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Impact at the Permo-Triassic boundary: a critical evaluation.

The recognition in 1980 of a signature of an extraterrestrial impact at the Cretaceous-Tertiary boundary and its apparent involvement with the mass extinction generated considerable enthusiasm for impacts at other mass extinctions. Numerous claims of impact evidence for the Permo-Triassic mass extinction (251.6 Ma), the largest of the Phanerozoic mass extinctions, have generally been rejected, found wanting, or been difficult to reproduce. Despite this lack of repeatable support, considerable available evidence is consistent with an impact, including the rapidity of extinction, coincident carbon shift, and evident correlation between terrestrial and marine extinctions. However attractive the hypothesis, the coincidence with the Siberian flood basalts and the complex nature of the carbon shift are in conflict with an impact. The most intriguing possibility is that the greatest mass extinction of the Phanerozoic left signals very similar to the end-Cretaceous mass extinction but was produced by entirely Earth-bound processes. If true, this would tell us far more about the nature of ecosystems and how they fail than would identification of another impact.

Carbon↗

Fast quantification of ethanol in whole blood specimens by the enzymatic alcohol dehydrogenase method. Optimization by experimental design.

A sensitive, fast, simple, and high-throughput enzymatic method for the quantification of ethanol in whole blood (blood) on Hitachi 917 is presented. Alcohol dehydrogenase (ADH) oxidizes ethanol to acetaldehyde using the coenzyme nicotinamide adenine dinucleotide (NAD), which is concurrently reduced to form NADH. Method development was performed with the aid of factorial design, varying pH, and concentrations of NAD+ and ADH. The linear range increased and reaction end point decreased with increasing NAD+ concentration and pH. The method was linear in the concentration range 0.0024-0.4220 g/dL. The limits of detection and quantification were 0.0007 g/dL and 0.0024 g/dL, respectively. Relative standard deviations for the repeatability and within-laboratory reproducibility were in the ranges 0.7-5.7% and 1.6-8.9%, respectively. The correlation coefficient when compared with headspace gas chromatography-flame ionization detection methods was 0.9903. Analysis of authentic positive blood specimens gave results that were slightly lower than those of the reference method.

Alcohol Dehydrogenase↗

An automated alcohol dehydrogenase method for ethanol quantification in urine and whole blood.

In a previous work, an automated alcohol dehydrogenase method for the quantification of ethanol in whole blood (blood) specimens was presented. In the present work the application of the method to urine specimens has been investigated. Also, method robustness to routine analysis of urine and blood specimens during a period of eight months is shown. The limits of detection and quantification for urine were 0.0012 g/dL and 0.0042 g/dL, respectively. Relative standard deviations for the repeatability and within-laboratory reproducibility were in the ranges 1.4-4.1% and 1.8-4.6%, respectively. The method was compared with two headspace gas chromatography-flame ionization detection methods using authentic forensic urine specimens (n = 305) and blood specimens (n = 3186). Passing-Bablok regression for the concentration range 0.01-0.48 g/dL (urine) and 0.002-0.40 g/dL (blood) showed a statistically significant difference, for urine y = 0.9313 (0.9250 - 0.9377)x + 0.0038 (0.0029-0.0044) and for blood y = 0.9493 (0.9491 - 0.9495)x + 0.0032 (0.00318-0.00323), at 95% confidence level. The results of the external quality control specimens were in accordance with the reported theoretical concentrations.

Alcohol Dehydrogenase↗

Clinical evaluation of the peritoneal equilibration test: a population-based study.

A total of 143 Peritoneal Equilibration Tests (PETs) were performed in 104 CAPD patients over a period of 18 months. A normal range (95% confidence limits) was constructed from 100 tests (68 consecutive new patients, 32 routine tests on problem-free patients) using a 2-dimensional plot of solute transfer (D/Pcreat) and ultrafiltration volume. These two parameters correlated inversely (r = -0.59, P < 0.0001) allowing calculation of a regression line. In the short term (< 3 months) D/Pcreat was stable across a wide range of values (0.45-0.98) with good correlation between tests indicating reproducibility (r = 0.94, P < 0.001). Repeated tests beyond 3 months were variable, explaining changes in the clinical picture, and in the majority of cases shifts in D/Pcreat and ultrafiltration parallelled the regression line for the whole population. Six of seven (85%) of patients with mechanical problems and 14 of 15 (93%) with poor ultrafiltration had at least one abnormal test, and these two problems could be distinguished in 90% of cases by 2-dimensional plotting. In five with ultrafiltration failure, dialysate volumes were less than predicted by solute clearance, and these patients failed continuous cycling peritoneal dialysis (CCPD). In contrast, a good response to CCPD was predicted correctly in five patients with high solute clearance. In nine patients with plasma creatinine > 1250 mumol/l the PET was normal. The PET is a useful adjunct to understanding and prescribing peritoneal dialysis, particularly when repeated tests are compared to a well-defined normal population.

Adolescent↗

Estimation of end-expiratory lung volume variations by optoelectronic plethysmography.

OBJECTIVE: To test the capability of optoelectronic plethysmography (OEP) to monitor positive end-expiratory pressure (PEEP)-induced changes of end-expiratory lung volume (EELV) changes in mechanically ventilated paralyzed patients. DESIGN: Laboratory and clinical investigation. SETTING: Intensive care unit of the Ospedale Maggiore Policlinico di Milano. PATIENTS: A total of eight patients with respiratory failure of various degrees, sedated and paralyzed. INTERVENTIONS: PEEP variations (+/-5 cm H2O) relative to the baseline PEEP of 10 cm H2O. MEASUREMENTS AND MAIN RESULTS: In the model protocol, we tested the reproducibility of the OEP by repeating volume measurements of a plastic torso model over a 21-hr period, every 30 mins. The variations of OEP measurements of the torso model (9337 mL value) were encountered in a range of 16 mL (sd = 4 mL). In the patient protocol, we measured the end-expiratory volume of the chest wall (EEVCW) breath-by-breath by OEP before, during, and after the PEEP increase/decrease and we compared its variations with the corresponding variations of EELV measured by helium dilution technique. The regression line between EELV changes measured by helium and EEVCW changes measured by OEP resulted very close to the identity line (slope 1.06, intercept -0.02 L, r(2) = 0.89) and their difference was not related to their absolute magnitude. After PEEP increase, the new steady state of EEVCW was reached approximately in 15 breaths; and, after PEEP decrease, in 3-4 breaths. The slow increase in EEVCW was mainly because of the abdominal compartment. CONCLUSION: OEP measurements of EEVCW accurately reflect the changes of EELV. Furthermore, OEP allows a continuous compartmental analysis, even during unsteady conditions.

Adult↗

Study of the electric activity of the liver with identification of a normal 'electrohepatogram' in a canine model.

OBJECTIVE: The electric activity of the liver was studied in 14 mongrel dogs, aiming at characterizing a normal electrohepatogram (EHG) that might act as a standard for abnormal recordings in hepatic pathological conditions. METHODS: Three monopolar silver-silver chloride electrodes were sutured to the anterior liver surface under anaesthesia. A recording session of 60 min was performed daily for 10 days. The effect of temporary clamping of the hepatic artery and portal vein, and of liver insult by irradiation, on the hepatic electric activity was also studied. RESULTS: Electric waves were recorded from the three electrodes. They were monophasic with a positive deflection, had identical frequency and amplitude from the three electrodes, and were reproducible when the test was repeated in the same animal. The mean frequency was 10.6 +/- 1.8 cycles/s and amplitude 63.7 +/- 11.4 microV. Fast activity spikes and abnormal waves were not encountered. Temporary occlusion of the hepatic artery and portal vein produced irregularities in the wave frequency and amplitude; the wave dysrhythmia disappeared after clamp release. The EHG following liver insult by irradiation exhibited two patterns: silent and dysrhythmic. CONCLUSION: A normal EHG could be characterized in the canine model. A dysrhythmic pattern was produced when liver vessels were clamped and following liver injury. It is suggested that in liver diseases, the normal EHG pattern is deranged and might thus act as a diagnostic tool in such conditions.

Animals↗

Quantitative in vitro assessment of N-alkyl sulphate-induced cytotoxicity in human keratinocytes (HaCaT). Comparison with in vivo human irritation tests.

A spontaneously immortalized human keratinocyte line, HaCaT, was used as an in vitro model to predict the cutaneous irritation of anionic surfactants. For this purpose, a number of sodium salts of N-alkyl sulphates with hydrocarbon chain lengths varying between C8 and C16 were studied for possible cytotoxic effects. The endpoints used to assess toxicity were uptake of the vital dye neutral red (NR) and cell morphology criteria 24 h after dosing. A linear proportionality between keratinocyte number and NR uptake was established. All tested surfactants had cytotoxic effects as demonstrated by a decreased NR uptake, which showed a clear dose-response relationship. Concentrations resulting in 50% inhibition of NR uptake (IC-50) ranged from 0.15 mmol (sodium lauryl sulphate, C12) to 1.23 mmol (sodium octyl sulphate, C8). The in vitro cytotoxicity data were highly reproducible when the test was repeated after several weeks. The cytotoxicity data from these assays were compared with the irritant responses (as evaluated by measurement of erythema and transepidermal water loss) obtained after 24 h application of the same compounds (300 microliters of 20 mmol aqueous solution) to the volar forearm of human volunteers. There were significant linear correlations between the IC-50 values and both barrier damage (transepidermal water loss) and erythema (as evaluated by skin colour reflectance measurements). For the test substances, however, the sensitivity of the in vitro system was between 10 and 100 times higher than that observed in human skin in vivo.(ABSTRACT TRUNCATED AT 250 WORDS)

Cell Line↗

Cytotoxicity of MTA and Portland cement on human ECV 304 endothelial cells.

AIM: To evaluate the cytotoxic effects of two brands of mineral trioxide aggregate (MTA) (Pro-Root MTA and MTA Angelus) and Portland cement (PC) on the human ECV 304 endothelial cell line. METHODOLOGY: Endothelial ECV 304 cells were incubated at 37 degrees C in an atmosphere of 95% air, 5% carbon dioxide and 100% humidity for 7 days and grown in F12 medium supplemented with 10% fetal bovine serum with 50 microg mL(-1) of gentamicin sulphate. Effects of the materials on mitochondrial functions were measured by a colorimetric assay. At each experimental time interval (24, 48 and 72 h), a dimethyl-thiazol-diphenyl tetrazolium bromid assay was conducted to measure cell viability. All assays were repeated three times to ensure reproducibility. Results were expressed as average absorbance (A(570/nm)) +/- SD and the data were analysed statistically by one-way analysis of variance and the Bonferroni post-test. A P-value < 0.05 was considered statistically significant. RESULTS: No statistically significant difference was shown between any of the experimental materials (P > 0.05). CONCLUSIONS: The two brands of MTA analysed, as well as the PC, initially showed a similar elevated cytotoxic effect that decreased gradually with time allowing the cell culture to become reestablished.

Aluminum Compounds↗

The short insulin tolerance test for determination of insulin sensitivity: a comparison with the euglycaemic clamp.

The glucose clamp technique is currently regarded as the standard test for measuring insulin sensitivity against which other methods are compared but is unsuitable for routine screening of patients outside a hospital base. There is thus a need for a simpler test to measure insulin sensitivity. We have therefore compared the glucose disappearance rate KITT in the first 15 min of the insulin tolerance test (ITT) with the M and M/I values derived from the standard euglycaemic clamp in nine normal subjects and eight subjects with Type 2 (non-insulin dependent) diabetes mellitus and coexisting obesity. All subjects underwent the ITT and euglycaemic clamp in random order. Nine subjects later had a repeat ITT to determine the reproducibility of the test. In the ITT, 0.1 U kg-1 body weight, human Actrapid insulin was given as an IV bolus and simultaneous arterialized and venous blood samples were obtained every minute for 15 min. The first order rate constant for the disappearance of glucose KITT over the period 3-15 min was taken as a measure of insulin sensitivity. The euglycaemic clamp was performed with an insulin infusion of 50 mU kg-1 h-1 for 120 min and a variable rate glucose infusion to maintain blood glucose concentration at 0.5 mmol l-1 below fasting level to minimize the effect of endogenous insulin secretion. The ratio of the mean rate of glucose infused (M, mumol kg-1 min-1) to the plasma insulin over the last 30 min of the clamp was taken as a measure of tissue sensitivity to insulin (M/I) assuming endogenous glucose output was suppressed.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

Effects of nicorandil on the recovery of reflex potentials after spinal cord ischaemia in cats.

1. The pathophysiological significance of ATP-sensitive K+ (KATP) channels in the central nervous system is not fully understood. In this study the effects of nicorandil (a hybrid vasodilator having a dual mechanism of action as a K+ channel opener and a nitrate) on the recovery of the spinal cord reflex potentials after spinal cord ischaemia were examined and compared with those of pinacidil and nitroprusside in anaesthetized spinal cats. 2. Spinal cord ischaemia was produced by occlusion of the thoracic aorta and the bilateral internal mammary arteries for 10 min. Regional blood flow in the spinal cord was continuously measured with a laser-Doppler flow meter. The monosynaptic (MSR) and polysynaptic reflex (PSR) potentials, elicited by electrical stimulation of the tibial nerve, were recorded from the lumbo-sacral ventral root. The recovery process of spinal reflex potentials was reproducible when the occlusion was repeated twice at an interval of 120 min. 3. Pretreatment with nicorandil (30-100 micrograms kg-1) accelerated the recovery of PSR potentials after spinal cord ischaemia. Such an accelerating effect on the recovery of PSR potentials was also shared by pinacidil (100 micrograms kg-1), another K+ channel opener. In addition, the accelerating effect of nicorandil (100 micrograms kg-1) on the recovery of PSR potentials was abolished by co-administration of glibenclamide (3 mg kg-1), a sulphonylurea KATP channel blocker. Nitroprusside (8 micrograms kg-1min-1) retarded rather than improved the recovery of PSR potentials after spinal cord ischaemia. All of these drugs failed to improve the spinal cord blood flow during ischaemia and reperfusion. 4 These results suggest that nicorandil promotes the recovery of polysynaptic reflex potentials after spinal cord ischaemia by opening the KATP channels of neurones rather than by increasing local bloodflow. K+ channel openers may exert a salutary effect on the functional recovery of the ischaemic spinal cord.

Animals↗

Development of esophageal hypersensitivity following experimental duodenal acidification.

OBJECTIVES: As visceral afferents from different regions of the gastrointestinal tract converge at the level of the spinal cord, we hypothesized that sensitization of one gut organ would induce visceral hypersensitivity in another gut organ, remote to the sensitizing stimulus. METHODS: Protocol 1: Eight healthy male volunteers, age 30 +/- 8.2 yr, underwent three studies on different days. Esophageal pain thresholds (PT) were recorded at 10-min intervals prior to and for 2 h following a 30-min duodenal infusion of either 0.15 M hydrochloric acid (HCl), saline, or no infusion. Five subjects repeated the study to demonstrate reproducibility. Protocol 2: Esophageal evoked potentials (EEP) were studied in six subjects on two occasions prior to and 1 h after a 30-min duodenal infusion of 0.15 M HCl or saline. RESULTS: Protocol 1: After acid infusion, there were reproducible reductions in esophageal PT (ICC = 0.88), which were maximal at 110 min (15.05 +/- 2.25 mA) (p < 0.002). Following saline infusion there was an increase in esophageal PT (ICC = 0.71), which was similar to the no-infusion condition (6.21 +/- 1.54 mA vs 8.5 + 7.6 mA; p > 0.05). Protocol 2: Esophageal sensation scores increased (p= 0.02) after acid, but not after saline infusion (p= 0.1). A comparison of the latencies of EEP components prior to and following acid and saline infusion revealed a reduction in the N1 (p= 0.02) and P2 components (p= 0.04). CONCLUSION: This study provides the first objective evidence that duodenal acidification can induce esophageal hypersensitivity associated with changes in sensitivity of the central visceral pain pathway. As the esophagus was remote from the sensitizing stimulus, central sensitization of spinal dorsal horn neurons is likely to have contributed to these changes.

Adult↗

Quantification of dental plaque on lingual tooth surfaces using image analysis: reliability and validation.

AIM: The aim of this study was to increase the versatility and further validate the method reported by Smith et al. (2001) by testing the reliability of plaque measurement against two well-known dental plaque quantification methodologies using image analysis in a clinical trial. METHOD: The teeth of 40 subjects were disclosed before digital images of the labial and lingual surfaces of their upper and lower incisors were acquired. The amount of plaque present was quantified using a modification of the method described by Smith et al. (2001). The method was modified for obtaining images of the lingual surfaces by incorporating the use of orthodontic occlusal mirrors and 5-mm pieces of moistened blue articulating paper used to enable calibration. Plaque measurements were made from 320 upper and lower anterior teeth from the 40 subjects by two operators. Fliess' coefficient of reliability was used to assess intra- and inter-operator reliability and the independent sample t test was used to assess statistical significance between test and control groups after checking the data for normality. For validation, measurements were recorded using the Turesky et al. (1970) (modification of the Quigley & Hein (1962) plaque index and the Addy et al. (1983) plaque area index. The results were compared with the image analysis method using Pearson's correlation coefficient. RESULTS: The results for reliability were within Fliess' range of "excellent" for both intra-operator repeatability and inter-operator reproducibility. Pearson's correlation coefficients showed highly significant values indicating the close similarity between all three methods. CONCLUSIONS: This method for the measurement of dental plaque on lingual surfaces of anterior teeth proved reliable. The combined results from the labial and lingual surfaces of anterior teeth using image analysis produced trial conclusions comparable with the alternate plaque quantification methods used, with less clinician time and further producing a permanent database of images for future use.

Adult↗

Phage-typing of coagulase-negative staphylococci. Factors influencing typability.

Factors influencing the phage-typability of coagulase-negative Micrococcaceae have been studied in 2,778 clinical isolates comprising A) 209 consecutive isolates from one laboratory, B) 2,107 clinical strains submitted for phage-typing for epidemiological reasons, and C) 462 strains representing all isolates of presumed clinical significance found in two laboratories during one month. The reproducibility was acceptable at duplicate repeated typing of the same strains as well as by typing epidemiologically-related pairs of strains from the same patient. Strains of Staphylococcus haemolyticus were seldom typable, whereas strains of S. epidermidis and S. hominis had a higher typability. Methicillin-resistant strains and other multiple-resistant strains were rarely typable (11-13%). The typability was higher among susceptible strains (36%) and strains resistant to penicillin only (43-50%). The typability of strains of the same species and antibiotic-resistance pattern differed between hospitals compared and decreased markedly over the years for multiple-resistant S. epidermidis isolates.

Bacteriophage Typing↗

Quantitation of noradrenaline nerve density in mouse iris by computer-assisted image analysis.

The density of noradrenaline (NA)-containing nerve fibres in mouse iris was measured with computer-assisted image analysis techniques both under normal conditions and during regeneration. Noradrenaline nerves were visualized by Falck-Hillarp formaldehyde condensation technique in whole-mount spread preparations of mouse irides. The samples were analysed in a fluorescence microscope connected to a commercially available image analysis system (IBAS/Kontron). A software program was developed for specific detection of fluorescence and the nerve density was determined by calculating the area covered by fluorescence in percentage of total measuring field. The method showed good reproducibility as observed both when repeated measurements were performed in the same measuring field or when consecutive measurements on the same set of irides were performed. Also the inter-assay variation between control values in the different experiments was low. Loading of the adrenergic nerves by incubation in alpha-methyl-NA or conditions leading to partial diffusion of the fluorophore had minor effects on the nerve density values. The regeneration of the NA nerve fibres after a selective toxic sympathectomy with 6-hydroxydopamine was also studied. The nerve fibre density values measured by image analysis correlated well with the uptake of [3H]NA; the endogenous NA levels recovered much more slowly, however. It thus seems that endogenous transmitter levels might be a somewhat insensitive index of nerve terminal regrowth, at least in early stages of regeneration. The results indicate that image analysis is a powerful tool to quantitate a transmitter-identified nerve terminal network in a histological preparation.

Adrenergic Fibers↗

Comparison of ROC and AFC methods in a visual detection task.

The visual observer performance for detecting low-contrast patterns by employing both the receiver operating characteristic (ROC) and the 9-alternative forced choice (9-AFC) method is investigated. For each observer and each experimental technique, the results were reproducible with respect to a repeated observation. When the results of different observers for each method are compared, a strong variation of values is found. From the ROC data obtained, the expected percentages of correct detection in a 9-AFC experiment is calculated and compared with the corresponding values of the actual 9-AFC experiment. For some observers, the latter values were significantly higher than the computed ones, whereas for others the results showed good agreement with predictions from theory.

Decision Making↗

Comparison of plasma virus loads among individuals infected with hepatitis C virus (HCV) genotypes 1, 2, and 3 by quantiplex HCV RNA assay versions 1 and 2, Roche Monitor assay, and an in-house limiting dilution method.

The accuracy of different methods for the quantitation of hepatitis C virus in plasma was measured with samples from individuals infected with different genotypes and by using RNA transcripts of predetermined concentrations. Highly reproducible results were observed upon repeat testing of samples by both the original version of the Chiron branched-DNA (bDNA) assay (Quantiplex RNA assay; bDNA-1) and the currently available version (Quantiplex HCV RNA 2.0 assay; bDNA-2). A greater variability was observed in the Roche Monitor assay (correlation coefficient of 0.537, compared with 0.942 and 0.964 for the bDNA-1 and bDNA-2 assays, respectively). Significant differences in the efficiency of detection of genotypes 1, 2, and 3 were observed for the bDNA-1 and Roche Monitor assays, whereas the bDNA-2 assay and nested PCR at limiting dilution were able to quantify genotypes with equal sensitivity. By quantifying RNA transcripts of different genotypes, the sensitivities of the Roche Monitor assay for sequences of the type 2 and type 3 transcripts were estimated to be 11 and 8% of those achieved for genotype 1. When correction factors based upon these results and those from quantitation of circulating viral RNA sequences in samples from blood donors were used, the genotype-specific differences in virus load in samples from blood donors were no longer observed, consistent with previous studies with corrected values from the bDNA-1 assay. These results suggest that many of the previous studies evaluating the effect of genotype and virus load on the response to interferon using methods such as the Roche Monitor assay and other competitive PCR methods require reinterpretation. Differences in efficiency of quantitation should be taken into account in future investigations of the relationship between genotype and virus load.

Genes, Viral↗

Gallbladder mucosal function: studies in absorption and secretion in humans and in dog gallbladder epithelium.

The gallbladder is conventionally regarded as an absorptive organ such that dilute hepatic bile is both stored and concentrated. We studied 35 patients who had recovered from a percutaneous transhepatic gallbladder drainage performed for acute cholecystitis. After an overnight fast, gallbladder bile was dark brown in color and had a wide scatter in the lipid composition. Two hours after a meal, the gallbladder bile was opalescent white in color and had the composition of an extracellular fluid. This phenomenon was uniformly observed in all 35 patients and was also consistently reproducible when five patients were repeatedly studied. We used normal dog gallbladder epithelial cell monolayers grown in culture and examined sodium flux. Control gallbladder cells absorbed sodium. When secretin (0.5-2.5 x 10(-7) M) was added, there was a prompt reversal of sodium flux, resulting in net secretion. We conclude that secretion is a physiological function of the gallbladder mucosa. After feeding, the neural and humoral factors divert stored and newly secreted bile into the duodenum and induce active de novo secretion thus producing a gallbladder bile that is opalescent white with no lipids. Our results also have important implications on the origin of the pathological "white bile," the pathogenesis and treatment of gallbladder sludge, as well as the kinetic analysis of compounds undergoing enterohepatic recirculation.

Absorption↗

Relative hysteresis of the airways and lung parenchyma in normal subjects.

We evaluated the mechanical properties of the airways sequentially from the glottis toward the main bronchi in 10 normal subjects. Plots of airway cross-sectional area vs. lung volume, measured during inspiration and expiration, were used to determine the relative magnitude of the airways vs. parenchymal hysteresis. Airway cross-sectional area was measured by means of the acoustic reflection technique. We found that the hysteresis of the proximal part of the trachea was greater than that of the lung parenchyma, whereas the hysteresis of the distal trachea and subcarinal segments of the airways was smaller than that of the lung parenchyma. The transition zone between the proximal and the more distal airway properties occurred 8-26 cm distal to the glottis. This transition zone was reproducible in its location on repeated testing in each subject but varied among subjects. To the extent that relative hysteresis of the airways depends on bronchomotor tone, our findings suggest that the bronchomotor tone is inhomogeneous, being maximal at the proximal part of the trachea and gradually decreasing toward the more distal trachea and subcarinal airway segments.

Acoustics↗