Understanding non-enzymatic aminophospholipid glycation and its inhibition. Polar head features affect the kinetics of Schiff base formation

Non-enzymatic aminophospholipid glycation is an especially important process because it alters the stability of lipid bilayers and interferes with cell function and integrity as a result. However, the kinetic mechanism behind this process has scarcely been studied. As in protein glycation, the process has been suggested to involve the formation of a Schiff base as the initial, rate-determining step. In this work, we conducted a comparative kinetic study of Schiff base formation under physiological conditions in three low-molecular weight analogues of polar heads in the naturally occurring aminophospholipids O-phosphorylethanolamine (PEA), O-phospho-DL-serine (PSer) and 2-aminoethylphenethylphosphate (APP) with various glycating carbonyl compounds (glucose, arabinose and acetol) and the lipid glycation inhibitor pyridoxal 5'-phosphate (PLP). Based on the results, the presence of a phosphate group and a carboxyl group in α position respect to the amino group decrease the formation constant for the Schiff base relative to amino acids. On the other hand, esterifying the phosphate group with a non-polar substituent in APP increases the stability of its Schiff base. The observed kinetic formation constants of aminophosphates with carbonyl groups were smaller than those for PLP. Our results constitute an important contribution to understanding the competitive inhibition effect of PLP on aminophospholipid glycation.     

Effectiveness of cognitive behaviour therapy for the treatment of catastrophisation in patients with fibromyalgia: a randomised controlled trial

Introduction  

No randomised, controlled trials have been conducted to date on the efficacy of psychological and pharmacological treatments of pain catastrophising (PC) in patients with fibromyalgia. Our aim in this study was to assess the effectiveness of cognitive-behaviour therapy (CBT) and the recommended pharmacological treatment (RPT) compared with treatment as usual (TAU) at the primary care level for the treatment of PC in fibromyalgia patients.     

Indirect DNA readout by an H-NS related protein: structure of the DNA complex of the C-terminal domain of Ler

Ler, a member of the H-NS protein family, is the master regulator of the LEE pathogenicity island in virulent Escherichia coli strains. Here, we determined the structure of a complex between the DNA-binding domain of Ler (CT-Ler) and a 15-mer DNA duplex. CT-Ler recognizes a preexisting structural pattern in the DNA minor groove formed by two consecutive regions which are narrower and wider, respectively, compared with standard B-DNA. The compressed region, associated with an AT-tract, is sensed by the side chain of Arg90, whose mutation abolishes the capacity of Ler to bind DNA. The expanded groove allows the approach of the loop in which Arg90 is located. This is the first report of an experimental structure of a DNA complex that includes a protein belonging to the H-NS family. The indirect readout mechanism not only explains the capacity of H-NS and other H-NS family members to modulate the expression of a large number of genes but also the origin of the specificity displayed by Ler. Our results point to a general mechanism by which horizontally acquired genes may be specifically recognized by members of the H-NS family.     

MASISH: a database for gene expression in maize seeds

Grass seeds are complex organs composed by multiple tissues and cell types that develop coordinately to produce a viable embryo. The identification of genes involved in seed development is of great interest, but systematic spatial analyses of gene expression on maize seeds at the cell level have not yet been performed. MASISH is an online database holding information for gene expression spatial patterns in maize seeds based on in situ hybridization experiments. The web-based query interface allows the execution of gene queries and provides hybridization images, published references and information of the analyzed genes. AVAILABILITY: http://masish.uab.cat/.     

Radiobiology of the acute radiation syndrome

Acute radiation syndrome or acute radiation sickness is classically subdivided into three subsyndromes: the hematopoietic, gastrointestinal and neurovascular syndrome but many other tissues can be damaged. The time course and severity of clinical signs and symptoms are a function of the overall body volume irradiated, the inhomogeneity of dose exposure, the particle type, the absorbed dose and the dose rate. Classical pathophysiology explain the failure of each of these organs and the timing of appearance of their signs and symptoms due to radiation-induced cytocidal effects of a great number of parenchymal cells of hierarchically organized tissues. Contemporaneously, many other radiation-induced effects has been described and all of them may lead to tissue injury with their corresponding signs and symptoms that can be expressed after short or long period of time. Radiation-induced multi-organ involvement is thought to be due to radiation-induced systemic inflammatory response mediated by released pro-inflammatory cytokines.     

Clinical relevance of tumor cells with stem-like properties in pediatric brain tumors

Background

Primitive brain tumors are the leading cause of cancer-related death in children. Tumor cells with stem-like properties (TSCs), thought to account for tumorigenesis and therapeutic resistance, have been isolated from high-grade gliomas in adults. Whether TSCs are a common component of pediatric brain tumors and are of clinical relevance remains to be determined.

Methodology/Principal Findings

Tumor cells with self-renewal properties were isolated with cell biology techniques from a majority of 55 pediatric brain tumors samples, regardless of their histopathologies and grades of malignancy (57% of embryonal tumors, 57% of low-grade gliomas and neuro-glial tumors, 70% of ependymomas, 91% of high-grade gliomas). Most high-grade glioma-derived oncospheres (10/12) sustained long-term self-renewal akin to neural stem cells (>7 self-renewals), whereas cells with limited renewing abilities akin to neural progenitors dominated in all other tumors. Regardless of tumor entities, the young age group was associated with self-renewal properties akin to neural stem cells (P = 0.05, chi-square test). Survival analysis of the cohort showed an association between isolation of cells with long-term self-renewal abilities and a higher patient mortality rate (P = 0.013, log-rank test). Sampling of low- and high-grade glioma cultures showed that self-renewing cells forming oncospheres shared a molecular profile comprising embryonic and neural stem cell markers. Further characterization performed on subsets of high-grade gliomas and one low-grade glioma culture showed combination of this profile with mesenchymal markers, the radio-chemoresistance of the cells and the formation of aggressive tumors after intracerebral grafting.

Conclusions/Significance

In brain tumors affecting adult patients, TSCs have been isolated only from high-grade gliomas. In contrast, our data show that tumor cells with stem cell-like or progenitor-like properties can be isolated from a wide range of histological sub-types and grades of pediatric brain tumors. They suggest that cellular mechanisms fueling tumor development differ between adult and pediatric brain tumors.     

Site-directed mutagenesis of mouse glutathione transferase P1-1 unlocks masked cooperativity, introduces a novel mechanism for 'ping pong' kinetic behaviour, and provides further structural evidence for participation of a water molecule in proton abstraction from glutathione

Mouse liver glutathione transferase P1-1 has three cysteine residues at positions 14, 47 and 169. We have constructed the single, double and triple cysteine to alanine mutants to define the behaviour of all three thiols. We confirm that C47 is the 'fast' thiol (pK 7.4), and define C169 as the alkaline reactive residue with a pK(a) of 8.6. Only a small proportion of C14 is reactive with 5,5'-dithiobis-(2-nitrobenoic acid) (DTNB) at pH 9 in the C47A/C169A double mutant. The native enzyme and the C169A mutant exhibited Michaelis-Menten kinetics, but all other thiol to alanine mutants exhibited sigmoidal kinetics to varying degrees. The C169A mutant exhibited 'ping pong' kinetics, consistent with a mechanism whereby liberation of a proton from a reduced enzyme-glutathione (GSH) complex to form an enzyme-GS(-) (unprotonated) complex is essentially irreversible. Intriguingly, similar behaviour has recently been reported for a mutant of the yeast prion Ure2p. This cooperative behaviour is 'mirrored' in the crystal structure of the C47A mutant, which binds the p-nitrobenzyl moiety of p-nitrobenzyglutathione in distinct orientations in the two crystallographic subunits. The asymmetry seen in this structure for product binding is associated with absence of a water molecule W0 in the standard wild-type conformation of product binding that is clearly identifiable in the new structure, which may represent a structural model for binding of incoming GSH prior to displacement of W0. Elimination of W0 as a hydroxonium ion may be the mechanism for the initial proton extrusion from the active site.