PTEN Phosphatase-Independent Maintenance of Glandular Morphology in a Predictive Colorectal Cancer Model System

Organotypic models may provide mechanistic insight into colorectal cancer (CRC) morphology. Three-dimensional (3D) colorectal gland formation is regulated by phosphatase and tensin homologue deleted on chromosome 10 (PTEN) coupling of cell division cycle 42 (cdc42) to atypical protein kinase C (aPKC). This study investigated PTEN phosphatase-dependent and phosphatase-independent morphogenic functions in 3D models and assessed translational relevance in human studies. Isogenic PTEN-expressing or PTEN-deficient 3D colorectal cultures were used. In translational studies, apical aPKC activity readout was assessed against apical membrane (AM) orientation and gland morphology in 3D models and human CRC. We found that catalytically active or inactive PTEN constructs containing an intact C2 domain enhanced cdc42 activity, whereas mutants of the C2 domain calcium binding region 3 membrane-binding loop (M-CBR3) were ineffective. The isolated PTEN C2 domain (C2) accumulated in membrane fractions, but C2 M-CBR3 remained in cytosol. Transfection of C2 but not C2 M-CBR3 rescued defective AM orientation and 3D morphogenesis of PTEN-deficient Caco-2 cultures. The signal intensity of apical phospho-aPKC correlated with that of Na/H exchanger regulatory factor-1 (NHERF-1) in the 3D model. Apical NHERF-1 intensity thus provided readout of apical aPKC activity and associated with glandular morphology in the model system and human colon. Low apical NHERF-1 intensity in CRC associated with disruption of glandular architecture, high cancer grade, and metastatic dissemination. We conclude that the membrane-binding function of the catalytically inert PTEN C2 domain influences cdc42/aPKC-dependent AM dynamics and gland formation in a highly relevant 3D CRC morphogenesis model system.

Characterization, crystallization and preliminary X—ray diffraction analysis of acutohaemolysin, a haemolytic toxin from Agkistrodon acutus venom

Acutohaemolysin, a phospholipase A2 (PLA2) from the venom of the snake Agkistrodon acutus, has been isolated and purified to homogeneity by anion-exchange chromatography on a DEAE-Sepharose column followed by cation-exchange chromatography on a CM-Sepharose column. It is an alkaline protein with an isoelectric point of 10.5 and is comprised of a single polypeptide chain of 13 938 Da. Its N-terminal amino-acid sequence shows very high similarity to Lys49-type PLA2 proteins from other snake venoms. Although its PLA2 enzymatic activity is very low, acutohaemolysin has a strong indirect haemolytic activity and anticoagulant activity. Acutohaemolysin crystals with a diffraction limit of 1.60 Å were obtained by the hanging-drop vapour-diffusion method. The crystals belong to the space group C2, with unit-cell parameters a = 45.30, b = 59.55, c = 46.13 Å, [beta] = 117.69°. The asymmetric unit contains one molecule

Thermal decomposition mechanism of levoglucosan during cellulose pyrolysis

Levoglucosan (1,6-anhydro-β-d-glucopyranose) decomposition is an important step during cellulose pyrolysis and for secondary tar reactions. The mechanism of levoglucosan thermal decomposition was studied in this paper using density functional theory methods. The decomposition included direct CO bond breaking, direct CC bond breaking, and dehydration. In total, 9 different pathways, including 16 elementary reactions, were studied, in which levoglucosan serves as a reactant. The properties of the reactants, transition states, intermediates, and products for every elementary reaction were obtained. It was found that 1-pentene-3,4-dione, acetaldehyde, 2,3-dihydroxypropanal, and propanedialdehyde can be formed from the CO bond breaking decomposition reactions. 1,2-Dihydroxyethene and hydroxyacetic acid vinyl ester can be formed from the CC bond breaking decomposition reactions. It was concluded that CO bond breaking is easier than CC bond breaking due to a lower activation energy and a higher released energy. During the 6 levoglucosan dehydration pathways, one water molecule which composed of a hydrogen atom from C3 and a hydroxyl group from C2 is the preferred pathway due to a lower activation energy and higher product stability. © 2012 Elsevier B.V. All rights reserved.

Kinetics study on thermal dissociation of levoglucosan during cellulose pyrolysis

(Chemical Equation Presented) The mechanisms and kinetics studies of the levoglucosan (LG) primary decomposition during cellulose pyrolysis have been carried out theoretically in this paper. Three decomposition mechanisms (C-O bond scission, C-C bond scission, and LG dehydration) including nine pathways and 16 elementary reactions were studied at the B3LYP/6-31 + G(D,P) level based on quantum mechanics. The variational transi-tion- state rate constants for every elementary reaction and every pathway were calculated within 298-1550 K. The first-order Arrhenius expressions for these 16 elementary reactions and nine pathways were suggested. It was concluded that computational method using transition state theory (TST) without tunneling correction gives good description for LG decomposition by comparing with the experimental result. With the temperature range of 667-1327 K, one dehydration pathway, with one water molecule composed of a hydrogen atom from C3 and a hydroxyl group from C2, is a preferred LG decomposition pathway by fitting well with the experimental results. The calculated Arrhenius plot of C-O bond scission mechanism is better agreed with the experimental Arrhenius plot than that of C-C bond scission. This C-O bond scission mechanism starts with breaking of C1-O5 and C6-O1 bonds with formation of CO molecule (C1-O1) simultaneously. C-C bond scission mechanism is the highest energetic barrier pathway for LG decomposition. © 2013 Elsevier Ltd. All rights reserved.

A belief revision framework for revising epistemic states with partial epistemic states

Belief revision performs belief change on an agent’s beliefs when new evidence (either of the form of a propositional formula or of the form of a total pre-order on a set of interpretations) is received. Jeffrey’s rule is commonly used for revising probabilistic epistemic states when new information is probabilistically uncertain. In this paper, we propose a general epistemic revision framework where new evidence is of the form of a partial epistemic state. Our framework extends Jeffrey’s rule with uncertain inputs and covers well-known existing frameworks such as ordinal conditional function (OCF) or possibility theory. We then define a set of postulates that such revision operators shall satisfy and establish representation theorems to characterize those postulates. We show that these postulates reveal common characteristics of various existing revision strategies and are satisfied by OCF conditionalization, Jeffrey’s rule of conditioning and possibility conditionalization. Furthermore, when reducing to the belief revision situation, our postulates can induce Darwiche and Pearl’s postulates C1 and C2.

Regioselective synthesis of pyrroloquinolines - Approaches to martinelline.

Indium trichloride catalysed Diels-Alder reaction of imines derived from anilines with cyclic enamides regioselectively gave the biologically important pyrroloquinoline nucleus, with a cis ring junction, in moderate yield. Although the euo:endo selectivity was in most cases poor, these isomers are readily separated by flash chromatography. The functionality tolerated at both C2 and C7 should allow further elaboration to Martinelline (C) 1999 Published by Elsevier Science Ltd. All rights reserved.

Sputtering of oxygen ice by low energy ions

Naturally occurring ices lie on both interstellar dust grains and on celestial objects, such as those in the outer Solar system. These ices are continuously subjected to irradiation by ions from the solar wind and/or cosmic rays, which modify their surfaces. As a result, new molecular species may form which can be sputtered off into space or planetary atmospheres. We determined the experimental values of sputtering yields for irradiation of oxygen ice at 10 K by singly (He+, C+, N+, O+ and Ar+) and doubly (C2 +, N2 + and O2 +) charged ions with 4 keV kinetic energy. In these laboratory experiments, oxygen ice was deposited and irradiated by ions in an ultra high vacuum chamber at low temperature to simulate the environment of space. The number of molecules removed by sputtering was observed by measurement of the ice thickness using laser interferometry. Preliminary mass spectra were taken of sputtered species and of molecules formed in the ice by temperature programmed desorption (TPD). We find that the experimental sputtering yields increase approximately linearly with the projectile ion mass (or momentum squared) for all ions studied. No difference was found between the sputtering yields for singly and doubly charged ions of the same atom within the experimental uncertainty, as expected for a process dominated by momentum transfer. The experimental sputter yields are in good agreement with values calculated using a theoretical model except in the case of oxygen ions. Preliminary studies have shown molecular oxygen as the dominant species sputtered and TPD measurements indicate ozone formation.

Comet C/2012 S1 (ISON): Observations of the Dust Grains from SOFIA and of the Atomic Gas from NSO Dunn and McMath-Pierce Solar Telescopes (Invited)

Comet C/2012 S1 (ISON) is unique in that it is a dynamically new comet derived from the Oort cloud reservoir of comets with a sun-grazing orbit. Infrared (IR) and visible wavelength observing campaigns were planned on NASA's Stratospheric Observatory For Infrared Astronomy (SOFIA) and on National Solar Observatory Dunn (DST) and McMath-Pierce Solar Telescopes, respectively. We highlight our early results. SOFIA (+FORCAST [1]) mid- to far-IR images and spectroscopy (~5-35 μm) of the dust in the coma of ISON are to be obtained by the ISON-SOFIA Team during a flight window 2013 Oct 21-23 UT (r_h≈1.18 AU). Dust characteristics, identified through the 10 μm silicate emission feature and its strength [2], as well as spectral features from cometary crystalline silicates (Forsterite) at 11.05-11.2 μm, and near 16, 19, 23.5, 27.5, and 33 μm are compared with other Oort cloud comets that span the range of small and/or highly porous grains (e.g., C/1995 O1 (Hale-Bopp) [3,4,5] and C/2001 Q4 (NEAT) [6]) to large and/or compact grains (e.g., C/2007 N4 (Lulin) [7] and C/2006 P1 (McNaught) [8]). Measurement of the crystalline peaks in contrast to the broad 10 and 20 μm amorphous silicate features yields the cometary silicate crystalline mass fraction [9], which is a benchmark for radial transport in our protoplanetary disk [10]. The central wavelength positions, relative intensities, and feature asymmetries for the crystalline peaks may constrain the shapes of the crystals [11]. Only SOFIA can look for cometary organics in the 5-8 μm region. Spatially resolved measurements of atoms and simple molecules from when comet ISON is near the Sun (r_h<0.4 AU, near Nov-20--Dec-03 UT) were proposed for by the ISON-DST Team. Comet ISON is the first comet since comet Ikeya-Seki (1965f) [12,13] suitable for studying the alkalai metals Na and K and the atoms specifically attributed to dust grains including Mg, Si, Fe, as well as Ca. DST's Horizontal Grating Spectrometer (HGS) measures 4 settings: Na I, K, C2 to sample cometary organics (along with Mg I), and [O I] as a proxy for activity from water [14] (along with Si I and Fe I). State-of-the-art instruments that will also be employed include IBIS [15], which is a Fabry-Perot spectral imaging system that concurrently measures lines of Na, K, Ca II, or Fe, and ROSA (CSUN/QUB) [16], which is a rapid imager that simultaneously monitors Ca II or CN. From McMath-Pierce, the Solar-Stellar Spectrograph also will target ISON (320-900 nm, R~21,000, r_h

THE SURFACE-STRUCTURE OF A C(2 X-2) POTASSIUM OVERLAYER ON CO(1010)

The surface structure of the clean Co{1010BAR} surface and a c(2 x 2) potassium overlayer have been determined by quantitative low energy electron diffraction. The Co{1010BAR} sample has been shown to be laterally unreconstructed with the surface being uniquely terminated by an outermost closely packed double layer (dz12 = 0.68 angstrom). A damped oscillatory relaxation of the outermost three atomic layers occurs, with relaxations DELTA-dz12 = -6.5 +/- 2% and DELTA-dz23 = +1.0 +/- 2%.

The c(2 x 2) overlayer formed at a coverage of 0.5 ML was subjected to a full I-V analysis. A range of adsorption sites were tested including fourfold hollow, on-top, and both long and short bridge sites in combination with both "long" and "short" cobalt interlayer terminations. A clear preference was found for adsorption in the maximal coordination fourfold hollow site. No switching of surface termination occurs. The potassium adatoms reside in the [1210BAR] surface channels directly above second layer cobalt atoms with a potassium to outermost cobalt interlayer separation of 2.44 +/- 0.05 angstrom. Potassium-cobalt bond lengths of 3.40 +/- 0.05 and 3.12 +/- 0.05 angstrom between the four (one) outermost (second) layer nearest-neighbour substrate atoms suggests a potassium effective radius of 1.87 +/- 0.05 angstrom, somewhat smaller than the Pauling covalent radius and considerably larger than the ionic radius (1.38 angstrom). The alkali-surface bonding is thus predominantly "covalent"/"metallic".

Development and Validation of a Novel Lateral Flow Immunoassay (LFIA) for the Rapid Screening of Paralytic Shellfish Toxins (PSTs) from Shellfish Extracts

A single-step lateral flow immunoassay (LFIA) was developed and validated for the rapid screening of paralytic shellfish toxins (PSTs) from a variety of shellfish species, at concentrations relevant to regulatory limits of 800 μg STX-diHCl equivalents/kg shellfish meat. A simple aqueous extraction protocol was performed within several minutes from sample homogenate. The qualitative result was generated after a 5 min run time using a portable reader which removed subjectivity from data interpretation. The test was designed to generate noncompliant results with samples containing approximately 800 μg of STX-diHCl/kg. The cross-reactivities in relation to STX, expressed as mean ± SD, were as follows: NEO: 128.9% ± 29%; GTX1&4: 5.7% ± 1.5%; GTX2&3: 23.4% ± 10.4%; dcSTX: 55.6% ± 10.9%; dcNEO: 28.0% ± 8.9%; dcGTX2&3: 8.3% ± 2.7%; C1&C2: 3.1% ± 1.2%; GTX5: 23.3% ± 14.4% (n = 5 LFIA lots). There were no indications of matrix effects from the different samples evaluated (mussels, scallops, oysters, clams, cockles) nor interference from other shellfish toxins (domoic acid, okadaic acid group). Naturally contaminated sample evaluations showed no false negative results were generated from a variety of different samples and profiles (n = 23), in comparison to reference methods (MBA method 959.08, LC-FD method 2005.06). External laboratory evaluations of naturally contaminated samples (n = 39) indicated good correlation with reference methods (MBA, LC-FD). This is the first LFIA which has been shown, through rigorous validation, to have the ability to detect most major PSTs in a reliable manner and will be a huge benefit to both industry and regulators, who need to perform rapid and reliable testing to ensure shellfish are safe to eat.

Age-Related Macular Degeneration-Associated Genes in Alzheimer Disease

Objectives: Given the clinical and pathological similarities between age-relatedmacular degeneration (AMD) and Alzheimer disease (AD), to assess whether AMDassociatedsingle nucleotide polymorphisms (SNPs), including those from complementrelatedgenes, are associated with AD. 
Design: A case-control association study-typedesign. 
Setting: A UK tertiary care dementia clinic. 
Participants: 322 cognitivelynormal participants and 258 cases with a clinical diagnosis of AD.
Measurements:Polymorphisms in the following genes were studied: CFH, ARMS2, C2/CFB, C3, CFI/PLA2G12a, SERPING1, TLR3, TLR4, CRP, APOE, and TOMM40. Haplotypes were analysedfor CFH, TOMM40, and APOE. Univariate analysis was performed for each geneticchange and case-comparator status, and then correction for multiple testing performed. 
Results: The presence of an ε4 APOE allele was significantly associated with AD. Noassociation was evident between CFH SNPs or haplotypes, or other AMD-associated SNPstested, and AD. The exceptions were TOMM40 SNPs, which were associated with AD evenafter correction for multiple comparisons. The associations disappeared, however, whenentered into a regression model including APOE genotypes. 
Conclusions: The resultsfor most SNPs tested, as well as CFH haplotypes, are novel. The functional effects ofabnormal complement activity in AD’s pathogenesis may be contradictory, butmethodological reasons may underlie the lack of association—for example, geneticchanges other than SNPs being involved.

Development of TMTP-1 targeted designer biopolymers for gene delivery to prostate cancer

Designer biopolymers (DBPs) represent state of the art genetically engineered biomacromolecules designed to condense plasmid DNA, and overcome intra- and extra- cellular barriers to gene delivery. Three DBPs were synthesized, each with the tumor molecular targeting peptide-1 (TMTP-1) motif to specifically target metastases. Each DBP was complexed with a pEGFP-N1 reporter plasmid to permit physiochemical and biological assay analysis. Results indicated that two of the biopolymers (RMHT and RM3GT) effectively condensed pEGFP-N1 into cationic nanoparticles< 100nm and were capable of transfecting PC-3 metastatic prostate cancer cells. Conversely the anionic RMGT DBP nanoparticles could not transfect PC-3 cells. RMHT and RM3GT nanoparticles were stable in the presence of serum and protected the cargo from degradation. Additionally it was concluded that cell viability could recover post-transfection with these DBPs, which were less toxic than the commercially available transfection reagent Lipofectamine® 2000. With both DBPs, a higher transfection efficacy was observed in PC-3 cells than in the moderately metastatic, DU145, and normal, PNT2-C2, cell lines. Blocking of the TMTP-1 receptors inhibited gene transfer indicating internalization via this receptor. In conclusion RMHT and RM3GT are fully functional DBPs that address major obstacles to gene delivery and target metastatic cells expressing the TMTP-1 receptor.

An R -matrix with pseudo-states calculation of electron-impact excitation in C 2 +

We have performed an R-matrix with pseudo-states (RMPS) calculation of electron-impact excitation in C2+.Collision strengths and effective collision strengths were determined for excitation between the lowest 24 terms, including all those arising from the 2s3l and 2s4l configurations. In the RMPS calculation, 238 terms (90 spectroscopic and 148 pseudo-state) were employed in the close-coupling (CC) expansion of the target. In order to investigate the significance of coupling to the target continuum and highly excited bound states, we compare the RMPS results with those from an R-matrix calculation that incorporated all 238 terms in the configuration- interaction expansion, but only the lowest 44 spectroscopic terms in the CC expansion. We also compare our effective collision strengths with those from an earlier 12-state R-matrix calculation (Berrington et al 1989 J. Phys. B: At.Mol. Opt. Phys. 22 665). The RMPS calculation was extremely large, involving (N +1)-electron Hamiltonian matrices of dimension up to 36 085, and required the use of our recently completed suite of parallel R-matrix programs. The full set of effective collision strengths fromourRMPS calculation is available at theOakRidgeNationalLaboratoryControlledFusion Atomic Data Center web site. 1.

Electron-impact ionization of C+ in both ground and metastable states

Electron-impact ionization cross sections are calculated for the ground and metastable states of C+. Com- parisons between perturbative distorted-wave and nonperturbative time-dependent close-coupling calculations find reductions in the peak direct ionization cross sections due to electron coupling effects of approximately 5% for ground state C+ and approximately 15% for metastable state C+. Fairly small excitation-autoionization contributions are found for ground state C+, while larger excitation-autoionization contributions are found for metastable state C+. Comparisons between perturbative distorted-wave and nonperturbative R-matrix with pseudostates calculations find reductions in the peak total ionization cross sections due to electron coupling effects of approximately 15–20 % for ground state C+ and approximately 25–35 % for metastable state C+. Finally, comparisons between theory and experiment find that present and previous C+ crossed-beam measure- ments are in excellent agreement with ground state nonperturbative R-matrix with pseudostates calculations for total ionization cross sections. Combined with previous non-perturbative calculations for C, C2+, and C3+, accurate ionization cross sections and rate coefficients are now available for the ground and metastable states of all carbon ion stages.