Proximity-accelerated chemical coupling reaction in the benzodiazepine-binding site of gamma-aminobutyric acid type A receptors: superposition of different allosteric modulators

Benzodiazepines are widely used drugs. They exert sedative/hypnotic, anxiolytic, muscle relaxant, and anticonvulsant effects and act through a specific high affinity binding site on the major inhibitory neurotransmitter receptor, the gamma-aminobutyric acid type A (GABA(A)) receptor. Ligands of the benzodiazepine-binding site are classified into three groups depending on their mode of action: positive and negative allosteric modulators and antagonists. To rationally design ligands of the benzodiazepine site in different isoforms of the GABA(A) receptor, we need to understand the relative positioning and overlap of modulators of different allosteric properties. To solve these questions, we used a proximity-accelerated irreversible chemical coupling reaction. GABA(A) receptor residues thought to reside in the benzodiazepine-binding site were individually mutated to cysteine and combined with a cysteine-reactive benzodiazepine site ligand. Direct apposition of reaction partners is expected to lead to a covalent reaction. We describe here such a reaction of predominantly alpha(1)H101C and also three other mutants (alpha(1)G157C, alpha(1)V202C, and alpha(1)V211C) with an Imid-NCS derivative in which a reactive isothiocyanate group (-NCS) replaces the azide group (-N(3)) in the partial negative allosteric modulator Ro15-4513. Our results show four contact points of imidazobenzodiazepines with the receptor, alpha(1)H101C being shared by classical benzodiazepines. Taken together with previous data, a similar orientation of these ligands within the benzodiazepine-binding pocket may be proposed.

The free radical scavenger alpha-phenyl-tert-butyl nitrone aggravates hippocampal apoptosis and learning deficits in experimental pneumococcal meningitis

The effect of adjuvant therapy with the radical scavenger alpha-phenyl-tert-butyl nitrone (PBN; 100 mg/kg given intraperitoneally every 8 h for 5 days) on brain injury and learning function was evaluated in an infant rat model of pneumococcal meningitis. Meningitis led to cortical necrotic injury (median, 3.97% [range, 0%-38.9%] of the cortex), which was reduced to a median of 0% (range, 0%-30.9%) of the cortex (P<.001) by PBN. However, neuronal apoptosis in the hippocampal dentate gyrus was increased by PBN, compared with that by saline (median score, 1.15 [range, 0.04-1.73] vs. 0.31 [range, 0-0.92]; P<.001). Learning function 3 weeks after cured infection, as assessed by the Morris water maze, was decreased, compared with that in uninfected control animals (P<.001). Parallel to the increase in hippocampal apoptosis, PBN further impaired learning in infected animals, compared with that in saline-treated animals (P<.02). These results contrast with those of an earlier study, in which PBN reduced cortical and hippocampal neuronal injury in group B streptococcal meningitis. Thus, in pneumococcal meningitis, antioxidant therapy with PBN aggravates hippocampal injury and learning deficits.

Good outcome for patients with few lymph node metastases after radical retropubic prostatectomy

BACKGROUND: Conflicting results exist regarding the value of an extended pelvic lymph node dissection (PLND) in node-positive patients undergoing radical retropubic prostatectomy (RRP) for clinically localized prostate cancer. OBJECTIVE: To assess the long-term outcome in node-positive patients who underwent extended PLND followed by RRP. DESIGN, SETTING, AND PARTICIPANTS: A consecutive series of 122 node positive patients with negative preoperative staging examinations, no neoadjuvant hormonal or radiotherapy, and who underwent extended PLND (>/=10 lymph nodes in the surgical specimen) followed by RRP were analyzed. None of the patients received immediate androgen deprivation therapy (ADT). INTERVENTION: All patients underwent extended PLND followed by RRP. MEASUREMENTS: Biochemical recurrence-free survival, cancer-specific, and overall survival were assessed using the Kaplan-Meier technique. RESULTS AND LIMITATIONS: Median prostate-specific antigen (PSA) was 16ng/ml. At pathological examination 76% of the 122 patients had pT3-pT4 tumours, 50% seminal vesicle infiltration. A median of 22 nodes were removed per patient. Median cancer-specific survival at 5 and 10 yr was 84.5% and 60.1%, respectively. In patients with /=3 positive nodes removed, median cancer-specific survival at 10 yr was 78.6% and 33.4%, respectively (p<0.001). After a median period of 33 mo, 61 of the 122 patients (50%) received ADT, particularly those (69%) with >/=3 positive nodes removed. This retrospective study includes a significant percentage of patients with high tumour burden, and therefore may not reflect current patient series. CONCLUSIONS: Patients with /=3 positive nodes, despite extended PLND and despite ADT in 69% of patients.

Radical Verses: Aku Päiviö's Literary Influence on the Finnish Working-Class

Aku Päiviö was one of the most influential voices of the Finnish labor movement in North America—a poet who also wrote plays and novels, and an editor who worked for a variety of newspapers across the United States and Canada. During the height of the Finnish socialist movement from around 1904-1916, Päiviö published a number of poems that identified with the actions and ideologies of the working-class. He also edited for newspapers such as Kansan Lehti and Raivaaja, further extending his literary reach. Despite his prodigious publications and influence, however, little of Päiviö’s writing has been translated into English. This paper celebrates Päiviö’s legacy with some English translations of his poems, specifically those commemorating the 1913-14 Michigan Copper Strike, and illuminates how various thematic and structural relationships in these poems relate to the ideologies and movements of the time.

Efficient coupling of transducin to monomeric rhodopsin in a phospholipid bilayer

G protein-coupled receptors (GPCRs) are seven transmembrane domain proteins that transduce extracellular signals across the plasma membrane and couple to the heterotrimeric family of G proteins. Like most intrinsic membrane proteins, GPCRs are capable of oligomerization, the function of which has only been established for a few different receptor systems. One challenge in understanding the function of oligomers relates to the inability to separate monomeric and oligomeric receptor complexes in membrane environments. Here we report the reconstitution of bovine rhodopsin, a GPCR expressed in the retina, into an apolipoprotein A-I phospholipid particle, derived from high density lipoprotein (HDL). We demonstrate that rhodopsin, when incorporated into these 10 nm reconstituted HDL (rHDL) particles, is monomeric and functional. Rhodopsin.rHDL maintains the appropriate spectral properties with respect to photoactivation and formation of the active form, metarhodopsin II. Additionally, the kinetics of metarhodopsin II decay is similar between rhodopsin in native membranes and rhodopsin in rHDL particles. Photoactivation of monomeric rhodopsin.rHDL also results in the rapid activation of transducin, at a rate that is comparable with that found in native rod outer segments and 20-fold faster than rhodopsin in detergent micelles. These data suggest that monomeric rhodopsin is the minimal functional unit in G protein activation and that oligomerization is not absolutely required for this process.

Hall-effect thruster-cathode coupling : the effect of cathode position and magnetic field topology

Hall-effect thruster (HET) cathodes are responsible for the generation of the free electrons necessary to initiate and sustain the main plasma discharge and to neutralize the ion beam. The position of the cathode relative to the thruster strongly affects the efficiency of thrust generation. However, the mechanisms by which the position affects the efficiency are not well understood. This dissertation explores the effect of cathode position on HET efficiency. Magnetic field topology is shown to play an important role in the coupling between the cathode plasma and the main discharge plasma. The position of the cathode within the magnetic field affects the ion beam and the plasma properties of the near-field plume, which explains the changes in efficiency of the thruster. Several experiments were conducted which explored the changes of efficiency arising from changes in cathode coupling. In each experiment, the thrust, discharge current, and cathode coupling voltage were monitored while changes in the independent variables of cathode position, cathode mass flow and magnetic field topology were made. From the telemetry data, the efficiency of the HET thrust generation was calculated. Furthermore, several ion beam and plasma properties were measured including ion energy distribution, beam current density profile, near-field plasma potential, electron temperature, and electron density. The ion beam data show how the independent variables affected the quality of ion beam and therefore the efficiency of thrust generation. The measurements of near-field plasma properties partially explain how the changes in ion beam quality arise. The results of the experiments show that cathode position, mass flow, and field topology affect several aspects of the HET operation, especially beam divergence and voltage utilization efficiencies. Furthermore, the experiments show that magnetic field topology is important in the cathode coupling process. In particular, the magnetic field separatrix plays a critical role in impeding the coupling between cathode and HET. Suggested changes to HET thruster designs are provided including ways to improve the position of the separatrix to accommodate the cathode.

Coupling of mutated Met variants to DNA repair via Abl and Rad51

Abnormal activation of DNA repair pathways by deregulated signaling of receptor tyrosine kinase systems is a compelling likelihood with significant implications in both cancer biology and treatment. Here, we show that due to a potential substrate switch, mutated variants of the receptor for hepatocyte growth factor Met, but not the wild-type form of the receptor, directly couple to the Abl tyrosine kinase and the Rad51 recombinase, two key signaling elements of homologous recombination-based DNA repair. Treatment of cells that express the mutated receptor variants with the Met inhibitor SU11274 leads, in a mutant-dependent manner, to a reduction of tyrosine phosphorylated levels of Abl and Rad51, impairs radiation-induced nuclear translocation of Rad51, and acts as a radiosensitizer together with the p53 inhibitor pifithrin-alpha by increasing cellular double-strand DNA break levels following exposure to ionizing radiation. Finally, we propose that in order to overcome a mutation-dependent resistance to SU11274, this aberrant molecular axis may alternatively be targeted with the Abl inhibitor, nilotinib.

Spin-strain coupling in a 3-D transition metal oxides

ab-initio Hartree Fock (HF), density functional theory (DFT) and hybrid potentials were employed to compute the optimized lattice parameters and elastic properties of perovskite 3-d transition metal oxides. The optimized lattice parameters and elastic properties are interdependent in these materials. An interaction is observed between the electronic charge, spin and lattice degrees of freedom in 3-d transition metal oxides. The coupling between the electronic charge, spin and lattice structures originates due to localization of d-atomic orbitals. The coupling between the electronic charge, spin and crystalline lattice also contributes in the ferroelectric and ferromagnetic properties in perovskites. The cubic and tetragonal crystalline structures of perovskite transition metal oxides of ABO3 are studied. The electronic structure and the physics of 3-d perovskite materials is complex and less well considered. Moreover, the novelty of the electronic structure and properties of these perovskites transition metal oxides exceeds the challenge offered by their complex crystalline structures. To achieve the objective of understanding the structure and property relationship of these materials the first-principle computational method is employed. CRYSTAL09 code is employed for computing crystalline structure, elastic, ferromagnetic and other electronic properties. Second-order elastic constants (SOEC) and bulk moduli (B) are computed in an automated process by employing ELASTCON (elastic constants) and EOS (equation of state) programs in CRYSTAL09 code. ELASTCON, EOS and other computational algorithms are utilized to determine the elastic properties of tetragonal BaTiO3, rutile TiO2, cubic and tetragonal BaFeO3 and the ferromagentic properties of 3-d transition metal oxides. Multiple methods are employed to crosscheck the consistency of our computational results. Computational results have motivated us to explore the ferromagnetic properties of 3-d transition metal oxides. Billyscript and CRYSTAL09 code are employed to compute the optimized geometry of the cubic and tetragonal crystalline structure of transition metal oxides of Sc to Cu. Cubic crystalline structure is initially chosen to determine the effect of lattice strains on ferromagnetism due to the spin angular momentum of an electron. The 3-d transition metals and their oxides are challenging as the basis functions and potentials are not fully developed to address the complex physics of the transition metals. Moreover, perovskite crystalline structures are extremely challenging with respect to the quality of computations as the latter requires the well established methods. Ferroelectric and ferromagnetic properties of bulk, surfaces and interfaces are explored by employing CRYSTAL09 code. In our computations done on cubic TMOs of Sc-Fe it is observed that there is a coupling between the crystalline structure and FM/AFM spin polarization. Strained crystalline structures of 3-d transition metal oxides are subjected to changes in the electromagnetic and electronic properties. The electronic structure and properties of bulk, composites, surfaces of 3-d transition metal oxides are computed successfully.