scholarly journals Surface Responses in Cultured Fibroblasts Elicited by Ethylenediaminetetraacetic Acid

1958 ◽  
Vol 4 (3) ◽  
pp. 243-250 ◽  
Author(s):  
Ernst J. Dornfeld ◽  
Alfred Owczarzak
Keyword(s):  
Ethylenediaminetetraacetic Acid ◽  
Divalent Cations ◽  
Chelating Agent ◽  
Time Lapse ◽  
Cellular Responses ◽  
Cultured Fibroblasts ◽  
Surface Activities ◽  
Dividing Cells ◽  
Chick Heart ◽  
And Migration

Cultures of chick heart fibroblasts were perfused with the chelating agent ethylenediaminetetraacetic acid (EDTA). Cellular responses were observed under phase optics and recorded by time-lapse cinemicrography. In interphasic fibroblasts, EDTA induces cellular contraction followed by continuous protrusion and retraction of ectoplasmic blebs ("surface bubbling"), formation of motile vermiform processes, and production of rotatory ectoplasmic swellings. The contraction and surface bubbling closely resemble the metaphase contraction and "anaphase bubbling" normally displayed by cultured fibroblasts. In dividing cells, EDTA does not affect metaphases, but anaphase bubbling appears and persists; telophasic expansion and migration of daughter cells are prevented. Initiation of new mitoses occurs during and after exposure to EDTA. No cellular responses are induced by calcium, magnesium, or ferrous chelates of EDTA. The EDTA elects are completely reversible on removal of the chelating agent, resulting in the restoration of the normal interphasic cell form and the normal expansion and migration of mitotic products. The EDTA effects are interpreted to result from the chelation and removal of divalent cations from the cell surface. Possible relations to surface activities observed in normal mitosis are considered, and an hypothesis is presented regarding the role of the developing spindle in cation transfer.

The release of soluble H-2 alloantigens during disaggregation of mouse embryo tissue by a chelating agent

Development ◽  
1966 ◽  
Vol 16 (3) ◽  
pp. 519-530
Author(s):  
Michael Edidin
Keyword(s):  
Molecular Weight ◽  
Ethylenediaminetetraacetic Acid ◽  
Divalent Cations ◽  
Weight Bearing ◽  
Chelating Agent ◽  
Cell Aggregation ◽  
Sponge Cell ◽  
Antigenic Sites ◽  
Dissociated Cells ◽  
Embryo Tissue

Treatment with chelating agents binding divalent cations has been found to effect the dissociation of a variety of tissues of both embryo and adult animals (reviewed in Steinberg, 1958). In the course of dissociation it appears that materials are released from cell surfaces which play a part in their specific adhesion, and which may be shown experimentally to promote selectively the re-aggregation of dissociated cells (Humphreys, 1963; Moscona, 1963). The extracted materials appear to be glycoprotein complexes (Humphreys, 1965), made up of fairly small subunits, estimated to be of 13000–20000 molecular weight (Margoliash et al. 1965). Units of about the same size appear to be the antigenic sites involved in the blocking of sponge cell aggregation by rabbit anti-sponge serum, specific for a given sponge species (MacLennan, 1963). I shall here present evidence that materials of similar molecular weight bearing immunological specificities of the H-2 alloantigen system are released from the tissues of certain mouse embryos during the course of their dissociation by the chelating agent Versene (ethylenediaminetetraacetic acid).

scholarly journals Cell-Projection Pumping of Fibroblast Contents into Osteosarcoma SAOS-2 Cells Correlates with Increased SAOS-2 Proliferation and Migration, as well as Altered Morphology

Biomolecules ◽  
2021 ◽  
Vol 11 (12) ◽  
pp. 1875
Author(s):  
Swarna Mahadevan ◽  
James A Cornwell ◽  
Belal Chami ◽  
Elizabeth Kelly ◽  
Hans Zoellner
Keyword(s):  
Cancer Cell ◽  
Cell Tracking ◽  
Time Lapse ◽  
Cell Phenotype ◽  
Phenotypic Change ◽  
Histopathological Diagnosis ◽  
Daughter Cells ◽  
Dividing Cells ◽  
And Migration ◽  
Cell Profile

We earlier reported that cell-projection pumping transfers fibroblast contents to cancer cells and this alters the cancer cell phenotype. Here, we report on single-cell tracking of time lapse recordings from co-cultured fluorescent fibroblasts and SAOS-2 osteosarcoma cells, tracking 5201 cells across 7 experiments. The fluorescent lipophilic marker DiD was used to label fibroblast organelles and to trace the transfer of fibroblast cytoplasm into SAOS-2 cells. We related SAOS-2 phenotypic change to levels of fluorescence transfer from fibroblasts to SAOS-2 cells, as well as what we term ‘compensated fluorescence’, that numerically projects mother cell fluorescence post-mitosis into daughter cells. The comparison of absolute with compensated fluorescence allowed us to deduct if the phenotypic effects in mother SAOS-2 cells were inherited by their daughters. SAOS-2 receipt of fibroblast fluorescence correlated by Kendall’s tau with cell-profile area and without evidence of persistence in daughter cells (median tau = 0.51, p < 0.016); negatively and weakly with cell circularity and with evidence of persistence (median tau = −0.19, p < 0.05); and very weakly with cell migration velocity and without evidence of persistence (median tau = 0.01, p < 0.016). In addition, mitotic SAOS-2 cells had higher rates of prior fluorescence uptake (median = 64.9 units/day) than non-dividing cells (median = 35.6 units/day, p < 0.016) and there was no evidence of persistence post-mitosis. We conclude that there was an appreciable impact of cell-projection pumping on cancer cell phenotype relevant to cancer histopathological diagnosis, clinical spread and growth, with most effects being ‘reset’ by cancer cell mitosis.

scholarly journals Antagonism between Bacteriostatic and Bactericidal Antibiotics Is Prevalent

2014 ◽  
Vol 58 (8) ◽  
pp. 4573-4582 ◽  
Author(s):  
Paolo S. Ocampo ◽  
Viktória Lázár ◽  
Balázs Papp ◽  
Markus Arnoldini ◽  
Pia Abel zur Wiesch ◽  
...  
Keyword(s):  
Combination Therapy ◽  
Bacterial Infections ◽  
Time Lapse ◽  
Cellular Responses ◽  
Inhibition Of Growth ◽  
Drug Classes ◽  
Dividing Cells ◽  
Evolution Of Resistance ◽  
Time Kill ◽  
Antagonistic Interactions

ABSTRACTCombination therapy is rarely used to counter the evolution of resistance in bacterial infections. Expansion of the use of combination therapy requires knowledge of how drugs interact at inhibitory concentrations. More than 50 years ago, it was noted that, if bactericidal drugs are most potent with actively dividing cells, then the inhibition of growth induced by a bacteriostatic drug should result in an overall reduction of efficacy when the drug is used in combination with a bactericidal drug. Our goal here was to investigate this hypothesis systematically. We first constructed time-kill curves using five different antibiotics at clinically relevant concentrations, and we observed antagonism between bactericidal and bacteriostatic drugs. We extended our investigation by performing a screen of pairwise combinations of 21 different antibiotics at subinhibitory concentrations, and we found that strong antagonistic interactions were enriched significantly among combinations of bacteriostatic and bactericidal drugs. Finally, since our hypothesis relies on phenotypic effects produced by different drug classes, we recreated these experiments in a microfluidic device and performed time-lapse microscopy to directly observe and quantify the growth and division of individual cells with controlled antibiotic concentrations. While our single-cell observations supported the antagonism between bacteriostatic and bactericidal drugs, they revealed an unexpected variety of cellular responses to antagonistic drug combinations, suggesting that multiple mechanisms underlie the interactions.

scholarly journals Cell-Projection Pumping of Fibroblast Contents into Osteosarcoma SAOS-2 Cells Correlates with Increased SAOS-2 Proliferation and Migration, and also with Altered Morphology

2021 ◽  
Author(s):  
Swarna Mahadevan ◽  
James A Cornwell ◽  
Belal Chami ◽  
Elizabeth Kelly ◽  
Hans Zoellner
Keyword(s):  
Cancer Cell ◽  
Cell Tracking ◽  
Time Lapse ◽  
Cell Phenotype ◽  
Phenotypic Change ◽  
Histopathological Diagnosis ◽  
Daughter Cells ◽  
Dividing Cells ◽  
And Migration ◽  
Cell Profile

We earlier reported that cell-projection pumping transfers fibroblast contents to cancer cells, and this alters cancer cell phenotype. We now report on single-cell tracking of time lapse recordings from co-cultured fluorescent fibroblasts and SAOS-2 osteosarcoma cells, tracking 5,201 cells across 7 experiments. The fluorescent lipophilic marker DiD was used to label fibroblast organelles, and to trace transfer of fibroblast cytoplasm into SAOS-2. We related SAOS-2 phenotypic change to levels of fluorescence transfer from fibroblasts to SAOS-2, and also to what we term 'compensated fluorescence', that numerically projects mother cell fluorescence post-mitosis, into daughter cells. Comparison of absolute with compensated fluorescence, allowed deduction if phenotypic effects in mother SAOS-2, were inherited by their daughters. SAOS-2 receipt of fibroblast fluorescence correlated by Kendall's tau: with cell-profile area, and without evidence for persistence in daughter cells (median tau = 0.51, p < 0. 016); negatively and weakly with cell circularity, and with evidence for persistence (median tau = -0.19, p < 0.05); and very weakly with cell migration velocity, and without evidence for persistence (median tau = 0.01, p < 0.016). Also, mitotic SAOS-2 had higher rates of prior fluorescence uptake (median = 64.9 units/day), compared with non dividing cells (median = 35.6 units/day, p < 0.016), and there was no evidence for persistence post-mitosis. We conclude there is appreciable impact of cell-projection pumping on cancer cell phenotype, relevant to cancer histopathological diagnosis, clinical spread, and growth, with most effects 'reset' by cancer cell mitosis.

scholarly journals Cytokinesis and Midzone Microtubule Organization inCaenorhabditis elegans Require the Kinesin-like Protein ZEN-4

1998 ◽  
Vol 9 (8) ◽  
pp. 2037-2049 ◽  
Author(s):  
William B. Raich ◽  
Adrienne N. Moran ◽  
Joel H. Rothman ◽  
Jeff Hardin
Keyword(s):  
Cell Division ◽  
Null Allele ◽  
Time Lapse ◽  
Equatorial Region ◽  
Microtubule Organization ◽  
Dividing Cells ◽  
Spindle Midzone ◽  
Cross Links ◽  
Complete Cell

Members of the MKLP1 subfamily of kinesin motor proteins localize to the equatorial region of the spindle midzone and are capable of bundling antiparallel microtubules in vitro. Despite these intriguing characteristics, it is unclear what role these kinesins play in dividing cells, particularly within the context of a developing embryo. Here, we report the identification of a null allele ofzen-4, an MKLP1 homologue in the nematodeCaenorhabditis elegans, and demonstrate that ZEN-4 is essential for cytokinesis. Embryos deprived of ZEN-4 form multinucleate single-celled embryos as they continue to cycle through mitosis but fail to complete cell division. Initiation of the cytokinetic furrow occurs at the normal time and place, but furrow propagation halts prematurely. Time-lapse recordings and microtubule staining reveal that the cytokinesis defect is preceded by the dissociation of the midzone microtubules. We show that ZEN-4 protein localizes to the spindle midzone during anaphase and persists at the midbody region throughout cytokinesis. We propose that ZEN-4 directly cross-links the midzone microtubules and suggest that these microtubules are required for the completion of cytokinesis.

scholarly journals THE CHEMICAL NATURE OF MACROPHAGE RNA-ANTIGEN COMPLEXES AND THEIR RELEVANCE TO IMMUNE INDUCTION

1969 ◽  
Vol 130 (3) ◽  
pp. 557-574 ◽  
Author(s):  
Georges E. Roelants ◽  
Joel W. Goodman
Keyword(s):  
Amino Acids ◽  
Complex Formation ◽  
Chemical Nature ◽  
Divalent Cations ◽  
Chelating Agent ◽  
Total Absence ◽  
Synthetic Polypeptides ◽  
Antigen Complex ◽  
Anionic Groups ◽  
Do So

10 different compounds, including natural and synthetic polypeptides, proteins, polysaccharides, amino acids, and steroid hormones, were assayed for their capacity to form complexes with peritoneal exudate cell RNA. Only molecules carrying negatively charged groups were able to do so. The formation of RNA-antigen complexes was unrelated to the immuno-potency of the "antigen," was not an enzyme-dependent reaction, did not require the synthesis of RNA following introduction of the antigen, did not seem to involve antigen-specific RNAs, was not specific for macrophages, since HeLa cells could be used as effectively, and occurred when purified RNA was mixed with antigen only in the presence of divalent cations. The complexes were very stable, once formed, but could be dissociated by exhaustive dialysis against buffers containing a chelating agent. The macrophage RNA-antigen complex therefore appears to be a chelate between anionic groups on the two components. Based on the total absence of a relationship between immunogenicity and the capacity to form such complexes, as well as the nonspecific nature of complex formation at every level examined, it appears unlikely that RNA-antigen complexes play a physiologically significant role in immune induction.

scholarly journals Removal of tetracyclines in wastewater : accumulation and distribution of chlortetracycline in bulk water and biomass compartments in activated sludge

2021 ◽  
Author(s):  
Ruba F. Farkh
Keyword(s):  
Microbial Community ◽  
Activated Sludge ◽  
Ethylenediaminetetraacetic Acid ◽  
Divalent Cations ◽  
Bulk Water ◽  
Microbial Cells ◽  
Initial Concentration ◽  
Anoxic Conditions ◽  
Polymeric Substance ◽  
The Impact

A study was conducted to examine the removal of chlortetracycline and its distribution and accumulation in three compartments; bulk water, extracellular polymeric substance (EPS) and the microbial cells in activated sludge. Also the effect of different environmental conditions on the distribution and accumulation in the three compartments was investigated. Effluent samples collected from a municipal activated sludge treatment system were set up in bath experiments to test the distribution and accumulation of chlortetracycline under aerobic and anoxic conditions for 14 days. In addition, the impact of the activity of the microbial community on the amassing of the antibiotic in the biomass was examined. The effect of divalent cations on import and accumulation of chlortetracycline was tested. Sorption in believed to be the main removal pathway in wastewater treatment systems for tetracyclines in general and chlortetracycline in particular. In this study that notion was confirmed, and it was found that the removal via sorption under anoxic condition (43.2%) is almost double of that under aerobic conditions (27.0%). The amount of what accumulated in the cells compared to that sorbed in the EPS is twice as much in the former and triple as much in the latter. These findings suggest that changes in the structure and charge of the EPS could be the reason of higher accumulation in the polymeric substance. The impact of microbial activity on the sorption and distribution of the chlortetracycline in the three compartments showed almost a similar behaviour to that under aerobic and anoxic conditions. It was clear that the more viable the microbial community, the less the antibiotic accumulated in the [sic] both biomass compartments; the EPS and microbial cells. Biomass with inhibited respiration accrued 90% of the initial concentration; where as the active microbial community was more resistant and only 24.2% of the initial concentration accumulated within the cells. The findings suggest that the antibiotic makes its way to the cells thus bypassing the EPS, and is trapped in the EPS as it is pumped out of the cells in an energy dependent mechanism. The presence of ethylenediaminetetraacetic acid (EDTA) which is a strong chelator had no import effect. Nevertheless it did indicate that the accumulation in the EPS could be attributed to the presence of cations since there was a high negative correlation (-0.98) between the disappearance of the antibiotic from the EPS compartment and the EDTA concentration used in incubation.

scholarly journals Changes in the Min Oscillation Pattern before and after Cell Birth

2010 ◽  
Vol 192 (16) ◽  
pp. 4134-4142 ◽  
Author(s):  
Jennifer R. Juarez ◽  
William Margolin
Keyword(s):  
Cell Division ◽  
Fluorescent Protein ◽  
Time Lapse ◽  
The Other ◽  
Cell Pole ◽  
Daughter Cells ◽  
Division Site ◽  
Before And After ◽  
Dividing Cells ◽  
Green Fluorescent

ABSTRACT The Min system regulates the positioning of the cell division site in many bacteria. In Escherichia coli, MinD migrates rapidly from one cell pole to the other. In conjunction with MinC, MinD helps to prevent unwanted FtsZ rings from assembling at the poles and to stabilize their positioning at midcell. Using time-lapse microscopy of growing and dividing cells expressing a gfp-minD fusion, we show that green fluorescent protein (GFP)-MinD often paused at midcell in addition to at the poles, and the frequency of midcell pausing increased as cells grew longer and cell division approached. At later stages of septum formation, GFP-MinD often paused specifically on only one side of the septum, followed by migration to the other side of the septum or to a cell pole. About the time of septum closure, this irregular pattern often switched to a transient double pole-to-pole oscillation in the daughter cells, which ultimately became a stable double oscillation. The splitting of a single MinD zone into two depends on the developing septum and is a potential mechanism to explain how MinD is distributed equitably to both daughter cells. Septal pausing of GFP-MinD did not require MinC, suggesting that MinC-FtsZ interactions do not drive MinD-septal interactions, and instead MinD recognizes a specific geometric, lipid, and/or protein target at the developing septum. Finally, we observed regular end-to-end oscillation over very short distances along the long axes of minicells, supporting the importance of geometry in MinD localization.

Regulation of Mitosis

1969 ◽  
Vol 5 (3) ◽  
pp. 745-755
Author(s):  
W. T. JACKSON
Keyword(s):  
Mitotic Spindle ◽  
Time Lapse ◽  
Polarization Microscopy ◽  
Animal Cells ◽  
Higher Plant ◽  
Competitive Antagonism ◽  
Dividing Cells ◽  
Spindle Microtubules ◽  
Giant Nuclei

Earlier studies on the effects of the herbicide isopropyl N-phenylcarbamate (IPC) on mitosis revealed blocked metaphases, multinucleate cells, giant nuclei and an increase in number of partly contracted chromosomes. It was assumed that IPC, like colchicine, was causing these effects by disruption of the spindle apparatus by destroying the spindle microtubules. The animal hormone melatonin causes an increase in birefringence of the mitotic spindle in animal cells, presumably by increasing the number of microtubules. We have studied the effects of IPC, melatonin, and combinations of the two on mitosis in dividing endosperm cells of the African blood lily (Haemanthus katherinae Baker) in vivo by phase-contrast and polarization microscopy. Both qualitative and quantitative data are presented. Interpretation of these results has been aided materially by a time-lapse cinemicrographic analysis of dividing cells subjected to 1 and 10 p.p.m. IPC (unpublished) and by an accompanying fine-structural analysis of untreated and IPC-treated cells. Mitosis was disrupted by 0.01-10 p.p.m. IPC, the severity of the effect depending on both concentration and stage of mitosis of the cell at the time of treatment. Concentrations of IPC that caused cessation of chromosome movement also caused loss of birefringence of the mitotic spindle. Melatonin increased birefringence of the mitotic spindle in these plant cells and partly nullified the adverse effects of IPC. The results of this study demonstrate that the herbicide IPC, under our conditions, causes disruption of mitosis and loss of birefringence of the spindle. And it has been established that an animal hormone is capable of increasing the birefringence, and presumably the number of microtubules, of the mitotic spindle in dividing endosperm cells of a higher plant. Although melatonin is capable of partly nullifying the effects of IPC, a competitive antagonism is not postulated.

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