Contraction waves in lymphocyte locomotion

1984 ◽  
Vol 68 (1) ◽  
pp. 227-241
Author(s):  
W.S. Haston ◽  
J.M. Shields
Keyword(s):  
Three Dimensional ◽  
Collagen Gel ◽  
Time Lapse ◽  
Collagen Gels ◽  
Collagen Network ◽  
Longitudinal Contraction ◽  
Constriction Ring ◽  
Motile Cells ◽  
Transmission Electron ◽  
Contraction Wave

In this paper we propose that the constriction ring, a prominent feature of moving leucocytes, is a major source of locomotive force. Analysis of time-lapse films of lymphocytes in suspension and moving through three-dimensional collagen gels, demonstrated that the constriction ring was the morphological manifestation of a wave of circular contraction that moved antero-posteriorly. In lymphocytes in suspension the wave moved, although the cells could not. Analysis of lymphocytes moving through a collagen gel revealed that the waves remained stationary with respect to the external environment while the cell appeared to move forward through them. Passage of a single equatorial contraction wave resulted in cell lengthening: a shortening of the region posterior to the constriction was observed in cells moving through collagen gels, but not in lymphocytes held in suspension, suggesting that attachment of cells to the collagen network was necessary for longitudinal contraction. Lymphocyte attachment to collagen gels was mediated through the rapid extension of bleb-like structures into the collagen network. Transmission electron microscopy (TEM) failed to demonstrate any organized structure at the constriction ring. NBD-Phallacidin staining of lymphocytes together with TEM demonstrated that F-actin was distributed evenly throughout the length of the cell. Cell polarity was clearly recognizable by the distribution of coated vesicles, microvilli, and all organelles to the rear, and Thy 1–2 to the front, of motile cells, but polarity could be reversed by the passage of a single contraction wave starting at the rear of the cell, without prior redistribution of these structures.

A three-dimensional collagen network for cartilage reconstruction

Development ◽  
1971 ◽  
Vol 26 (2) ◽  
pp. 157-167
Author(s):  
S. Moskalewski ◽  
M. Kamiński ◽  
A. Dukwicz
Keyword(s):  
Three Dimensional ◽  
Collagen Gel ◽  
Collagen Gels ◽  
Collagen Network ◽  
Chondrocyte Cultures ◽  
Dimensional Network ◽  
Ehrlich Ascites ◽  
Cartilage Reconstruction ◽  
Ehrlich Ascites Tumour ◽  
Ascites Tumour Cells

To obtain a three-dimensional network which would support cells during culture, cold collagen solution was mixed with cells and converted into a gel at 37 °C. Gelation of collagen did not influence cell viability. The development of chondrocyte cultures in collagen gels depended on the distance between the cells. Single chondrocytes were surrounded by a mucopolysaccharide ring. The collagen fibres in parts of the cultures with moderate cell density were strongly alcian-blue-positive. Chondrocytes in crowded areas of cultures formed a cartilage matrix. Collagen gel with Ehrlich ascites tumour cells and liver cells remained unchanged after cultivation. Cultures of kidney cells and some chondrocyte cultures shrivelled, owing to partial collagen digestion. The delicate primary collagen network in some cultures partially transformed to much thicker, long fibres or to distinct capsules around chondrocytes.

F-actin distribution in polymorphonuclear leucocytes

1987 ◽  
Vol 88 (4) ◽  
pp. 495-501
Author(s):  
W.S. Haston
Keyword(s):  
Three Dimensional ◽  
Time Lapse ◽  
Polymorphonuclear Leucocytes ◽  
Two Dimensional ◽  
Collagen Gels ◽  
Motile Cells ◽  
Low Concentrations ◽  
Chemotactic Factors ◽  
Basic Features ◽  
Cytoskeletal Elements

The polarization and amoeboid locomotion of neutrophil leucocytes is stimulated by chemotactic factors, which initiate waves of contraction in both adherent and non-adherent neutrophils. These cyclical contractile events have previously been analysed by time-lapse filming but the mechanisms involved in the coordination of the cytoskeleton during locomotion have not been elucidated, one reason being because of the problems involved in fixing motile cells. In this paper we show that improved fixation of motile neutrophils with low concentrations of glutaraldehyde followed by glycine quenching demonstrated significant differences in the pattern of staining with TRITC-phalloidin in neutrophils moving on different substrata. Previous film analysis had shown the basic features of locomotion to be similar on all substrata. A prominent feature of leucocyte locomotion on two-dimensional substrata (e.g. protein-coated glass), on three-dimensional collagen gels or in motile cells floating in suspension, is the wave of contraction that passes antero-posteriorly along the length of the cell. The organization of the cytoskeletal elements has not been demonstrated at contraction waves, but light fixation with glutaraldehyde followed by staining with TRITC-phalloidin demonstrated prominent bands of F-actin in neutrophils inside collagen gels. These bands were not present in neutrophils either in suspension or moving on a two-dimensional substratum. Although all motile neutrophils had brightly stained anterior lamellipodia, the cells moving on the two-dimensional substratum had very extensively ruffled leading lamellae stained very brightly with TRITC-phalloidin. The reasons for the absence of consistent bands of F-actin at contraction waves are discussed.

Effect of shear stress on microvessel network formation of endothelial cells with in vitro three-dimensional model

2004 ◽  
Vol 287 (3) ◽  
pp. H994-H1002 ◽  
Author(s):  
Akinori Ueda ◽  
Masaki Koga ◽  
Mariko Ikeda ◽  
Susumu Kudo ◽  
Kazuo Tanishita
Keyword(s):  
Endothelial Cells ◽  
Shear Stress ◽  
Network Formation ◽  
Three Dimensional ◽  
Collagen Gel ◽  
Flow Chamber ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Collagen Gels

Shear stress stimulus is expected to enhance angiogenesis, the formation of microvessels. We determined the effect of shear stress stimulus on three-dimensional microvessel formation in vitro. Bovine pulmonary microvascular endothelial cells were seeded onto collagen gels with basic fibroblast growth factor to make a microvessel formation model. We observed this model in detail using phase-contrast microscopy, confocal laser scanning microscopy, and electron microscopy. The results show that cells invaded the collagen gel and reconstructed the tubular structures, containing a clearly defined lumen consisting of multiple cells. The model was placed in a parallel-plate flow chamber. A laminar shear stress of 0.3 Pa was applied to the surfaces of the cells for 48 h. Promotion of microvessel network formation was detectable after ∼10 h in the flow chamber. After 48 h, the length of networks exposed to shear stress was 6.17 (±0.59) times longer than at the initial state, whereas the length of networks not exposed to shear stress was only 3.30 (±0.41) times longer. The number of bifurcations and endpoints increased for networks exposed to shear stress, whereas the number of bifurcations alone increased for networks not exposed to shear stress. These results demonstrate that shear stress applied to the surfaces of endothelial cells on collagen gel promotes the growth of microvessel network formation in the gel and expands the network because of repeated bifurcation and elongation.

The Effect of Mechanical Constraints on Gelatin Samples under Pulsatile Flux

2012 ◽  
Vol 706-709 ◽  
pp. 449-454
Author(s):  
Eugenia Blangino ◽  
Martín A. Cagnoli ◽  
Ramiro M. Irastorza ◽  
Fernando Vericat
Keyword(s):  
Mechanical Properties ◽  
Tissue Engineering ◽  
Collagen Gel ◽  
Biological Tissues ◽  
Mechanical Stimuli ◽  
Collagen Gels ◽  
Collagen Network ◽  
Mechanical Constraints ◽  
Preparation Conditions ◽  
Biological Compatibility

It is of great interest in tissue engineering the role of collagen gel-based structures (scaffolds, grafts and-by cell seeded and maturation-tissue equivalents (TEs) for several purposes). It is expected the appropriate biological compatibility when the extracellular matrix (ECM) is collagen-based. Regarding the mechanical properties (MP), great efforts in tissue engineering are focused in tailoring TE properties by controlling ECM composition and organization. When cells are seeded, the collagen network is remodeled by cell-driven compaction and consolidation, produced mainly through the mechanical stimuli that can be directed selecting the geometry and the surfaces exposed to the cells. Collagen gels have different (chemical and mechanical) properties depending on their origin and preparation conditions. The MP of the collagen network are derived from the degree of cross-linking (CLD) which can be modified by different treatments. One of the techniques to evaluate MP in the network is by ultrasound (US). In this work we analyse the effect of several mechanical constraints (similar to that imposed to promote cell growth on certain sample surfaces, when seeded) on samples of gelatin with a specific geometry (thick walls cylinders) under loading conditions of pulsatile flow. We checked US parameters and estimates evolution of the network structure for different restrictions in the sample mobility. It was implemented by adapting devices specially built to measure elastic properties of biological tissues by US. The material (origin and purity) and the preparation conditions for the gelatin were selected in order to compare the results with those of literature.

Three-dimensional reconstruction of cell morphology using intermediate voltage electron microscopy and computer graphics

1987 ◽  
Vol 45 ◽  
pp. 590-591 ◽  
Author(s):  
Julian P. Heath ◽  
Ming Hsiu Ho ◽  
Lee D. Peachey
Keyword(s):  
Electron Microscopy ◽  
Three Dimensional ◽  
University Of Pennsylvania ◽  
Stereoscopic Display ◽  
Collagen Gels ◽  
Biomedical Image ◽  
Voltage Electron ◽  
Specimen Holder ◽  
Transmission Electron ◽  
Dimensional Reconstruction

The Intermediate Voltage Electron Microscopy and Biomedical Image Analysis facility at the University of Pennsylvania is a National Resource supported by NIH and provides users with facilities for transmission electron microscopy and digital image processing. The facility comprises a JEOL 4000EX transmission electron microscope with a 360 degree goniometer specimen holder, a VAX 11/750 computer, and a Raster Technologies Inc. Model One/380 Graphics work station with two high resolution 1280x1024 pixel color video display monitors for stereoscopic display. We are using this facility to examine the morphology of fibroblasts migrating through fibrillar collagen gels: these matrices closely model the environments encountered by migratory cells during embryonic morphogenesis.The identification and three dimensional localization of structures in stereoscopic electron micrographs of thick sections of cells can be hampered by diffuse boundaries and by the superposition of details with similar electron density.

Effects of culture conditions on the proliferation, morphology and migration of bovine aortic endothelial cells

1983 ◽  
Vol 62 (1) ◽  
pp. 267-285
Author(s):  
A.M. Schor ◽  
S.L. Schor ◽  
T.D. Allen
Keyword(s):  
Endothelial Cells ◽  
Three Dimensional ◽  
Collagen Gel ◽  
Culture Conditions ◽  
Apical Surface ◽  
Collagen Gels ◽  
Aortic Endothelial Cells ◽  
Plating Density ◽  
Adhesive Interactions ◽  
And Migration

Various culture conditions, such as the presence of ascorbic acid, initial plating density and the nature of the substratum (plastic, gelatin or native collagen gels), influenced the growth, morphology and migration of three cloned populations of adult bovine aorta endothelial cells. Aorta endothelial cells showed two distinctive and reversible morphological phenotypes. Cells presenting a free apical surface were polygonal and formed sheets of overlapping or non-overlapping cells, depending on the culture conditions. When the cells were able to establish adhesive interactions over their entire cell surface they adopted an elongated shape and formed meshworks of interconnected ‘sprouting’ cells. The cells were capable of migrating into a collagen gel from both their basal and apical surfaces. Once in the gel, they formed characteristic, compact, three-dimensional meshworks.

Sodium nitroprusside augments human lung fibroblast collagen gel contraction independently of NO-cGMP pathway

2000 ◽  
Vol 278 (5) ◽  
pp. L1032-L1038 ◽  
Author(s):  
X. D. Liu ◽  
C. M. Skold ◽  
T. Umino ◽  
J. R. Spurzem ◽  
D. J. Romberger ◽  
...  
Keyword(s):  
Type I Collagen ◽  
Human Lung ◽  
Three Dimensional ◽  
Collagen Gel ◽  
Lung Fibroblast ◽  
Type I ◽  
Collagen Gels ◽  
Human Lung Fibroblast ◽  
Collagen Gel Contraction ◽  
Cgmp Pathway

Nitric oxide (NO) relaxes vascular smooth muscle in part through an accumulation of cGMP in the target cells. We hypothesized that a similar effect may also exist on collagen gel contraction mediated by human fetal lung (HFL1) fibroblasts, a model of wound contraction. To evaluate this, HFL1 cells were cultured in three-dimensional type I collagen gels and floated in serum-free DMEM with and without various NO donors. Gel size was measured with an image analyzer. Sodium nitroprusside (SNP, 100 μM) significantly augmented collagen gel contraction by HFL1 cells (78.5 ± 0.8 vs. 58.3 ± 2.1, P < 0.01), whereas S-nitroso- N-acetylpenicillamine, 5-amino-3-(4-morpholinyl)-1,2,3-oxadiazolium chloride, NONOate, and N G-monomethyl-l-arginine did not affect the contraction. Sodium ferricyanide, sodium nitrate, or sodium nitrite was not active. The augmentory effect of SNP could not be blocked by 1 H-[1,2,4]-oxadiazolo-[4,3- a]-quinoxalin-1-one, whereas it was partially reversed by 8-(4-chlorophenylthio) (CPT)-cGMP. To further explore the mechanisms by which SNP acted, fibronectin and PGE2 production were measured by immunoassay after 2 days of gel contraction. SNP inhibited PGE2 production and increased fibronectin production by HFL1 cells in a concentration-dependent manner. CPT-cGMP had opposite effects on fibronectin and PGE2 production. Addition of exogenous PGE2 blocked SNP-augmented contraction and fibronectin production by HFL1 cells. Therefore, SNP was able to augment human lung fibroblast-mediated collagen gel contraction, an effect that appears to be independent of NO production and not mediated through cGMP. Decreased PGE2 production and augmented fibronectin production may have a role in this effect. These data suggest that human lung fibroblasts in three-dimensional type I collagen gels respond distinctly to SNP by mechanisms unrelated to the NO-cGMP pathway.

scholarly journals Epithelia suspended in collagen gels can lose polarity and express characteristics of migrating mesenchymal cells.

1982 ◽  
Vol 95 (1) ◽  
pp. 333-339 ◽  
Author(s):  
G Greenburg ◽  
E D Hay
Keyword(s):  
Epithelial Cells ◽  
Cell Polarity ◽  
Three Dimensional ◽  
Collagen Gel ◽  
Mesenchymal Cells ◽  
Apical Surface ◽  
Collagen Gels ◽  
Mesenchymal Transformation

This study of epithelial-mesenchymal transformation and epithelial cell polarity in vitro reveals that environmental conditions can have a profound effect on the epithelial phenotype, cell shape, and polarity as expressed by the presence of apical and basal surfaces. A number of different adult and embryonic epithelia were suspended within native collagen gels. Under these conditions, cells elongate, detach from the explants, and migrate as individual cells within the three-dimensional lattice, a previously unknown property of well-differentiated epithelia. Epithelial cells from adult and embryonic anterior lens were studied in detail. Elongated cells derived from the apical surface develop pseudopodia and filopodia characteristic of migratory cells and acquire a morphology and ultrastructure virtually indistinguishable from that of mesenchymal cells in vivo. It is concluded from these experiments that the three-dimensional collagen gel can promote dissociation, migration, and acquisition of secretory organelles by differentiated epithelial cells, and can abolish the apical-basal cell polarity characteristic of the original epithelium.

scholarly journals ROCK-generated contractility regulates breast epithelial cell differentiation in response to the physical properties of a three-dimensional collagen matrix

2003 ◽  
Vol 163 (3) ◽  
pp. 583-595 ◽  
Author(s):  
Michele A. Wozniak ◽  
Radhika Desai ◽  
Patricia A. Solski ◽  
Channing J. Der ◽  
Patricia J. Keely
Keyword(s):  
Epithelial Cells ◽  
Three Dimensional ◽  
Collagen Gel ◽  
Collagen Matrix ◽  
Breast Epithelial Cell ◽  
Collagen Gels ◽  
Culture Dish ◽  
Breast Epithelial Cells ◽  
3D Collagen ◽  
Breast Epithelial

Breast epithelial cells differentiate into tubules when cultured in floating three-dimensional (3D) collagen gels, but not when the cells are cultured in the same collagen matrix that is attached to the culture dish. These observations suggest that the biophysical properties of collagenous matrices regulate epithelial differentiation, but the mechanism by which this occurs is unknown. Tubulogenesis required the contraction of floating collagen gels through Rho and ROCK-mediated contractility. ROCK-mediated contractility diminished Rho activity in a floating 3D collagen gel, and corresponded to a loss of FAK phosphorylated at Y397 localized to 3D matrix adhesions. Increasing the density of floating 3D collagen gels also disrupted tubulogenesis, promoted FAK phosphorylation, and sustained high Rho activity. These data demonstrate the novel finding that breast epithelial cells sense the rigidity or density of their environment via ROCK-mediated contractility and a subsequent down-regulation of Rho and FAK function, which is necessary for breast epithelial tubulogenesis to occur.

scholarly journals Skin fibroblasts obtained from cancer patients display foetal-like migratory behaviour on collagen gels

1985 ◽  
Vol 73 (1) ◽  
pp. 235-244
Author(s):  
S.L. Schor ◽  
A.M. Schor ◽  
P. Durning ◽  
G. Rushton
Keyword(s):  
Cell Density ◽  
Normal Skin ◽  
Three Dimensional ◽  
Collagen Gel ◽  
Collagen Matrix ◽  
Skin Fibroblasts ◽  
Collagen Gels ◽  
Normal Range ◽  
Migratory Response ◽  
Fibroblast Migration

When plated on the surface of collagen gel substrata, all types of fibroblasts rapidly begin to migrate down into the three-dimensional collagen matrix. We have previously demonstrated that normal (adult and foreskin), foetal and transformed fibroblasts may be distinguished from each other by virtue of their differential migratory response to changes in cell density. The effects of cell density on fibroblast migration into the gel may be expressed by a single numerical value, the ‘cell density migration index’ (CDMI). We now present evidence that ostensibly normal skin fibroblasts obtained from the majority of patients we examined with carcinoma of the breast, malignant melanoma, familial polyposis coli, retinoblastoma and Wilms' tumours display aberrant CDMI values falling within the foetal range. Skin fibroblasts obtained from the majority of patients examined with genetic or chronic diseases (e.g. rheumatoid arthritis, Duchenne muscular dystrophy) displayed CDMI values falling within the normal range.

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