Effects of aluminium on polyphosphate mobilization of the ectomycorrhizal fungus Suillus bovinus

Aluminium toxicity may be an important factor in the decline in vitality of many forest trees and the associated ectomycorrhizal fungal flora. In this study, comparative in vivo ³¹P NMR investigations on Al-adapted and non-Aladapted fungus of Suillus bovinus in pure culture have produced interesting new data. With respect to intracellular compartments, ³¹P NMR spectroscopy showed the spectra to differ in a peak-6 ppm appearing in the spectra of the A1-adapted fungus indicating terminal phosphate groups of mobile polyphosphate. Thus, in the Al-adapted fungus the average chain length of mobile polyphosphate is considerably shorter than in the non-Al-adapted fungus. A special method of cyclic phosphate supply followed by block averaging of the NMR spectra was used to determine the kinetic behaviour of phosphate uptake, storage and incorporation into polyphosphate at a constant external pH 3.5. While the Al-adapted fungus showed resistance to Al, an irreversible break-down in phosphate metabolism of the non-Al-adapted fungus by exposure to Al was caused. In comparison with the non-Al-adapted fungus supplied by nutrient solutions omitting Al, the Al-adapted fungus showed higher levels in both phosphate uptake and mobile polyphosphate concentration. As a consequence of these results a de-toxification of freely mobile Al-ions into a stable and insoluble complex in the Al-adapted fungus is considered to be due to a capture of intracellular Al by mobile polyphosphate of shorter chain length.     

The Ethiopian community in Israel: Segregation and the creation of a racial cleavage

This study examines the processes that led to the creation of a segregated Ethiopian community in Israel and its implications for the development of a racially-based stratification system. Interviews with thirty Ethiopian immigrants reveal that ethnic ties played an important role in creating a segregated ethnic community. Many families were pushed to the neighbourhood by co-ethnic realtors or were influenced by relatives and friends who had already settled there. However, no ethnic networks were found in the domain of employment. This absence coupled with low human capital made joblessness an acute problem for many Ethiopian immigrants. Overall, Ethiopian immigrants’ high residential concentration was perceived as a barrier for the successful integration of the young generation. Considering Ethiopian immigrants’ lack of social and economic resources, there is concern that their segregation will be exacerbated in the future, leading to social marginalization and the creation of a racial cleavage in Israeli society.     

Determination of allantoin in bovine milk by high-performance liquid chromatography

Determination of allantoin in bovine milk based on high-performance liquid chromatography (HPLC) is described. Following dilution and filtration, milk samples were analysed directly. Separation and quantification of allantoin was achieved using a Spherisorb 5 NH₂ column (250×4.6 mm ID), acetonitrile–water (90:10, v/v) mobile phase at a flow-rate of 2.0 ml min⁻¹, temperature 20°C and monitoring the effluent at 214 nm. Total analysis time was 10 min. Recovery of allantoin added to milk was 97 (±3.7, n=30)%. Lowest detectable concentration was 1 μmol l⁻¹. Within-day and between-day variability were less than 3%. Advantages of improved retention and separation of allantoin, and less complicated sample preparation exist over current methods.     

Do stable nitroxide radicals catalyze or inhibit the degradation of hyaluronic acid?

Reactive oxygen-derived species and particularly OH radicals can degrade hyaluronic acid (HA), resulting in a loss of viscosity and a subsequent decrease in its effectiveness as a joint-lubricating agent. The production of OH in the vicinity of HA can be catalyzed by bound redox-active metals, which participate in the Haber-Weiss reaction. Damage to HA can also occur as a result of hypochlorite formed by myeloperoxidase (MPO). The protective reagents commonly used to inhibit oxidative stress-induced degradation of HA include antioxidative enzymes, such as SOD and catalase, chelators that coordinate metal ions rendering them redox-inactive, and scavengers of radicals, such as OH, as well as nonradical reactive species. In recent years, stable cyclic nitroxides have also been widely used as effective antioxidants. In many cases, nitroxide antioxidants operate catalytically and mediate their protective effect through an exchange between their oxidized and reduced forms. It was anticipated, therefore, that nitroxides would protect HA from oxidative degradation as well. On the other hand, nitroxides serve as catalysts in many oxidation reactions of alcohols, sugars and polysaccharides, including hyalouronan. Such opposite effects of nitroxides on oxidative degradation are particularly intriguing and the aim of the present study was to examine their effect on HA when subjected to diverse forms of oxidative stress. The results indicate that nitroxides protect HA from OH radicals generated enzymatically or radiolytically. The protective effect is attributable neither to the scavenging of OH nor to the oxidation of reduced metal, but to the reaction of nitroxides with secondary carbohydrate radicals-most likely peroxyl radicals.     

The pro-oxidative activity of SOD and nitroxide SOD mimics.

Native Cu,Zn-SOD and synthetic SOD mimics sometimes demonstrate an apparently anomalous bell-shaped dose-response relationship when protecting various biological systems from oxidative stress. Several mechanisms have been proposed to account for such an effect, including: overproduction of H(2)O(2), peroxidative activity of SOD, and opposing roles played by O(2)(*-) in both initiation and termination of radical chain reactions. In the present study, ferrocyanide and thiols, which are susceptible to one-electron and two-electron oxidation, respectively, were subjected to a flux of superoxide in the presence and absence of SOD or SOD mimics. The results show that 1) either O(2)(*-)/HO(2)(*) or H(2)O(2) alone partially inactivates papain, whereas when combined they act synergistically; 2) nitroxide SOD mimics, but not SOD, exhibit a bell-shaped dose-response relationship in protecting papain from inactivation; 3) SOD, which at low dose inhibits superoxide-induced oxidation of ferrocyanide, loses its antioxidative effect as its concentration increases. These findings offer an additional explanation for the pro-oxidative activity of SOD and SOD mimics without invoking any dual activity of O(2)(*-) or a combined effect of SOD and H(2)O(2). The most significant outcome of an increase in SOD level is a decrease of [O(2)(*-)](steady state), rather than any notable elevation of [H(2)O(2)](steady state). As a result, the reaction kinetics of the high oxidation state of each catalyst is altered. In the presence of ultra-low [O(2)(*-)](steady state), the oxidized form of SOD [Cu(II),Zn-SOD] or SOD mimic (oxo-ammonium cation) does not react with O(2)(*-) but rather oxidizes the target molecule that it was supposed to have protected. Consequently, these catalysts exert an anti- or pro-oxidative effect depending on their concentration.     

Impaired myocardial perfusion reserve in experimental hypercholesterolemia is independent of myocardial neovascularization

Our objective was to investigate the functional role of hypercholesterolemia-associated myocardial neovascularization in early atherosclerosis using the antiangiogenic thalidomide. Experimental atherosclerosis is characterized by myocardial neovascularization, associated with a decrease in myocardial perfusion response to challenge, coronary endothelial dysfunction, and high oxidative stress. However, the functional significance of these neovessels is not known. Three groups of pigs (n = 6 each) were studied after 12 wk of normal or hypercholesterolemic diet without (HC) or with thalidomide (HC + Thal). Myocardial perfusion and permeability were assessed at baseline and in response to cardiac challenge, using electron beam computed tomography, and coronary endothelial function was assessed using organ chambers. Myocardial samples were scanned ex vivo with a three-dimensional microscopic computed tomography scanner, and the spatial density of the myocardial microvessels was quantified. Growth factors and oxidative stress were measured in the myocardial tissue. As a results of these procedures, myocardial perfusion response to adenosine and dobutamine was blunted in both HC and HC + Thal pigs compared with normal pigs (P < 0.05, HC and HC + Thal vs. normal) as was the coronary endothelial function. Myocardial permeability response to adenosine was increased in both HC and HC + Thal pigs compared with normal pigs (P < 0.05, HC and HC + Thal vs. normal, and HC + Thal vs. HC). The microvascular density was increased in HC pigs compared with normal pigs but normalized in HC + Thal pigs (P < 0.001 HC vs. normal and HC + Thal). HC + Thal pigs showed decreased expression of Flk-1 and basic FGF but increased expression of VEGF compared with normal and HC pigs. Oxidative stress was increased in both HC and HC + Thal pigs compared with normal pigs. In conclusion, chronic administration of thalidomide attenuates myocardial neovascularization in experimental HC pigs without affecting myocardial perfusion response to stimulation. This suggests that the myocardial neovascularization may not contribute to the attenuated myocardial perfusion response in hypercholesterolemia.     

In Vivo Experiments Reveal the Good,the Bad and the Ugly Faces of sFlt-1 in Pregnancy

Objective

Soluble fms-like tyrosine kinase (sFlt)-1-e15a, a primate-specific sFlt-1-isoform most abundant in the human placenta in preeclampsia, can induce preeclampsia in mice. This study compared the effects of full-length human (h)sFlt-1-e15a with those of truncated mouse (m)sFlt-1(1-3) used in previous preeclampsia studies on pregnancy outcome and clinical symptoms in preeclampsia.

Methods

Mice were injected with adenoviruses or fiber-mutant adenoviruses overexpressing hsFlt-1-e15a, msFlt-1(1-3) or control GFP under the CMV or CYP19A1 promoters on gestational day 8 (GD8) and GD11. Placentas and pups were delivered by cesarean section, and dams were monitored postpartum. Blood pressure was telemetrically recorded. Urine samples were collected with cystocentesis and examined for albumin/creatinine ratios. Tissue specimens were evaluated for transgene as well as endogenous mFlt-1 and msFlt-1-i13 expression. H&E-, Jones- and PAS-stained kidney sections were histopathologically examined. Placental GFP expression and aortic ring assays were investigated with confocal microscopy.

Results

Mean arterial blood pressure (MAP) was elevated before delivery in hsFlt-1-e15a-treated mice compared to controls (GD18: ΔMAP = 7.8 mmHg, p = 0.009), while ΔMAP was 12.8 mmHg (GD18, p = 0.005) in msFlt-1(1-3)-treated mice. Urine albumin/creatinine ratio was higher in hsFlt-1-e15a-treated mice than in controls (GD18, p = 0.04; PPD8, p = 0.03), and msFlt-1(1-3)-treated mice had marked proteinuria postpartum (PPD8, p = 4×10⁻⁵). Focal glomerular changes were detected in hsFlt-1-e15a and msFlt-1(1-3)-treated mice. Aortic ring microvessel outgrowth was decreased in hsFlt-1-e15a (p = 0.007) and msFlt-1(1-3)-treated (p = 0.02) mice. Full-length msFlt-1-i13 expression was unique for the placenta. In hsFlt-1-e15a-treated mice, the number of pups (p = 0.046), total weight of living pups (p = 0.04) and maternal weights (p = 0.04) were higher than in controls. These differences were not observed in truncated msFlt-1(1-3)-treated mice.

Conclusions

Truncated msFlt-1(1-3) simulated the preeclampsia-promoting effects of full-length hsFlt-1. MsFlt-1(1-3) had strong effect on maternal endothelium but not on placentas and embryos. In contrast, hsFlt-1-e15a induced preeclampsia-like symptoms; however, it also increased litter size. In accord with the predominant placental expression of hsFlt-1-e15a and msFlt-1-i13, full-length sFlt-1 may have a role in the regulation of embryonic development. These observations point to the difference in the biological effects of full-length and truncated sFlt-1 and the changes in the effect of full-length sFlt-1 during pregnancy, and may have important implications in the management of preeclampsia.     

Skip residues correlate with bends in the myosin tail

Sharp bends have previously been observed in the tail of the skeletal myosin molecule at well-defined positions 44, 75 and 135 nm from the head-tail junction, and in vertebrate smooth myosin at two positions about 45 and 96 nm from this junction. The amino acid sequence of the heavy chain does not straightforwardly account for such bending on the original model of the tail in which an invariant proline residue is present at the head-tail junction and the repeating seven amino acid pattern of hydrophobic residues lies entirely in the tail. Recently, a revised model has been proposed by Rimm et al. in which the first seven to eight heptads lie in the heads. It is shown here that with this model the observed bends in the tail of skeletal myosin coincide with three of the four additional (skip) residues that interrupt the heptad repeat. It is concluded that the skip residues, by causing localized instability of the coiled-coil, are responsible for the bends. Smooth myosin lacks the second of these skip residues explaining the absence of a bend at 75 nm.