnormalities. Retinoic acid application leads to visible changes in bud shape and this allows dissection of the way in which patterning is co-ordinated with morphogenesis. Results of recombination experiments and studies of changes in the apical ridge and proliferation in the mesenchyme suggest the following scheme: retinoic acid is involved in specification of position of mesenchyme cells; this specification determines their local interaction with the ridge that controls ridge morphology; the thickened apical ridge permits local proliferation in the underlying mesenchyme. The recent advances in molecular biology that permit analysis of the expression of various interesting genes in developing limbs hold out the promise that further investigation may soon allow a complete account of the patterning process in one part of the vertebrate embryo.

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