Non-reciprocal contact inhibition of locomotion of chick embryonic choroid fibroblasts by pigmented retina epithelial cells

1978 ◽  
Vol 33 (1) ◽  
pp. 103-120
Author(s):  
E.K. Parkinson ◽  
J.G. Edwards
Keyword(s):  
Epithelial Cells ◽  
Light And Electron Microscopy ◽  
Leading Edge ◽  
Time Lapse ◽  
Contact Inhibition ◽  
Cultured Fibroblasts ◽  
Lateral Contact ◽  
Fibroblastic Cells

Using light and electron microscopy, we have confirmed an earlier observation that chick embryonic pigmented retina epithelial cells (PRE cells) seeded in vitro on cultured sheets of choroid fibroblasts, are able to spread. Spreading is as rapid (and shows the same dependence on lateral contact between PRE cells) as on a serum-coated culture substrate. After 1 h most cells are spreading on the upper surface of the choroid sheet, but after 4 h, some PRE cells can be found sandwiched between overlapping choroid cells, and thus have invaded the sheet. Choroid fibroblasts underlie PRE in vivo, but the ability of PRE cells to spread on cultured fibroblasts is not specific for choroid, since PRE cells spread also on BKH21 hamster kidney fibroblasts, and on fibroblasts from chick embryonic heart. As reported by others for various fibroblastic cells, choroid cells seeded on to choroid sheets or on to cultured PRE are unable to spread. A possible explanation is that spreading of adherent cells is contact-inhibited by the cells in the sheet, just as their leading edges are paralysed on contact, and thus locomotion is inhibited, when fibroblasts collide on a plane substratum. If spreading of seeded cells and cell locomotion are inhibited by the same mechanism, PRE cells should contact-inhibit choroid fibroblasts with which they collide, but not themselves be so inhibited. Using time-lapse cinemicrography, we have found this to be the case. We first established that in homotypic collisions, choroid fibroblasts do show contact inhibition of locomotion, despite the criss-cross (not well monolayered) appearance of confluent cultures. In heterotypic collisions between choroid and PRE we found the predicted nonreciprocal behaviour: the choroid leading edge is paralysed on collision, and the cell subsequently retracts, whereas the active PRE margin appears to be completely unaffected. Speed measurements from a series of such collisions show that the speed of choroid cells is markedly reduced on collision with PRE, whereas the slight slowing of PRE is not statistically significant. We have observed similar behaviour in heterotypic collisions between various epithelial and fibroblastic cells, and so it seems possible that non-reciprocity may prove general for this interaction. If so, it has important implications for the role of contact inhibition of locomotion in phenomena such as morphogenesis, wound healing and the invasiveness of carcinoma cells. On the one hand, non-reciprocal contact-inhibition of locomotion may permit the spreading of epithelia over mesenchymal cells, thus generating or restoring an epithelial bounding membrane. On the other hand, in the absence of other interactions, it would fail to inhibit the invasion of mesenchymal territory by aberrant epithelial cells, or presumably by epithelial free edges.

Epithelial-fibroblastic organization in cultures grown from human embryonic kidney: its significance for morphogenesis in vivo

1979 ◽  
Vol 39 (1) ◽  
pp. 291-298
Author(s):  
J.B. Bard
Keyword(s):  
Free Surface ◽  
Epithelial Cells ◽  
Functional Properties ◽  
Human Embryonic Kidney ◽  
Tissue Differences ◽  
Cultured Epithelia ◽  
Fibroblastic Cells ◽  
Different Tissues

The morphological behaviour of explants of human embryonic kidney has been studied in order to investigate the rules governing interactions between epithelial and fibroblastic cells from the same tissue. When the fragments are cultured, epithelia migrate out first and are followed, a few days later, by cables which grow out from the fragments and which are composed of multilayered fibroblasts. These cables extend through the epithelia to reach the substratum, to which they adhere. The epithelia maintain an upper surface free of spread fibroblasts and are unable to multilayer, although occasional rounded-up cells adhere to this surface. Fibroblasts, however, not only multilayer in the cables but can act as a substratum for epithelial cells which migrate on the cable surface. Fibroblasts and epithelia from kidney thus follow the same behavioural rules that govern the interactions between kidney epithelia and fibroblasts from different tissues. The suggestion that these rules derive from tissue differences and that cells from the same tissue are more tolerant of one another is not borne out. These observations and those reported by others are interpreted in terms of the functional properties of the cells in vivo. It is further pointed out that only those epithelia that maintain a free surface in vivo would be expected to show this property in vitro. Finally, the implications of cells with essentially the same properties generating more than one structure are considered.

Locomotory invasion of human cervical epithelium and avian fibroblasts by HeLa cells in vitro

1982 ◽  
Vol 57 (1) ◽  
pp. 293-314 ◽  
Author(s):  
E.M. Stephenson
Keyword(s):  
Epithelial Cells ◽  
Hela Cells ◽  
Time Lapse ◽  
Tumour Cells ◽  
Contact Inhibition ◽  
Type I ◽  
Original Direction ◽  
Epithelial Sheet ◽  
Invasion Index

The locomotory invasive ability of HeLa cells was tested against: (a) embryonic chick heart fibroblasts (CHF); and (b) normal epithelial cells from human cervix (HCE) in explant confrontations. Data for analyses were obtained from replicate cultures fixed 24 h after junction and from 24-h time-lapse films. The mean invasion index for HeLa versus CHF did not indicate significant obstruction but analyses of hourly radial advance and orientation frequencies showed that obstruction eventually developed as postjunctional incubation time increased. Early contacts between HeLa and CHF demonstrated non-reciprocity of type I contact inhibition of locomotion by the tumour cells, which continued moving in their original direction to underlap contact-inhibited fibroblasts and eventually to occupy spaces vacated by them. When CHF population density increased and free space diminished, HeLa cells displayed directional and probably substrate-dependent contact inhibition. The high invasion index of HeLa versus HCE was largely due to occupation of previous HCE territory by tumour cells and only occasionally to actual infiltration of the epithelial sheet. After contact with HeLa, ruffling substrate-adherent marginal epithelial cells displayed contractile, type I contact inhibition of locomotion. After orientation changes, they gradually retreated. Against HCE, HeLa cells exhibited non-reciprocity of type I contact inhibition and continued radially forward, following the retreating epithelial margin. They did not move onto exposed upper surfaces of epithelial cells and did not underlap marginal cells firmly adherent to the substratum. Invasion of the epithelial sheet was seen only when initial access beneath a cell with a non-adherent margin was available. The contact relationships of isolated invading HeLa cells with their epithelial neighbours suggested successive non-reciprocal contact inhibition reactions.

scholarly journals Regulation of Myosin II Dynamics by Phosphorylation and Dephosphorylation of Its Light Chain in Epithelial Cells

2007 ◽  
Vol 18 (2) ◽  
pp. 605-616 ◽  
Author(s):  
Toshiyuki Watanabe ◽  
Hiroshi Hosoya ◽  
Shigenobu Yonemura
Keyword(s):  
Epithelial Cells ◽  
Light Chain ◽  
Fluorescent Protein ◽  
Myosin Ii ◽  
Time Lapse ◽  
Stress Fibers ◽  
Interacting Protein ◽  
Cytoskeleton Organization

Nonmuscle myosin II, an actin-based motor protein, plays an essential role in actin cytoskeleton organization and cellular motility. Although phosphorylation of its regulatory light chain (MRLC) is known to be involved in myosin II filament assembly and motor activity in vitro, it remains unclear exactly how MRLC phosphorylation regulates myosin II dynamics in vivo. We established clones of Madin Darby canine kidney II epithelial cells expressing MRLC-enhanced green fluorescent protein or its mutants. Time-lapse imaging revealed that both phosphorylation and dephosphorylation are required for proper dynamics of myosin II. Inhibitors affecting myosin phosphorylation and MRLC mutants indicated that monophosphorylation of MRLC is required and sufficient for maintenance of stress fibers. Diphosphorylated MRLC stabilized myosin II filaments and was distributed locally in regions of stress fibers where contraction occurs, suggesting that diphosphorylation is involved in the spatial regulation of myosin II assembly and contraction. We further found that myosin phosphatase or Zipper-interacting protein kinase localizes to stress fibers depending on the activity of myosin II ATPase.

scholarly journals DENND2B activates Rab13 at the leading edge of migrating cells and promotes metastatic behavior

2015 ◽  
Vol 208 (5) ◽  
pp. 629-648 ◽  
Author(s):  
Maria S. Ioannou ◽  
Emily S. Bell ◽  
Martine Girard ◽  
Mathilde Chaineau ◽  
Jason N.R. Hamlin ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Cancer Progression ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Leading Edge ◽  
Cell Periphery ◽  
Nucleotide Exchange ◽  
Migrating Cells

The small guanosine triphosphatase Rab13 functions in exocytic vesicle trafficking in epithelial cells. Alterations in Rab13 activity have been observed in human cancers, yet the mechanism of Rab13 activation and its role in cancer progression remain unclear. In this paper, we identify the DENN domain protein DENND2B as the guanine nucleotide exchange factor for Rab13 and develop a novel Förster resonance energy transfer–based Rab biosensor to reveal activation of Rab13 by DENND2B at the leading edge of migrating cells. DENND2B interacts with the Rab13 effector MICAL-L2 at the cell periphery, and this interaction is required for the dynamic remodeling of the cell’s leading edge. Disruption of Rab13-mediated trafficking dramatically limits the invasive behavior of epithelial cells in vitro and the growth and migration of highly invasive cancer cells in vivo. Thus, blocking Rab13 activation by DENND2B may provide a novel target to limit the spread of epithelial cancers.

Invasive locomotory behaviour between malignant human melanoma cells and normal fibroblasts filmed in vitro

1978 ◽  
Vol 32 (1) ◽  
pp. 389-418
Author(s):  
E.M. Stephenson ◽  
N.G. Stephenson
Keyword(s):  
Human Fibroblasts ◽  
Leading Edge ◽  
Time Lapse ◽  
Melanoma Cells ◽  
Primary Objective ◽  
Human Melanoma ◽  
Contact Inhibition ◽  
Human Malignant Melanoma ◽  
Human Melanoma Cells

Explants of human malignant melanoma (MM96), normal adult human skin fibroblasts (HSF) and embryonic chick ventricle were confronted in pairs. The 2 outwandering populations in each confrontation eventually met in a situation where each could potentially invade the other. The human explants were artificially prepared from dissociated cells. The primary objective of the study was to compare the relative invasive capacities of the malignant and nonmalignant human cell populations against a standard population of chick heart fibroblasts (CHF). Relative invasiveness was also compared for (a) malignant human melanoma cells against human and avian fibroblasts; (b) the 2 different fibroblast populations against MM96; and (c) the 2 fibroblast populations against each other. Time-lapse films were prepared for each confrontation. Each cell population was also filmed in a free or unconfronted state. The films were analysed in terms of (a) cell speeds in relation to numbers of contacts; (b) the net radial outward velocity; (c) directional frequencies of movements; and (d) cell behavior resulting from heterologous collisions. Replicate cultures were fixed and stained 24–40 h after junction. Measurements from these indicated relative distances travelled by each population towards the opposing population and towards free space. Nuclear overlap and population density measurements were also recorded. MM96 cells invaded the standard chick fibroblast population but eventually encountered moderate obstruction. Invasion was mainly by oriented movement between successive pairs of chick cells. Contact paralysis of ruffling of MM96 was not seen. Human fibroblasts did not invade the CHF population. They were contact inhibited by chick cells and typically reversed their direction of movement. MM96 cells invaded the human fibroblast population without obstruction. They were not usually inhibited by HSF and several different pathways for locomotory invasion, including overlapping, were involved. Cells of both fibroblast populations were contact inhibited by leading-edge collisions with MM96 cells. Fibroblast invasion of MM96 was by underlapping of orthogonally oriented, bipolar melanocytes or dendritic processes. Human fibroblasts were less obstructed than chick fibroblasts during invasion of MM96. Initiation of invasion of HSF by chick cells was apparently orientation dependent. Invasion by CHF was assisted by the alignment, morphology and contact-mediated withdrawal of the human cells. Heterologous contact inhibition was not apparently defective for either cell type. In all confrontations involving both malignant and non-malignant cells, the extent and pattern of invasion could be related to apparent deficiencies in heterologous contact inhibition of locomotion.

Ultrastructure of melanocyte-keratinocyte interactions and pigment transfer in vitro

1978 ◽  
Vol 36 (2) ◽  
pp. 650-651
Author(s):  
Raul I. Garcia ◽  
Evelyn A. Flynn ◽  
George Szabo
Keyword(s):  
Direct Injection ◽  
Skin Pigmentation ◽  
Freeze Fracture ◽  
Pigment Granule ◽  
Time Lapse ◽  
Ultrastructural Level ◽  
Lanthanum Tracer ◽  
A Cell

Skin pigmentation in mammals involves the interaction of epidermal melanocytes and keratinocytes in the structural and functional unit known as the Epidermal Melanin Unit. Melanocytes(M) synthesize melanin within specialized membrane-bound organelles, the melanosome or pigment granule. These are subsequently transferred by way of M dendrites to keratinocytes(K) by a mechanism still to be clearly defined. Three different, though not necessarily mutually exclusive, mechanisms of melanosome transfer have been proposed: cytophagocytosis by K of M dendrite tips containing melanosomes, direct injection of melanosomes into the K cytoplasm through a cell-to-cell pore or communicating channel formed by localized fusion of M and K cell membranes, release of melanosomes into the extracellular space(ECS) by exocytosis followed by K uptake using conventional phagocytosis. Variability in methods of transfer has been noted both in vivo and in vitro and there is evidence in support of each transfer mechanism. We Have previously studied M-K interactions in vitro using time-lapse cinemicrography and in vivo at the ultrastructural level using lanthanum tracer and freeze-fracture.

Three-dimensional Growth of Renal Epithelial Cells in Vitro: A Tool in Toxicity Testing

1993 ◽  
Vol 21 (2) ◽  
pp. 191-195 ◽  
Author(s):  
Knut-Jan Andersen ◽  
Erik Ilsø Christensen ◽  
Hogne Vik
Keyword(s):  
Epithelial Cells ◽  
Three Dimensional ◽  
Toxicity Testing ◽  
Renal Tissue ◽  
Epithelial Cell Line ◽  
Multicellular Spheroids ◽  
Renal Epithelial Cell ◽  
Renal Epithelial Cells

The tissue culture of multicellular spheroids from the renal epithelial cell line LLC-PK1 (proximal tubule) is described. This represents a biological system of intermediate complexity between renal tissue in vivo and simple monolayer cultures. The multicellular structures, which show many similarities to kidney tubules in vivo, including a vectorial water transport, should prove useful for studying the potential nephrotoxicity of drugs and chemicals in vitro. In addition, the propagation of renal epithelial cells as multicellular spheroids in serum-free culture may provide information on the release of specific biological parameters, which may be suppressed or masked in serum-supplemented media.

ErbB-2 Induces the Cyclin D1 Gene in Prostate Epithelial Cells In vitro and In vivo

2007 ◽  
Vol 67 (9) ◽  
pp. 4364-4372 ◽  
Author(s):  
Mathew Casimiro ◽  
Olga Rodriguez ◽  
Llana Pootrakul ◽  
Maral Aventian ◽  
Nadia Lushina ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Cyclin D1 ◽  
Prostate Epithelial Cells
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