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PubMed1997

Virulence plasmid instability in Shigella flexneri 2a is induced by virulence gene expression.

Expression of the predominantly plasmid-encoded virulence regulon of Shigella flexneri 2a is induced by growth at 37 degrees C and repressed by growth at 30 degrees C. During growth at 37 degrees C, spontaneous S. flexneri mutants arise which have undergone virulence plasmid curing or rearrangement and no longer display the virulent phenotype. In the laboratory, the unstable nature of the virulence plasmid causes complete loss of virulence in a growing population. We have undertaken an analysis of virulence plasmid instability, classifying events which produced individual avirulent derivatives within a virulent population and identifying the factor(s) which controlled conversion. Multiplex PCR analysis of DNA obtained from spontaneous avirulent derivatives indicated that virF and virB were deleted or otherwise inactivated in over 97% of the isolates. The virF and virB loci encode regulatory proteins required for transcriptional activation of the virulence regulon. Inactivation of these key regulatory loci in the vast majority of avirulent derivatives which arose during growth at 37 degrees C suggested that virulence gene expression induced virulence plasmid instability. Consistent with this hypothesis, we observed stable virulence plasmid maintenance during growth of a wild-type strain at 30 degrees C where virulence gene expression was repressed. The virulence plasmid was also stably maintained in virF and virB mutants grown at 37 degrees C. Conversely, virulence plasmid destabilization was induced at 30 degrees C and accelerated at 37 degrees C through expression of VirF or VirB from multicopy plasmids. These results indicate that exposure of S. flexneri to conditions favoring induction of the virulent phenotype also favor its loss. The significance of this paradox of Shigella pathogenicity is discussed.

Bacterial Proteins↗
PubMed2003

Pre-steady-state analysis of the nucleoside hydrolase of Trypanosoma vivax. Evidence for half-of-the-sites reactivity and rate-limiting product release.

The nucleoside hydrolase (NH) of the Trypanosoma vivax parasite catalyzes the hydrolysis of the N-glycosidic bond in ribonucleosides according to the following reaction: beta-purine (or pyrimidine) nucleoside + H(2)O --> purine (pyrimidine) base + ribose. The reaction follows a highly dissociative nucleophilic displacement reaction mechanism with a ribosyl oxocarbenium-like transition state. This paper describes the first pre-steady-state analysis of the conversion of a number of purine nucleosides. The NH exhibits burst kinetics and behaves with half-of-the-sites reactivity. The analysis suggests that the NH of T. vivax follows a complex multistep mechanism in which a common slow step different from the chemical hydrolysis is rate limiting. Stopped-flow fluorescence binding experiments with ribose indicate that a tightly bound enzyme-ribose complex accumulates during the enzymatic hydrolysis of the common purine nucleosides. This is caused by a slow isomerization between a tight and a loose enzyme-ribose complex forming the rate-limiting step on the reaction coordinate.

Animals↗
PubMed1993

Adsorption and conversion of prothrombin on a rotating disc.

In most flow systems, the rate of protein transfer from bulk solution to a macroscopic surface is site-dependent. In studies on surface-mediated protein conversion, this hampers the comparison of a proposed expression for the conversion process, such as the Michaelis-Menten equation, which actually measured overall conversion rates. However, the rotating disc is a classical example of a uniformly accessible surface and therefore was used for a quantitative analysis of prothrombin conversion by the phospholipid-bound factor Xa/factor Va complex (prothrombinase complex). A simple design of a rotating disc, adapted for ellipsometric measurement of protein adsorption, is presented. Agreement between experiment and theory was obtained for the influence of rotation velocity on the initial, transport-limited, adsorption rates of lysozyme, prothrombin, and fibrinogen. After coverage of the disc with a 20% phosphatidylserine/80% phosphatidylcholine bilayer and preadsorption of factor Va, addition of excess factor Xa and prothrombin resulted in effective conversion of prothrombin. For high (10 fmol.cm-2) surface coverage of prothrombinase, the rate of conversion equals the transport limited adsorption rate of prothrombin. For low (0.1 to 0.5 fmol.cm-2) surface concentrations of prothrombinase, the conversion rate dropped below the transport limit and the intrinsic kinetic parameters could be estimated at Km = 7.1 +/- 1.2 nM and kcat = 25 +/- 1.0 s-1 (20 degrees C). At these low surface activities of prothrombinase, the effect of the rotation rate (6 to 225 rad.s-1) on prothrombin conversion could be explained by the rotation-rate dependent prothrombin transport. This indicates that the fluid shear rate has no drastic influence on the intrinsic kinetics of prothrombin conversion.

Adsorption↗
PubMed1999

Gene duplication and gene conversion in the Caenorhabditis elegans genome.

A comprehensive analysis of duplication and gene conversion for 7394 Caenorhabditis elegans genes (about half the expected total for the genome) is presented. Of the genes examined, 40% are involved in duplicated gene pairs. Intrachromosomal or cis gene duplications occur approximately two times more often than expected. In general the closer the members of duplicated gene pairs are, the more likely it is that gene orientation is conserved. Gene conversion events are detectable between only 2% of the duplicated pairs. Even given the excesses of cis duplications, there is an excess of gene conversion events between cis duplicated pairs on every chromosome except the X chromosome. The relative rates of cis and trans gene conversion and the negative correlation between conversion frequency and DNA sequence divergence for unconverted regions of converted pairs are consistent with previous experimental studies in yeast. Three recent, regional duplications, each spanning three genes are described. All three have already undergone substantial deletions spanning hundreds of base pairs. The relative rates of duplication and deletion may contribute to the compactness of the C. elegans genome.

Animals↗
PubMed1983

Epithelial differentiation in the fetal rat colon. I. Plasma membrane phosphatase activities.

During the last week of gestation of the fetal rat, the epithelium of the colon is rapidly remodeled. At 16 days a primitive stratified epithelium surrounds a small central lumen. Over the next 3 days, the main lumen extends narrow clefts down to the basal cell layer and small secondary lumina appear within the stratified epithelium between these clefts. At 19 and 20 days, secondary lumina enlarge but remain discrete; an infusion of cationic ferritin into the main lumen does not enter secondary lumina. During the 2 days prior to birth (21-22), the secondary lumina join the main lumen as superficial cells are sloughed, and the epithelium becomes simple columnar. Freeze-fracture replicas indicate that luminal and nonluminal membrane domains of epithelial cell plasma membranes are separated by continuous tight junctions throughout the conversion process. Cytochemical analysis of tissue slices from 16- to 22-day fetal colon demonstrated the appearance and segregation of two phosphatases on apical and basolateral membrane domains during epithelial conversion. Cysteine-sensitive pH 9.0 (alkaline) phosphatase activity was first detected along the luminal membranes of cells bordering both primary and secondary lumina at 18 days gestation and increased to a maximum at 20-21 days; weaker activity was present on basolateral membranes. Phosphatase activity at pH 8.0 also appeared at 18 days and increased thereafter, but was localized primarily on nonluminal membranes. At pH 8.0, reaction product appeared on both inner and outer sides of the membrane, and was only partially abolished by omission of K+ or addition of ouabain; thus the reaction may be only partially due to K+-dependent ATPase activity. Biochemical analysis of the cytochemical media confirmed the appearance of phosphatase activities at 18 days. Thus, plasma membrane phosphatase activities appear while the epithelium is still stratified, but are segregated to luminal and nonluminal membrane domains at the onset of activity. Segregation is maintained throughout the process of conversion of a simple columnar epithelium.

Animals↗
PubMed2003

Inhibition of UV-induced G1 arrest by exposure to 50 Hz magnetic fields in repair-proficient and -deficient yeast strains.

PURPOSE: To assess the possibility that extremely low frequency (ELF) magnetic fields obstruct the damage repair process, the gene conversion frequency and cell cycle kinetics in a DNA repair-proficient and nucleotide excision repair (NER)-deficient strain of diploid yeast Saccharomyces cerevisiae. MATERIALS AND METHODS: DNA repair- or NER-deficient cells were irradiated with sublethal doses of ultraviolet light (UV) radiation followed by exposure to 50 Hz magnetic fields up to 30 mT for 48 h. After exposure, colony-forming ability was scored as revertants in which gene conversion had restored the functional allele of the ARG4 gene conversion hotspot. Cell cycle analysis was performed using flow cytometry. RESULTS: Gene conversion rate was increased by the combined exposure in DNA repair-proficient cells, whereas it remained unchanged between UV alone and the combined exposure in NER-deficient cells. The UV-induced G1 arrest was inhibited by exposure to 30 mT ELF magnetic fields in both repair-proficient and -deficient cells. CONCLUSIONS: The results suggest that exposure to high-density (30 mT) ELF magnetic fields decreases the efficiency of NER by suppressing G1 arrest, which in turn led to enhancement of the UV-induced gene conversion.

Cell Cycle↗
PubMed2006

Differential regulation of short- and long-tract gene conversion between sister chromatids by Rad51C.

The Rad51 paralog Rad51C has been implicated in the control of homologous recombination. To study the role of Rad51C in vivo in mammalian cells, we analyzed short-tract and long-tract gene conversion between sister chromatids in hamster Rad51C(-/-) CL-V4B cells in response to a site-specific chromosomal double-strand break. Gene conversion was inefficient in these cells and was specifically restored by expression of wild-type Rad51C. Surprisingly, gene conversions in CL-V4B cells were biased in favor of long-tract gene conversion, in comparison to controls expressing wild-type Rad51C. These long-tract events were not associated with crossing over between sister chromatids. Analysis of gene conversion tract lengths in CL-V4B cells lacking Rad51C revealed a bimodal frequency distribution, with almost all gene conversions being either less than 1 kb or greater than 3.2 kb in length. These results indicate that Rad51C plays a pivotal role in determining the "choice" between short- and long-tract gene conversion and in suppressing gene amplifications associated with sister chromatid recombination.

Animals↗
PubMedDate not supplied

Conversion of HMOs and hospitals: what's at stake?

Because for-profit conversions of nonprofit organizations are regulated under trust law at the state level, their health policy implications have generally not been part of the process. This paper provides a health policy framework for assessing conversions of hospitals and health maintenance organizations (HMOs). It begins with basic differences in ownership forms and identifies considerations on both sides of the conversion question. The analysis turns on the extent of the social benefits of nonprofits: the regulatory tool provided by tax exemptions, trustworthiness in the presence of informational asymmetries, and community benefit activities. The analysis and the evidence suggest that the nonprofit form continues to hold significant advantages in health care that bear consideration by policymakers faced with conversion proposals.

Health Facility Merger↗
PubMed1991

The protocols and codes of practice used for the determination of absorbed dose in megavoltage photon and electron beams.

Air kerma-to-dose and exposure-to-dose conversion factors for megavoltage photon beams and electron beams are derived for a variety of commonly used ionization chambers, using the various dosimetry protocols and codes of practice currently in use. It is found that differences in the resultant values are present for both photon and electron beams. For example, for a NE2571 graphite-walled thimble ionization chamber, the maximum variation in the air kerma-to-dose conversion factor for photons, Cw,lambda, occurs at a "nominal" energy of 25 MV, when the difference between the maximum and minimum value of Cw,lambda is 5.4%, whereas the maximum difference between different protocols in the air kerma-to-dose conversion factor for electrons, Cw,e, occurs at Eo = 9 MeV, when the spread between maximum and minimum Cw,e values is 2.7%. An analysis of the conversion factors is undertaken to examine if the numerical or theoretical data are the cause of the discrepancies, and it is found to be a combination of both, although the major contribution to this difference arises from the use of different sets of stopping power ratios in the different protocols.

Calibration↗
PubMed2004

Ten-year experience with laparoscopy on a gynecologic oncology service: analysis of risk factors for complications and conversion to laparotomy.

OBJECTIVE: The purpose of this study was to analyze our initial 10-year experience with laparoscopy and to determine risk factors for complications and conversions to laparotomy for technical difficulty. STUDY DESIGN: We reviewed the charts of all laparoscopic procedures from January 1991 through December 2000 and divided the procedures into 4 levels on the basis of the degree of difficulty: level I, diagnostic; level II, procedures on the uterus and/or adnexa; level III, second look operations for malignancy; and level IV, lymphadenectomies/other complex procedures. Complications were graded from 1 (mild) to 5 (death). Standard univariate and multivariate analyses were performed. RESULTS: We identified 1451 evaluable procedures. The number of complications was as follows: grades 1 to 5, 129 complications (9%); grades 3 to 5, 36 complications (2.5%). On multivariate analysis, older age ( P = .03), previous radiation ( P = .03), and malignancy ( P = .006) were associated with an increased risk of complications grades 3 to 5. Complication rates for grades 3 to 5 for patients with malignancy versus benign disease was 4% versus 1%, respectively. Technical difficulty led to conversion to laparotomy in 105 cases (7%). Previous abdominal surgery ( P < .001) significantly increased the rate of conversion to laparotomy; more complex, higher procedure levels were associated with a significant decrease in conversions ( P = .005). CONCLUSION: Both simple and complex laparoscopic procedures can be performed by a gynecologic oncology service with a low rate of complications and conversions to laparotomy. Older age, malignancy, previous radiation therapy, and previous abdominal surgery were identified as significant risk factors for complications and/or conversion and should be taken into account in patient selection, preoperative counseling, and surgical planning.

Adolescent↗
PubMedDate not supplied

Do violations of the axioms of expected utility theory threaten decision analysis?

Research demonstrates that people violate the independence principle of expected utility theory, raising the question of whether expected utility theory is normative for medical decision making. The author provides three arguments that violations of the independence principle are less problematic than they might first appear. First, the independence principle follows from other more fundamental axioms whose appeal may be more readily apparent than that of the independence principle. Second, the axioms need not be descriptive to be normative, and they need not be attractive to all decision makers for expected utility theory to be useful for some. Finally, by providing a metaphor of decision analysis as a conversation between the actual decision maker and a model decision maker, the author argues that expected utility theory need not be purely normative for decision analysis to be useful. In short, violations of the independence principle do not necessarily represent direct violations of the axioms of expected utility theory; behavioral violations of the axioms of expected utility theory do not necessarily imply that decision analysis is not normative; and full normativeness is not necessary for decision analysis to generate valuable insights.

Bias↗
PubMed1996

Cleavage of DNA by the insulin-mimetic compound, NH4[VO(O2)2(phen)].

The kinetics and mechanism of cleavage of DNA by the insulin-mimetic peroxo-vanadate NH4[VO(O2)2(phen)], pV, are described. In the presence of low energy UV radiation or biologically common reducing agents, pV decomposes into the monomer, dimer, and tetramer of vanadate and an uncharacterized compound of V4+ as shown by 51V NMR, ESR, and absorption spectra. The rate of photodecomposition of pV is reduced in the presence of calf thymus DNA, indicating that a decomposition product of the peroxo-vanadate, that is important in the destruction pathway of the complex, is interacting with DNA. This species, probably a short-lived complex of V4+, may also be responsible for the observed catalytic decomposition of pV in the absence of DNA by ascorbate. If closed circular pBR322 DNA is present when the peroxo-vanadate is destroyed by either UV radiation or reducing agents, the polymer may have its sugar-phosphate backbone broken. Closed circular DNA (form I) is converted into nicked circular DNA (form II) and linear DNA (form III). The amounts of the various forms produced as a function of irradiation time and peroxo-vanadate concentration were fit to a kinetic model to derive rate constants for the conversions. The kinetic analysis shows that pV is a single-strand nicking agent which exhibits some base and/or sequence preference. Furthermore, the pH dependences of the rates for conversion of form I to form II and for conversion of form II to form III are different, indicating that the nature of the chemistry at the site of cleavage on DNA influences further cutting by activated pV. Reduced amounts of DNA breakage in the presence of various salts and metal binding ligands indicate that a short-lived reactive complex of V4+, not the V4+ species detected by ESR at long irradiation times, is important in the cleavage process. The susceptibility of pV to decomposition by biologically common reducing agents suggests that metabolites of the agent, and not the compound itself, are responsible for its insulin-mimetic effects.

Ascorbic Acid↗
PubMed2002

[Development of an image processing scheme for chest radiographs using a dot printer].

The performance of dot printers has recently been improved. Images output by dot printers can provide simple, economical medical reference images if important diagnostic information is not lost. We developed an image processing scheme for chest radiographs that employed a dot printer. We used two types of pixel value conversions, a nonlinear pixel value correction using a lookup table and a linear pixel value conversion using histogram analysis. The density distribution of chest radiographs was analyzed and classified into high-density and low-density images. The two types of pixel value conversions were used depending on the density of chest radiographs. Converted pixel value had density characteristics that were adapted to the output image of the dot printer, and thus the density distributions of the lungs of radiographs became comparable. In addition, an adaptive unsharp masking technique with processing parameters optimized for each of the lungs and mediastinum was applied. An ROC study for the detection of lung nodules was carried out to evaluate the performance of dot printer images. The area under the ROC curve (A(z)) for dot printer images was 0.816, while sensitivity and specificity were 77.6% and 75.2%, respectively. The performance indicated the usefulness of our image processing scheme.

Humans↗
PubMed2002

Factors associated with conversion to laparotomy in patients undergoing laparoscopic appendectomy.

BACKGROUND: Laparoscopic appendectomy (LA) has been increasingly adopted for its advantages over the open technique, but there is a possibility of conversion to open appendectomy (OA) if complications occur or the extent of inflammation prohibits successful dissection. This study aimed to identify the preoperative predictors for conversion from laparoscopic to open appendectomy. STUDY DESIGN: Medical records of 705 consecutive patients who underwent surgery for suspected appendicitis were reviewed retrospectively. LA was attempted in 595 patients by 25 different surgeons. Factors evaluated were age, gender, body mass index, previous abdominal surgery, previous appendicitis attack, pain, nausea, vomiting, fever, duration of symptoms, local or diffuse tenderness, leukocyte count and surgeon's experience in LA. RESULTS: Conversion to OA occurred in 58 patients (9.7%). The most common reason for conversion was dense adhesions due to inflammation, followed by localized perforation and diffuse peritonitis. Based on 261 patients evaluated by CT scan preoperatively, significant factors in the final multivariate analysis associated with conversion to OA were age > or = 65 [Odds ratio (OR) = 3.78, 95% CI:1.11-12.84], diffuse tenderness on physical examination (OR = 11.32, 95% CI: 1.32-96.62), and a surgeon with less experience in LA (< or = 10 operations, OR = 3.38, 95% CI:1.02-11.17). The presence of significant fat stranding associated with fluid accumulation, inflammatory mass or localized abscess in CT scan also significantly increased the possibility of conversion (OR = 5.60, 95% CI:2.48-12.65). CONCLUSIONS: Identifying the potential factors for conversion preoperatively may assist the surgeons in making decisions concerning the management of patients with appendicitis and in the judicious use of LA.

Adult↗
PubMed1999

Economic analysis of selected lignocellulose-to-ethanol conversion technologies.

The objective of this case study was to examine the economics of three lignocellulose-to-ethanol conversion technologies: fast pyrolysis integrated with a fermentation step, simultaneous saccharification and fermentation (SSF), and dilute sulfuric acid hydrolysis and fermentation. All technologies were assumed to have an annual production rate of 25 million gallons of ethanol. The three technologies were compared in terms of capital costs, operating costs, and ethanol production costs. Sensitivity analyses were carried out to study the uncertainties of wood costs and ethanol production rates on ethanol production costs. Final economic analysis showed that fast pyrolysis integrated with a fermentation step is comparable with the other two processes and suggests that it should be considered for further development.

Journal Article↗
PubMed2005

Optimal initial dose of orally administered cyclosporine following intravenous cyclosporine therapy.

Conversion of the main immunosuppressive agent from tacrolimus to cyclosporine (CyA) is a valuable rescue therapy for the severe adverse effects of tacrolimus. We report our experience of the conversion in living donor liver transplantation, in which tacrolimus therapy was converted to CyA. There are few reports describing the optimal initial dose of orally administered CyA (PO CyA) after intravenously administered CyA (IV CyA). We retrospectively investigated the ratio of CyA blood concentrations at 12 hours after the initiation of PO CyA to that just before the conversion, and the ratio of the initial dose of PO CyA to the total dose of IV CyA administered during the 12 hours before the conversion. Linear regression analysis revealed a close correlation between these ratios (y = 2.7 + 5.8x, r = 0.73; P = .002). We recommend that, to maintain a similar trough level of CyA during conversion in liver transplantation, Neoral should be initiated at a ninefold greater dose than the total dose of IV CyA administered during the 12 hours before conversion.

Administration, Oral↗
PubMed1985

High-performance liquid chromatographic studies on non-mercapt in equilibrium with mercapt conversion of human serum albumin. II.

High-performance liquid chromatographic (HPLC) analysis of human serum albumin (HSA) on a GS-520 column with 0.03 M sodium phosphate buffer-0.15 M sodium sulphate (pH 6.87) showed three peaks, the principal component corresponding to human mercaptalbumin (HMA) and the secondary and tertiary components to nonmercaptalbumin (HNA). Using HPLC analysis, the nonmercapt----mercapt conversion of HSA during haemodialysis and the mercapt----nonmercapt conversion after haemodialysis in chronic renal failure were re-confirmed, indicating that HMA is a covalent carrier protein for sulphur-containing amino acids. Fractions of HMA in various liver diseases were significantly lower than those of healthy male adults.

Adult↗
PubMed1994

Gene cluster controlling conversion to alginate-overproducing phenotype in Pseudomonas aeruginosa: functional analysis in a heterologous host and role in the instability of mucoidy.

Conversion to mucoidy, caused by the overproduction of the exopolysaccharide alginate in laboratory and cystic fibrosis strains of Pseudomonas aeruginosa, can occur via frameshift or nonsense mutations in the second gene of the algU mucA mucB cluster. The first gene of the cluster, algU, encodes a putative alternative sigma factor required for algD transcription. The algD gene encodes a critical alginate biosynthetic enzyme and is invariably activated in mucoid P. aeruginosa cells. To investigate the function of the genes controlling conversion to mucoidy, the wild-type algU mucA mucB cluster from the standard genetic strain PAO1 was used to reconstitute algD transcription in Escherichia coli. Transcription of an algD-lacZ chromosomal fusion in E. coli was detected upon introduction of plasmid-borne algU mucA mucB. Moreover, insertional inactivation of either mucA or mucB resulted in further stimulation of transcriptional activity from the algD promoter. This activation was dependent on algU, since a double algU mucA mutation abrogated transcription of algD. These experiments suggest that the phenotypic manifestations of muc mutations, i.e., increased algD expression and mucoid phenotype, depend on the presence of an active algU gene and that this regulator and the factors encoded by the downstream genes interact. Further support for these conclusions came from the investigations of the mechanism of reversion to nonmucoidy in P. aeruginosa, a phenomenon frequently referred to as the instability of mucoid phenotype. Spontaneous nonmucoid derivatives of the mucoid strain PAO578 carrying the mucA22 mutation were examined for the presence of alterations within the algU mucA mucB locus. Point mutations which inactivated algU were detected in some, but not all, nonmucoid revertants. No reversion of the original mucA22 mutation (a deletion of one C) was observed in any of the investigated strains. This observation suggests that the process of conversion to nonmucoidy ban be explained, at least partially, by second-site suppressor mutations and that a fraction of such mutations occurs in algU.

Alginates↗