Factor analysis and discriminant analysis of wastewater quality in Vidyaranyapuram sewage treatment plant, Mysore, India: a case study.
Wastewater treatment plant monitoring is an essential part of effective wastewater management. The analysis of eight physico-chemical parameters of untreated wastewater was carried out at Vidyaranyapuram sewage treatment plant, Mysore, India. Factor analysis (FA) was applied to the untreated wastewater data matrix, and pollution was found to be the most contributing factor, explaining 22.31% of the total variance (chloride, biochemical oxygen demand, chemical oxygen demand and total dissolved solids). The second most contributing factor was found to be nitrification which explained 21.11% of the total variance (pH and nitrate), whereas the salinization factor contributed 16.98% of the total variance (total solids and total suspended solids). FA regression scores could not satisfactorily classify the data matrix with respect to the seasonal variations. Discriminant analysis (DA) was used to find the seasonal variations in the data matrix, and the standard mode DA explained 66.6% of total variance by grouping the cases with respect to seasons.


Drosophila Gene Expression Pattern Annotation through Multi-Instance Multi-Label Learning.
The Berkeley Drosophila Genome Project (BDGP) has produced a large number of gene expression patterns, many of which have been annotated textually with anatomical and developmental terms. These terms spatially correspond to local regions of the images; however, they are attached collectively to groups of images, such that it is unknown which term is assigned to which region of which image in the group. This poses a challenge to the development of the computational method to automate the textual description of expression patterns contained in each image. In this paper, we show that the underlying nature of this task matches well with a new machine learning framework, Multi-Instance Multi-Label learning (MIML). We propose a new MIML support vector machine to solve the problems that beset the annotation task. Empirical study shows that the proposed method outperforms the state-of-the-art Drosophila gene expression pattern annotation methods.


A pillar[5]arene-based multiple-stimuli responsive metal-organic gel was constructed for facile removal of mercury ions.
A thioacetohydrazide functionalized pillar[5]arene was synthesized, which could further assemble into a linear supramolecular metal-organic polymer upon adding Zn2+. Furthermore, the obtained linear supramolecular metal-organic polymer could self-assemble to form a fluorescent supramolecular metal-organic gel at high concentration. When TBAOH was added to the viscous solution at high temperature, the obtained solution could not form a supramolecular metal-organic gel upon cooling. More importantly, when Hg2+ ions are added to the metal-organic gel, the strong blue fluorescence is clearly quenched, and this metal-organic gel (xerogel) could effectively remove Hg2+ from water. Simultaneously, a thin film based on the metal-organic gel was prepared, which was confirmed to be a convenient test kit for detecting Hg2+.


MYB Translocation Status in Salivary Gland Epithelial-Myoepithelial Carcinoma: Evaluation of Classic, Variant, and Hybrid Forms.
Epithelial-myoepithelial carcinoma (EMC) is a malignant salivary gland neoplasm comprised of a biphasic arrangement of inner luminal ductal cells and outer myoepithelial cells. Adenoid cystic carcinoma (AdCC) is also a biphasic tumor comprised of ductal and myoepithelial cells, but these components tend to be arranged in a more cribriform pattern. The occurrence of "hybrid carcinomas" that show mixed patterns of EMC and AdCC raises questions about the relationship of these morphologically overlapping but clinically distinct tumors. AdCCs frequently harbor MYB-NFIB gene fusions. Mapping of EMCs (including hybrid forms with an AdCC component) for this fusion could help clarify the true nature of EMC as a distinct entity or simply as some variant form of AdCC. Twenty-nine cases of EMC were evaluated including 15 classic low-grade EMCs, 7 intermediate-grade EMCs, 2 EMCs with myoepithelial anaplasia, 1 EMC with high-grade transformation, and 4 hybrid EMCs with an AdCC component. Break apart fluorescence in situ hybridization for MYB was performed, as was MYB immunohistochemistry. For the hybrid carcinomas and those with high-grade transformation, the divergent tumor components were separately analyzed. A MYB translocation was identified in 5 of 28 (18%) tumors including 3 of 4 (75%) hybrid carcinomas and 2 of 7 (29%) intermediate-grade EMCs. For the positive hybrid carcinomas, the fusion was detected in both the EMC and AdCC components. The MYB fusion was not detected in any of the classic EMCs (0/15) or in any of the EMCs with myoepithelial anaplasia (0/2) or high-grade transformation (0/1). The fluorescence in situ hybridization assay was unsuccessful in 1 case. MYB immunostaining was seen in 5 of 5 fusion-positive cases, and also 9 of 23 fusion-negative tumors. Classic low-grade EMCs are genetically distinct from AdCCs in that they do not harbor MYB fusions. The presence of a MYB fusion in EMCs showing hybrid features of AdCC or exhibiting highly infiltrative growth points to a subset of these tumors that may well be true AdCCs masquerading as EMCs.


Protein Supplementation Has Minimal Effects on Muscle Adaptations during Resistance Exercise Training in Young Men: A Double-Blind Randomized Clinical Trial.
To our knowledge the efficacy of soy-dairy protein blend (PB) supplementation with resistance exercise training (RET) has not been evaluated in a longitudinal study. Our aim was to determine the effect of PB supplementation during RET on muscle adaptation. In this double-blind randomized clinical trial, healthy young men [18-30 y; BMI (in kg/m(2)): 25 ± 0.5] participated in supervised whole-body RET at 60-80% 1-repetition maximum (1-RM) for 3 d/wk for 12 wk with random assignment to daily receive 22 g PB (n = 23), whey protein (WP) isolate (n = 22), or an isocaloric maltodextrin (carbohydrate) placebo [(MDP) n = 23]. Serum testosterone, muscle strength, thigh muscle thickness (MT), myofiber cross-sectional area (mCSA), and lean body mass (LBM) were assessed before and after 6 and 12 wk of RET. All treatments increased LBM (P < 0.001). ANCOVA did not identify an overall treatment effect at 12 wk (P = 0.11). There tended to be a greater change in LBM from baseline to 12 wk in the PB group than in the MDP group (0.92 kg; 95% CI: -0.12, 1.95 kg; P = 0.09); however, changes in the WP and MDP groups did not differ. Pooling data from combined PB and WP treatments showed a trend for greater change in LBM from baseline to 12 wk compared with MDP treatment (0.69 kg; 95% CI: -0.08, 1.46 kg; P = 0.08). Muscle strength, mCSA, and MT increased (P < 0.05) similarly for all treatments and were not different (P > 0.10) between treatments. Testosterone was not altered. PB supplementation during 3 mo of RET tended to slightly enhance gains in whole-body and arm LBM, but not leg muscle mass, compared with RET without protein supplementation. Although protein supplementation minimally enhanced gains in LBM of healthy young men, there was no enhancement of gains in strength. This trial was registered at clinicaltrials.gov as NCT01749189.


Facilitating chlamydia testing among young people: a randomised controlled trial in cyberspace.
Chlamydia notifications have been rising in Australia for over a decade and are highest in young people. This study aimed to evaluate the impact of an internet-based intervention on chlamydia testing among young people 16-25 years. In this randomised controlled trial, recruitment, data collection, study interventions and follow-up occurred entirely in cyberspace, facilitated by a website. Eligible participants were aged 16-25 years and resided in Australia. The intervention group received personalised emails inviting interaction about chlamydia testing, while the control group received regular impersonal emails. Primary outcome was self-reported chlamydia testing at 6-month follow-up; secondary outcomes were condom use and changes in knowledge and attitudes. 704 young people completed baseline information, 40 were excluded and five withdrew prior to follow-up. The follow-up rate was 47.3% overall. In the intervention group, 40.6% (95% CI 30.7% to 51.1%) reported having had a chlamydia test at follow-up compared with 31.0% (95% CI 24.8% to 37.2%) in the control group (p=0.07). A per-protocol analysis found that those who engaged in email interaction were more likely to report chlamydia test uptake compared with those in the control group (52.5%, 95% CI 39.3 to 65.4% cf 31.0%, 95% CI 24.8% to 37.2%, p=0.002). There were no differences in secondary outcomes between groups. This is the first randomised controlled trial undertaken in cyberspace to promote chlamydia testing. E-technology may be useful in promoting chlamydia testing and healthcare seeking behaviour in young people.


Terrestrial plant methane production and emission.
In this minireview, we evaluate all experimental work published on the phenomenon of aerobic methane (CH(4) ) generation in terrestrial plants and plant. Clearly, despite much uncertainty and skepticism, we conclude that the phenomenon is true. Four stimulating factors have been observed to induce aerobic plant CH(4) production, i.e. cutting injuries, increasing temperature, ultraviolet radiation and reactive oxygen species. Further, we analyze rates of measured emission of aerobically produced CH(4) in pectin and in plant tissues from different studies and argue that pectin is very far from the sole contributing precursor. In consequence, scaling up of aerobic CH(4) emission needs to take into consideration other potential sources than pectin. Due to the large uncertainties related to effects of stimulating factors, genotypic responses and type of precursors, we conclude that current attempts for upscaling aerobic CH(4) into a global budget is inadequate. Thus it is too early to draw the line under the aerobic methane emission in plants. Future work is needed for establishing the relative contribution of several proven potential CH(4) precursors in plant material.


Differential susceptibility to activation-induced apoptosis among peripheral Th1 subsets: correlation with Bcl-2 expression and consequences for AIDS pathogenesis.
It has been proposed that HIV infection is associated with an imbalance in Th1 and Th2 subsets. Recent reports indicate that Th1 and Th2 effectors differ in their susceptibility to activation-induced apoptosis. To determine whether increased T cell apoptosis in HIV-infected patients contributes to alterations in cytokine synthesis, we performed single-cell analysis of type 1 and type 2 cytokine production by CD4 and CD8 T cells, simultaneously with detection of apoptosis. We demonstrate that a differential alteration in representation of Th1 subsets, rather than commitment of T cells to secrete Th2 cytokines, occurs throughout HIV infection. A significant decrease in the number of IL-2- or TNF-alpha-producing T cells was observed, whereas those producing IFN-gamma remained preserved. Furthermore, there is a gradient of susceptibility to activation-induced apoptosis (IL-2 < IFN-gamma < TNF-alpha) among the different Th1 subsets. This gradient was detected in both CD4 and CD8 subsets, as well as in control donors and HIV-infected patients, in whom the susceptibility to apoptosis of IL-2 and IFN-gamma producers was increased compared with controls. This differential intrinsic apoptosis susceptibility of Th1 effectors was found to be tightly regulated by Bcl-2 expression. In HIV-infected persons, disappearance of IL-2-producing T cells was a good indicator of disease progression and was correlated with the progressive shrinkage of the CD4+ CD45RA+ T cell compartment and a gradual increased susceptibility to activation-induced apoptosis of the IL-2-producing subset. This close relationship between the CD45RA/CD45R0 ratio, the level of type 1 cytokine production, and susceptibility to apoptosis should be considered in HIV-infected patients under antiviral or immune-based therapies.


Electrophysiological evidence for retrieval mode immediately after a task switch.
It has been suggested that retrieving episodic information can involve adopting a cognitive state or set: retrieval mode. In a series of studies, an event-related potential (ERP) index of retrieval mode has been identified in designs which cue participants on a trial-by-trial basis to switch between preparing for and then completing an episodic or non-episodic retrieval task. However, a confound in these studies is that along with task type the content of what is to be retrieved has varied. Here we examined whether the ERP index of retrieval mode remains when the contents of an episodic and non-episodic task are highly similar - both requiring a location judgement. In the episodic task participants indicated the screen location where words had been shown in a prior study phase (left/right/new); whereas in the perceptual task they indicated the current screen location of the word (top/middle/bottom). Consistent with previous studies the ERPs elicited while participants prepared for episodic retrieval were more positive-going at right-frontal sites than when they prepared for the perceptual task. This index was observed, however, on the first trial after participants had switched tasks, rather than on the second trial, as has been observed previously. Potential reasons for this are discussed, including the critical manipulation of similarity in contents between tasks, as well as the use of a predictable cue sequence.


COX-2 promoter polymorphisms and the association with prostate cancer risk in South African men.
Cyclooxygenase-2 (COX-2) converts arachidonic acid to prostaglandins, which are important mediators of cell proliferation and inflammation. Evidence indicates that COX-2 plays a role in carcinogenesis and that it is over-expressed in prostate tumours. We investigated the role of COX-2 variants in prostate cancer in a case-control study of South African Coloured men, consisting of 151 cases and 134 controls. The genotype frequencies of four single-nucleotide polymorphisms (SNPs) in the COX-2 promoter were initially determined in 50 control subjects. One SNP, rs20417 (-899G>C), was monomorphic and excluded from further investigation. Three SNPs, rs3918304 (-1285A>G), rs20415 (-1265C>T) and rs5270 (-297C>G), were genotyped in all the case patients and control subjects. The -1285 G-allele and -1265 T-allele were associated with increased risk of prostate cancer [odds ratio (OR) = 3.53; confidence interval (CI) = 2.14-5.90; P < 0.0001 and OR = 3.01; CI = 1.82-5.02; P < 0.0001] after adjusting for age. Haplotype GTC conferred increased risk of prostate cancer in South African Coloured men (OR = 3.54 versus ACC; CI = 2.12-5.92; P < 0.0001). These findings in conjunction with findings in other populations of African descent might suggest a common causal variant for prostate cancer in COX-2, or a variant in a nearby gene.


Common SNP-based haplotype analysis of the 4p16.3 Huntington disease gene region.
Age at the onset of motor symptoms in Huntington disease (HD) is determined largely by the length of a CAG repeat expansion in HTT but is also influenced by other genetic factors. We tested whether common genetic variation near the mutation site is associated with differences in the distribution of expanded CAG alleles or age at the onset of motor symptoms. To define disease-associated single-nucleotide polymorphisms (SNPs), we compared 4p16.3 SNPs in HD subjects with population controls in a case:control strategy, which revealed that the strongest signals occurred at a great distance from the HD mutation as a result of "synthetic association" with SNP alleles that are of low frequency in population controls. Detailed analysis delineated a prominent ancestral haplotype that accounted for ∼50% of HD chromosomes and extended to at least 938 kb on about half of these. Together, the seven most abundant haplotypes accounted for ∼83% of HD chromosomes. Neither the extended shared haplotype nor the individual local HTT haplotypes were associated with altered CAG-repeat length distribution or residual age at the onset of motor symptoms, arguing against modification of these disease features by common cis-regulatory elements. Similarly, the 11 most frequent control haplotypes showed no trans-modifier effect on age at the onset of motor symptoms. Our results argue against common local regulatory variation as a factor influencing HD pathogenesis, suggesting that genetic modifiers be sought elsewhere in the genome. They also indicate that genome-wide association analysis with a small number of cases can be effective for regional localization of genetic defects, even when a founder effect accounts for only a fraction of the disorder.


Interplay between Foxd3 and Mitf regulates cell fate plasticity in the zebrafish neural crest.
Pigment cells of the zebrafish, Danio rerio, offer an exceptionally tractable system for studying the genetic and cellular bases of cell fate decisions. In the zebrafish, neural crest cells generate three types of pigment cells during embryogenesis: yellow xanthophores, iridescent iridophores and black melanophores. In this study, we present evidence for a model whereby melanophores and iridophores descend from a common precursor whose fate is regulated by an interplay between the transcription factors Mitf and Foxd3. Loss of mitfa, a key regulator of melanophore development, resulted in supernumerary ectopic iridophores while loss of foxd3, a mitfa repressor, resulted in fewer iridophores. Double mutants showed a restoration of iridophores, suggesting that one of Foxd3's roles is to suppress mitfa to promote iridophore development. Foxd3 co-localized with pnp4a, a novel marker of early iridophore development, and was necessary for its expression. A considerable overlap was found between iridoblast and melanoblast markers but not xanthoblast markers, which resolved as cells began to differentiate. Cell lineage analyses using the photoconvertible marker, EosFP, revealed that both melanophores and iridophores develop from a mitfa+ precursor. Taken together, our data reveal a Foxd3/mitfa transcriptional switch that governs whether a bi-potent pigment precursor will attain either an iridophore or a melanophore fate.


A total-evidence approach to dating with fossils, applied to the early radiation of the hymenoptera.
Phylogenies are usually dated by calibrating interior nodes against the fossil record. This relies on indirect methods that, in the worst case, misrepresent the fossil information. Here, we contrast such node dating with an approach that includes fossils along with the extant taxa in a Bayesian total-evidence analysis. As a test case, we focus on the early radiation of the Hymenoptera, mostly documented by poorly preserved impression fossils that are difficult to place phylogenetically. Specifically, we compare node dating using nine calibration points derived from the fossil record with total-evidence dating based on 343 morphological characters scored for 45 fossil (4--20 complete) and 68 extant taxa. In both cases we use molecular data from seven markers (∼5 kb) for the extant taxa. Because it is difficult to model speciation, extinction, sampling, and fossil preservation realistically, we develop a simple uniform prior for clock trees with fossils, and we use relaxed clock models to accommodate rate variation across the tree. Despite considerable uncertainty in the placement of most fossils, we find that they contribute significantly to the estimation of divergence times in the total-evidence analysis. In particular, the posterior distributions on divergence times are less sensitive to prior assumptions and tend to be more precise than in node dating. The total-evidence analysis also shows that four of the seven Hymenoptera calibration points used in node dating are likely to be based on erroneous or doubtful assumptions about the fossil placement. With respect to the early radiation of Hymenoptera, our results suggest that the crown group dates back to the Carboniferous, ∼309 Ma (95% interval: 291--347 Ma), and diversified into major extant lineages much earlier than previously thought, well before the Triassic. [Bayesian inference; fossil dating; morphological evolution; relaxed clock; statistical phylogenetics.].


Composition of the Survival Motor Neuron (SMN) Complex in Drosophila melanogaster.
Spinal Muscular Atrophy (SMA) is caused by homozygous mutations in the human survival motor neuron 1 (SMN1) gene. SMN protein has a well-characterized role in the biogenesis of small nuclear ribonucleoproteins (snRNPs), core components of the spliceosome. SMN is part of an oligomeric complex with core binding partners, collectively called Gemins. Biochemical and cell biological studies demonstrate that certain Gemins are required for proper snRNP assembly and transport. However, the precise functions of most Gemins are unknown. To gain a deeper understanding of the SMN complex in the context of metazoan evolution, we investigated its composition in Drosophila melanogaster Using transgenic flies that exclusively express Flag-tagged SMN from its native promoter, we previously found that Gemin2, Gemin3, Gemin5, and all nine classical Sm proteins, including Lsm10 and Lsm11, co-purify with SMN. Here, we show that CG2941 is also highly enriched in the pulldown. Reciprocal co-immunoprecipitation reveals that epitope-tagged CG2941 interacts with endogenous SMN in Schneider2 cells. Bioinformatic comparisons show that CG2941 shares sequence and structural similarity with metazoan Gemin4. Additional analysis shows that three other genes (CG14164, CG31950 and CG2371) are not orthologous to Gemins 6-7-8, respectively, as previously suggested. In D.melanogaster, CG2941 is located within an evolutionarily recent genomic triplication with two other nearly identical paralogous genes (CG32783 and CG32786). RNAi-mediated knockdown of CG2941 and its two close paralogs reveals that Gemin4 is essential for organismal viability.


Furrow-associated microtubule arrays are required for the cohesion of zebrafish blastomeres following cytokinesis.
During the first few cleavages of the zebrafish embryo, daughter blastomeres are loosely associated immediately after furrow ingression, but then gradually cohere. Cohesion appears to be cadherin-dependent, as cadherin and beta-catenin are found at the membrane between cohering blastomeres, and blastomeres fail to cohere in calcium-free medium. Cadherin and beta-catenin are not initially found on the blastomere surface, but are deposited specifically at the furrow surface. An array of parallel microtubules is present on either side of the furrow tip during ingression, as seen by confocal microscopy of alpha-tubulin labelled embryos. Transient incubation of embryos in 1 microg/ml nocodazole at the start of furrowing, which causes a loss of the furrow array, inhibits the localization of beta-catenin to the furrow surface but does not prevent furrow ingression. During ingression, intracellular membranes are transported to the furrow, as shown by labelling with DiD or DiOC6(3). Concentration of these membranes near the furrow surface is microtubule-dependent. These findings suggest that microtubules are required for cohesion of blastomeres because they mediate trafficking of intracellular membranes to the furrow surface, where they are exocytosed and allow cohesion via cadherins.


Characterization and function of histamine receptors in corpus cavernosum.
Widespread clinical use of H2 antagonists for peptic ulcer disease has been associated with reports of impotence. Histamine is a vasoactive amine which is endogenously produced in many organs including the penis. To date, 3 histamine receptor subtypes (H1, H2 and H3) have been identified. However, the role and function of histamine in the corpus cavernosal physiology are poorly understood. This study evaluates the in vitro functional characteristics of the 3 histamine receptor subtypes in the isolated corpus cavernosal strips from New Zealand White rabbits. The isometric responses to histamine and specific histamine receptor subtype agonists were assessed, following cyclooxygenase (indomethacin, 10(-5) M.), adrenergic (guanethidine, 5 x 10(-6) M.) and cholinergic (atropine, 5 x 10(-6) M.) blockade, at resting tension and after submaximal precontraction with norepinephrine (NE, 2 x 10(-5) M.). Histamine (10(-8) M. to 10(-3) M.) produced concentration-dependent contraction from basal and precontracted states and did not relax precontracted tissue. The H1 agonist, 2-(2-thiazolyl)ethylamine (10(-8) M. to 10(-3) M.), produced a contractile response from both basal and precontracted states, while the corporal tissue did not respond to either dimaprit, an H2 agonist (10(-8) M. to 10(-5) M.) or R(-)-alpha-methyl-histamine, an H3 agonist (10(-8) M. to 10(-5) M.). The response to histamine was progressively attenuated by an H1 antagonist (mepyramine; 10(-8) M. to 10(-5) M.), while neither an H2 antagonist (cimetidine; 10(-4)M.) nor an H3 antagonist (thioperamide; 10(-4)M.) had any inhibitory effects. H1 antagonism enhanced relaxation induced by electrical field stimulation (neurally mediated). Such relaxation increased after preincubation with 10(-6) M. or greater of mepyramine (p < 0.05). This study suggests that the principal histamine receptor subtype that mediates smooth muscle cell contraction in the corpus cavernosum is the H1 subtype. Since histamine H1 receptor antagonism increased NANC neurally mediated corporal relaxation, it possesses potential as an intracavernosal pharmacotherapeutic agent for the treatment of erectile dysfunction. This study, therefore, strongly indicates that H2 receptor antagonists are unlikely to have direct effects on penile erection.


Characterization of spatial patterns of flow within the reperfused myocardium by myocardial contrast echocardiography. Implications in determining extent of myocardial salvage.
Since myocardial blood flow changes dynamically after reperfusion and since both hyperemia and impairment in microvascular function exist within the acutely reperfused bed, we sought to investigate the role of myocardial contrast echocardiography (MCE) in (1) defining the temporal variability in perfusion patterns after reflow and relating these to microsphere-derived blood flow; (2) differentiating viable from infarcted tissue during different periods of reflow; and (3) defining spatial perfusion patterns within the infarct bed in response to exogenously induced maximal vasodilation and relating these to infarct size and extent of myocardial salvage. Twenty-one dogs with 3 hours of left anterior descending coronary artery occlusion and 2 to 3 hours of reflow were studied. MCE was performed at 15 and 45 minutes and 2 and 3 hours after reflow. It was also performed at either 2 or 3 hours after reflow in the presence of 0.56 mg/kg of dipyridamole. Radiolabeled microsphere-derived blood flow was measured at 15 minutes and 2 and 3 hours after reflow and during dipyridamole effect. Infarct size was measured at the end of the experiment by use of triphenyl tetrazolium chloride. MCE data were processed with color-coding schemes that highlighted differences in myocardial videointensities in proportion to the concentration of microbubbles within the microvasculature. There was significant variability in MCE-defined perfusion patterns after reflow, with contrast defects noted mainly within the endocardium. There were fair and significant (P < .05) correlation (r = -.73 to r = -.55) between MCE defect size and normalized endocardial blood flow. Except at 15 minutes after reflow, there was poor correlation (r = .31 to r = .51) between MCE defect and infarct sizes. Even at 15 minutes after reflow, MCE defect size underestimated infarct size by 50%. In comparison, in the presence of dipyridamole, MCE defect size correlated strongly (r = .87, P < .001) with infarct size and reasonably well with normalized transmural blood flow (r = -.62, P = .04). Moreover, the topography of the MCE perfusion defect reflected the topography of the infarct. MCE revealed striking temporal heterogeneity in the spatial distribution of myocardial perfusion during postischemia reflow and either significantly underestimated or did not correlate with infarct size during reperfusion. Because of abnormalities in coronary vascular reserve specific to infarcted tissue, MCE in conjunction with intravenous dipyridamole depicted, in vivo, the actual topography of the infarct with remarkable accuracy.


Heat strain attenuation while wearing NBC clothing: dry-ice vest compared to water spray.
While wearing impermeable nuclear, biological, and chemical (NBC) clothing, reduction of thermal stress is of primary importance. We compared the effect between two cooling methods on the attenuation of heat strain. There were six male subjects who were divided into two groups of three and exposed on two consecutive days to 125 min of exercise in a high heat load (40 degrees C, 40% RH) wearing NBC clothing. They were cooled by one of two different cooling methods: an active cooling vest (CV) based on the sublimation of dry ice, or tap water spraying (TP). After 2 h, rectal temperature (Tre) was significantly higher for the CV compared with the TP (38.1 +/- 0.04 degrees C vs. 37.7 +/- 0.10 degrees C, respectively). Skin temperature (Tsk) was significantly higher for the CV compared with the TP (36.60 +/- 0.54 degrees C vs. 34.90 +/- 0.35 degrees C, respectively). In the second hour, heart rate (HR) was significantly higher for CV compared with TP (118 +/- 13 bpm vs. 104 +/- 64 bpm, respectively). Heat storage was significantly higher after the first and second hours for the CV compared with the TP. The physiological strain index (PSI) was higher for CV compared with TP in the second hour. Sweat rate (msw) was significantly higher for CV compared with TP (560 +/- 45 g x h(-1) vs. 409 +/- 84 g x h(-1), respectively). Subjective thermal comfort was not significantly different. TP was more effective than the CV in reducing heat strain under the conditions used in the study. Until a significant breakthrough in reducing heat strain while wearing NBC clothing in field conditions can be found, TP appears to be an effective and recommended cooling method.


The human bronchial epithelial cell line 16HBE14o- as a model system of the airways for studying drug transport.
The 16HBE14o- cell line, which forms polarised cell layers in vitro, provides a promising opportunity to develop a convenient epithelial cell culture model in which respiratory drug transport can be evaluated in vitro. This study investigated the effect of cell seeding density, collagen substratum and time in culture on the development of barrier properties in this cell line, after which the permeability of the 16HBE14o- cell layers to a series of solutes was studied. Seeding cells at a density of 2.5 x 10(5) cells per cm(2) on a monofibrillar Vitrogen-100 collagen substratum, followed by culture at an air-liquid interface for 6 days resulted in cell layers with a transepithelial electrical resistance (TER) of 247+/-47 omegacm(2) and an apparent permeability coefficient of 2.5 x 10(-6)cms(-1) for mannitol. The permeability of the 16HBE14o- cells to hydrophilic molecules (logP<1.9) was of an order of magnitude greater than that of typical alveolar cell cultures, possibly reflecting barrier properties more representative of the airways. More lipophilic drugs showed higher permeabilities indicating a sigmoidal relationship between permeability and lipophilicity similar to that observed for solute transport across primary cultured epithelial cell layers. These results indicate that under appropriate culture conditions, 16HBE14o- cell layers provide a discriminatory barrier to solute transport.


Changes in collagen and albumin mRNA in liver tissue of mice infected with Schistosoma mansoni as determined by in situ hybridization.
We have employed in situ hybridization to evaluate the molecular mechanisms responsible for hypoalbuminemia and increased liver collagen content in murine schistosomiasis. Results were compared using a simplified method of hybridizing isolated hepatocytes from Schistosoma mansoni-infected and normal mouse liver with mouse albumin (pmalb-2) and chick pro-alpha 2(l) collagen (pCg45) probes. Whereas hepatocytes from infected mice showed significantly less albumin mRNA than hepatocytes from control, there were more grains of procollagen mRNA in hepatocytes from infected as compared with control liver. Hybridization of infected liver tissue sections with the collagen probe showed more grains per field in granulomas than in liver regions, whereas with the albumin probe there was more hybridization in liver tissue than in granulomas. These results suggest that in murine schistosomiasis a reduction in albumin mRNA sequence content may be associated with decreased albumin synthesis and ultimately leads to hypoalbuminemia. In addition, although the granuloma seems to be the primary source of type I collagen synthesis, hepatocytes are also capable of synthesizing collagen, especially under fibrogenic stimulation.


Chitosan in mucoadhesive drug delivery: focus on local vaginal therapy.
Mucoadhesive drug therapy destined for localized drug treatment is gaining increasing importance in today's drug development. Chitosan, due to its known biodegradability, bioadhesiveness and excellent safety profile offers means to improve mucosal drug therapy. We have used chitosan as mucoadhesive polymer to develop liposomes able to ensure prolonged residence time at vaginal site. Two types of mucoadhesive liposomes, namely the chitosan-coated liposomes and chitosan-containing liposomes, where chitosan is both embedded and surface-available, were made of soy phosphatidylcholine with entrapped fluorescence markers of two molecular weights, FITC-dextran 4000 and 20,000, respectively. Both liposomal types were characterized for their size distribution, zeta potential, entrapment efficiency and the in vitro release profile, and compared to plain liposomes. The proof of chitosan being both surface-available as well as embedded into the liposomes in the chitosan-containing liposomes was found. The capability of the surface-available chitosan to interact with the model porcine mucin was confirmed for both chitosan-containing and chitosan-coated liposomes implying potential mucoadhesive behavior. Chitosan-containing liposomes were shown to be superior in respect to the simplicity of preparation, FITC-dextran load, mucoadhesiveness and in vitro release and are expected to ensure prolonged residence time on the vaginal mucosa providing localized sustained release of entrapped model substances.


Pharmacodynamic evaluation and safety assessment of treatment with antibodies to serum amyloid P component in patients with cardiac amyloidosis: an open-label Phase 2 study and an adjunctive immuno-PET imaging study.
In a Phase I study treatment with the serum amyloid P component (SAP) depleter miridesap followed by monoclonal antibody to SAP (dezamizumab) showed removal of amyloid from liver, spleen and kidney in patients with systemic amyloidosis. We report results from a Phase 2 study and concurrent immuno-positron emission tomography (PET) study assessing efficacy, pharmacodynamics, pharmacokinetics, safety and cardiac uptake (of dezamizumab) following the same intervention in patients with cardiac amyloidosis. Both were uncontrolled open-label studies. After SAP depletion with miridesap, patients received ≤ 6 monthly doses of dezamizumab in the Phase 2 trial (n = 7), ≤ 2 doses of non-radiolabelled dezamizumab plus [89Zr]Zr-dezamizumab (total mass dose of 80 mg at session 1 and 500 mg at session 2) in the immuno-PET study (n = 2). Primary endpoints of the Phase 2 study were changed from baseline to follow-up (at 8 weeks) in left ventricular mass (LVM) by cardiac magnetic resonance imaging and safety. Primary endpoint of the immuno-PET study was [89Zr]Zr-dezamizumab cardiac uptake assessed via PET. Dezamizumab produced no appreciable or consistent reduction in LVM nor improvement in cardiac function in the Phase 2 study. In the immuno-PET study, measurable cardiac uptake of [89Zr]Zr-dezamizumab, although seen in both patients, was moderate to low. Uptake was notably lower in the patient with higher LVM. Treatment-associated rash with cutaneous small-vessel vasculitis was observed in both studies. Abdominal large-vessel vasculitis after initial dezamizumab dosing (300 mg) occurred in the first patient with immunoglobulin light chain amyloidosis enrolled in the Phase 2 study. Symptom resolution was nearly complete within 24 h of intravenous methylprednisolone and dezamizumab discontinuation; abdominal computed tomography imaging showed vasculitis resolution by 8 weeks. Unlike previous observations of visceral amyloid reduction, there was no appreciable evidence of amyloid removal in patients with cardiac amyloidosis in this Phase 2 trial, potentially related to limited cardiac uptake of dezamizumab as demonstrated in the immuno-PET study. The benefit-risk assessment for dezamizumab in cardiac amyloidosis was considered unfavourable after the incidence of large-vessel vasculitis and development for this indication was terminated. Trial registration NCT03044353 (2 February 2017) and NCT03417830 (25 January 2018).


The treatment of neuroleptic malignant syndrome. Are dantrolene and bromocriptine useful adjuncts to supportive care?
In a prospective study of 20 patients with NMS, those who received dantrolene (2 patients) or bromocriptine (2 patients) or both (4 patients) tended to have a more prolonged course of illness and a greater incidence of sequelae than those treated with supportive care alone. These results do not indicate a useful role for dantrolene or bromocriptine in the treatment of NMS.


Radiosurgery for Unruptured Brain Arteriovenous Malformations: An International Multicenter Retrospective Cohort Study.
The role of intervention in the management of unruptured brain arteriovenous malformations (AVM) is controversial. To analyze in a multicenter, retrospective cohort study, the outcomes following radiosurgery for unruptured AVMs and determine predictive factors. We evaluated and pooled AVM radiosurgery data from 8 institutions participating in the International Gamma Knife Research Foundation. Patients with unruptured AVMs and ≥12 mo of follow-up were included in the study cohort. Favorable outcome was defined as AVM obliteration, no postradiosurgical hemorrhage, and no permanently symptomatic radiation-induced changes. The unruptured AVM cohort comprised 938 patients with a median age of 35 yr. The median nidus volume was 2.4 cm 3 , 71% of AVMs were located in eloquent brain areas, and the Spetzler-Martin grade was III or higher in 57%. The median radiosurgical margin dose was 21 Gy and follow-up was 71 mo. AVM obliteration was achieved in 65%. The annual postradiosurgery hemorrhage rate was 1.4%. Symptomatic and permanent radiation-induced changes occurred in 9% and 3%, respectively. Favorable outcome was achieved in 61%. In the multivariate logistic regression analysis, smaller AVM maximum diameter ( P = .001), the absence of AVM-associated arterial aneurysms ( P = .001), and higher margin dose ( P = .002) were found to be independent predictors of a favorable outcome. A margin dose ≥ 20 Gy yielded a significantly higher rate of favorable outcome (70% vs 36%; P < .001). Radiosurgery affords an acceptable risk to benefit profile for patients harboring unruptured AVMs. These findings justify further prospective studies comparing radiosurgical intervention to conservative management for unruptured AVMs.


Chronic fatigue syndrome defies the mind-body-schism of medicine. New perspectives on a multiple realisable developmental systems disorder.
The article maintains that chronic fatigue syndrome can be properly understood only by taking an integrated perspective in which evolutionary, developmental and ecological aspects are considered. The integrative approach, supplemented by a complexity theory and psychoneuroimmunological research, is capable of explaining why there are so few structural aberrations to be found in chronic fatigue syndrome and why specific treatment is so difficult to establish. A major outcome of the investigation, that all individuals with chronic fatigue syndrome are diseased in their own way, emphasises the need to study the development of personalised life histories. It also highlights an ethical dimension; personalised disease defies essentialist thinking on patient management. Another major outcome, which follows from the developmental systems perspective, is the dissolution of ontological mind-body dualism. This in turn allows for a methodological complementation of the biological and phenomenological approaches to knowledge. New research strategies that may help to resolve chronic fatigue syndrome, grounded in the revised perspective on individual development, are suggested.


Human liver chimeric mice provide a model for hepatitis B and C virus infection and treatment.
A paucity of versatile small animal models of hepatitis B virus (HBV) and hepatitis C virus (HCV) infection has been an impediment to both furthering understanding of virus biology and testing antiviral therapies. We recently described a regulatable system for repopulating the liver of immunodeficient mice (specifically mice lacking fumaryl acetoacetate hydrolase [Fah], recombination activating gene 2 [Rag2], and the gamma-chain of the receptor for IL-2 [Il-2rgamma]) with human hepatocytes. Here we have shown that a high transplantation dose (3 x 106 to 5 x 106 human hepatocytes/mouse) generates a higher rate of liver chimerism than was previously obtained in these mice, up to 95% human hepatocyte chimerism. Mice with a high level of human liver chimerism propagated both HBV and HCV, and the HCV-infected mice were responsive to antiviral treatment. This human liver chimeric mouse model will expand the experimental possibilities for studying HBV and HCV infection, and possibly other human hepatotropic pathogens, and prove useful for antiviral drug testing.


Terahertz transmission control using polarization-independent metamaterials.
We present terahertz (THz) transmission control by several uniquely designed patterns of nano-slot antenna array. Collinearly aligned slot antenna arrays have been usually applied to THz filters with frequency band tunability by their geometry. Normally the amplitude in transmission (reflection) in the collinear alignment case can be varied via rotating the azimuthal angle with a sinusoidal trend, which can limit their utilization and performance only at fixed angle between the alignment of the resonant antennas and incident beam polarization. To pursue a variety of metamaterial uses, here, we present polarization-independent THz filters using variously aligned antenna array (asterisk, chlorophyll, and honeycomb patterns) in such counter-intuitive aspects. Besides, unprecedented multi resonance behaviors were observed in chlorophyll and honeycomb patterns, which can be explained with interferences by adjacent structures. The measured spectra were analyzed by harmonic oscillator model with simplified coupling between slots and their adjacent.


Regulation of expression of antioxidant enzymes by vitamin E and curcumin in L-thyroxine-induced oxidative stress in rat renal cortex.
The present study investigates the antioxidative effects of vitamin E and curcumin against L-thyroxine (T(4))-induced oxidative stress in renal cortex of adult male rats. Rats were made hyperthyroid by administration of L-thyroxine (0.0012%) in their drinking water for 30 days. Vitamin E (200 mg/kg body weight/day) and curcumin (30 mg/kg body weight/day) were supplemented singly or in combination orally for 30 days along with L-thyroxine treatment. The elevated level of oxidative stress parameters (lipid peroxidation and protein carbonylation) and decline level of small antioxidant molecules (reduced glutathione and ascorbic acid) in renal cortex of T(4)-treated rats were restored back by supplementation of vitamin E or/and curcumin. Increased superoxide dismutase and catalase activities in kidney cortex of T(4)-treated rats were ameliorated in response to vitamin E or/and curcumin treatment. The elevated translated product of Cu/Zn-SOD, Mn-SOD and catalase in T(4)-treated rats were differentially reduced by the administration of vitamin E and curcumin independently or in combination. Cu/Zn-SOD expression was ameliorated by both vitamin E and curcumin independently or in combination, whereas Mn-SOD expression was ameliorated by the supplementation of vitamin E or curcumin independently. However, the expression of catalase was alleviated by only supplementation of vitamin E to T(4)-treated rats. The results suggest that both vitamin E and curcumin may play an important role in protecting T(4)-induced oxidative stress in rat renal cortex by differentially modulating the activities of antioxidant enzymes and oxidative stress parameters.


Effects of the beta-adrenoceptor agonist isoprenaline on insulin-sensitivity in soleus muscle of the rat.
The interactions between a beta-adrenoceptor agonist (isoprenaline) and insulin on rates of hexose transport, glucose phosphorylation, glycogen synthesis and glycogenolysis were investigated in the incubated stripped soleus-muscle preparation of the rat. In the presence of 1 microM-isoprenaline, insulin was less effective in stimulating glucose phosphorylation and glycogen synthesis. The stimulation of glycogenolysis by isoprenaline was only slightly decreased even at high (10000 microunits/ml) concentrations of insulin. Insulin-stimulated phosphorylation of 2-deoxyglucose was decreased by isoprenaline. It is suggested that this decrease in the rate of glucose phosphorylation is caused by the observed elevated concentration of glucose 6-phosphate, which inhibits hexokinase activity. This conclusion is supported by the fact that isoprenaline had no effect on the stimulation of 3-O-methylglucose transport by insulin.


Birthweight, HIV exposure and infant feeding as predictors of malnutrition in Botswanan infants.
A better understanding of the nutritional status of infants who are HIV-Exposed-Uninfected (HEU) and HIV-Unexposed-Uninfected (HUU) during their first 1000 days is key to improving population health, particularly in sub-Saharan Africa. A cross-sectional study compared the nutritional status, feeding practices and determinants of nutritional status of HEU and HUU infants residing in representative selected districts in Botswana during their first 1000 days of life. Four hundred and thirteen infants (37.3% HIV-exposed), aged 6-24 months, attending routine child health clinics, were recruited. Anthropometric, 24-h dietary intake and socio-demographic data was collected. Anthropometric Z-scores were calculated using 2006 World Health Organization growth standards. Modelling of the determinants of malnutrition was undertaken using logistic regression. Overall, the prevalences of stunting, wasting and being underweight were 10.4%, 11.9% and 10.2%, respectively. HEU infants were more likely to be underweight (15.6% versus 6.9%), (P < 0.01) and stunted (15.6% versus 7.3%), (P < 0.05) but not wasted (P = 0.14) than HUU infants. HEU infants tended to be formula fed (82.5%), whereas HUU infants tended to breastfeed (94%) for the first 6 months (P < 0.001). Significant predictors of nutritional status were HIV exposure, birthweight, birth length, APGAR (appearance, pulse, grimace, activity and respiration) score and mother/caregiver's education with little influence of socio-economic status. HEU infants aged 6-24 months had worse nutritional status compared to HUU infants. Low birthweight was the main predictor of undernutrition in this population. Optimisation of infant nutritional status should focus on improving birthweight. In addition, specific interventions should target HEU infants aiming to eliminate growth disparity between HEU and HUU infants.


Using zeta potential to study the ionisation behaviour of polymers employed in modified-release dosage forms and estimating their pKa.
A range of enteric polymers is used in pharmaceutical industry for developing gastro-resistant formulations. It is generally implied that these coatings are interchangeable due to similar dissolution pH thresholds reported by suppliers. Despite rapid dissolution in compendial phosphate buffers, these products can take up to 2 h to disintegrate in-vivo in the human small intestine. The factors primarily responsible for such variability in dissolution of these polymeric coatings are the differences in ionisation of acidic functional groups on polymer chains and their interplay with ions and buffer species present in gastrointestinal fluids. In this study, we aim to develop a novel, simple and inexpensive technique that can be used under various in-vitro conditions to study the ionisation behaviour of commonly used polymers (EUDRAGIT-E100, L100, S100, HPMC AS-LF, AS-HF, HP-50, HP-55) and to estimate their pKa. Moreover, this method was successfully applied to study the ionisation behaviour of a range of natural polymers (Guar, Tara, locust bean, Konjac gums, gum Arabic, citrus pectin, chitosan and alginate) and their pKa was also estimated. The proposed method would allow a better understanding of the dissolution behaviour of these polymers within gastrointestinal tract and will aid rational design of modified release dosage forms.


Neutralizing antibody affords comparable protection against vaginal and rectal simian/human immunodeficiency virus challenge in macaques.
Passive administration of broadly neutralizing antibodies has been shown to protect against both vaginal and rectal challenge in the simian/human immunodeficiency virus (SHIV)/macaque model of HIV transmission. However, the relative efficacy of antibody against the two modes of exposure is unknown and, given differences in the composition and immunology of the two tissue compartments, this is an important gap in knowledge. To investigate the significance of the challenge route for antibody-mediated protection, we performed a comparative protection study in macaques using the highly potent human monoclonal antibody, PGT126. Animals were administered PGT126 at three different doses before challenged either vaginally or rectally with a single dose of SHIVSF163P3. Viral loads, PGT126 serum concentrations, and serum neutralizing titers were monitored. In vaginally challenged animals, sterilizing immunity was achieved in all animals administered 10 mg/kg, in two of five animals administered 2 mg/kg and in one of five animals administered 0.4 mg/kg PGT126. Comparable protection was observed for the corresponding groups challenged rectally as sterilizing immunity was achieved in three of four animals administered 10 mg/kg, in two of four animals administered 2 mg/kg and in none of four animals administered 0.4 mg/kg PGT126. Serological analysis showed similar serum concentrations of PGT126 and serum neutralization titers in animals administered the same antibody dose. Our data suggest that broadly neutralizing antibody-mediated protection is not strongly dependent on the mucosal route of challenge, which indicates that a vaccine aimed to induce a neutralizing antibody response would have broadly similar efficacy against both primary transmission routes for HIV.


The base excision repair pathway is required for efficient lentivirus integration.
An siRNA screen has identified several proteins throughout the base excision repair (BER) pathway of oxidative DNA damage as important for efficient HIV infection. The proteins identified included early repair factors such as the base damage recognition glycosylases OGG1 and MYH and the late repair factor POLß, implicating the entire BER pathway. Murine cells with deletions of the genes Ogg1, Myh, Neil1 and Polß recapitulate the defect of HIV infection in the absence of BER. Defective infection in the absence of BER proteins was also seen with the lentivirus FIV, but not the gammaretrovirus MMLV. BER proteins do not affect HIV infection through its accessory genes nor the central polypurine tract. HIV reverse transcription and nuclear entry appear unaffected by the absence of BER proteins. However, HIV integration to the host chromosome is reduced in the absence of BER proteins. Pre-integration complexes from BER deficient cell lines show reduced integration activity in vitro. Integration activity is restored by addition of recombinant BER protein POLß. Lentiviral infection and integration efficiency appears to depend on the presence of BER proteins.


Acetic Acid Bacteria in the Food Industry: Systematics, Characteristics and Applications.
The group of Gram-negative bacteria capable of oxidising ethanol to acetic acid is called acetic acid bacteria (AAB). They are widespread in nature and play an important role in the production of food and beverages, such as vinegar and kombucha. The ability to oxidise ethanol to acetic acid also allows the unwanted growth of AAB in other fermented beverages, such as wine, cider, beer and functional and soft beverages, causing an undesirable sour taste. These bacteria are also used in the production of other metabolic products, for example, gluconic acid, l-sorbose and bacterial cellulose, with potential applications in the food and biomedical industries. The classification of AAB into distinct genera has undergone several modifications over the last years, based on morphological, physiological and genetic characteristics. Therefore, this review focuses on the history of taxonomy, biochemical aspects and methods of isolation, identification and quantification of AAB, mainly related to those with important biotechnological applications.


Living with an aging parent: "It was a beautiful invitation".
In the United States and globally, increasing numbers of older parents are living with their adult children. Making the decision to live together requires careful thought and planning; particularly when the decision means the children will be responsible for their elder parents' care, the physicians of all parties should be consulted. More than one-third of caregivers state that they seek advice from their physician or other clinician when information is needed about this transition. Using the case of a couple receiving care from their daughter in her own home, we review the prevalence and epidemiology of adult children caring for a parent in the adult child's home, important issues to consider, and a framework for clinicians to help guide their patients through this transition. We describe the physician's ongoing role in caring for and advising both the older parent and the adult children during common stages of multigenerational living: before the move and during coresidency and subsequent care transitions.


Tacit knowledge as the unifying factor in evidence based medicine and clinical judgement.
The paper outlines the role that tacit knowledge plays in what might seem to be an area of knowledge that can be made fully explicit or codified and which forms a central element of Evidence Based Medicine. Appeal to the role the role of tacit knowledge in science provides a way to unify the tripartite definition of Evidence Based Medicine given by Sackett et al: the integration of best research evidence with clinical expertise and patient values. Each of these three elements, crucially including research evidence, rests on an ineliminable and irreducible notion of uncodified good judgement. The paper focuses on research evidence, drawing first on the work of Kuhn to suggest that tacit knowledge contributes, as a matter of fact, to puzzle solving within what he calls normal science. A stronger argument that it must play a role in research is first motivated by looking to Collins' first hand account of replication in applied physics and then broader considerations of replication in justifying knowledge claims in scientific research. Finally, consideration of an argument from Wittgenstein shows that whatever explicit guidelines can be drawn up to guide judgement the specification of what counts as correctly following them has to remain implicit.Overall, the paper sets out arguments for the claim that even though explicit guidelines and codifications can play a practical role in informing clinical practice, they rest on a body of tacit or implicit skill that is in principle ineliminable. It forms the bedrock of good judgement and unites the integration of research, expertise and values.


Aging of the human lens: changes in lens shape upon accommodation and with accommodative loss.
Accommodation in the human eye occurs through controlled changes in crystalline lens shape, thickness, and refractive surface placement relative to the cornea. The changes in lens curvatures, whether surface or internal, have been characterized as a function of accommodation and subject age by use of quantitative analysis of Scheimpflug slit-lamp photographic images. Radii of curvature of the major lens refractive surfaces--the external and nuclear boundaries--decrease linearly with increasing accommodation in all eyes that are capable of accommodation. The rates at which they change with accommodation are age dependent, decreasing steadily toward zero with increased age. For the curves visible in each lens half, arising from boundaries between adjacent zones of discontinuity, radius of curvature and location are linearly related over the entire accommodative range for a given lens and over the age range for the population. The slope of this relationship changes with both accommodation and age, decreasing linearly in both cases. The relationship between these geometric changes and the loss of accommodative amplitude is discussed.


A highly sensitive ultra-high performance liquid chromatography/tandem mass spectrometry method with in-source fragmentation for rapid quantification of raspberry ketone.
Raspberry ketone (RK) is the characteristic aromatic compound in raspberry (Rubus idaeus L.) with wide applications as food additive and anti-obesity agent. However, quantification of RK has presented difficulties in MS detection and reliable LC-MS method for RK analysis in literature is in limit to date. In order to facilitate quality control of raspberry derived products and RK metabolomics study, this study aimed to develop a validated and sensitive UHPLC-MS/MS method. Strong in-source fragmentation was noted and the fragmental ion of 107 m/z produced was selected as the precursor ion for MRM detection, and as such the electrospray ionization performance was optimized by fractional factorial design to accommodate such ion-source dissociation behavior as well as its moderate volatility. A pathway involving the formation of quinone-like structure with strong conjugation was proposed to explain the intense in-source fragmentation. The MRM transition was optimized with product ion of 77 m/z selected as the quantifier ion. The method featured low limit of quantification of ∼2 ng/mL and allowed for rapid detection of RK in fresh raspberries following direct sample preparation. RK contents were found to be higher from locally grown and harvested farm sources compared to commercial products shipped into the state, and higher in those at late-stage compared with early-stage maturity. No correlations in RK content between organic and non-organic labels were noted.


Successful chemical ablation of an intraorbital cyst caused by an eyelid injury and iatrogenic ankyloblepharon formation in a duck.
CASE DESCRIPTION A client-owned 2-year-old 1.8-kg (4-lb) male pet Rouen duck (Anas platyrhynchos domesticus) was evaluated because of severe swelling around the left eye following traumatic injury to the upper and lower eyelids and 2 associated surgeries that resulted in the removal of the entire upper and lower eyelid margins. CLINICAL FINDINGS At initial evaluation, ankyloblepharon of the left eye was observed, with no upper or lower eyelid margins and a large, round, fluctuant subcutaneous mass over the left orbit. Orbital exploration and histologic examination revealed a benign cyst consisting of fibrous tissue, conjunctiva, and skeletal muscle bundles. Bacterial culture of cystic fluid yielded few Staphylococcus delphini. TREATMENT AND OUTCOME Excision of the cyst and evisceration of the left globe were performed, and once daily treatment with orally administered enrofloxacin suspension (12.6 mg/kg [5.7 mg/lb]) and meloxicam (1 mg/kg [0.45 mg/lb]) was initiated. Over the next 4 days, the cyst redeveloped and progressively enlarged. Accumulated fluid was aspirated from the cyst, and 20 mg of gentamicin was injected intraorbitally with ultrasound guidance. Over the subsequent 27-month period, no recurrence of clinical signs or adverse effects were reported by the owner. CLINICAL RELEVANCE To the authors' knowledge, this is the first report of cyst formation after adnexal injury and evisceration in birds and its successful treatment with intralesional gentamicin injection. Findings emphasized the importance of preserving lacrimal puncta during adnexal or eye removal surgeries in birds. Intralesional injection of gentamicin with the goal of destroying fluid-producing cells may be a safe and effective way to treat intraorbital cysts in birds and other species, although additional research would be required to confirm this.


Genomic analysis of atypical fibroxanthoma.
Atypical fibroxanthoma (AFX), is a rare type of skin cancer affecting older individuals with sun damaged skin. Since there is limited genomic information about AFX, our study seeks to improve the understanding of AFX through whole-exome and RNA sequencing of 8 matched tumor-normal samples. AFX is a highly mutated malignancy with recurrent mutations in a number of genes, including COL11A1, ERBB4, CSMD3, and FAT1. The majority of mutations identified were UV signature (C>T in dipyrimidines). We observed deletion of chromosomal segments on chr9p and chr13q, including tumor suppressor genes such as KANK1 and CDKN2A, but no gene fusions were found. Gene expression profiling revealed several biological pathways that are upregulated in AFX, including tumor associated macrophage response, GPCR signaling, and epithelial to mesenchymal transition (EMT). To further investigate the presence of EMT in AFX, we conducted a gene expression meta-analysis that incorporated RNA-seq data from dermal fibroblasts and keratinocytes. Ours is the first study to employ high throughput sequencing for molecular profiling of AFX. These data provide valuable insights to inform models of carcinogenesis and additional research towards tumor-directed therapy.


Genetic diversity of Spiroplasma citri strains from different regions, hosts, and isolation dates.
Spiroplasma citri, a phloem-limited pathogen, causes citrus stubborn disease (CSD). Losses due to CSD in California orchards have grown over the past decade. To investigate the possibility of introduction or emergence of a new strain, a study of genetic diversity among S. citri strains from various locations was conducted using random amplified polymorphism DNA-polymerase chain reaction (RAPD-PCR) of 35 strains cultured from 1980 to 1993, and of 35 strains cultured from 2005 to 2006. Analysis using 20 primer pairs revealed considerable diversity among strains. However, no unique genetic signatures were associated with recently collected strains compared with those collected 15 to 28 years ago, and no geographically associated pattern was distinguishable. S. citri strains from carrot and daikon radish contain some unique DNA fragments, suggesting some host plant influence. Multiple strains from single trees also showed genetic diversity. Sequencing of five RAPD bands that differed among strains showed that diversity-related gene sequences include virus fragments, and fragments potentially encoding a membrane lipoprotein, a DNA modification enzyme, and a mobilization element. No differences in colony morphology were observed among the strains. The lack of correlation between PCR patterns and isolation date or collection site is inconsistent with the hypothesis that recent infections are due to the introduction or emergence of novel pathogen strains.


Crystallites of magnetic charges in artificial spin ice.
Artificial spin ice is a class of lithographically created arrays of interacting ferromagnetic nanometre-scale islands. It was introduced to investigate many-body phenomena related to frustration and disorder in a material that could be tailored to precise specifications and imaged directly. Because of the large magnetic energy scales of these nanoscale islands, it has so far been impossible to thermally anneal artificial spin ice into desired thermodynamic ensembles; nearly all studies of artificial spin ice have either treated it as a granular material activated by alternating fields or focused on the as-grown state of the arrays. This limitation has prevented experimental investigation of novel phases that can emerge from the nominal ground states of frustrated lattices. For example, artificial kagome spin ice, in which the islands are arranged on the edges of a hexagonal net, is predicted to support states with monopolar charge order at entropies below that of the previously observed pseudo-ice manifold. Here we demonstrate a method for thermalizing artificial spin ices with square and kagome lattices by heating above the Curie temperature of the constituent material. In this manner, artificial square spin ice achieves unprecedented thermal ordering of the moments. In artificial kagome spin ice, we observe incipient crystallization of the magnetic charges embedded in pseudo-ice, with crystallites of magnetic charges whose size can be controlled by tuning the lattice constant. We find excellent agreement between experimental data and Monte Carlo simulations of emergent charge-charge interactions.


Cardiac remote ischaemic preconditioning reduces periprocedural myocardial infarction for patients undergoing percutaneous coronary interventions: a meta-analysis of randomised clinical trials.
To establish the cardioprotective effect of remote ischaemic preconditioning (RIPC) in patients undergoing percutaneous coronary intervention (PCI). Pubmed (MEDLINE), Cochrane and Embase were systematically searched for randomised controlled trials of RIPC in patients undergoing PCI. Periprocedural myocardial infarction (PMI) was the primary endpoint (defined as troponin elevation >3 times upper reference limit) and C-reactive protein (CRP) was a secondary endpoint. Five studies with 731 patients were included. The median age of the patients was 62 (59-68) years old, 25% were female (23-33), 29% (25-33) had diabetes mellitus, and 26.5% (19-31) presented with multivessel disease. RIPC significantly reduced the incidence of PMI (odds ratio: 0.58 [0.36, 0.93]; I2 43%), with a greater benefit when performed using the lower limb (0.21 [0.07-0.66]) compared to the upper limb (0.67 [0.46-0.99]). This reduction was enhanced for patients with multivessel disease (beta -0.05 [-0.09;-0.01], p=0.01) and with type C lesion (beta -0.014 [-0.04;-0.010], p=0.01) and did not vary according to age, female gender, diabetes mellitus, use of beta-blockers and of angiotensin converting enzyme inhibitors. Absolute risk difference was -0.10 [-0.19, -0.02], with a number needed to treat of 10 [6-50] patients to avoid one event. CRP -0.69 [-1.69, 0.31] was not significantly reduced by RIPC. RIPC reduced the incidence of PMI following PCI, especially when performed in the lower limb and for patients with multivessel disease and complex lesions.


Impact of CYP2B6 983T>C polymorphism on non-nucleoside reverse transcriptase inhibitor plasma concentrations in HIV-infected patients.
The aim of this study was to investigate the frequency of CYP2B6 polymorphisms (according to ethnicity) and the influence of heterozygosity and homozygosity on plasma concentrations of efavirenz and nevirapine. Following written informed consent, 225 Caucasians and 146 Blacks were recruited from the German Competence Network for HIV/AIDS. Plasma concentrations of efavirenz and nevirapine were assessed by HPLC, and genotyping for 516G>T, 983T>C and 1459T>C polymorphisms in CYP2B6 was conducted by real-time PCR-based allelic discrimination. The minor allele frequency for 516G>T, 983T>C and 1459T>C was 0.29, 0 and 0.08 in Caucasians and 0.34, 0.07 and 0.02 in Blacks, respectively. Two Black patients with the 983C allele receiving efavirenz were identified and both were withdrawn from therapy within 1 week of sampling due to toxicity. In multivariate analyses, efavirenz and nevirapine plasma concentrations were significantly associated with 983T>C (P < 0.0001 and P = 0.02, respectively), 516G>T (P < 0.0001 and P = 0.002, respectively) and time of drug analysis post-dose (P < 0.0001 for both). Body mass index was independently related to efavirenz (P = 0.04) but not nevirapine concentrations, and age was related to nevirapine (P = 0.05) but not efavirenz concentrations. Consistent with other studies, 1459C>T was not associated with plasma concentrations of either drug (P > 0.05 for both drugs). This is the first report that the 983T>C genotype (part of the CYP2B6*18 haplotype) impacts on nevirapine plasma concentrations and the first study to assess the impact of 983C homozygosity on efavirenz concentrations. These data have implications for administration of non-nucleoside reverse transcriptase inhibitors to Black patients.


Stacked and H-Bonded Cytosine Dimers. Analysis of the Intermolecular Interaction Energies by Parallel Quantum Chemistry and Polarizable Molecular Mechanics.
Until now, atomistic simulations of DNA and RNA and their complexes have been executed using well calibrated but conceptually simple pair-additive empirical potentials (force fields). Although such simulations provided many valuable results, it is well established that simple force fields also introduce errors into the description, underlying the need for development of alternative anisotropic, polarizable molecular mechanics (APMM) potentials. One of the most abundant forces in all kinds of nucleic acids topologies is base stacking. Intra- and interstrand stacking is assumed to be the most essential factor affecting local conformational variations of B-DNA. However, stacking also contributes to formation of all kinds of noncanonical nucleic acids structures, such as quadruplexes or folded RNAs. The present study focuses on 14 stacked cytosine (Cyt) dimers and the doubly H-bonded dimer. We evaluate the extent to which an APMM procedure, SIBFA, could account quantitatively for the results of high-level quantum chemistry (QC) on the total interaction energies, and the individual energy contributions and their nonisotropic behaviors. Good agreements are found at both uncorrelated HF and correlated DFT and CCSD(T) levels. Resorting in SIBFA to distributed QC multipoles and to an explicit representation of the lone pairs is essential to respectively account for the anisotropies of the Coulomb and of the exchange-repulsion QC contributions.


Effect of Clinical Variables on the Volume of Blood Collected for Blood Cultures in an Adult Patient Population.
OBJECTIVE To identify clinical variables that influence blood culture volume recovery DESIGN Retrospective chart review and linear model analysis SETTING A 621-bed Academic Medical Center with a Clinical Laboratory that processes 20,000+ blood cultures annually and dedicated phlebotomy staff for venipuncture PATIENTS Consecutive patients requiring blood culture METHODS Over a 6-day period, blood volume was determined in 568 culture bottles from 128 unique adult patients, and clinical data from the time of phlebotomy were extracted from hospital electronic medical records. Conditional hierarchical linear models with random effects for patient and phlebotomy occasion were utilized to analyze correlations between values collected from the same patient and during the same phlebotomy occasion. RESULTS Blood samples obtained from a central venous catheter yielded, on average, 2.53 mL more blood (95% CI, 1.63-3.44 mL; P<.001) than those from peripheral venipuncture, and aerobic bottles contained 0.38 mL more blood (95% CI, 0.1-0.67 mL; P=.009) than the anaerobic bottles. The remaining clinical variables (eg, hospital department, patient age, body mass index, gender, mean arterial pressure, concomitant systemic antibiotic use, and Charlson comorbidity index score) failed to reach statistical significance (P<.05) in relation to volume. CONCLUSIONS Blood cultures obtained from central venous catheters contain significantly greater volume than those obtained via peripheral venipuncture. These data highlight the clinically significant issue of low culture volume recovery, indicate that diagnostic and prognostic tools that rely on volume-dependent phenomena (ie, time to positivity) may require further validation under usual clinical practice circumstances, and suggest goals for future institutional performance improvement. Infect Control Hosp Epidemiol 2017;38:1493-1497.


The effect of modified constraint-induced movement therapy on spasticity and motor function of the affected arm in patients with chronic stroke.
The purpose of this study was to explore the effect of modified constraint-induced movement therapy (CIMT) in a real-world clinical setting on spasticity and functional use of the affected arm and hand in patients with spastic chronic hemiplegia. A prospective consecutive quasi-experimental study design was used. Twenty patients with spastic hemiplegia (aged 22-67 years) were tested before and after 2-week modified CIMT in an outpatient rehabilitation clinic and at 6 months. The Modified Ashworth Scale (MAS), active range of motion (AROM), grip strength, Motor Activity Log (MAL), Sollerman hand function test, and Box and Block Test (BBT) were used as outcome measures. Reductions (p<0.05-0.001) in spasticity (MAS) were seen both after the 2-week training period and at 6-month follow-up. Improvements were also seen in AROM (median change of elbow extension 5°, dorsiflexion of hand 10°), grip strength (20 Newton), and functional use after the 2-week training period (MAL: 1 point; Sollerman test: 8 points; BBT: 4 blocks). The improvements persisted at 6-month follow-up, except for scores on the Sollerman hand function test, which improved further. Our study suggests that modified CIMT in an outpatient clinic may reduce spasticity and increase functional use of the affected arm in spastic chronic hemiplegia, with improvements persisting at 6 months.


Therapies for multiple sclerosis: translational achievements and outstanding needs.
In recent years, multiple sclerosis (MS) research has progressed on several fronts, prompting numerous clinical trials, primarily for immunotherapeutics. Although several new therapies have been disappointing and some were revealed to have devastating side effects, others have shown benefits and all have generated valuable knowledge about the progression of MS, the key contributors to pathogenesis, and on natural surveillance mechanisms for brain infections. This makes now a useful time to take stock of recent advances in developing MS treatments and consider new approaches for adding information where the gaps are greatest - mainly in understanding the degenerative processes responsible for most of the long-term disability. Here, we summarize currently accepted therapeutic principles and the drugs in late stages of development, as well as spotlighting potential novel openings for future research.


What you see may not be what you get: beware of patients bearing gifts.
The professional boundaries of practice for nurses and midwives are specifically addressed in professional codes of ethics and conduct, employer policies and guidelines and more frequently in the professional literature. There are many forms in which boundary violations and boundary crossings may present or circumstances in which they may occur. This column considers a recent case (involving a bequest to a registered nurse under a will) which came before the Queensland Nursing Tribunal. The decisions of the tribunal, the District Court and the Court of Appeal provide practical guidance to nurses and midwives on the importance of establishing and maintaining the professional boundaries essential to the therapeutic relationship with their patients and clients.


Tumor necrosis factor alpha is a potent synergistic factor for the proliferation of primitive human hematopoietic progenitor cells and induces resistance to transforming growth factor beta but not to interferon gamma.
Since tumor necrosis factor (TNF)-alpha, interferon (IFN)-gamma, and transforming growth factor (TGF)-beta have all been shown to be specific inhibitors of early human hematopoiesis, we wanted to investigate the interactions of these three cytokines on very primitive human adult bone marrow CD34++CD38- hematopoietic progenitor cells, using a pre-colony-forming cell (pre-CFC) assay, which detects the effects of these cytokines on the initial phases of the differentiation of these primitive progenitors, which are unresponsive to interleukin (IL) 3 alone. Surprisingly, TNF-alpha was a very potent stimulator of the proliferation of CD34++CD38- cells and was the most potent synergistic factor for the IL-3-induced proliferation of these cells of all cytokines tested (IL-1, IL-6, granulocyte colony-stimulating factor, kit ligand). TNF-alpha was the only cytokine that, as a single added factor, induced substantial proliferation in CD34++CD38- cells in the presence of IL-3, except for kit ligand, which induced very limited proliferation. TNF-alpha, moreover, induced a high degree of resistance to the inhibitory effects of TGF-beta in a dose-dependent way. The inhibitory effects of IFN-gamma, however, were not affected by the presence of TNF-alpha. We hypothesize that in situations of the hematopoietic stress, TNF-alpha may abrogate the inhibitory effect of ambient TGF-beta in the bone marrow microenvironment to allow primitive stem cells to proliferate and differentiate in response to an increased demand for mature blood cells.


Sex trafficking and sexual exploitation in settings affected by armed conflicts in Africa, Asia and the Middle East: systematic review.
Sex trafficking and sexual exploitation has been widely reported, especially in conflict-affected settings, which appear to increase women's and children's vulnerabilities to these extreme abuses. We conducted a systematic search of ten databases and extensive grey literature to gather evidence of sex trafficking and sexual exploitation in conflict-affected settings. International definitions of "sexual exploitation" and "sex trafficking" set the indicator parameters. We focused on sexual exploitation in forms of early or forced marriage, forced combatant sexual exploitation and sexual slavery. We extracted prevalence measures, health outcomes and sexual exploitation terminology definitions. The review adhered to PRISMA guidelines and includes quality appraisal. The search identified 29 eligible papers with evidence of sex trafficking and sexual exploitation in armed conflict settings in twelve countries in Africa, Asia, and the Middle East. The evidence was limited and not generalizable, due to few prevalence estimates and inconsistent definitions of "sexual exploitation". The prevalence estimates available indicate that females were more likely than males to be victims of sexual exploitation in conflict settings. In some settings, as many as one in four forced marriages took place before the girls reached 18 years old. Findings suggest that the vast majority of former female combatants were sexually exploited during the conflict. These studies provided various indicators of sexual exploitation compatible to the United Nation's definition of sex trafficking, but only 2 studies identified the exploitation as trafficking. None of the studies solely aimed to measure the prevalence of sex trafficking or sexual exploitation. Similar descriptions of types of sexual exploitation and trafficking were found, but the inconsistent terminology or measurements inhibited a meta-analysis. Findings indicate there are various forms of human trafficking and sexual exploitation in conflict-affected settings, primarily occurring as early or forced marriage, forced combatant sexual exploitation, and sexual slavery. The studies highlight the extraordinary vulnerability of women and girls to these extreme abuses. Simultaneously, this review suggests the need to clarify terminology around sex trafficking in conflict to foster a more cohesive future evidence-base, and in particular, robust prevalence figures from conflict-affected and displaced populations.


Cytological maps of lampbrush chromosomes of European water frogs (Pelophylax esculentus complex) from the Eastern Ukraine.
Hybridogenesis (hemiclonal inheritance) is a kind of clonal reproduction in which hybrids between parental species are reproduced by crossing with one of the parental species. European water frogs (Pelophylax esculentus complex) represent an appropriate model for studying interspecies hybridization, processes of hemiclonal inheritance and polyploidization. P. esculentus complex consists of two parental species, P. ridibundus (the lake frog) and P. lessonae (the pool frog), and their hybridogenetic hybrid - P. esculentus (the edible frog). Parental and hybrid frogs can reproduce syntopically and form hemiclonal population systems. For studying mechanisms underlying the maintenance of water frog population systems it is required to characterize the karyotypes transmitted in gametes of parental and different hybrid animals of both sexes. In order to obtain an instrument for characterization of oocyte karyotypes in hybrid female frogs, we constructed cytological maps of lampbrush chromosomes from oocytes of both parental species originating in Eastern Ukraine. We further identified certain molecular components of chromosomal marker structures and mapped coilin-rich spheres and granules, chromosome associated nucleoli and special loops accumulating splicing factors. We recorded the dissimilarities between P. ridibundus and P. lessonae lampbrush chromosomes in the length of orthologous chromosomes, number and location of marker structures and interstitial (TTAGGG)n-repeat sites as well as activity of nucleolus organizer. Satellite repeat RrS1 was mapped in centromere regions of lampbrush chromosomes of the both species. Additionally, we discovered transcripts of RrS1 repeat in oocytes of P. ridibundus and P. lessonae. Moreover, G-rich transcripts of telomere repeat were revealed in association with terminal regions of P. ridibundus and P. lessonae lampbrush chromosomes. The constructed cytological maps of lampbrush chromosomes of P. ridibundus and P. lessonae provide basis to define the type of genome transmitted within individual oocytes of P. esculentus females with different ploidy and from various population systems.


Principal angles separate subject illumination spaces in YDB and CMU-PIE.
The theory of illumination subspaces is well developed and has been tested extensively on the Yale Face Database B (YDB) and CMU-PIE (PIE) data sets. This paper shows that if face recognition under varying illumination is cast as a problem of matching sets of images to sets of images, then the minimal principal angle between subspaces is sufficient to perfectly separate matching pairs of image sets from nonmatching pairs of image sets sampled from YDB and PIE. This is true even for subspaces estimated from as few as six images and when one of the subspaces is estimated from as few as three images if the second subspace is estimated from a larger set (10 or more). This suggests that variation under illumination may be thought of as useful discriminating information rather than unwanted noise.


Fractal geometry and the pharmacometrics of micafungin in overweight, obese, and extremely obese people.
The majority of Americans are overweight, and the incidence of obesity continues to increase. This trend predisposes people to a number of deleterious consequences, including the metabolic syndrome and other conditions that lead to a greater number of hospital admissions. Invasive candidiasis is an important nosocomial infection that results from these admissions. Echinocandins such as micafungin are indicated for treatment. We have previously demonstrated that overweight patients exhibit higher micafungin systemic clearance (SCL) than leaner patients. We hypothesized that obese and extremely obese people would show even higher SCL than merely overweight patients. To test this, we performed a prospective study of 36 adult volunteers randomized to receive a single dose of either 100 mg or 300 mg of micafungin whose body mass index fell within one of the following categories: <25, 25 to 40, and >40 kg/m(2). The male-to-female ratio was 1:1. The minimum weight was 43 kg, the median 97 kg, and the maximum weight 155 kg. A two-compartment model was examined using the maximum likelihood solution via the expectation-maximization algorithm. Men had a higher median SCL of 1.53 liters/h versus 1.29 liters/h (P = 0.01) in the Mann-Whitney U-test. The typical SCL was 1.04 liters/h but increased by a factor of (weight/66)(0.75) as weight increased above 66 kg. Thus, the relationship between micafungin SCL and weight in adults is best described by fractal-geometry-based laws. Furthermore, micafungin SCL continues to increase as weight increases, with no obvious plateau. This leads to a requirement for strategies to determine individualized dosing levels for obese and extremely obese patients.


Process evaluation of the systematic medical appraisal, referral and treatment (SMART) mental health project in rural India.
Availability of basic mental health services is limited in rural areas of India. Health system and individual level factors such as lack of mental health professionals and infrastructure, poor awareness about mental health, stigma related to help seeking, are responsible for poor awareness and use of mental health services. We implemented a mental health services delivery model that leveraged technology and task sharing to facilitate identification and treatment of common mental disorders (CMDs) such as stress, depression, anxiety and suicide risk in rural areas of the state of Andhra Pradesh, India. The intervention was delivered by lay village health workers (Accredited Social Health Activists - ASHAs) and primary care doctors. An anti-stigma campaign was implemented prior to this activity. This paper reports the process evaluation of the intervention using mixed methods. A mixed methods pre-post evaluation assessed the intervention using quantitative service usage analytics from the server, and qualitative interviews with different stakeholders. Barriers and facilitators in implementing the intervention were identified. Health service use increased significantly at post-intervention, ASHAs could followup 78.6% of those who had screened positive, and 78.6% of the 1243 Interactive Voice Response System calls made, were successful. Most respondents were aware of the intervention. They indicated that knowledge received through the intervention empowered them to approach ASHAs and share their mental health symptoms. ASHAs and doctors opined that EDSS was useful and easy to use. Medical camps organized in villages to increase access to the doctor were received positively by all. However, some aspects or facilitators of the intervention need to be improved, including network connectivity, booster training, anti-stigma campaigns, quality of mental health services provided by doctors, provision of psychotropic medications at primary health centers and frequency of health camps. The respondents' views helped to understand the barriers and facilitators for improving the likely effectiveness of the intervention using Andersen's Modified Behavioral Model of Health Services Use, and identify the mechanisms by which those factors affected mental health services uptake in the community. The study is registered with Clinical Trials Registry India (Applied - 16/07/14-Ref2014/07/007256; registration received - 04/10/17-CTRI/2017/10/009992 ).


Emerging Toxicity Models from Emerging Scientists.
Toxicology has seen a recent influx of new talents from other fields attracted by the application of their expertise to pressing questions of toxicological and environmental relevance. This transition has opened the door to innovative and exciting scientific opportunities but has also generated a new set of questions and challenges. In this viewpoint article, I will highlight some of the drivers and hurdles encountered by the recent new breeds of toxicologists.


Resetting of aortic baroreceptors with non-myelinated afferent fibers in spontaneously hypertensive rats.
The phenomenon of resetting and the possible mechanisms responsible for it were investigated in aortic baroreceptors having unmyelinated axons. Recordings were obtained from 31 aortic baroreceptor C-fibers in 20 normotensive Wistar Kyoto rats (WKY) and 37 baroreceptor C-fibers in 11 spontaneously hypertensive rats (SHR). The recordings were obtained from the left aortic nerve of an in vitro aortic nerve-aortic arch preparation. The thresholds for activation of baroreceptor C-fibers were not different between SHR's and WKY's at 16 weeks of age (139 +/- 7 and 133 +/- 5 mmHg, respectively). However, the thresholds for activation in SHR at 36 weeks of age were considerably higher (163 +/- 4 mmHg) indicating resetting of the receptors in the older rats. The pressure-discharge curves were not different between SHR's and WKY's at 16 weeks of age and WKY's at 35 weeks of age, but were shifted to the right for SHR's at 36 weeks of age. The receptors showed greater responses to pulsatile pressures than to steady state pressures at the same mean level of pressure. We found that aortic C-fibers were not reset during the early established phase of hypertension in SHR's because of enhanced receptor strain sensitivity. This is likely to be of importance for tonic vasomotor control at this time. In later stages of hypertension and despite enhanced strain sensitivity the endings were reset to higher pressures probably due to an increased collagen content in the aortic wall.


Factors predicting complication rates following total knee replacement.
The purpose of this investigation was to expand on previous studies by more fully examining the role of a variety of patient and hospital characteristics in determining adverse outcomes following total knee replacement. With use of data from all hospital admissions in California from 1991 through 2001, multiple logistic regression was performed on the information regarding patients treated with total knee replacement. Rates of mortality and readmission due to infection and pulmonary embolism during the first ninety days after discharge were regressed against a variety of independent variables, including demographic factors (age, gender, race, ethnicity, and insurance type), burden of comorbid disease (Charlson comorbidity index), and provider variables (hospital size, teaching status, and surgical volume). A separate baseline probability analysis was then performed to compare the relative importance of all predictor variables. The sample size for this analysis was 222,684. A total of 1176 deaths (rate, 0.53%), 1586 infections (0.71%), and 914 pulmonary emboli (0.41%) occurred within the first ninety days after discharge. The average age of the patients at the time of surgery was sixty-nine years. Sixty-two percent of the patients were women, and 32% had a Charlson comorbidity index of >0. The significant predictors for complications (p < 0.05) included age, gender, race/ethnicity, Charlson comorbidity index, insurance type, and hospital volume. A baseline probability analysis was performed with the base case considered to be a white woman who was over the age of sixty-five years, had a Charlson comorbidity index of 0, had Medicare insurance, and was treated at a high-volume, non-teaching hospital. For a patient with the baseline case characteristics, the probability of death was 31/10,000, the probability of infection was 59/10,000, and the probability of pulmonary embolism was 41/10,000 in the first ninety days after discharge. Altering the base case by assuming that care was received at a low-volume hospital increased the expected mortality rate by a factor of 26%. Increasing the Charlson comorbidity index to 1 increased the mortality rate by 170%, whereas decreasing the age to younger than sixty-five years lowered the mortality rate by 73%. Hospital volume, comorbidity, and age had similar effects on the expected rates of readmission due to infection and pulmonary embolism. The effects of age and the Charlson comorbidity index on the baseline probability of adverse outcomes following total knee replacement were shown to be similar to or greater than the effect of hospital volume. This study elucidates and compares the relative importance of the effects of several different factors on outcome. This information is important when considering the conclusions and implications of this type of policy-relevant outcomes research.


Bound water structure and polymorphic amino acids act together to allow the binding of different peptides to MHC class I HLA-B53.
The structure of the human MHC class I molecule HLA-B53 complexed to two nonameric peptide epitopes (from the malaria parasite P. falciparum and the HIV2 gag protein) has been determined by X-ray crystallography at 2.3 angstrom resolution. The structures reveal the architecture of a Pro-specific B pocket common to many HLA-B alleles. Relative to other alleles, the B53 peptide-binding groove is widened by a significant (up to 1.25 angstrom) shift in the position of the alpha 1 helix. Within this groove, bound water molecules, acting in concert with the side chains of polymorphic residues, provide the functional malleability of the MHC, which enables the high affinity/low specificity binding of multiple peptide epitopes.


Genetic relationships determined by a DNA heteroduplex mobility assay: analysis of HIV-1 env genes.
The genetic diversity of human immunodeficiency virus (HIV) is a major concern thought to impact on immunologic escape and eventual vaccine efficacy. Here, simple and rapid methods are described for the detection and estimation of genetic divergence between HIV strains on the basis of the observation that DNA heteroduplexes formed between related sequences have a reduced mobility in polyacrylamide gels proportional to their degree of divergence. Reliable phylogenetic subtypes were assigned for HIV-1 strains from around the world. Relationships between viruses were closest when derived from the same or epidemiologically linked individuals. When derived from epidemiologically unlinked individuals, the relationships between viruses in a given geographic region correlated with the length of time HIV-1 had been detected in the population and the number of strains initiating widespread infection. Heteroduplex mobility analysis thus provides a tool to expedite epidemiological investigations by assisting in the classification of HIV and is readily applicable to the screening and characterization of other infectious agents and cellular genes.


Discovery of stimulator binding to a conserved pocket in the heme domain of soluble guanylyl cyclase.
Soluble guanylyl cyclase (sGC) is the receptor for nitric oxide and a highly sought-after therapeutic target for the management of cardiovascular diseases. New compounds that stimulate sGC show clinical promise, but where these stimulator compounds bind and how they function remains unknown. Here, using a photolyzable diazirine derivative of a novel stimulator compound, IWP-051, and MS analysis, we localized drug binding to the β1 heme domain of sGC proteins from the hawkmoth Manduca sexta and from human. Covalent attachments to the stimulator were also identified in bacterial homologs of the sGC heme domain, referred to as H-NOX domains, including those from Nostoc sp. PCC 7120, Shewanella oneidensis, Shewanella woodyi, and Clostridium botulinum, indicating that the binding site is highly conserved. The identification of photoaffinity-labeled peptides was aided by a signature MS fragmentation pattern of general applicability for unequivocal identification of covalently attached compounds. Using NMR, we also examined stimulator binding to sGC from M. sexta and bacterial H-NOX homologs. These data indicated that stimulators bind to a conserved cleft between two subdomains in the sGC heme domain. L12W/T48W substitutions within the binding pocket resulted in a 9-fold decrease in drug response, suggesting that the bulkier tryptophan residues directly block stimulator binding. The localization of stimulator binding to the sGC heme domain reported here resolves the longstanding question of where stimulators bind and provides a path forward for drug discovery.