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PubMed2002

Exponential velocity tails in a driven inelastic Maxwell model.

The problem of the steady-state velocity distribution in a driven inelastic Maxwell model of shaken granular material is revisited. Numerical solution of the master equation and analytical arguments show that the model has bilateral exponential velocity tails [P(v) approximately e(-|v|/sqrt[D])], where D is the amplitude of the noise. Previous study of this model predicted Gaussian tails [P(v) approximately e(-av(2))].

Journal Article↗
PubMed1999

Gene expression profiling of fibroblasts resistant toward oncogene-mediated transformation reveals preferential transcription of negative growth regulators.

The signal-transducing Ras proteins are important driving forces of diverse cellular processes such as proliferation, neoplastic transformation, differentiation and growth inhibition. As a step toward understanding the complex mechanisms underlying cellular responses, gene expression patterns were examined in two phenotypically normal fibroblast lines which differ in their sensitivity toward oncogene-mediated transformation. Suppression subtractive hybridization (SSH) was used to establish a subtracted cDNA library specific for the REF52 cell line which, like normal diploid fibroblasts, is refractory toward neoplastic transformation induced by mutated HRAS oncogenes. In contrast, rat 208F control cells can be efficiently transformed by HRAS. The nucleotide sequence of 549 subtracted cDNA clones ('REF52 minus 208F') was determined. We identified 93 preferentially expressed gene fragments in resistant REF52 cells as compared to 208F cells. Seventeen of the 52 known genes (32.6%) are capable of inhibiting cell proliferation or of adversely affecting oncogenic signal transduction pathways. These results suggest that the anti-oncogenic properties of resistant REF52 cells are determined by multiple negative growth regulators. The preneoplastic state expressed in 208F cells is characterized by impairment of unexpectedly redundant control mechanisms. Our results also demonstrate that SSH is a powerful method for identifying specific transcriptional patterns in closely related cell types.

Cell Division↗
PubMed2001

Bioavailability and bioequivalence: an FDA regulatory overview.

Bioavailability and/or bioequivalence studies play a key role in the drug development period for both new drug products and their generic equivalents. For both, these studies are also important in the postapproval period in the presence of certain manufacturing changes. Like many regulatory studies, the assessment of bioavailability and bioequivalence can generally be achieved by considering the following three questions. What is the primary question of the study? What are the tests that can be used to address the question? What degree of confidence is needed for the test outcome? This article reviews the regulatory science of bioavailability and bioequivalence and provides FDA's recommendations for drug sponsors who intend to establish bioavailability and/or demonstrate bioequivalence for their pharmaceutical products during the developmental process or after approval.

Biological Availability↗
PubMed1995

Radioimmunoassay for hexarelin, a peptidic growth hormone secretagogue, and its pharmacokinetic studies.

A radioimmunoassay (RIA) method for hexarelin, a peptidic growth hormone secretagogue, has been developed and applied to pharmacokinetic studies in dogs following an IV dose of 1 microgram/kg, and three SC doses of 1, 10, and 100 micrograms/kg. The sensitivity of the assay was determined to be 1.34 fmol/assay. Cross-reactivity of the antiserum with nine hexarelin analogues was less than 1% upon modification of positions 3, 4, or 5 of the peptide. No apparent cross-reaction with endogenous hexarelin metabolites were observed. Intra- and interassay coefficients of variation were less than 3% and 4%, respectively. Intravenous bolus pharmacokinetics of hexarelin displayed a high terminal half-life of 120 min, a fractional plasma clearance of 4.28 ml/min/kg, and a volume of distribution at steady state of 387.7 ml/kg. Following SC administration of hexarelin, despite the increase in dose administered, both clearance (3.93-5.17 ml/min/kg) and volume of distribution (316-544 ml/kg) parameters remained constant over the dose range studied.

Amino Acid Sequence↗
PubMed1999

Community and hospital outcome from ruptured abdominal aortic aneurysm within the catchment area of a regional vascular surgical service.

OBJECTIVE: The objective of this study was to examine patterns of referral, management, and outcome of patients with ruptured abdominal aortic aneurysm (RAAA) within the catchment area of this regional vascular unit (RVU). METHODS: Referral, management, and outcome data regarding 972 consecutive patients admitted to the hospital or certified deceased in the community because of RAAA between January 1, 1989, and December 31, 1995, were retrieved from prospectively gathered computerized national and local databases. RESULTS: Of 381 (39.2%) patients admitted to this unit, 316 (82.9%) underwent surgery, and of those, 188 (59.5%) survived. There was no significant difference in overall mortality between patients who were admitted directly to this unit (152 of 310, 49%) and those who were transferred from elsewhere (41 of 71, 58%). Surgical patients traveled significantly farther to the RVU than nonsurgical patients (P <.001), but there was no significant difference in traveling distance between surgical patients who survived and those who did not. Of 372 (38%) patients who were admitted to other units and not transferred, 24 (6.4%) underwent surgery and 14 (3.8%) survived. Of 972 patients, the overall community mortality from RAAA was 770 (79%). CONCLUSION: Transferring patients from outlying units did not appear to prejudice operative outcome in this RVU. However, less than half of all RAAA patients were transferred, and only a small minority of those not transferred underwent surgery. Although the overall community mortality from RAAA was similar to that reported in earlier studies from other regions and countries where centralization has not occurred, centralization of vascular surgical services may be associated with an inappropriately low operation and survival rate for those patients who are not transferred to the regional center. The effect of centralization on the community outcome of emergent vascular surgical conditions requires further investigation.

Aged↗
PubMed2004

High-dose intravenous proton pump inhibition following endoscopic therapy in the acute management of patients with bleeding peptic ulcers in the USA and Canada: a cost-effectiveness analysis.

BACKGROUND: The efficacy of high-dose intravenous proton pump inhibition has recently been shown, yet its cost-effectiveness remains poorly studied. AIM: To assess the cost-effectiveness of this approach separately for American and Canadian health care settings. METHODS: A validated decision model included patients with bleeding ulcers after successful endoscopic haemostasis. Probabilities were determined from the literature, and charges and lengths of stay from national databases. A third-party payer perspective was adopted over a 30-day time horizon. RESULTS: Re-bleeding rates were 5.9% for patients who received high-dose intravenous proton pump inhibition and 22.9% for those who did not. Hospitalization costs for patients with and without re-bleeding were 11,802 US dollars and 7993 US dollars, and 5220 Canadian dollars and 2696 Canadian dollars, respectively. High-dose intravenous proton pump inhibition was more effective and less costly than the alternative of not administering it. The cost-effectiveness ratios for high-dose and no high-dose intravenous proton pump inhibition were 9112 US dollars and 11,819 US dollars (3293 dollars and 4284 dollars for the Canadian case), respectively. Sensitivity and threshold analyses showed that the results were robust across a wide range of clinically relevant assumptions. CONCLUSION: In the USA and Canada, administering high-dose intravenous proton pump inhibition for 3 days is both more effective and less costly than not doing so for patients with bleeding ulcers after successful endoscopic haemostasis.

Anti-Ulcer Agents↗
PubMed2006

Search for the Higgs boson in H --> WW(*) decays in pp collisions at square root of 1.96 TeV.

We present a search for the standard model Higgs boson in H --> WW(*) decays with e+e-, e+/-mu-/+, and mu+mu- final states in pp collisions at a center-of-mass energy of square root of s = 1.96 TeV. The data, collected from April 2002 to June 2004 with the D0 detector, correspond to an integrated luminosity of 300-325 pb(-1), depending on the final state. The number of events observed is consistent with the expectation from backgrounds. Limits from the combination of all three channels on the Higgs boson production cross section times branching ratio sigma x BR(H --> WW(*) are presented.

Journal Article↗
PubMed2003

Critical assessment of methods of protein structure prediction (CASP)-round V.

This article provides an introduction to the special issue of the journal Proteins dedicated to the fifth CASP experiment to assess the state of the art in protein structure prediction. The article describes the conduct, the categories of prediction, and the evaluation and assessment procedures of the experiment. A brief summary of progress over the five CASP experiments is provided. Related developments in the field are also described.

Computational Biology↗
PubMed2005

Essential roles of sphingosine-1-phosphate and platelet-derived growth factor in the maintenance of human embryonic stem cells.

Human embryonic stem cells (hESCs) have great potential for use in research and regenerative medicine, but very little is known about the factors that maintain these cells in the pluripotent state. We investigated the role of three major mitogenic agents present in serum--sphingosine-1-phosphate (S1P), lysophosphatidic acid (LPA), and platelet-derived growth factor (PDGF)--in maintaining hESCs. We show here that although LPA does not affect hESC growth or differentiation, coincubation of S1P and PDGF in a serum-free culture medium successfully maintains hESCs in an undifferentiated state. Our studies indicate that signaling pathways activated by tyrosine kinase receptors act synergistically with those downstream from lysophospholipid receptors to maintain hESCs in the undifferentiated state. This study is the first demonstration of a role for lysophospholipid receptor signaling in the maintenance of stem cell pluri-potentiality.

Cell Culture Techniques↗
PubMed2006

The component polypeptide chains of bovine insulin nucleate or inhibit aggregation of the parent protein in a conformation-dependent manner.

Amyloid fibrils are typically rigid, unbranched structures with diameters of approximately 10 nm and lengths up to several micrometres, and are associated with more than 20 diseases including Alzheimer's disease and type II diabetes. Insulin is a small, predominantly alpha-helical protein consisting of 51 residues in two disulfide-linked polypeptide chains that readily assembles into amyloid fibrils under conditions of low pH and elevated temperature. We demonstrate here that both the A-chain and the B-chain of insulin are capable of forming amyloid fibrils in isolation under similar conditions, with fibrillar morphologies that differ from those composed of intact insulin. Both the A-chain and B-chain fibrils were found to be able to cross-seed the fibrillization of the parent protein, although these reactions were substantially less efficient than self-seeding with fibrils composed of full-length insulin. In both cases, the cross-seeded fibrils were morphologically distinct from the seeding material, but shared common characteristics with typical insulin fibrils, including a very similar helical repeat. The broader distribution of heights of the cross-seeded fibrils compared to typical insulin fibrils, however, indicates that their underlying protofilament hierarchy may be subtly different. In addition, and remarkably in view of this seeding behavior, the soluble forms of the A-chain and B-chain peptides were found to be capable of inhibiting insulin fibril formation. Studies using mass spectrometry suggest that this behavior might be attributable to complex formation between insulin and the A-chain and B-chain peptides. The finding that the same chemical form of a polypeptide chain in different physical states can either stimulate or inhibit the conversion of a protein into amyloid fibrils sheds new light on the mechanisms underlying fibril formation, fibril strain propagation and amyloid disease initiation and progression.

Amyloid↗
PubMed1995

[Discovery of the pacemaker and heart conduction system in mammals. Fantasy and truth].

The first report about experimental investigations on the origin of the rhythms in the mammalian heart were published in 1888 by McWilliam. Similar experiments were described by Hering in several publications starting from 1900. The results led to the hypothesis of the myogenic genesis of the rhythms. However, in 1907, Hering stated that the origin of the stimulus in the mammal heart was probably a nervous function. In 1898, 1903, and 1906 Wenckebach described the function and the functional localization of the pacemaker in the heart of the intact man, and in 1906, he described the morphology of the pacemaker and its blood supply in the post mortem human heart. In the same year, Adam analyzed the localization of the "natural" pacemaker and the regions of the secondary pacemakers in the mammal heart. In 1907, Keith and Flack introduced the term "sino-auricular node". They quoted the findings of Wenckebach and Hering, and they mentioned McWilliam. A muscular connection between the right atrium and both ventricles of the mammal heart was found by Kent and by His in 1893. In 1907, Tawara called the structure of the origin of the a-v connection "atrio-ventricular node". It was, however, the scientific discovery of Tawara that the a-v pathway was connected with the muscles of the ventricles by the Purkinje fibers. He discovered that the pathway consists of muscles which are histologically divided and functionally different from the other heart muscles. It is a system of specialized muscles which conduct the stimulus for the contraction without contracting themselves.

Animals↗
PubMed2001

The human papillomavirus type 16 E5 protein alters vacuolar H(+)-ATPase function and stability in Saccharomyces cerevisiae.

The human papillomavirus 16 (HPV-16) E5 oncoprotein is a small integral membrane protein that binds to the 16-kDa subunit of the vacuolar H(+)-ATPase (v-ATPase). Conservation within the family of v-ATPases prompted us to look to Saccharomyces cerevisiae as a potential model organism for E5 study. The E5 open reading frame, driven by a galactose-inducible promoter, was integrated into the yeast genome, and the resulting strain demonstrated a nearly complete growth arrest at neutral pH, consistent with defects associated with yeast v-ATPase mutants. Furthermore, this strain demonstrated a severe reduction in pH-dependent and v-ATPase-dependent vacuolar localization of fluorescent markers. Overexpression of the yeast 16-kDa subunit homolog partially suppressed E5-associated growth defects. E5 expression was correlated with a disassociation of the integral (V(o)) and peripheral (V(i)) v-ATPase sub-complexes, as well as a dramatic reduction of the steady-state levels of one mature V(o) subunit and the concomitant accumulation of its major proteolytic fragment, with unchanged levels of two V(i) subunits. Similar analyses of selected E5 mutants in yeast demonstrated a correlation between E5 biology and v-ATPase disruption. Our observations suggest that wild-type HPV-16 E5 acts during the assembly of the v-ATPase to inhibit, either directly or indirectly, V(o) stability and complex formation.

Enzyme Stability↗
PubMed1982

Voltage-clamp analysis of muscarinic excitation in hippocampal neurons.

Pyramidal cells in the CA1 field of guinea pig hippocampal slices were voltage-clamped using a single microelectrode, at 23-30 degrees C. Small inwardly relaxing currents triggered by step hyperpolarizations from holding potentials of -80 to -40 mV were investigated. Inward relaxations occurring for negative steps between -40 mV and -70 mV resembled M-currents of sympathetic ganglion cells: they were abolished by addition of carbachol, muscarine or bethanechol, as well as by 1 mM barium; the relaxations appeared to invert at around -80 mV; they became faster at more negative potentials; and the inversion potential was shifted positively by raising external K+ concentration. Inward relaxations triggered by steps negative to -80 mV, in contrast, appeared to reflect passage of another current species, which has been labelled IQ. Thus IQ did not invert negative to -80 mV, it was insensitive to muscarinic agonists or to barium, and it was blocked by 0.5-3 mM cesium (which does not block IM). Turn-on of IQ causes the well known droop in the hyperpolarizing electrotonic potential in these cells. The combined effects of IQ and IM make the steady-state current-voltage relation of CA1 cells slightly sigmoidal around rest potential. It is suggested that activation of cholinergic septal inputs to the hippocampus facilitates repetitive firing of pyramidal cells by turning off the M-conductance, without much change in the resting potential of the cell.

Animals↗
PubMed1995

Bacterial expression, purification and preliminary kinetic description of the kinase domain of v-fps.

The gene coding for the tyrosine protein kinase domain of v-fps was subcloned into a plasmid vector expressing glutathione-S-transferase (GST). This new vector expresses a fusion protein in Escherichia coli composed of the kinase domain linked with GST at the N-terminus (GST-kin). A portion of the total expressed protein was soluble upon cell lysis and was purified by affinity chromatography using glutathione cross-linked agarose. GST-kin (M(r) 57,000) is a phosphoprotein as judged by 32P autoradiography, consistent with the known autophosphorylation site within the kinase core [Weinmaster et al. (1984) Cell, 37, 559-568]. Cleavage of the fusion protein with thrombin and purification on phosphocellulose resin yielded the pure kinase domain (M(r) 33,000). The activity of the kinase domain is indistinguishable from that of GST-kin using the peptide substrate EEEIYEEIE, indicating that N-terminal fusion has no effect on the kinase domain. GST-kin phosphorylates a second peptide, EAEIYEAIE, with improved catalytic efficiency. Initial velocity data are consistent with a random bireactant mechanism with no substrate synergism observed in the ternary complex. Steady-state kinetic analyses reveal that this peptide is phosphorylated, with a kcat of 3.6 s-1, a Kpeptide of 500 microM and a KATP of 250 microM. The expression, purification and preliminary kinetic analysis of the kinase domain of v-fps provide the first step in the application of structure-function studies for this oncoprotein.

Amino Acid Sequence↗
PubMed1993

[Modified radical mastectomy in local anesthesia].

The authors report their experience on the use of local anesthesia for modified radical mastectomy. They have used this modality in five patients who could not undergo general anesthesia because of underlying severe medical illness and in a sixth one who adamantly refused general anesthesia. They describe in detail the technique used, stressing the importance of a thorough knowledge of regional neuroanatomy. There were no complications in their small series. The authors conclude by stating that the use of local anesthesia may expand the indications for "toilet mastectomy" to include patients in poor medical condition, unsuitable for general anesthesia.

Aged↗