Bioelectrochemistry and bioenergetics (Lausanne, Switzerland)最新文献

Differential pulse polarography (DPP) and electron spin resonance (ESR) were used to study the influence of substituents and of the pH of the medium on DPP peak potentials (electrochemical reduction) resp. k_reduction (chemical reduction) of nitroxyl free radicals. The DPP peak potentials can be used to select the appropriate nitroxide spin label for relevant biochemical and biophysical applications.

In Hevea, rubber synthesis is confined to the cytosol of the highly differentiated laticifer cells. Agronomic and biochemical studies showed that this process uses high amounts of sugars that are efficiently imported into the laticifer. A H⁺–sugar symport system located in the plasma membrane is involved in sugar uptake into laticifers. Laticifer protoplasts were prepared and used in electrophysiological and labeling experiments to test the capacity of this system to transport a variety of sugars such as oligosaccharides from the raffinose family, trace compounds in rubber. Translocation of sugars known to be transported with different efficiency across the plasma membrane of plant cells was also tested. A 1 mM sucrose affinity was found for the symport. All the sugars tested, except palatinose induce membrane depolarization indicating that they were actively absorbed by the laticifer network. This reveals the wide capacity of this peculiar sink for the uptake of sugars.

The voltammetric behavior of human mammalian cells was studied by choosing human leukemia cells (HL60) and human erythroleukemia cells (HEL). The voltammetric response of the cells was found having relation with cell metabolic viability in culture course. For example, the fluctuations of peak currents of HL60 were parallel with the nutrients replenished or not, which can reflect cell health state; the voltammetric response of HL60 regulated by the anti-metabolic drug 5F-Uriacil (5F-U) in culture course behaved in a much decreased manner, by which a voltammetric method for evaluating cytotoxicity is proposed. In this paper, the relation between HEL cell metabolism and the activation of receptor Mpl by its ligand TPO was also studied. Moreover, the mechanism of cell voltammetric behavior was discussed.

A model eukaryotic cell system was used to explore the effect of a weak pulsed magnetic field (PMF) on time-varying physiological parameters. Dictyostelium discoideum cells (V12 strain) were exposed to a pulsed magnetic field (PMF) of flux density 0.4 mT, generated via air-cored coils in trains of 2 ms pulses gated at 20 ms. This signal is similar to those used to treat non-uniting fractures. Samples were taken over periods of 20 min from harvested suspensions of amoebae during early aggregation phase, extracted and derivatised for HPLC fluorescent assay of adenine nucleotides. Analysis of variance showed a significant athermal damping effect (P<0.002, n=22) of the PMF on natural adenine nucleotide oscillations and some consistent changes in phase relationships. The technique of nonlinear dielectric spectroscopy (NLDS) revealed a distinctive effect of PMF, caffeine and EGTA in modulating the cellular harmonic response to an applied weak signal. Light scattering studies also showed altered frequency response of cells to PMF, EGTA and caffeine. PMF caused a significant reduction of caffeine induced cell contraction (P<0.0006, n=19 by paired t-test) as shown by Malvern particle size analyser, suggesting that intracellular calcium may be involved in mediating the effect of the PMF.

In order to detect and model the effect of functional chain heterogeneity on Nernst plots for heme proteins, we examined the redox properties of various myoglobins (Mbs) and their mixtures using an improved spectroelectrochemical method. Specific redox responses and formal half potentials (E_1/2) were obtained for Aplysia, horse, and sperm whale Mbs, as well as 1:1 mixtures of Mbs consisting of Aplysia/sperm whale, sperm whale/horse, and horse/Aplysia. Linear Nernst plots with slopes near unity were observed for horse, sperm whale, and Aplysia Mbs, with E_1/2 values of 14, 19, and 96 mV (vs. NHE) respectively, consistent with previous reports using indirect methods. The Nernst plot responses for mixtures of some of these Mbs allowed us to evaluate and model the non-Nernstian behavior that results from intrinsically different values of E_1/2 and from incomplete spectral overlap. The data demonstrate that increasing the E_1/2 differences between the components of a Mb mixture increases the changes in shape of the resulting Nernst plots, the dominant effect being a decrease in the observed Nernst coefficient (n_Nernst). Comparison of Nernst plots for redox data with Hill plots for O₂ binding data shows that the redox process is more affected by the structural differences in the distal heme pockets of the Mbs studied than is O₂ binding. Similar effects of chain heterogeneity may give rise to disproportionate reductions in the slopes of Nernst and Hill plots for hemoglobins (Hbs). This possibility is discussed in relation to Hbs investigated for redox and O₂ binding activity in our laboratories where we find n_Nernst to be commonly less than n_Hill over a range of experimental conditions.

Homocysteine differs from cysteine in electrochemical behaviour due to slight differences in hydrophobicity and in structure of their complexes with metals. This shows in adsorptivity, in anodic reactions with mercury, in catalytic reductions of oxygen mediated by mercurous thiolates and of thiol-cobalt(II) complexes, but most markedly in catalytic evolution of hydrogen from solutions containing cobalt ions.

Electrochemotherapy is a method for cancer treatment consisting in combining intratumor injection of cytotoxic agent with the application of intensive electrical stimuli. Thus transient cell membranes permeabilization is created, allowing the agent to better exercise its destroying effect. Positive results have been published in the treatment of cutaneous malignant formations and other types of cancer are under consideration. The electrode configurations presently used are based mainly on empirical treatment results. In vivo imaging of stimulation currents was attempted in animal models. A preliminary study revealed that having in view the relatively high voltages and currents, there was a virtually resistive load to the electrical source. Assuming a homogeneous medium, potential and current distributions were modeled and studied. The results could help in selection of specific electrode designs, depending on tumor size and location. An optimization of the voltages and/or currents by different electrode arrays can lead to obtaining desired field distribution.

The effect of the initial biochemical or metabolic state of a cell membrane target pathway on its sensitivity to exogenous electromagnetic (EMF) fields is considered. It is shown that the resting or initial transmembrane voltage can affect the frequency response of the membrane pathway and substantially alter the signal to thermal noise threshold (SNR) of the target. EMF sensitivity is examined using a model which describes the response to applied fields of both single cells and cells in gap junction contact via a distributed parameter electrical circuit analog, wherein a voltage-dependent membrane impedance, relating to the initial biochemical state of the target cell(s), is considered. Application of the Hodgkin–Huxley K⁺-conduction pathway membrane to this model results, at a given transmembrane voltage, in a preferential array response to applied field frequencies in the 1–100 Hz range, centered at approximately 16 Hz for 1–10 mm array lengths. Extension of the model to consider the voltage dependence of the Hodgkin–Huxley K⁺ pathway results in a significant modulation of array frequency response with changing membrane resting potential. The result is EMF sensitivity (SNR) depends upon the initial state of the target tissue, providing a possible explanation of why, e.g., repairing, rather than resting, bone exhibits a physiologically relevant response to certain weak EMF signals.

The photoinduced electrical events at energy-conserving chloroplast membranes can be studied in whole plastids using suction electrodes. In chloroplasts of Peperomia metallica the kinetic profile of photocurrent contains a minor outward component that occurs prior to and differs in polarity from the main component. The origin of this outward current was analyzed using single-turnover flashes in combination with prolonged light exposures and differential physicochemical treatments of tip-located (internal) and the exposed parts of a chloroplast. The outward current signal was higher after 10- to 20-s preillumination and gradually reduced in darkness. The relative amplitude of the outward peak current was enhanced when photosystem II (PS II) was excited by flashes given in the presence of far-red background light (λ=712 nm). The outward current was small or absent under conditions promoting activity of photosystem I (cyclic electron transport supported by artificial redox mediators in the presence of diuron) and was particularly high in the presence of PS II electron acceptors (e.g., p-phenylenediamine). This indicates the predominant association of the outward current with activity of PS II. The external application of diuron strongly inhibited the inward current, giving rise to a temporal increase in the outward current. On the contrary, when diuron was added into the suction pipette, the outward current was inhibited soon after sealing. The data suggest that the outward current originated in the tip-located portions of the thylakoid membrane that have orientation opposite to the exposed part of `whole thylakoid'. These tip-located membrane portions are least accessible for inhibitors added into the outer medium and are highly sensitive to inhibitors (diuron), ionophores (gramicidin D), and detergents (Triton X-100) added into the pipette. Differential involvement of two photosystems in generation of the outward current may be caused by uneven structural distribution of photosystems I and II between appressed (granal) and nonappressed (stromal) thylakoids and by different recording configurations for these thylakoids.

The kinetics of an electromagnetic field (EMF) target pathway are used to estimate frequency windows for EMF bioeffects. Ion/ligand binding is characterized via first order kinetics from which a specific electrical impedance can be derived. The resistance/capacitance properties of the binding pathway impedance, determined by the kinetics of the rate-determining step, define the frequency range over which the target pathway is most sensitive to external EMF. Applied signals may thus be configured such that their spectral content closely matches that of the target, using evaluation of the signal to thermal noise ratio to optimize waveform parameters. Using the approach proposed in this study, a pulsed radio frequency (PRF) waveform, currently employed clinically for soft tissue repair, was retuned by modulation of burst duration, producing significant bioeffects at substantially reduced signal amplitude. Application is made to Ca²⁺/Calmodulin-dependent myosin phosphorylation, for which the binding time constants may be estimated from reported kinetics, neurite outgrowth from embryonic chick dorsal root explants and bone repair in a fracture model. The results showed that the retuned signal produced increased phosphorylation rates, neurite outgrowth and biomechanical strength that were indistinguishable from those produced by the clinical signal, but with a tenfold reduction in peak signal amplitude, ≈800-fold reduction in average amplitude and ≈10⁶-fold reduction in average power.