Enhancing the fungicidal activity of amphotericin B via vacuole disruption by benzyl isothiocyanate,a cruciferous plant constituent

Amphotericin B (AmB), a typical polyene macrolide antifungal agent, is widely used to treat systemic mycoses. In the present study, we show that the fungicidal activity of AmB was enhanced by benzyl isothiocyanate (BITC), a cruciferous plant-derived compound, in the budding yeast, Saccharomyces cerevisiae. In addition to forming a molecular complex with ergosterol present in fungal cell membranes to form K⁺-permeable ion channels, AmB has been recognized to mediate vacuolar membrane disruption resulting in lethal effects. BITC showed no effect on AmB-induced plasma membrane permeability; however, it amplified AmB-induced vacuolar membrane disruption in S. cerevisiae. Furthermore, the BITC-enhanced fungicidal effects of AmB significantly decreased cell viability due to the disruption of vacuoles in the pathogenic fungus Candida albicans. The application of the combinatorial antifungal effect of AmB and BITC may aid in dose reduction of AmB in clinical antifungal therapy and consequently decrease side effects in patients. These results also have significant implications for the development of vacuole-targeting chemotherapy against fungal infections.     

Interacting Effects of Discharge and Channel Morphology on Transport of Semibuoyant Fish Eggs in Large,Altered River Systems

Habitat fragmentation and flow regulation are significant factors related to the decline and extinction of freshwater biota. Pelagic-broadcast spawning cyprinids require moving water and some length of unfragmented stream to complete their life cycle. However, it is unknown how discharge and habitat features interact at multiple spatial scales to alter the transport of semi-buoyant fish eggs. Our objective was to assess the relationship between downstream drift of semi-buoyant egg surrogates (gellan beads) and discharge and habitat complexity. We quantified transport time of a known quantity of beads using 2–3 sampling devices at each of seven locations on the North Canadian and Canadian rivers. Transport time was assessed based on median capture time (time at which 50% of beads were captured) and sampling period (time period when 2.5% and 97.5% of beads were captured). Habitat complexity was assessed by calculating width∶depth ratios at each site, and several habitat metrics determined using analyses of aerial photographs. Median time of egg capture was negatively correlated to site discharge. The temporal extent of the sampling period at each site was negatively correlated to both site discharge and habitat-patch dispersion. Our results highlight the role of discharge in driving transport times, but also indicate that higher dispersion of habitat patches relates to increased retention of beads within the river. These results could be used to target restoration activities or prioritize water use to create and maintain habitat complexity within large, fragmented river systems.     

Hypophysiotropic neurons in the goldfish hypothalamus demonstrated by retrograde transport of horseradish peroxidase

Summary The horseradish-peroxidase (HRP) technique was used to visualize the cell bodies of axons projecting to the goldfish pituitary. Following intravenous injections of HRP, HRP reaction products were observed in axons of the rostral pars distalis, proximal pars distalis, neurointermediate lobe, pituitary stalk and in axons coursing from the pituitary into the hypothalamus. HRP-labelled cells in the brain were localized in two regions only — the nucleus preopticus (NPO) pars magnocellularis and pars parvocellularis, and the nucleus lateralis tuberis (NLT) of the hypothalamus. These observations suggest that the NPO and NLT are the source of the neurosecretory innervation of the goldfish pituitary.     

Tissue distribution and molecular heterogeneity of bovine thiol:protein-disulphide oxidoreductase (disulphide interchange enzyme)

1. The distribution of thiol:protein-disulphide oxidoreductase (disulphide interchange enzyme) in 17 bovine tissue extracts was determined by rocket immunoelectrophoresis and by measuring the reductive cleavage of insulin. 2. The relative concentration (per mg total protein) was found to be in the order: Pancreas greater than liver greater than lymph node greater than testes, fat tissue greater than parotid gland, brain, spleen, lung greater than small intestine, spinal cord, large intestine, kidney greater than paunch, aorta greater than skeletal muscle greater than heart. 3. The distribution of specific activity showed a similar pattern, irrespectively of whether glutathione or L-cysteine was used as cosubstrate. 4. The concentration varied 200-fold and the specific activity 400-fold between pancreas and heart muscle, respectively. 5. Crossed immunoelectrophoresis demonstrated that a fast-migrating form of the enzyme was the only one present in almost all tissues, but 15% of the enzyme in liver was a slow-migrating form and 50% in heart muscle a medium-migrating form. 6. The lung contains a species having partial immunological identity to the enzyme. 7. Purified enzyme from bovine liver has a somewhat lower mobility than the fast-migrating form in extract. 8. The results seem to support the general view that the enzyme is involved in synthesis of disulphide-bonded extracellular proteins, although the presence of the enzyme in tissues like fat, brain, spinal cord, skeletal muscle and heart indicates other cellular functions as well.     

Mechanism of the change in erythrocyte osmotic resistance in rats exposed to valinomycin: the features seen in spontaneous hypertension

Introduction of valinomycin into erythrocyte incubation medium increased the cell stability to water-induced hemolysis. In these conditions the erythrocytes of spontaneously hypertensive and normotensive (control) rats release 63.2 +/- 1.5% and 80.9 +/- 1.6%, respectively, of the total hemoglobin content. Valinomycin effect is completely abolished with K+ substitution for Na+ and is independent of extracellular Ca2+ concentration. Valinomycin had no effect on human erythrocyte osmotic stability. It has been shown that valinomycin-induced kinetics of Na+ and K+ redistribution was different in human and rat erythrocytes. The distinctions are thought to be related to specific anion transport mediated by the third band protein--the main component of membrane cytoskeleton.     

Mechanism of the binding of 2-(4'-hydroxyphenylazo)benzoic acid to bovine serum albumin

The mechanism of the binding of 2-(4'-hydroxyphenylazo)benzoic acid (HABA) to bovine serum albumin was studied by relaxation methods as well as the binding isotherm using gel chromatography. A single relaxation was observed over a wide range of HABA concentration except at the extremes of high concentration where another slow process was observed. The concentration dependence of the reciprocal relaxation time of the fast process decreased monotonically with increase in concentration of HABA at constant polymer concentration. The data were analyzed on the basis of Brown's domain structure model and were found to be consistent with a sequential binding mechanism. The azohydrazon tautomerism of HABA was identified with the intramolecular step of the complex. The activation parameters of the step, determined from the temperature dependence of the relaxation time of the fast process, showed that this step is rate limited by an enthalpy barrier in both forward and backward directions. Comparison of the activation parameters with those of other serum albumin-ligand systems suggests that there is an enthalpy-entropy compensation in the activation process of the intramolecular step with the compensation temperature at about 270 K; the enthalpy-entropy compensation is thought to be related to the hydrophobic nature of the ligand.