Time-based changes in fibroblast three-dimensional locomotory characteristics and phenotypes

Three-dimensional locomotory trajectories have been determined for an embryonic fibroblast population and also for these cells after a 20-day period in tissue culture (C-20 cells). Differences in locomotory characteristics between these two cell populations are reported. Applying factor analysis to the distribution of angle changes between vectors in a cell's trajectory reveals the possibility that different locomotory phenotypes exist. Of the three phenotypes detected in the embryonic population only one continues in the C-20 population while a new phenotype appears in the latter. These results document changes in locomotory characteristics and phenotypes in an embryonic population with time.     

Running worms: C. elegans self-sorting by electrotaxis

The nematode C. elegans displays complex dynamical behaviors that are commonly used to identify relevant phenotypes. Although its maintenance is straightforward, sorting large populations of worms when looking for a behavioral phenotype is difficult, time consuming and hardly quantitative when done manually. Interestingly, when submitted to a moderate electric field, worms move steadily along straight trajectories. Here, we report an inexpensive method to measure worms crawling velocities and sort them within a few minutes by taking advantage of their electrotactic skills. This method allows to quantitatively measure the effect of mutations and aging on worm's crawling velocity. We also show that worms with different locomotory phenotypes can be spatially sorted, fast worms traveling away from slow ones. Group of nematodes with comparable locomotory fitness could then be isolated for further analysis. C. elegans is a growing model for neurodegenerative diseases and using electrotaxis for self-sorting can improve the high-throughput search of therapeutic bio-molecules.     

Locomotory characteristics of Treponema denticola

Locomotion of pathogenic spirochetes has been suggested as a virulence factor in their pathogenesis. Little is known of the locomotory characteristics of oral anaerobic spirochetes. We have determined the optimal conditions for motility of seven strains of Treponema denticola in menstrua of different viscosities. The viscosity for optimum motility for all strains was found to be 9.57 centipoises at 25 degrees C. Under these conditions the average speeds for each strain was computed from the motility tracks as recorded by timed exposures under dark-field microscopy. Differences in speeds were found between the various strains. In addition, we have determined the "persistence" (direct distance/actual pathlength travelled) of cell movement of each strain. Interstrain differences were also noted. These locomotory characteristics contribute to the locomotory phenotypes of the various strains and therefore may aid in their characterization and provide an insight into locomotion as a virulence factor in periodontitis.     

Motility of murine lymphocytes during transit through cell cycle. Analysis by a new in vitro assay

The relationship between the basal (spontaneous) motility of murine lymphocytes and their position in the cell cycle was examined in a new collagen gel motility assay system. Concanavalin A-stimulated or control lymphocytes were allowed to locomote into slabs of type I collagen gel. The assay configuration permitted extraction of both total populations and locomotory subpopulations as viable, single-cell suspensions suitable for phenotypic and cell analysis. Concanavalin A stimulation resulted in a significant increase in the mean distance traveled by the leading cell front in 4 hr, from 23 microns (controls) to 67 microns. The estimated percentage of motile cells increased from 0.9 to 2.8%. Similar increases were observed after 18 hr of locomotion. The SIg+, Thy-1+, L3T4+, and Ly-2+ subsets exhibited equivalent increases in motility. Total populations and locomotory subpopulations were allowed to incorporate 5-bromo-2'-deoxyuridine, and their cell cycle profiles were compared by dual parameter anti-5-bromo-2'-deoxyuridine, propidium iodide fluorescence analysis. Total population and locomotory subpopulations did not differ significantly with respect to the ratio G0/G1:S, indicating that lymphocytes in these two phases exhibited approximately equal motility. Cells in late S and G2 + M were significantly less motile; locomotory subpopulations contained 60 to 75% fewer G2 + M cells than the total populations from which they were derived. Taken together, the results indicate that the concanavalin A-induced increase in motility commences before S phase and that motility diminishes shortly before or during G2 + M.     

Locomotory competence and laminin-specific cell surface binding sites are lost during myoblast differentiation

The specific interaction of embryonal cells with the extracellular matrix (ECM) is one of the principal forces influencing embryonal development (Hay, 1984; Trinkaus, 1984). We used a muscle satellite cell line (MM14dy) to determine the relationship between locomotory response to laminin and the expression of specific cell surface binding sites for it. Time lapse videomicroscopic analysis was used to study the locomotory response and radioligand binding assays and cell attachment assays were used to follow the expression levels of binding sites for laminin and its subfragments E8 and E1-4. We report here the novel finding that the ability of MM14dy to locomote over laminin diminishes and finally vanishes as the cells differentiate. The simultaneous drop in expression of binding sites for laminin is interpreted as being of potential significance during development and repair.     

Modulation of swimming speed in the pteropod mollusc,Clione limacina: Role of a compartmental serotonergic system

In locomotory systems, the central pattern generator and motoneuron output must be modulated in order to achieve variability in locomotory speed, particularly when speed changes are important components of different behavior acts. The swimming system of the pteropod molluscClione limacina is an excellent model system for investigating such modulation. In particular, a system of central serotonergic neurons has been shown to be intimately involved in regulating output of the locomotory pattern generator and motor system ofClione. There are approximately 27 pairs of serotonin-immunoreactive neurons in the central nervous system ofClione, with about 75% of these identified. The majority of these identified immunoreactive neurons are involved in various aspects of locomotory speed modulation. A symmetrical cluster of pedal serotonergic neurons serves to increase wing contractility without affecting wing-beat frequency or motoneuron activity. Two clusters of cerebral cells produce widespread responses that lead to an increase in pattern generator cycle frequency, recruitment of swim motoneurons, activation of the pedal serotonergic neurons and excitation of the heart excitor neuron. A pair of ventral cerebral neurons provides weak excitatory inputs to the swimming system, and strongly inhibits neurons of the competing whole-body withdrawal network. Overall, the serotonergic system inClione is compartmentalized so that each subsystem (usually neuron cluster) can act independently or in concert to produce variability in locomotory speed.     

Immunologic characterization of T-lymphoproliferative disorders by monoclonal antibodies

A series of 11 mouse monoclonal antibodies reactive with human T lymphoid cells at different stages of differentiation was used for immunological classification of leukaemic cells of 16 patients with T cell lymphoproliferative disorders by using a fluorescence assay. The majority of T-ALL cells had an immature or early thymic phenotype and T lymphoblastic lymphoma had phenotypes corresponding to different levels of more mature stages of T cell differentiation, Two cases of T-CLL and one adult patient with mycosis fungoides had mature T cell phenotypes being T-3+, T-4-, T-8+, cytotoxic/suppressor cell types and one case of T-CLL had T-3+, T-4+, T-8-, "helper/inducer" cell type, too. These results suggested that surface marker analysis in T cell lymphoproliferative disorders may be used as a highly reproducible immunological classification system that will provide additional information about phenotypes of leukaemic cells in connection with morphological analysis and clinical diagnosis.     

Effect of a mistletoe extract (Iscador® QuFrF) on viability and migratory behavior of human peripheral CD4+ and CD8+ T lymphocytes in three-dimensional collagen lattices

Summary Migration and tissue distribution of immunocompetent cells may be critical prerequisites for efficient immune surveillance. The effect of various concentrations of the mistletoe extract Iscador? QuFrF on the locomotory behavior and viability of immunomagnetically isolated human CD4⁺ and CD8⁺ T lymphocytes within three-dimensional collagen gels was investigated. Although variation in baseline activities of spontaneously migrating T cells was donor-dependent, a dose-dependent stimulation of the locomotory activity in both CD4⁺ and CD8⁺ T cells for noncytotoxic concentrations of Iscador QuFrF (0.25–1.25 μg/ml) was detected. The optimal concentration of mistletoe extract and time of maximal response were specific for each donor. As shown by cell tracking and subsequent data analysis, CD4⁺ T cells exposed to the mistletoe extract displayed a significant increase in mean velocity and time locomoting; total distance migrated was nearly doubled. In contrast, CD8⁺ T cells showed less pronounced changes in these critical parameters. Cytotoxic effects of the mistletoe preparation on T lymphocytes, which could at least partially be attributed to the induction of apoptosis, were drastically reduced in the presence of fetal calf serum in the culture system. Our data suggest that the direct stimulation of T-cell migration in the presence of mistletoe components may modulate in a dose-dependent manner the system of immune surveillance and recognition in patients under mistletoe therapy.     

Behavior of cultured cells on substrata of variable adhesiveness

The locomotory behavior of tissue cells cultured on various artificial substrata was studied by time-lapse cinemicrography. Cells were able to spread more completely on certain more wettable substrata and to accumulate preferentially on these substrata according to a consistent hierarchy of cell-substratum affinity, which was the same for all cell types. Cell responses to variations in substrata suggest that substratum adhesiveness is the determining factor, but that cells accumulate on more adhesive substrata as the result of unequal competition between several actively locomotory ruffled lamellae around their margin. The increased overlapping between cells cultured on less adhesive substrata was found to be attributable to factors other than a decrease of contact inhibition of locomotion.     

Glycosylation genes expressed in seam cells determine complex surface properties and bacterial adhesion to the cuticle of Caenorhabditis elegans

The surface of the nematode Caenorhabditis elegans is poorly understood but critical for its interactions with the environment and with pathogens. We show here that six genes (bus-2, bus-4, and bus-12, together with the previously cloned srf-3, bus-8, and bus-17) encode proteins predicted to act in surface glycosylation, thereby affecting disease susceptibility, locomotory competence, and sexual recognition. Mutations in all six genes cause resistance to the bacterial pathogen Microbacterium nematophilum, and most of these mutations also affect bacterial adhesion and biofilm formation by Yersinia species, demonstrating that both infection and biofilm formation depend on interaction with complex surface carbohydrates. A new bacterial interaction, involving locomotory inhibition by a strain of Bacillus pumilus, reveals diversity in the surface properties of these mutants. Another biological property--contact recognition of hermaphrodites by males during mating--was also found to be impaired in mutants of all six genes. An important common feature is that all are expressed most strongly in seam cells, rather than in the main hypodermal syncytium, indicating that seam cells play the major role in secreting surface coat and consequently in determining environmental interactions. To test for possible redundancies in gene action, the 15 double mutants for this set of genes were constructed and examined, but no synthetic phenotypes were observed. Comparison of the six genes shows that each has distinctive properties, suggesting that they do not act in a linear pathway.     

Cytoskeletal organization in locomoting cells of the V2 rabbit carcinoma

The relationship between the organization of cytoskeletal elements and locomotory activity was studied in single cells of the V2 rabbit carcinoma. Like migratory fibroblasts, and unlike colony-forming epithelial cells, these cells show a pronounced horizontal polarization, and develop a large lamella at their leading front. With affinity-purified antibodies and a combination of light and electron microscopic techniques, actin and alpha-actinin (but not myosin and tropomyosin) were found highly concentrated within the marginal region of the leading lamella, both in ruffles and in the underlying zone of contacts with the substratum. Close contacts prevailed in the locomotory cells and small focal contacts developed only in cells detaching from others. Focal contacts always contained small microfilament bundles. Reorganization of actin filaments is suggested as the fundamental event for the dynamic contact formation of the leading lamella. Large microfilament bundles (stress fibers) were absent in all stages of locomotion.Since locomotory behavior and shape changes of V2 cells are the same on glass as on the surface of a natural membrane, the rabbit mesentery, organization and distribution of contractile elements of cultured V2 cells probably reflect the in vivo situation.     

Evolutionary and functional substitution of extrinsic musculature in Solifugae (Arachnida)

The locomotory system of Solifugae is distinct from that of other Arachnida in several ways. Only three pairs of legs are involved in locomotion, while the first pair function as sensory appendages. Morphologically, the proximal region of the locomotory system in Solifugae is characterized by fused coxae. Within the prosoma of Solifugae, an endosternite is missing: in other Arachnida, this endosternite serves as the proximal attachment site for a portion of the extrinsic musculature. How then do these skeletal modifications influence the muscular anatomy in the proximal region of the locomotory system? To answer this question, we studied the skeletomuscular anatomy of Galeodes granti at the interface between the prosoma and legs, reinvestigating the complex muscular anatomy of this body region for the first time in over 80 years and—for the first time—using detailed micro-computed tomography scans to analyze the skeletomuscular morphology. Specimens of three further species were checked for comparison. The analysis revealed differences in the number and composition of coxa-trochanter muscles in each of the four pairs of legs. These are compared in the light of serial homology. The comparison between the proximal locomotory system of Solifugae and that of other Arachnida unveils a series of analogies. Primarily, the coxa-trochanter joint is the most proximal joint to move the leg relative to the prosoma. Therefore, we argue that from a morpho-functional point of view, the coxa-trochanter muscles in Solifugae should be considered secondary extrinsic musculature. Thus, the legs gain a stable, articulated joint in the most proximal region of the leg to the prosoma, which might be advantageous for agile locomotion.     

The respiratory pattern in Drosophila melanogaster selected for desiccation resistance is not associated with the observed evolution of decreased locomotory activity

We examined spontaneous locomotory behavior and respiratory pattern in replicate outbred populations of Drosophila melanogaster selected for desiccation resistance or starvation resistance, as well as their control and ancestral populations. Use of these populations allows us to compare evolved behavioral changes in response to different stress selections. We also reasoned that previously observed changes in respiratory patterns following selection for increased desiccation resistance might be associated with or even caused by changes in locomotory behavior. We measured spontaneous locomotory behavior using video recordings and a computer-based tracking system while simultaneously measuring patterns of CO(2) release from single fruit flies. Statistically significant differences in behavior were observed to be correlated with selection regime. Reduced levels of spontaneous locomotory activity were observed in moist air in both desiccation- and starvation-selected populations compared with their controls. Interestingly, in dry air, only the desiccation-selected flies continue to show reduced spontaneous locomotory activity. No correlation was found between the level of locomotory activity of individual flies and the respiratory patterns of those flies, indicating that the reduced activity levels that have evolved in these flies did not directly cause the documented changes in their respiratory pattern.     

Locomotory behavior of fibroblasts in “sail-sheets”

Summary The locomotory behavior of fibroblasts in two-dimensional cultures (e.g., on culture dishes, cover slips, etc.) was first reported by Abercrombie’s group. This paper describes some of the features of movement of fibroblasts in a unique culture system known as sail-sheet cultures (SSCs). Cells in SSCs grow mostly on one another in a three-dimensional form that resembles, to some extent, the in vivo situation. We grew chicken heart fibroblasts (CHFs) as SSCs and studied their locomotory behavior by time-lapse filming extended for periods ranging from 12 h to several days. It was found that CHFs grown as sail-sheets exhibit many features of movement as observed in conventional two-dimensional cultures (CCs). However, we observed that CHFs in SSCs, like those in vivo, lack leading lamella directing their movement. Furthermore, locomotion is significantly slower in SSCs than in CCs. Based on data on the movement of CHFs within the mesh holes of inert grids, we suggest that the mesenchymal cells in SSCs, in addition to their individual movement, move in sheets and that their movement in sheets results in the closure of the mesh holes, a situation that resembles the phenomenon of healing of wounds. Thus SSCs provide a model system for the study of healing of wounds. The presence of collagenlike extracellular matrix (CLECM) between cellular layers in SSCs suggests that CLECM may be involved in guiding the locomotory behavior of CHFs in SSCs.     

Analysis of cell three-dimensional locomotory vectors

A method is presented for analysing the vectors of cells locomoting within three-dimensional collagen gels. The method detected differences in locomotory vector patterns between cells locomoting in two different gel formats. The potential of this analytical method for determining the role of the extracellular matrix in modulating cell locomotory behaviour is discussed.     

The hydraulic system in a burrowing worm, Priapulus caudatus

The function of the coelom as a locomotory hydraulic system in Priapulus caudatus has been examined and discussed. Pressure changes within the coelom during burrowing have been measured and used to monitor the activities of the muscular system.     

蕨类植物精细胞运动器和细胞骨架的研究

介绍了近年来蕨类植物游动精子运动器和细胞骨架的研究进展。游动精子由配子体精子器中的非运动细胞发育形成,其分化过程包括了运动器官和细胞骨架的合成和组装。精子发生过程中形成的运动器的各部分结构包括鞭毛、基体、多层结构及附属结构;基体是细胞中新形成的结构,在不同类群的蕨类植物中分别由双中心粒、分支生毛体和生毛体产生。鞭毛、基体和多层结构中的微管带形成了游动精子三个独特的微管列阵,由于微管蛋白的后修饰作用这些微管列阵十分稳定;centrin是运动器中的重要成分, 但功能尚不清楚,可能和细胞骨架及运动器的构建有关。     

Cells can use their transferrin receptors for locomotion.

Cells of the B lymphoblastoid cell line JY attach to substrata made of antibodies to the transferrin receptor. Many of these attached cells migrate considerable distances. JY cells also attach to an anti-integrin substratum (anti LFA-1), but on this surface they do not migrate. These results suggest that a circulating receptor--the transferrin receptor--can be used for locomotory purposes, whereas LFA-1, which is not endocytosed in these cells, cannot be used for locomotion. This indicates that the endocytotic cycle can drive cell locomotion.     

ANALYSIS OF MICROTUBULE STRUCTURE IN EUGLENA GRANULATA

The microtubules associated with the pellicle, reservoir and locomotory flagellum of Euglena granulata were subjected to rotational analysis using the technique of Markham et al. The results indicate that the microtubules have 11-14 subunits when viewed in transverse section. Rotational analysis of the entire locomotory flagellum was also conducted to better demonstrate the nature of the arms extending from the peripheral microtubules.     

A comparison of the distribution of muscle type in the tadpole tails of Xenopus laevis and Rana temporaria: an histological and ultrastructural study

The types and the distribution of muscle fibres were analysed and compared in the tails of Xenopus laevis and Rana temporaria tadpoles. The filter feeding tadpoles of X. laevis were found to have both white muscle fibres adjacent to the notochord used for normal locomotory swimming and a superficial layer of small red fibres. The red fibres are probably used for the continuous flickering movement of the tail associated with the maintenance of the mid-water filter feeding position. R. temporaria, a grazing detritus feeding tadpole, was found to have only white muscle fibres used for normal locomotory swimming. Smaller superficial fibres were not red fibres but were thought to be immature white fibres.