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Biomedical subjects

B Ehrenberg

Publications and source records attributed to B Ehrenberg.

At least 37 records · Page 2Linked to original sources
PubMed1990

The bactericidal activity of a deuteroporphyrin-hemin mixture on gram-positive bacteria. A microbiological and spectroscopic study.

The combined antibacterial activity of various porphyrins with hemin on Gram-positive bacteria was studied. Protoporphyrin, hematoporphyrin derivative and deuteroporphyrin show only a marginal inhibitory effect in the dark. However, hemin has a strong cytotoxic effect which is independent of illumination and is equally strong in the dark. The disadvantage of hemin treatment is that it is temporary. In this study, we have demonstrated that a combination of deuteroporphyrin and hemin has a unique cytotoxic activity on Staphylococcus aureus, Streptococcus faecalis and Bacillus cereus. The effect of the combined compound is stronger than that of the separate constituents, and is as strong in the dark as in the light. Only 0.005% of the initial S. aureus population survive after a 2 h treatment. Absorption and fluorescence spectra of hemin-deuteroporphyrin mixtures in water and liposomes suggest the formation of a species with spectroscopic properties which are different from those of the two constituents.

Bacillus cereus↗
PubMed1989

The partition and distribution of porphyrins in liposomal membranes. A spectroscopic study.

Spectroscopic techniques were employed to establish the first comparative evaluation of the partition of the photosensitizers hematoporphyrin derivative (HPD) and photofrin II (PF-II) into phosphatidylcholine (PC) and PC/cholesterol liposomes. The fluorescence of the dyes was monitored while they were titrated with liposomes, yielding the dyes' effective binding constants to the membranes. The binding constants of HPD and PF-II to PC liposomes are 12.2 +/- 0.3 (mg/ml)-1 and 9.2 +/- 0.8 (mg/ml)-1 and to PC/cholesterol (50% w/w) liposomes they are 6.7 +/- 0.9 (mg/ml)-1 and 8.0 +/- 0.8 (mg/ml)-1, respectively. The vertical distribution of the dyes in the bilayer was determined by quenching their fluorescence with spin-labeled stearic acids. PF-II was found to reside deeper in the membrane than HPD. Cholesterol was found to modulate the distribution of the two dyes to a greater extent then DPPC and DMPC. The modulation mechanism is discussed.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1989

Inactivation of erythrocytic, lymphocytic and myelocytic leukemic cells by photoexcitation of endogenous porphyrins.

The photodynamic sensitization of leukemic cells (erythrocytic, myelocytic and lymphocytic) via light activation of endogenous porphyrins is described. Human myelocytic-erythrocytic K562 cells and murine Friend erythroleukemia (FELC) and T-cell lymphoma Eb-Esb cells were stimulated to synthesize and accumulate porphyrins. K562 cells accumulated high amounts of protoporphyrin by stimulation with 5-aminolevulinic acid (ALA) plus sodium butyrate or hemin. For Friend and Eb-Ebs cells ALA was an adequate stimulator. The high-metastatic Esb lymphoma cells accumulated comparatively more porphyrin than the low-metastatic Eb cell line. Maximal porphyrin accumulation produced mortality rates of more than 99% after 10 min of photoactivation of the three leukemic lines. Thymidine incorporation was inhibited by the photodynamic effect depending on porphyrin concentration. These results confirm the photodynamic ability of endogenous porphyrins to inactivate cancer cells of different origins.

Aminolevulinic Acid↗
PubMed1989

Analysis of the effect of medium and membrane conductance on the amplitude and kinetics of membrane potentials induced by externally applied electric fields.

The kinetics and amplitudes of membrane potential induced by externally applied electric field pulses are determined for a spherical lipid bilayer using a voltage-sensitive dye. Several experimental parameters were systematically varied. These included the incorporation of gramicidin into the membrane to alter its conductivity and the variation of the external electrolyte conductivity via changes in salt concentration. The ability of the solution to Laplace's equation for a spherical dielectric shell to quantitatively describe the membrane potential induced on a lipid bilayer could thus be critically evaluated. Both the amplitude and the kinetics of the induced potential were consistent with the predictions of this simple model, even at the extremes of membrane conductance or electrolyte concentration. The success of the experimental approach for this system encourages its application to more complex problems such as electroporation and the influences of external electric fields in growth and development.

Cell Membrane↗
PubMed1989

Evoked potential assessment: utility in prognosis of chronic head injury.

Brainstem auditory evoked potentials (BAEP) and somatosensory evoked potentials (SSEP) were performed on 29 patients an average of 12.4 months after traumatic brain injury (TBI). The study purpose was to predict long-term outcome in chronic TBI patients by using multimodality evoked potentials (MEP), the Rancho Los Amigos Scale (RLAS), and other clinical parameters. Neither the BAEP nor SSEP correlated significantly with the cognitive level on the RLAS at the MEP study approximately one year after TBI (RLAS1). Only 11.7% of RLAS1 could be predicted by the combined study of BAEP and SSEP. BAEP and SSEP obtained about one year after TBI jointly had a 15% predictive power of the long-term follow-up RLAS score obtained 18 months after performance of the MEP (RLAS2). Stepwise regression analysis showed that the best predictive indicator of the status of long-term outcome was RLAS1 which alone can predict 60% of long-term outcome. The predictive value of combinations of RLAS1 and age improved prediction of long-term outcome to 66.8%, and the combination of RLAS1, age, and SSEP further increased the value to 72%. Perhaps the MEPs were relatively insensitive in reflecting the patient's adaptation to fixed neuronal damage since patients can perform higher cognitive function by adaptation through behavioral modification and cognitive retraining despite little structural improvement. This adaptation would result in a discrepancy between MEP and RLAS scores in the late chronic phase.

Adolescent↗
PubMed1988

Membrane potential can be determined in individual cells from the nernstian distribution of cationic dyes.

The distribution of a selection of cationic fluorescent dyes can be used to measure the membrane potential of individual cells with a microfluorometer. The essential attributes of these dyes include membrane permeability, low membrane binding, spectral properties which are insensitive to environment, and, of course, strong fluorescence. A series of dyes were screened on HeLa cells for their ability to meet these criteria and several commercially available dyes were found to be satisfactory. In addition, two new dyes were synthesized for this work by esterification of tetramethyl rhodamine. The analysis of the measured fluorescent intensities requires correction for fluorescence collected from outside the plane of focus of the cell and for nonpotentiometric binding of the dye. The measurements and analysis were performed on three different cell types for which there exists a body of literature on membrane potential; the potentials determined in this work were always within the range of literature values. The rhodamine esters are nontoxic, highly fluorescent dyes which do not form aggregates or display binding-dependent changes in fluorescence efficiency. Thus, their reversible accumulation is quantitatively related to the contrast between intracellular and extracellular fluorescence and allows membrane potentials in individual cells to be continuously monitored.

Animals↗
PubMed1987

Effects of membrane physical parameters on hematoporphyrin-derivative binding to liposomes: a spectroscopic study.

Physical parameters of membrane bilayers were studied for their effect on the binding of hematoporphyrin derivative (Hpd), which is used as a sensitizer in photodynamic therapy of cancerous tissues. The purpose of this study was to clarify which parameters were relevant, under physiological conditions, to the selectivity of Hpd binding to cancer cells. Fluorescence spectroscopy was used to measure the relative partitioning of the dye between the lipid and aqueous media. Increasing the microviscosity of the liposomes' membranes by various bilayer additives results in a strong reduction of Hpd binding, to an extent independent of the specific additive. The effect of temperature near the physiological value as well as the effect of cross membrane potential are small. Surface potential does not affect the binding constant, indicating that the binding species does not carry a net electric charge.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1987

Membrane potential induced by external electric field pulses can be followed with a potentiometric dye.

A potential-sensitive dye was recently used to measure the spatial variation in the membrane potential induced by an externally applied electric field. In this work, we demonstrate that the time course of these induced potentials can also be followed. Two experimental systems were explored. Dye fluorescence from HeLa cells could be modulated by a train of field pulses; the relative fluorescence change measured with a lock-in amplifier was linear with the field and similar to the fluorescence responses obtained in the static measurements. A model membrane system consisting of a hemispherical bilayer allowed convenient measurement of the dye absorbance change as a function of the bathing solution conductivity. The charging time of the membrane was inversely related to the aqueous conductance as predicted by the theoretical solution to Laplace's equation.

Cell Membrane↗
PubMed1985

Protective effect of cholesterol on Friend leukemic cells against photosensitization by hematoporphyrin derivative.

The cytotoxic effects of hematoporphyrin derivative (HPD) on Friend erythroleukemic cells were studied. Upon binding of the porphyrin to the cells, the fluorescence spectra was shifted from 613 to 633 nm regarding the main band and from 676 to 667 nm concerning the secondary band. The kinetics of HPD binding was then determined. Maximum binding already occurred at 60 s after exposure of the cells to HPD. It could be demonstrated that the effect of the photoactivated HPD on cell viability was drug, dose, and light fluorescence dependent. Cellular protein synthesis and Friend virus complex release from the cells were equally inhibited by the photodynamic sensitization of the drug, indicating no specific effect on virus maturation. Since cholesterol affects the fluidity of cell membranes, it was important to study the effect of cholesterol enrichment on the photodynamic sensitization by HPD. It was found that, while a 50% reduction in protein synthesis was monitored following treatment with 20 micrograms of HPD per ml and illumination by a 6-milliwatt white light for 60 s, no inhibition was observed following preenrichment of the cells with 0.5, 1, or 2% of cholesterol hemisuccinate. The same trend of cholesterol protection was demonstrated with longer illumination periods up to 10 min. The protective effect of cholesterol hemisuccinate was also seen using scanning electron microscopy. It is thus concluded that the cholesterol hemisuccinate content of Friend erythroleukemic cell membranes is an important factor in regulating the cytotoxicity of photoactivated HPD.

Animals↗
PubMed1984

Surface potential on purple membranes and its sidedness studied by a resonance Raman dye probe.

A new technique for the measurement of membrane surface potential is proposed and demonstrated. The method is based on the fact that a positively charged styryl dye molecule aggregates when present at high concentration in the Debye layer near a membrane bearing a negative surface potential. The dye in its aggregated form exhibits marked differences in its resonance Raman spectrum relative to the free dye molecules. This method was used to study the potential on the surfaces of the purple membrane that contains the pigment bacteriorhodopsin. A value of -29.5 mV was found for membranes with bacteriorhodopsin in its relaxed, light-adapted state, and the potential decreased to -34.5 mV when most of the bacteriorhodopsin was converted to the M412 intermediate. Because the dye probe does not diffuse through the lipid bilayer, it can be used to probe the potential on the external or internal surface of a vesicle. Thus, we found that the potential on the purple membrane was asymmetric and was localized mainly on the surface that faces the cytoplasm in the cell.

Bacteriorhodopsins↗
PubMed1980

Resonance Raman spectroscopy of chemically modified and isotopically labelled purple membranes. I. A critical examination of the carbon-nitrogen vibrational modes.

Resonance Raman spectra of bacteriorhodopsin are compared to the spectra of this protein modified in the following ways: (1) selective deuteration at the C-15 carbon atom of retinal, (2) full deuteration of the retinal, (3) the addition of a conjugated double bond in the beta-ionone ring (3-dehydroretinal), (4) full deuteration of the protein and lipid components, (5) 15N enrichment of the entire membrane and (6) deuteration of the entire membrane (including the retinal). A detailed comparison of the 15N-enriched membrane and naturally occurring purple membrane from 800 cm-1 to 1700 cm-1 reveals that 15N enrichment affects the frequency of only two vibrational modes. These occur at 1642 cm-1 and 1620 cm-1 in naturally occurring purple membrane and at 1628 cm-1 and 1615 cm-1 in the 15N-enriched samples. Therefore, this pair of bands reflects the states of protonation of the Schiff base. However, our data also indicate that neither of these modes are simple, localized C=N-H or C=N stretching vibrations. In the case of the 1642 cm-1 band motions of the retinal chain beyond C-15 are not significantly involved. On the other hand, in the 1620 cm-1 band atomic motions in the isoprenoid chain beyond C-15 are involved.

Bacteriorhodopsins↗
PubMed1980

Resonance Raman spectroscopy of chemically modified and isotopically labelled purple membranes. II. Kinetic studies.

Kinetic resonance Raman spectra of native and isotopically labelled purple membranes are compared. Using these data and the assignments of the previous paper in this sequence, we have confirmed that the Schiff base is deprotonated at times that are short in comparison to M412 evolution. In addition, by monitoring the kinetic resonance Raman spectra in 2H2O with 488.0 nm excitation we have been able to characterize in more detail the vibrational features associated with this unprotonated intermediate that precedes M412. Furthermore, the kinetic spectra of fully deuterated purple membrane in H2O have allowed us to assign the 1465 cm-1 band in these spectra to the C=C stretching frequency of BR570 and the 1512 cm-1 band to the C=C stretching frequency of M412. These spectra have also provided an indication of a Raman spectral feature associated with O640 and, finally, our kinetic spectra have provided evidence that there is a significant alteration in the rate constants for the evolution of the various intermediates when the non-exchangeable protons on the membrane are replaced by deuterons.

Bacteriorhodopsins↗