Hsp70 and Hsp40 attenuate formation of spherical and annular polyglutamine oligomers by partitioning monomer

Protein conformational changes that result in misfolding, aggregation and amyloid fibril formation are a common feature of many neurodegenerative disorders. Studies with beta-amyloid (Abeta), alpha-synuclein and other amyloid-forming proteins indicate that the assembly of misfolded protein conformers into fibrils is a complex process that may involve the population of metastable spherical and/or annular oligomeric assemblies. Here, we show by atomic force microscopy that a mutant huntingtin fragment with an expanded polyglutamine repeat forms spherical and annular oligomeric structures reminiscent of those formed by Abeta and alpha-synuclein. Notably, the molecular chaperones Hsp70 and Hsp40, which are protective in animal models of neurodegeneration, modulate polyglutamine aggregation reactions by partitioning monomeric conformations and disfavoring the accretion of spherical and annular oligomers.     

Hippophae rhamnoides L.: chromatographic methods to determine chemical composition, use in traditional medicine and pharmacological effects

There is an increasing interest in the usage of chromatographic methods on the analysis of chemical compounds present in Hippophae rhamnoides L. In this paper, the chromatographic techniques applied for the determination, separation and identification of chemical compounds of H. rhamnoides L. are reviewed. We examined the existing chromatographic methods based on separations by paper and thin-layer chromatography, gas chromatography, high-performance liquid chromatography and capillary electrophoresis and also methods of detection by ultraviolet absorption, fluorescence, refractive index, electrochemical and mass spectrometry. Biological properties of the plant and its pharmacological effects and use in traditional medicine have also been reviewed.     

Comparison of the effects of specific and nonspecific inhibition of nitric oxide synthase on morphine analgesia,tolerance and dependence in mice

The effects of L-Canavanine, a selective inducible nitric oxide synthase (NOS) inhibitor and N(G)-nitro-L-arginine methyl ester (L-NAME), a nonselective NOS inhibitor, on pain threshold and morphine induced analgesia, tolerance and dependence in mice were investigated and compared. Morphine was administered by subcutaneous implantation of a pellet containing 40 mg free base and withdrawal was precipitated by intraperitoneal (i.p.) injection of naloxone (2 mg/kg). L-Canavanine (200 mg/kg, i.p.) did not affect the pain threshold, morphine-induced analgesia and the induction and expression phases of morphine tolerance and dependence. L-NAME (20 mg/kg, i.p.) significantly (p < 0.05) enhanced the pain threshold, potentiated morphine-induced analgesia and attenuated the expression phase of morphine dependence which has been characterized by withdrawal signs and body weight loss, but did not modify the induction phase of morphine tolerance and dependence. It is concluded that constitutive NOS isoforms which were inhibited by L-NAME may be involved specifically in the mechanisms of morphine induced analgesia, tolerance and dependence.     

Dantrolene protects erythrocytes against oxidative stress during whole-body irradiation in rats

In our study, we examined the radioprotective effects of dantrolene against gamma irradiation-induced damage of blood cells after total body irradiation of rats. Rats were divided into three groups of eight rats each. The first group was the control group receiving no dantrolene or irradiation, the second group received total body irradiation (RT) with 5 Gy of gamma irradiation only, and the third group received dantrolene at a dose of 5 mg x kg(-1) plus RT. Dantrolene was given intraperitoneally 30 min before RT. All groups were sacrificed 2 h after RT, and blood samples were taken. Leukocyte, and thrombocyte counts and hemoglobin levels were measured. Furthermore, malondialdehyde (MDA) levels in plasma and erythrocytes and superoxide dismutase (SOD) and glutathione peroxidase activities (GSH-Px) in erythrocytes were determined. It was found that pretreatment with dantrolene at a dose of 5 mg x kg(-1) significantly reduced the MDA levels and increased the antioxidant SOD and GSH-Px activities, and prevented the decrease in leukocyte and thrombocyte counts. We conclude that dantrolene has clear antioxidant properties when given prior to radiation exposure and the protective effect of dantrolene against damage inflicted by radiation, depends, at least in part, on the decrease in lipid peroxidation and increase in the activity of antioxidant enzymes SOD and GSH-Px.     

The in vitro and in vivo inhibitory effects of some sulfonamide derivatives on rainbow trout (Oncorhynchus mykiss) erythrocyte carbonic anhydrase activity

The in vitro and in vivo inhibitory effects of 5-(3alpha, 12alpha-dihydroxy-5-beta-cholanamido)-1,3,4-thiadiazole-2-sulfonamide (1), 5-(3alpha, 7alpha, 12alpha-trihydroxy-5-beta-cholanamido)-1,3,4-thiadiazole-2-sulfonamide (2), 5-(3alpha, 7alpha, 12alpha-triacetoxy-5-beta-cholanamido)-1,3,4-thiadiazole-2-sulfonamide (3) and acetazolamide on rainbow trout (Oncorhynchus mykiss) (RT) erythrocyte carbonic anhydrase (CA) were investigated. The RT erythrocyte CA was obtained by affinity chromatography with a yield of 20.9%, a specific activity of 422.5EU/mg protein and a purification of 222.4-fold. The purity of the enzyme was confirmed by SDS-PAGE. Inhibitory effects of the sulfonamides and acetazolamide on the RT erythrocyte CA were determined using the CO2-Hydratase method in vitro and in vivo studies. From in vitro studies, it was found that all the compounds inhibited CA. The obtained I50 value for the sulfonamides (1), (2) and (3) and acetazolamide were 0.83, 0.049, 0.82 and 0.052 microM, respectively. From in vivo studies, it was observed that CA was inhibited by the sulfonamides (1), (2) and (3) and acetazolamide.     

The kinetic basis of a general method for the investigation of active site content of enzymes and catalytic antibodies: first-order behaviour under single-turnover and cycling conditions

The theoretical foundation has been laid for the investigation of catalytic systems using first-order kinetics and for a general kinetic method of investigation of the active site content, E(a), of enzymes, catalytic antibodies, and other enzyme-like catalysts. The method involves a combination of steady-state and single-turnover kinetics to provide Vmax and Km and k(lim)(obs) and K(app)(m), respectively. The validity of the method is shown to remain valid for two extensions of the simple two-step enzyme catalysis model (a) when the catalyst preparation contains molecules (Eb) that bind substrate but fail to catalyse product formation and (b) when the catalyst itself binds substrate non-productively as well as productively. The former is a particularly serious complication for polyclonal catalytic antibodies and the latter a potential complication for all catalysts. For the simple model and for (b) Vmax/k(lim)(obs) provides the value of [Ea]T and for (a) its upper limit. This can be refined by consideration of the relative values of Km and the equilibrium dissociation constant of EbS. For the polyclonal catalytic antibody preparation investigated, the fact that K(app/m) > Km demonstrates for the first time the presence of a substrate-binding but non-catalytic component in a polyclonal preparation. First-order behaviour in catalytic systems occurs not only with a large excess of catalyst over substrate but also with lower catalyst/substrate ratios, including the equimolar condition, when K(app)(m) > [S]0, a phenomenon that is not widely appreciated.     

Permeable Endothelium and the Interstitial Space of Brain

1. Fenestrated vessels can be reversibly induced in brain by agents that stimulate urokinase production. This plasminogen activator, like vascular endothelial growth factor and metalloproteinases, is secreted by tumor cells and may account for induction of fenestrated vessels. Why only some of the brain's barrier vessels are converted to fenestrated vessels is unknown.2. The structures responsible for the filtering of solutes by fenestrated vessels may be the same as those of continuous, less permeable vessels: the glycocalyx on the surfaces of the endothelial cells and the subendothelial basal lamina.3. Solutes leaving the cerebral ventricles immediately enter the interstitial clefts between the cells lining the ventricles. A fraction of a variety of solutes, injected into CSF compartments, is retained by subendothelial basal lamina, from which the solutes may be released in a regulated way.4. The brain's CSF and interstitial clefts are the conduits for nonsynaptic volume transmission of diffusible signals, e.g., ions, neurotransmitters, and hormones. This type of transmission could be abetted by a parallel, cell-to-cell volume transmission mediated by gap junctions between astrocytes bordering CSF compartments and parenchymal astrocytes.5. The width and contents of the interstitial clefts in fetal brain permit cell migration and outgrowth of neurites. The contents of the narrower and different interstitial clefts of mature brain permit solute convection but must be enzymatically degraded in order for cells to migrate through it.     

Cadmium-induced changes in epithelial cells of the rat stomach

The aim of this study was to determine the changes in the function and fine structure of the gastric mucosa following exposure to high cadmium (Cd) for 30 d in rats. In the present study, control antimals were fed with normal food and tap water and the remaining animals received Cd (15 ppm CdCl₂) in drinking water for the same period. Receiving Cd for 30 d increased the mean blood (p<0.01) and mucosa (p<0.001) Cd levels, while decreased mucus thickness, mucin content (p<0.01) significantly. Basal acid output fell significantly (p<0.01). Light and electron microscopic examination revealed the following: (1) Cd decreases the mean number of surface mucous, isthmic-neck, parietal cells (p<0.05) and chief cells (p<0.001) per unit from the control value and (2) in some cells of zymogenic unit, the Cd-induced alterations were characterized with dilated Golgi cisternae, focal enlarged endoplasmic reticulum, broken tubulovesicles, degenerated mitochondria, dense nuclei, as well as lysosomal structures. We concluded that Cd augments the elimination rate of zymogenic unit’s cells by increasing the alteration rate, and the reduced basal acid output, mucin content, and mucus thickness can be explained easily with the loss of zymogenic unit’s cell population.     

Global gene expression profiling in congenital diaphragmatic hernia (CDH) patients

Functional & Integrative Genomics - Congenital diaphragmatic hernia (CDH) is an anomaly characterized by a defect in the diaphragm, leading to the passage of intra-abdominal organs into the...     

Quantitative analysis of an anaphase B switch: predicted role for a microtubule catastrophe gradient

Anaphase B in Drosophila embryos is initiated by the inhibition of microtubule (MT) depolymerization at spindle poles, which allows outwardly sliding interpolar (ip) MTs to drive pole-pole separation. Using fluorescence recovery after photobleaching, we observed that MTs throughout the preanaphase B spindle are very dynamic and display complete recovery of fluorescence, but during anaphase B, MTs proximal to the poles stabilize and therefore display lower recovery than those elsewhere. Fluorescence microscopy of the MT tip tracker EB1 revealed that growing MT plus ends localize throughout the preanaphase B spindle but concentrate in the overlap region of interpolar MTs (ipMTs) at anaphase B onset. None of these changes occurred in the presence of nondegradable cyclin B. Modeling suggests that they depend on the establishment of a spatial gradient of MT plus-end catastrophe frequencies, decreasing toward the equator. The resulting redistribution of ipMT plus ends to the overlap zone, together with the suppression of minus-end depolymerization at the poles, could constitute a mechanical switch that initiates spindle elongation.