Oligonucleotide-directed mutagenesis of psbB, the gene encoding CP47, employing a deletion mutant strain of the cyanobacterium Synechocystis sp. PCC 6803.
A mutant strain of the cyanobacterium Synechocystis sp. PCC (Pasteur Culture Collection) 6803 has been developed in which psbB, the gene coding for the chlorophyl alpha-binding protein CP47 in Photosystem II (PSII), has been deleted. This deletion mutant can be used for the reintroduction of modified psbB into the cyanobacterium. To study the role of a large hydrophilic region in CP47, presumably located on the lumenal side of the thylakoid membrane between the fifth and sixth membrane-spanning regions, specific deletions have been introduced in psbB coding for regions within this domain. One psbB mutation leads to deletion of Gly-351 to Thr-365 in CP47, another psbB mutation was targeted towards deletion of Arg-384 to Val-392 in this protein. The deletion from Gly-351 to Thr-365 results in a loss of PSII activity and of photoautotrophic growth of the mutant, but the deletion between Arg-384 and Val-392 retains PSII activity and the ability to grow photoautotrophically. The mutant strain with the deletion from Gly-351 to Thr-365 does not assemble a stable PSII reaction center complex in its thylakoid membranes, and exhibits diminished levels of CP47 and of the reaction center proteins D1 and D2. In contrast to the Arg-384 to Val-392 portion of this domain, the region between Gly-351 and Thr-365 appears essential for the normal structure and function of photosystem II.


Differentiating structural isomers of sialylated glycans by matrix-assisted laser desorption/ionization time-of-flight/time-of-flight tandem mass spectrometry.
Using model acidic glycans, we demonstrate the benefits of permethylation for matrix-assisted laser desorption/ionization time-of-flight/time-of-flight (MALDI/TOF-TOF) tandem mass spectrometry. With both the linear and branched structures, extensive cross-ring fragmentation product ions were generated, yielding valuable information on sugar linkages. Elimination of the negative charges commonly associated with sialylated structures through permethylation allowed their structural analysis in the positive ion mode. Extensive A- and X-type ions were observed for the linear structures, and slightly weaker signals for the branched sialylated structures. The diagnostic cross-ring fragments, permitting a distinction between alpha2-3 and alpha2-6 linkages of the sialic acid residues, were seen in abundance. Importantly, the cross-ring fragmentation with the branched structures provides adequate information to assign sialic acid residues, with a specific linkage, to a particular antenna.


Nonsteroidal anti-inflammatory drugs for pain in women with endometriosis.
Endometriosis is a common gynaecological condition that affects women and can lead to painful symptoms and infertility. It greatly affects women's quality of life, impacting their careers, everyday activities, sexual and nonsexual relationships and fertility. Nonsteroidal anti-inflammatory drugs (NSAIDs) are most commonly used as first-line treatment for women with pain associated with endometriosis. To assess effects of NSAIDs used for management of pain in women with endometriosis compared with placebo, other NSAIDs, other pain management drugs or no treatment. We searched the Cochrane Gynaecology and Fertility Group Specialised Register of Controlled Trials (October 2016), published in the Cochrane Central Register of Controlled Trials (CENTRAL) in the Cochrane Library, as well as MEDLINE (January 2008 to October 2016), Embase (date limited from 1 January 2016 to 19 October 2016, as all earlier references are included in CENTRAL output as a result of the Embase project), registers of ongoing trials and the reference lists of relevant publications. We identified no new randomised controlled trials. Unless we identify new evidence in the future, we will not update this review. We included all randomised controlled trials (RCTs) describing use of NSAIDs for management of pain associated with endometriosis in women of all ages. In the 2009 update of this review, two review authors (CA and SH) independently read and extracted data from each of the included studies. We analysed cross-over trials using the inverse variance method of RevMan to calculate odds ratios for binary outcomes. We identified no new trials for the 2016 update. This review includes two trials, but we included only one trial, with 24 women, in the analysis.The overall risk of bias was unclear owing to lack of methodological detail. Using the GRADE method, we judged the quality of the evidence to be very low. We downgraded evidence for risk of bias and for imprecision (wide confidence intervals and evidence based on a single small trial).Comparison of NSAIDs (naproxen) versus placebo revealed no evidence of a positive effect on pain relief (odds ratio (OR) 3.27, 95% confidence interval (CI) 0.61 to 17.69; one trial, 24 women; very low-quality evidence) in women with endometriosis. Evidence indicating whether women taking NSAIDs (naproxen) were less likely to require additional analgesia (OR 0.12, 95% CI 0.01 to 1.29; one trial, 24 women; very low-quality evidence) or to experience side effects (OR 0.46, 95% CI 0.09 to 2.47; one trial, 24 women; very low-quality evidence) when compared with placebo was inconclusive.Studies provided no data on quality of life, effects on daily activities, absence from work or school, need for more invasive treatment or participant satisfaction with treatment. Owing to lack of high-quality evidence and lack of reporting of outcomes of interest for this review, we can make no judgement as to whether NSAIDs (naproxen) are effective in managing pain caused by endometriosis. No evidence shows whether any individual NSAID is more effective than another. As shown in other Cochrane reviews, women taking NSAIDs must be aware that these drugs may cause unintended effects.


Extract of corn silk (stigma of Zea mays) inhibits the tumour necrosis factor-alpha- and bacterial lipopolysaccharide-induced cell adhesion and ICAM-1 expression.
Treatment of human endothelial cells with cytokines such as tumour necrosis factor-alpha (TNF) or E. coli lipopolysaccharide (LPS) induces the expression of several adhesion molecules and enhances leukocyte adhesion to endothelial cell surface. Interfering with this leukocyte adhesion or adhesion molecules upregulation is an important therapeutic target for the treatment of bacterial sepsis and various inflammatory diseases. In the course of screening marketed European anti-inflammatory herbal drugs for TNF antagonistic activity, a crude ethanolic extract of corn silk (stigma of Zea mays) exhibited significant activity. The extract at concentrations of 9-250 micrograms/ml effectively inhibited the TNF- and LPS-induced adhesiveness of EAhy 926 endothelial cells to monocytic U937 cells. Similar concentration ranges of corn silk extract did also block the TNF and LPS but not the phorbol 12-myristate 13-acetate-induced ICAM-1 expression on EAhy 926 endothelial cell surface. The extract did not alter the production of TNF by LPS-activated macrophages and failed to inhibit the cytotoxic activity of TNF. It is concluded that corn silk possesses important therapeutic potential for TNF- and LPS-mediated leukocyte adhesion and trafficking.


Characterization of supported membranes on topographically patterned polymeric elastomers and their applications to microcontact printing.
This article describes the fluorescence microscopy and imaging ellipsometry-based characterization of supported phospholipid bilayer formation on elastomeric substrates and its application in microcontact printing of spatially patterned phospholipid bilayers. Elastomeric stamps, displaying a uniformly spaced array of square wells (20, 50, and 100 mum linear dimensions), are prepared using poly(dimethyl)siloxane from photolithographically derived silicon masters. Exposing elastomeric stamps, following UV/ozone-induced oxidation, to a solution of small unilamellar phospholipid vesicles results in the formation of a 2D contiguous, fluid phospholipid bilayers. The bilayer covers both the elevated and depressed regions of the stamp and exhibits a lateral connectivity allowing molecular transport across the topographic boundaries. Applications of these bilayer-coated elastomeric stamps in microcontact printing of lipid bilayers reveal a fluid-tearing process wherein the bilayer in contact regions selectively transfers with 75-90% efficiency, leaving behind unperturbed patches in the depressed regions of the stamp. Next, using cholera-toxin binding fluid POPC bilayers that have been asymmetrically doped with ganglioside Gm1 ligand in the outer leaflets, we examine whether the microcontact transfer of bilayers results in the inversion of the lipid leaflets. Our results suggest a complex transfer process involving at least partial bilayer reorganization and molecular re-equilibration during (or upon) substrate transfer. Taken together, the study sheds light on the structuring of lipid inks on PDMS elastomers and provides clues regarding the mechanism of bilayer transfer. It further highlights some important differences in stamping fluid bilayers from the more routine applications of stamping in the creation of patterned self-assembled monolayers.


Alleviating sample imbalance in water quality assessment using the VAE-WGAN-GP model.
Water resources are essential for sustaining human life and promoting sustainable development. However, rapid urbanization and industrialization have resulted in a decline in freshwater availability. Effective prevention and control of water pollution are essential for ecological balance and human well-being. Water quality assessment is crucial for monitoring and managing water resources. Existing machine learning-based assessment methods tend to classify the results into the majority class, leading to inaccuracies in the outcomes due to the prevalent issue of imbalanced class sample distribution in practical scenarios. To tackle the issue, we propose a novel approach that utilizes the VAE-WGAN-GP model. The VAE-WGAN-GP model combines the encoding and decoding mechanisms of VAE with the adversarial learning of GAN. It generates synthetic samples that closely resemble real samples, effectively compensating data of the scarcity category in water quality evaluation. Our contributions include (1) introducing a deep generative model to alleviate the issue of imbalanced category samples in water quality assessment, (2) demonstrating the faster convergence speed and improved potential distribution learning ability of the proposed VAE-WGAN-GP model, (3) introducing the compensation degree concept and conducting comprehensive compensation experiments, resulting in a 9.7% increase in the accuracy of water quality assessment for multi-classification imbalance samples.


Expression of human myoglobin in H9c2 cells enhances toxicity to added hydrogen peroxide.
Hydrogen peroxide (H2O2) is implicated in cardiac myocyte (CM) damage during myocardial ischemia-reperfusion (IR) injury. Myoglobin (Mb) is present in CM at significant concentrations and reacts with H2O2 to yield one- and two-electron oxidants that may promote myocardial injury. Paradoxically, hearts from mice lacking Mb are more susceptible to H2O2-induced dysfunction than the corresponding controls [U. Flogel, A. Godecke, L.O. Klotz, J. Schrader, Role of myoglobin in the anti-oxidant defense of the heart, FASEB J. 18 (2004) 1156-1158]. We have overexpressed wild-type or Y103F variant of human Mb in cultured CMs to test whether Mb protects against H2O2 insult. Contrary to expectation, cells expressing WT or the Y103F Mb show increased mitochondrial dysfunction and apoptosis, and decreased ATP in response to H2O2 that follows the order native < Y103F Mb < WT human Mb consistent with the increasing pro-oxidant activity for these proteins. These data indicate that (i) Mb promotes oxidative damage to cultured CM and (ii) Mb may be a useful target for the design of inhibitors of myocardial IR injury.


Vector competence of northern and southern European Culex pipiens pipiens mosquitoes for West Nile virus across a gradient of temperatures.
In Europe, West Nile virus (WNV) outbreaks have been limited to southern and central European countries. However, competent mosquito vectors and susceptible bird hosts are present in northern Europe. Differences in temperature and vector competence of mosquito populations may explain the absence of WNV outbreaks in northern Europe. The aim of the present study was to directly compare vector competence of northern and southern European Culex pipiens (Cx. p.) pipiens mosquitoes for WNV across a gradient of temperatures. WNV infection and transmission rates were determined for two Cx. p. pipiens populations originating from The Netherlands and Italy, respectively. Mosquitoes were orally exposed by providing an infectious bloodmeal, or by injecting WNV (lineage 2) in the thorax, followed by 14-day incubation at 18, 23, or 28 °C. No differences in infection or transmission rates were found between the Cx. p. pipiens populations with both infection methods, but WNV transmission rates were significantly higher at temperatures above 18 °C. The absence of WNV outbreaks in northern Europe cannot be explained by differences in vector competence between Cx. p. pipiens populations originating from northern and southern Europe. This study suggests that low temperature is a key limiting factor for WNV transmission.


[Availability of essential medications at primary level units of the Health Department of Tamaulipas, Mexico].
To describe the availability of some essential drugs at the primary health care units of the Ministry of Health of Tamaulipas, Mexico. Between September and October 1998, all first level healthcare units of Tamaulipas' three sanitary jurisdictions were surveyed. Drug availability was assessed. The measurement instrument was a checklist of 56 drugs and 10 different supplies. For each drug and input the absolute number and the proportion of units with this drug or input was calculated. In the units where the drugs were available, the medians were calculated. The median of the total number of drugs available in all units was used as a global indicator. This same exercise was developed for each unit. Comparisons between the availability of these inputs in the units and stockrooms were also done. Stata 5.0 was used for statistical analysis. None of the inspected units had full availability of all checklist drugs. The highest percentage of drug availability was 84% and the lowest was 32%. There was limited availability of antibiotics, antihypertensive, hypoglycemic, and iron deficiency drugs. The availability of oral rehydration salts and contraceptive and vaccine agents was acceptable. Healthcare organizations must find alternative ways to improve access to drugs nationwide, in general, and availability of essential drugs in first level healthcare units, in particular. Two recent initiatives provide an optimistic outlook: decentralization of health services for the uninsured and the Generic Exchangeable Drugs Program, established nationwide in 1998.


Rate of hydrolysis in ATP synthase is fine-tuned by α-subunit motif controlling active site conformation.
Computer-designed artificial enzymes will require precise understanding of how conformation of active sites may control barrier heights of key transition states, including dependence on structure and dynamics at larger molecular scale. F(o)F(1) ATP synthase is interesting as a model system: a delicate molecular machine synthesizing or hydrolyzing ATP using a rotary motor. Isolated F(1) performs hydrolysis with a rate very sensitive to ATP concentration. Experimental and theoretical results show that, at low ATP concentrations, ATP is slowly hydrolyzed in the so-called tight binding site, whereas at higher concentrations, the binding of additional ATP molecules induces rotation of the central γ-subunit, thereby forcing the site to transform through subtle conformational changes into a loose binding site in which hydrolysis occurs faster. How the 1-Å-scale rearrangements are controlled is not yet fully understood. By a combination of theoretical approaches, we address how large macromolecular rearrangements may manipulate the active site and how the reaction rate changes with active site conformation. Simulations reveal that, in response to γ-subunit position, the active site conformation is fine-tuned mainly by small α-subunit changes. Quantum mechanics-based results confirm that the sub-Ångström gradual changes between tight and loose binding site structures dramatically alter the hydrolysis rate.


LIVER TRANSPLANTATION IN PATIENTS WITH PRIMARY SCLEROSING CHOLANGITIS: A MULTICENTRIC STUDY.
The prevalence of primary sclerosing cholangitis (PSC) in the general population has not yet been clearly established. The management of PSC should focus on delaying the progression of the disease and restraining its complications. The only curative therapy for the disease remains liver transplantation (LT). PSC is currently the fifth most common indication for LT and corresponds to 5% of all LT indications in adults. Our objective is to evaluate the indications and outcomes of PSC patients undergoing LT in three liver transplantation centers in southern Brazil - Hospital Santa Isabel in Blumenau, Santa Catarina state, and Hospital das Clínicas and Hospital Nossa Senhora das Graças, in Curitiba, Parana state). This is a longitudinal observational study of patients with PSC who underwent LT in three major Brazilian medical centers. Electronic medical records and study protocols of all patients subjected to LT from January 2011 to December 2021 were retrospectively reviewed. Of the 1,362 transplants performed in the three medical centers, 37 were due to PSC. Recurrence of PSC occurred in three patients (8.1%) in 3.0±2.4 years (range, 1-4 years). The 1-year and 5-year survival rates after the first LT were 83.8 and 80.6%, respectively. The 1-year and 5-year graft survival rates were, respectively, 83.8 and 74.8%. Our experience with LT in patients with PSC demonstrated good patient and graft survival results. Most deaths were due to common factors in patients undergoing LT.


Computational and Biochemical Discovery of RSK2 as a Novel Target for Epigallocatechin Gallate (EGCG).
The most active anticancer component in green tea is epigallocatechin-3-gallate (EGCG). Protein interaction with EGCG is a critical step for mediating the effects of EGCG on the regulation of various key molecules involved in signal transduction. By using computational docking screening methods for protein identification, we identified a serine/threonine kinase, 90-kDa ribosomal S6 kinase (RSK2), as a novel molecular target of EGCG. RSK2 includes two kinase catalytic domains in the N-terminal (NTD) and the C-terminal (CTD) and RSK2 full activation requires phosphorylation of both terminals. The computer prediction was confirmed by an in vitro kinase assay in which EGCG inhibited RSK2 activity in a dose-dependent manner. Pull-down assay results showed that EGCG could bind with RSK2 at both kinase catalytic domains in vitro and ex vivo. Furthermore, results of an ATP competition assay and a computer-docking model showed that EGCG binds with RSK2 in an ATP-dependent manner. In RSK2+/+ and RSK2-/- murine embryonic fibroblasts, EGCG decreased viability only in the presence of RSK2. EGCG also suppressed epidermal growth factor-induced neoplastic cell transformation by inhibiting phosphorylation of histone H3 at Ser10. Overall, these results indicate that RSK2 is a novel molecular target of EGCG.


Na intake, renal renin, and the severity of myohemoglobinuric renal failure in rats.
We have examined the role of dietary Na and renal cortical renin concentration (RCRC) on the severity of glycerol-induced acute renal failure (ARF) in the rat. Na loading (1% NaCl to drink) and Na deprivation (Na-free diet) of 5-6 wk duration decreased and increased RCRC, respectively, with respect to control. As assessed by blood urea nitrogen measured 48 h after glycerol injection, the severity of ARF was inversely related to RCRC. However, this was not a causal relationship. Either 6- or 48-h access to 1% NaCl did not lower RCRC of previously Na-deprived rats, but did decrease the severity of ARF. Conversely, 6- or 48-h access to tap water did not increase RCRC of previously Na-loaded rats, but did increase severity of ARF. A significant inverse correlation was found between severity of ARF and Na excretion prior to glycerol injection, a finding consistent with the hypothesis that solute diuresis has a protective effect.


CD74/SLC34A2-ROS1 fusion variants involving the transmembrane region predict poor response to crizotinib in non-small cell lung cancer independent of TP53 mutations.
Variable partners and breakpoints have been reported in ROS1-rearranged non-small cell lung cancer (NSCLC) patients. Here, we investigated the association of fusion partners and breakpoints with crizotinib efficacy in NSCLCs with common ROS1 fusions. DNA/RNA next-generation sequencing (NGS) and immunohistochemistry (IHC) were performed to characterize ROS1 fusions. Using DNA NGS, we identified ROS1 fusions in 210 cases, comprising 171 common (CD74/EZR/TPM3/SDC4/SLC34A2-ROS1) and 39 uncommon (variants identified in <5%) ROS1 fusion cases. DNA NGS detected variable ROS1 genomic breakpoints in common ROS1 fusions, whereas RNA NGS found ROS1 breakpoints mainly occurring in exons 32 and 34/35, resulting in long (exon 32) and short (exon 34/35) ROS1 fusions. ROS1 IHC showed that membranous and cytoplasmic staining was predominant in long ROS1 fusions, whereas cytoplasmic staining was predominant in short ROS1 fusions (P=0.006). For patients who received first-line crizotinib, median progression-free survival (mPFS) was lower in patients with long ROS1 fusions than those with short ROS1 fusions (8.0 versus 24.0 months; P=0.006). Moreover, mPFS for patients with and without TP53 mutations was 8.0 and 19.0 months, respectively (P=0.159); mPFS for patients with and without BIM deletion polymorphism was 5.0 and 22.0 months, respectively (P=0.003). When analyzing together with fusion partners, patients with long CD74/SLC34A2-ROS1 fusions showed shorter PFS than those with other ROS1, regardless of the presence or absence of TP53 mutations (P<0.001 and P=0.002, respectively). Long CD74/SLC34A2-ROS1 fusions, which retain transmembrane regions in ROS1 and fusion partners, are associated with poor response to crizotinib independent of TP53 mutations.


Assessment of rival males through the use of multiple sensory cues in the fruitfly Drosophila pseudoobscura.
Environments vary stochastically, and animals need to behave in ways that best fit the conditions in which they find themselves. The social environment is particularly variable, and responding appropriately to it can be vital for an animal's success. However, cues of social environment are not always reliable, and animals may need to balance accuracy against the risk of failing to respond if local conditions or interfering signals prevent them detecting a cue. Recent work has shown that many male Drosophila fruit flies respond to the presence of rival males, and that these responses increase their success in acquiring mates and fathering offspring. In Drosophila melanogaster males detect rivals using auditory, tactile and olfactory cues. However, males fail to respond to rivals if any two of these senses are not functioning: a single cue is not enough to produce a response. Here we examined cue use in the detection of rival males in a distantly related Drosophila species, D. pseudoobscura, where auditory, olfactory, tactile and visual cues were manipulated to assess the importance of each sensory cue singly and in combination. In contrast to D. melanogaster, male D. pseudoobscura require intact olfactory and tactile cues to respond to rivals. Visual cues were not important for detecting rival D. pseudoobscura, while results on auditory cues appeared puzzling. This difference in cue use in two species in the same genus suggests that cue use is evolutionarily labile, and may evolve in response to ecological or life history differences between species.


Arginine-vasopressin activates the JAK-STAT pathway in vascular smooth muscle cells.
JAK (Janus-activated kinase)-STAT (signal transducers and activators of transcription) signaling is a major signal transduction pathway in mammalian cells. Different growth factors and cytokines were reported as activators of the JAK-STAT pathway in various cell types. Interestingly, arginine-vasopressin (AVP) was never reported as an inducer of the JAK-STAT pathway. In the present study, we show for the first time that AVP stimulation of vascular smooth muscle cells (VSMCs) induces STAT3 tyrosine and serine phosphorylation, followed by nuclear translocation of the phosphorylated STAT3. In addition, we found that AVP induced JAK2 tyrosine phosphorylation. Taken together, these results demonstrate that AVP activates the JAK-STAT pathway in VSMCs. Furthermore, our results indicate that AVP-induced STAT3 tyrosine phosphorylation requires both JAK2 and c-Src tyrosine kinases. The present study also implicates that extracellular signal-regulated kinase (ERK1/2), which are serine/threonine kinases, are the mediators of STAT3 serine phosphorylation upon AVP stimulation. We further suggest that AVP-induced STAT3 serine phosphorylation negatively modulates AVP-induced STAT3 tyrosine phosphorylation. Finally, our results implicate a novel role for the JAK-STAT pathway, mediating AVP-induced VSMC hypertrophy.


Gold Nanoparticles as Colorimetric Sensors for the Detection of DNA Bases and Related Compounds.
Results regarding interaction of colloidal gold solutions with nucleobases, including uracil (U), as well as its sulfur derivatives, 2-thiouracil (2TU) and 4-thiouracil (4TU), cytosine (C), adenine (A), and guanine (G), as well as urea and thiourea (TU), are reported. Anionic stabilized citrate gold nanoparticles (AuNPs) were synthesized by reducing the tetrachloroaurate (III) trihydrate with trisodium citrate. The surface plasmon resonance (SPR) band was used in the characterization of synthesized AuNPs, as well as transmission electron microscope (TEM) imaging, which was used in the characterization of dispersed and aggregated gold nanoparticles. Interactions of nucleobases with the gold surface was analyzed by following the plasmon absorbance band red shift of the AuNPs. The sulfur-containing compounds adsorbed to the nanoparticle surfaces by chemisorption-type interactions; with TU and 4TU, the process is accompanied by a sudden change in color; in contrast, 2TU forms stable functionalized gold nanoparticles. Urea and U do not adsorb to nanoparticle surfaces, but the other heterocyclic bases containing nitrogen interact effectively with the gold surface, causing the assembly of nanoparticles, even though the interparticle self-aggregation process was slower than that mediated by either TU or 4TU. The method is efficient in the colorimetric detection of nucleobases and derivatives at concentration levels on the order of 1 µM.


Laser-assisted in-situ keratomileusis (LASIK) with a mechanical microkeratome compared to LASIK with a femtosecond laser for LASIK in adults with myopia or myopic astigmatism.
Laser-assisted in-situ keratomileusis (LASIK) is a surgical procedure that corrects refractive errors. This technique creates a flap of the outermost parts of the cornea (epithelium, bowman layer, and anterior stroma) to expose the middle part of the cornea (stromal bed) and reshape it with excimer laser using photoablation. The flaps can be created by a mechanical microkeratome or a femtosecond laser. To compare the effectiveness and safety of mechanical microkeratome versus femtosecond laser in LASIK for adults with myopia. We searched CENTRAL (which contains the Cochrane Eyes and Vision Trials Register) (2019, Issue 2); Ovid MEDLINE; Embase; PubMed; LILACS; ClinicalTrials.gov and the World Health Organization (WHO) International Clinical Trials Registry Platform (ICTRP). We used no date or language restrictions. We searched the reference lists of included trials. We searched the electronic databases on 22 February 2019. We included randomized controlled trials (RCTs) of LASIK with a mechanical microkeratome compared to a femtosecond laser in people aged 18 years or older with more than 0.5 diopters of myopia or myopic astigmatism. We used standard methodological procedures expected by Cochrane. We included 16 records from 11 trials enrolling 943 adults (1691 eyes) with spherical or spherocylindrical myopia, who were suitable candidates for LASIK. Five hundred and forty-seven participants (824 eyes) received LASIK with a mechanical microkeratome and 588 participants (867 eyes) with a femtosecond laser. Each trial included between nine and 360 participants. In six trials, the same participants received both interventions. Overall, the trials were at an uncertain risk of bias for most domains. At 12 months, data from one trial (42 eyes) indicates no difference in the mean uncorrected visual acuity (logMAR scale) between LASIK with a mechanical microkeratome and LASIK with a femtosecond laser (mean difference (MD) -0.01, 95% confidence interval (CI) -0.06 to 0.04; low-certainty evidence). Similar findings were observed at 12 months after surgery, regarding participants achieving 0.5 diopters within target refraction (risk ratio (RR) 0.97, 95% CI 0.85 to 1.11; 1 trial, 79 eyes; low-certainty evidence) as well as mean spherical equivalent of the refractive error 12 months after surgery (MD 0.09, 95% CI -0.01 to 0.19; 3 trials, 168 eyes [92 participants]; low-certainty evidence). Based on data from three trials (134 eyes, 66 participants), mechanical microkeratome was associated with lower risk of diffuse lamellar keratitis compared with femtosecond laser (RR 0.27, 95% CI 0.10 to 0.78; low-certainty evidence). Thus, diffuse lamellar keratitis was a more common adverse event with femtosecond laser than with mechanical microkeratome, decreasing from an assumed rate of 209 per 1000 people in the femtosecond laser group to 56 per 1000 people in the mechanical microkeratome group. Data from one trial (183 eyes, 183 participants) indicates that dry eye as an adverse event may be more common with mechanical microkeratome than with femtosecond laser, increasing from an assumed rate of 80 per 1000 people in the femtosecond laser group to 457 per 1000 people in the mechanical microkeratome group (RR 5.74, 95% CI 2.92 to 11.29; low-certainty evidence). There was no evidence of a difference between the two groups for corneal haze (RR 0.33, 95% CI 0.01 to 7.96; 1 trial, 43 eyes) and epithelial ingrowth (RR 1.04, 95% CI 0.11 to 9.42; 2 trials, 102 eyes [50 participants]). The certainty of evidence for both outcomes was very low. Regarding the visual acuity outcomes, there may be no difference between LASIK with mechanical microkeratome and LASIK with femtosecond laser. Dry eye and diffuse lamellar keratitis are likely adverse events with mechanical microkeratome and femtosecond laser, respectively. The evidence is uncertain regarding corneal haze and epithelial ingrowth as adverse events of each intervention. The limited number of outcomes reported in the included trials, some with potentially significant risk of bias, makes it difficult to draw a firm conclusion regarding the effectiveness and safety of the interventions investigated in this review.


Changes in serum potassium and blood glucose concentrations after trimethaphan administration in man.
Blood glucose and serum potassium (K+) concentrations were measured before, during, and 60 minutes after operation in two groups of 10 patients during nitrous oxide/halothane/d-tubocurarine anesthesia for major orthopedic surgery. In the control group, arterial blood pressure was maintained within normal range, while in the study group trimethaphan camsylate was administered as an intravenous infusion (average, 218 mg.) to maintain a systolic blood pressure of 60 to 65 torr. In the normotensive group, blood glucose rose significantly during operation and early postoperatively and serum K+ was essentially unchanged. In the hypotensive group, trimethaphan caused a striking modification of surgically induced hyperglycemia, together with a small significant decrease in serum K+ intraoperatively. The observed increase in blood glucose is part of the autonomic response to surgical stress. Hormonal factors (growth hormone, cortisol and glucagon) may conceivably be involved. The decrease in serum K+ is probably caused by decreased hepatic glycogenolysis and attenuation of the suppressive effect of catecholamines on insulin release, both effects being secondary to the ganglionic blocking property of trimethaphan. These results indicate that trimethaphan, in contrast to other ganglionic blocking drugs, does not cause hypoglycemia and suggest that serum K+ concentration should be monitored whenever these drugs are used.


Immunotherapy in gastrointestinal cancers.
Gastrointestinal (GI) cancers such as gastric, esophageal, pancreas, hepatobiliary, colorectal and anal cancers are a major cause of cancer related mortality worldwide. Traditional treatment options such as chemotherapy, surgery, radiation therapy, monoclonal antibodies and anti-angiogenic agents have been the backbone of treatment of GI cancers in various stages. Current cancer research is moving forward to incorporate immunotherapies in the treatment of GI cancers either as single agent or in combination with current available treatment modalities. This review summarizes the existing and ongoing immunotherapies in the treatment of GI cancers.


Effect of hindered internal rotation treatments on predicting the thermodynamic properties of alkanes.
When considering hindered internal rotation, we usually have several options, including (i) single structure harmonic oscillator (SS-HO) approximation that considers the lowest-energy conformer only and approximates all molecular vibrations as harmonic oscillations, (ii) one-dimensional (1-D) internal rotation treatment that replaces the corresponding vibrational mode with one-dimensional torsion, and (iii) the multistructural method with torsional anharmonicity (MS-T) that considers the multiple-structure and torsional anharmonicity. These methods differ greatly in computational cost and accuracy. To evaluate the effect of different treatments on predicting thermodynamic properties, we calculated enthalpy, entropy, and heat capacity for a series of normal and branched alkanes using six different methods, including the SS-HO treatment, three 1-D methods, the MS-T method, and the group additivity (GA) method. The comparison of the computational results with experimental data shows that GA and two 1-D methods proposed in this study are more suitable for reliable and rapid predictions of thermodynamic properties for large hydrocarbons with many carbon-carbon single bonds.


Analytical Guidelines for co-fractionation Mass Spectrometry Obtained through Global Profiling of Gold Standard Saccharomyces cerevisiae Protein Complexes.
Co-fractionation MS (CF-MS) is a technique with potential to characterize endogenous and unmanipulated protein complexes on an unprecedented scale. However this potential has been offset by a lack of guidelines for best-practice CF-MS data collection and analysis. To obtain such guidelines, this study thoroughly evaluates novel and published Saccharomyces cerevisiae CF-MS data sets using very high proteome coverage libraries of yeast gold standard complexes. A new method for identifying gold standard complexes in CF-MS data, Reference Complex Profiling, and the Extending 'Guilt-by-Association' by Degree (EGAD) R package are used for these evaluations, which are verified with concurrent analyses of published human data. By evaluating data collection designs, which involve fractionation of cell lysates, it is found that near-maximum recall of complexes can be achieved with fewer samples than published studies. Distributing sample collection across orthogonal fractionation methods, rather than a single high resolution data set, leads to particularly efficient recall. By evaluating 17 different similarity scoring metrics, which are central to CF-MS data analysis, it is found that two metrics rarely used in past CF-MS studies - Spearman and Kendall correlations - and the recently introduced Co-apex metric frequently maximize recall, whereas a popular metric-Euclidean distance-delivers poor recall. The common practice of integrating external genomic data into CF-MS data analysis is also evaluated, revealing that this practice may improve the precision and recall of known complexes but is generally unsuitable for predicting novel complexes in model organisms. If studying nonmodel organisms using orthologous genomic data, it is found that particular subsets of fractionation profiles (e.g. the lowest abundance quartile) should be excluded to minimize false discovery. These assessments are summarized in a series of universally applicable guidelines for precise, sensitive and efficient CF-MS studies of known complexes, and effective predictions of novel complexes for orthogonal experimental validation.


[Prevalence of optic atrophy and associated ocular and systemic diseases in a department of paediatric ophthalmology].
Optic atrophy is one of the most common causes of severe visual impairment in children. So far an analysis of ocular and systemic findings comparing patients with and without optic atrophy has not performed. Ocular and systemic findings of a total of 1042 patients (examination of all patients in May 1995 [N = 485] and May 2001 [N = 557]) of the department of paediatric ophthalmology in Homburg/Saar were retrospectively evaluated and diagnoses of patients with and without optic atrophy were compared. Optic atrophy was diagnosed ophthalmoscopically and in 1/3rd of the patients by VEP as well. 18 % of all patients (N = 188; 87.2 % [N = 164] were children) had optic atrophy. Nearly half of these children were prematurely born (46.7 %). 53.2 % of patients with optic atrophy (N = 88) showed nystagmus (without atrophy: 10.7 %), especially sensory defect nystagmus. Median of visual acuity level was 0.2 with optic atrophy and 0.8 without. Albinism and buphthalmia were common findings. 69.5 % of all patients with optic atrophy suffered from systemic diseases (without atrophy: 25.2 %), especially mental retardation, neurologic findings and oculocutaneous albinism. In 10.4 % more than two systemic diagnoses could be found. 55.3 % of all the patients with optic atrophy were disabled, 31.9 % multiply disabled. Sequelae of prematurity, peripartal asphyxia and congenital brain damages are the main findings in patients with optic nerve atrophy. Such children are worst-case patients of any paediatric ophthalmology department with a high prevalence of severe visual impairment, mental retardation and multiple disability. Treatment in a specialised center is therefore necessary for an efficient therapy. In addition, the statistical survey shows that in children who survived the critical phases of prematurity secondary damages can lead to persistent impairment and to alterations of visual and general development.


Hormonal and metabolic response to physical exercise in hyperinsulinemic and non-hyperinsulinemic type 2 diabetics.
In order to find out the influence of hyperinsulinaemia and initial blood glucose levels on glucose homeostasis during physical exercise, 6 Type 2 diabetic patients with basal hyperinsulinemia (0.209 nmol/l) (group A) and 10 Type 2 diabetics without basal hyperinsulinemia (0.046 nmol/l) (group B) took part in a study on metabolic effects of exercise. Mean bodymass was higher in group A (101 kg) than in group B (71.7 kg). Exercise was performed on a bicycle-ergometer for 1 hr. Work load was adjusted to a pulse-rate of 120/min with a mean of 48 watt (W) in group A and 52 W in group B. Blood glucose (BG), insulin (IRI), glucagon (G), growth hormone (HGH), cortisol (C), epinephrine (E), norepinephrine (NE), lactate (L), pyruvate (P) and free fatty acids (NEFA) were measured during 3 hr. BG and IRI were also documented for the following 7 hr. Both groups showed a small but significant decrease of BG during exercise (group A from 11.54-10.38 mmol/l, p less than 0.01, and group B from 8.71-7.22 mmol/l, p less than 0.005). IRI decreased insignificantly in group A (from 0.209-0.174 nmol/l, p less than 0.15) and significantly in group B (from 0.046-0.032 nmol/l, p less than 0.01). G increased significantly in both groups (from 62.4-75.2 pmol/l in A, p less than 0.05, and from 38.8-47.1 pmol/l in B, p less than 0.01). HGH rose from 0.018-0.149 nmol/l in A, p less than 0.01, and from 0.077-0.320 nmol/l in B, p less than 0.01.(ABSTRACT TRUNCATED AT 250 WORDS)


Knowledge Graph Based Hard Drive Failure Prediction.
The hard drive is one of the important components of a computing system, and its failure can lead to both system failure and data loss. Therefore, the reliability of a hard drive is very important. Realising this importance, a number of studies have been conducted and many are still ongoing to improve hard drive failure prediction. Most of those studies rely solely on machine learning, and a few others on semantic technology. The studies based on machine learning, despite promising results, lack context-awareness such as how failures are related or what other factors, such as humidity, influence the failure of hard drives. Semantic technology, on the other hand, by means of ontologies and knowledge graphs (KGs), is able to provide the context-awareness that machine learning-based studies lack. However, the studies based on semantic technology lack the advantages of machine learning, such as the ability to learn a pattern and make predictions based on learned patterns. Therefore, in this paper, leveraging the benefits of both machine learning (ML) and semantic technology, we present our study, knowledge graph-based hard drive failure prediction. The experimental results demonstrate that our proposed method achieves higher accuracy in comparison to the current state of the art.


Thrombotic microangiopathy in primary antiphospholipid syndrome is linked to stroke and less deep venous thrombosis.
To compare clinical and laboratory data obtained from patients with primary Antiphospholipid Syndrome (pAPS) with and without Thrombotic Microangiopathy (TMA). A cross-sectional study with 66 (83.3% female) pAPS patients was performed. Demographic, clinical, drug use, antiphospholipid antibodies data were evaluated. Patients were subdivided into one of two groups: pAPS with TMA and pAPS without TMA and were compared. In this sample, 5/66 (7.6%) of patients had TMA. Primary APS with TMA group exhibited a higher frequency of arterial events (100% vs. 54.1%, p=0.02), stroke (100% vs. 32.8%, p=0.001) and a lower frequency of deep venous thrombosis (0 vs. 68.9%, p=0.0009) compared to the patients without TMA. Analysis of therapy used in these patients showed a higher frequency of current (40% vs. 6.6%, p=0.0006) and previous glucocorticoid use (80% vs. 36%, p=0.0007) and statin use (50% vs. 22.9%, p=0.037) in the first group. The two groups exhibited no differences in the frequency of positive autoantibodies, except for higher IgG anticardiolipin titers (86 ± 52 vs. 34.5 ± 39 GPL, p=0.003). Patients with pAPS and TMA have distinct clinical and laboratory spectra from those without TMA, that is characterized by an increased frequency of arterial events, stroke, and higher titers of IgG anticardiolipin; they have deep venous thrombosis less frequently.


An immunohistochemical study of perianal Paget's disease. Possible origins and clinical implications.
The histogenesis of perianal Paget's disease is controversial. A clinical and pathologic study was done of a patient with a history of adenocarcinoma of the rectum for whom a subsequent diagnosis of perianal Paget's disease was the sole manifestation of recurrent rectal cancer. Immunohistochemical techniques were used to compare and contrast the original rectal adenocarcinoma with the subsequent perianal skin recurrence confined to the epidermis. Both the rectal adenocarcinoma and the Paget's cells were positive for cytokeratin, epithelial membrane antigen, B72.3, and carcinoembryonic antigen and negative for gross cystic disease fluid protein-15, Leu-M1, CA 125, and S-100 protein. These findings, their relevance to the histogenesis of perianal Paget's disease, and the possible clinical implications are discussed.


Innate defenses of the intestinal epithelial barrier.
The innate immune system plays a crucial role in maintaining the integrity of the intestine and protecting the host against a vast number of potential microbial pathogens from resident and transient gut microflora. Mucosal epithelial cells and Paneth cells produce a variety of antimicrobial peptides (defensins, cathelicidins, crytdinrelated sequence peptides, bactericidal/permeabilityincreasing protein, chemokine CCL20) and bacteriolytic enzymes (lysozyme, group IIA phospholipase A2) that protect mucosal surfaces and crypts containing intestinal stem cells against invading microbes. Many of the intestinal antimicrobial molecules have additional roles of attracting leukocytes, alarming the adaptive immune system or neutralizing proinflammatory bacterial molecules. Dysfunction of components of the innate immune system has recently been implicated in chronic inflammatory bowel diseases such as Crohn's disease and ulcerative colitis, illustrating the pivotal role of innate immunity in maintaining the delicate balance between immune tolerance and immune response in the gut.


Comparative genomic analysis and evolution of the T cell receptor loci in the opossum Monodelphis domestica.
All jawed-vertebrates have four T cell receptor (TCR) chains: alpha (TRA), beta (TRB), gamma (TRG) and delta (TRD). Marsupials appear unique by having an additional TCR: mu (TRM). The evolutionary origin of TRM and its relationship to other TCR remain obscure, and is confounded by previous results that support TRM being a hybrid between a TCR and immunoglobulin locus. The availability of the first marsupial genome sequence allows investigation of these evolutionary relationships. The organization of the conventional TCR loci, encoding the TRA, TRB, TRG and TRD chains, in the opossum Monodelphis domestica are highly conserved with and of similar complexity to that of eutherians (placental mammals). There is a high degree of conserved synteny in the genomic regions encoding the conventional TCR across mammals and birds. In contrast the chromosomal region containing TRM is not well conserved across mammals. None of the conventional TCR loci contain variable region gene segments with homology to those found in TRM; rather TRM variable genes are most similar to that of immunoglobulin heavy chain genes. Complete genomic analyses of the opossum TCR loci continue to support an origin of TRM as a hybrid between a TCR and immunoglobulin locus. None of the conventional TCR loci contain evidence that such a recombination event occurred, rather they demonstrate a high degree of stability across distantly related mammals. TRM, therefore, appears to be derived from receptor genes no longer extant in placental mammals. These analyses provide the first genomic scale structural detail of marsupial TCR genes, a lineage of mammals used as models of early development and human disease.


Unusual foreign bodies in the orofacial region.
Foreign bodies may be deposited in the oral cavity either by traumatic injury or iatrogenically. Among the commonly encountered iatrogenic foreign bodies are restorative materials like amalgam, obturation materials, broken instruments, needles, and so forth. The discovery of foreign bodies in the teeth is a special situation, which is often diagnosed accidentally. Detailed case history, clinical and radiographic examinations are necessary to come to a conclusion about the nature, size, location of the foreign body, and the difficulty involved in its retrieval. It is more common to find this situation in children as it is a well-known fact that children often tend to have the habit of placing foreign objects in the mouth. Sometimes the foreign objects get stuck in the root canals of the teeth, which the children do not reveal to their parents due to fear. These foreign objects may act as a potential source of infection and may later lead to a painful condition. This paper discusses the presence of unusual foreign bodies-a tip of the metallic compass, stapler pin, copper strip, and a broken sewing needle impregnated in the gingiva and their management.


Phase II trial of short-course R-CHOP followed by 90Y-ibritumomab tiuxetan in previously untreated high-risk elderly diffuse large B-cell lymphoma patients.
This study aimed to evaluate the efficacy and safety of the treatment with (90)Y-ibritumomab tiuxetan following a short-course of rituximab with cyclophosphamide-adriamycin-vincristine-prednisone (R-CHOP) in high-risk elderly patients with previously untreated diffuse large B-cell lymphoma (DLBCL). From December 2006 to October 2008, 55 high-risk elderly (age > or =60 years) untreated DLBCL patients were treated in seven Italian institutions with a short-course of chemotherapy consisting of four cycles of R-CHOP21 followed by (90)Y-ibritumomab tiuxetan 6 to 10 weeks later. Of the 55 patients, 48 underwent radioimmunotherapy. The overall response rate to the entire treatment regimen was 80%, including 73% complete remissions and 7% partial remissions. Eight (50%) of the 16 patients who achieved less than a complete response with CHOP improved their remission status after (90)Y-ibritumomab tiuxetan administration. With a median follow-up of 18 months, the 2-year progression-free survival was estimated to be 85%, with a 2-year overall survival of 86%. (90)Y-ibritumomab tiuxetan toxicity consisted of grade 3 to 4 hematologic toxicity in 28 of 48 patients, mainly neutropenia (23 patients) and thrombocytopenia (15 patients). Red cells and/or platelets transfusions were given to three patients. This study evaluated the feasibility, efficacy, and safety of a short-course R-CHOP21 regimen followed by (90)Y-ibritumomab tiuxetan in high-risk elderly DLBCL patients.


Incidence of Stroke and Transient Ischemic Attack in Croatia: A Population Based Study.
The aim of this population based neuroepidemiological study was to establish the real incidence rates of acute cerebrovascular disease (CVD): stroke and transient ischemic attack (TIA) in the Republic of Croatia. Multicentric study included 89 501 persons of all ages in four regional centres in Croatia: Zagreb, Osijek + Slavonski Brod, Rijeka and Split. The following incidence rates of stroke, expressed at population of 100 000, have been established: Zagreb 290.52, Osijek + Slavonski Brod 302.14, Rijeka 219.65, Split 195.82. Incidence rate of stroke for the Republic of Croatia is 251.39. The following incidence rates of TIA, expressed at population of 100,000, have been established: Zagreb 87.15, Osijek + Slavonski Brod 156.53, Rijeka 90.11, Split 59.10. Incidence rate of TIA for the Republic of Croatia is 100.55. In the continental part of Croatia (Zagreb, Osijek + Slavonski Brod) incidence rate of stroke is higher by 45%, while incidence rate of TIA is higher by 82% than in the coastal part of Croatia, probably due to different lifestyle and environmental factors. The study has shown relatively high incidence rates of acute CVD (stroke and TIA) in the Republic of Croatia, which proves that CVD are a great public health problem.


A diagnostic procedure to detect departures from local independence in item response theory models.
Item response theory (IRT) is a widely used measurement model. When considering its use in education, health outcomes, and psychology, it is likely to be one of the most impactful psychometric models in existence. IRT has many advantages over classical test theory-based measurement models. For these advantages to hold in practice, strong assumptions must be satisfied. One of these assumptions, local independence, is the focus of the work described here. Local independence is the assumption that, conditional on the latent variable(s), item responses are unrelated to one another (i.e., independent). Stated another way, local independence implies that the only thing causing items to covary is the modeled latent variable(s). Violations of this assumption, quite aptly titled local dependence, can have serious consequences for the estimated parameters. A new diagnostic is proposed, based on parameter stability in an item-level jackknife resampling procedure. We review the ideas underlying the new diagnostic and how it is computed before covering some simulated and real examples demonstrating its effectiveness. (PsycINFO Database Record


Guanine quadruplexes in the RNA genome of the tick-borne encephalitis virus: their role as a new antiviral target and in virus biology.
We have identified seven putative guanine quadruplexes (G4) in the RNA genome of tick-borne encephalitis virus (TBEV), a flavivirus causing thousands of human infections and numerous deaths every year. The formation of G4s was confirmed by biophysical methods on synthetic oligonucleotides derived from the predicted TBEV sequences. TBEV-5, located at the NS4b/NS5 boundary and conserved among all known flaviviruses, was tested along with its mutated variants for interactions with a panel of known G4 ligands, for the ability to affect RNA synthesis by the flaviviral RNA-dependent RNA polymerase (RdRp) and for effects on TBEV replication fitness in cells. G4-stabilizing TBEV-5 mutations strongly inhibited RdRp RNA synthesis and exhibited substantially reduced replication fitness, different plaque morphology and increased sensitivity to G4-binding ligands in cell-based systems. In contrast, strongly destabilizing TBEV-5 G4 mutations caused rapid reversion to the wild-type genotype. Our results suggest that there is a threshold of stability for G4 sequences in the TBEV genome, with any deviation resulting in either dramatic changes in viral phenotype or a rapid return to this optimal level of G4 stability. The data indicate that G4s are critical elements for efficient TBEV replication and are suitable targets to tackle TBEV infection.


Growth rate control of flagellar assembly in Escherichia coli strain RP437.
The flagellum is a rotary motor that enables bacteria to swim in liquids and swarm over surfaces. Numerous global regulators control flagellar assembly in response to cellular and environmental factors. Previous studies have also shown that flagellar assembly is affected by the growth-rate of the cell. However, a systematic study has not yet been described under controlled growth conditions. Here, we investigated the effect of growth rate on flagellar assembly in Escherichia coli using steady-state chemostat cultures where we could precisely control the cell growth-rate. Our results demonstrate that flagellar abundance correlates with growth rate, where faster growing cells produce more flagella. They also demonstrate that this growth-rate dependent control occurs through the expression of the flagellar master regulator, FlhD4C2. Collectively, our results demonstrate that motility is intimately coupled to the growth-rate of the cell.


Minimal invasive surgery for atrial fibrillation: an updated review.
Despite its proven efficacy, the Cox-Maze III procedure did not gain widespread acceptance for the treatment of stand-alone atrial fibrillation (SA-AF) because of its complexity and technical difficulty. Surgical ablation for SA-AF can now be successfully performed utilizing minimally invasive surgery (MIS). This study provides an overview of state-of-the-art MIS for the treatment of SA-AF. Studies selected for this review were identified on PUBMED and exclusion and inclusion criteria were applied to select the publication to be screened. Twenty-eight studies were included; 27 (96.4%) were observational in nature whereas 1 was prospective non-randomized. The total number of patients was 1051 (range 14-114). Mean age ranged from 45.3 to 67.1 years. Suboptimal results were obtained when employing microwave and high focused ultrasound energies. In contrast, MIS ablation of SA-AF achieved satisfactory 1-year results when the bipolar radiofrequency was employed as energy source, with antiarrhythmic drug-free success rate comparable to percutaneous catheter ablation (PCA). The success rate in paroxysmal was even higher than in PCA. In contrast, ganglionated plexi ablation and left atrial appendage removal seem not to influence the recurrence of AF and the occurrence of postoperative thromboembolic events. Minimally invasive surgery ablation of SA-AF achieved satisfactory 1-year results when the bipolar radiofrequency was employed. Nevertheless, the relatively high complication rate reported suggest that such techniques require further refinement. Finally, the preliminary results of the hybrid approach are promising but they need to be confirmed.


Predictive Factors and ACE-2 Gene Polymorphisms in Susceptibility to Long COVID-19 Syndrome.
Long COVID-19 syndrome is present in 5-10% of patients infected with SARS-CoV-2, and there is still little information on the predisposing factors that lead to its development. The purpose of the study was to evaluate the predictive factors in early symptoms, clinical features and the role of Angiotensin-Converting Enzyme-2 (ACE-2) c.513-1451G>A (rs2106806) and c.15643279T>C (rs6629110) polymorphisms in the susceptibility to developing Long COVID-19 syndrome subsequent to COVID-19 infectionA total of 29 patients who suffered COVID-19 were recruited in a descriptive longitudinal study of two groups: Long COVID-19 (n = 16) and non-Long COVID-19 (n = 13). Early symptoms and clinical features during COVID-19 were classified by a medical service. ACE-2 polymorphisms were genotyped by using a Single Nucleotide Primer Extension (SNPE). Of the early symptoms, fatigue, myalgia and headache showed a high risk of increasing Long COVID-19 susceptibility. Clinical features such as emergency care, SARS-CoV-2 reinfection, previous diseases, respiratory disease and brain fog also had a high risk of increasing Long COVID-19 susceptibility. The A allele in the rs2106806 variant was associated with an odds ratio (OR) of 4.214 (95% CI 2.521-8.853; p < 0.001), and the T allele in the rs6629110 variant was associated with an OR of 3.754 (95% CI 1.785-6.105; p = 0.002) of increasing Long COVID-19 susceptibility. This study shows the risk of ACE-2 polymorphisms, different early symptoms and clinical features during SARS-CoV-2 infection in susceptibility to Long COVID-19.


[The influence of radiation of the child mental development].
The paper presents the results of implementation of the first stage of WHO programme concerning consequences of the Chernobyl accident in terms of influence of radiation on mental health of children whose mothers were exposed to radiation. 4210 children born in the period from April 26, 1986 to February 1, 1987 in Byelorussia, Ukraine, Russia in contaminated and non-polluted (control) regions. In radiation-contaminated regions there was higher frequency of mental retardation and other mental disorders as well as deterioration of both mental and physic health of their parents. A preliminary character of the data obtained is emphasized as well as the necessity of more detailed follow-up of these groups.


Pharmacokinetics and safety of epinephrine use in liposuction.
Patients are routinely exposed to high-dose epinephrine infiltration during large-volume liposuction. Because of the serious cardiovascular side-effect profile of catecholamine overdose, the authors examined the safety of larger-volume liposuction by assessing epinephrine pharmacokinetics. Five female volunteers with American Society of Anesthesiologists physical status of I or II, aged 29 to 40 years and weighing 75.9 to 95 kg, underwent liposuction. The wetting solution contained 7.3 mg (SEM, 0.7 mg) of epinephrine, corresponding to 0.09 mg/kg (0.04 mg/kg). Total plasma epinephrine and norepinephrine concentrations were assessed by high-performance liquid chromatography. Approximate exogenous epinephrine absorption was calculated after correction for estimated endogenous epinephrine production. Pharmacokinetic assessments were performed using standard equations. The total plasma epinephrine peak occurred at the final intraoperative reading (5 hours after induction) and was 323 pg/ml (24.8 pg/ml), three to four times maximum baseline resting levels. The norepinephrine level was slightly elevated throughout the study period, with a reversal of the normal epinephrine/norepinephrine ratio (<0.5:1) demonstrated intraoperatively (>5:1). Estimated time to peak exogenous epinephrine level ranged from 1 to 4 hours from the start of infiltration. Area under the plasma concentration versus time curve was approximately 2089 to 2610 pg x hour/ml. Peak exogenous epinephrine concentration was estimated to be 286 to 335 pg/ml. Clearance was 764,508 ml/hour and volume of distribution was 0.4 liter/kg (0.006 liter/kg). Total absorbed epinephrine was estimated, 1.8 mg to 2.2 mg, equivalent to 25 to 32 percent of the infiltrated dose. The reversal of the normal epinephrine/norepinephrine ratio and the fact that norepinephrine levels were within normal range implied that the majority of plasma epinephrine measured was exogenously infiltrated and not endogenously synthesized. On the basis of these observations, pharmacokinetic analyses were performed. Although unequivocal toxic epinephrine levels were not demonstrated, epinephrine peaks were three to four times the maximum observed in normal resting patients. Peak levels were comparable to those observed during major physiologic stresses, such as exercising to exhaustion, open abdominal surgery, or cross-clamping the aorta during surgical repair. Furthermore, epinephrine has been associated with myocardial infarction, arrhythmias, and fatal asystole in susceptible patients at these levels. Patients should be carefully screened for clinical evidence of hemodynamic and cardiac pathology before larger-volume liposuction is undertaken, as it may result in unnecessary high risk for patients who have preexisting cardiovascular disorders. Healthy American Society of Anesthesiologists physical status I or II patients should have sufficient cardiac reserve to tolerate these catecholamine levels.


High-performance variants of plant diacylglycerol acyltransferase 1 generated by directed evolution provide insights into structure function.
Diacylglycerol acyltransferase 1 (DGAT1) catalyzes the acyl-CoA-dependent biosynthesis of triacylglycerol, the predominant component of seed oil. In some oil crops, including Brassica napus, the level of DGAT1 activity can have a substantial effect on triacylglycerol production. Structure-function insights into DGAT1, however, remain limited because of the lack of a three-dimensional detailed structure for this membrane-bound enzyme. In this study, the amino acid residues governing B. napus DGAT1 (BnaDGAT1) activity were investigated via directed evolution, targeted mutagenesis, in vitro enzymatic assay, topological analysis, and transient expression of cDNA encoding selected enzyme variants in Nicotiana benthamiana. Directed evolution revealed that numerous amino acid residues were associated with increased BnaDGAT1 activity, and 67% of these residues were conserved among plant DGAT1s. The identified amino acid residue substitution sites occur throughout the BnaDGAT1 polypeptide, with 89% of the substitutions located outside the putative substrate binding or active sites. In addition, cDNAs encoding variants I447F or L441P were transiently overexpressed in N. benthamiana leaves, resulting in 33.2 or 70.5% higher triacylglycerol content, respectively, compared with native BnaDGAT1. Overall, the results provide novel insights into amino acid residues underlying plant DGAT1 function and performance-enhanced BnaDGAT1 variants for increasing vegetable oil production.


Germline RET Leu56Met Variant Is Likely Not Causative of Multiple Endocrine Neoplasia Type 2.
Activating variants in the receptor tyrosine kinase REarranged during Transfection (RET) cause multiple endocrine neoplasia type 2 (MEN 2), an autosomal dominantly inherited cancer-susceptibility syndrome. The variant c.166C>A, p.Leu56Met in RET was recently reported in two patients with medullary thyroid cancer (MTC). The presence of a pheochromocytoma in one of the patients, suggested a possible pathogenic role of the variant in MEN 2A. Here, we present clinical follow up of a Danish RET Leu56Met cohort. Patients were evaluated for signs of MEN 2 according to a set of predefined criteria. None of the seven patients in our cohort exhibited evidence of MEN 2. Furthermore, we found the Leu56Met variant in our in-house diagnostic cohort with an allele frequency of 0.59%, suggesting that it is a common variant in the population. Additionally, none of the patients who harbored the allele were listed in the Danish MTC and MEN 2 registries. In conclusion, our findings do not support a pathogenic role of the Leu56Met variant in MEN 2.


Opportunities and challenges of current electrophysiology research: a plea to establish 'translational electrophysiology' curricula.
Cardiac electrophysiology has evolved into an important subspecialty in cardiovascular medicine. This is in part due to the significant advances made in our understanding and treatment of heart rhythm disorders following more than a century of scientific discoveries and research. More recently, the rapid development of technology in cellular electrophysiology, molecular biology, genetics, computer modelling, and imaging have led to the exponential growth of knowledge in basic cardiac electrophysiology. The paradigm of evidence-based medicine has led to a more comprehensive decision-making process and most likely to improved outcomes in many patients. However, implementing relevant basic research knowledge in a system of evidence-based medicine appears to be challenging. Furthermore, the current economic climate and the restricted nature of research funding call for improved efficiency of translation from basic discoveries to healthcare delivery. Here, we aim to (i) appraise the broad challenges of translational research in cardiac electrophysiology, (ii) highlight the need for improved strategies in the training of translational electrophysiologists, and (iii) discuss steps towards building a favourable translational research environment and culture.


Temporal muscle flap in reconstruction of maxillo-facial tissues.
Presents our experience in reconstruction of postsurgical defects with use of temporal muscle flap. In the years 1996-2010 we treated 10 patients with malignant neoplasms of maxillo-facial region. Following tumor removal the fascia-muscle flaps of temporal muscle were used for reconstruction. Temporal muscle flaps were applied in 5 cases for reconstructing the defect of eye socket tissue after extensive resections and in the other 5 patients was used for reconstructing the defect of palate following maxillary resections. All the flaps of temporal muscle that were used for reconstructing eye sockets incorporated with no local reaction. In one case of flap applied into the oral cavity for tightly separate from nasal, partial necrosis of flap's edge and oro-nasal fistula occurred. Fascia-muscular flap of temporal muscle is a simple, easy and effective method of tissue reconstruction in maxillo-facial region following extensive oncologic procedures.


Biological Effects of Anti-Granulocyte-Macrophage Colony-Stimulating Factor (GM-CSF) Antibody Formation in Patients Treated With GM-CSF (Sargramostim) as Adjuvant Therapy of Melanoma.
We investigated the development of binding and neutralizing antibodies to granulocyte-macrophage colony-stimulating factor (GM-CSF) in patients receiving prolonged therapy with GM-CSF as adjuvant therapy of melanoma and the impact of these antibodies on biological effects. Fifty-three patients with high-risk melanoma that had been surgically excised were treated with GM-CSF, 125 μg/m daily for 14 days every 28 days for 1 year after surgical resection of disease. Serum samples for antibodies to GM-CSF were measured before treatment and on study days 155 and 351. Blood draws for testing biological effects were keyed to GM-CSF administration: days 0 (before), 15 (after 14 d on GM-CSF), 29 (after 14 d off GM-CSF), 155, and 351 (after 14 d on GM-CSF in the sixth and 13th cycle of treatment). Of 53 patients enrolled, 43 were evaluable for the development of anti-GM-CSF antibodies. Of these, 93% developed binding antibodies and 42% developed both binding and neutralizing antibodies. The increase in the white blood cell count, percent eosinophils, or neopterin levels engendered by GM-CSF administration was abrogated or markedly decreased in patients with neutralizing antibodies but not in patients who developed only binding antibodies. Ninety-three percent of patients with melanoma treated with GM-CSF as adjuvant therapy develop antibodies to GM-CSF. In those with neutralizing antibodies, a diminution of the biological effects of GM-CSF was observed. The development of neutralizing antibodies might also abrogate the potential clinical benefit of this treatment and should be considered in the design of future clinical trials.


'Just helping': Children living with a parent with young onset dementia.
While the consequences of caring for younger people with dementia have been a growing area of research, little is known about the children of these individuals. This study aimed to discover whether children of younger people with dementia can be compared to other young carers, the impact of their caring on mood, burden and resilience and what could promote coping. In-depth interviews were carried out with 12 participants aged 11-18. A grounded theory methodology was used, supplemented with three quantitative measures. Four higher order categories were identified focusing on: discovering dementia; developing a new relationship; learning to live with it and going through it together. Few participants showed depressive symptomatology, but more than half showed high levels of burden and most showed moderate levels of resilience. A three-stage process model of adapting to dementia is proposed, with children moving through grief to emotional detachment and increased maturity. This study has implications for service development in light of the National Dementia Strategy and proposals for whole family approaches for young carers. Whole family working by both dementia and children's services could reduce the burden on children of people with young onset dementia.


Myosin-X and talin modulate integrin activity at filopodia tips.
Filopodia assemble unique integrin-adhesion complexes to sense the extracellular matrix. However, the mechanisms of integrin regulation in filopodia are poorly defined. Here, we report that active integrins accumulate at the tip of myosin-X (MYO10)-positive filopodia, while inactive integrins are uniformly distributed. We identify talin and MYO10 as the principal integrin activators in filopodia. In addition, deletion of MYO10's FERM domain, or mutation of its β1-integrin-binding residues, reveals MYO10 as facilitating integrin activation, but not transport, in filopodia. However, MYO10's isolated FERM domain alone cannot activate integrins, potentially because of binding to both integrin tails. Finally, because a chimera construct generated by swapping MYO10-FERM by talin-FERM enables integrin activation in filopodia, our data indicate that an integrin-binding FERM domain coupled to a myosin motor is a core requirement for integrin activation in filopodia. Therefore, we propose a two-step integrin activation model in filopodia: receptor tethering by MYO10 followed by talin-mediated integrin activation.


Commentary: Breaking the mold of normative clinical decision making: is it adaptive, suboptimal, or somewhere in between?
Two articles in this issue, "Contextual decision making and the implementation of clinical guidelines" by Falzer and Garman, and " Uses and misuses of thresholds in diagnostic decision making" by Warner et al, take very different approaches to the issue of variation among physicians in diagnostic and therapeutic decision making. Falzer and Garman critically examine the well-known phenomenon of poor compliance with practice guidelines. They view this as a reflection of the mismatch between the guideline and the characteristics and needs of individual patients, and as a consequence of the adaptive judgment of the physician. They go on to show that, as the match between the individual patient and the hypothetical patient in the guideline increases, adherence with guidelines increases. Warner and colleagues take a more theoretical position on the larger issue of physician decision making, through the concept of "decision thresholds" originally advanced by Pauker and Kassirer, and attempt to model an example of clinical judgments through this conceptualization. The two articles represent an intriguing contrast on many levels, most fundamentally in that the first views any departure from normative behavior as goal oriented and adaptive and seeks to understand it; the second views it as suboptimal and seeks to minimize it.


[EEG coherence analysis in music listening].
Changes in EEG coherence were studied in subjects without special musical education when they were listening to low classic music (below 30 dB). The numbers of significant changes in coherence function in the left and right hemispheres were different. Predominant increase in coherence in the alpha 2, beta 1, and beta 2 frequency ranges was observed in the right temporal-central and parietal-occipital cortical areas. The left-hemispheric coherence was predominantly, decreased on the account of its decrease in the alpha range.


The (1)H, (13)C, (15)N resonance assignment, solution structure, and residue level stability of eosinophil cationic protein/RNase 3 determined by NMR spectroscopy.
Eosinophil cationic protein (ECP)/human RNase 3, a member of the RNase A family, is a remarkably cytotoxic protein implicated in asthma and allergies. These activities are probably due to ECP's ability to interact with and disrupt membranes and depend on two Trp, 19 Arg, and possibly an extremely high conformational stability. Here, we have used NMR spectroscopy to assign essentially all (1)H, (15)N, and backbone (13)C resonances, to solve the 3D structure in aqueous solution and to quantify its residue-level stability. The NMR solution structure was determined on the basis of 2316 distance constraints and is well-defined (backbone RMSD = 0.81 A). The N-terminus and the loop composed of residues 114-123 are relatively well-ordered; in contrast, conformational diversity is observed for the loop segments 17-22, 65-68, and 92-95 and most exposed sidechains. The side chain NH groups of the two Trp and 19 Arg showed no significant protection against hydrogen/deuterium exchange. The most protected NH groups belong to the first and last two beta-strands, and curiously, the first alpha-helix. Analysis of their exchange rates reveals a strikingly high global stability of 11.8 kcal/mol. This value and other stability measurements are used to better quantify ECP's unfolding thermodynamics.


Dystrophin is required for the formation of stable muscle attachments in the zebrafish embryo.
A class of recessive lethal zebrafish mutations has been identified in which normal skeletal muscle differentiation is followed by a tissue-specific degeneration that is reminiscent of the human muscular dystrophies. Here, we show that one of these mutations, sapje, disrupts the zebrafish orthologue of the X-linked human Duchenne muscular dystrophy (DMD) gene. Mutations in this locus cause Duchenne or Becker muscular dystrophies in human patients and are thought to result in a dystrophic pathology through disconnecting the cytoskeleton from the extracellular matrix in skeletal muscle by reducing the level of dystrophin protein at the sarcolemma. This is thought to allow tearing of this membrane, which in turn leads to cell death. Surprisingly, we have found that the progressive muscle degeneration phenotype of sapje mutant zebrafish embryos is caused by the failure of embryonic muscle end attachments. Although a role for dystrophin in maintaining vertebrate myotendinous junctions (MTJs) has been postulated previously and MTJ structural abnormalities have been identified in the Dystrophin-deficient mdx mouse model, in vivo evidence of pathology based on muscle attachment failure has thus far been lacking. This zebrafish mutation may therefore provide a model for a novel pathological mechanism of Duchenne muscular dystrophy and other muscle diseases.


Thyroxine signal transduction in liver cells involves phospholipase C and phospholipase D activation. Genomic independent action of thyroid hormone.
Numerous investigations demonstrate a novel role of thyroid hormone as a modulator of signal transduction. Protein kinase C (PKC) is critical to the mechanism by which thyroid hormones potentiate both the antiviral and immunomodulatory actions of IFNgamma in different cells and regulate the exchange of signalling phospholipids in hepatocytes. Because nothing is known about accumulation of PKC modulator - diacylglycerol in cells treated with T4, we examined the nongenomic effect of thyroid hormones on DAG formation and phospholipase activation in liver cells. The results obtained provide the first demonstration of phospholipase C, phospholipase D and protein kinase C nongenomic activation and diacylglycerol (DAG) accumulation by L-T4 in liver cells. The experiments were performed in either the [14C]CH3COOH-labeled rat liver slices or isolated hepatocytes pre-labeled by [14C]oleic acid. L-T4 activates the DAG production in a concentration- and time-dependent manner. DAG formation in stimulated cells is biphasic and short-lived event: there is an initial, rapid rise in DAG concentration and then a slower accumulation that can be sustained for a few minutes. The early phase of L-T4 generated DAG only is accompanied by phosphatidylinositol 4,5-bisphosphate level decrease and inositol 1,4,5-trisphosphate formation while the second phase is abolished by PKC inhibitor l,(5-isoquinolinesulphonyl)2methylpiperasine dihydrochloride (H7) and propranolol. The second phase of DAG production is accompanied by free choline release, phosphatidylcholine content drop and phosphatidylethanol (Peth) formation. Inhibitor of phospholipase-C-dependent phosphoinositide hydrolysis, neomycin sulfate, reduced the Peth as well as the DAG response to L-T4. The present data have indicated the DAG signaling in thyroid hormone-stimulated liver cells. L-thyroxine activates a dual phospholipase pathway in a sequential and synchronized manner: phospholipase C initiates the DAG formation, and PKC mediates the integration of phospholipase D into the signaling response during the sustained phase of agonist stimulation.


Challenges and psychosocial growth for older volunteers giving intensive humanitarian service.
We conducted a qualitative study of 38 mid-late life volunteers in intensive humanitarian service to ascertain the challenges, personal changes, and benefits they experienced from their volunteer activities. Intensive volunteering was defined as service done on a 24-hr a day basis at a location away from home. In-depth interviews were conducted by phone or in person with each participant. Couples serving together were interviewed conjointly. Data were analyzed using McCracken's 5-step process for long interviews. Participants described and resolved many challenges including adjustment to new situations and cultures, work-related challenges, and readjustment to home. Consistent with Erikson's descriptions of care, volunteers described benefits of expanded social networks, increased closeness to spouses, and increased compassion and empathy for others. In terms of wisdom, they described altered personal perspectives, with lessened materialism and self-focus, greater appreciation of cultural differences, and finding existential meaning in service. Because of population aging and social service organizational contraction, humanitarian service organizations may rely more on older volunteers to fulfill their goals. A deeper understanding of the challenges, ways in which they were met, and perceived benefits of later life intensive volunteering can inform organizational efforts to recruit older adults and support them in diverse settings.


Polyunsaturated fatty acid supplementation in peripheral neuropathy.
This study is an attempt to assess the role of dietary supplementation in the treatment and prevention of diabetic peripheral neuropathy. The authors developed an animal model system to study this problem. Animals given streptozotocin to induce a type I diabetic state showed elevated glucose levels and decreased body weight. Analysis of the sciatic nerve revealed a decrease in nerve conduction velocity and Na(+)-K(+)-ATPase activity. The activity of protein kinase C, another component of the nerve transmission process, was also affected by the diabetic state. The dietary intervention of polyunsaturated fatty acids seemed to revert some of these changes toward normal.


Recurrent Hyperparathyroidism Following Successful Four-Gland Parathyroidectomy and Normalization of Parathyroid Hormone in a Renal Transplant Patient.
Chronic kidney disease has an estimated prevalence of 10% in Australia and is predicted to rise in the coming years. Secondary hyperparathyroidism resulting from chronic kidney disease is an important cause of morbidity in these patients; and screening for secondary hyperparathyroidism is recommended in international guidelines. We present the case of a chronic kidney disease patient who developed recurrent hyperparathyroidism despite previous "total" parathyroidectomy and subsequent renal transplant. After targeted investigations he was diagnosed with an accessory parathyroid gland in his thorax, causing the recurrent hyperparathyroidism. He was managed with a thoracoscopic excision with a resultant drop in parathyroid hormone consistent with surgical cure. This case highlights the rare phenomenon of supernumerary and ectopic parathyroid glands. Cross sectional thoracic imaging can and should be used to detect and localize supernumerary glands not apparent at the time of original surgery.


Therapists' responses to training in brief supportive psychotherapy.
There is little research addressing supportive psychotherapy training. This article describes training clinicians in a form of brief supportive psychotherapy (BSP) for a multisite depression study, and reports on a survey of therapist attitudes toward BSP. We hypothesized that while most therapists would report acclimating to BSP, cognitive-behavioral therapy (CBT)-trained therapists would report greater frustration with BSP. Sixteen (89%) of 18 therapists completed a brief questionnaire. Therapists reported gaining comfort with supportive concepts and interventions. Therapists with cognitive behavior therapy orientations did not report significantly greater frustration with intervention restrictions. All practitioners indicated they were already using or were planning to use BSP outside the study, and that BSP training had altered their appreciation of psychotherapy. Most study therapists lacked prior supportive therapy training but reported adapting to BSP and appreciating its strengths and limitations.


Preterm's Nutrition from Hospital to Solid Foods: Are We Still Navigating by Sight?
As preterm birth rates are globally increasing, together with research on preterms' peculiar needs, neonatologists are still facing the challenge of how to properly feed them. The need to strike a balance between excessive catch-up growth and extrauterine growth retardation, both leading to adverse outcomes, is made even more difficult by the broad range of preterms' needs. Although mother's fresh milk is undoubtedly the best nourishment, its availability during hospital stay is often lower than recommended, and its fortification at discharge is still an open issue. Formula milks are available as an alternative to breast milk. However, choosing the right formula requires a thorough evaluation of the infant's perinatal history and targets. Last but not least, adequate timing and initiation of weaning in premature babies are still a poorly explored matter. This narrative review aims at evaluating the multitude of issues to consider when feeding preterms in the three stages of their first life: in-hospital care, discharge, and, eventually, weaning. Given the current absence of internationally shared guidelines, understanding the potential pitfalls of preterms' nutrition could help us trace the right path for the right preterm.


Comprehensive analysis of 66 complete molluscum contagiosum virus (MOCV) genomes: characterization and functional annotation of 47 novel complete MOCV genomes, including the first genome of MOCV genotype 3, and a proposal for harmonized MOCV genotyping indexing.
Four molluscum contagiosum virus (MOCV) genotypes (MOCV1-4) and four subtype variants (MOCV1p, MOCV1va, MOCV1vb, and MOCV1vc) were partially characterized using restriction enzyme profiling in the early 1980s/1990s. However, complete genome sequences of only MOCV1 and MOCV2 are available. The evolutionary pathways of MOCV genotypes and subtype variants with unavailable sequences remain unclear, and also whether all MOCV genotypes/subtype variants can be reliably detected and appropriately categorized using available PCR-based protocols. We de novo fully characterized and functionally annotated 47 complete MOCV genomes, including two putative non-MOCV1/2 isolates, expanding the number of fully characterized MOCV genomes to 66. To ascertain the placement of any putative novel MOCV sequence into the restriction profiling typing scheme, we developed an original framework for extracting complete MOCV genome sequence-based restriction profiles and matching them with reference restriction profiles. We confirmed that two putative non-MOCV1/2 isolates represent the first complete genomes of MOCV3. Comprehensive phylogenomic, recombination, and restriction enzyme recognition site analysis of all 66 currently available MOCV genomes showed that they can be agglomerated into six phylogenetic subgroups (PG1-6), corresponding to the subtype variants from the pioneering studies. PG5 was a novel subtype variant of MOCV2, but no PGs corresponded to the subtype variants MOCV1vb or MOCV4. We showed that the phylogenetic subgroups may have diverged from the prototype MOCV genotype lineages following large-scale recombination events and hinted at partial sequence content of MOCV4 and direction of recombinant transfer in the events that spawned PG5 and the yet undetected subtype variant MOCV1vb.IMPORTANCEFour molluscum contagiosum virus (MOCV) genotypes (MOCV1-4) and four subtype variants were partially characterized using restriction enzyme profiling in the 1980s/1990s, but complete genome sequences of only MOCV1 and MOCV2 are available. The evolutionary pathways whereby genotypes/subtype variants with unavailable sequences emerged and whether all MOCVs can be detected using current diagnostic approaches remain unclear. We fully characterized 47 novel complete MOCV genomes, including the first complete MOCV3 genome, expanding the number of fully characterized genomes to 66. For reliably classifying the novel non-MOCV1/2 genomes, we developed and validated a framework for matching sequence-derived restriction maps with those defining MOCV subtypes in pioneering studies. Six phylogenetic subgroups (PG1-6) were identified, PG5 representing a novel MOCV2 subtype. The phylogenetic subgroups diverged from the prototype lineages following large-scale recombination events and hinted at partial sequence content of MOCV4 and direction of recombinant transfer in the events spawning PG5 and yet undetected MOCV1vb variant.


Barriers to diaphragm use: the views of advanced practice nurses.
Women have used the contraceptive diaphragm for decades. Although use has recently declined, the diaphragm may find a new role in STI/HIV and dual-prevention programmes when microbicides become available. We developed a questionnaire to examine seven provider issues identified as possible barriers to diaphragm use among advanced practice nurses (APNs) specialising in women's health. The perceived degree to which each issue represented a barrier was examined. Non-parametric correlations were calculated between diaphragm fitting history, demographic and practice characteristics, and the response ratings for each issue. Responses were analysed for 204 APNs who averaged 15 years' experience in women's health care; 87% had fitted a diaphragm at least once, but 40% had not prescribed one in the past year. The degree to which each issue was perceived as a barrier varied. Based on respondents' ratings of a 'more than moderate barrier,' diaphragm non-promotion by women's health providers, effectiveness doubts, unfamiliarity and lack of access to educational materials were more often perceived as impeding diaphragm use. Other results indicated that APNs with recent diaphragm fitting history perceived five of the seven issues to be less of a barrier: non-promotion by women's health providers, lack of access to educational materials and to a fitting set, unfamiliarity, and inadequate reimbursement. Formulation of successful strategies to reintroduce the diaphragm will depend on better identification and understanding of provider-perceived barriers. This paper offers new insights about such barriers and guidance for the development of strategies for diaphragm reintroduction.


The ontogenesis of GABA mediated inhibition of growth hormone release in the sheep.
To investigate the ontogenesis of gamma-aminobutyric acid (GABA) mediated regulation of growth hormone release, the GABA agonist muscimol (250-300 micrograms/kg, i.v.) or antagonist, picrotoxin (300-500 micrograms/kg, i.v.) was administered to the chronically catheterized ovine fetus, infant lamb or adult ewe. Neither agent affected growth hormone release in fetuses younger than 105 days of gestation or in infant lambs or adult sheep. In fetuses between 115 and 140 days of gestation muscimol inhibited (n = 5, P less than 0.02) and picrotoxin stimulated (n = 5, P less than 0.001) fetal growth hormone secretion. These observations indicate the presence of tonic GABA mediated inhibition of growth hormone release in the late gestation fetus and are evidence that fetal GH secretion is not entirely unrestrained. Compared to other neurotransmitter systems potentially regulating growth hormone secretion, the GABA-ergic system is intermediate in its appearance. The presence of a growth hormone response to GABA agonists and antagonists in the fetal but not in the postnatal period is further evidence of the particularly prominent role of GABA as a neurotransmitter in the immature brain.


A multistep cytological approach for patients with jaundice and biliary strictures of indeterminate origin.
Fluorescence in situ hybridisation (FISH) increases the sensitivity for detecting pancreatobiliary tract cancer over routine cytology. In this study, diagnostic accuracy and costs of cytology and FISH in detecting cancer in patients with jaundice with biliary strictures were assessed. Brushing specimens from 109 patients with jaundice were obtained during endoscopic retrograde cholangiopancreatography and examined by cytology and FISH. The specimens were considered FISH-positive for malignancy if at least five polysomic cells or 10 cells with homozygous or heterozygous 9p21/p16 deletion were detected. Definitive diagnosis of the stricture as benign or malignant relied on surgical pathology (45 cases) or clinical-radiological follow-up >18 months (64 cases). We calculated costs of cytology and FISH based on the reimbursement from the Piedmont region, Italy (respectively, €33 and €750). Ninety of 109 patients had evidence of malignancy (44 pancreatic carcinomas, 36 cholangiocarcinomas, 5 gallbladder carcinomas, 5 other cancers), while 19 had benign strictures. Routine cytology showed 42% sensitivity, but 100% specificity for the diagnosis of malignancy, while FISH-polysomy showed 70% sensitivity with 100% specificity and FISH-polysomy plus homozygous or heterozygous 9p21/p16 deletion showed 76% sensitivity with 100% specificity. The cost per additional correct diagnosis of cancer obtained by FISH, in comparison with cytology, was €1775 using a sequential cytological approach (ie, performing FISH only in patients with negative or indeterminate cytology). FISH should be recommended as the second step in detecting cancer in patients with jaundice with pancreatobiliary tract strictures and cytology negative or indeterminate for malignancy.


Mechanical torque measurement in the proximal femur correlates to failure load and bone mineral density ex vivo.
Knowledge of local bone quality is essential for surgeons to determine operation techniques. A device for intraoperative measurement of local bone quality has been developed by the AO-Research Foundation (Densi - Probe®). We used this device to experimentally measure peak breakaway torque of trabecular bone in the proximal femur and correlated this with local bone mineral density (BMD) and failure load. Bone mineral density of 160 cadaver femurs was measured by ex situ dualenergy X-ray absorptiometry. The failure load of all femurs was analyzed by side-impact analysis. Femur fractures were fixed and mechanical peak torque was measured with the DensiProbe® device. Correlation was calculated whereas correlation coefficient and significance was calculated by Fisher's Ztransformation. Moreover, linear regression analysis was carried out. The unpaired Student's t-test was used to assess the significance of differences. The Ward triangle region had the lowest BMD with 0.511 g/cm(2) (±0.17 g/cm(2)), followed by the upper neck region with 0.546 g/cm(2) (±0.16 g/cm(2)), trochanteric region with 0.685 g/cm(2) (±0.19 g/cm(2)) and the femoral neck with 0.813 g/cm(2) (±0.2 g/cm(2)). Peak torque of DensiProbe® in the femoral head was 3.48 Nm (±2.34 Nm). Load to failure was 4050.2 N (±1586.7 N). The highest correlation of peak torque measured by Densi Probe® and load to failure was found in the femoral neck (r=0.64, P<0.001). The overall correlation of mechanical peak torque with T-score was r=0.60 (P<0.001). A correlation was found between mechanical peak torque, load to failure of bone and BMD in vitro. Trabecular strength of bone and bone mineral density are different aspects of bone strength, but a correlation was found between them. Mechanical peak torque as measured may contribute additional information about bone strength, especially in the perioperative testing.


Joint frequencies of alleles determined by separate formulations for the mating and mutation systems.
A method to calculate joint gene frequencies, which are the probabilities that two neutral genes taken at random from a population have certain allelic states, is developed taking into account the effects of the mating system and the mutation scheme. We assume that the mutation rates are constant in the population and that the mating system does not depend on allelic states. Under either--the condition that mutation rates are symmetric or that the mating unit is large and the mutation rate is small--the general formula is represented by two terms, one for the mating system and the other for the mutation scheme. The term for the mating system is expressed using the coancestry coefficient in the infinite allele model, and the term for the mutation scheme is a function of the eigenvalues and the eigenvectors of the mutation matrix. Several examples are presented as applications of the method, including homozygosity in a stepping-stone model with a symmetric mutation scheme.


Expression of ENaC subunits, chloride channels, and aquaporins in ovine fetal lung: ontogeny of expression and effects of altered fetal cortisol concentrations.
Transition of the epithelium of the fetal lung from fluid secretion to fluid reabsorption requires changes in the expression of ion channels. Corticosteroids regulate expression of several of these channels, including the epithelium sodium channel (ENaC) subunits and aquaporins (AQP). We investigated the ontogenetic changes in these ion channels in the ovine fetal lung during the last half of gestation, a time of increasing adrenal maturation. Expression of the mRNAs for the chloride channels, cystic fibrosis transmembrane conductance regulator (CFTR), and chloride channel 2 (CLCN2) decreased with age. Expression of mRNAs for AQP1, AQP5, and for subunits of ENaC (alpha, beta, gamma) increased with age. In the fetal sheep the expression of ENaCbeta mRNA was dramatically higher than the expression of ENaCalpha or ENaCgamma, but expression of ENaCbeta protein decreased with maturation, although the ratio of the mature (112 kDa) to immature (102 kDa) ENaCbeta protein increased with age, particularly in the membrane fraction. In contrast, ENaCalpha mRNA and protein both increase with maturation, and the mature form of ENaCalpha (68 kDa) predominates at all ages. A modest increase in fetal cortisol, within the range expected to occur naturally in late gestation but prior to active labor, increased ENaCalpha mRNA but not ENaCbeta, ENaCgamma, or AQP mRNAs. We conclude that in the ovine fetal lung, appearance of functional sodium channels is associated with induction of ENACalpha and ENaCgamma, and that ENaCalpha expression may be induced by even small, preterm increases in fetal cortisol.


US grower perceptions and experiences with glyphosate-resistant weeds.
A survey of 400 growers of maize, soybeans and cotton was made in the United States to determine perceptions, experiences and management practices with glyphosate-resistant weeds. The survey included growers in the north (corn belt) and south (cotton belt) of the USA in spring 2006. Interestingly, 24 and 39% of northern and southern growers, respectively, assumed they had glyphosate weed resistance on their farm. Of the 200 southern growers interviewed, 67% had planted continuous glyphosate-resistant (GR) crops for a period of 3-5 years. According to the survey respondents, the key method for managing glyphosate-resistant weeds was to rotate to other herbicides. Growers do value GR crop technology but are adopting measures to manage resistance only as needed.