Antioxidant Properties of Galanin and Its N-Terminal Fragments in in vitro and in vivo Oxidative Stress Modeling

Biochemistry (Moscow) - Antioxidant properties of rat galanin GWTLNSAGYLLGPHAIDNHRSFSDKHGLT-NH2 (Gal), N-terminal fragment of galanin (2-15 aa) WTLNSAGYLLGPHA (G1), and its modified analogue...     

Atomic resolution crystallography reveals how changes in pH shape the protein microenvironment

Hydrogen atoms are a vital component of enzyme structure and function. In recent years, atomic resolution crystallography (>or=1.2 A) has been successfully used to investigate the role of the hydrogen atom in enzymatic catalysis. Here, atomic resolution crystallography was used to study the effect of pH on cholesterol oxidase from Streptomyces sp., a flavoenzyme oxidoreductase. Crystallographic observations of the anionic oxidized flavin cofactor at basic pH are consistent with the UV-visible absorption profile of the enzyme and readily explain the reversible pH-dependent loss of oxidation activity. Furthermore, a hydrogen atom, positioned at an unusually short distance from the main chain carbonyl oxygen of Met122 at high pH, was observed, suggesting a previously unknown mechanism of cofactor stabilization. This study shows how a redox active site responds to changes in the enzyme's environment and how these changes are able to influence the mechanism of enzymatic catalysis.     

Pharmacokinetic and tissue distribution study of [14C]fluasterone in male Beagle dogs following intravenous,oral and subcutaneous dosing routes

The purpose of this work was to compare the pharmacokinetics (PK) and tissue distribution of [¹⁴C]fluasterone following intravenous (iv), subcutaneous (sc) and oral (po) administration in male Beagle dogs. The main goal of the investigation was to discover if non-oral routes would alter parameters observed in this study following the administration of [¹⁴C]fluasterone. The oral formulation had a lower bioavailability (47%) compared to the sc formulation (84%). Po and sc administration resulted in a similar t_max; however, the observed C_max following sc dosing was less than half of that after oral dosing. The sc route had the greatest overall exposure (AUC_0–∞). Tissue distribution analysis 2 h post-intravenous dosing showed that connective tissue (adipose and bone), liver, and skeletal muscle accumulated relatively high levels of fluasterone. The majority of the dose was retained during the first 24 h. Elimination of [¹⁴C]fluasterone-derived radioactivity following intravenous dosing resulted in urine and feces containing 7.6% and 28%, respectively, of the total dose over the first 24 h. Elimination of [¹⁴C]fluasterone-derived radioactivity following subcutaneous dosing resulted in 4.6% in urine and 7.8% in feces of the total dose over the first 24 h. Following oral dosing, elimination resulted in 3.8% in urine and 36% in feces over the first 24 h. In conclusion, the sc route of administration offers some advantages to po and iv due to the prolonged release and increased retention through 24 h.     

Direct determination of selenium in rat blood plasma by Zeeman atomic absorption spectrometry

The method was developed to be applied for direct determination of selenium in rat plasma by graphite-furnace atomic absorption spectrometry with Zeeman background correction. Blood was obtained from CD rats of both sexes 2h after dosing in weeks 7 and 13 in order to acquire data on the levels of selenium in these animals during 13-week gavage administration of l-seleno-methylselenocysteine (SeMC), a new candidate chemopreventive agent under development. Application of the commonly used method of standard addition was found to be unsuitable to calculate the selenium content in rat plasma (within-run and between-run accuracy and precision parameters were less than 85%). Therefore, a new analytical method was developed. In this method, samples of rat plasma (50 microL) were diluted 10-fold with a reducing agent containing l-ascorbic acid, a modifier solution containing palladium chloride and Triton X-100. Samples were atomized in pyrolytically coated graphite tubes and peak height signals were measured. Selenium concentrations were determined by linear least squares regression analysis based on the standard curve generated in pooled rat blank plasma. Since selenium is normally present in plasma, a three-step approach was used to calculate selenium plasma levels. Initially selenium levels were determined based on the standard curve with selenium-spiked pool plasma. In the second step, background selenium levels in the pooled plasma were determined based on the same standard curve. In the third step, background level was added to the previously derived number. The relative errors were in the range from -4.6 to 11.4% (intra-day assay) and from -0.4 to 8.8% (inter-day assay) which proved good accuracy. The relative standard deviations were in the range from 1.88 to 4.70% (intra-day precision) and from 3.28 to 5.38% (inter-day precision). In rat plasma, the following dose-dependent selenium levels (mean+/-S.D.) in males and females, respectively, were observed at 13 weeks: 655.5+/-48.8 and 595.8+/-43.9 ng/mL (control group), 927.9+/-85.3 and 859.3+/-164.3 ng/mL (0.4 mg/kg per day dose group), 1238.9+/-182.4 and 1169.9+/-112.6 ng/mL (0.8 mg/kg per day dose group), and 1476.5+/-138.1 and 1320.1+/-228.6 ng/mL (2.0mg/kg per day dose group). No significant sex differences in selenium plasma levels were seen in the SeMC-treated groups. No significant differences in selenium plasma levels were seen between mean plasma levels at 7 and 13 weeks. The described method is simple, rapid, accurate, precise and can be easily applied in other laboratories for a large number of samples.     

Pathologic high shear stress induces apoptosis events in human platelets

Recently, it has been discovered that apoptosis of anucleate platelets can be induced by chemical agonists. Other studies demonstrated that mechanical forces (shear stresses) stimulate platelet activation and signaling in the absence of exogenous chemical stimuli. We analyzed whether shear stresses can trigger platelet apoptosis, a question that has not yet been studied. Using a cone-and-plate viscometer, we exposed human platelet-rich plasma to different shear stresses, ranging from physiologic arterial and arteriole levels (10-44 dyn/cm2) to pathologic high levels (117-388 dyn/cm2) occurring in stenotic vessels. We found that pathologic shear stresses induce not only platelet activation (P-selectin upregulation and GPIbalpha downregulation) but also trigger apoptosis events, including mitochondrial transmembrane potential depolarization, caspase 3 activation, phosphatidylserine exposure, and platelet shrinkage and fragmentation, whereas physiological shear stresses are not effective.This novel finding suggests that shear-induced platelet apoptosis can be mediated by mechanoreceptors, does not require nuclear participation, and may affect platelet clearance.     

Estimating the elastic properties of the eye from differential tonometry by the Schiøtz tonometer: Analysis of the measurement procedure on the basis of a two-component mathematical model

Biophysics - The procedure of differential tonometry by the Schiøtz tonometer is modeled using a two-component model of the eyeball in which the cornea is represented by a momentless...     

Mechanisms of inhibitory effects of polycyclic aromatic hydrocarbons in photosynthetic primary processes in pea leaves and thylakoid preparations

• Inhibitory effects of polycyclic aromatic hydrocarbons (PAHs) on plants were studied in pea leaves in order to elucidate the mechanisms of action of PAHs such as naphthalene (Naph) and phenanthrene (Phen) on activity of photosystem II (PSII).
• The changes in different Chl fluorescence parameters were calculated on the basis of Chl fluorescence induction curves. H₂O₂ content was measured in leaf homogenates with the luminol‐dependent chemiluminescence method.
• We demonstrated that following PAH treatment, total energy dissipation (DI₀/ABS) and amount of Q_B‐non‐reducing complexes of PSII significantly increased. Non‐photochemical quenching (NPQ) also increased, when weak oxidative stress after PAH application developed. In leaves, a two‐step increase in H₂O₂ was found with time of incubation in the presence of PAHs, which may be associated with damage to the lipid bilayer of the plasma membrane and then violation of lipid bilayer membranes of cell organelles.
• A hypothesis for the mode of action of PAHs is provided that involves the role of ROS, membrane permeability and associated functional changes in PSII.

     

The nuts and bolts of ring-translocase structure and mechanism

Ring-shaped, oligomeric translocases are multisubunit enzymes that couple the hydrolysis of Nucleoside TriPhosphates (NTPs) to directed movement along extended biopolymer substrates. These motors help unwind nucleic acid duplexes, unfold protein chains, and shepherd nucleic acids between cellular and/or viral compartments. Substrates are translocated through a central pore formed by a circular array of catalytic subunits. Cycles of nucleotide binding, hydrolysis, and product release help reposition translocation loops in the pore to direct movement. How NTP turnover allosterically induces these conformational changes, and the extent of mechanistic divergence between motor families, remain outstanding problems. This review examines the current models for ring-translocase function and highlights the fundamental gaps remaining in our understanding of these molecular machines.     

Mathematical Modeling of Applanation and Impression Tonometry after Surgical Correction of Vision

Biophysics - Abstract—Mathematical modeling of applanation and impression tonometry (according to Maklakoff and Schiøtz, respectively) was performed for the eye with artificially induced...     

Photoregulation of the Cytoplasmic PGA Dehydrogenase Complex in Wheat Leaves

Russian Journal of Plant Physiology - Experiments were performed with 10-day-old seedlings of wheat (Triticum aestivum L.) grown on sand saturated with Knop’s solution at a temperature of...