Okadaic acid-induced lens epithelial cell apoptosis requires inhibition of phosphatase-1 and is associated with induction of gene expression including p53 and bax

We report the 2.2 A resolution structure of the Drosophila engrailed homeodomain bound to its optimal DNA site. The original 2.8 A resolution structure of this complex provided the first detailed three-dimensional view of how homeodomains recognize DNA, and has served as the basis for biochemical studies, structural studies and molecular modeling. Our refined structure confirms the principal conclusions of the original structure, but provides important new details about the recognition interface. Biochemical and NMR studies of other homeodomains had led to the notion that Gln50 was an especially important determinant of specificity. However, our refined structure shows that this side-chain makes no direct hydrogen bonds to the DNA. The structure does reveal an extensive network of ordered water molecules which mediate contacts to several bases and phosphates (including contacts from Gln50), and our model provides a basis for detailed comparison with the structure of an engrailed Q50K altered-specificity variant. Comparing our structure with the crystal structure of the free protein confirms that the N and C termini of the homeodomain become ordered upon DNA-binding. However, we also find that several key DNA contact residues in the recognition helix have the same conformation in the free and bound protein, and that several water molecules also are "preorganized" to contact the DNA. Our structure helps provide a more complete basis for the detailed analysis of homeodomain-DNA interactions.     

Exploring the genetic and environmental etiology of high general cognitive ability in fourteen- to thirty-six-month-old twins

Two definitions of normality ("isolated" or "correlated") are considered. The boundaries of "isolated" normality were determined by a statistical procedure, whereas the "correlated" approach was related to a clinical or predictive definition. In the latter case, the biological variations were considered abnormal if they implied a hazard with some significant future ailment as a risk factor. In this pragmatic approach, the upper limit of normal/abnormal variations is the point beyond which medical strategy is related to the most expected benefit when applied to a definite population or to an individual patient. The capacity of a diagnostic test to discriminate between patients with a defined risk and those without risk depends strictly on the value of the parameter chosen. In medical care for the prevention of vascular complications in diabetic patients or with foetal risks in pregnant women, the limits of the so-called normal range of glycaemia and other parameters should be determined according to the objective of the preventive and/or therapeutic measures to be prescribed.     

Prophylactic efficacy of tilmicosin for bovine respiratory tract disease

The prophylactic administration of injectable tilmicosin for pneumonia in weaned beef calves was investigated in 1,806 animals. Comparisons were made among calves receiving an "on-arrival" injection of tilmicosin, calves receiving a single injection of long-acting oxytetracycline, and calves receiving no prophylaxis. Morbidity and mortality attributable to pneumonia, morbidity and mortality attributable to all causes, and case fatality were significantly lower in the group of calves that received tilmicosin, compared with calves that received long-acting oxytetracycline and calves that received no prophylactic antibiotic. Mean time to initial pneumonia treatment was significantly extended in calves that received prophylaxis, compared with those that received no antibiotic on arrival at the feedlot. Calves that received tilmicosin gained significantly more weight than calves that received oxytetracycline. Calves that were not treated for pneumonia during the trial period gained significantly more weight than did those calves that were treated for pneumonia regardless of experimental group. The majority of mortalities were attributable to fibrinous pneumonia (31/34). Important bacterial isolates (Pasteurella spp, Haemophilus somnus, Actinomyces pyogenes) obtained at necropsy did not have resistance to tilmicosin in association with administration of tilmicosin as prophylaxis for pneumonia. However, bacterial resistance to trimethoprim/sulfonamide and to oxytetracycline were commonly found in these postmortem isolates.     

Application of confocal scanning laser microscopy in experimental pathology

Confocal scanning laser microscopy (CSLM) represents an exciting new tool for scientific disciplines which focus on mechanistic studies such as experimental pathology. Enhanced resolution in the specimen plane and rejection of out-of-focus fluorescence flare allow analysis of specific nucleic acid sequences, enzymes, structural macromolecules, and cellular homeostasis utilizing fluorescent probes. Four different experimental applications are discussed which utilize CSLM to evaluate pathological processes at the subcellular, cellular, and tissue levels. Programmed cell death, or apoptosis, is a natural process of significance both during development and as a response to toxic stimuli. CSLM-imaging of nuclei of human B lymphoblastoid cells following exposure to a monofunctional alkylating agent suggests that the degradation of chromatin characteristic of apoptosis may occur in asymmetric patterns. Surfactant apoprotein-A is the major non-serum protein component of pulmonary surfactant and is essential for the extracellular function of surfactant. CSLM of alveolar type II cells suggests that apoprotein-A is present in both the cytoplasm, predominantly in lamellar bodies, and in the nucleus. The tumor promoter, phorbol myristate acetate, rapidly stimulated the formation of vacuoles in human neutrophils. CSLM using Lucifer Yellow as a probe suggests that cylindrical vacuoles are formed by fluid-phase pinocytosis. The blood-nerve barrier (BNB) in peripheral nerves may be an important target during toxin-induced neuropathies. Ricin-induced permeability of the BNB in the rat was rapidly visualized by CSLM as leakage of fluorescein isothiocynate (FITC)-dextran into the endoneurial compartment.     

Dishwashing behaviour of European consumers with regard to the acceptance of long programme cycles

Since the introduction of the energy label for household dishwashers in the EU, manufacturers have been incentivised to reduce resource consumption and increase the energy efficiency of their appliances. Technological progress has led to very efficient programmes with cleaning cycles of 3 to 4 h or longer. The European Commission recently initiated a revision of the energy label and Ecodesign requirements, leading to their adjustment to the state of the art and to actual usage patterns. The University of Bonn was tasked with investigating dishwashing habits in Europe. An online survey was conducted in 11 countries of the EU with more than 5000 participants. The survey focused on the choice of programme, attitudes towards energy-saving programmes and practices and the willingness to apply them. It appears that consumers are willing to apply energy-saving practices and to use energy-saving programmes, but the acceptance of long cycles that take more than 2 h is low, which stands in contradiction to the fact that 19% of all dishwashing cycles are run in the Eco programme, which takes more than 2 h in most cases. The percentage of people who understand that long cycles can be energy-efficient is smaller than the percentage of those who do not believe this. The statements of the participants are contradictory regarding the importance of saving energy and of programme duration. The results of the survey point out the importance of better consumer education and better communication by manufacturers, consumer organisations and legislation.     

Role of the liver and gut in systemic diphenhydramine clearance in adult nonpregnant sheep

We investigated the contribution of the liver and gut to systemic diphenhydramine (DPHM) clearance in adult nonpregnant sheep in two separate studies. In the first study, a simultaneous 50-mg bolus each of DPHM and its deuterium-labeled analog ([2H10]DPHM) was administered to five sheep via the femoral (i.v.) and the portal venous (p.v.) routes in a randomized manner. Arterial plasma concentrations of DPHM, [2H10]DPHM, and their deaminated metabolites, DPMA (diphenylmethoxyacetic acid) and [2H10]DPMA, were measured using gas chromatography-mass spectrometry. The hepatic first-pass extraction of DPHM after p.v. administration was 94.2 +/- 3.7%. However, the area under the plasma concentration versus time profile of the metabolite after i.v. dosing was only 32.5 +/- 14.0% relative to that after p.v. administration. Thus, only approximately 32.5% of the i.v. dose is metabolized in the liver and a significant extrahepatic systemic clearance component is evident. Using the calculated total hepatic blood flow values, it was found that 98.6 +/- 9.2% of the i.v. dose eventually was delivered to the "hepatoportal" system. Because the drug delivered to the hepatoportal system is almost completely eliminated in a single pass (hepatic extraction approximately 94%), this indicates a lack of any significant pulmonary drug uptake. Also, because only approximately 32.5% of the i.v. dose is metabolized in liver, the gut is most likely responsible for the clearance of the remainder. This gut contribution to systemic DPHM clearance was confirmed in a separate direct study in four sheep where the steady-state DPHM gut extraction ratio was 49.0 +/- 3.0%. Thus, gut accounts for a significant proportion (>/=50%) of DPHM systemic clearance in sheep in spite of a very high hepatic drug extraction efficiency.     

Enhancement of the surface expression of tumor necrosis factor alpha (TNFalpha) but not the p55 TNFalpha receptor in the THP-1 monocytic cell line by matrix metalloprotease inhibitors

The monocytic cell line THP-1 can be induced to express and release tumor necrosis factor alpha (TNFalpha) and both TNFalpha receptors (p55 and p75) upon exposure to bacterial lipopolysaccharide (LPS). The broad-spectrum matrix metalloprotease (MMP) inhibitors [4-(N-hydroxyamino)-2R-isobutyl-3S-(phenylthiomethyl)succinyl]-L-p henylalanine-N-methylamide (GI-129471) and marimastat [2S-[N4(R*),2R*,3S*]]-N4[2,2-dimethyl-1-[(methylamino)carbonyl]propyl]-N 1,2-dihydroxy-3-(2-methylpropyl)butanediamide (BB-2516) were effective inhibitors of LPS-induced TNFalpha (soluble) release with IC50 values of 0.2 and 4.0 microM, respectively. Upon LPS stimulation, the expression of pro-TNFalpha (membrane associated) on the cell surface (FACS analysis) could not be observed. However, in the presence of GI-129471, a concentration-dependent increase in TNFalpha surface expression was observed. Peak expression (percentage of cells expressing pro-TNFalpha and mean fluorescence units) in the presence of GI-129471 was at 2 hr, and steadily declined to return to near control levels by 8 hr. This time course was similar to TNFalpha release, which also peaked at 2-4 hr after LPS exposure and then declined. Stimulation of THP-1 cells with LPS + phorbol myristate acetate increased the percentage of cells expressing pro-TNFalpha by 10-fold. In the presence of GI-129471, these increases were augmented further and peaked between 2 and 4 hr, but also returned to near control levels of expression by 24 hr. This was in contrast to the release of soluble TNFalpha, which continued to accumulate over a 24-hr time course. TNFalpha receptor I (p55, TNFRI) and II (p75, TNFRII) shedding was also inhibited by GI-129471 (IC50 = 1.5 and 3.1 microM, respectively) and BB-2516 (IC50 = 14 and 15 microM, respectively). Unlike pro-TNFalpha surface expression, surface expression of both TNFalpha receptors steadily increased over 72 hr. In contrast to pro-TNFalpha surface expression, TNFRI surface expression was not augmented by these MMP inhibitors in THP-1 cells after LPS stimulation. Surface expression of TNFRII was augmented by these MMP inhibitors. These results suggest that even in the continued presence of LPS stimulation and an inhibitor of TNFalpha processing, the augmented surface expression of TNFalpha is transient. The potential "deleterious" implications of high levels of surface pro-TNFalpha expression in the presence of these inhibitors may be lessened by its transient nature.     

Cerebral emboli and serum S100beta during cardiac operations

BACKGROUND: The glial protein S100beta has been used to estimate cerebral damage in a number of clinical settings. The purpose of this investigation was to determine the correlation between cerebral microemboli and S100beta levels during cardiac operations. METHODS: Transcranial Doppler ultrasonography was used to measure emboli in the right middle cerebral artery. Emboli counts (n = 111) were divided into five time periods: (1) incision to aortic cannulation; (2) aortic cannulation to cross-clamp onset; (3) cross-clamp onset to cross-clamp release; (4) cross-clamp release to decannulation; and (5) decannulation to chest closure. The level of S100beta (n = 156) was measured at baseline, at the end of cardiopulmonary bypass, then 150 and 270 minutes after cross-clamp release. RESULTS: The level of S100beta correlated with age, cardiopulmonary bypass time, cross-clamp time, and number of emboli at time period 2. Although cardiopulmonary bypass time was univariately associated with S100beta level, it became nonsignificant in a multivariable model that included age and cross-clamp time. CONCLUSIONS: The correlation of S100beta level with emboli measured during cannulation (time period 2) supports the hypothesis that cannulation is a high-risk time period for cerebral injury.     

A complication of T-fasteners in percutaneous endoscopic gastrostomy (PEG) placement

Two patients with sinus tracts from retained T-fasteners following PEG tube placement are reported. Both patients had the PEG tubes subsequently removed and presented with purulent discharge and granulations near well-healed gastrostomy sites. The management of this complication and a possible method of prevention are discussed.