Direct analysis of inorganic anions in samples with high salt content by capillary zone electrophoresis.
(57/714)
Trace-level inorganic anions in seawater are separated efficiently by capillary zone electrophoresis using direct UV detection. The carrier electrolyte is 50 mM borate at pH 9.3 and contained 1.5M NaCl. This buffer solution is adopted to prevent interference from high concentrations of the chloride ion in seawater. No electro-osmotic flow reverser is used to shorten the analysis time. The experimental conditions such as the concentration of NaCl in the carrier electrolyte, capillary inner diameter, applied current, and temperature are optimized. Linear plots are obtained in the concentration range of 0.1 to 20 microg/mL. The quantitation limits of the anions are in the order of 0.02 to 0.1 microg/mL. The proposed method may be applicable to the determination of inorganic anions in other environmental samples and effluents of a power plant. (+info)
Studies on the structure of the rabbit kidney brush border.
(58/714)
The effects of salts and non-ionic detergents on renal brush borders have been studied. 2 M sodium chloride, iodide or thiocyanate dissociated up to 40% of the protein from the brush borders, destroying the core filaments and resulting in the formation of membrane vesicles; EDTA had a similar effect on structure but released little protein. Triton X-100 and Nonidet P-40 extracted up to 60% of the protein including the major membrane glycoproteins and the enzymes trehalase, maltase and aminopeptidase (microsomal). Triton exhibited a selective effect on lipids removing phosphatidylserine, phosphatidylethanolamine and sphingomyelin but not the bulk of the phosphatidylcholine or cholesterol. The residual structures after Triton extraction comprised the core filaments associated with vesicles of lipid containing alkaline phosphatase and several other proteins. Treatment of these core-vesicle complexes with 2 M sodium chloride dissociated the filaments, releasing the vesicles which could be recovered as a pellicle on centrifugation. It is suggested that the proteins found in the vesicles might serve to interconnect the core filaments with the lipid bilayer. (+info)
Telomerase activity of the Lugol-stained and -unstained squamous epithelia in the process of oesophageal carcinogenesis.
(59/714)
Up-regulation of telomerase has been reported in many cancers. Our aim was to characterize telomerase activity in various states of the oesophagus to facilitate better understanding of carcinogenesis of oesophageal squamous cell carcinoma. During endoscopic examinations, we obtained 45 Lugol-stained normal epithelia, 31 Lugol-unstained epithelia (14 oesophagitis, 7 mild dysplasia, 5 severe dysplasia and 5 intramucosal cancer) and 9 advanced cancer. Telomerase activity was semi-quantified by a telomeric repeat amplification protocol using enzyme-linked immunosorbent assay, and expression of human telomerase reverse transcriptase mRNA was examined by in situ hybridization. In the Lugol-stained normal epithelia, telomerase activity increased in proportion to the increase of severity of the accompanying lesions, with a rank order of advanced cancer, intramucosal cancer, mild dysplasia and oesophagitis. In the Lugol-unstained lesions and advanced cancer, telomerase activity was highest in advanced cancer. Up-regulation of telomerase in normal squamous epithelium may be a marker of progression of oesophageal squamous cell carcinoma. (+info)
Role of CFTR's PDZ1-binding domain, NBF1 and Cl(-) conductance in inhibition of epithelial Na(+) channels in Xenopus oocytes.
(60/714)
The cystic fibrosis transmembrane conductance regulator (CFTR) inhibits epithelial Na(+) channels (ENaC). Evidence has accumulated that both Cl(-) transport through CFTR Cl(-) channels and the first nucleotide binding domain (NBF1) of CFTR are crucial for inhibition of ENaC. A PDZ binding domain (PDZ-BD) at the C-terminal end links CFTR to scaffolding and cytoskeletal proteins, which have been suggested to play an important role in activation of CFTR and eventually inhibition of ENaC. We eliminated the PDZ-BD of CFTR and coexpressed Na(+)/H(+)-exchange regulator factors together with CFTR and ENaC. The results do not support a role of PDZ-BD in inhibition of ENaC by CFTR. However, inhibition of ENaC was closely linked to Cl(-) currents generated by CFTR and was observed in the presence of Cl(-), I(-) or Br(-) but not gluconate. Therefore, functional NBF1 and Cl(-) transport are required for inhibition of ENaC in Xenopus oocytes, while the PDZ-BD is not essential. (+info)
Activation of G551D CFTR channel with MPB-91: regulation by ATPase activity and phosphorylation.
(61/714)
We have designed and synthesized benzo[c]quinolizinium derivatives and evaluated their effects on the activity of G551D cystic fibrosis transmembrane conductance regulator (CFTR) expressed in Chinese hamster ovary and Fisher rat thyroid cells. We demonstrated, using iodide efflux, whole cell patch clamp, and short-circuit recordings, that 5-butyl-6-hydroxy-10-chlorobenzo[c]quinolizinium chloride (MPB-91) restored the activity of G551D CFTR (EC(50) = 85 microM) and activated CFTR in Calu-3 cells (EC(50) = 47 microM). MPB-91 has no effect on the ATPase activity of wild-type and G551D NBD1/R/GST fusion proteins or on the ATPase, GTPase, and adenylate kinase activities of purified NBD2. The activation of CFTR by MPB-91 is independent of phosphorylation because 1) kinase inhibitors have no effect and 2) the compound still activated CFTR having 10 mutated protein kinase A sites (10SA-CFTR). The new pharmacological agent MPB-91 may be an important candidate drug to ameliorate the ion transport defect associated with CF and to point out a new pathway to modulate CFTR activity. (+info)
Diffusion of nitric oxide into low density lipoprotein.
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A key early event in the development of atherosclerosis is the oxidation of low density lipoprotein (LDL) via different mechanisms including free radical reactions with both protein and lipid components. Nitric oxide (( small middle dot)NO) is capable of inhibiting LDL oxidation by scavenging radical species involved in oxidative chain propagation reactions. Herein, the diffusion of ( small middle dot)NO into LDL is studied by fluorescence quenching of pyrene derivatives. Selected probes 1-(pyrenyl)methyltrimethylammonium (PMTMA) and 1-(pyrenyl)-methyl-3-(9-octadecenoyloxy)-22,23-bisnor-5-cholenate (PMChO) were chosen so that they could be incorporated at different depths of the LDL particle. Indeed, PMTMA and PMChO were located in the surface and core of LDL, respectively, as indicated by changes in fluorescence spectra, fluorescence quenching studies with water-soluble quenchers and the lifetime values (tau(o)) of the excited probes. The apparent second order rate quenching constants of ( small middle dot)NO (k(NO)) for both probes were 2.6-3.8 x 10(10) m(-1) s(-1) and 1.2 x 10(10) m(-1) s(-1) in solution and native LDL, respectively, indicating that there is no significant barrier to the diffusion of ( small middle dot)NO to the surface and core of LDL. Nitric oxide was also capable of diffusing through oxidized LDL. Considering the preferential partitioning of ( small middle dot)NO in apolar milieu (6-8 for n-octanol:water) and therefore a larger ( small middle dot)NO concentration in LDL with respect to the aqueous phase, a corrected k(NO) value of approximately 0.2 x 10(10) m(-1) s(-1) can be determined, which still is sufficiently large and consistent with a facile diffusion of ( small middle dot)NO through LDL. Applying the Einstein-Smoluchowsky treatment, the apparent diffusion coefficient (D(')NO) of ( small middle dot)NO in native LDL is on average 2 x 10(-5) cm(2) s(-1), six times larger than that previously reported for erythrocyte plasma membrane. Thus, our observations support that ( small middle dot)NO readily traverses the LDL surface accessing the hydrophobic lipid core of the particle and affirm a role for ( small middle dot)NO as a major lipophilic antioxidant in LDL. (+info)
Phenotypes associated with replacement of His by Arg in the Pendred syndrome gene.
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BACKGROUND: Pendred syndrome is often associated with inner ear malformations, especially enlarged vestibular aqueduct (EVA). Recently, mutations in the Pendred syndrome gene (PDS) have been reported in patients with EVA, in addition to those with classical Pendred syndrome. OBJECTIVE: The aim of this study was to investigate the genotype-phenotype correlations of PDS. METHODS: Each of the 21 exons and flanking splice regions of PDS was analysed by direct DNA sequencing in nine patients with EVA; allele-specific amplification was performed to confirm the mutation. Genetic analyses were compared with thyroid function tests, perchlorate discharge tests, thyroid volume and pure-tone audiogram. Magnetic resonance imaging was used to determine the volume of the endolymphatic duct and sac of each patient. RESULTS: A missense mutation, H723R, was identified in the homozygous state in three patients and in the heterozygous state in another three. Although none of the patients had goitre, increased serum thyroglobulin and an abnormal degree of iodide release were correlated with the number of mutant alleles identified. However, there was no relationship between the degree of hearing loss and the number of mutant alleles. CONCLUSION: The present study reveals that the number of mutant alleles correlates with the degree of subclinical thyroid abnormality, but not with the degree of hearing loss in Japanese patients with the PDS missense mutation H723R. (+info)
Hemolysis and iodination of erythrocyte components by a myeloperoxidase-mediated system.
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Erythrocytes are hemolyzed by myeloperoxidase, an H2O2-generating system (glucose + glucose oxidase; hypoxanthine + xanthine oxidase) and an oxidizable cofactor (chloride, iodide, thyroxine, triiodothyronine). The combined effect of chloride and either iodide or the thyroid hormones is greater than additive. Myeloperoxidase can be replaced by lactoperoxidase in the iodide-, thyroxine and triiodothyronine-dependent, but not in the chloride-dependent, systems. Hemolysis is is inhibited by the peroxidase inhibitors, azide and cyanide, and by catalase and is stimulated by superoxide dismutase when the xanthine oxidase system is employed as the source of H2O2. Hemolysis by the iodide-dependent system is associated with the iodination of erythrocyte components. (+info)