Efficient time-independent wave packet scattering calculations within a Lanczos subspace: H+O-2 (J=0) state-to-state reaction probabilities

An efficient Lanczos subspace method has been devised for calculating state-to-state reaction probabilities. The method recasts the time-independent wave packet Lippmann-Schwinger equation [Kouri , Chem. Phys. Lett. 203, 166 (1993)] inside a tridiagonal (Lanczos) representation in which action of the causal Green's operator is affected easily with a QR algorithm. The method is designed to yield all state-to-state reaction probabilities from a given reactant-channel wave packet using a single Lanczos subspace; the spectral properties of the tridiagonal Hamiltonian allow calculations to be undertaken at arbitrary energies within the spectral range of the initial wave packet. The method is applied to a H+O-2 system (J=0), and the results indicate the approach is accurate and stable. (C) 2002 American Institute of Physics.

Lanczos subspace time-independent wave packet calculations of S(D-1)+H-2 reactive scattering

In this paper. we present the results of quantum dynamical simulations of the S (D-1) + H-2 insertion reaction on a newly developed potential energy surface (J. Chem. Phys. 2001, 114, 320). State-to-state reaction probabilities. product state distributions, and initial-state resolved cumulative reaction probabilities from a given incoming reactant channel are obtained from a time-independent wave packet analysis, performed within a single Lanczos subspace. Integral reaction cross sections are then estimated by J-shifting method and compared with the results from molecular beam experiment and QCT calculations.

Characterization of the retinoid orphan-related receptor-alpha coactivator binding interface: A structural basis for ligand-independent transcription

The retinoid orphan-related receptor-alpha (RORalpha) is a member of the ROR subfamily of orphan receptors and acts as a constitutive activator of transcription in the absence of exogenous ligands. To understand the basis of this activity, we constructed a homology model of Rill using the closely related TRbeta as a template. Molecular modeling suggested that bulky hydrophobic side chains occupy the RORa ligand cavity leaving a small but distinct cavity that may be involved in receptor stabilization. This model was subject to docking simulation with a receptor-interacting peptide from the steroid receptor coactivator, GR-interacting protein-1, which delineated a coactivator binding surface consisting of the signature motif spanning helices 3-5 and helix 12 [activation function 2 (AF2)]. Probing this surface with scanning alanine mutagenesis showed structural and functional equivalence between homologous residues of RORalpha and TRbeta. This was surprising (given that Rill is a ligand-independent activator, whereas TRbeta has an absolute requirement for ligand) and prompted us to use molecular modeling to identify differences between Rill and TRbeta in the way that the All helix interacts with the rest of the receptor. Modeling highlighted a nonconserved amino acid in helix 11 of RORa (Phe491) and a short-length of 3.10 helix at the N terminus of AF2 which we suggest i) ensures that AF2 is locked permanently in the holoconformation described for other liganded receptors and thus 2) enables ligand-independent recruitment of coactivators. Consistent with this, mutation of RORa Phe491 to either methionine or alanine (methionine is the homologous residue in TRbeta), reduced and ablated transcriptional activation and recruitment of coactivators, respectively. Furthermore, we were able to reconstitute transcriptional activity for both a deletion mutant of Ill lacking All and Phe491 Met, by overexpression of a GAL-AF2 fusion protein, demonstrating ligand-independent recruitment of AF2 and a role for Phe491 in recruiting AF2.

Prospective independent validation of APACHE III models in an Australian tertiary adult intensive care unit

Evaluation of the performance of the APACHE III (Acute Physiology and Chronic Health Evaluation) ICU (intensive care unit) and hospital mortality models at the Princess Alexandra Hospital, Brisbane is reported. Prospective collection of demographic, diagnostic, physiological, laboratory, admission and discharge data of 5681 consecutive eligible admissions (1 January 1995 to 1 January 2000) was conducted at the Princess Alexandra Hospital, a metropolitan Australian tertiary referral medical/surgical adult ICU. ROC (receiver operating characteristic) curve areas for the APACHE III ICU mortality and hospital mortality models demonstrated excellent discrimination. Observed ICU mortality (9.1%) was significantly overestimated by the APACHE III model adjusted for hospital characteristics (10.1%), but did not significantly differ from the prediction of the generic APACHE III model (8.6%). In contrast, observed hospital mortality (14.8%) agreed well with the prediction of the APACHE III model adjusted for hospital characteristics (14.6%), but was significantly underestimated by the unadjusted APACHE III model (13.2%). Calibration curves and goodness-of-fit analysis using Hosmer-Lemeshow statistics, demonstrated that calibration was good with the unadjusted APACHE III ICU mortality model, and the APACHE III hospital mortality model adjusted for hospital characteristics. Post hoc analysis revealed a declining annual SMR (standardized mortality rate) during the study period. This trend was present in each of the non-surgical, emergency and elective surgical diagnostic groups, and the change was temporally related to increased specialist staffing levels. This study demonstrates that the APACHE III model performs well on independent assessment in an Australian hospital. Changes observed in annual SMR using such a validated model support an hypothesis of improved survival outcomes 1995-1999.

Identification of T cell epitopes on the 33-kDa fragment of Plasmodium yoelii merozoite surface protein 1 and their antibody-independent protective role in immunity to blood stage malarial

Merozoite surface protein 1 (MSP1) of malaria parasites undergoes proteolytic processing at least twice before invasion into a new RBC. The 42-kDa fragment, a product of primary processing, is cleaved by proteolytic enzymes giving rise to MSP1(33), which is shed from the merozoite surface, and MSP1(19), which is the only fragment carried into a new RBC. In this study, we have identified T cell epitopes on MSP1(33) of Plasmodium yoelii and have examined their function in immunity to blood stage malaria. Peptides 20 aa in length, spanning the length of MSP1(33) and overlapping each other by 10 aa, were analyzed for their ability to induce T cell proliferation in immunized BALB/c and C57BL/6 mice. Multiple epitopes were recognized by these two strains of mice. Effector functions of the dominant epitopes were then investigated. Peptides Cm15 and Cm21 were of particular interest as they were able to induce effector T cells capable of delaying growth of lethal P. yoelii YM following adoptive transfer into immuno-deficient mice without inducing detectable Ab responses. Homologs of these epitopes could be candidates for inclusion in a subunit vaccine.

GPI-anchored proteins are delivered to recycling endosomes via a distinct cdc42-regulated, clathrin-independent pinocytic pathway

Endocytosis of cell-surface proteins via specific pathways is critical for their function. We show that multiple glycosylphosphatidylinositol-anchored proteins (GPI-APs) are endocytosed to the recycling endosomal compartment but not to the Golgi via a nonclathrin, noncaveolae mediated pathway. GPI anchoring is a positive signal for internalization into rab5-independent tubular-vesicular endosomes also responsible for a major fraction of fluid-phase uptake; molecules merely lacking cytoplasmic extensions are not included. Unlike the internalization of detergent-resistant membrane (DRM)-associated interleukin 2 receptor, endocytosis of DRM-associated GPI-APs is unaffected by inhibition of RhoA or dynamin 2 activity. Inhibition of Rho family GTPase cdc42, but not Rac1, reduces fluid-phase uptake and redistributes GPI-APs to the clathrin-mediated pathway. These results describe a distinct constitutive pinocytic pathway, specifically regulated by cdc42.

Independent and conjugate eye movements during optokinesis in teleost fish

In response to movements involving a large part of the visual field, the eyes of vertebrates typically show an optokinetic nystagmus, a response in which both eyes are tightly yoked. Using a comparative approach, this study sets out to establish whether fish with independent spontaneous eye movements show independent optokinetic nystagmus in each eye. Two fish with independent spontaneous eye movements, the pipefish Corythoichthyes intestinalis and the sandlance Limnichthyes fasciatus were compared with the butterflyfish Chaetodon rainfordi, which exhibits tightly yoked eye movements. In the butterflyfish a single whole-field stimulus elicits conjugate optokinesis, whereas the sandlance and pipefish show asynchronous optokinetic movements. In a split drum experiment, when both eyes were stimulated in opposite directions with different speeds, both the sandlance and the pipefish compensated independently with each eye. The optokinetic response in the butterflyfish showed some disconjugacy but was generally confused. When one eye was occluded, the seeing eye was capable of driving the occluded eye in both the butterflyfish and the pipefish but not in the sandlance. Monocular occlusion therefore unmasks a link between the two eyes in the pipefish, which is overridden when both eyes receive visual input. The sandlance never showed any correlation between the eyes during optokinesis in all stimulus conditions. This suggests that there are different levels of linkage between the two eyes in the oculomotor system of teleosts, depending on the visual input.

Mechanisms for the inhibition of amiloride-sensitive Na+ absorption by extracellular nucleotides in mouse trachea

Purinergic stimulation of airway epithelial cells induces Cl- secretion and modulates Na+ absorption by an unknown mechanism. To gain insight into this mechanism, we used a perfused micro-Ussing chamber to assess transepithelial voltage (V-te) and amiloride-sensitive short-circuit current (Isc-Amil) in mouse trachea. Exposure to apical ATP or UTP (each 100 mumol/l) caused a large initial increase in lumen negative V-te and I-sc corresponding to a transient Cl- secretion, while basolateral application of ATP/UTP induced only a small secretory response. Luminal, but not basolateral, application of nucleotides was followed by a sustained and reversible inhibition of Isc-Amil that was independent of extracellular Ca2+ or activation of protein kinase C and was not induced by carbachol (100 mumol/l) or the Ca2+ ionophore ionomycin (1 mumol/l). Removal of extracellular Cl- or exposure to 200 muM DIDS reduced UTP-mediated inhibition of Isc-Amil Substantially. The phospholipase inhibitor U73122 (10 mumol/l) and pertussis toxin (PTX 200 ng/ml) both attenuated UTP-induced Cl- secretion and inhibition of Isc-Amil. Taken together, these data imply a contribution of Cl- conductance and PTX-sensitive G proteins to nucleotide-dependent inhibition of the amiloride-sensitive Na+ current in the mouse trachea.

Inhibition of rat platelet aggregation by the diazeniumdiolate nitric oxide donor MAHMA NONOate

1 Inhibition of rat platelet aggregation by the nitric oxide (NO) donor MAHMA NONOate (Z-1-{N-methyl-N-[6-(N-methylammoniohexyl)amino]}diazen-l-ium-1,2-diolate) was investigated. The aims were to compare its anti-aggregatory effect with vasorelaxation, to determine the effects of the soluble guanylate cyclase inhibitor, ODQ (1H-[1,2,4]oxadiazolo[4,3-ajquinoxalin-1-one), and to investigate the possible role of activation of sarco-encloplasmic reticulum calcium-ATPase (SERCA), independent of soluble guanylate cyclase, using thapsigargin. 2 MAHMA NONOate concentration-dependently inhibited sub-maximal aggregation responses to collagen (2 - 10 mug ml(-1)) and adenosine diphosphate (ADP; 2 mum) in platelet rich plasma. It was (i) more effective at inhibiting aggregation induced by collagen than by ADP, and (ii) less potent at inhibiting platelet aggregation than relaxing rat pulmonary artery. 3 ODQ (10 mum) caused only a small shift (approximately half a log unit) in the concentration-response curve to MAHMA NONOate irrespective of the aggregating agent. 4 The NO-independent activator of soluble guanylate cyclase, YC-1 (3-(5'-hydroxymethyl-2'-furyl)-1-benzy] indazole; 1 - 100 mum), did not inhibit aggregation. The cGMP analogue, 8-pCPT-cGMP (8-(4-chlorophenylthio)guanosine 3'5' cyclic monophosphate; 0.1 - 1 mm), caused minimal inhibition. 5 On collagen-aggregated platelets responses to MAHMA NONOate (ODQ 10 PM present) were abolished by thapsigargin (200 nm). On ADP-aggregated platelets thapsigargin caused partial inhibition. 6 Results with S-nitrosoglutathione (GSNO) resembled those with MAHMA NONOate. Glyceryl trinitrate and sodium nitroprusside were poor inhibitors of aggregation. 7 Thus inhibition of rat platelet aggregation by MAHMA NONOate (like GSNO) is largely ODQ-resistant and, by implication, independent of soluble guanylate cyclase. A likely mechanism of inhibition is activation of SERCA.

Impulse-response function of splanchnic circulation with model-independent constraints: theory and experimental validation

Modeling physiological processes using tracer kinetic methods requires knowledge of the time course of the tracer concentration in blood supplying the organ. For liver studies, however, inaccessibility of the portal vein makes direct measurement of the hepatic dual-input function impossible in humans. We want to develop a method to predict the portal venous time-activity curve from measurements of an arterial time-activity curve. An impulse-response function based on a continuous distribution of washout constants is developed and validated for the gut. Experiments with simultaneous blood sampling in aorta and portal vein were made in 13 anesthetized pigs following inhalation of intravascular [O-15] CO or injections of diffusible 3-O[ C-11] methylglucose (MG). The parameters of the impulse-response function have a physiological interpretation in terms of the distribution of washout constants and are mathematically equivalent to the mean transit time ( T) and standard deviation of transit times. The results include estimates of mean transit times from the aorta to the portal vein in pigs: (T) over bar = 0.35 +/- 0.05 min for CO and 1.7 +/- 0.1 min for MG. The prediction of the portal venous time-activity curve benefits from constraining the regression fits by parameters estimated independently. This is strong evidence for the physiological relevance of the impulse-response function, which includes asymptotically, and thereby justifies kinetically, a useful and simple power law. Similarity between our parameter estimates in pigs and parameter estimates in normal humans suggests that the proposed model can be adapted for use in humans.

Sorting and convergence of primary olfactory axons are independent of the olfactory bulb

Primary olfactory axons expressing the same odorant receptor gene sort out and converge to fixed sites in the olfactory bulb. We examined the guidance of axons expressing the P2 odorant receptor when they were challenged with different cellular environments in vivo. In the mutant extratoes mouse, the olfactory bulb is lacking and is replaced by a fibrocellular mass. In these animals, primary olfactory axons form glomerular-like loci despite the absence of normal postsynaptic targets. P2 axons are able to sort out from other axons in this fibrocellular mass and converge to form loci of like axons. The sites of these loci along mediolateral and ventrodorsal axes were highly variable. Similar convergence was observed for larger subpopulations of axons expressing the same cell surface carbohydrates. The sorting out and convergence of like axons also occurred during regeneration following bulbectomy. Olfactory axon behaviour in these models demonstrates that sorting and convergence of axons are independent of the target, which instead provides distinct topographic cues for guidance. (C) 2003 Wiley-Liss, Inc.

The Wilms' Tumour Suppressor Protein, WT1, Undergoes CRM1-independent Nucleocytoplasmic Shuttling

The Wilms' tumour suppressor gene (WT1) encodes a zinc finger-containing nuclear protein essential for kidney and urogenital development. Initially considered a transcription factor, there is mounting evidence that WT1 has a role in post-transcriptional processing. Using the interspecies heterokaryon assay, we have demonstrated that WT1 can undergo nucleocytoplasmic shuttling. We have also mapped the region responsible for nuclear export to residues 182-324. Our data add further complexity to the role of WT1 in trancriptional and post-transcriptional regulation. (C) 2003 Published by Elsevier B.V. on behalf of the Federation of European Biochemical Societies.

Abnormal left ventricular filling with increasing age reflects abnormal myocardial characteristics independent of ischemia or hypertrophy

Abnormal left ventricular (IV) filling may occur with increasing age despite apparently normal IV size and function, and is usually attributed to IV hypertrophy and coronary artery disease. The purpose of this study was to determine whether myocardial abnormalities could be identified in 67 such patients (36 men, mean age 57 +/- 9 years) whose IV hypertrophy and coronary artery disease were excluded by dobutamine echocardiography. All patients underwent gray scale and color tissue Doppler imaging from 3 apical views, which were stored and analyzed off line. Disturbances in structure and function were assessed by averaging the cyclic variation of integrated backscatter, strain rate, and peak systolic strain from each myocardial segment. Calibrated integrated backscatter (corrected for pericardial backscatter intensity) was measured in the septum and posterior wall from the parasternal long-axis view. Abnormal IV filling was present in 36 subjects (54%). Subjects with and without abnormal IV filling had similar IV mass, but differed in age (p <0.01), cyclic variation (p = 0.001), strain rate (p <0.01), and peak systolic strain (p <0.001). Multivariate logistic regression analysis demonstrated that age (p = 0.016) and cyclic variation (p = 0.042) were the most important determinants of abnormal IV filling in these apparently normal subjects. (C) 2003 by Excerpta Medica, Inc.

Independent contribution of plaque complexity to myocardial ischemia during Dobutamine stress echocardiography

The influence of complex plaque morphology on the extent of demand-induced ischemia in unselected patients is not well defined. We sought to investigate the functional significance of lesion morphology in patients who underwent coronary angiography and dobutamine stress echocardiography (DSE).,Angiography and DSE were performed within a 6-month period (mean 1 +/- 1 month) in 196 patients. Angiographic assessments involved quantification of stenosis severity, assessment of the extent of jeopardized myocardium, and categorization of plaque morphology according to the Ambrose classification. DSE was interpreted by separate investigators with respect to wall motion score index (WMSI) and number of coronary territories involved. A general linear model was constructed to assess,the independent contribution of patient characteristics and angiographic and DSE results with respect to extent of ischemic myocardium. Complex lesion morphology was seen in 62 patients (32%). Patients with complex lesions were more likely to have had prior myocardial infarction (p < 0.001) and be current smokers (p = 0.03). During angiography, they exhibited a trend toward a greater number of diseased vessels, had a greater coronary jeopardy score (p < 0.001) and more frequent collateral flow (p = 0.03). During echocardiography, patients had a higher stress WMSI (p < 0.001) and were more likely to show ischemia in all 3 arterial territories (p < 0.01). On multivariate regression, the coronary artery jeopardy score and the presence of complex plaque morphology were independent predictors of the extent of ischemic myocardium (R 2 = 34%, p < 0.001). Thus, patients with complex plaque morphology are older, more likely to smoke, and more likely to have had prior myocardial. infarction. They exhibit more extensive disease with higher coronary jeopardy scores and a higher resting and peak stress WMSI. Despite these differences, complex plaque morphology remains an independent predictor of the extent of ischemia during stress. (C) 2003 by Excerpta Medica, Inc.

Muscarinic and nicotinic ACh receptor activation differentially mobilize Ca2+ in rat intracardiac ganglion neurons

The origin of intracellular Ca2+ concentration ([Ca2+](i)) transients stimulated by nicotinic ( nAChR) and muscarinic ( mAChR) receptor activation was investigated in fura-2-loaded neonatal rat intracardiac neurons. ACh evoked [Ca2+](i) increases that were reduced to similar to 60% of control in the presence of either atropine ( 1 muM) or mecamylamine ( 3 muM) and to < 20% in the presence of both antagonists. Removal of external Ca2+ reduced ACh-induced responses to 58% of control, which was unchanged in the presence of mecamylamine but reduced to 5% of control by atropine. The nAChR-induced [Ca2+](i) response was reduced to 50% by 10 μM ryanodine, whereas the mAChR-induced response was unaffected by ryanodine, suggesting that Ca2+ release from ryanodine-sensitive Ca2+ stores may only contribute to the nAChR-induced [Ca2+](i) responses. Perforated-patch whole cell recording at - 60 mV shows that the rise in [Ca2+](i) is concomitant with slow outward currents on mAChR activation and with rapid inward currents after nAChR activation. In conclusion, different signaling pathways mediate the rise in [Ca2+](i) and membrane currents evoked by ACh binding to nicotinic and muscarinic receptors in rat intracardiac neurons.