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Biomedical subjects

D S Dimitrov

Publications and source records attributed to D S Dimitrov.

At least 73 records · Page 4Linked to original sources
PubMed1992

Interactions of CD4+ plasma membrane vesicles with HIV-1 and HIV-1 envelope glycoprotein-expressing cells.

To study interactions between the human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein (gp120-gp41) and the receptor in the target membrane, CD4, a new experimental system utilizing CD4-carrying plasma membrane vesicles (CD4 PMVs) was developed. CD4 PMVs were prepared by hypotonic lysis of HeLa cells expressing CD4 after infection with recombinant vaccinia virus containing the CD4 cDNA. The CD4 PMVs carried up to 680 CD4 molecules per vesicle. Their fusion with cells expressing gp120-gp41 after infection with recombinant vaccinia virus was monitored by fluorescence video microscopy by using lipophilic fluorescent dyes. Fluorescence changes as a result of fusion occurred within 30 min at 37 degrees C, and little fluorescence changes were seen with cells expressing the noncleaved HIV-1 envelope glycoprotein (gp160). The preincubation of CD4 PMVs with HIV-1 reduced its infectivity 10-fold. The CD4 PMVs were more effective in inhibiting syncytia formation than sCD4. These results demonstrate that CD4 PMVs could be used to study the mechanisms of HIV-1 envelope-mediated fusion and have the potential to inactivate HIV-1.

Animals↗
PubMed1991

Initial stages of HIV-1 envelope glycoprotein-mediated cell fusion monitored by a new assay based on redistribution of fluorescent dyes.

Membrane fusion is an essential step in the infection of permissive cells with human immunodeficiency virus (HIV). Infected cells frequently fuse with each other, and then progress to form multinucleated giant cells (syncytia). To gain insight into mechanisms of HIV env-mediated membrane fusion, we developed a new assay for studying the initial events. The assay is based on the redistribution of fluorescent markers between membranes and cytoplasm of adjacent cells examined by means of fluorescence video microscopy. Membrane fusion between HIV-1 envelope glycoprotein (gp120/41) expressing effector cells and CD4+ target cells was observed 90 min after the association of cells, whereas the first syncytia only became apparent after 5 h. Moreover, membrane fusion events were observed under conditions where no syncytia were detected, for example, when the effector:target cell ratio was greater than 100:1, or less than 1:100. A significant number of cells with fused membranes were not involved in the syncytia. In order to determine whether quantitative differences in receptor expression might influence the extent of membrane fusion, we used laboratory-selected variants of CEM cells that differ in their expression of CD4. We found that CD4 is required on the target membrane for HIV env-mediated membrane fusion, but its extent is only partially dependent on CD4 surface concentration. The ability of those CEM variants to take part in HIV env-mediated membrane fusion did not correlate with their capacity to form syncytia. These findings indicate that additional steps are needed to form syncytia after membrane fusion.

CD4 Antigens↗
PubMed1990

A delay in membrane fusion: lag times observed by fluorescence microscopy of individual fusion events induced by an electric field pulse.

Low light level video microscopy of the fusion of DiI- (1,1'-dihexadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate) labeled rabbit erythrocyte ghosts with unlabeled rabbit erythrocyte ghosts, held in stable apposition by dielectrophoresis in sodium phosphate buffers, showed reproducible time intervals (delays) between the application of a single fusogenic electric pulse and the earliest detection of fluorescence in the unlabeled adjacent membranes. The delay increased over the range 0.3-4 s with a decrease in (i) the electric field strength of the fusion-inducing pulse from 1000 to 250 V/mm, (ii) the decay half-time of the fusogenic pulse in the range 1.8-0.073 ms, and (iii) the dielectrophoretic force which brings the membranes into close apposition. A change in the buffer viscosity from 1.8 to 10 mP.s caused the delay to increase from 0.36 to 3.7 s (in glycerol solutions) or to 5.2 s (in sucrose solutions). The delay decreased 2-3 times with an increase in temperature from 21 to 37 degrees C. It did not differ significantly for "white" ghosts [0.013 mM hemoglobin (Hb)] or "red" ghosts (0.15 mM Hb) or buffer strength over the range 5-60 mM (sodium phosphate, pH 8.5). The calculated activation energy, 17 kcal/mol, does not depend on the field strength. The yield of fused cells was high when the delay was short. The delay in electrofusion resembles the delays in pH-dependent fusion of vesicular stomatitis viruses with erythrocyte ghosts [Clague, M. J., Schoch, C., Zech, L., & Blumenthal, R. (1990) Biochemistry 29, 1303-1308] and of fibroblasts expressing influenza hemagglutinin and red blood cells [Morris, S. J., Sarkar, D.P., White, J. M., & Blumenthal, R. (1989) J. Biol. Chem. 264, 3972-3978].(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

Attraction, deformation and contact of membranes induced by low frequency electric fields.

The force of attraction between erythrocyte ghosts induced by low frequency electric fields (60 Hz) was measured as a function of the intermembrane separation. It varied from 10(-14) N for separation of the order of the cell diameter to 10(-12) N for close approach and contact in 20 mM sodium phosphate buffers (conductivity 260 mS/m, pH 8.5). For large separations the interaction force followed a dependence on separation as predicted for dipole-dipole interactions. For small separation an empirical formula was obtained. The membranes deformed at close approach (less than 1 microns) before making contact. The contact area increased with time until reaching the final equilibrium state. The ghosts separated reversibly after switching off the electric field. The membrane tension induced by the ghost interaction at contact was estimated to be of the order of 0.1 mN/m. These first quantitative measurements of the force/separation dependence for intermembrane interactions induced by low frequency electric fields indicate that attractive forces, membrane deformation and contact area of cells depend strongly on intermembrane separation and field strength. The quantitative relationship between them are important for measuring membrane surface and mechanical properties, intermembrane forces and understanding mechanisms of membrane adhesion, instability and fusion in electric fields and in general.

Cell Aggregation↗
PubMed1990

Membrane electroporation--fast molecular exchange by electroosmosis.

Human and rabbit erythrocyte ghosts loaded with FITC-dextran (mol. mass = 10 kDa) and NBD-glucosamine (mol. mass = 342 Da) in buffers of different ionic strength and composition were subjected to electric pulses (intensity 0.7 kV/mm and decay half-time 1 ms) at 7-10 degrees C and 20-24 degrees C. The transfer of the fluorescent dyes from the interior of the ghosts through the electropores was observed by low light level video microscopy. The pulses caused the fluorescence to appear outside the membranes as a transient cylindrical cloud directed toward the negative electrode during the first video frame (17 ms). It was similar in both rabbit and human erythrocyte ghosts and at both temperatures but differs for the two dyes, the fluorescence cylinder is long and tall for the FITC-dextran and relatively short and thick for the NBD-glucosamine. The molecular exchange was 2-3 orders of magnitude faster within the first 17 ms after the pulse than the diffusional exchange. It decreased with increasing ionic strength. Formulae for the transfer of molecules by electroosmotic flow through the pores are in agreement with these observations. They allow estimation of the total area of pores with radii larger than that of the fluorescent dye during the pulse. The major conclusion is that electroosmosis is the dominating mechanism of molecular exchange in electroporation of erythrocyte ghosts.

4-Chloro-7-nitrobenzofurazan↗
PubMed1989

Electrofusion of fragile mutants of Saccharomyces cerevisiae.

This communication presents experimental evidence that intact fragile (osmotic sensitive) yeasts can be electrofused and give viable hybrids. The yield increases with one order of magnitude for electrofusion of intact fragile yeasts with protoplasts of non-fragile ones. The yield of viable hybrids, obtained by electrofusion of protoplasts of fragile and non-fragile yeasts, is one order of magnitude higher than the yield from protoplasts of non-fragile yeasts. The destabilized cell wall and plasma membrane of the mutant yeasts could be a possible explanation for this phenomenon.

Electricity↗
PubMed1987

[Histological characteristics of the intraorbital glands of pheasants with tuberculosis].

For the first time in the country are investigated histologically lacrimal and Harderian glands of pheasants, affected by tuberculosis, raised in pens. By histological analysis were reported the results of the cell reactivity in the affected by the tubercular process tissues and organs. Under the influence of the mycobacteria and their disintegration products the tissues react specifically. In pheasants, affected by tuberculosis are established destructive processes in the intraorbital glands. Mainly in the connective tissue of the glands are observed tubercular knots, which morphologically differ from these in the hen. In the interlobular tissue, in the interacinar and peritubular connective tissue exist strongly expressed infiltration with cells of the lymphoid series, phagocytes and heterophils, as a result of the affected-tissue reactivity, on account of the local and diffusional spreading of the tubercular bacillus in the glands.

Animals↗
PubMed1986

Mechanical aspects of the lungs as cancer cell-killing organs during hematogenous metastasis.

A substantial proportion of many different types of circulating cancer cells appear to be killed during their interactions with the pulmonary microcirculation. Different tensions exist during respiration within alveolar units, and hence the pulmonary capillaries. We have calculated the effects of these tensions on the entry and subsequent fate of circulating cancer cells. Our calculations indicate that during expiration, when tension in the capillary walls is low, cancer cells can enter and travel along the capillaries without damage, because the vessels are deformed by the cells and the hydrodynamic field surrounding them. During normal inspiration when the alveoli are stretched, the increased tension within the capillary walls serves to compress the contained cancer cells. This compression, together with previously calculated blood pressure differentials between the ends of the cells, is thought in some cases, to increase their membrane tensions above the critical level for rupture, resulting in cytolysis, in accord with experimental observations. In deep inspiration, when a very substantial increase in capillary wall tension occurs, cancer cells already within the capillaries, entering them and in transit along them are expected to develop membrane tensions greatly exceeding the critical values for rupture. It is suggested that these respiration-induced effects may act as an important rate-regulating step in the metastatic process, where the development of pulmonary metastases plays a central role. Furthermore, induced deep inspiration may conceivably be utilized in the inhibition of pulmonary metastasis.

Animals↗
PubMed1986

Red blood cell dielectrophoresis in axisymmetric fields.

Dielectrophoretic velocities of human red blood cells in an axisymmetric field were measured as a function of the applied voltage and the distance from the axis of symmetry. The voltage of the alternating electric field (frequency 2 MHz), applied between two concentric cylindrical metal electrodes (outer and inner radii 0.24 and 1 mm, respectively), was varied up to 19 V. Two kinds of mediums were used: (a) 90% of 2.1% glycine solution and 10% of 5.5% glucose solution and (b) 5.4% sorbitol solution. The results have shown that in both mediums the cell velocities are proportional to the square of the applied voltage and inversely proportional to the cube of the distance from the axis of symmetry, as predicted by the theory. The coefficient of proportionality (dielectrophoretic coefficient) is on the order of 10(-25) A2S4kg-1. It depends on the donor of red blood cells and might be used for diagnostic purposes. These results will be used in future investigations of membrane adhesion, stability and fusion.

Cell Movement↗
PubMed1985

Stability of membrane systems modeled as multilayered viscoelastic films.

We have modeled interacting plane-parallel membranes as viscoelastic multilayered films. A linear dynamic analysis was used. It gave the general conditions for stability of membrane systems having an arbitrary number of membranes and films. The particular cases of one, two and three interacting layers were considered in detail. The results suggest a strong dependence of the stability of membrane systems on the interfacial tensions and the thickness of individual membranes and films and rather weak dependence on their viscosities and elasticities. The theoretical predictions are in semi-quantitative agreement with data on electrically induced membrane fusion in dielectrophoresis.

Cell Membrane↗
PubMed1985

The hemodynamic destruction of intravascular cancer cells in relation to myocardial metastasis.

A variety of observations in humans and experimental animals indicate that large numbers of circulating cancer cells are killed in the microvasculature. It is suggested that this occurs when friction or adhesion between individual cancer cells and capillary walls results in an increase of tension in the cancer cell peripheries above a critical level because of (blood) pressure differentials between their free ends. Hemodynamic and anatomic data relating to the myocardial circulation and deformability measurements on four types of rat cancer cells have been reported previously by others. Novel calculations based on these data suggest that the increased tension at the peripheries of cancer cells passing through the myocardial capillaries will exceed the critical levels for rupture. Analysis of autopsy data for solid tumors reveals a low (less than 3%) incidence of myocardial metastases in the absence of lung metastases and a higher (15%) incidence in their presence. One explanation for these observations is that, in the absence of lung metastases, relatively few of the cancer cells enter the coronary arteries from primary tumors with systemic venous drainage because many are retained or destroyed in transit through the pulmonary vasculature, and most of those delivered to the myocardium then suffer hemodynamic destruction. In the presence of pulmonary metastases, large numbers of viable cancer cells are liberated directly into the pulmonary venules and subsequently are delivered to the myocardium without prior exposure to the arterial side of the microcirculation. The combined effects of increased delivery and the protective effects of arrested cells on those preceding them in files along the capillaries account for the higher incidence of myocardial metastases. It is proposed that hemodynamic destruction of circulating cancer cells may be an important underlying cause of metastatic inefficiency, together with other cytocidal mechanisms.

Biophysical Phenomena↗
PubMed1984

Membrane stability.

Rupture and buckling of artificial and biological membranes is an important part of many biological processes. In this review, we present some of the main experimental facts and their analysis. Recent theoretical work, in particular thin film models and nucleation mechanisms of membrane instability, are discussed in detail. Possible applications to membrane adhesion and fusion are pointed out. Attempts are made to explain biological phenomena and experimental results for biological membranes based on a rigorous physicochemical approach developed previously for thin films in colloid systems.

Animals↗
PubMed1984

Electric field-induced breakdown of lipid bilayers and cell membranes: a thin viscoelastic film model.

A simple viscoelastic film model is presented, which predicts a breakdown electric potential having a dependence on the electric pulse length which approximates the available experimental data for the electric breakdown of lipid bilayers and cell membranes (summarized in the reviews of U. Zimmermann and J. Vienken, 1982, J. Membrane Biol. 67:165 and U. Zimmermann, 1982, Biochim. Biophys. Acta 694:227). The basic result is a formula for the time tau of membrane breakdown (up to the formation of pores): tau = alpha (mu/G)/(epsilon 2m epsilon 2oU4/24 sigma Gh3 + T2/sigma Gh-1), where alpha is a proportionality coefficient approximately equal to ln(h/2 zeta o), h being the membrane thickness and zeta o the amplitude of the initial membrane surface shape fluctuation (alpha is usually of the order of unity), mu represents the membrane shear viscosity, G the membranes shear elasticity modules, epsilon m the membrane relative permittivity, epsilon o = 8.85 X 10(-12) F/m, U the electric potential across the membrane, sigma the membrane surface tension and T the membrane tension. This formula predicts a critical potential Uc; Uc = (24 sigma Gh3/epsilon 2m epsilon 2o)1/4 (for tau = infinity and T = 0). It is proposed that the time course of the electric field-induced membrane breakdown can be divided into three stages: (i) growth of the membrane surface fluctuations, (ii) molecular rearrangements leading to membrane discontinuities, and (iii) expansion of the pores, resulting in the mechanical breakdown of the membrane.

Cell Membrane↗
PubMed1984

A fluid mechanical analysis of the velocity, adhesion, and destruction of cancer cells in capillaries during metastasis.

Metastasis, a multistep process by which cancer disseminates through the body, mainly by intravascular routes, constitutes a major problem in cancer. When cancer cells are injected directly into the veins of animals, they are apparently arrested in the vascular bed of the first organ encountered and gradually released over the next 24 h. These interactions with the microvasculature are often associated in some manner with the death of many cancer cells, and are thought to contribute to the inefficiency of the metastatic process. We have made a theoretical analysis of cancer cells deformed into capillaries with respect to their intravascular velocity, adhesion to the vascular endothelium and intravascular destruction, in terms of the dynamics of the thin liquid film separating the surfaces of the blood vessels and cancer cells. Our calculations, which are based on previously reported experimental observations, indicate that the transit of cancer cells through the microvasculature is discontinuous, being interrupted by adhesions between the two. In addition, in some cases cell membrane rupture (and cell death) will occur when the critical membrane tension of the cancer cells is exceeded by the sum of their initial equilibrium membrane tension and the increased tension in the cancer cell membranes caused by friction generated as they move over the intraluminal surfaces of the capillaries. Our calculations on membrane rupture are consistent with previously unexplained observations by Sato and Suzuki relating cancer cell deformability to death on transpulmonary passage, and constitute a novel mechanism for "metastatic inefficiency" in terms of intravascular cancer cell death.

Animals↗
PubMed1981

A hydrodynamic theory of bilayer membrane formation.

A simple formula for the expansion rate U of the bilayer membrane perimeter was derived by the solution of the hydrodynamic and the Laplace equations at small rates and small contact angles: U = theta delta/6 micron ln (h/e 3/2 h0), where theta is the equilibrium contact angle between the bilayer membrane and the Plateau-Gibbs border, delta the excess membrane tension, micron the bulk viscosity, h the thickness of the thick liquid film, and h0 the thickness of the liquid core in the membrane. This formula coincides with the available experimental data, and explains the observed independence of the expansion rate on the membrane radius. The significance of the theory for the full description of the cell adhesion kinetics as well as the possible ways for its generalization are discussed.

Animals↗
PubMedDate not supplied

Surface pressure hysteresis of mixed lipid/protein monolayers: applications to the alveolar dynamics.

Experimental data on surface pressure-surface area hysteresis of mixed serum albumin/dipalmitoyl lecithin/sphingomyelin monolayers in the Langmuir trough are presented. Several possible physicochemical mechanisms of the hysteresis are discussed: Marangoni effect, surface pressure relaxations, bulk-to-surface diffusion interchange, and collapse. Depending on the concrete conditions each of these mechanisms can be important. Possible applications of these results to the alveolar dynamics are presented and discussed on the basis of the balloon model of the alveolus. The main conclusions of biological importance are that 1) the alveolar stability depends on the DPL/SM ratio as well as on the protein content. Under normal breathing conditions the surface pressure hysteresis is small and does not play a decisive role in the alveolar dynamics. 2) At large extent of compression the collapse predominates in determining the hysteretic behavior of the alveolar surface.

Animals↗
PubMedDate not supplied

Kinetics of calcium-induced fusion of cell-size liposomes with monolayers in solutions of different osmolarity.

The effects of osmolarity, calcium concentration and cell-size liposomes in the subphase on the surface tension of phospholipid monolayers were investigated. The monolayers were spread from chloroform solutions of phosphatidic acid at air/water solution interface. The liposomes (of average diameter 3 micron) were formed from phosphatidic acid/egg lecithin (1:2) mixtures in water or 0.1 M water solutions of sucrose. For this system there were critical concentrations of calcium ions to produce a large reduction of the monolayer surface tension. The threshold calcium concentrations depended upon the sucrose concentration in the subphase. Without sucrose the threshold calcium concentration was 8 mM, while for isoosmotic sucrose solutions (0.1/0.1 M in/out of liposome) it was 14 mM. It sharply increased to 28 mM CaCl2 at sucrose concentration difference across the liposome membrane 0.02 M and decreased to 26 mM, 19 mM, and 18 mM with further increase of that difference to 0.04 M, 0.06 M, and 0.08 M, respectively. The rate of monolayer surface tension decrease was measured as a function of time at 30 mM CaCl2 and different sucrose concentrations in the subphase solution. The initial rates at first decreased with increasing the osmotic pressure and after that they increased. The minimum occurred at sucrose concentration gradient across the liposome membrane 0.02 M, i.e., at the point of maximum threshold calcium concentration required for large decrease of the monolayer surface tension. These facts may be explained by recent theories of dynamics of adhesion, instability and fusion of membranes modeled as thin films.

Calcium↗