ATP-dependent K+ channels in renal ischemia reperfusion injury

ATP-dependent K+ channels (K-ATP) account for most of the recycling of K+ which enters the proximal tubules cell via Na, K-ATPase. In the mitochondrial membrane, opening of these channels preserves mitochondrial viability and matrix volume during ischemia. We examined KATP channel modulation in renal ischemia-reperfusion injury (IRI), using an isolated perfused rat kidney (IPRK) model, in control, IRI, IRI + 200 muM diazoxide (a K-ATP opener), IRI + 10 muM glibenclamide (a K-ATP blocker) and IRI + 200 muM diazoxide + 10 muM glibenclamide groups. IRI was induced by 2 periods of warm ischemia, followed by 45 min of reperfusion. IRI significantly decreased glomerular filtration rate (GFR) and increased fractional excretion of sodium (FENa) (p < 0.01). Neither diazoxide nor glibenclamide had an effect on control kidney function other than an increase in renal vascular resistance produced by glibenclamide. Pretreatment with 200 muM diazoxide reduced the postischemic increase in FENa (p < 0.05). Adding 10 muM glibenclamide inhibited the diazoxide effect on postischemic FENa (p < 0.01). Histology showed that kidneys pretreated with glibenclamide demonstrated an increase in injure in the thick ascending limb of outer medulla (p < 0.05). Glibenclamide significantly decreased post ischemic renal vascular resistance (p < 0.05). but had no significant effect on other renal function parameters. Our results suggest that sodium reabsorption is improved by K-ATP activation and blockade of K-ATP channels during IRI has an injury enhancing effect on renal epithelial function and histology. This may be mediated through K-ATP modulation in cell and or mitochondrial inner membrane.

Complete genomic sequence of the Australian south-west genotype of Sindbis virus: Comparisons with other Sindbis strains and identification of a unique deletion in the 3 '-untranslated region

Our previous studies have shown that two distinct genotypes of Sindbis (SIN) virus occur in Australia. One of these, the Oriental/Australian type, circulates throughout most of the Australian continent, whereas the recently identified south-west (SW) genetic type appears to be restricted to a distinct geographic region located in the temperate south-west of Australia. We have now determined the complete nucleotide and translated amino acid sequences of a SW isolate of SIN virus (SW6562) and performed comparative analyses with other SIN viruses at the genomic level. The genome of SW6562 is 11,569 nucleotides in length, excluding the cap nucleotide and poly (A) tail. Overall this virus differs from the prototype SIN virus (strain AR339) by 23% in nucleotide sequence and 12.5% in amino acid sequence. Partial sequences of four regions of the genome of four SW isolates were determined and compared with the corresponding sequences from a number of SIN isolates from different regions of the World. These regions are the non-structural protein (nsP3), the E2 gene, the capsid gene, and the repeated sequence elements (RSE) of the 3'UTR. These comparisons revealed that the SW SIN viruses were more closely related to South African and European strains than to other Australian isolates of SIN virus. Thus the SW genotype of SIN virus may have been introduced into this region of Australia by viremic humans or migratory birds and subsequently evolved independently in the region. The sequence data also revealed that the SW genotype contains a unique deletion in the RSE of the 3'UTR region of the genome. Previous studies have shown that deletions in this region of the SIN genome can have significant effects on virus replication in mosquito and avian cells, which may explain the restricted distribution of this genotype of SIN virus.

Structure of the HERG K+ channel S5P extracellular linker - Role of an amphipathic alpha-helix in C-type inactivation

The HERG K+ channel has very unusual kinetic behavior that includes slow activation but rapid inactivation. These features are critical for normal cardiac repolarization as well as in preventing lethal ventricular arrhythmias. Mutagenesis studies have shown that the extracellular peptide linker joining the fifth transmembrane domain to the pore helix is critical for rapid inactivation of the HERG K+ channel. This peptide linker is also considerably longer in HERG K+ channels, 40 amino acids, than in most other voltage-gated K+ channels. In this study we show that a synthetic 42-residue peptide corresponding to this linker region of the HERG K+ channel does not have defined structural elements in aqueous solution; however, it displays two well defined helical regions when in the presence of SDS micelles. The helices correspond to Trp(585)-Ile(593) and Gly(604)-Tyr(611) of the channel. The Trp(585)-Ile(593) helix has distinct hydrophilic and hydrophobic surfaces. The Gly(604)-Tyr(611) helix corresponds to an N-terminal extension of the pore helix. Electrophysiological studies of HERG currents following application of exogenous S5P peptides show that the amphipathic helix in the S5P linker interacts with the pore region of the channel in a voltage-dependent manner.

Compositional changes of Australia-grown Western Schley pecans [Carya illinoinensis (Wangenh.) K. Koch] during maturation

Changes in composition during the maturation of Western Schley pecans [Carya illinoinensis (Wangenh.) K. Koch] grown in Australia were investigated. Pecans of different maturity levels were collected at monthly intervals between March and June in. 1999 and 2000 and analyzed for the concentrations of moisture, total lipid, sucrose, raffinose, protein, and the minerals aluminum, boron, calcium, copper, iron, potassium, magnesium, manganese, sodium, phosphorus, sulfur, and zinc. Moisture, total lipid, and calcium contents changed significantly (p < 0.05) with harvest time and maturity, whereas the other components did not. Western Schley pecans grown in Australia should be harvested after the shuck has opened and it is either green or brown in color to maximize total lipid content and quality. This occurred after May 11 in 1999 and after May 17 in 2000.

Comparative study of the radiation-induced grafting of styrene onto poly(tetrafluoroethylene-co-perfluoropropylvinyl ether) and polypropylene substrates. I: Kinetics and structural investigation

A comparative study has been made of the radiation grafting of styrene onto poly(tetrafluoroethylene-co-perfluoropropyl vinyl ether) (PFA) and polypropylene (PP) substrates, using the simultaneous irradiation method. Effects of grafting conditions such as monomer concentrations, type of solvent, dose rate and irradiation dose on the grafting yield were investigated. Under the same grafting conditions it was found that a higher degree of grafting of styrene was obtained using a mixture of dichloromethane/methanol solvents for PFA and methanol for PP and the degree of grafting was higher in PP than in PFA at all doses. However, the micro-Raman spectroscopy analysis of the graft revealed that, for the same degree of grafting, the penetration depth of the grafted polystyrene into the substrate was higher in PFA than in PP substrates. In both polymers the crystallinity was hardly affected by the grafting process and the degree of crystallinity decreased slightly with grafting dose. The dependence of the initial rate of grafting on the dose rate and the monomer concentration was found to be 0.6 and 1.4 order for PFA and 0.15 and 2.2 for PP, respectively. The degree of grafting increased with increasing radiation dose in both polymers. However, the grafting yield decreased with an increase in the dose rate. The increase in the overall grafting yield for PFA and PP was accompanied by a proportional increase in the penetration depth of the graft into the substrates. (C) 2003 Society of Chemical Industry.

G-values for scission and crosslinking on γ-radiolysis of Ultem at 303 K

The radiation chemical yields G(S) and G(X) for H-linking and Y-linking models for Ultem have been calculated from molecular weight analysis by gel permeation chromatography. These G-values have been compared with the G-values obtained from analysis of soluble fractions above the gel dose, Which have been reported in previous works. An analysis of the molecular weight data in terms of H-linking and Y-linking mechanisms yielded values of G(S-H) = 1.0 x 10(-3) and G(H) = 6.0 x 10(-3) and G(S-Y) = 1.3 x 10(-2) and G(Y) = 1.8 x 10(-2). The corresponding values obtained from the solubility data were G(SH) = 0.53 x 10(-2), G(H) = 1.39 x 10(-2), G(S-Y) = 4.2 x 10(-2) and G(Y) - 4.6 x 10(-2). The origin of the disagreement between the molecular weight and solubility values is not clear, but it could arise as a result of observed microgel formation below the reported gel dose of 0.13 MGy. Whether the crosslink mechanism proceeds by an H-linking or Y-linking process is also unclear and will require direct observation of the crosslinking structures.

High energy radiation grafting of fluoropolymers

Fluoropolymers are known as chemically inert materials with good high temperature resistance, so they are often the materials of choice for harsh chemical environments. These properties arise because the carbon-fluorine bond is the strongest of all bonds between other elements and carbon, and, because of their large size, fluorine atoms can protect the carbon backbone of polymers such as poly(tetrafluoroethylene), PTFE, from chemical attack. However, while the carbon-fluorine bond is much stronger than the carbon hydrogen bond, the G values for radical formation on high energy radiolysis of fluoropolymers are roughly comparable to those of their protonated counterparts. Thus, efficient high energy radiation grafting of fluoropolymers is practical, and this process can be used to modify either the surface or bulk properties of a fluoropolymer. Indeed, radiation grafted fluoropolymers are currently being used as separation membranes for fuel cells, hydrophilic filtration membranes and matrix substrate materials for use in combinatorial chemistry. Herein we present a review of recent studies of the high energy radiation grafting of fluoropolymers and of the analytical methods available to characterize the grafts. (C) 2003 Elsevier Ltd. All rights reserved.

Investigation of the vapor-phase grafting of styrene onto PFA

Poly(tetrafluoroethylene-co-perfluoropropyI vinyl ether), PFA, was grafted with styrene from the vapor phase using a simultaneous radiation grafting method. The graft yields were measured as a function of the dose and dose rate and were found to be initially linearly dependent on the dose and independent of the dose rate up to dose rates of similar to3 kGy/h. However, at a dose rate of 6.2 kGy/h, the slope of the yield-grafting time plot decreased. Raman depth profiles of the grafts showed that the polystyrene concentrations were greatest near the surface of the grafted samples and decreased with depth. The maximum penetration depth of the graft depended on the radiation dose for a fixed dose rate. Fmoc-Rink loading tests showed that the grafts displayed superior loading compared to grafts prepared from bulk styrene or from styrene solutions other than methanol.

Cross-linking of PFA by electron beam irradiation

The effect of electron beam radiation on a perfluoroalkoxy (PFA) resin was examined using solid-state high-speed magic angle spinning F-19 NMR spectroscopy and FT-IR spectroscopy. Samples were prepared for analysis by subjecting them to electron beam radiation in the dose range 0.5-2.0 MGy at 633 K, which is above the crystalline melting temperature. The new structures were identified and include new saturated chain ends, short and long branches, unsaturated groups, and cross-links. The radiation chemical yield (G value) of new long branch points was greater than the G value of new chain ends, suggesting that cross-linking is the net radiolytic process. This conclusion was supported by an observed decrease in the crystallinity and an increase in the optical clarity of the polymer.

Ventricular performance and Na+-K+ ATPase activity are reduced early and late after myocardial infarction in rats

Myocardial infarction leads to compensatory ventricular remodeling. Disturbances in myocardial contractility depend on the active transport of Ca2+ and Na+, which are regulated by Na+-K+ ATPase. Inappropriate regulation of Na+-K+ ATPase activity leads to excessive loss of K+ and gain of Na+ by the cell. We determined the participation of Na+-K+ ATPase in ventricular performance early and late after myocardial infarction. Wistar rats (8-10 per group) underwent left coronary artery ligation (infarcted, Inf) or sham-operation (Sham). Ventricular performance was measured at 3 and 30 days after surgery using the Langendorff technique. Left ventricular systolic pressure was obtained under different ventricular diastolic pressures and increased extracellular Ca2+ concentrations (Ca2+e) and after low and high ouabain concentrations. The baseline coronary perfusion pressure increased 3 days after myocardial infarction and normalized by 30 days (Sham 3 = 88 ± 6; Inf 3 = 130 ± 9; Inf 30 = 92 ± 7 mmHg; P < 0.05). The inotropic response to Ca2+e and ouabain was reduced at 3 and 30 days after myocardial infarction (Ca2+ = 1.25 mM; Sham 3 = 70 ± 3; Inf 3 = 45 ± 2; Inf 30 = 29 ± 3 mmHg; P < 0.05), while the Frank-Starling mechanism was preserved. At 3 and 30 days after myocardial infarction, ventricular Na+-K+ ATPase activity and contractility were reduced. This Na+-K+ ATPase hypoactivity may modify the Na+, K+ and Ca2+ transport across the sarcolemma resulting in ventricular dysfunction.

Acúmulo e liberação de P, K, Ca e Mg em crotalária e milheto solteiros e consorciados

As características das leguminosas e gramíneas usadas como plantas de cobertura são bastante conhecidas, mas a velocidade com que os nutrientes associados à biomassa vegetal tornam-se disponíveis às culturas é ainda pouco estudada. O objetivo deste trabalho foi determinar o acúmulo e a taxa de liberação de nutrientes de crotalária e milheto solteiros ou consorciados. O delineamento experimental adotado foi blocos ao acaso, com quatro repetições. Os tratamentos foram crotalária, milheto, crotalária + milheto e vegetação espontânea. O acúmulo de P e Mg foi influenciado pela produção de massa, com valores elevados na crotalária, enquanto o acúmulo de Ca resultou tanto do maior teor quanto da maior produção de massa nos tratamentos com a leguminosa. A liberação de nutrientes dos resíduos apresentou dinâmica semelhante, com duas fases distintas: a primeira com taxas mais elevadas nos primeiros 15 dias e a segunda mais lenta a partir dessa fase. As taxas de liberação de K, Ca e Mg foram semelhantes, independentemente dos tratamentos. Os resíduos da crotalária apresentaram maior liberação de P do que as da vegetação espontânea, com meia vida de 11 dias e a vegetação espontânea de 17 dias. O Ca foi liberado mais lentamente do que os demais nutrientes, e o Mg mais rapidamente. As plantas espontâneas e o milheto apresentam importante potencial na reciclagem de K, e a crotalária se destacou na reciclagem de Ca e Mg.

Fiber average size and distribution dependence on the electrospinning parameters of poly(vinylidene fluoride trifluoroethylene)) membranes for biomedical applications

Poly(vinylidene fluoride-trifluoethylene) electrospun membranes were obtained from a blend of dimethylformamide (DMF) and methylethylketone (MEK) solvents. The inclusion of the MEK to the solvent system promotes a faster solvent evaporation allowing complete polymer crystallization during the jet travelling between the tip and the grounded collector. Several processing parameters were systematically changed to study their influence on fiber dimensions. Applied voltage and inner needle diameter do not have large influence on the electrospun fiber average diameter but in the fiber diameter distribution. On the other hand, the increase of the distance between the needle tip to collector results in fibers with larger average diameter. Independently on the processing conditions, all mats are produced in the electroactive phase of the polymer. Further, MC-3T3-E1cell adhesion was not inhibited by the fiber mats preparation, indicating their potential use for biomedical applications.

Osteoblast, fibroblast and in vivo biological response to poly(vinylidene fluoride) based composite materials

Electroactivematerials can be taken to advantage for the development of sensors and actuators as well as for novel tissue engineering strategies. Composites based on poly(vinylidenefluoride),PVDF,have been evaluated with respect to their biological response. Cell viability and proliferation were performed in vitro both with Mesenchymal Stem Cells differentiated to osteoblasts and Human Fibroblast Foreskin 1. In vivo tests were also performed using 6-week-old C57Bl/6 mice. It was concluded that zeolite and clay composites are biocompatible materials promoting cell response and not showing in vivo pro-inflammatory effects which renders both of them attractive for biological applications and tissue engineering, opening interesting perspectives to development of scaffolds from these composites. Ferrite and silver nanoparticle composites decrease osteoblast cell viability and carbon nanotubes decrease fibroblast viability. Further, carbon nanotube composites result in a significant increase in local vascularization accompanied an increase of inflammatory markers after implantation.

Influence of electrospinning parameters on Poly(hydroxybutyrate) electrospun membranes fiber size and distribution

Poly(hydroxybutyrate) (PHB) obtained from sugar cane was dissolved in a blend of chloroform and dimethylformamide (DMF) and electrospun at 40 ºC. By adding DMF to the solution, the electrospinning process for the PHB polymer becomes more stable, allowing complete polymer crystallization during the jet travelling between the tip and the grounded collector. The influence of processing parameters on fiber size and distribution was systematically studied. It was observed that an increase of tip inner diameter promotes a decrease of the fiber average size and a broader distribution. On the other hand, an increase of the electric field and flow rate produces an increase of fiber diameter until a maximum of ~2.0 m, but for electric fields higher than 1.5 kV.cm-1, a decrease of the fiber diameter was observed. Polymer crystalline phase seems to be independent of the processing conditions and a crystallinity degree of 53 % was found. Moreover, thermal degradation of the as-spun membrane occurs in single step degradation with activation energy of 91 kJ/mol. Furthermore, MC-3T3-E1 cell adhesion was not inhibited by the fiber mats preparation, indicating their potential use for biomedical applications.

Fatigue Prediction on Fibrin Poly-ε-caprolactone Macroporous Scaffolds

Tissue engineering applications rely on scaffolds that during its service life, either for in-vivo or in vitro applications, are under mechanical solicitations. The variation of the mechanical condition of the scaffold is strongly relevant for cell culture and has been scarcely addressed. Fatigue life cycle of poly-ε-caprolactone, PCL, scaffolds with and without fibrin as filler of the pore structure were characterized both dry and immersed in liquid water. It is observed that the there is a strong increase from 100 to 500 in the number of loading cycles before collapse in the samples tested in immersed conditions due to the more uniform stress distributions within the samples, the fibrin loading playing a minor role in the mechanical performance of the scaffolds