Delayed Slater determinant update algorithms for high efficiency quantum Monte Carlo.

Within ab initio Quantum Monte Carlo simulations, the leading numerical cost for large systems is the computation of the values of the Slater determinants in the trial wavefunction. Each Monte Carlo step requires finding the determinant of a dense matrix. This is most commonly iteratively evaluated using a rank-1 Sherman-Morrison updating scheme to avoid repeated explicit calculation of the inverse. The overall computational cost is, therefore, formally cubic in the number of electrons or matrix size. To improve the numerical efficiency of this procedure, we propose a novel multiple rank delayed update scheme. This strategy enables probability evaluation with an application of accepted moves to the matrices delayed until after a predetermined number of moves, K. The accepted events are then applied to the matrices en bloc with enhanced arithmetic intensity and computational efficiency via matrix-matrix operations instead of matrix-vector operations. This procedure does not change the underlying Monte Carlo sampling or its statistical efficiency. For calculations on large systems and algorithms such as diffusion Monte Carlo, where the acceptance ratio is high, order of magnitude improvements in the update time can be obtained on both multi-core central processing units and graphical processing units.

Evidence of complementarity between targeted and non-targeted analysis based on liquid and gas-phase chromatography coupled to mass spectrometry for screening halogenated persistent organic pollutants in environmental matrices.

This study explored the complementarity between targeted (TS) and non-targeted screening (NTS) based on liquid and gas-phase chromatography coupled to (high-resolution) mass spectrometry (LC-/GC-(HR)MS) for the comprehensive characterization of organohalogen fingerprints within a set of Lake Ontario lake trout samples. The concentrations of 86 legacy, emerging and novel halogenated compounds (HCs), were determined through 4 TS approaches involving no less than 6 hyphenated systems. In parallel, an innovative NTS strategy, involving both LC and GC-Q-Orbitrap, was implemented to specifically highlight halogenated signals. Non-targeted HRMS data were processed under the HaloSeeker software based on Cl and Br isotopic ratio and mass defect to extend the screening to unsuspected and unknown HCs. A total of 195 halogenated mass spectral features were characterized in the Lake Ontario lake trout, including well known HCs (PCBs, PBDEs, PBBs, DDT and their degradation products), emerging HCs (novel brominated flame retardants, short-, medium- and long-chain chlorinated paraffins) or suggested molecular formula (mainly polychlorinated ones). Among the 122 HCs highlighted by TS, only 21 were identified by NTS. These results fueled a discussion on the potential and limitations of both approaches, and the current position of NTS within environmental and health monitoring programs.

A model of technical variation of microarray signals.

We describe a mathematical model of signal from single-channel direct hybridization microarray platforms. The model establishes a linear relationship between microarray signals and their standard deviations from a minimum set of assumptions. We use the model to precisely define important microarray quality characteristics: resolved fold change and dynamic range. The definitions lead to closed form expressions relating these characteristics to physical parameters of the microarray experiment in the case when both specific and nonspecific binding of target to probe are governed by the Langmuir hybridization isotherm. The predictions of the model are in close agreement to data obtained from spike-in experiments. Given the generality of the model, the introduced definitions of dynamic range and resolved concentration fold-change can be used to conduct cross-platform comparisons and to guide improvement of the microarray platform.

Altered messenger RNA and unique mutational profiles of p53 and Rb in human esophageal carcinomas.

Seventy-nine esophageal carcinoma patients were studied for genetic abnormalities in the p53 and Rb tumor suppressor genes. Single-strand conformation polymorphism analysis and DNA sequencing were used to detect p53 point mutations, Northern blotting was used to examine abnormal expression of p53 and Rb, and polymerase chain reaction and Southern blotting were used to analyze allelic loss. Twenty-five cases were analyzed by DNA sequencing to detect mutations in p53. Fourteen samples contained mutations within exons 5 through 9 of p53; seven had missense mutations giving rise to single amino acid substitutions. The remaining seven (50%) contained nonsense mutations leading to premature termination, five due to single base pair substitutions, and two that were the result of frameshift mutations. In other human tumors, p53 mutations are predominantly missense mutations, but our data as well as those from other groups show that nonsense mutations are common in human esophageal cancer. All but one of the constitutionally heterozygous samples containing mutations also manifested loss of the normal p53 allele; the one exception without allelic loss contained a silent mutation, which should not have had any affect on the p53 protein product. In addition, Northern blotting analysis revealed abnormalities (altered transcript size or mRNA levels) in 5 of 7 cases involving p53 and in 2 of 7 cases analyzed for Rb. Thirty-four cases were informative for allelic loss studies of both p53 and Rb; of these, 25 (74%) lost heterozygosity of p53, Rb, or both. When point mutations and mRNA expression abnormalities were also considered, 33 of 45 (73%) tumors informative for allelic loss assays of both genes as well as for mRNA or point mutation studies showed one or more abnormalities in p53 or Rb. Our results strongly suggest that a unique profile of molecular alterations involving p53 and Rb characterizes human esophageal cancer and that these specific genetic lesions are important in the development and/or progression of most human esophageal carcinomas.

Loss of heterozygosity affecting the p53, Rb, and mcc/apc tumor suppressor gene loci in dysplastic and cancerous ulcerative colitis.

Allelic deletions of tumor suppressor genes have been observed frequently in a variety of human tumors. These losses are believed to contribute to the development of human cancer. Three of the most frequently deleted chromosomal loci contain the tumor suppressor genes p53, retinoblastoma (Rb), and mcc/apc. In order to detect loss of heterozygosity (LOH) within these genes in dysplastic and cancerous ulcerative colitis, we used an application of the polymerase chain reaction. LOH affecting p53 was observed in 8 of 17 (47%) of heterozygous patients, while LOH of Rb and the mcc/apc locus was observed in 9 of 27 (33%) and 13 of 39 (33%) of heterozygotes, respectively. Among 35 patients heterozygous at 2 or more loci, LOH of p53, Rb, and/or mcc/apc was observed in 18 (51%). LOH was more common in left-sided neoplasms. These data suggest that allelic deletion of p53, Rb, mcc, and/or apc is involved in the pathogenesis and/or progression of at least a subset of colonic dysplasias and carcinomas occurring in the setting of ulcerative colitis.

Frequent loss of heterozygosity at the retinoblastoma locus in human esophageal cancers.

Abnormalities in the retinoblastoma tumor suppressor gene (Rb) have been observed in a large number of human cancers. Loss of heterozygosity is a common mode of allelic inactivation of Rb and other tumor suppressor genes. We investigated DNA from 61 primary human esophageal tumors for loss of heterozygosity at the Rb locus using a polymerase chain reaction-based restriction fragment length polymorphism assay. Of informative cases, we found loss of heterozygosity in 14 of 26 (54%) squamous cell carcinomas and 5 of 14 (36%) adenocarcinomas. These data support the hypothesis that Rb inactivation is involved in the pathogenesis and/or progression of esophageal cancer.

Loss of heterozygosity involves multiple tumor suppressor genes in human esophageal cancers.

Loss of heterozygosity occurring on various chromosomes has been described in the majority of human tumors. The targets of frequent or consistent subchromosomal deletions are believed to be tumor suppressor genes. We examined 72 esophageal tumors (46 squamous cell carcinomas and 26 adenocarcinomas) for loss of heterozygosity at the p53, Rb, APC, MCC, and DCC loci. Inclusion of these tumor suppressor genes in the allelic deletions was directly ascertained by performing polymerase chain reaction at polymorphic sites within the genes. Loss of heterozygosity occurred in 55% of informative cases at p53, in 48% of informative cases at Rb, in 66% at APC, in 63% at MCC, and in 24% at DCC. Ninety-three % of tumors informative at all loci (fully informative) lost heterozygosity of at least one locus. A high percentage of fully informative tumors (71%) also lost heterozygosity at more than one locus. There were no significant differences among histological types in the prevalence of loss of heterozygosity at any locus. There were correlations of losses involving MCC versus DCC, Rb, and p53. These data suggest that (a) allelic deletions including these tumor suppressor genes are important in the formation and/or progression of most esophageal cancers; (b) allelic deletions involving MCC may not occur independently of deletions involving other tumor suppressor genes; and (c) the accumulation of multiple allelic deletions involving specific tumor suppressor genes may be important in most esophageal tumorigenesis or tumor evolution.

Direct radioactive labeling of unpurified PCR products using Klenow fragment.

We describe a method for rapid radioactive labeling of PCR product. The method, employing the Klenow fragment of DNA polymerase I, consumes little product, requires no product purification and takes under 30 minutes.

Effect of chronic near-ultraviolet radiation on the gray squirrel lens in vivo.

The effects of ambient exposure to near-ultraviolet (near-UV) radiation (300-400 nm) on the ocular lens of the diurnal squirrel (Sciurus carolinensis) are reported. Gray squirrels lived in cages illuminated for 12 hr a day with near-UV light (6 mW/cm2, 365 nm) for 1 yr. The non-UV-exposed controls were housed separately. In the lenses of UV-exposed animals, anterior pole changes occurred. Central epithelial cells swelled, disappeared, or underwent proliferation. A band of disoriented degenerating fiber cells was seen in the midcortex, with a degree of liquefaction. When lens protein compartments were separated by centrifugation, water-insoluble but urea-soluble fractions were enhanced in the outer and inner cortex and the nucleus. Both high-performance liquid chromatography and polyacrylamide gel electrophoresis revealed that proteins mainly in the midcortex and nucleus were altered considerably. Evidence of a loss of sulfhydryl compounds (by chemical and Raman spectroscopic analyses) and an increase of protein-thiol mixed disulfides (chemically) was also observed. These data prove that repetitive ambient exposure of diurnal animals to near-UV radiation at subsolar levels damages the lens by interfering with the maintenance of epithelial cells and altering the structural proteins; some of this may be due to the conversion of sulfhydryls to mixed disulfides.

Intraocular transplantation of E1A-immortalized retinal precursor cells.

The purpose of this study was to examine the effect of the ocular environment on the survival, tumorigenicity, and phenotypic marker expression of immortalized retinal precursor cells transplanted into immunocompetent adult and neonatal Sprague-Dawley rats. EIA-NR.3, a rat immortalized retinal precursor cell culture, was used as an inexhaustible source of experimental graft material. These cells were prelabeled with the fluorescent marker dil (1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate) and transplanted intravitreally (50,000 cells per microL) into 11 adult and 31 neonatal Sprague-Dawley rat eyes. At 1 mo posttransplant, animals were sacrificed and retinal tissue sections examined histologically for the presence of grafted cells, signs of tumor formation, and retinal phenotypic marker expression. No obvious signs of tumor formation or rejection were seen in a total of 42 eyes in the immunocompetent hosts. Our results indicate that EIA-NR.3 cells survive at least 1 month in vivo, and can migrate from the vitreous into neuroretinal cell layers. Subpopulations of surviving grafted cells were seen to express photoreceptor markers rhodopsin and recoverin comparably between in vitro and in vivo conditions. However, the number of cells immunoreactive for vimentin and E1A decreased significantly under in vivo conditions. This report represents the first experimental intravitreal transplantation of E1A-immortalized retinal precursor cells into adult and neonatal rats. The intraocular location and environment appears to affect phenotypic expression of surviving grafted cells, especially with respect to vimentin and E1A expression. The fact that E1A-NR.3 cells survived intraocularly at least 1 mo without tumor formation suggests that the cells may continue to be useful for further in vivo studies of experimental retinal transplantation, and effects of histological location on retinal cell phenotype and histogenesis in immunocompetent hosts.

Insertion site of the locus of enterocyte effacement in enteropathogenic and enterohemorrhagic Escherichia coli differs in relation to the clonal phylogeny of the strains.

The locus of enterocyte effacement pathogenicity island confers the attaching and effacing histopathology on epithelial cells infected with enteropathogenic and enterohemorrhagic Escherichia coli. We investigated the site of insertion of the locus of enterocyte effacement in E. coli strains in relation to their evolution based on conservation of housekeeping proteins in these strains. The results indicate that the insertion site of the locus of enterocyte effacement varies according to the evolutionary lineage, suggesting that it has inserted at multiple times and sites during the evolution of these pathogens.

Ice incubation step allows adherence of archival histological specimens on gelatin-coated slides for TUNEL staining.

Gelatin-coated slides provide poor tissue adherence during histological procedures which require 37 degreesC incubations, such as terminal deoxynucleotidyl transferase nick-end labelling (TUNEL) analysis. We encountered this difficulty during attempts to analyze archival ocular tissue sections which had been previously sectioned and mounted on gelatin-coated slides. The solution to this problem turned out to be relatively straightforward: Immediately after the 37 degreesC terminal deoxynucleotide transferase step, we incubated the slides on ice for 30 min before continuing with the remainder of the protocol. This ice-incubation step re-solidified the melted gelatin, which allowed for continued adherence of the tissue specimen for further manipulations. This modified TUNEL staining protocol has been used successfully to analyze 14 archival specimens thus far, which would have been nearly impossible to accomplish otherwise. We believe that this ice re-solidification step for gelatin-coated slides has broad applications for procedures which require 37 degreesC incubations, including TUNEL staining, as well as other in situ hybridization and immunohistochemistry protocols.

Structural and functional changes in catalase induced by near-UV radiation.

Part one of this study shows that exposure of purified beef liver catalase in buffered solutions to BL lamps that provide a mixture of 99% UVA and 1% UVB (to be labeled UVA) alters its chemistry and enzymatic activity. Thus, its spectral absorbance lost detail, it aggregated and exhibited a lower isoelectric point and its enzymatic activity was substantially reduced. These photochemically induced changes were increased by irradiation in phosphate buffer or in physiological medium (minimal essential medium) containing riboflavin and tryptophan. Neither alpha-tocopherol nor deferoxamine were protective against these UVA-induced changes in pure catalase. We further investigated the effect of UVA radiation on the activity of catalase in cultured lens epithelial cells and the protective effects of antioxidants. Cultured lens epithelial cells of rabbits and squirrels were exposed to near-UV radiation with representation in the UVA region of 99% and 1% UVB. Catalase assays were done on homogenate supernatants of cells kept dark or UV exposed. In some instances, cells were cultured in medium containing alpha-tocopherol or deferoxamine prior to UV radiation. Comparisons were made between UV-exposed lens cell catalase activity when exposure was done with or without the antioxidants. The UVA radiation was strongly inhibitory to both rabbit and squirrel lens epithelial cell catalase activities. The range of fluxes of near UV radiation was compatible with that which could reach the lens from the sunlit environment. Catalase inactivation was lessened in cells preincubated with alpha-tocopherol and deferoxamine. This suggests that both singlet oxygen and hydroxyl radical formation may be involved in near-UV damage to lens epithelial cell catalase. Such inhibition of catalase by near-UV would enhance H2O2 toxicity and stimulate SH oxidation so as to damage the lens.

Direct radioactive labeling of polymerase chain reaction products.

Radioactively labeled polymerase chain reaction (PCR) products are being used in an increasing number of molecular biology research techniques. Among these are PCR-based polymorphism assays such as linkage analysis (1) and detecting allelic loss in cancer cells (2). Other uses of radioactive PCR include generating probes for Southern and Northern blotting (3) and screening for polymorphisms and point mutations by the single-strand conformation polymorphism (SSCP) technique (4).

Effects of intermittent UVA exposure on cultured lens epithelial cells.

PURPOSE: This paper addresses the question of whether a single non-lethal dose of UVA radiation or the same dose divided three daily one-third doses have like or unlike effects on the growth and the catalase activity of cultured rabbit epithelial (RLE) and immortalized human epithelial (HLE) cells. METHODS: Non-confluent cultured RLE and HLE cells were used to study the effects of UVA radiation on their growth. The effects of three doses of 3 J/cm(2) given one day apart on cell growth (i.e. live cell number) over a three day period were compared with those of a single 9 J/cm(2 ) exposure. Estimation of live cell numbers was done 24 hrs after each exposure by counting trypan blue exclusive cells using a hemocytometer. Confluent cultures of RLE cells were used to study the effects of UVA radiation on their catalase activity. The effects of three doses of 1.25 J/cm(2) each given one day apart on catalase activity (breakdown of H( 2) O( 2) measured spectro-photometrically at 240 nm) were compared with that of a single 3.75 J/cm(2) exposure. RESULTS: A single 9 J/cm(2) dose of UVA reduced the cell growth to only 10% of controls after 24 hrs. By three days after a single 9 J/cm( 2) exposure, the number of live cells was only 70% of controls. Three intermittent exposures of 3 J/cm(2) each for three consecutive days reduced the cell number to 76% of controls. Similar results were obtained for HLE cells. Little if any recovery of cell numbers occurred after three intermittent exposures. A single dose of 3.7 J/cm(2) reduced RLE catalase activity to 20% of controls by one day. By three days after exposure, catalase activity returned to 90% of controls. After three intermittent exposures to 1.25 J/cm(2) each, over three days, RLE catalase activity was reduced to 75% of the controls. There appeared to be no recovery within two additional days. CONCLUSIONS: While single below lethal doses of UVA allow no recovery of RLE or HLE cell growth, partial recovery does occur after several daily intermittent exposures. Recovery of the catalase activity by RLE cells does occur after single sub lethal exposures, but intermittent exposures allow no recovery. The recovery of lens epithelial cell growth and catalase activity from UVA damage varies with the exposure regimen.

Cloning of the RDEC-1 locus of enterocyte effacement (LEE) and functional analysis of the phenotype on HEp-2 cells.

EPEC Escherichia coli are an important cause of epidemic diarrhea in infants. The disease is characterized by attaching and effacing (A/E) lesions where the bacteria attach intimately to the enterocyte surface resulting in localized destruction of microvilli. A 35-kb chromosomal locus termed LEE (locus of enterocyte effacement) in an EPEC strain (E2348/69) has recently been found and is thought to contain all the necessary genes for A/E lesions. RDEC-1 is a strain of E. coli that causes diarrhea in rabbits by a similar mechanism and serves as a model for human EPEC disease. We report 1) the cloning of the RDEC-1 LEE, 2) show that the RDEC-1 LEE is similar in size to the LEE in E2348/69, 3) the RDEC-1 LEE possesses all four regions of the E2348/69 LEE, 4) there are restriction site polymorphisms between the RDEC-1 LEE and that of E2348/69, and 5) the RDEC-1 LEE clone is functionally similar to E2348/69 in its fluorescent actin staining test and suggests that the LEE may be sufficient for the production of A/E lesions by these strains.

Genetics of virulence of enteropathogenic E. coli.

Enteropathogenic E. coli (EPEC) are a major cause of diarrhea in infants throughout the world. Although this pathogen was described 50 years ago, it is only recently that the pathogenic mechanisms employed by this organism have been elucidated. The characteristic histopathology induced by this organism, called "attaching and effacing", consists of intimate adherence of the bacterium to the epithelial cell with marked cytoskeletal changes including effacement of microvilli. A 35 kb region of chromosomal DNA, called the LEE for locus of enterocyte effacement has recently been described which contains all known genes necessary for production of this characteristic histopathology. Within this region is the eae gene encoding intimin, a 94 kDa OMP involved in intimate adherence. Also within this region are genes encoding proteins secreted extracellularly by EPEC (esp) and a type III secretion apparatus (sep) which shares homology with similar systems in Yersinia, Shigella, and Salmonella. Additional genes on a 60 MDa plasmid encode a type IV pilus (BFP) and a positive transcriptional activator (per) of multiple chromosomal and plasmid virulence genes.

Assessing patient willingness to reveal health history information.

A survey was designed to determine how willingly patients reveal personal health information on dental health history forms. After giving informed consent, 107 patients at a university dental hygiene program completed a 10-item survey privately and anonymously. The results suggest that a significant number of patients provide inaccurate or incomplete information to questions routinely asked on the dental health history form.

Effects of sonic toothbrush use on permanent dental luting cements.

The aim of this project was to assess the effects of sonic toothbrushes on commonly used permanent luting cements. While results showed differences between the tensile bond strengths of the three cements, the differences were similar between the two groups: sonic and nonsonic toohbrush exposure. These findings suggest that the sonic toothbrush had no significant effect on the tensile bond strengths of any of the three tested cements.

Effects of toothbrush design and brushing proficiency on plaque removal.

A single-blind, short-term cross-over clinical trial compared the plaque removal performance of three commercially available manual toothbrushes. A sample of 25 dental hygiene students, 19 to 42 years old, served as participants. On 3 separate occasions, participants were instructed to refrain from toothbrushing or flossing for 24 hours before clinical trials. A prebrushing plaque index using disclosing solution was performed on each participant. One of the 3 test brushes was then randomly dispensed to each participant, and they were allowed to brush for 90 seconds without the aid of a mirror. A postbrushing plaque index was then performed on each participant. This procedure was repeated 2 more times at 2-week intervals so that each participant was tested with all 3 toothbrushes. Previous studies of this kind using random participant samples have suggested that brush design does indeed affect the efficacy of plaque removal. This study used participants who were well versed on efficient toothbrushing technique to determine if improved brushing skills would overshadow advantages in toothbrush design. No significant differences in performance were detected between the test brushes for any of the three scored areas.