Effect of disulfiram on turnover of 5-hydroxytryptamine in rat brain.

Effect of disulfiram on 5-hydroxytryptamine (5-HT) turnover was studied. Treatment with disulfiram caused increases in 5-HT and 5-hydroxyindoleacetic acid (5-HIAA) in rat brain. Under the same condition, activity of brain mitochondrial aldehyde dehydrogenase was reduced, however, supernatant aldehyde dehydrogenase and monoamine oxidase activities remained unchanged. Disulfiram had no effect on synthesis rate of 5-HT, but decreased metabolism of 5-HT. Moreover, disulfiram impaired transport of 5-HIAA from brain tissue.     

Simultaneous radioimmunoassay of testosterone,dihydrotestosterone, 5α-androstane-3α, 17β-diol and 5α-androstane-3β, 17β-diol in the plasma of adult male rats

A single thin layer chromatography and three antibodies were used for the specific radioimmunoassay of four androgens in pooled rat plasma (Sprague-Dawley adult males). The following values were found (pg/ml ± SD). Testosterone : 3, 138 ± 173; dihydrotestosterone : 374 ± 20; 5α-androstane-3α 17β-diol : 284 ± 24; 5α-androstane-3β, 17β-diol : 223 ± 11.     

Crystal and molecular structure of the sulfhydryl protease calotropin DI at 3·2 Å resolution

The three-dimensional structure of the sulfhydryl protease calotropin DI from the madar plant, Calotropis gigantea, has been determined at 3·2 Å resolution using the multiple isomorphous replacement method with five heavy atom derivatives. A Fourier synthesis based on protein phases with a mean figure of merit of 0·857 was used for model building. The polypeptide backbone of calotropin DI is folded to form two distinct lobes, one of which is comprised mainly of α-helices, while the other is characterized by a system of all antiparallel pleated sheets. The overall molecular architecture closely resembles those found in the sulfhydryl proteases papain and actinidin.Despite the unknown amino acid sequence of calotropin DI a number of residues around its active center could be identified. These amino acid side-chains were found in a similar arrangement as the corresponding ones in papain and actinidin. The polypeptide chain between residues 1 and 18 of calotropin DI folds in a unique manner, providing a possible explanation for the unusual inability of calotropin DI to hydrolyze those synthetic substrates that papain and actinidin act upon.     

Alterations of Inositol Lipid Metabolism of Rat Sciatic Nerve in Streptozotocin-Induced Diabetes

Abstract: The composition and metabolism of rat sciatic nerve phospholipids were studied 20 weeks after induction of chronic diabetes by intraperitoneal injection of streptozotocin (50 mg/kg). On a wet weight basis the nerves from the diabetic animals showed a 7% decrease in total phospholipid from that of controls and a relative decrease in phosphatidylinositol. Incubations of isolated sciatic nerves of diabetic rats in a medium containing [³³P]orthophosphate gave decreased labeling of phosphatidylinositol and substantial changes in the labeling pattern of phosphatidylinositol phosphate and 4,5-bisphosphate from that of controls. The ratio of label in these polyphosphoinositides decreased from 2.5 for normal nerve to about 1.0 for diabetic nerve within a 2-h incubation period. These metabolic alterations were not observed in acutely diabetic animals 5 days after streptozotocin (100 mg/kg) administration. Because polyphosphoinositides may be involved in the control of membrane permeability during axonal conduction, alterations in their relative amounts or turnover rates could be related to the physiological changes of early diabetic neuropathy.     

MA 160 and EB 33 cell lines: HeLa cell contaminants,hybrids or prostatic epithelial cells?

Summary Studies of acid phosphates produced by cell lines MA 160 and EB 33 demonstrated immunochemically their prostatic origin. MA 160 and EB 33, rather than being HeLa contaminants, may be hybrids of prostatic epithelial and HeLa cells or true prostatic cell lines with chromosomal changes common to all long-term cultivated cell lines. This research was supported by NIH (Cancer) Research Grants Nos. 18748 and 16426; and Detroit General Hospital Research Corporation.     

Airway epithelial cells initiate the allergen response through transglutaminase 2 by inducing IL-33 expression and a subsequent Th2 response

Background

Transglutaminase 2 (TG2) is a post-translational protein-modifying enzyme that catalyzes the transamidation reaction, producing crosslinked or polyaminated proteins. Increased TG2 expression and activity have been reported in various inflammatory conditions, such as rheumatoid arthritis, inflammation-associated pulmonary fibrosis, and autoimmune encephalitis. In particular, TG2 from epithelial cells is important during the initial inflammatory response in the lung. In this study, we evaluated the role of TG2 in the pathogenesis of allergic asthma, particularly whether TG2 affects initial activation signaling leading to Th2 differentiation against antigens.

Methods

We induced allergic asthma by ovalbumin sensitization and intranasal challenge in wild-type (WT) BALB/c and TG2-deficient mice. Broncheoalveolar lavage fluid cells and intracellular cytokine production were analyzed by flow cytometry. Interleukin (IL)-33 and TG2 expression in lung epithelial cells was detected by confocal microscopy.

Results

Airway responsiveness was attenuated in TG2-deficient mice compared to that in the WT control. In addition, recruitment of eosinophils and Th2 and Th17 differentiation decreased in TG2-deficient mice. Treatment with cysteamine, a transglutaminase inhibitor, also reduced airway hypersensitivity, inflammatory cell recruitment, and T helper cell differentiation. TG2-deficient mice showed reduced IL-33 expression following induction of allergic asthma compared to those in the WT control.

Conclusions

We found that pulmonary epithelial cells damaged by allergens triggered TG2-mediated IL-33 expression leading to type 2 responses by recruiting both innate and adaptive arms of the immune system.     

Functional and structural roles of the N-terminal extension in Methanosarcina acetivorans protoglobin

Functional and structural properties of protoglobin from Methanosarcina acetivorans, whose Cys(101)E20 residue was mutated to Ser (MaPgb*), and of mutants missing either the first 20 N-terminal amino acids (MaPgb*-ΔN20 mutant), or the first 33 N-terminal amino acids [N-terminal loop of 20 amino acids and a 13-residue Z-helix, preceding the globin fold A-helix; (MaPgb*-ΔN20Z mutant)] have been investigated. In keeping with the MaPgb*-ΔN20 mutant crystal structure, here reported at 2.0 Å resolution, which shows an increased exposure of the haem propionates to the solvent, the analysis of ligand binding kinetics highlights high accessibility of ligands to the haem pocket in ferric MaPgb*-ΔN20. CO binding to ferrous MaPgb*-ΔN20 displays a marked biphasic behavior, with a fast binding process close to that observed in MaPgb* and a slow carbonylation process, characterized by a rate-limiting step. Conversely, removal of the first 33 residues induces a substantial perturbation of the overall MaPgb* structure, with loss of α-helical content and potential partial collapse of the protein chain. As such, ligand binding kinetics are characterized by very slow rates that are independent of ligand concentration, this being indicative of a high energy barrier for ligand access to the haem, possibly due to localized misfolding. This article is part of a Special Issue entitled: Oxygen Binding and Sensing Proteins.     

Multitargeted therapies for multiple myeloma

Multiple myeloma (MM) comprises 1% of all malignancies and 10% of all hematological malignancies. MM is a malignancy of plasma cells in the bone marrow where complex and dynamic interactions with the bone marrow microenvironment lead to tumor progression, skeletal destruction and angiogenesis. Despite the discovery of several novel treatments against MM, including the proteasome inhibitor bortezomib, it is considered to be an incurable disease with an average 4–5 years overall survival.