Time-dependent Losses Research Articles

Degradation of rat hepatic cytochrome P-450 heme by 3,5-dicarbethoxy-2, 6-dimethyl-4-ethyl-1,4-dihydropyridine to irreversibly bound protein adducts

Administration of 3,5-dicarbethoxy-2,6-dimethyl-4-ethyl-1,4-dihydropyridine (DDEP) (a structural analog of the dihydropyridine Ca 2+ antagonists) to untreated, phenobarbital-, or dexamethasone-pretreated rats results in time-dependent losses of hepatic cytochrome P-450 content. Functional markers for various cytochrome P-450 isozymes have permitted the identification of P-450 h, P-450 PB-1/k, and P-450 p as the isozymes inactivated preferentially by the drug. DDEP-mediated cytochrome P-450 destruction may be reproduced in vitro, is most prominent after pretreatment of rats with dexamethasone, pregnenolone 16α-carbonitrile or phenobarbital, and is blocked by triacetyloleandomycin. These findings together with the observation that DDEP markedly inactivates hepatic 2β- and 6β-testosterone hydroxylase and erythromycin N-demethylase tend to indict the steroid-inducible P-450 p isozyme as a key protagonist in this event. The precise mechanism of such DDEP-mediated P-450 p heme destruction is unclear, but involves prosthetic heme alkylation of the apocytochrome at its active site in what appears to be a novel mechanism-based “suicide” inactivation. Such inactivation appears to involve fragmentation of the heme to reactive metabolites that irreversibly bind to the protein, but the chemical structure of the heme-protein adducts is yet to be established. Intriguingly, such DDEP-mediated P-450 p destruction in vivo also results in accelerated loss of immunochemically detectable apocytochrome P-450 p. It remains to be determined whether or not this loss is due to enhanced proteolysis triggered by the structural modification of the apocytochrome.

Studies on aromatase inhibition with 4-androstene-3,6,17-trione: Its 3β-reduction and time-dependent irreversible binding to aromatase with human placental microsomes

The metabolism of 4-androstene-3,6,17-trione (AT), previously described as a suicide substrate for aromatase, and its irreversible binding to aromatase were studied by using human placental microsomes. AT was rapidly converted into 3β-reduced metabolite (3-OHAT) with an enzyme other than aromatase in the microsomes in the presence of NADPH under either aerobic or anaerobic conditions. The conversion was efficiently prevented by a steroid 5α-reductase inhibitor. 3-OHAT was characterized as a competitive ( K i = 6.5 μM) and irreversible inhibitor of aromatase. Both 14C-labeled AT and 3-OHAT were demonstrated to be irreversibly bound to aromatase probably through a sulfur atom of the enzyme in time-dependent manners in the presence of NADPH, being accompanied with time-dependent losses of the enzyme activity. It was shown that the process of an apparent time-dependent loss of aromatase activity caused by AT even under conditions allowing its 3β-reduction should principally depend on the action of the parent inhibitor AT itself and not on that of the metabolite 3-OHAT.

Determining Time Dependent Losses Of Prestress with Due Consideration of Aging Coefficient and Percentage of Steel

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Time-dependent changes in H1 content, H1 turnover, DNA elongation, and the survival of cells blocked in early S phase by hydroxyurea, aphidicolin, or 5-fluorodeoxyuridine.

Cells were synchronized in G1 by isoleucine deprivation and then released into medium containing 1 mM hydroxyurea (HU), 5 micrograms mL-1 aphidicolin (APC), or 1 microgram mL-1 5-fluorodeoxyuridine (fl5dU). Coulter volume, content of histone H1 per unit DNA, turnover of histone H1, the extent of DNA elongation, and the survival of cells were measured as functions of time after release into the presence of the drugs. At the concentrations used in the experiments, the drug differ in their toxicity (fl5dU greater than HU greater than APC), induction of unbalanced cell growth, and the distribution of new DNA fragment sizes allowed during block, but they all (1) allow cells to enter S phase, (2) cause similar time-dependent losses of histone H1 per unit DNA, which begin as synchronized G1 cells begin to enter S phase, (3) retard DNA elongation beyond replicon size, and (4) retard the turnover of histone H1. The results indicate that loss of histone H1, inhibition of histone turnover, the retarded ligation of newly replicated DNA into bulk chromatin, and chromatin structural changes may be part of the cell's general response to inhibition of DNA replication. Since transient S phase block increases the frequencies of gene amplification [Mariani, B. D., & Schimke, R. T. (1984) J. Biol. Chem. 259, 1901-1910] and sister chromatid exchanges (SCE) [Rainaldi, G., Sessa, M. R., & Mariani, T. (1984) Chromosoma 90, 46-49], the observed changes in H1 content and chromatin organization may also be essential features of gene amplification and SCE.

A comparative study of intensity pulsations in a double heterostructure AlxGa1?xAs laser with and without an external resonator present

This paper describes the pulsing behaviour of AlxGa1−x As double heterostructure (DH) lasers with and without an external dispersive resonator present. We have investigated only those lasers which show self-sustained oscillations under dc excitation. For the solitary diode we have found pulses possessing a width at half-power points, Δτp, where Δτp=0.9 ns and a repetition frequency,fos, ranging from 176 to 775 MHz depending on injection currentJ. Introducing a correlation time,tc, as a measure of the stability of the pulse height we have obtained 5 <tc < 40 ns. The maximum value oftc occurs in the vicinity of the crossing point offos2 andτc−2, whereτc is the effective carrier lifetime. Output pulsations are accompanied by spectral shifts of the longitudinal diode cavity modes. The frequency halfwidth, ΔνHw, ranging from 30 to 90 GHz is closely related to the maximum pulse power,Pmax. Coupling the laser diode to an external resonator, pulse widths, oscillation frequencies and correlation times have been determined as functions ofJ/Jth (Jth is the threshold injection current), provided the photon roundtrip time is nearly equal toτc and the equivalent reflection factorR* of the external resonator isR* ∼ 0.87. By evaluating the experimental findings pulsing under dc excitation may be described, without a detailed knowledge of the reasons leading thereto, by using the concepts of a nonstationary quality factorQns and time-dependent resonator losses characterized by ΔνHw/δτp. Oscillations become self-sustained forQns=const., i.e. the minimum photon density with which the pulses start to grow has to be time and excitation independent, in this casefos2 is proportional toJ/Jth as well as Δτp to (J/Jth−1)−1/2.

Stabilization of glucocorticoid-receptor complexes in rat thymus cytosol by a factor from WEHI-7 cells

Using a variety of physico-chemical techniques we have recently characterized three distinct forms of glucocorticoid-receptor complexes present in the cytosol from rat thymus cells incubated with glucocorticoid; the relative proportions of these complexes are dependent on the conditions to which the cells or cytosols are exposed. Two of these complexes correspond to the well established nonactivated and activated receptor forms, while the third has properties consistent with mero-receptor. Based on their differential affinities for DNA- and DEAE-cellulose we have developed a rapid mini-column chromatographie procedure for separating these three forms and have used it to examine the stability of complexes in cytosol preparations. We have found that activated glucocorticoid-receptor complexes from rat thymus cells are relatively unstable under cell-free conditions in that they undergo time-dependent losses in DNA binding and are converted to mero-receptor. In contrast, cytosolic glucocorticoid-receptor complexes prepared from WEHI-7 mouse thymoma cells are remarkably stable under similar conditions. Mixing experiments with equal portions of rat thymus and WEHI-7 cytosol revealed that the difference between the two tissues cannot be accounted for merely by differences in amounts of proteolytic enzymes, since addition of rat thymus cytosol to WEHI-7 cytosol containing activated glucocorticoid-receptor complexes does not result in their conversion to mero-receptor. However, the WEHI-7 cytosol affords considerable protection to activated glucocorticoid-receptor complexes in thymus cytosol. The stabilizing factor from WEHI-7 cytosol is heat stable (survives 100°C for 30 min), insensitive to pH over a wide range (4.0–10.0), and appears to be macromolecular. It does not inhibit activation, and thus appears distinct from the previously described endogenous glucocorticoid receptor stabilizing factor responsible for stabilization of thymocyte receptor binding capacity (Leach et al., J. Biol. Chem. 257: 381–388, 1982). We propose that the factor is an endogenous inhibitor of the protease(s) responsible for mero-receptor formation.

Dual picosecond dye lasers synchronously pumped by a mode locked cw yag laser

The performance of a novel dual dye laser system synchronously pumped by the frequency doubled output of a mode-locked CW-YAG laser is evaluated in relation to pulsewidth, pulse substructure, pulse spectral width and timing jitter. The behavior of the system is adequately described by a theoretical model which includes the time dependent gain and losses due to frequency bandwidth, cavity length mismatch and output coupler. The jitter is significantly reduced from that obtained with CW gas laser pumping as a result of the shorter pump pulse (50 ns instead of ≈100 ps). A routine operating condition uses 2-plate birefringen filters, 0.8 W pump power at 532 nm, to yield two 2.0 ps pulses having a cross correlation width of 3.8 ps, and 30 mW average power from each laser.

Dual activity at an enzyme active site: 3 beta,20 alpha-hydroxysteroid oxidoreductase from fetal blood.

An enzyme exhibiting both 3 beta and 20 alpha steroid reductase activities from calf fetal red blood cells was purified to homogeneity as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. 3 beta,20 alpha-Hydroxysteroid oxidoreductase (3 beta,20 alpha-HSD) was found to be a single-stranded polypeptide with a molecular weight of 55 000 +/- 1 000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Sephadex G-100 chromatography. The amino acid composition of 3 beta,20 alpha-HSD was obtained. 17 beta-Hydroxy-5 alpha-androstan-3-one and progesterone were substrates for the enzyme's 3 beta and 20 alpha reductase activities, respectively, which required NADPH for both 3 beta [Km = 9.4 microM; Vmax = 2.4 nmol min-1 (nmol of enzyme)-1] and 20 alpha [Km = 2.5 microM; Vmax = 2.4 nmol min-1 (nmol of enzyme)-1] reductase activities. 17 beta-Hydroxy-5 alpha-androstan-3-one competitively inhibited (Ki = 35 microM) 20 alpha reduction of progesterone. Incubating 3 beta,20 alpha-HSD with 19-nortestosterone 17-bromoacetate at pH 7.0 and 25 degrees C caused simultaneous, time-dependent, and irreversible losses of 3 beta and 20 alpha activities by a first-order kinetic process. Similar incubations with either of the 3 beta or 20 alpha substrates present at concentrations equal to their respective Km values practically doubled the time required for loss of 3 beta and 20 alpha enzyme activities. These data lead us to conclude that the active site of 3 beta,20 alpha-HSD contains 3 beta and 20 alpha dual activity.

A critical evaluation of the effect of electric fields on the residual structure of vapor-deposited metal films

Thin films of Al, Pb, Bi, and Sn representing a range of bulk crystal structures and low-melting-point metals were vacuum-vapor deposited onto air-cleaved (001) NaCl substrates in the presence of dc electric fields applied in the plane of the deposit (using detached electrodes) ranging between 102 and 103 V/cm even considering displacement current effects during evaporation. Such experiments, conducted at room temperature (25 °C) within short vaporization periods in order to avoid thermal-induced enhancement of bulk and surface conductivities in the NaCl substrate, and time-dependent gradient losses, showed no significant or detectable effects on residual microstructures or crystal structure upon examination and comparison of representative film samples that have an approximate thickness of 1200 A by transmission electron microscopy.

The formation of plasma equilibrium in a Q-machine with radial potential discontinuities

A potassium-ion plasma forms in a Q-machine with mirror magnetic fields at the hotplates. Discontinuity in the thermoelectric potential at the hotplate edge produces a strong localized radial electric field that both electrostatically confines the ions and creates a high-velocity rotating annulus. Neglecting time-dependent losses, the radial equilibrium in the collisional regime with the column radius comparable to the ion gyro radius is adequately described by an isothermal single-fluid model, including both ion viscosity and rotational inertia. The radial density profiles are consistent with classical diffusion at the column edge.

The effect of thermal blooming in a liquid filter on the emission of a Nd: Glass laser

The spiking behaviour of a Nd: glass laser operating reproducibly in TEM 00-mode operation was investigated with a liquid filter inside the cavity. It was found that thermal blooming in the liquid causes time-dependent losses of the order of 1%, and therefore a substantial decrease of spike intensity.

Pulse Delays in TEA CO2 Lasers

A novel method is reported for studying the time-dependent gain and cavity losses in a TEA CO2 laser. The technique involves an investigation of the time delay of the onset of lasing with respect to the current pulse in such a laser. Detailed measurements of this time delay as both the laser gain and the cavity loss are varied, in conjunction with a simple theoretical analysis, permit the evaluation of parameters describing the time-dependent gain and the cavity loss. In particular, good values for the peak gain, the rise and fall times of the gain, and the average cavity loss per pass can be obtained. The parameters measured in this way are in agreement with other reported data, where available. The method of investigation reported avoids many of the experimental problems which are involved in the direct measurement of the time-dependent gain using cw CO2 laser techniques. Furthermore, the measurement method described also provides an in situ method of measuring the laser cavity loss.