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Biomedical subjects

G A Dunn

Publications and source records attributed to G A Dunn.

At least 37 records · Page 2Linked to original sources
PubMed1991

A new direct-viewing chemotaxis chamber.

A new form of chamber for studying chemotaxis, similar in principle to the Zigmond chamber, allows the behaviour of the cells in a linear concentration gradient to be observed directly. The chamber was developed mainly for studying chemotaxis in fibroblasts using interferometric microscopy and the main design criteria were that it should have better optical characteristics, a higher dimensional precision and better long-term stability than the Zigmond chamber. It is made entirely from glass by grinding a blind circular well centrally in the counting platform of a Helber bacteria counting chamber. This procedure leaves an annular 'bridge', approximately 1 mm wide, between the new inner circular well and the original outer annular well. This bridge fulfils the same function as the linear bridge of the Zigmond chamber but the precise construction of the counting chamber ensures that a gap of 20 microns between bridge and coverslip can be accurately and repeatedly achieved when the chamber is assembled. It is envisaged that the improved optical clarity, dimensional accuracy and long-term stability of the new chamber will be advantageous in other applications, particularly in studies requiring critical microscopy or a precise knowledge of the gradient and in studies of cells, such as fibroblasts, that move much more slowly than neutrophils.

Chemotaxis↗
PubMed1989

A quantitative analysis of alterations in the shape of cultured fibroblasts induced by tumour-promoting phorbol ester.

Quantitative analysis of cell size and shape has shown that the tumour promotor 12-o-tetradecanoylphorbol-13-acetate (TPA) alters one of three parameters of cell shape, dispersion. It has no effect on cell area or elongation. This reflects the fact that TPA causes cells to form long narrow processes extending from the cell body. The ability to measure alterations in specific parameters of cell shape is important for correlating changes in the observed behaviour of cells with effects on cytoskeletal organisation.

Animals↗
PubMed1989

Microinterferometry of the movement of dry matter in fibroblasts.

We describe the use of interferometric microscopy coupled with a novel application of Sénarmont compensation for detecting and quantifying the distribution of dry matter in cultured cells. In conjunction with video techniques and digital image processing, a two-dimensional, calibrated map of the dry mass distribution in an isolated cell can be obtained and digitally recorded. We have called the technique Digitally Recorded Interferometric Microscopy with Analyser Shift (DRIMAS). The method greatly facilitates the automatic recognition of cells by computer. Recorded time-lapse sequences can be used to establish a database of the growth and motility of specific cells in given experimental conditions. Databases of this type can be analysed to reveal the patterns of growth and locomotory behaviour of individual cells. We describe a systematic method of obtaining parameters of cell size, shape, spreading, intracellular motility and translocation. Auto-correlations and cross-correlations between these parameters can be detected and quantified using time series analysis, revealing potential cause/effect relationships in the mechanisms of growth and motility. Besides characterizing the overall pattern of cell behaviour, these data can also yield information about the instantaneous pattern of intracellular motility. We describe the use of finite element analysis to reveal the dynamics of the intracellular transport of dry matter. This yields the pattern of the minimum flow of dry matter required to account for the changes in its distribution. Most of this flux is not associated with the movement of visible structures and possibly represents the transport of dissociated components of the cytoskeleton. In chick heart fibroblasts, surprisingly high velocities of nearly 2.0 microns s-1 were detected during the period of increased motility following tail detachment. The total kinetic energy associated with the dry mass flux is a single parameter, which characterizes the instantaneous motility of the cell. We found that the kinetic energy of intracellular motility can be several hundred times greater than the kinetic energy of translocation. Kinetic energy may prove to be a very informative single measure of intracellular motility for assessing the effects of malignant transformation, genetic manipulations, and other experimental treatments on the locomotory machinery of the cell.

Animals↗
PubMed1987

A unified approach to analysing cell motility.

The quantitative analysis of cell motility in culture has several important functions. First, it gives a concise and accurate description of the motile process and can detect subtle differences in motility due to different genetic makeup or experimental conditions. Second, its objectivity means that results can be communicated precisely and used unambiguously to test hypotheses about motility. Third, it may be used to derive a mathematical model with the same statistical properties as the motile process and thus elucidate the mechanism of motility. In this paper, we introduce a general procedure for analysing cell motility in a wide variety of circumstances. We describe a pilot project for the analysis of simple geometrical data obtained from randomly moving fibroblasts. Finally, as an example, we show how an analysis of the translocation of the fibroblasts can lead to insights into the mechanism of motility that are arguably not obtainable by any other approach.

Animals↗
PubMed1986

Analysing collisions between fibroblasts and fibrosarcoma cells: fibrosarcoma cells show an active invasionary response.

We describe a direct way of measuring contact inhibition of locomotion by analysing the changes in motion of pairs of colliding cells. This allows values to be assigned to each type of cell in mixed collisions and will enable certain hypotheses about the relationship between contact inhibition and invasion in culture to be tested critically. We find that fibrosarcoma (FS9) cells, on colliding with chick heart fibroblasts, show a reversed contact-inhibition response that we call contact promotion of locomotion. We also describe a measure of the lateral changes in motion that result from collisions between cells and show that this is dependent on the type of colliding cell but, unlike contact inhibition, it does not appear to be dependent on the type of cell with which it collides for the types studied here. Finally, we analyse how the total response is dependent on the dispositions and motions of the cells before collision and we find that FS9 cells, on colliding with fibroblasts, tend to turn towards the point of initial marginal contact. We conclude that the FS9 cells show a pronounced response on colliding with the fibroblasts, which is in contrast to the subjective impression that the FS9 cells do not respond much. These findings support the thesis of Abercrombie and colleagues, that the infiltration of a population of normal cells by a population of invasive cells in culture is dependent on the nature of the response of each cell type to collision with the other and that the invasive cells fail to show contact inhibition in these heterotypic collisions; but the findings further suggest that these invasive cells show an active invasionary response as opposed to merely failing to show contact inhibition.

Animals↗
PubMed1986

Alignment of fibroblasts on grooved surfaces described by a simple geometric transformation.

The response of chick heart fibroblasts to grooved substrata was studied using microfabricated grooves and new measures of shape and alignment derived from the moments of cell shapes. Cell shape and alignment were measured on 23 different sets of regular, parallel grooves, which ranged in width from 1.65 to 8.96 micron, and in repeat spacing from 3.0 to 32.0 micron. The grooves were of constant depth, 0.69 micron. Digitized video images were analysed to extract the zero-, first- and second-order moments of the cell shapes, from which were calculated three measures of cell shape, and three measures of cell alignment. Regression analyses of the measures against parameters of the substratum such as groove width, repeat spacing and the ridge width between grooves show that ridge width is the main parameter affecting cell alignment (alignment being inversely proportional to ridge width), although groove width has a small additional effect. All the differences in cell shape between the different grooves can be summarized to a very good approximation as simple geometrical stretch transformations of the shapes of cells on planar surfaces. Our principal measure of cell alignment, paraxial elongation, is a measure of the necessary transformation. This finding has the interesting biological implication that the shape and orientation adopted by cells, in response to the grooves, are not governed by independent cellular mechanisms.

Animals↗
PubMed1985

A simple chamber for observing microscopic specimens in both top and side views.

A method is described which permits the observation of microscopic specimens in both top and side views, a rapid change between the two views being possible without disturbing the specimen. The system requires the introduction of simple optical components into a culture chamber, does not restrict the types of microscopy that may be used, and does not necessitate modification of the microscope itself. The technique is discussed in relation to the study of tissue cells in culture, but may find applications in other areas.

Culture Techniques↗
PubMed1984

Chemokinetic accumulation of human neutrophils on immune complex-coated substrata: analysis at a boundary.

The locomotory behavior of human blood neutrophil leukocytes was studied at a boundary between two surfaces with different chemokinetic properties. This was achieved by time-lapse cinematography of neutrophils moving on coverslips coated with BSA, then part-coated with immune complexes by adding anti-BSA IgG with a straight-line boundary between the BSA and the immune complexes. Cell locomotion was filmed in microscopic fields bisected by the boundary, and kinetic behavior was assessed by comparing speed (orthokinesis), turning behavior (klinokinesis), and the rate of diffusion of the cells on each side of the boundary, using a recently described mathematical analysis of kinesis. In the absence of serum or complement, the proportion of motile cells and their speed and rate of diffusion were greater on BSA than on antiBSA, but there was no consistent difference in turning behavior between cells on the two surfaces. The immune complexes were therefore negatively chemokinetic in comparison with BSA, and this resulted from a negative orthokinesis with little or no contribution from klinokinesis. As would be predicted theoretically, this resulted in gradual accumulation of cells on the immune complexes even in the absence of a chemotactic factor. In further studies, a parallel plate flow chamber was used to show that, under conditions of flow, neutrophils accumulated much more rapidly on a surface coated with BSA-anti-BSA than on BSA alone. Moreover, neutrophils on immune complex-coated surfaces lost their ability to form rosettes with IgG-coated erythrocytes. This suggests that neutrophils on immune complex-coated surfaces redistribute their Fc receptors (RFc gamma) to the under surface, and that the lowered speed of locomotion is due to tethering of neutrophils by substratum-bound IgG-Fc.

Antigen-Antibody Complex↗
PubMed1982

Analysing the motile behaviour of cells: a general approach with special reference to pairs of cells in collision.

As a measure of the change in motion of locomoting cells, acceleration has several advantages over more conventional measures such as rate of change in speed and rate of change in direction. In particular the instantaneous acceleration vector associated with a specific event, such as the collision of the cell with another, may be correlated with directional information contained in the structure of the cell and in the geometrical configuration of the event. It is hoped that this will prove to be a generally useful method for examining the relation between cell structure and motion during both normal and aberrant behaviour. Preliminary results obtained with this method to examine the changes in motion of a normal cell and a malignant cell on colliding with each other are already of interest for elucidating the mechanism of contact inhibition of locomotion and its failure during malignant invasion.

Animals↗