Simultaneous labeling of vagal innervation of the gut and afferent projections from the visceral forebrain with dil injected into the dorsal vagal complex in the rat.
The vagal innervation of the different layers of the rat gastrointestinal wall was identified with the fluorescent carbocyanine dye Dil, injected into the dorsal motor nucleus of the vagus (dmnX). Multiple, bilateral injections were used to label all dmnX preganglionic motoneurons, and as a consequence, most of the vagal primary afferents that terminate in the adjacent nucleus of the solitary tract (nts) were also retrogradely and transganglionically labeled. With Fluorogold used to label the enteric nervous system completely and specifically, the Dil-labeled vagal profiles could be visualized and quantified in their anatomical relation to the neurons of the myenteric and submucous ganglia. In the myenteric plexus, vagal fibers and terminals were found throughout the gastrointestinal tract as far caudal as the descending colon, but there was a general decreasing proximodistal gradient in the density of vagal innervation. All parts of the gastric myenteric plexus (fundus, corpus, antrum), as well as the proximal duodenum, were extremely densely innervated, with vagal fibers and terminals in virtually every ganglion and connective. Further caudally, both the percentage of innervated myenteric ganglia and the average density of label within the ganglia rapidly decreased, with the exception of the cecum and proximal colon, where up to 65% of the ganglia were innervated. In the gastric and duodenal submucosa very few and in the mucosa no vagal fibers and terminals were found. With both normal epifluorescence and laser scanning confocal microscopy, highly varicose or beaded terminal structures of various size and geometry could be identified. The Dil injections, which impregnated the dmnX as well as the adjacent nts, resulted in retrograde and anterograde labeling of all the previously reported forebrain connections with the dorsal vagal complex. We conclude that the myenteric plexus is the primary target of vagal innervation throughout the gastrointestinal tract, and that its innervation is more complete than previously assumed. In contrast, vagal afferent (and efferent) innervation of mucosa and submucosa seems conspicuously sparse or absent. Furthermore, the use of more focal injections of Dil offers the prospect to simultaneously identify specific subsets of vagal preganglionics and their central nervous inputs.


Calcium and cAMP activate different chloride channels in the apical membrane of normal and cystic fibrosis epithelia.
The genetic disease cystic fibrosis (CF) causes decreased Cl- transport in several epithelia. cAMP-dependent regulation of apical membrane Cl- channels is defective in CF airway epithelia; as a result, CF epithelia fail to secrete Cl-. In contrast, Ca(2+)-stimulated Cl- secretion is intact in CF airway epithelia and thus has the potential to bypass the CF Cl- secretory defect. For a Cl- channel to govern Cl- secretion, it must be located in the apical membrane. To specifically investigate apical membrane Cl- channels, we studied cells grown on permeable filter supports and measured Cl- currents across the apical membrane. We found that Ca2+ and cAMP activate different Cl- channels in the apical membrane. (i) Ca(2+)-activated Cl- channels were present in the apical membrane of airway but not in intestinal epithelia. (ii) cAMP- but not Ca(2+)-activated Cl- channels were defective in CF airway epithelia. (iii) Ca(2+)- but not cAMP-activated Cl- channels were blocked by 4,4'-diisothiocyanato-2,2'-stilbenedisulfonate. (iv) Ca(2+)- and cAMP-activated apical channels had different anion permeabilities. (v) An increase in both second messengers produced an additive increase in Cl- current. These results also explain the puzzling observation that Ca(2+)-stimulated Cl- secretion is defective in CF intestine: the Ca(2+)-activated Cl- channels that could circumvent the Cl- secretory defect in CF airway are missing from the apical membrane of intestinal epithelia.


Mechanism of cis/trans equilibration of alkenes via iodine catalysis.
Iodine is commonly used to speed the equilibration of Wittig cis/trans alkene products. This study uses computational chemistry to study the catalyzed isomerization mechanism in detail for seven different examples of 1,2-disubstituted alkenes. We find that the iodo intermediates of the conventional three-step reaction path are weakly stable, bound by less than 7 kJ mol(-1) in five cases and nonexistent in the other two. These variations in relative stability appear to be closely related to the degree of conjugation interruption in the alkene upon attachment of iodine. The rate-determining reaction barrier always occurs in the middle step, the internal rotation of the iodo intermediate, and the variations in the barrier heights are dictated by varying levels of steric hindrance in the seven cases. Regiospecificity of I-atom addition and noticeable hyperconjugative effects are discussed. Comparisons between various theoretical approximations are performed to demonstrate the great difficulty in obtaining accurate results for iodine-atom bond-forming and bond-breaking energies.


One hundred years of X rays in biological research.
The history of X rays in biological research began almost simultaneously with Roentgen's discovery of his new rays. The history may be unique because of the remarkable collaboration of physicists, chemists, biologists and clinicians--collaborations which have produced and are continuing to produce major contributions to both biological and medical science. These contributions include the use of X rays to investigate molecular structure and function, the first demonstration of induced mutagenesis, the delineation of the cell cycle, the initiation of in vitro and in vivo cloning of mammalian cells, and original studies in DNA repair. The following is a personal overview of the history of some of these developments and their relationship to areas of current biological research. In each case an attempt has been made to trace developments from an early observation or observations to the current day. The history has been divided into two segments, each of approximately 50 years. This division seems appropriate because the separation occurs at approximately the same time as developments which were to play a major role in determining the future of radiation research.


StaphVAX (Nabi).
StaphVAX is a bivalent polysaccharide- and protein-conjugated vaccine, directed against capsular Staphylococcus aureus types 5 and 8, which are associated with 80 to 90% of S aureus clinical infections. The vaccine is being developed by Nabifor the potential treatment of infections in kidney patients who are receiving peritoneal dialysis and are prone to serious staphylococcal infections [193495], [221403], [222643], [283114]. In February 2001, Nabi revealed that it was movingforward during the second quarter of 2001 with previously stated plans to conduct a boosting study of StaphVAX in patients with end-stage renal disease (ESRD). This study would be conducted in patients who were enrolled in the first phase III trial and the company expected completion by early 2002 [283114]. The company was also progressing with scale-up of the manufacturing process for commercial production of the vaccine. Nabi met with the FDA in December 2000 to review results from its phase III trial of StaphVAX in patients with ESRD. The FDA stated that a definitive demonstration of preventative efficacy in another well-designed, randomized and controlled clinical study would be required for licensing. At this time, Nabi was making plans for this second phase III trial, despite plans to appeal the FDA's decision [397729].


Determination of metformin in rat plasma by HILIC-MS/MS combined with Tecan automation and direct injection.
Metformin is a well-known oral antihyperglycemic drug used in treatment of type II diabetes. Analysis of metformin in biological fluids is a challenge owing to its high polarity and small molecular size, which lead to poor retention of metformin on reversed-phase liquid chromatographic columns. A high-throughput method was developed and validated for the determination of metformin in rat plasma in support of preclinical toxicology studies, using hydrophilic interaction liquid chromatography tandem mass spectrometry (HILIC-MS/MS) and Tecan automated sample preparation. Extracted samples were directly injected onto the unbounded silica column with an aqueous-organic mobile phase. This HILIC-MS/MS method was validated for accuracy, precision, sensitivity, stability, matrix effect, recovery and calibration range. Acceptable intra-run and inter-run assay precision (coefficient of variation ≤ 3.9%) and accuracy (99.0-101.8%) were achieved over a linear range of 50-50,000 ng/mL. Metformin is stable in rat plasma for at least 6 h at room temperature, 147 days at -70°C and through three freeze (-70°C) and thaw cycles. Metformin is also stable in rat whole blood for at least 2 h at room temperature and in an ice-water bath. The validated method was successfully used in support of several preclinical studies where metformin is dosed together with an investigational drug substance. The ruggedness of the validated method was demonstrated by the incurred sample reproducibility test.


Role of endogenous cholecystokinin in the regeneration of pancreatic tissue after acute hemorrhagic pancreatitis in rats.
The aim of this study was to investigate the effect of endogenous cholecystokinin (CCK) on pancreatic regeneration after acute hemorrhagic pancreatitis. Acute hemorrhagic pancreatitis was induced in rats by two intraperitoneal cerulein injection (20 micrograms/kg BW) with 5h water-immersion stress once a day for successive 3 days. After the cessation of repetition of acute pancreatitis the rats were treated with successive feeding with 0.1% camostat-containing diet or SC injection of CR-1505 (CCK receptor antagonist, 50 mg/kg BW x 2/day) for 7 days. Zymogen enzymes and protein contents per DNA in pancreatic tissue were significantly higher in rats treated with camostat compared with control rats, and plasma CCK level was elevated. To the contrary, pancreatic regeneration was retarded in the rats treated with CR 1505. It is concluded that endogenous CCK has a trophic effect during regeneration after acute hemorrhagic pancreatitis.


Comparing land surface phenology of major European crops as derived from SAR and multispectral data of Sentinel-1 and -2.
The frequent acquisitions of fine spatial resolution imagery (10 m) offered by recent multispectral satellite missions, including Sentinel-2, can resolve single agricultural fields and thus provide crop-specific phenology metrics, a crucial information for crop monitoring. However, effective phenology retrieval may still be hampered by significant cloud cover. Synthetic aperture radar (SAR) observations are not restricted by weather conditions, and Sentinel-1 thus ensures more frequent observations of the land surface. However, these data have not been systematically exploited for phenology retrieval so far. In this study, we extracted crop-specific land surface phenology (LSP) from Sentinel-1 and Sentinel-2 of major European crops (common and durum wheat, barley, maize, oats, rape and turnip rape, sugar beet, sunflower, and dry pulses) using ground-truth information from the "Copernicus module" of the Land Use/Cover Area frame statistical Survey (LUCAS) of 2018. We consistently used a single model-fit approach to retrieve LSP metrics on temporal profiles of CR (Cross Ratio, the ratio of the backscattering coefficient VH/VV from Sentinel-1) and NDVI (Normalized Difference Vegetation Index from Sentinel-2). Our analysis revealed that LSP retrievals from Sentinel-1 are comparable to those of Sentinel-2, particularly for winter crops. The start of season (SOS) timings, as derived from Sentinel-1 and -2, are significantly correlated (average r of 0.78 for winter and 0.46 for summer crops). The correlation is lower for end of season retrievals (EOS, r of 0.62 and 0.34). Agreement between LSP derived from Sentinel-1 and -2 varies among crop types, ranging from r = 0.89 and mean absolute error MAE = 10 days (SOS of dry pulses) to r = 0.15 and MAE = 53 days (EOS of sugar beet). Observed deviations revealed that Sentinel-1 and -2 LSP retrievals can be complementary; for example for winter crops we found that SAR detected the start of the spring growth while multispectral data is sensitive to the vegetative growth before and during winter. To test if our results correspond reasonably to in-situ data, we compared average crop-specific LSP for Germany to average phenology from ground phenological observations of 2018 gathered from the German Meteorological Service (DWD). Our study demonstrated that both Sentinel-1 and -2 can provide relevant and at times complementary LSP information at field- and crop-level.


Nafamostat mesilate promotes endothelium-dependent vasorelaxation via the Akt-eNOS dependent pathway.
Nafamostat mesilate (NM), a synthetic serine protease inhibitor, has anticoagulant and anti-inflammatory properties. The intracellular mediator and external anti-inflammatory external signal in the vascular wall have been reported to protect endothelial cells, in part due to nitric oxide (NO) production. This study was designed to examine whether NM exhibit endothelium dependent vascular relaxation through Akt/endothelial nitric oxide synthase (eNOS) activation and generation of NO. NM enhanced Akt/eNOS phosphorylation and NO production in a dose- and time-dependent manner in human umbilical vein endothelial cells (HUVECs) and aorta tissues obtained from rats treated with various concentrations of NM. NM concomitantly decreased arginase activity, which could increase the available arginine substrate for NO production. Moreover, we investigated whether NM increased NO bioavailability and decreased aortic relaxation response to an eNOS inhibitor in the aorta. These results suggest that NM increases NO generation via the Akt/eNOS signaling pathway, leading to endothelium-dependent vascular relaxation. Therefore, the vasorelaxing action of NM may contribute to the regulation of cardiovascular function.


Liver fatty acid binding protein is the mitosis-associated polypeptide target of a carcinogen in rat hepatocytes.
Hepatocytes in normal rat liver were found previously to contain a cytoplasmic 14,000-dalton polypeptide (p14) that is associated with mitosis and is the principal early covalent target of activated metabolites of the carcinogen N-2-fluorenylacetamide (2-acetylaminofluorene). The level of immunohistochemically detected p14 was low when growth activity of hepatocytes was low, was markedly elevated during mitosis in normal and regenerating livers, but was very high throughout interphase during proliferation of hyperplastic and malignant hepatocytes induced in rat liver by a carcinogen (N-2-fluorenylacetamide or 3'-methyl-4-dimethylaminoazobenzene). We report here that p14 is the liver fatty acid binding protein. The nucleotide sequence of p14 cDNA clones, isolated by screening a rat liver cDNA library in bacteriophage lambda gt11 using p14 antiserum, was completely identical to part of the sequence reported for liver fatty acid binding protein. Furthermore, the two proteins shared the following properties: size of mRNA, amino acid composition, molecular size according to NaDodSO4 gel electrophoresis, and electrophoretic mobilities in a Triton X-100/acetic acid/urea gel. Their pI values overlapped in 2-dimensional isoelectric focusing/NaDodSO4 gel electrophoresis and showed the same response to delipidation. Either polypeptide reacted with and blocked the antiserum raised against the other polypeptide. The two polypeptides bound oleic acid similarly. Finally, identical elevations of cytoplasmic immunostain were detected specifically in mitotic hepatocytes with either antiserum. The collected findings are suggestive that liver fatty acid binding protein may carry ligands that promote hepatocyte division and may transport certain activated chemical carcinogens.


Study of influential factors on oligosaccharide formation by fructosyltransferase activity during stachyose hydrolysis by Pectinex ultra SP-L.
The influence of reaction conditions for oligosaccharide synthesis from stachyose using a commercial enzymatic preparation from Aspergillus aculeatus (Pectinex Ultra SP-L) was studied. Oligosaccharides were analyzed by gas chromatography with flame ionization detection (GC-FID) and matrix-assisted laser desorption/ionization-time-of-flight-mass spectrometry (MALDI-TOF-MS). Galactosyl-melibiose (DP(3)) was synthesized as a result of fructosidase activity, whereas fructosyl-stachyose (DP(5)) and difructosyl-stachyose (DP(6)) were formed as a consequence of the fructosyltransferase activity of Pectinex Ultra SP-L. The optimal reaction conditions for the synthesis of penta- and hexasaccharides were 60 °C, pH 5.5, 600 mg/mL stachyose, and 34 U/mL enzyme. Reaction time played an important role in oligosaccharide mixture composition constituted by 20% DP(5), 0.7% DP(6), 55% stachyose, 21% galactosyl-melibiose, and 1% monosaccharides after 1 h and 16% DP(5), 4% DP(6), 27% stachyose, 44% galactosyl-melibiose, and 2% monosaccharides after 3 h. In conclusion, stachyose could be used as a substrate for the enzymatic synthesis of new oligosaccharides that may open new opportunities in the development of future prebiotics.


A likelihood framework for estimating phylogeographic history on a continuous landscape.
Due to lack of an adequate statistical framework, biologists studying phylogeography are abandoning traditional methods of estimating phylogeographic history in favor of statistical methods designed to test a priori hypotheses. These new methods may, however, have limited descriptive utility. Here, we develop a new statistical framework that can be used to both test a priori hypotheses and estimate phylogeographic history of a gene (and the statistical confidence in that history) in the absence of such hypotheses. The statistical approach concentrates on estimation of geographic locations of the ancestors of a set of sampled organisms. Now we use (2) to derive the likelihood of the ancestral geographic coordinates and the value of the scaled dispersal parameter, given the observed geographic coordinates (assuming known topology and branch lengths). Using a maximum likelihood approach, which is implemented in the new program PhyloMapper, we apply this statistical framework to a 246-taxon mitochondrial genealogy of North American chorus frogs, focusing in detail on one of these species. We demonstrate three lines of evidence for recent northward expansion of the mitochondrion of the coastal clade of Pseudacris feriarum: higher per-generation dispersal distance in the recently colonized region, a noncentral ancestral location, and directional migration. After illustrating one method of accommodating phylogenetic uncertainty, we conclude by discussing how extensions of this framework could function to incorporate a priori ecological and geological information into phylogeographic analyses.


A nine-long non-coding RNA signature for prognosis prediction of patients with lung squamous cell carcinoma.
Lung squamous cell carcinoma (LUSC) is malignant disease with poor therapeutic response and unfavourable prognosis. This study aims to develop a long non-coding RNA (lncRNA) signature for survival prediction in patients with LUSC. We obtained lncRNA expression profiles of 493 LUSC cases from The Cancer Genome Atlas, and randomly divided the samples into a training set (n= 296) and a testing set (n= 197). Univariate Cox regression and random survival forest algorithm were performed to select optimum survival-related lncRNAs. A lncRNA-focused risk score model was then constructed for prognosis prediction in the training set and further validated in the testing set and the entire set. Finally, bioinformatics analysis was carried out to explore the potential signaling pathways associated with the prognostic lncRNAs. A set of 9 lncRNAs were found to be strongly correlated with overall survival of LUSC patients. These 9 lncRNAs were integrated into a prognostic signature, which could separate patients into high- and low-risk groups with significantly different survival times in the training set (median: 30.5 vs. 80.5 months, log-rank P< 0.001). This signature was also confirmed in the testing set and the entire set. Besides, the prognostic value of the 9-lncRNA signature was independent of clinical features and maintained stable in stratified analyses. Functional enrichment study suggested that the 9 lncRNAs may be mainly involved in metabolism-related pathways, phosphatidylinositol signaling system, p53 signaling pathway, and notch signaling pathway. Our study demonstrated the potential clinical implication of the 9-lncRNA signature for survival prediction of LUSC patients.


ASSESSMENT OF RADIOLOGICAL HAZARDS OF USING PETROLEUM RAW MATERIALS AND THEIR WASTE.
Activity concentrations of 238U, 232Th and 40K in raw and waste petroleum materials (Egypt and Kuwait) were measured using gamma ray spectrometer. The average values of 226Ra, 232Th and 40K were 21.1 ± 3.2, 7.6 ± 1.3 and 88.4 ± 8.2 Bq kg-1 for Egyptian samples while for Kuwaiti samples, they were 25.2 ± 3.4, 6.1 ± 2.2 and 67.8 ± 6.4 Bq kg-1, respectively. All samples had activity less than the exemption level recommended by the International Atomic Energy Agency. Moreover, radiological indices of radium equivalent, external, internal, alpha and gamma indices and radiation dose as well were calculated and their values were lower than the recommended regulatory limits. Thus, radiation exposure to petroleum materials did not present a significant radiological hazard.


Gene expression heterogeneities in embryonic stem cell populations: origin and function.
Stem and progenitor cells are populations of cells that retain the capacity to populate specific lineages and to transit this capacity through cell division. However, attempts to define markers for stem cells have met with limited success. Here we consider whether this limited success reflects an intrinsic requirement for heterogeneity with stem cell populations. We focus on Embryonic Stem (ES) cells, in vitro derived cell lines from the early embryo that are considered both pluripotent (able to generate all the lineages of the future embryo) and indefinitely self renewing. We examine the relevance of recently reported heterogeneities in ES cells and whether these heterogeneities themselves are inherent requirements of functional potency and self renewal.


Self-reported periodontitis in cannabis club members in Montevideo, Uruguay. An exploratory study.
In 2013, the Oriental Republic of Uruguay enacted a law regulating the cannabis market, and since then, an increasing number of users has been registered. Previous reports based on data from other countries link cannabis smokers to worse periodontal health status. The aim of this study is to describe self-reported gingival and periodontal health status and estimate the prevalence of periodontitis among Uruguayan cannabis club members. A cross-sectional study was conducted on a convenience sample of cannabis club members in Uruguay. A survey was used to gather sociodemographic data and information on oral hygiene, use of cannabis and other drugs, and self-reported gingival and periodontal health status. Eight questions validated in Spanish were used to estimate the prevalence of total periodontitis and severe periodontitis, based on two previously implemented predictive models. The survey was completed by 50 people, of whom 68% were male and 78% were in the 20-40 year age range. Based on the first model, estimated prevalence was 12% for total periodontitis and 10% for severe periodontitis. Based on the second model, estimated prevalence was 36% for total periodontitis and 12% for severe periodontitis. These findings suggest that smoking cannabis could potentially have a detrimental effect on oral health status.


Clinical research in developing countries: recent moral arguments.
During the 1990s, bioethicists raised questions about certain clinical trials conducted in developing countries. These inquiries led to revisions in the Declaration of Helsinki and recommendations from the US National Bioethics Advisory Commission. This article raises doubts about the original questions and subsequent recommendations. It is possible that impractical solutions have been proposed for nonexistent ethical problems.


Development of a Survey of Sunscreen Use and Attitudes among Adults in Two Coastal States, 2019.
Skin cancer is the most common form of cancer in the United States, and regular use of broad-spectrum sunscreens can prevent skin cancer. However, a new law in Hawaii that limits sunscreen choices due to the belief that some UV (ultraviolet) filters may damage coral reefs may reduce sunscreen use and increase skin-cancer risk. Because of this, there is a need for measurement tools to help understand consumer behavior and determinants of sunscreen purchase and use. The objectives of this study were (1) to test new questionnaire measures relevant to the Hawaii Sunscreen Ban; and (2) to assess adults' knowledge, attitudes, and habits related to sunscreen in two other coastal states. This survey of adult residents of California and Florida was conducted in the summer of 2019. Newly developed scales addressed beliefs about effects of sunscreens on aquatic/marine environments and awareness of the Hawaii sunscreen ban. Respondents completed the survey twice to evaluate the test-retest reliability. Respondents (n = 162) were mainly female, White, and college-educated. New scales had moderate-to-high internal consistency and high test-retest reliability. Sunscreen use was high, sunburn was common, and knowledge and attitudes about sunscreen were modest. Most respondents did not know the specifics of the Hawaii Sunscreen Ban. In multivariate models, significant predictors of sunscreen use were being older, female, and having higher sunscreen knowledge. Sunscreen beliefs were not significantly associated with sunscreen use or sunburn. The findings support the use of the newly developed survey and suggest that more education about sunscreen and sunscreen ingredients is needed.


Immunodepression following neurosurgical procedures.
To determine the influence of a selective, sterile central nervous system surgery on immune reactivity, particularly whether a decrease of monocytic human leukocyte antigen-DR expression, indicating immunodepression, occurs after neurosurgery and if this measurement is useful for identification of patients with a high risk of infection. Prospective study. Department of neurosurgery and intensive care unit in a university hospital. Blood samples were obtained from 46 patients at least once during the first 3 days after undergoing sterile central nervous system surgery. Fourteen of these patients developed infectious complications as defined by clinical and microbiological criteria. In ten of 46 patients, paired samples of blood and cerebrospinal fluid were collected from a ventricle drain at the following times: 1 day before surgery; several times on the day of surgery; and every day after surgery for at least 6 days. Monocytic human leukocyte antigen-DR expression, as measured by flow cytometry on days 1 through 3 after surgery in 46 patients, was lower in 14 patients who developed infection after neurosurgery (p < .0001). In all ten closely monitored patients, monocytic human leukocyte antigen-DR expression decreased temporarily after surgery. Of these patients, only one patient showed a persistent and considerably decreased monocytic human leukocyte antigen-DR expression. This patient was the only patient in this subgroup who developed sepsis syndrome. In order to assess whether the monocytic human leukocyte antigen-DR decrease was associated with a preceding inflammatory response, local and systemic concentrations of interleukin (IL)-1 beta, IL-6, IL-8, tumor necrosis factor-alpha, and interferon-gamma were measured in this subgroup. These cytokines were not detectable in plasma during the first days after surgery. In contrast, considerable increases of IL-6 and IL-8 concentrations were detectable in cerebrospinal fluid within hours after surgery. A decrease of monocytic human leukocyte antigen-DR expression occurs after neurosurgery and is associated with a preceding, strong, intracranial (but not systemic) inflammatory response. A very low monocytic human leukocyte antigen-DR expression (< 30% positive monocytes) suggests the possibility of infection. Measurement of monocytic human leukocyte antigen-DR expression could help to detect patients with a high risk of infection after neurosurgery. Our results suggest that even sterile central nervous system surgery may contribute to general immunodepression. The local intracranial inflammatory response may be involved in this process.


Standardization of calibration and quality control using surface enhanced laser desorption ionization-time of flight-mass spectrometry.
Protein profiling by surface enhanced laser desorption ionization-time of flight-mass spectrometry (SELDI-TOF-MS) is gaining importance as a diagnostic tool for a whole range of diseases. This report describes a QC procedure, which acts prospectively by checking the calibration before starting profiling experiments. A well-defined protocol for calibration of the Protein Biosystem IIc instrument was established, using a commercial QC sample containing independent certified standards and by determination of acceptance criteria. Instrument calibration was performed externally every week with the standards provided by the manufacturer. QC was performed for the period of 5 months. According to the acceptance criteria defined in this study, data points should be in the established range of the process mean+/-2 standard deviations for the mass-to-charge ratios (m/z values), peak intensities, signal-to-noise ratios (S/N), and peak resolutions for insulin and apomyoglobin in the QC sample. Moreover, it was demonstrated that the pipetting variability in the handling of the QC sample significantly contributed to systematic errors and that spotting of a larger volume of QC sample resulted in a better reproducibility. Stringent quality control of the calibration part of SELDI-TOF-MS experiments prevents unreliable data acquisition from the very start.


Emotion, working memory, and cognitive control in patients with first-onset and previously untreated minor depressive disorders.
To explore working memory and the ability to process different emotional stimuli in patients with first-onset and untreated minor (mild or moderate) depression. Patients with first-onset and previously untreated minor depression, and healthy controls, were enrolled. Using a modified Sternberg working memory paradigm to investigate the combined effects of emotional stimuli with working memory, participants were exposed to experimental stimuli comprising pictures that represented positive, neutral and negative emotions. Working memory ability was measured using reaction time and accuracy, and emotion-processing ability was measured using pupil diameter. Out of 36 participants (18 patients with minor depression and 18 controls), there were no statistically significant between-group differences in response time and accuracy. Positive stimuli evoked changes in pupil diameter that were significantly smaller in patients with minor depression versus controls, but changes in pupil diameter evoked by negative stimuli were not significantly different between the two groups. Healthy subjects showed a stronger emotional response to positive emotional stimuli than patients with first onset and previously untreated minor depression, but there were no differences in response to negative emotions. There were no statistically significant between-group differences in terms of speed of cognitive response, but this may have been due to the relatively small samples sizes assessed. Studies with larger sample populations are required to further investigate these results.


A Treasure of Bioactive Compounds from the Deep Sea.
The deep-sea environment is a unique, challenging extreme habitat where species have had to adapt to the absence of light, low levels of oxygen, high pressure and little food. In order to survive such harsh conditions, these organisms have evolved different biochemical and physiological features that often have no other equivalent in terrestrial habitats. Recent analyses have highlighted how the deep sea is one of the most diverse and species-rich habitats on the planet but less explored compared to more accessible sites. Because of their adaptation to this extreme environment, deep-sea species have the potential to produce novel secondary metabolites with potent biological activities. Recent advances in sampling and novel techniques in microorganism culturing and chemical isolation have promoted the discovery of bioactive agents from deep-sea organisms. However, reports of natural products derived from deep-sea species are still scarce, probably because of the difficulty in accessing deep-sea samples, sampling costs and the difficulty in culturing deep-sea organisms. In this review, we give an overview of the potential treasure represented by metabolites produced by deep marine species and their bioactivities for the treatment and prevention of various human pathologies.


Clinical efficacy of icotinib in patients with advanced nonsquamous non-small cell lung cancer with unknown EGFR mutation status that failed to respond to second-line chemotherapy.
Evaluation of the clinical efficacy and safety of icotinib in advanced nonsquamous non-small cell lung cancer (NSCLC) patients with an unknown EGFR mutation who failed to respond to second-line chemotherapy. Seventy-six cases of advanced nonsquamous NSCLC were involved in this study from seven hospitals from the Hubei province of China. Patients with an unknown EGFR mutation status were treated with Icotinib, at an oral dosage of 125 mg three times daily. All patients were followed up for at least 1 year to observe the efficacy, adverse reactions, and 1-year survival. The patients' overall objective response rate (ORR) was 34.2%, the disease control rate (DCR) was 75.0%, the clinical benefit rate (CBR) was 80.2%, the median progression-free survival (PFS) was 11.0 months, the median overall survival (OS) was 16.9 months, and the 1-year OS rate was 63.2%. Gender and smoking history were associated with the DCR (P<0.05). Both PFS and OS were significantly higher in groups that had pre-accepted ≤6 cycles of chemotherapy than in groups that had pre-accepted >6 cycles. Our results demonstrated that icotinib had a better DCR or clinical benefits for treating the patients with unknown EGFR mutation who failed to respond to second-line chemotherapy in advanced nonsquamous NSCLC, and the adverse effects are tolerable.


DNA replication stress drives fragile site instability.
DNA replication stress is one of the early drivers enabling the ongoing acquisition of genetic changes arising during tumorigenesis. As such, it is a feature of most pre-malignant and malignant cells. In this review article, we focus on the early events initiating DNA replication stress and the preferential sensitivity of common fragile sites (CFSs) to this stress. CFSs are specific genomic regions within the normal chromosomal structure, which appear as gaps and breaks in the metaphase chromosomes of cells grown under mild replication stress conditions. The main characteristics predisposing CFSs to instability include late replication timing, delayed replication completion, failure to activate additional origins, origin paucity along large genomic regions, collision between replication and transcription complexes along large genes, and the presence of AT-dinucleotide rich sequences. The contribution of these features to instability at CFSs during early cancer development is discussed.


Analysis of murine HOXA-2 activity in Drosophila melanogaster.
The murine HOXA-2 protein shares amino acid sequence similarity with Drosophila Proboscipedia (PB). In this paper, we test whether HOXA-2 and PB are functionally equivalent in Drosophila. In Drosophila, PB inhibits SCR activity required for larval T1 beard formation and adult tarsus formation and is required for maxillary palp and proboscis formation. HOXA-2 expressed from a heat-shock promoter weakly suppressed SCR activity required for T1 beard formation. But interestingly neither PB nor HOXA-2 expressed from a heat-shock promoter suppressed murine HOXA-5 activity, the murine SCR homologue, from inducing ectopic T1 beards in T2 and T3, indicating that HOXA-5 does not interact with PB. HOXA-2 activity expressed from the Tubulin alpha 1 promoter modified the pb null phenotype resulting in a proboscis-to-arista transformation, indicating that HOXA-2 was able to suppress SCR activity required for tarsus formation. However, HOXA-2 expressed from a Tubulin alpha 1 promoter was unable to direct maxillary palp determination when either ectopically expressed in the antenna or in the maxillary palp primordia of a pb null mutant. HOXA-2 was also unable to rescue pseudotrachea formation in a pb null mutant. These results indicate that the only activity that PB and HOXA-2 weakly share is the inhibition of SCR activity, and that murine HOXA-5 and Drosophila SCR do not share inhibition by PB activity.


Carbonic anhydrase activities from the rainbow trout lens correspond to the development of acute gas bubble disease.
Dissolved gas supersaturation is hazardous to fish and can result in gas bubble disease (GBD). Signs of GBD typically include bubbles in the eyes, fins, skin, lateral line, and gill filaments. Ocular abnormalities in diseased salmonids typically occur after aberrant gas production in the eyes. In this study, freshwater rainbow trout Oncorhynchus mykiss were exposed experimentally to percent total gas pressure (TGP%) levels of 104% (control) and 115%. No mortalities occurred during the 7-d experimental period. Effects of GBD were observed externally as a darkened skin, exophthalmia, localized hemorrhage in the eye, and gas bubbles on the operculum. Additional signs included increased swimming activity and, more frequently, panic episodes. Carbonic anhydrase (CA) enzyme activities from the lens and retina were determined at days 0, 1, 3, 5, and 7 of the study. Venous blood gases were also measured on day 7. Retinal pH did not differ between normal and affected fish, but blood characteristics such as the partial pressure of O2, partial pressure of CO2, carboxyhemoglobin level, and bicarbonate ion concentration were significantly elevated in affected fish relative to normal fish. Venous blood pH and oxyhemoglobin levels were not significantly different between affected and normal fish. Patterns of response to total dissolved gas levels differed between the lens and the retina. Mean CA activities in the lenses of fish exposed to a TGP% level of 115% were significantly below those of control fish. However, retinal CA activities did not significantly differ between the two groups over the course of the experiment. These findings show that dissolved gas supersaturation reduces CA activity in the rainbow trout lens.


Procedure-Specific Complications Associated with Ultrasound-Guided Erector Spinae Plane Block for Lumbar Spine Surgery: A Retrospective Analysis of 342 Consecutive Cases.
Presumed benefits of erector spinae plane blocks (ESPB) include an enhanced safety profile and few complications. There are few large series, which report the incidence of complications associated with ESPB on a procedure-specific basis. The objective of this retrospective cohort study was to estimate the incidence of complications of ESPB in a large series of patients undergoing lumbar spine surgery. We included 342 consecutive patients who underwent any primary lumbar spine surgery via posterior approach (November 2018-July 2020). All patients received bilateral ultrasound-guided ESPB. The primary study outcome was the incidence of any perioperative complication, defined a priori as sensory, motor, hematologic, hemodynamic or respiratory complication consistent with plausible contribution from the ESPB. Secondary outcomes included the incidence of numeric rating scale (NRS) pain scores ≥7 in the post anesthesia care unit (PACU) and risk factors associated with NRS ≥7 (age, sex, ASA class, BMI, opioid tolerance, surgical type, and duration). We did not identify any pre-specified complications associated with ESPB. There was one unilateral pneumothorax, in one patient, deemed unlikely to have been related to ESPB. NRS ≥7 was found in 17/342 patients (5%) and was independent of any background differences or risk factors assessed. Ultrasound guided ESPB for lumbar spine surgery was associated with zero complications, no interference with intraoperative neuromonitoring or the early postoperative neurological examination, and low incidence of poorly controlled pain in the PACU. These results help to establish procedure-specific risks and benefits of ESPB for spine surgery.


Insights from mixture cure modeling of molecular markers for prognosis in breast cancer.
With the ultimate aim of improving clinical management of breast cancer, investigators have sought to identify molecular genetic markers that stratify newly diagnosed patients into subtypes differing in short- or long-term prognosis. Conventional survival models can fail to describe adequately the relationship between subtype and disease recurrence, particularly when there is a substantial proportion of long-term disease-free survivors. The observed patterns of disease-free survival in an undifferentiated patient cohort may be explained by an underlying mixture of two subgroups: patients who will remain free of disease in the long term (ie, cured), and those who will experience disease recurrence within their lifetime (ie, susceptible.) In this article, we review the concepts and methods of the mixture cure models and apply them in the analysis of molecular genetic prognostic factors for disease-free survival and time to disease recurrence in a cohort of patients with axillary lymph node-negative breast cancer.


[In vitro experiment showing that chemotherapy of tuberculosis by aminoglycoside antibiotics may influence successive chemotherapy with rifampicin].
When Mycobacterium tuberculosis strain H37Rv was cultivated in Ogawa egg medium containing 5 micrograms/ml streptomycin and/or 10 micrograms/ml kanamycin, which were considered as subinhibitory, it was observed that growing bacterial population contained several times more rifampicin-resistant mutants than did the parent strain. The ratio of isoniazid-resistant mutants did not change by the above treatment. The finding suggests that, even without the use of rifampicin, the bacterial population of patients becomes more resistant to rifampicin by chemotherapy in the past with streptomycin or kanamycin. Furthermore, it was shown that the ratio of streptomycin-resistant mutants increased by pre-treatment with kanamycin and the ratio of kanamycin-resistant mutants by pre-treatment with streptomycin. Parent, susceptible bacterial population develops 4R-phenotype mutants which are resistant to four drugs, high concentrations of kanamycin, lividomycin and paraomomycin and a low concentration of capreomycin, whereas streptomycin-resistant mutant population does not develop the 4R mutants but develops only mutants with the KR phenotype which is almost mono-resistant to kanamycin.


On the origins of the salt-concentration-dependent instability and lateral nanoscale heterogeneities of weak polyelectrolyte brushes: gradient brush experiment and Flory-type theoretical analysis.
We investigated, by experiment and theory, the lateral structure of a weak polyelectrolyte brush at various added salt concentrations and chain grafting densities. Model poly(2-(dimethylamino)ethyl methacrylate) (PDMAEMA) brushes with grafting density gradients were developed for this study by using a novel "Langmuir-Blodgett-deposition-under-compression" (LB\C) method. Fluid AFM images of these brushes indicate that the lateral structure of the brush system is sensitive to both added salt concentration and grafting density. Under low salt conditions, 0-20 mM NaCl, the brush structure shows strong microscopic lateral heterogeneities at high grafting densities; both the width and height of the heterogeneities increase with increasing grafting density but are independent of the salt concentration. As the bulk salt concentration is increased to an intermediate regime, 60-100 mM NaCl, these heterogeneities become smaller in size and number, coexisting with smooth homogeneous regions. At high enough concentrations, 300-500 mM NaCl, the entire surface becomes homogeneous. A simple free energy-based Flory-type argument is presented which explains the essential features of the thermodynamic behavior of the brush system. In the zero-salt limit, relatively few monomers are charged, and the hydrophobicity of the backbone chain drives the collapse/aggregation of the chains. At high salt concentrations, the brush chains become sufficiently charged to overcome the hydrophobic nature of the monomers and stabilize the homogeneous state. However, at intermediate salt concentrations, it is found that the osmotic pressure of the counterions surrounding the charged polymer moieties can be decreased by collapsing the chain structure while simultaneously decreasing the number of charges along the backbone and releasing small ions into the bulk solution. This effect, which we term "osmotic instability", serves to destabilize the homogeneous brush configuration.


Seasonal dynamics of bacterial community structure and composition in cold and hot drinking water derived from surface water reservoirs.
In temperate regions, seasonal variability of environmental factors affects the bacterial community in source water and finished drinking water. Therefore, the bacterial core community and its seasonal variability in cold and the respective hot drinking water was investigated. The bacterial core community was studied by 16S rRNA-based SSCP fingerprint analyses and band sequencing of DNA and RNA extracts of cold and hot water (60 °C). The bacterial communities of cold and hot drinking water showed a highly different structure and phylogenetic composition both for RNA and DNA extracts. For cold drinking water substantial seasonal dynamics of the bacterial community was observed related to environmental factors such as temperature and precipitation affecting source and drinking water. Phylogenetic analyses of the cold water community indicated that the majority of phylotypes were very closely affiliated with those detected in former studies of the same drinking water supply system (DWSS) in the preceding 6 years, indicating a high stability over time. The hot water community was very stable over time and seasons and highly distinct from the cold water with respect to structure and composition. The hot water community displayed a lower diversity and its phylotypes were mostly affiliated with bacteria of high temperature habitats with high growth rates indicated by their high RNA content. The conversion of the cold to the hot water bacterial community is considered as occurring within a few hours by the following two processes, i) by decay of most of the cold water bacteria due to heating, and ii) rapid growth of the high temperature adapted bacteria present in the hot water (co-heated with the cold water in the same device) using the nutrients released from the decaying cold water bacteria. The high temperature adapted bacteria originated partially from low abundant but beforehand detected members of the cold water; additionally, the rare members ("seed bank ") of the cold water are considered as a source.


Combining ecological, eco-cultural, and environmental justice parameters to create Eco-EJ indicators to monitor cultural and environmental justices for diverse communities around contaminated sites.
Assessing environmental quality often requires selection of indicators that can be employed over large spatial scales and over long-time periods to assess the health and well-being of species, natural communities, and ecosystems, and to detect changes warranting intervention. Typically, the ecologic environment and the human environment are evaluated separately and selection of indicators and monitoring approaches are not integrated even though ecological indicators may also provide information on risk to human consumers from contaminants (e.g., eco-cultural indicators) or because of disease levels. This paper is a call for ecologists and managers to consider diverse cultural and environmental injustice disparities and health issues when selecting indicators for environmental assessment and monitoring. There is an opportunity for managers and community members to work together to preserve ecological and cultural resources and heritages. We propose a paradigm that selects indicators and monitoring approaches that lend themselves to the integration of human-diversity and uniqueness in the same manner that the selection of ecological indicators and monitoring approaches consider biological species diversity and uniqueness. The proposed paradigm builds on ecological risk assessment techniques, developing analogous endpoints for neighboring communities. For example, identification and protection of human communities, particularly culturally diverse and environmental justice communities, identification of contaminant corridors (e.g., through water or green corridors) into communities, and eco-monitoring of vulnerable communities are not routine at contaminated sites. Green corridors refers to a width of wild habitat (forest, grasslands) that connects other similar habitat paths (usually a corridor runs through an urban or suburban habitat). We coin the term Eco-EJ indicators for these endpoints, including examination of (1) unique cultural relationships to resources; (2) connectedness of on-site and off-site resources and habitats; (3) health of threatened, rare, and unique cultures and communities; and (4) linkages between ecological, eco-cultural, and public health for monitoring and assessment. We also propose that assessment and monitoring include these Eco-EJ indicators, especially for communities near facilities that have extensive chemical or radiological contamination.Developing these indicators to assess risk to culturally diverse and environmental justice communities would be an equivalent goal to reducing risk for significant ecological resources (e.g., endangered species, species of special concern). These Eco-EJ indicators are complementary to the usual human health-risk assessments, would include surveys of neighboring vulnerable communities, and require time and re-organization of current data and additional data collection at site boundaries and in adjacent communities, as well as rethinking the human component of indicators. This approach lends itself to addressing some diverse cultural and environmental justice issues with current indicator selection and biomonitoring, and helps identify specific hotspots of unique ecosystem risk and environmental justice community risk. We briefly discuss ecological and eco-cultural monitoring already on-going at three Department of Energy sites to illustrate how the addition of these indicators might work and add value to environmental management and to their relationships with surrounding communities. We recommend that managers of contaminated sites convene people from culturally diverse communities, environmental justice communities, local and federal government, Tribes, resource trustees, managers, and other stakeholders to develop appropriate site-specific indicators to address environmental inequities around contaminated facilities.


Variations in methadone treatment practices. Results from a national study.
To examine the extent to which outpatient methadone maintenance treatment units are engaging in treatment practices that previous research indicates are ineffective (eg, inadequate dose levels); to examine factors that may be related to variation in methadone treatment practices. Survey of unit directors and clinical supervisors. The study includes units that vary in terms of ownership (public, private for-profit, or private not-for-profit) and setting (eg, hospital-based, mental health center-based, or free-standing facility). A national random sample of 172 units participated, for an 82% response rate; the data were weighted to ensure that they were nationally representative. Clients' awareness of and influence on doses; units' use of take-home dosages; upper limits on doses; average dose levels; unit emphasis on decreasing dosages; time when clients are encouraged to detoxify; average length of treatment. The data indicate that many units have treatment practices such as low average dose levels that are not effective according to the majority of previous studies. Units with higher average dose levels have longer average lengths of time in treatment. Steps should be taken to monitor and, if necessary, change the treatment practices of methadone units that are providing inadequate dose levels with little client input.


'Farmer's ear': sudden sensorineural hearing loss due to Chlamydia psittaci infection.
A case of sudden sensorineural hearing loss in association with a Chlamydia psittaci pneumonia is reported. Rapid recovery was seen when the patient was treated with high dose steroids and appropriate antibiotics. This is the first such case report in the literature.


Passive fluidic micro-sensor with all-optical readout realized using a direct laser writing technique.
In this Letter, we report on design and realization of solvent-sensitive microstructures based on three-dimensional periodic lattices fabricated in a polymeric photoresist. Sensing is based on reversible size change in polymeric microstructures upon immersion in wetting and non-wetting solvents. Its readout is achieved purely optically by observing modification of a Moiré pattern formed by grating-like deformable and rigid polymeric structures. A compact micro-sensor using these principles was realized using a direct laser writing technique in the photoresist. High sensitivity and easy optical readout of the sensor were demonstrated. In the future, sensors based on similar principles may find applications in microfluidic systems, such as lab-on-a-chip.


Informed consent: a risk management view. Council on Insurance.
In the best of all possible worlds, rules could be established that, if strictly followed, would eliminate litigation. Regrettably, that world does not exist. When dealing with informed consent many issues remain uncertain. Doctors must use their best professional judgment and hope that the jury will support it. Some risk management suggestions would include: Personally discuss the benefits and risks of the proposed treatment with the patient as well as the alternatives, and the consequences of nontreatment. Give the patient an opportunity to ask questions. Get a commitment from the patient to proceed. Use lay terms. Give no guarantees. If in doubt, disclose the risks. Use a signed informed consent form. Keep well-documented records. Consult your attorney about developing informed consent procedures that reflect state laws.


Incidental microvesicular steatosis due to valproic acid anticonvulsant therapy.
Valproic acid has been implicated in at least 100 cases of fatal acute liver failure. Most cases have occurred in patients less than 10 yr old; however, at least seven have involved adults. Microvesicular steatosis has been uniformly observed, but its incidence in less severe liver disease and in asymptomatic patients receiving valproate is unknown. We report two patients receiving maintenance valproate, one with resolving acute hepatitis C and the other with chronic persistent hepatitis C, with incidental microvesicular steatosis demonstrated on oil-red O stains. We conclude that microvesicular steatosis does not necessarily signify hepatotoxicity in patients on chronic valproic acid, and should not lead to discontinuation of the drug until other causes of acute or chronic liver disease have been excluded.


Robust oscillations within the interlocked feedback model of Drosophila circadian rhythm.
A mechanism for generating circadian rhythms has been of major interest in recent years. After the discovery of per and tim, a model with a simple feedback loop involving per and tim has been proposed. However, it is recognized that the simple feedback model cannot account for phenotypes generated by various mutants. A recent report by Glossop, Lyons & Hardin [Science286, 766 (1999)] on Drosophila suggests involvement of another feedback loop by dClk that is interlocked with per-tim feedback loop. In order to examine whether interlocked feedback loops can be a basic mechanism for circadian rhythms, a mathematical model was created and examined. Through extensive simulation and mathematical analysis, it was revealed that the interlocked feedback model accounts for the observations that are not explained by the simple feedback model. Moreover, the interlocked feedback model has robust properties in oscillations.


Colonic mucosubmucosal elongated polyp: a clinicopathologic study of 13 cases and review of the literature.
Colonic mucosubmucosal elongated polyp (CMSEP) is a distinctive non-neoplastic colorectal polyp characterized by pedunculated, elongated shape and is composed mainly of expanded submucosa with a normal mucosal lining. Only a small number of these polyps have been reported, exclusively from Japan. We report the clinicopathologic characteristics of 13 CMSEPs occurring in 11 patients, mostly from European ancestry. Ten of these polyps were resected during colonoscopy, and 3 were diagnosed in a patient who underwent sigmoid resection for diverticular disease. Among patients who had undergone a colonoscopy, 4 had altered bowel habit, and 1 suffered from abdominal discomfort; the other 5 patients had routine screening colonoscopy. Eight polyps were located in the sigmoid colon, 3 in the right colon, 1 in the rectosigmoid junction, and 1 in the descending colon. Polyp size ranged from 10 to 150 mm. Histologically, CMSEPs were characterized by unremarkable large bowel mucosa and submucosal stalk containing dilated thick-walled veins running parallel to the long axis of the polyp. Mucosal inflammation or fibromuscular proliferation characteristic of mucosal prolapse was absent. The pathogenesis of CMSEP may involve mechanical traction of the mucosa and the superficial submucosa during peristalsis in a fragile area of the colon. Despite the occasional large size, CMSEP is a benign lesion seldom leading to clinical complications.


Food-Related Impulsivity in Obesity and Binge Eating Disorder-A Systematic Update of the Evidence.
The specific eating pattern of Binge Eating Disorder (BED) patients has provoked the assumption that BED might represent a phenotype within the obesity spectrum that is characterized by increased impulsivity. Following the guidelines of the PRISMA statement (preferred reporting items for systematic reviews and meta-analyses), we here provide a systematic update on the evidence on food-related impulsivity in obese individuals, with and without BED, as well as normal-weight individuals. We separately analyzed potential group differences in the impulsivity components of reward sensitivity and rash-spontaneous behavior. Our search resulted in twenty experimental studies with high methodological quality. The synthesis of the latest evidence consolidates conclusions drawn in our initial systematic review that BED represents a distinct phenotype within the obesity spectrum that is characterized by increased impulsivity. Rash-spontaneous behavior in general, and specifically towards food, is increased in BED, while food-specific reward sensitivity is also increased in obese individuals without BED, but potentially to a lesser degree. A major next step for research entails the investigation of sub-domains and temporal components of inhibitory control in BED and obesity. Based on the evidence of impaired inhibitory control in BED, affected patients might profit from interventions that address impulsive behavior.


Regulation of bicarbonate-dependent ductular bile secretion assessed by lumenal micropuncture of isolated rodent intrahepatic bile ducts.
While intrahepatic bile duct epithelial cells secrete bile through transport of ions and water, the physiological mechanisms regulating ductular bile secretion are obscure, in part because of the lack of suitable experimental models. We report here the successful micropuncture of the lumen of isolated intrahepatic bile ducts and direct measurements of ductular ion secretion. Intact, polarized bile duct units (BDUs) were isolated from livers of normal rats by enzymatic digestion and microdissection. BDUs were cultured and mounted on a microscope in bicarbonate-containing buffer, and the lumens were microinjected with 2',7'-bis(2-carboxyethyl)-5-(and -6)carboxyfluorescein (BCECF)-dextran. Lumenal pH was measured by ratio imaging of BCECF fluorescence using digitized video fluorescent microscopy. After 36 hr in culture, the ends of BDUs sealed, forming closed compartments. After lumenal microinjection of BCECF-dextran, fluorescence was stable at the pH-insensitive wavelength, indicating no dye leakage. Serial changes in pH of extralumenal buffers containing pH-gradient collapsing ionophores allowed us to establish reliable standard curves relating fluorescence ratio to lumenal pH (r = 0.99; P < 0.001). By this approach, the basal pH inside the lumen of BDUs was 7.87 +/- 0.08 units (n = 9), 0.47 unit higher (P < 0.001) than the bathing buffer pH. Addition of 100 microM forskolin increased (P = 0.02) the lumenal pH from 7.78 +/- 0.06 to 7.97 +/- 0.06 units (n = 5); the forskolin effect was completely abolished by incubation of BDUs in HCO3-/CO2-free buffer. Moreover, forskolin caused a 50-fold increase in cAMP levels in BDUs. The observations are consistent with cAMP-dependent, active lumenal HCO3- secretion by BDUs. Furthermore, they demonstrate the suitability of the BDU model for studying regulatory and mechanistic aspects of ductular bile secretion.


Making strides in women's mental health care delivery in rural Ethiopia: demographics of a female outpatient psychiatric cohort at Jimma University Specialized Hospital (2006-2008).
This paper presents the delivery of mental health care to a sample of women living in Jimma, rural Ethiopia, and their access to mental health services. A total of 226 psychiatric charts were reviewed for women seen at Jimma University Specialized Hospital. The mental health charts included documentation ranging from one paragraph to a full note. No psychiatric chart recorded medication status, detailed substance abuse history, or a history of violence. Rendering appropriate mental health care for women requires concerted efforts by multiple stake holders. Using our results, we advance concrete and practical suggestions for improving women's mental health in rural Ethiopia. We point out that the health care system needs to be responsive, allowing for change starting with gender rights, so that rural women have access to basic mental health services.


A comparison of the bronchodilating effects of oxitropium bromide and fenoterol in patients with chronic obstructive pulmonary disease.
Oxitropium bromide is a novel anticholinergic bronchodilator agent. The purpose of this study was to compare the bronchodilating and cardiovascular effects of oxitropium (0.2 mg), fenoterol (0.4 mg), combined oxitropium and fenoterol (0.2 mg and 0.4 mg, respectively) over a 10-h test period. Fourteen patients with chronic obstructive pulmonary disease (COPD) (FEV1, 0.95 +/- 0.38L) were studied in a randomized, double-blind, placebo-controlled trial. Combined oxitropium and fenoterol produced significantly greater improvements in FEV1 over a time span of 15 min to 10 h and in the area under the time-FEV1 curve (AUC) than either oxitropium or fenoterol alone. The effects of oxitropium on both FEV1 and AUC values were similar to those of fenoterol. Oxitropium resulted in a greater increase in FEV1 than the placebo even after 10 h. In contrast; fenoterol produced a significant improvement in the FEV1 for only 15 min to 4 h. Oxitropium showed no adverse cardiovascular effects, whereas fenoterol was associated with an increased heart rate at 15 min and 1 h after the administration. We conclude that oxitropium bromide is an effective and safe bronchodilator for even elderly patients with COPD.


[From mice to men: the threat of hantavirus infections].
Hantaviruses are naturally hosted by small mammals. Virus transmission to humans can result in the development of Hemorrhagic Fever with Renal Syndrome (Asia, Europe) or Hantavirus Cardiopulmonary Syndrome (The Americas). Certain hantaviruses can cause life-treatening infections with case fatality indices up to about 50%. Different virus types have evolved which use different species of small mammals (mainly but not exclusively rodent species) as natural reservoirs. It is still unknown why these different hantaviruses exhibit a different virulence when transmitted to humans as accidental hosts. This paper reviews the nature and molecular epidemiology of hantaviruses as well as strategies to identify new virus types.


Quantitative multistandard RT-PCR assay using interspecies polymorphism.
The use of RT-PCR to quantify mRNA is often compromised by the variability of reverse-transcription and amplification reactions as well as by the difficulty of assessing the amount and/or the integrity of input RNA. Use of a competitor RNA or the coamplification of an endogenous standard are widespread methods of monitoring these steps. Taking advantage of both sequence conservation between homologous genes in related animal species and interspecific polymorphism, a protocol that may be regarded as a compromise between these two methods is described here. Total RNA samples, extracted from even minute amounts of tissue belonging to a first animal species, were supplemented with a constant amount of total RNA prepared from a second animal species, which thus acts as a multistandard source. The mixture was reverse-transcribed using hexa-random primers. Separate PCRs were then undertaken so that, for each mRNA of interest, products from both origins could be distinguished. Since the ratio between amplified mRNAs is constant in the standard preparation, an accurate normalization in the assay samples of most variations inherent to PCR is obtained. This protocol allows quantification of several mRNAs species, whose amounts may be very different, in a single cDNA preparation.


The effect of scapular protraction on isometric shoulder rotation strength in normal subjects.
To measure the effect of scapular protraction on isometric shoulder rotation strength, 20 normal subjects completed 2 maximal isometric internal and external rotation contractions in 2 scapular positions (scapula neutral [SN] and scapula protracted [SP]) from 3 arm positions (90 degrees internal rotation [IR], 45 degrees internal rotation [MR], and 90 degrees external rotation [ER]). Scapular protraction reduced shoulder rotation strength in 5 of 6 test positions (P < .0004), with significant interactions between scapular position and arm position (P < .001) and between scapular position and contraction type (P < .0001). Protraction significantly reduced IR strength by 13% to 24% relative to SN. The effect of SP on ER strength was more position-dependent, increasing strength by 6% in the IR position and decreasing it by 7% in the MR position and 20% in the ER position. In conclusion, acute changes in scapular position affect shoulder isometric IR and ER strength. The potential adverse effects of scapular protraction on shoulder rotation strength should be considered during the evaluation and treatment of shoulder pain.


A Novel Ferroptosis-Related Gene Signature to Predict Prognosis in Patients with Head and Neck Squamous Cell Carcinoma.
The clinical TNM staging system is currently used to evaluate the prognosis of head and neck squamous cell carcinoma (HNSCC). The 5-year survival rate for patients with HNSCC is less than 50%, which is attributed to the lack of reliable prognostic biomarkers. Ferroptosis-related genes (FRGs) regulate cancer initiation and progression. Therefore, we analyzed the correlation between FRGs and the clinical outcomes of patients with HNSCC. A typical prognostic model of FRGs for HNSCC was constructed using bioinformatics tools and data from public databases, including The Cancer Genome Atlas (TCGA), Gene Expression Omnibus (GEO), and GeneCards. The model was generated based on the following six FRGs: ATG5, PRDX6, OTUB1, FTH1, SOCS1, and MAP3K5. The accuracy of model prediction was analyzed systematically. The overall survival (OS) of the high-risk group was significantly lower than that of the low-risk group. The AUC for 1-year, 3-year, and 5-year survival were 0.645, 0.721, and 0.737, respectively, in the training set (TCGA cohort) and 0.726, 0.620, and 0.584, respectively, in the validation set (GSE65858). The multivariate Cox regression analysis revealed that the risk score was an independent prognostic factor for HNSCC. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses revealed that six FRGs were enriched in the ferroptosis pathway. A novel FRG prognostic signature model was established for HNSCC. The findings of this study reveal that FRGs are potential biomarkers for HNSCC.


Automated phantom analysis for gamma cameras and SPECT: A methodology for use in a clinical setting.
We introduce an automated, quantitative image analysis package for gamma camera and single photon emission computed tomography quality control. Our focus was to produce consistent methods that are feasible in clinical settings and use standard phantoms. Four gamma cameras were used to acquire planar images of four-quadrant bar phantoms and projection views of an American College of Radiology (ACR) phantom as part of a standard gamma camera quality control program. Images were sent to QC-Track® (Atirix Medical Systems, Inc., Minneapolis, MN, USA), which automatically placed predetermined regions of interest (ROIs) and performed analysis. For the bar phantom, a standard deviation (SD)-based modulation transfer function was calculated for a circular ROI in each quadrant. The bar widths at various MTF values were reported using linear interpolation as applicable. For the ACR phantom, the contrast-to-noise ratio (CNR) for each sphere, a modulation for each rods section, and a percent deviation for uniformity ROIs was calculated. Spheres corresponding to a CNR of 3, and the rod size at various modulations were also reported using linear interpolation. Visual analysis was performed by three medical physicists to evaluate interobserver variability and correlation to quantitative values. Analysis of the bar phantom showed predictable differences with changes in matrix size and bar width and showed consistency over similar acquisitions over the course of the study. Analysis of the ACR Phantom showed increasing CNR and modulation with increasing sphere and rod diameter, as expected. For both phantoms, quantitative values from linear interpolation correlated well with visual analysis. Our automated method for quantitative image analysis is consistent and shows increased precision and sensitivity when compared to standard visual methods. Thresholds correspond well with visual analysis and previous guidelines for observer visibility (e.g., Rose criterion), making our framework suitable for routine use in a nuclear medicine department.


Retained Digital Flexible Ureteroscope During Percutaneous Nephrolithotomy.
Background: During percutaneous nephrolithotomy, retrograde flexible ureteroscopy can be utilized to facilitate repositioning of stones, enable direct vision percutaneous access, minimize radiation exposure, reduce operating times, and improve stone-free rates. Although advancements in technique and flexible ureteroscope technology for the past decades have rendered complications rare, herein we report a case of a retained ureteroscope during percutaneous nephrolithotomy that was effectively managed endoscopically. Case Presentation: A 59-year-old Caucasian gentleman with a history of recurrent bilateral nephrolithiasis presents for a left-sided percutaneous nephrolithotomy for a large stone burden >4 cm. A ureteral access sheath was used and retrograde ureteroscopy was performed to first reposition several stones into the renal pelvis. During manipulation, we were unexpectedly unable to retract the ureteroscope from the access sheath. We describe procedural details leading up to the event and subsequent intraoperative management using an antegrade approach. Conclusion: A retained flexible ureteroscope is a rare but serious intraoperative complication that may require invasive open surgical management. However, careful endoscopic management may be feasible in select cases, allowing for preservation of ureter and instrument.


In Vitro Biocompatibility of Several Children's Toothpastes on Human Gingival Fibroblasts.
The European Academy of Paediatric Dentistry has recommended fluoride toothpastes from the eruption of the first teeth in children. Toothpastes stay in the mouth in contact with human gingival fibroblasts (hGFs) for a long time. Thus, the objective of this study was to compare the cytotoxicity of five different commonly used children's toothpastes: Oral B Kids +3 (Procter & Gamble, Alicante, Spain), Fluor Kin Calcium (Kin, Madrid, Spain), PHB Junior (PHB, Barcelona, Spain), Colgate +3 (Colgate Palmolive, Madrid, Spain) and Vitis Kids (Dentaid, Valencia, Spain) on hGFs. The children's toothpastes were exposed to hGFs at different concentrations (1:1, 1:2, 1:4). Afterwards, several tests were performed: MTT assays, cell cycle analyses, cell cytoskeleton staining assays, apoptosis/necrosis assays, and ICP-MS and ion chromatography. Oral B displayed the lowest cytotoxicity and was the toothpaste with the highest fluoride ion release; meanwhile, the other toothpastes were cytotoxic (*** p < 0.0001); Fluor Kin being the one with the lowest fluoride ion release. Among all the toothpastes analyzed, Oral B exhibited the best results in vitro in terms of biocompatibility. Future evaluations, both in vitro and in vivo, are required to confirm the biocompatibility of sodium lauryl sarcosinate and sodium lauryl sulfate containing toothpastes.


[Triple therapy regimens involving H2 blockaders for therapy of Helicobacter pylori infections].
Comparison of ranitidine and lansoprazole in short-term low-dose triple therapy for Helicobacter pylori infection. To evaluate the efficacy and safety of two 1-week low-dose triple-therapy drug regimens involving antisecretory drugs for Helicobacter pylori infection, 99 patients with H. pylori infection were treated with either lansoprazole (LPZ) or ranitidine (RNT) used together with clarithromycin (CAM) and metrinidazole (MTZ). The drug combination and administration periods in the PPI group were LPZ 30 mg, CAM 400 mg, MTZ 500 mg (LCM group). The ranitidine group received RNT 300 mg, CAM 400 mg, MTZ 500 mg (RCM group). The cure rate of H. pylori infection was 88% in the LCM group; 95% CI 79-97 and 92% in the RCM group; 95% CI 84-99.


A comparison of multiple methods for the identification of children with reading disabilities.
There has been considerable discussion of the methods used for the identification of children with reading disabilities. This study examined three different methods that could be used in the identification of children with reading disabilities and their consistency with teacher ratings of behaviors believed to be associated with reading disabilities. Standardized, norm-referenced measures of achievement, phonological processing measures, and curriculum-based measures of reading fluency were used with 40 children in Grades 1 and 2. Comparisons were made to determine which measures, if any, differentiated between children referred for reading disabilities (n = 20) and children who were progressing typically in reading in their general education classroom settings (n = 20). The results indicated significant between-group differences on standardized, norm-referenced measures of reading recognition, word attack, and comprehension; phonological measures of blending nonwords and elision; and reading fluency. Teacher ratings on the Dyslexia Screening Instrument were consistent with teacher beliefs regarding children's progress in reading. All measures were found to correlate significantly with each other. However, correlations were generally in the moderate range, suggesting that the measures used did not measure the same reading skills or, in the case of phonological processing, the underlying abilities believed to be necessary for reading. Given the differences in tasks and the moderate correlations, it is likely that the choice of measures may affect the conclusions reached regarding a student's reading ability.


Tumor Progression of Non-Small Cell Lung Cancer Controlled by Albumin and Micellar Nanoparticles of Itraconazole, a Multitarget Angiogenesis Inhibitor.
Itraconazole (ITA), an old and widely prescribed antifungal drug with excellent safety profile, has more recently been demonstrated to be a multitarget antiangiogenesis agent affecting multiple angiogenic stimulatory signals and pathways, including vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), vascular endothelial growth factor receptor 2 (VEGFR2) glycosylation, and mammalian target of rapamycin (mTOR). In this study, we developed two nanoparticle formulations, i.e., polymer micelles (IP2K) and albumin nanoparticles (IBSA), to solubilize the extremely hydrophobic and insoluble ITA to allow intravenous administration and pharmacokinetics (PK)/pharmacodynamics (PD) comparisons. Although none of the formulations showed strong antiproliferation potency against non-small cell lung cancer (NSCLC) cells in vitro, when administrated at the equivalent ITA dose to a NSCLC patient-derived xenograft (PDX) model, IBSA retarded while IP2K accelerated the tumor growth. We attributed the cause of this paradox to formulation-dependent PK and vascular manipulation: IBSA demonstrated a more sustained PK with a Cmax of 60-70% and an AUC ∼2 times of those of IP2K, and alleviated the tumor hypoxia presumably through vascular normalization. In contrast, the high Cmax of IP2K elevated tumor hypoxia through a strong angiogenesis inhibition, which could have aggravated cancer aggressiveness and accelerated tumor growth. Furthermore, IBSA induced minimal hepatic and hematologic toxicities compared to IP2K and significantly enhanced the in vivo tumor inhibition activity of paclitaxel albumin nanoparticles when used in combination. These findings suggest that formulation and pharmacokinetics are critical aspects to be considered when designing the ITA angiogenesis therapy, and IBSA could potentially be assessed as a novel and safe multitarget angiogenesis therapy to be used in combination with other anticancer agents.


Transcranial direct current stimulation (tDCS) of the primary motor cortex and robot-assisted arm training in chronic incomplete cervical spinal cord injury: A proof of concept sham-randomized clinical study.
After cervical spinal cord injury, current options for treatment of upper extremity motor functions have been limited to traditional approaches. However, there is a substantial need to explore more rigorous alternative treatments to facilitate motor recovery. To demonstrate whether anodal-primary motor cortex (M1) excitability enhancement (with cathodal-supra orbital area) (atDCS) combined with robot-assisted arm training (R-AAT) will provide greater improvement in contralateral arm and hand motor functions compared to sham stimulation (stDCS) and R-AAT in patients with chronic, incomplete cervical spinal cord injury (iCSCI). In this parallel-group, double-blinded, randomized and sham-controlled trial, nine participants with chronic iCSCI (AIS C and D level) were randomized to receive 10 sessions of atDCS or stDSC combined with R-AAT. Feasibility and tolerability was assessed with attrition rate and occurrence of adverse events, Changes in arm and hand function were assessed with Jebson Taylor Hand Function Test (JTHFT). Amount of Use Scale of Motor Activity Log (AOU-MAL), American Spinal Injury Association Upper Extremity Motor Score and Modified Ashworth Scale (MAS) at baseline, after treatment, and at two-month follow-up. None of the participants missed a treatment session or dropped-out due to adverse events related to the treatment protocol. Participants tended to perform better in JTHFT and AOU-MAL after treatment. Active group at post-treatment and two-month follow-up demonstrated better arm and hand performance compared to sham group. These preliminary findings support that modulating excitatory input of the corticospinal tracts on spinal circuits may be a promising strategy in improving arm and hand functions in persons with incomplete tetraplegia. Further study is needed to explore the underlying mechanisms of recovery.


Effects of crystallization and non-lattice oxygen atoms on Cu(x)O-based resistive switching memory.
In this work, the effects of crystallization and non-lattice oxygen atoms on the Cu(x)O-based memory device are investigated. The 150 degrees C-deposited Cu(x)O film possesses a larger amount of non-lattice oxygen atoms than those deposited at the higher temperatures, leading to the formation of AIOy interface layer during the sputtering deposition of Al top electrode. Resistive switching occurring within the interface layer is easily controlled, so the set and reset voltages are decreased. In addition, it is demonstrated that the set and reset processes agree with the formation and rupture of a conductive filament in the Cu(x)O film. The 150 degrees C-deposited Cu(x)O-based memory device with good non-volatility is possibly used in the next-generation non-volatile memory.


Evaluation of immunogenicity of Campylobacter strains isolated from ovine abortions by laboratory test systems.
A microagglutination test and in vitro serum bactericidal assay were used for testing cross-reactions and cross-bactericidal activities of 15 Campylobacter strains isolated from ovine abortions. Antisera against certain strains of C. fetus subsp. fetus and C. jejuni showed higher microagglutination titres and bactericidal activity against homologous and/or heterologous strains. In both tests, cross-reactions and bactericidal activity were higher among the strains of homologous species than among those of heterologous species. The pregnant guinea-pig model was used for testing the efficacy of experimental campylobacter vaccines. Monovalent bacterins prepared with either C. fetus subsp. fetus or C. jejuni did not produce immunity to cross-challenge with the opposite species. Guinea-pigs immunized with the bivalent bacterin were found to be immune to both groups of Campylobacter when subsequently challenged. In vaccinated guinea-pigs, their in vivo ability to overcome infection is reflected in the in vitro bactericidal capacity of homologous rabbit antiserum.


Influence of epinastine hydrochloride, an H1-receptor antagonist, on the function of mite allergen-pulsed murine bone marrow-derived dendritic cells in vitro and in vivo.
There is established concept that dendritic cells (DCs) play essential roles in the development of allergic immune responses. However, the influence of H(1) receptor antagonists on DC functions is not well defined. The aim of the present study was to examine the effect of epinastine hydrochloride (EP), the most notable histamine H(1) receptor antagonists in Japan, on Dermatophagoides farinae (Der f)-pulsed mouse bone marrow-derived DCs in vitro and in vivo. EP at more than 25 ng/mL could significantly inhibit the production of IL-6, TNF-alpha and IL-10 from Der f-pulsed DCs, which was increased by Der f challenge in vitro. On the other hand, EP increased the ability of Der f-pulsed DCs to produce IL-12. Intranasal instillation of Der f-pulsed DCs resulted in nasal eosinophilia associated with a significant increase in IL-5 levels in nasal lavage fluids. Der f-pulsed and EP-treated DCs significantly inhibited nasal eosinophila and reduced IL-5. These results indicate that EP inhibits the development of Th2 immune responses through the modulation of DC functions and results in favorable modification of clinical status of allergic diseases.


Characterization of an HIV-1 isolate displaying an apparent absence of virion-associated reverse transcriptase activity.
In characterizing a group of independent human immunodeficiency virus (HIV-1) isolates, we noted that certain isolates had anomolously low levels of virion-associated reverse transcriptase activity. In an attempt to understand the basis of this phenomenon, we examined in detail one such isolate, HIV-1G. We found correctly processed forms of the viral reverse transcriptase in virions as well as processed forms of other viral proteins, suggesting that viral proteins are both expressed and properly processed. We have detected a nuclease activity associated with the outer face of the HIV-1G envelope. This nuclease degrades the DNA product generated during the reverse transcription assay. The nuclease activity is more sensitive to mild protein denaturation than is the viral reverse transcriptase, and it is stimulated by the presence of Ca2+. The amount of virion-associated nuclease activity relative to reverse transcriptase activity varies between virus isolates and can vary also for one isolate during virus spread through a culture. The origin of the nuclease activity is unknown but is presumed to be cellular. The variability in amount of nuclease activity may reflect variability in the interaction of the virus with different cellular components during maturation.


Thinking of England and taking care: family building strategies and infant mortality in England and Wales, 1891-1911.
"Debates concerning the origins and development of the late nineteenth- to early twentieth-century declines in marital fertility and infant mortality in England and Wales have been centred largely on the material provided by answers to the ¿special' questions in the 1911 census. In their published form these figures have restricted researchers to an examination of large scale geographic and social class differences in the levels and rates of decline of the two phenomena. This paper outlines research conducted on a sample of individual census returns from the 1911 census. From this data it becomes clear that for Victorian and Edwardian England ¿where one lived' was rather more important than ¿who one was' in determining both family building strategies and the survival of those children born."


Fibromyalgia Patients Are Not Only Hypersensitive to Painful Stimuli But Also to Acoustic Stimuli.
Fibromyalgia is a chronic widespread pain syndrome associated with hypersensitivity to nociceptive stimuli. This increased sensitivity of FM patients has been associated with central sensitization of dorsal horn neurons. Increasing evidence, however, suggests that the mechanisms of FM hypersensitivity not only affect pain but include light, smell, and sound. We hypothesized that supraspinal augmentation of sensory input including sound represent a hallmark of FM. We tested 23 FM patients and 28 healthy controls (HC) for sensory augmentation of nociceptive and non-nociceptive sensations: For assessment of nociceptive augmentation we used sensitivity adjusted mechanical and heat ramp & hold stimuli and for assessment of sound augmentation, we applied wideband noise stimuli using a random-staircase design. Quantitative sensory testing demonstrated increased heat and mechanical pain sensitivity in FM participants (P < .001). The sound pressures needed to report mild, moderate, and intense sound levels were significantly lower in FM compared to HC (P < .001), consistent with auditory augmentation. FM patients are not only augmenting noxious sensations but also sound, suggesting that FM augmentation mechanisms are not only operant in the spinal cord but also in the brain. Whether the central nervous system mechanisms for auditory and nociceptive augmentation are similar, needs to be determined in future studies. PERSPECTIVE: This study presents QST evidence that the hypersensitivity of FM patients is not limited to painful stimuli but also to innocuous stimuli like sound. Our results suggest that abnormal brain mechanisms may be responsible for the increased sensitivity of FM patients.


[Home hospice care at a clinic].
For the people who want to stay at home until their last day, the primary doctor and clinic where they were diagnosed will be the most reliable supports. We have been operating a 19 bed clinic since 1996. In these three years, we have established what we call a "combination palliative care system." A team composed of two doctors, 13 nurses, 3 care aids, a social worker, and a counselor provides home care services as well as outpatient and inpatient care. From April, 1998 to March, 1999, 59 patients died of cancer. Among them, 25 patients died at home. Their primary cancers were lung (7), colon (3), pancreatic (2), breast (2), ovarian (2), brain (1), stomach (1), hepatoma (1), neck (1) and others. First of all, sufficient consultation with patients and family makes this care successful. Through this, the patient can choose his style of care. The whole staff is involved in this care in turn, so that all of us become acquainted with each patient. Home care includes: 1) medical and nursing service available 24 hours a day, 2) activation of social resources for the support of the patient user, 3) constructive cooperation with relevant institutions, 4) relieving the patient's physical and mental suffering, 5) aroma therapy, oil massage, hair cuts and music therapy, and 6) support by volunteers. In this way, as a neighborhood clinic, the combination palliative care system is valuable.


The role of research in molecular entomology in the fight against malaria vectors.
The text summarizes the principal current fields of investigation and the recent achievements of the research groups presently contributing to the Molecular Entomology Cluster of the Italian Malaria Network. Particular emphasis is given to the researches with a more direct impact on the fight against malaria vectors.


Comparison between Aptima Assays (Hologic) and the Allplex STI Essential Assay (Seegene) for the diagnosis of Sexually transmitted infections.
Sexually transmitted infections (STIs) remain a worldwide problem and a severe threat to public health. The purpose of this study was to compare Aptima® Assays (Hologic®) and the Allplex™ STI Essential Assay (Seegene®) for the simultaneous detection of Chlamydia trachomatis, Neisseria gonorrhoeae, Trichomonas vaginalis and Mycoplasma genitalium in clinical practice. The Aptima® assays (Hologic®) are based on a transcription-mediated amplification (TMA) method. The Allplex™ STI Essential assay (Seegene®) is based on a multiplex Real-Time PCR (RT-PCR) method. A total of 622 clinical samples from different anatomical sites were tested using both methods. A total of 88 (14.1%) and 66 (10.6%) positive samples were found for any of the TMA assays used and for the RT-PCR assay, respectively. Aptima® assays showed a slightly higher rate of positive results for all pathogens except for T. vaginalis, the results of which were similar to those obtained with Allplex™. The most commonly detected pathogen was C. trachomatis (37 samples; 5.9% using TMA assays) and the anatomical site with the highest prevalence of microorganisms was a non-urogenital site, the pharynx (27 positive samples; 4.3%). Using the Aptima® assays as reference method, the comparison showed that the average specificity of multiplex RT-PCR was 100.0% for the four pathogens. However an average sensitivity of 74.5% was observed, showing 95.2% (CI95%; 93.6-96.9) of overall concordance (κ = 0.80). In conclusion, the Aptima® assays show a higher sensitivity on a wide range of sample types compared to the Allplex™ assay.


Instrument to detect syncope and the onset of shock.
Currently the diagnosis of hemorrhagic shock is essentially clinical, relying on the expertise of nurses and doctors. One of the first measurable physiological changes that marks the onset of hemorrhagic shock is a decrease in capillary blood flow. Diffuse correlation spectroscopy (DCS) quantifies this decrease. DCS collects and analyzes multiply scattered, coherent, near infrared light to assess relative blood flow. This work presents a preliminary study using a DCS instrument with human subjects undergoing a lower body negative pressure (LBNP) protocol. This work builds on previous successful DCS instrumentation development and we believe it represents progress toward understanding how DCS can be used in a clinical setting.


Alternative approaches for estimating prevalence in epidemiologic surveys with two waves of respondents.
Estimates of prevalence in epidemiologic surveys are prone to bias due to selective response. Therefore, much effort is devoted to reduce the number of nonrespondents. For example, individuals who do not respond in the first round of recruitment in mail surveys are usually contacted a second (or even third or fourth) time yielding consecutive waves of responses. Yet this sequence of waves is often neglected in epidemiologic analyses in that prevalence is simply estimated as the proportion of trait-positive individuals among the total group of respondents. This paper investigates alternative estimates of prevalence that might be used in surveys with two waves of respondents. The estimates are based on different assumptions on the relation of response rates with the trait of interest. As this relation is likely to vary from survey to survey depending on the specific circumstances under which the recruitment of participants is conducted, none of the estimates is universally preferable. The performance of the different estimates is assessed in a variety of hypothetical and empirical examples, and strategies are discussed to make the best use of the different estimates in the analysis of epidemiologic studies.


Three new diterpene alkaloids from the roots of Aconitum nagarum var. lasiandrum.
Three new diterpene alkaloids, 16,17-dihydro-12 beta,16 beta-epoxynapelline (1), N-deethyl- N-methyl-12-epi-napelline (2) and 11- epi-16 alpha,17-dihydroxylepenine (3), along with twenty-six known alkaloids (4 - 29) were isolated from the roots of Aconitum nagarum var. lasiandrum. The chemical structures of the new compounds were established by HR-MS as well as 1D- and 2D-NMR spectroscopic analysis. The absolute stereochemistry of 1 was confirmed by X-ray crystallography.


Differential proteomics analysis of the surface heterogeneity of dextran iron oxide nanoparticles and the implications for their in vivo clearance.
In order to understand the role of plasma proteins in the rapid liver clearance of dextran-coated superparamagnetic iron oxide (SPIO) in vivo, we analyzed the full repertoire of SPIO-binding blood proteins using novel two-dimensional differential mass spectrometry approach. The identified proteins showed specificity for surface domains of the nanoparticles: mannan-binding lectins bound to the dextran coating, histidine-rich glycoprotein and kininogen bound to the iron oxide part, and the complement lectin and contact clotting factors were secondary binders. Nanoparticle clearance studies in knockout mice suggested that these proteins, as well as several previously identified opsonins, do not play a significant role in the SPIO clearance. However, both the dextran coat and the iron oxide core remained accessible to specific probes after incubation of SPIO in plasma, suggesting that the nanoparticle surface could be available for recognition by macrophages, regardless of protein coating. These data provide guidance to rational design of bioinert, long-circulating nanoparticles.


Antitumor activity of pimasertib, a selective MEK 1/2 inhibitor, in combination with PI3K/mTOR inhibitors or with multi-targeted kinase inhibitors in pimasertib-resistant human lung and colorectal cancer cells.
The RAS/RAF/MEK/MAPK and the PTEN/PI3K/AKT/mTOR pathways are key regulators of proliferation and survival in human cancer cells. Selective inhibitors of different transducer molecules in these pathways have been developed as molecular targeted anti-cancer therapies. The in vitro and in vivo anti-tumor activity of pimasertib, a selective MEK 1/2 inhibitor, alone or in combination with a PI3K inhibitor (PI3Ki), a mTOR inhibitor (everolimus), or with multi-targeted kinase inhibitors (sorafenib and regorafenib), that block also BRAF and CRAF, were tested in a panel of eight human lung and colon cancer cell lines. Following pimasertib treatment, cancer cell lines were classified as pimasertib-sensitive (IC50 for cell growth inhibition of 0.001 µM) or pimasertib-resistant. Evaluation of basal gene expression profiles by microarrays identified several genes that were up-regulated in pimasertib-resistant cancer cells and that were involved in both RAS/RAF/MEK/MAPK and PTEN/PI3K/AKT/mTOR pathways. Therefore, a series of combination experiments with pimasertib and either PI3Ki, everolimus, sorafenib or regorafenib were conducted, demonstrating a synergistic effect in cell growth inhibition and induction of apoptosis with sustained blockade in MAPK- and AKT-dependent signaling pathways in pimasertib-resistant human colon carcinoma (HCT15) and lung adenocarcinoma (H1975) cells. Finally, in nude mice bearing established HCT15 and H1975 subcutaneous tumor xenografts, the combined treatment with pimasertib and BEZ235 (a dual PI3K/mTOR inhibitor) or with sorafenib caused significant tumor growth delays and increase in mice survival as compared to single agent treatment. These results suggest that dual blockade of MAPK and PI3K pathways could overcome intrinsic resistance to MEK inhibition.