Modeling metastasis biology and therapy in real time in the mouse lung.
Pulmonary metastasis remains the leading ca use of death for cancer patients. Opportunities to improve treatment outcomes for patients require new methods to study and view the biology of metastatic progression. Here, we describe an ex vivo pulmonary metastasis assay (PuMA) in which the metastatic progression of GFP-expressing cancer cells, from a single cell to the formation of multicellular colonies, in the mouse lung microenvironment was assessed in real time for up to 21 days. The biological validity of this assay was confirmed by its prediction of the in vivo behavior of a variety of high- and low-metastatic human and mouse cancer cell lines and the discrimination of tumor microenvironments in the lung that were most permissive to metastasis. Using this approach, we provide what we believe to be new insights into the importance of tumor cell interactions with the stromal components of the lung microenvironment. Finally, the translational utility of this assay was demonstrated through its use in the evaluation of therapeutics at discrete time points during metastatic progression. We believe that this assay system is uniquely capable of advancing our understanding of both metastasis biology and therapeutic strategies.


The interplay of genetic and environmental factors in shaping well-being across the lifespan: Evidence from the serotonin transporter gene.
Converging evidence suggests that well-being plays an important role in promoting and maintaining mental health across the life span. It has been shown that well-being has a considerable heritable component, but little is known about the specific genes involved. In this study, we investigated a healthy sample (N = 298) that was genotyped for the serotonin transporter-linked polymorphic region (5-HTTLPR). We hypothesized that 5-HTTLPR gene variation would influence well-being, and additionally investigated interaction effects with age and the environmental influence of early life stress (ELS). Using multiple regression, our results showed a significant three-way interaction between genotype, ELS, and age. Exploration of this interaction showed that young subjects had decreased levels of well-being if they were exposed to ELS and homozygous for the short variant of 5-HTTLPR. This relationship was reversed in old age: subjects that were exposed to ELS and carried the long variant of 5-HTTLPR had decreased levels of well-being. Our results indicate that genetic and environmental factors have joint effects on well-being that are susceptible to profound changes across the life span.


The ways that people talk about natural resources: discursive strategies as barriers to environmentally sustainable practices.
In this paper, we analyse talk about water and energy use taken from nine interviews with citizens of Perth, Western Australia. Participants' talk offered representations of water as a scarce, shared, natural resource that must not be wasted, whereas talk about energy use focused on the environmental impacts of different technologies for generating electricity, rather than on energy as a consumable resource. Participants accounted for their water-use habits by positioning themselves as caught between a personal desire to conserve water and an (incompatible) social obligation to maintain the appearance of their gardens in keeping with the aesthetic appeal of the suburbs in which they lived. We identify several discursive strategies by which people construct the environmental impact of their actions as minimal or unavoidable. These constitute a barrier to the promotion of more environmentally sustainable practices. Potential implications for environmental policy development are discussed, as are the wider issues associated with the development of 'applied' discourse analysis.


Assortment and packaging of the segmented rotavirus genome.
The rotavirus (RV) genome comprises 11 segments of double-stranded RNA (dsRNA) and is contained within a non-enveloped, icosahedral particle. During assembly, a highly coordinated selective packaging mechanism ensures that progeny RV virions contain one of each genome segment. Cis-acting signals thought to mediate assortment and packaging are associated with putative panhandle structures formed by base-pairing of the ends of RV plus-strand RNAs (+RNAs). Viral polymerases within assembling core particles convert the 11 distinct +RNAs to dsRNA genome segments. It remains unclear whether RV +RNAs are assorted before or during encapsidation, and the functions of viral proteins during these processes are not resolved. However, as reviewed here, recent insights gained from the study of RV and two other segmented RNA viruses, influenza A virus and bacteriophage Φ6, reveal potential mechanisms of RV assortment and packaging.


Totally laparoscopic Roux-en-Y duct-to-mucosa pancreaticojejunostomy after middle pancreatectomy: a consecutive nine-case series at a single institution.
To present the results of a series of laparoscopic middle pancreatectomies with roux-en-Y duct-to-mucosa pancreaticojejunostomy. Middle pancreatectomy makes it possible to preserve pancreatic parenchyma in the resection of lesions that traditionally have been treated by distal splenopancreatectomy or cephalic duodenopancreatectomy. The laparoscopic approach could minimize the invasiveness of the procedure and enhance the benefits of middle pancreatectomy. From March 2005 to October 2007, 9 consecutive patients with benign or low malignant potential lesions in the pancreatic neck or body underwent surgery. Laparoscopic middle pancreatectomy with a roux-en-Y duct-to-mucosa pancreaticojejunostomy was planned on all of them. In the first 2 patients, the pancreas was transected by endostapler; in the last 7, the staple line was reinforced with absorbable polymer membrane. The intervention was concluded laparoscopically in every case except 1 (laparoscopic-assisted) in which pancreaticojejunostomy was performed by means of minilaparotomy. Mortality was 0% and perioperative morbidity was 33%, (fistula of the cephalic stump in the first 2 patients (22%)). The pancreaticojejunostomy fistula rate was 0%. The median postoperative hospital stay was 5 days (range, 3-41). In the last 7 patients, in which pancreas was transected with staple line reinforcement material there were no stump fistulas; morbidity decreased to 14% and the median hospital stay was 4 days (range, 3-30). Laparoscopic middle pancreatectomy is feasible and safe. Duct-to-mucosa pancreaticojejunostomy can be performed safely using this approach. The method of pancreatic transection seems to be decisive in the incidence of cephalic stump fistulas.


Nuclear-specific gene expression in heterokaryons of the filamentous ascomycete Neurospora tetrasperma.
Heterokaryosis is a system in which genetically distinct nuclei coexist within the same cytoplasm. While heterokaryosis dominates the life cycle of many fungal species, the transcriptomic changes associated with the transition from homokaryosis to heterokaryosis is not well understood. Here, we analyse gene expression profiles of homokaryons and heterokaryons from three phylogenetically and reproductively isolated lineages of the filamentous ascomycete Neurospora tetrasperma. We show that heterokaryons are transcriptionally distinct from homokaryons in the sexual stage of development, but not in the vegetative stage, suggesting that the phenotypic switch to fertility in heterokaryons is associated with major changes in gene expression. Heterokaryon expression is predominantly defined by additive effects of its two nuclear components. Furthermore, allele-specific expression analysis of heterokaryons with varying nuclear ratios show patterns of expression ratios strongly dependent on nuclear ratios in the vegetative stage. By contrast, in the sexual stage, strong deviations of expression ratios indicate a co-regulation of nuclear gene expression in all three lineages. Taken together, our results show two levels of expression control: additive effects suggest a nuclear level of expression, whereas co-regulation of gene expression indicate a heterokaryon level of control.


Quinol oxidation by c-type cytochromes: structural characterization of the menaquinol binding site of NrfHA.
Membrane-bound cytochrome c quinol dehydrogenases play a crucial role in bacterial respiration by oxidizing menaquinol and transferring electrons to various periplasmic oxidoreductases. In this work, the menaquinol oxidation site of NrfH was characterized by the determination of the X-ray structure of Desulfovibrio vulgaris NrfHA nitrite reductase complex bound to 2-heptyl-4-hydroxyquinoline-N-oxide, which is shown to act as a competitive inhibitor of NrfH quinol oxidation activity. The structure, at 2.8-A resolution, reveals that the inhibitor binds close to NrfH heme 1, where it establishes polar contacts with two essential residues: Asp89, the residue occupying the heme distal ligand position, and Lys82, a strictly conserved residue. The menaquinol binding cavity is largely polar and has a wide opening to the protein surface. Coarse-grained molecular dynamics simulations suggest that the quinol binding site of NrfH and several other respiratory enzymes lie in the head group region of the membrane, which probably facilitates proton transfer to the periplasm. Although NrfH is not a multi-span membrane protein, its quinol binding site has several characteristics similar to those of quinone binding sites previously described. The data presented here provide the first characterization of the quinol binding site of the cytochrome c quinol dehydrogenase family.


The antiproliferative effect of mesenchymal stem cells is a fundamental property shared by all stromal cells.
Although it has been widely demonstrated that human mesenchymal stem cells exert potent immunosuppressive effects, there is little information as to whether more mature mesenchymal stromal cells (SC) share the same property. Accordingly, we set out to test the ability of SC from different human tissues to inhibit the proliferation of PBMC following polyclonal stimuli. Chondrocytes, as well as fibroblasts from synovial joints, lung, and skin, were used as a source of SC. Irrespective of their differentiation potential and/or content of progenitor cells, SC from all tissues exhibited antiproliferative functions. This was in marked contrast to parenchymal cells. Although SC did not interfere with early T lymphocyte activation, they arrested stimulated T cells in the G(0)/G(1) phase of the cell cycle and rescued them from apoptosis. In addition, IFN-gamma and TNF-alpha production were reduced. We observed that the inhibitory effect is ultimately mediated by soluble factors, the production of which requires SC to be licensed in an inflammatory environment by cell contact. We conclude that the immunosuppressive effect of mesenchymal cells is not confined to multipotent stem cells, but is a fundamental characteristic of all stroma. Our data suggest that SC, appropriately licensed, regulate T cell homeostasis.


The prolongation of the lifespan of rats by repeated oral administration of [60]fullerene.
Countless studies showed that [60]fullerene (C(60)) and derivatives could have many potential biomedical applications. However, while several independent research groups showed that C(60) has no acute or sub-acute toxicity in various experimental models, more than 25 years after its discovery the in vivo fate and the chronic effects of this fullerene remain unknown. If the potential of C(60) and derivatives in the biomedical field have to be fulfilled these issues must be addressed. Here we show that oral administration of C(60) dissolved in olive oil (0.8 mg/ml) at reiterated doses (1.7 mg/kg of body weight) to rats not only does not entail chronic toxicity but it almost doubles their lifespan. The effects of C(60)-olive oil solutions in an experimental model of CCl(4) intoxication in rat strongly suggest that the effect on lifespan is mainly due to the attenuation of age-associated increases in oxidative stress. Pharmacokinetic studies show that dissolved C(60) is absorbed by the gastro-intestinal tract and eliminated in a few tens of hours. These results of importance in the fields of medicine and toxicology should open the way for the many possible -and waited for- biomedical applications of C(60) including cancer therapy, neurodegenerative disorders, and ageing.


Structural insights into broadly neutralizing antibodies elicited by hybrid immunity against SARS-CoV-2.
ABSTRACTIncreasing spread by SARS-CoV-2 Omicron variants challenges existing vaccines and broadly reactive neutralizing antibodies (bNAbs) against COVID-19. Here we determine the diversity, potency, breadth and structural insights of bNAbs derived from memory B cells of BNT162b2-vaccinee after homogeneous Omicron BA.1 breakthrough infection. The infection activates diverse memory B cell clonotypes for generating potent class I/II and III bNAbs with new epitopes mapped to the receptor-binding domain (RBD). The top eight bNAbs neutralize wildtype and BA.1 potently but display divergent IgH/IgL sequences and neuralization profiles against other variants of concern (VOCs). Two of them (P2D9 and P3E6) belonging to class III NAbs display comparable potency against BA.4/BA.5, although structural analysis reveals distinct modes of action. P3E6 neutralizes all variants tested through a unique bivalent interaction with two RBDs. Our findings provide new insights into hybrid immunity on BNT162b2-induced diverse memory B cells in response to Omicron breakthrough infection for generating diverse bNAbs with distinct structural basis.


Release of calcium from endolysosomes increases calcium influx through N-type calcium channels: Evidence for acidic store-operated calcium entry in neurons.
Neurons possess an elaborate system of endolysosomes. Recently, endolysosomes were found to have readily releasable stores of intracellular calcium; however, relatively little is known about how such 'acidic calcium stores' affect calcium signaling in neurons. Here we demonstrated in primary cultured neurons that calcium released from acidic calcium stores triggered calcium influx across the plasma membrane, a phenomenon we have termed "acidic store-operated calcium entry (aSOCE)". aSOCE was functionally distinct from store-operated calcium release and calcium entry involving endoplasmic reticulum. aSOCE appeared to be governed by N-type calcium channels (NTCCs) because aSOCE was attenuated significantly by selectively blocking NTCCs or by siRNA knockdown of NTCCs. Furthermore, we demonstrated that NTCCs co-immunoprecipitated with the lysosome associated membrane protein 1 (LAMP1), and that aSOCE is accompanied by increased cell-surface expression levels of NTCC and LAMP1 proteins. Moreover, we demonstrated that siRNA knockdown of LAMP1 or Rab27a, both of which are key proteins involved in lysosome exocytosis, attenuated significantly aSOCE. Taken together our data suggest that aSOCE occurs in neurons, that aSOCE plays an important role in regulating the levels and actions of intraneuronal calcium, and that aSOCE is regulated at least in part by exocytotic insertion of N-type calcium channels into plasma membranes through LAMP1-dependent lysosome exocytosis.


Visual field preservation after multisession cyberknife radiosurgery for perioptic lesions.
The restricted radiation tolerance of the anterior visual pathways represents a unique challenge for ablating adjacent lesions with single-session radiosurgery. Although preliminary studies have recently demonstrated that multisession radiosurgery for selected perioptic tumors is both safe and effective, the number of patients in these clinical series was modest and the length of follow-up limited. The current retrospective study is intended to help address these shortcomings. Forty-nine consecutive patients with meningioma (n = 27), pituitary adenoma (n = 19), craniopharyngioma (n = 2), or mixed germ cell tumor (n = 1) situated within 2 mm of a "short segment" of the optic apparatus underwent multisession image-guided radiosurgery at Stanford University Medical Center. Thirty-nine of these patients had previous subtotal surgical resection, and six had previously been treated with conventional fractionated radiotherapy (6). CyberKnife radiosurgery was delivered in two to five sessions to an average tumor volume of 7.7 cm3 and a cumulative average marginal dose of 20.3 Gy. Formal visual testing and clinical examinations were performed before treatment and at follow-up intervals beginning at 6 months. After a mean visual field follow-up of 49 months (range, 6-96 mo), vision was unchanged postradiosurgery in 38 patients, improved in eight (16%), and worse in three (6%). In each instance, visual deterioration was accompanied by tumor progression that ultimately resulted in patient death. However, one of these patients, who had a multiply recurrent adrenocorticotropic hormone-secreting pituitary adenoma, initially experienced early visual loss without significant tumor progression after both a previous course of radiotherapy and three separate sessions of radiosurgery. After a mean magnetic resonance imaging follow-up period of 46 months, tumor volume was stable or smaller in all other cases. Two patients died of unrelated nonbrain causes. Multisession radiosurgery resulted in high rates of tumor control and preservation of visual function in this group of perioptic tumors. Ninety-four percent of patients retained or improved preradiosurgical vision. This intermediate-term experience reinforces the findings from earlier studies that suggested that multisession radiosurgery can be a safe and effective alternative to either surgery or fractionated radiotherapy for selected lesions immediately adjacent to short segments of the optic apparatus.


Microcalorimetry does not predict the cellular phagocytosis of latex microspheres.
Current literature highlights the potential suitability of microcalorimetry for the investigation of cell-drug interactions. Previous work using bacteria or antigens derived from infectious organisms yielded conclusions that heat production is a quantitative means of measuring phagocytosis. In this study we evaluated the potential of flow-through microcalorimetry as a method of quantifying the phagocytosis of microsphere particulates. The technique avoids the need to incorporate radioactive or fluorescent markers into the particulate formulation, and would be widely applicable in biopharmaceutical research. Using the monocyte cell line Mono Mac 6 a power output of 9.00 microW per million cells was increased significantly on addition of zymosan, lipopolysaccaride (LPS) and phorbol myristate acetate but not following exposure to FITC labelled latex microspheres (LM). TNFalpha production increased on exposure to zymosan, LPS and LPS-phorbol myristate acetate, though not on exposure to LB. An assay was developed which allowed the quantification of internalised particulates in phagocytic cells using fluorescent activated cell sorting (FACS). In contrast to the microcalorimetric and TNFalpha data FACS revealed that 20% of the MM6 population phagocytosed a mean of 1.35 LM. Microcalorimetry and measurements of TNFalpha production are assays of cellular activation a phenomenon not necessarily associated with phagocytosis. FACS, however, serves as a specific and quantitative measure of phagocytosis. Microcalorimetry may not be a suitable technique for the quantitative assessment of the phagocytosis of drug delivery particulates.


Identification of Novel Disease-Relevant Genes and Pathways in the Pathogenesis of Type 1 Diabetes: A Potential Defect in Pancreatic Iron Homeostasis.
Multiple pathways contribute to the pathophysiological development of type 1 diabetes (T1D); however, the exact mechanisms involved are unclear. We performed differential gene expression analysis in pancreatic islets of NOD mice versus age-matched congenic NOD.B10 controls to identify genes that may contribute to disease pathogenesis. Novel genes related to extracellular matrix development and glucagon and insulin signaling/secretion were changed in NOD mice during early inflammation. During "respective" insulitis, the expression of genes encoding multiple chemosensory olfactory receptors were upregulated, and during "destructive" insulitis, the expression of genes involved in antimicrobial defense and iron homeostasis were downregulated. Islet inflammation reduced the expression of Hamp that encodes hepcidin. Hepcidin is expressed in β-cells and serves as the key regulator of iron homeostasis. We showed that Hamp and hepcidin levels were lower, while iron levels were higher in the pancreas of 12-week-old NOD versus NOD.B10 mice, suggesting that a loss of iron homeostasis may occur in the islets during the onset of "destructive" insulitis. Interestingly, we showed that the severity of NOD disease correlates with dietary iron intake. NOD mice maintained on low-iron diets had a lower incidence of hyperglycemia, while those maintained on high-iron diets had an earlier onset and higher incidence of disease, suggesting that high iron exposure combined with a loss of pancreatic iron homeostasis may exacerbate NOD disease. This mechanism may explain the link seen between high iron exposure and the increased risk for T1D in humans.


Targeted polymersome delivery of siRNA induces cell death of breast cancer cells dependent upon Orai3 protein expression.
Polymersomes, polymeric vesicles that self-assemble in aqueous solutions from block copolymers, have been avidly investigated in recent years as potential drug delivery agents. Past work has highlighted peptide-functionalized polymersomes as a highly promising targeted delivery system. However, few reports have investigated the ability of polymersomes to operate as gene delivery agents. In this study, we report on the encapsulation and delivery of siRNA inside of peptide-functionalized polymersomes composed of poly(1,2-butadiene)-b-poly(ethylene oxide). In particular, PR_b peptide-functionalized polymer vesicles are shown to be a promising system for siRNA delivery. PR_b is a fibronectin mimetic peptide targeting specifically the α(5)β(1) integrin. The Orai3 gene was targeted for siRNA knockdown, and PR_b-functionalized polymer vesicles encapsulating siRNA were found to specifically decrease cell viability of T47D breast cancer cells to a certain extent, while preserving viability of noncancerous MCF10A breast cells. siRNA delivery by PR_b-functionalized polymer vesicles was compared to that of a current commercial siRNA transfection agent, and produced less dramatic decreases in cancer cell viability, but compared favorably in regards to the relative toxicity of the delivery systems. Finally, delivery and vesicle release of a fluorescent encapsulate by PR_b-functionalized polymer vesicles was visualized by confocal microscopy, and colocalization with cellular endosomes and lysosomes was assessed by organelle staining. Polymersomes were observed to primarily release their encapsulate in the early endosomal intracellular compartments, and data may suggest some escape to the cytosol. These results represent a promising first generation model system for targeted delivery of siRNA.


CD24 offers a therapeutic target for control of bladder cancer metastasis based on a requirement for lung colonization.
Metastasis is lethal in most bladder cancer patients. Expression of CD24, a glycosyl phosphatidylinositol (GPI)-linked sialoglycoprotein and cancer stem cell marker, is associated with metastatic progression in multiple cancer types, yet the role of CD24 in this process remains unclear. While developing a murine model of human metastatic bladder cancer, we observed that tumor cell CD24 expression correlated with a propensity to metastasize to the lung. Our immunohistochemical evaluation of 60 paired primary and metastatic human bladder cancer samples revealed increased intensity (P < 0.001) and frequency (P < 0.001) of CD24 expression in metastases. To directly evaluate the role of CD24 in metastatic colonization, we manipulated CD24 expression in human bladder cancer cell lines using short hairpin RNA depletion, cDNA overexpression, and fluorescence-activated cell sorting selection. Although suppression of CD24 reduced acute tumor cell retention in the lungs of mice inoculated intravenously with cancer cells, this differential retention was no longer apparent after 24 hours, prompting us to evaluate the role of CD24 in lung colonization. Here, CD24 was found necessary for subsequent development of lung metastases. We next treated clinically detectable lung metastases in mice with anti-CD24 antibody and observed reduced tumor growth and prolonged survival. These findings suggest that CD24 is a lynchpin of metastatic progression and a promising therapeutic target for antimetastatic therapy.


A systematic review of antiepileptic drug initiation and withdrawal.
The proper clinical context to initiate or discontinue antiepileptic drug (AED) treatment has been extensively studied over the past 40 years. Differences in study design and methodology have led to reports of seizure recurrence rates of 7% to 96% in drug initiation trials and seizure relapse rates of 12% to 67% in drug withdrawal trials. A number of different historical and clinical factors have been cited in various studies as altering seizure risk. Our objective was to review the data from large, well-conducted, prospective studies. We performed an electronic search of 3 medical and scientific databases for articles on initiating or withdrawing antiepileptic medication. Our review focused primarily on articles meeting specific inclusion and exclusion criteria. We discuss the risk of seizure recurrence after a first seizure and the factors associated with an elevated risk of recurrence. The risks of treatment, including life-threatening idiosyncratic reactions and dose-related physical and cognitive side effects are reviewed. We also summarize the data on the seizure relapse risk after drug discontinuation and the factors associated with an increased relapse risk. Benefits of successful AED withdrawal are highlighted. The special considerations of treatment initiation and withdrawal in the pediatric population are explored. We propose general guidelines to assist the clinician in evaluating the risk/benefit ratio of initiating and withdrawing antiepileptic drugs. Drug initiation after a first seizure decreases early seizure recurrence, but does not affect the long-term prognosis of developing epilepsy. Medication withdrawal after a period of seizure remission increases the risk of relapse, but the benefits of successful AED discontinuation may be substantial. In the end, the decision of whether to initiate treatment after a single seizure and whether to withdraw AED therapy in patients enjoying a prolonged period of seizure freedom should be made on an individual case basis, which balances the risk of seizure relapse and subsequent disability against the likely impact of medication-related physical, cognitive, and psychologic adverse effects.


Who sleeps best? Longitudinal patterns and covariates of change in sleep quantity, quality, and timing across four university years.
This study tracked change over time in sleep quantity, disturbance, and timing, and sleep's covariations with living situation, stress, social support, alcohol use, and grade point average (GPA) across four years of university in 186 Canadian students. Women slept longer as they moved through university, and men slept less; rise times were later each year. Students reported sleeping fewer hours, more sleep disturbances, and later rise times during years with higher stress. In years when students lived away from home, they reported more sleep disturbances, later bedtimes, and later rise times. Living on campus was associated with later bedtimes and rise times. Alcohol use was higher and GPA was lower when bedtimes were later. The implications of these observed patterns for understanding the correlates and consequences of university students' sleep are discussed.


Groundwater stress and vulnerability in rural coastal aquifers under competing demands: a case study from Sri Lanka.
Rural coastal aquifers are undergoing rapid changes due to increasing population, high water demand with expanding agricultural and domestic uses, and seawater intrusion due to unmanaged water pumping. The combined impact of these activities is the deterioration of groundwater quality, public health concerns, and unsustainable water demands. The Kalpitiya peninsula located northwest of Sri Lanka is one area undergoing such changes. This land area is limited and surrounded almost completely by sea and lagoon. This study consists of groundwater sampling and analysis, and vulnerability assessment using the DRASTIC method. The results reveal that the peninsula is experiencing multiple threats due to population growth, seawater intrusion, land use exploitation for intensive agriculture, groundwater vulnerability from agricultural and domestic uses, and potential public health impacts. Results show that nitrate is a prevalent and serious contaminant occurring in large concentrations (up to 128 mg/l NO(3)-N), while salinity from seawater intrusion produces high chloride content (up to 471 mg/l), affecting freshwater sources. High nitrate levels may have already affected public health based on limited sampling for methemoglobin. The two main sources of nitrogen loadings in the area are fertilizer and human excreta. The major source of nitrogen results from the use of fertilizers and poor management of intense agricultural systems where a maximum application rate of up to 11.21 metric tons N/km(2) per season is typical. These findings suggest that management of coastal aquifers requires an integrated approach to address both the prevalence of agriculture as an economic livelihood, and increasing population growth.


Post-testicular sperm maturation and identification of an epididymal protein in the Japanese quail (Coturnix coturnix japonica).
The role of the avian epididymis in post-testicular development and capacitation was examined to assess whether avian spermatozoa undergo any processes similar to those characteristic of mammalian sperm development. We found no evidence of a need for quail sperm to undergo capacitation and 90% of testicular sperm could bind to a perivitelline membrane and acrosome react. However, computer-assisted sperm analysis showed that 20% of testicular sperm from the quail were capable of movement and only about 12% of the motile sperm would have a curvilinear velocity greater than the mean for sperm from the distal epididymis. Nevertheless, epididymal transit was associated with increases in mean sperm velocity and the proportion of motile sperm. Together, these findings explain why earlier workers have achieved some fertilizations following inseminations of testicular spermatozoa and also demonstrate the need for some epididymal maturation of avian spermatozoa. Analysis of the electrophoretic profile of quail epididymal luminal proteins revealed that only one major protein (∼16 kDa) is secreted by the epididymis and it was virtually the only protein secreted by the ipsilateral epididymis following unilateral orchidectomy. Mass spectrometry showed that this protein is hemoglobin; this finding was confirmed using anti-hemoglobin antibodies. It is suggested that hemoglobin may support sperm metabolism in the quail epididymis, aid in motility, and/or serve as an antioxidant.


Are the metabolic components of crassulacean acid metabolism up-regulated in response to an increase in oxidative burden?
In the halophytic species Mesembryanthemum crystallinum, crassulacean acid metabolism (CAM) may be induced by a range of abiotic factors including drought, salinity, high light intensity, low temperature, and anoxia. A key biotic consequence of all these environmental changes is the generation of reactive oxygen species in planta that can elicit potentially damaging oxidative reactions and/or act as signals for engaging mechanisms that alleviate oxidative stress. However, induction of CAM per se also has the potential for increasing the oxidative burden via the enhanced internal O2 concentrations that develop behind closed stomata during daytime decarboxylation. The aim of this paper was to test two hypotheses. The first one, that reactive oxygen species are key signals for up-regulating the major genes and proteins required for the operation of CAM as part of an integrated strategy for alleviating oxidative burden, was tested using gaseous ozone to increase the oxidative burden at a cellular level. The second hypothesis, that CAM potentially increases oxidative load, was tested using a CAM-deficient mutant of M. crystallinum. The data indicate that ozone, like salinity, elicits an increase in the transcript and protein abundance of myo-inositol o-methyl transferase (a key enzyme of cyclitol synthesis), together with phosphoenolpyruvate carboxylase and other 'CAM-related' enzymes. However, ozone, unlike salinity, does not induce functional CAM, implying that the various metabolic components required for CAM respond to different signals. Comparing the activities of different subcellular isoforms of superoxide dismutase in wild-type and CAM-deficient mutants of M. crystallinum suggests that the induction of CAM potentially curtails the oxidative load in planta.


The Role of Stewardship in Addressing Antibacterial Resistance: Stewardship and Infection Control Committee of the Antibacterial Resistance Leadership Group.
Antibacterial resistance is increasing globally and has been recognized as a major public health threat. Antibacterial stewardship is the coordinated effort to improve the appropriate use of antibiotics with the aim to decrease selective pressure for multidrug-resistant organisms in order to preserve the utility of antibacterial agents. This article describes the activities of the Antibacterial Resistance Leadership Group (ARLG) in the area of antibacterial stewardship. To date, the ARLG has focused intensely on development of rapid diagnostic tests, which (when coupled with educational and institutional initiatives) will enable the robust stewardship that is needed to address the current crisis of antibacterial resistance. In addition to exploring the effectiveness of stewardship techniques in community hospitals, the ARLG has also developed strategy trials to assess the feasibility of reducing antibacterial usage while preserving patient outcome.


Lorazepam and meprobamate dose effects in humans: behavioral effects and abuse liability.
On a residential research ward, the acute effects of placebo, lorazepam (LZ) (1.5-9.0 mg) and meprobamate (MEP) (600-3600 mg) were examined using a within-subject double-blind Latin Square design in nine male subjects with histories of drug abuse. Drug effects were assessed with objective performance tasks, subject estimates of performance, staff ratings of drug effects and subject ratings of drug effects, sleep, mood, drug liking and monetary street value. Generally, both LZ and MEP produced comparable dose-related effects; LZ had a more rapid onset of action and on several measures showed a more shallow dose-response curve than MEP. With LZ, but not MEP, subjects under-estimated the degree to which their performance was impaired and under-rated drug effects as compared to analogous staff ratings. Both drugs produced sedation-like subject ratings of mood and sleep but generally did not produce tranquilization-like ratings. MEP produced subject ratings of drug liking and monetary street value which were equal to or in some cases greater than those of LZ. Relative potency estimations showed that LZ was 510 to 783 times more potent than MEP in producing performance impairment but was only 370 times more potent than MEP in producing subject ratings of drug liking. Overall, these data indicate that the likelihood of abuse of MEP is at least equal to if not greater than that of LZ although LZ may have a greater likelihood of producing adverse behavioral effects such as a performance impairment and under-estimates of the degree of impairment. These data in conjunction with previous results from this laboratory show that the behavioral effects of benzodiazepines can be differentiated from those of other types of sedative/anxiolytics and that MEP displays characteristics of a barbiturate-like profile of action.


Fluorescent pseudo-peptide linear vasopressin antagonists: design, synthesis, and applications.
Fluoresceinyl and rhodamyl groups have been coupled by an amide link to side-chain amino groups at positions 1, 6, and 8 of pseudo-peptide linear vasopressin antagonists (Manning et al. Int. J. Pept. Protein Res. 1992, 40, 261-267) through different positions on the fluorophore, to give tetraethylrhodamyl-DTyr(Me)-Phe-Gln-Asn-Arg-Pro-Arg-Tyr-NH2 (2), 4-HOPh(CH2)2CO-DTyr(Me)-Phe-Gln-Asn-Lys(5-carboxyfl uoresceinyl)-Pro-A rg-NH2 (4), 4-HOPh(CH2)2CO-DTyr(Me)-Phe-Gln-Asn-Lys(5- or 6-carboxytetramethylrhodamyl)-Pro-Arg-NH2 (5, 6), 4-HOPh(CH2)2CO-DTyr(Me)-Phe-Gln-Asn-Arg-Pro-Lys(5- or 6- carboxyfluoresceinyl)-NH2 (8, 9), and 4-HOPh(CH2)2CO-DTyr(Me)-Phe-Gln-Asn-Arg-Pro-Lys(5- or 6- carboxytetramethylrhodamyl)-NH2 (10, 11). The closer to the C-terminus the fluorophore, the higher the affinities of the fluorescent derivatives for the human vasopressin V1a receptor transfected in CHO cells. The compound 10 has a Ki of 70 pM, as determined by competition experiments with [125I]-4-HOPhCH2CO-DTyr(Me)-Phe-Gln-Asn-Arg-Pro-Arg-NH2. It showed a good selectivity for human V1a receptor versus human OT (Ki = 1.2 nM), human vasopressin V1b (Ki approximately 27 nM), and human vasopressin V2 (Ki > 5000 nM) receptor subtypes. All fluorescent analogues were antagonists as shown by the inhibition of vasopressin induced inositol phosphate accumulation. These fluorescent ligands are efficient for labeling cells expressing the human V1a receptor subtype, as shown by flow cytofluorometric experiments or fluorescence microscopy. They are also appropriate tools for structural analysis of the vasopressin receptors by fluorescence.


Induced cell aggregation and mating in Streptococcus faecalis: evidence for a bacterial sex pheromone.
Recipient strains of Streptococcus faecalis produce a trypsin sensitive, heat resistant, nuclease resistant factor, designated clumping-inducing agent (CIA) which causes strains carrying certain conjugative plasmids to aggregate. RNA and protein synthesis but not DNA synthesis are required for aggregation to occur. Recipient filtrates that contain CIA activity also induce donors to mate at high frequencies. Introduction of a transferable plasmid into strains producing CIA dramatically reduces the amount of CIA activity produced by the strain but allows the strain to respond to exogenously added CIA. Our data suggest that CIA represents a bacterial sex hormone (pheromone).


Transvenous approach for the treatment of direct carotid cavernous fistula following Pipeline embolization of cavernous carotid aneurysm: a report of two cases and review of the literature.
Flow diverters are increasingly used for the endovascular treatment of cerebral aneurysms. A rare complication from flow diversion is delayed aneurysm rupture, which can lead to carotid-cavernous fistula (CCF) in the setting of cavernous carotid aneurysms (CCAs). Direct CCFs pose unique management challenges, given the lack of transarterial access to the fistula. We present two cases of direct CCFs following treatment of CCAs with the Pipeline embolization device (PED). Case 1 was a middle-aged patient with a symptomatic 10 mm wide-necked left CCA. Six weeks after PED treatment the patient developed diplopia secondary to direct CCF. Case 2 was a middle-aged patient with a symptomatic 17 mm left CCA treated with PED. One-month follow-up angiography demonstrated a direct CCF. Both patients underwent successful coil embolization of the CCF through a transvenous approach. Direct CCF formation following PED deployment for CCA is a rare complication. Parent vessel sacrifice may be avoided with transvenous occlusion of the fistula.


A compact DD neutron generator-based NAA system to quantify manganese (Mn) in bone in vivo.
A deuterium-deuterium (DD) neutron generator-based neutron activation analysis (NAA) system has been developed to quantify metals, including manganese (Mn), in bone in vivo. A DD neutron generator with a flux of up to 3*10(9) neutrons s(-1) was set up in our lab for this purpose. Optimized settings, including moderator, reflector, and shielding material and thickness, were selected based on Monte Carlo (MC) simulations conducted in our previous work. Hand phantoms doped with different Mn concentrations were irradiated using the optimized DD neutron generator irradiation system. The Mn characteristic γ-rays were collected by an HPGe detector system with 100% relative efficiency. The calibration line of the Mn/calcium (Ca) count ratio versus bone Mn concentration was obtained (R(2) = 0.99) using the hand phantoms. The detection limit (DL) was calculated to be about 1.05 μg g(-1) dry bone (ppm) with an equivalent dose of 85.4 mSv to the hand. The DL can be reduced to 0.74 ppm by using two 100% HPGe detectors. The whole body effective dose delivered to the irradiated subject was calculated to be about 17 μSv. Given the average normal bone Mn concentration of 1 ppm in the general population, this system is promising for in vivo bone Mn quantification in humans.


Age-group differences in risk perceptions of non-communicable diseases among adults in Diepsloot township, Johannesburg, South Africa: A cross-sectional study based on the Health Belief Model.
Non-communicable diseases (NCDs) in South Africa (SA) occur simultaneously with an ageing HIV-positive population, resulting in premature deaths in persons <70 years of age. Poor risk perception of NCDs results in poor adoption practices of NCD preventive measures. There is a gap in age-related research regarding risk perceptions of NCDs among the SA population. To investigate age-group differences in risk perceptions of NCDs based on the Health Belief Model. This cross-sectional design used secondary data obtained from Community AIDS Response (CARe), Johannesburg, SA. Data were collected by means of a cross-sectional survey in Extension 2 (Blocks I, J, K and L) of Diepsloot township, Johannesburg, SA. The Pearson χ2 test of independence was used to examine the relationship between age groups and risk perceptions of NCDs. A p<0.05 value was considered statistically significant. A total of 2 135 participants were included in the analysis, of whom 71.5% were young adults (18 - 35 years). The mean age of the study participants was 32.1 (standard deviation 9.87) years. Significant associations were found between age groups and risk perceptions of NCDs. More middle-aged adults than young adults and older-aged adults perceived family history (74.00% v. 72.74% v. 62.39%, p=0.045) and smoking (83.80% v. 77.20% v. 74.31%, p=0.004) as risk factors that would increase their risk of NCDs. A higher proportion of older-aged adults than young adults and middle-aged adults perceived effects on life and family (89.91% v. 77.39% v. 75.40%, p=0.004) as risks of NCD morbidities. More middle-aged adults than young adults and older-aged adults perceived the usefulness of not smoking (84.60% v. 81.06% v. 74.31%, p=0.028) as an effective NCD preventive measure. More young adults than middle-aged and older-aged adults considered health check-ups (59.31% v. 58.00% v. 41.28%, p=0.001) as a time-consuming process to prevent risks of NCDs. Young adults had poorer risk perceptions of NCDs than middle-aged and older-aged adults in Diepsloot township, resulting in poor practice of NCD preventive measures among young adults in the area. This may be due to the misunderstanding of the concept of invulnerability, possibly resulting from the limited access and exposure to NCD-related information among young adults compared with middle-aged and older-aged groups. This highlights the need to expand public health education programmes to increase outreach to the young adult population and increase accessibility to information relating to NCD risks, and encourage adoption of NCD preventive measures.


Protein expression in a transformed trabecular meshwork cell line: proteome analysis.
Characterization of the human trabecular meshwork (TM) proteome is hindered by the small mass of intact tissue and the slow growth of cultured cell strains. We have previously characterized a transformed TM cell strain (GTM3) that demonstrates many of the same protein expression and cell signaling systems of nontransformed cell strains. The aim of this study was to initiate a proteomic survey of GTM3 cells as the initial step toward characterization of the complete human TM proteome. GTM3 cells were cultured to confluence, harvested and solubilized in urea/Nonidet. The protein extract (600 mug) was focused in immobilized isoelectric focusing (IEF) strips, separated by 10% SDS PAGE, and visualized with colloidal Coomassie Blue. Spots of interest were excised, destained, and the contained proteins subjected to in-gel reduction, derivatization, and tryptic digestion. Tryptic peptides were extracted and analyzed by electrospray LC/MS/MS. Protein identification was made using the TurboSequest search algorithm and a recent version of the nonredundant human protein database downloaded from the National Center for Biotechnology Information (NCBI). Eighty-seven (87) primary proteins and 93 variants of these proteins were identified. A website was created (TM proteome) that combines data such as graphic spot location within the gel, peptide sequence, apparent and calculated pI, apparent and calculated mass, percentage of coverage, and protein informatic website links. Proteomic analysis of a transformed human TM cell line has been initiated combining preparative two-dimensional PAGE separation, LC/MS/MS analysis of major proteins, and bioinformatic cataloging of the data. Further investigation of data from the transformed cell strain will be used in a comparative fashion for spot identification of analytical proteomic gels of human TM tissue and cultured normal cells. These initial data will form the base from which the characterization of protein expression in the normal and glaucomatous TM can be accomplished.


Karymsky volcano eruptive plume properties based on MISR multi-angle imagery, and volcanological implications.
Space-based, operational instruments are in unique positions to monitor volcanic activity globally, especially in remote locations or where suborbital observing conditions are hazardous. The Multi-angle Imaging SpectroRadiometer (MISR) provides hyper-stereo imagery, from which the altitude and microphysical properties of suspended atmospheric aerosols can be derived. These capabilities are applied to plumes emitted at Karymsky volcano from 2000 to 2017. Observed plumes from Karymsky were emitted predominantly to an altitude of 2-4 km, with occasional events exceeding 6 km. MISR plume observations were most common when volcanic surface manifestations, such as lava flows, were identified by satellite-based thermal anomaly detection. The analyzed plumes predominantly contained large (1.28 µm effective radius), strongly absorbing particles indicative of ash-rich eruptions. Differences between the retrievals for Karymsky volcano's ash-rich plumes and the sulfur-rich plumes emitted during the 2014-2015 eruption of Holuhraun (Iceland) highlight the ability of MISR to distinguish particle types from such events. Observed plumes ranged from 30 to 220 km in length, and were imaged at a spatial resolution of 1.1 km. Retrieved particle properties display evidence of downwind particle fallout, particle aggregation and chemical evolution. In addition, changes in plume properties retrieved from the remote-sensing observations over time are interpreted in terms of shifts in eruption dynamics within the volcano itself, corroborated to the extent possible with suborbital data. Plumes emitted at Karymsky prior to 2010 display mixed emissions of ash and sulfate particles. After 2010, all plumes contain consistent particle components, indicative of entering an ash-dominated regime. Post-2010 event timing, relative to eruption phase, was found to influence the optical properties of observed plume particles, with light-absorption varying in a consistent sequence as each respective eruption phase progressed.


Analysis of allopolyploidy-induced rapid genetic and epigenetic changes and their relationship in wheat.
We used the conventional and methylation-sensitive randomly amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR) analyses to assess genome-wide changes and explore the relationships between genetic and epigenetic variations among individuals of a newly synthesized allohexaploid wheat line whose genomic constitution is identical to that of the natural common wheat, compared with its parent plants and a natural counterpart named Chinese Spring. We found rapid, extensive, and predominantly consistent non-Mendelian changes in the form of genetic and DNA methylation variations in the allohexaploid individuals. Specifically, at least 30-40% of the epigenetic component was truly independent of genetic changes, which answered a critical question, i.e. its autonomy in relation to the genetic context. Striking correlations were detected between genetic and epigenetic changes. Interestingly, as previously reported, the paternally donated nuclear genomes showed more genetic changes than the maternally donated ones; the loss of paternal bands was significantly correlated with the hypomethylation of CG or CHG sequences, suggesting an unknown link between genetic instability and hypomethylation. Sequence analysis indicated that most variations occurred in the cellular genes and sequences related to transposable elements. Based on these findings, the possible mechanisms and effects of the genomic changes in allopolyploid speciation and evolution were discussed.


Hypoxia in the eye: a two-sided coin.
Tissue oxygenation in general and hypoxia in particular are important regulators of retinal physiology and pathophysiology. Reduced oxygen tension and hypoxia-inducible transcription factors along with some of their target genes are critically involved in retinal development, and especially in the generation of a normal retinal vasculature. Well-timed hypoxia is thus vital for the young eye to establish proper retinal function and vision. However, when hypoxia is ill-timed, reduced oxygen tension may be associated with the development of retinal pathologies, including retinopathy of prematurity, diabetic retinopathy, glaucoma, age-related macular degeneration, or high altitude retinopathy. Here, reduced oxygen tension activates a hypoxic response that culminates in an increased expression of vascular endothelial growth factor. This causes pathological neovascularization of the delicate neuronal retina, a process that may ultimately lead to loss of vision. In contrast, preconditioning by well-defined and controlled short-term hypoxia is not devastating for the retina but instead induces a molecular response that provides protection to neuronal cells. Detailed investigation of hypoxic mechanisms during development and adulthood may thus reveal factors, which may be targeted by therapeutic approaches to save and preserve vision in patients.


The interaction of secreted phospholipase A2-IIA with the microbiota alters its lipidome and promotes inflammation.
Secreted phospholipase A2-IIA (sPLA2-IIA) hydrolyzes phospholipids to liberate lysophospholipids and fatty acids. Given its poor activity toward eukaryotic cell membranes, its role in the generation of proinflammatory lipid mediators is unclear. Conversely, sPLA2-IIA efficiently hydrolyzes bacterial membranes. Here, we show that sPLA2-IIA affects the immune system by acting on the intestinal microbial flora. Using mice overexpressing transgene-driven human sPLA2-IIA, we found that the intestinal microbiota was critical for both induction of an immune phenotype and promotion of inflammatory arthritis. The expression of sPLA2-IIA led to alterations of the intestinal microbiota composition, but housing in a more stringent pathogen-free facility revealed that its expression could affect the immune system in the absence of changes to the composition of this flora. In contrast, untargeted lipidomic analysis focusing on bacteria-derived lipid mediators revealed that sPLA2-IIA could profoundly alter the fecal lipidome. The data suggest that a singular protein, sPLA2-IIA, produces systemic effects on the immune system through its activity on the microbiota and its lipidome.


The Oxford classification of IgA nephropathy: rationale, clinicopathological correlations, and classification.
IgA nephropathy is the most common glomerular disease worldwide, yet there is no international consensus for its pathological or clinical classification. Here a new classification for IgA nephropathy is presented by an international consensus working group. The goal of this new system was to identify specific pathological features that more accurately predict risk of progression of renal disease in IgA nephropathy, thus enabling both clinicians and pathologists to improve individual patient prognostication. In a retrospective analysis, sequential clinical data were obtained on 265 adults and children with IgA nephropathy who were followed for a median of 5 years. Renal biopsies from all patients were scored by pathologists blinded to the clinical data for pathological variables identified as reproducible by an iterative process. Four of these variables: (1) the mesangial hypercellularity score, (2) segmental glomerulosclerosis, (3) endocapillary hypercellularity, and (4) tubular atrophy/interstitial fibrosis were subsequently shown to have independent value in predicting renal outcome. These specific pathological features withstood rigorous statistical analysis even after taking into account all clinical indicators available at the time of biopsy as well as during follow-up. The features have prognostic significance and we recommended they be taken into account for predicting outcome independent of the clinical features both at the time of presentation and during follow-up. The value of crescents was not addressed due to their low prevalence in the enrolled cohort.


A non-canonical, interferon-independent signaling activity of cGAMP triggers DNA damage response signaling.
Cyclic guanosine monophosphate-adenosine monophosphate (cGAMP), produced by cyclic GMP-AMP synthase (cGAS), stimulates the production of type I interferons (IFN). Here we show that cGAMP activates DNA damage response (DDR) signaling independently of its canonical IFN pathways. Loss of cGAS dampens DDR signaling induced by genotoxic insults. Mechanistically, cGAS activates DDR in a STING-TBK1-dependent manner, wherein TBK1 stimulates the autophosphorylation of the DDR kinase ATM, with the consequent activation of the CHK2-p53-p21 signal transduction pathway and the induction of G1 cell cycle arrest. Despite its stimulatory activity on ATM, cGAMP suppresses homology-directed repair (HDR) through the inhibition of polyADP-ribosylation (PARylation), in which cGAMP reduces cellular levels of NAD+; meanwhile, restoring NAD+ levels abrogates cGAMP-mediated suppression of PARylation and HDR. Finally, we show that cGAMP also activates DDR signaling in invertebrate species lacking IFN (Crassostrea virginica and Nematostella vectensis), suggesting that the genome surveillance mechanism of cGAS predates metazoan interferon-based immunity.


Preference reversal in multiattribute choice.
A central puzzle for theories of choice is that people's preferences between options can be reversed by the presence of decoy options (that are not chosen) or by the presence of other irrelevant options added to the choice set. Three types of reversal effect reported in the decision-making literature, the attraction, compromise, and similarity effects, have been explained by a number of theoretical proposals. Yet a major theoretical challenge is capturing all 3 effects simultaneously. We review the range of mechanisms that have been proposed to account for decoy effects and analyze in detail 2 computational models, decision field theory (Roe, Busemeyer, & Townsend, 2001) and leaky competing accumulators (Usher & McClelland, 2004), that aim to combine several such mechanisms into an integrated account. By simulating the models, we examine differences in the ways the decoy effects are predicted. We argue that the LCA framework, which follows on Tversky's relational evaluation with loss aversion (Tversky & Kahneman, 1991), provides a more robust account, suggesting that common mechanisms are involved in both high-level decision making and perceptual choice, for which LCA was originally developed.


Evolving mobile robots able to display collective behaviors.
We present a set of experiments in which simulated robots are evolved for the ability to aggregate and move together toward a light target. By developing and using quantitative indexes that capture the structural properties of the emerged formations, we show that evolved individuals display interesting behavioral patterns in which groups of robots act as a single unit. Moreover, evolved groups of robots with identical controllers display primitive forms of situated specialization and play different behavioral functions within the group according to the circumstances. Overall, the results presented in the article demonstrate that evolutionary techniques, by exploiting the self-organizing behavioral properties that emerge from the interactions between the robots and between the robots and the environment, are a powerful method for synthesizing collective behavior.


Correlation of serological markers and platelet count in the diagnosis of Dengue virus infection.
The dengue virus causes one of the most important mosquito-borne viral diseases. Annually, it affects up to 100 million people. Detection of the secreted NS1 protein represents a new approach to the diagnosis of acute dengue infection. Platelet count is the only non-dengue parameter that can support the diagnosis of the dengue shock syndrome (DSS) and dengue hemorrhagic fever (DHF). This study was done to correlate the platelet count and dengue parameters detected by the immunochromatographic test (ICT). Serum samples collected from patients presenting with dengue-like illness and for whom an anti-dengue antibody test was requested between August 2010 and August 2012, were included in this study. A total of 520 serum samples were collected from the suspected dengue fever patients. The samples were tested for NS1 antigen, IgM, and IgG antibodies, using the ICT kit. The platelet count was recorded in dengue parameter-positive and -negative cases. A total of 520 serum samples were collected from the suspected dengue fever patients. Sixty-two samples tested positive for one or more dengue-specific parameters. Out of the 62 samples, 39 (62.9%) were positive for the NS1 antigen, only seven (11.3%) were positive for IgM, and only three (4.9%) were positive for IgG. A platelet count < 1,00,000/ml was observed in 32 cases (51.6%). When the platelet count was done in 100 dengue parameter-negative fever patients (controls), thrombocytopenia was observed in 30% of the cases. Association of thrombocytopenia in dengue parameter-positive cases was highly significant (Z = 2.76, P = 0.006) when compared to thrombocytopenia in dengue parameter-negative patients.


Isolated loss of the AUTS2 long isoform, brain-wide or targeted to Calbindin -lineage cells, generates a specific suite of brain, behavioral and molecular pathologies.
Rearrangements within the AUTS2 region are associated with a rare syndromic disorder with intellectual disability, developmental delay and behavioral abnormalities as core features. In addition, smaller regional variants are linked to wide range of neuropsychiatric disorders, underscoring the gene's essential role in brain development. Like many essential neurodevelopmental genes, AUTS2 is large and complex, generating distinct long (AUTS2-l) and short (AUTS2-s) protein isoforms from alternative promoters. Although evidence suggests unique isoform functions, the contributions of each isoform to specific AUTS2- linked phenotypes have not been clearly resolved. Furthermore, Auts2 is widely expressed across the developing brain, but cell populations most central to disease presentation have not been determined. In this study, we focused on the specific roles of AUTS2-l in brain development, behavior, and postnatal brain gene expression, showing that brain-wide AUTS2-l ablation leads to specific subsets of the recessive pathologies associated with C-terminal mutations that disrupt both isoforms. We identify downstream genes that could explain expressed phenotypes including hundreds of putative direct AUTS2- l target genes. Furthermore, in contrast to C-terminal Auts2 mutations which lead to dominant hypoactivity, AUTS2-l loss-of-function is associated with dominant hyperactivity, a phenotype exhibited by many human patients. Finally, we show that AUTS2-l ablation in Calbindin 1 -expressing cell lineages is sufficient to yield learning/memory deficits and hyperactivity with abnormal dentate gyrus granule cell maturation, but not other phenotypic effects. These data provide new clues to in vivo AUTS2-l functions and novel information relevant to genotype-phenotype correlations in the human AUTS2 region.


Using Tibia Proximal Cut Autograft in Advanced Varus Knee Deformity in Total Knee Arthroplasty; Outcomes Compared to the Control Group.
The purpose of this study was to compare the outcomes of Total Knee Arthroplasty (TKA) performed for advanced varus knee deformity, which is performed by using tibia proximal cut bone for the reconstruction of the tibia proximal medial bone defects, with a control group consisting of TKAs which did not need reconstruction. Patients in the present study underwent total knee arthroplasty between 2009 and 2015. 12 patients with advanced varus deformity who had undergone reconstruction with tibia proximal cut autograft and 15 patients who were randomly selected from patients who did not need reconstruction, were compared clinically and radiographically. The mean follow-up period of the patients was found to be 73.1 ± 19.7 (36-108) months in the reconstruction group and 73.2 ± 12.3 (39-107) months in the control group. (p> 0.05) In both groups, significant improvement was observed postoperatively. In both groups, there was no evidence of loosening the required revision. WOMAC score was 32.4 ± 13.3 (8-64) in the reconstruction group and 28.9 ± 17.2 (6-70) in the control group at the last control visit. There was no difference between the groups when comparing the WOMAC scores at the last control visit. In the reconstruction group, the Hip-Knee-Ankle (HKA) angle was 26.1 ± 4.9 ° varus preoperatively and 1.3 ± 2.3 ° valgus postoperatively; and in the control group 10.1 ± 2.1 ° varus preoperatively and 2.7 ± 3.4 ° valgus postoperatively. (p> 0.05). In the present study, clinical and radiographic results of total knee arthroplasty patients, who suffered from advanced knee varus deformity and whose proximal tibia medial defects were reconstructed by using tibial proximal cut autograft, have been found to be successful when compared to the control group.


Epidermal Growth Factor Receptor targeting in non-small cell lung cancer: revisiting different strategies against the same target.
Epidermal Growth Factor Receptor (EGFR) tyrosine kinase inhibitors (TKIs) have changed the paradigm of treatment in non-small cell lung cancer (NSCLC). The molecular biology study of EGFR has led to clinical trials that select patients more accurately, regarding the presence of EGFR activating mutations. Nonetheless, a lack of response or a temporary condition of the response has been detected in patients on EGFR TKIs. This has urged to study potential resistance mechanisms underneath. The most important ones are the presence of secondary mutations in EGFR, such as T790M, or the overexpression of mesenchymal-epithelial transition factor (MET) that may explain why patients who initially respond to EGFR TKIs, may ultimately become refractory. Several approaches have been taken and new drugs both targeting EGFR resistance-mutation or MET are currently being developed. Here we review and update the EGFR biological pathway as well as the clinical data leading to approval of the EGFR TKIs currently in the market. New compounds under investigation targeting resistance mutations or dually targeting EGFR and other relevant receptors are also reviewed and discussed.


Comparison of Fecal Calprotectin Methods for Predicting Relapse of Pediatric Inflammatory Bowel Disease.
Background. Pediatric inflammatory bowel disease (IBD) is on the rise worldwide. Endoscopies are necessary for IBD assessment but are invasive, expensive, and inconvenient. Recently, fecal calprotectin (FCal) was proposed as a noninvasive and specific marker of gut inflammation. We evaluated the analytical performance of three FCal assays and their clinical performance in predicting relapse in pediatric IBD. Methods. This study used 40 pediatric IBD and 40 random non-IBD patients' fecal samples. Two automated ELISAs (Bühlmann and PhiCal® Calprotectin-EIA) and an EliA (Phadia 250 EliA-Calprotectin) were used to evaluate the analytical performance. The clinical performance was assessed by PhiCal Calprotectin-EIA, EliA-Calprotectin, and Bühlmann immunochromatographic point-of-care test (POCT). Results. All assays displayed acceptable analytical performance below and above the medical decision cut-off [imprecision (CV < 10% intra-assay; <15% interassay); linearity (overall mean % deviation < 16.5%)]. The agreement with PhiCal Calprotectin-EIA was 100% and 78.6% for Bühlmann (95% CI, 87.5-100; Kappa: 1) and EliA-Calprotectin (95% CI, 60.5-89.8; Kappa: 0.32), respectively, and 63.6% between Bühlmann and EliA-Calprotectin (95% CI, 46.6-77.8; Kappa: 0.16). All assays evaluated had similar clinical performance [AUC: 0.84 (EliA-Calprotectin); 0.83 (POCT and PhiCal Calprotectin-EIA)]. Conclusion. FCal levels determined using the same method and assay together with clinical history would be a noninvasive and useful tool in monitoring pediatric IBD.


Can Insertion Sequences Proliferation Influence Genomic Plasticity? Comparative Analysis of Acinetobacter baumannii Sequence Type 78, a Persistent Clone in Italian Hospitals.
Acinetobacter baumannii is a known opportunistic pathogen, dangerous for public health mostly due to its ability to rapidly acquire antibiotic-resistance traits. Its genome was described as characterized by remarkable plasticity, with a high frequency of homologous recombinations and proliferation of Insertion Sequences (IS). The SMAL pulsotype is an A. baumannii strain currently isolated only in Italy, characterized by a low incidence and a high persistence over the years. In this present work, we have conducted a comparative genomic analysis on this clone. The genome of 15 SMAL isolates was obtained and characterized in comparison with 24 other assemblies of evolutionary related isolates. The phylogeny highlighted the presence of a monophyletic clade (named ST78A), which includes the SMAL isolates. ST78A isolates have a low rate of homologous recombination and low gene content variability when compared to two related clades (ST78B and ST49) and to the most common A. baumannii variants worldwide (International Clones I and II). Remarkably, genomes in the ST78A clade present a high number of IS, including classes mostly absent in the other related genomes. Among these IS, one copy of IS66 was found to interrupt the gene comEC/rec2, involved in the acquisition of exogenous DNA. The genomic characterization of SMAL isolates shed light on the surprisingly low genomic plasticity and the high IS proliferation present in this strain. The interruption of the gene comEC/rec2 by an IS in the SMAL genomes brought us to formulate an evolutionary hypothesis according to which the proliferation of IS is slowing the acquisition of exogenous DNA, thus limiting genome plasticity. Such genomic architecture could explain the epidemiological behavior of high persistence and low incidence of the clone and provides an interesting framework to compare ST78 with the highly epidemic International Clones, characterized by high genomic plasticity.


Italian cancer figures, report 2013: Multiple tumours.
This collaborative study, based on data collected by the network of Italian association of cancer registries (AIRTUM), provides updated estimates on the incidence risk of multiple primary cancer (MP). The objective is to highlight and quantify the bidirectional associations between different oncological diseases. The quantification of the excess or decreased risk of further cancers in cancer patients, in comparison with the general population, may contribute to understand the aetiology of cancer and to address clinical follow-up. Data herein presented were provided by AIRTUM population-based cancer registries, which cover nowadays 48% of the Italian population. This monograph utilizes the AIRTUM database (December 2012), considering all malignant cancer cases diagnosed between 1976 and 2010. All cases are coded according to ICD-O-3. Non-melanoma skin cancer cases, cases based on death certificate only, cases based on autopsy only, and cases with follow-up time equal to zero were excluded. To define multiple primaries, IARC-IACR rules were adopted (http://www.iacr.com.fr/MPrules_july2004.pdf). Data were subjected to standard quality control procedures (described in the AIRTUM data management protocol) and specific quality control checks defined for the present study. A cohort of cancer patients was followed over time from first cancer diagnosis until the date of second cancer diagnosis, death, or the end of follow-up, to evaluate whether the number of observed second cancer cases was greater than expected. Person years at risk (PY) were computed by first cancer site, geographic area (North, Centre, South and Islands), attained age, and attained calendar-year group. All second cancers diagnosed in the cohort's patients were included in the observed numbers of cases. The expected number of cancer cases was computed multiplying the accumulated PY by the expected rates, calculated from the AIRTUM database stratified by cancer site, geographic area, age, and calendar-year group. The Standardized Incidence Ratio (SIR) was calculated as the ratio of observed to expected cancer cases. The Excess Absolute Risk (EAR) beyond the expected amount were calculated subtracting the expected number of subsequent cancers from the observed number of cancer cases; the difference was then divided by the PY and the number of cancer cases in excess (or deficit) was expressed per 1,000 PY. Confidence intervals were stated at 95%. The two months (60 days) after first cancer diagnosis were defined as "synchronicity period", and in the main analysis observed and expected cases during this period were excluded. It was estimated the excess risk in the period after first diagnosis (≥ 0 months), excluding the synchronicity period (≥ 2 months), and during the following periods: 2-11, 12-59, 60-119 and 120 months after diagnosis. First-cancer-site-and-gender-specific sheets are presented, reporting both SIRs and EARs. For 5,979,338 person-years a cohort of 1,635,060 cancer patients (880,361 males and 754,699 females) diagnosed between 1976 and 2010 was followed. The mean follow-up length was 14 years. Overall, 85,399 metachronous (latency ≥2 months) cancers were observed, while 77,813 were expected during the study period: SIR: 1.10 (95%CI 1.09-1.10), EAR: 1.32 x 1,000 person-years (95%CI 1.19 - 1.46). The SIR was 1.08 (95%CI 1.08-1.09) for men (54,518 observed and 50,260 expected) and 1.12 (95%CI 1.11-1.13) for women (30,881/27,553), and the EAR 1.61 (95%CI 1.37-1.84) and 1.08 x 1,000 person-years (95%CI 0.93-1.24), respectively.Moreover, during the first two months after first cancer diagnosis (synchronous period) 14,807 cancers were observed while 3,536 were expected (SIR: 4.16; 95%CI 4.09-4.22); the SIR was 4.08 (95%CI 4.00-4.16) for men and 4.32 (95%CI 4.20-4.45) for women.The mean age of patients at first cancer diagnosis was 67.0 years among males and 65.8 among females.The risk of MP was related to age being higher for younger patients and lower for older ones. In relation to the time of first cancer diagnosis, the SIR was very high at the beginning and then decreased, although remaining constantly over 1, and then rose over time. No strong differences were evident across the different incidence periods, which all showed an increased MP risk.Women had higher SIRs than expected for 18 cancer sites, men for 12. The statistically significantly SIRs lower than 1 were 2 and 8, respectively. Increased overall MP risk was observed for patients of both sexes with a first primary in the oral cavity (SIR men: 1.93; SIR women: 1.48), pharynx (SIR men: 2.13; SIR women: 1.99), larynx (SIR men: 1.57; SIR women: 1.79), oesophagus (SIR men: 1.45; SIR women: 1.41), lung (SIR men: 1.09; SIR women: 1.13), kidney (SIR men: 1.14; SIR women: 1.15), urinary bladder (SIR men: 1.29; SIR women: 1.22), thyroid (SIR: 1.22 in both sexes), Hodgkin lymphoma (SIR men: 1.59; SIR women: 1.94), and non-Hodgkin lymphoma (SIR men: 1.13; SIR women: 1.12), and for the heterogeneous group "other sites" (SIR men: 1.09; SIR women: 1.07). Moreover, men had a higher MP risk if the first cancer was in the testis (SIR: 1.24), while the same was true for women with gallbladder (SIR: 1.21), skin melanoma (SIR: 1.17), bone (SIR: 1.41), breast (SIR: 1.12), cervix uteri (SIR: 1.23) and corpus uteri (SIR: 1.23), and ovarian cancer (SIR: 1.18). On the contrary, a first liver or pancreas cancer were associated with a decreased MP risk in both sexes (liver SIR: 0.86 and 0.81 for men and women, respectively; pancreas SIR: 0.70 and 0.78 for men and women, respectively), as were those of colon (SIR: 0.93), rectum (SIR: 0.83), gallbladder (SIR: 0.80), prostate (SIR: 0.93), mesothelioma (SIR: 0.65), and central nervous system (SIR: 0.82) among men. Among the cancers for which the EAR is statistically significant, those with higher Excess Absolute Risk of MP were those of the oral cavity (EAR: 16.0 x 1,000 person-years in men and 5.4 in women), pharynx (17.6 and 9.1), larynx (11.4 and 8.8), and oesophagus (8.5 and 4.8). This descriptive study provides quantitative information on the risk of developing a second cancer in an Italian population-based cohort of approximately 1.65 million cancer patients, compared to the risk of the general population. During the follow-up time (on average 14 years) cancer patients had an MP risk that was 10% higher in comparison to the general population and an Excess Absolute Risk of 1.32 x 1,000 person-years. Study of MPs and their risk measures are dependent on methods used in the calculation. The definition of MP is not univocal and using different rules can greatly change the number of cancers in a patient with MPs. However, the AIRTUM cancer registries adopt the same recommendations for MP definition. This monograph was therefore made possible by the shared rules and standards used by AIRTUM registries. The cancer site-specific sheets, which represent the core of the monograph, can be useful to highlight and quantify the bidirectional associations among different diseases and therefore provide indications for clinical follow-up. Lifestyle changes in more healthful directions can have a positive effect in the cancer patient population and should always be recommended.


Integration of Membrane Distillation with solar photo-Fenton for purification of water contaminated with Bacillus sp. and Clostridium sp. spores.
Although Membrane Distillation (MD) has been extensively studied for desalination, it has other applications like removing all kinds of solutes from water and concentrating non-volatile substances. MD offers the possibility of producing a clean stream while concentrating valuable compounds from waste streams towards their recovery, or emerging contaminants and pathogens present in wastewater in order to facilitate their chemical elimination. This paper analyses the elimination of bacterial spores from contaminated water with MD and the role of MD in the subsequent treatment of the concentrate with photo-Fenton process. The experiments were performed at Plataforma Solar de Almería (PSA) using a plate and frame bench module with a Permeate Gap Membrane Distillation (PGMD) configuration. Tests were done for two different kinds of spores in two different water matrixes: distilled water with 3.5wt% of sea salts contaminated with spores of Bacillus subtilis (B. subtilis) and wastewater after a secondary treatment and still contaminated with Clostridium sp. spores. An analysis of the permeate was performed in all cases to determine its purity, as well as the concentrated stream and its further treatment in order to assess the benefits of using MD. Results showed a permeate free of spores in all the cases, demonstrating the viability of MD to treat biological contaminated wastewater for further use in agriculture. Moreover, the results obtained after treating the concentrate with photo-Fenton showed a shorter treatment time for the reduction of the spore concentration in the water than that when only photo-Fenton was used.


Suicide-associated comorbidity among US males and females: a multiple cause-of-death analysis.
To describe physical and mental comorbidity among male and female suicides in the US. This research replicates a seminal Australian study, which permits inference on comparative cause-of-death data quality. National cross-sectional study of suicides and unintentional injury decedents (comparison group) using Multiple Cause of Death (MCOD) public use files for 1999-2003. Prevalence of medical conditions; relative odds of suicide-associated comorbidity. For 14% of male suicides and 19% of female suicides, comorbidity was shown on their death certificates. Respective prevalences for mental and physical comorbidity were 7.2% and 7.1% for males and 10.0% and 9.3% for females. Mean numbers of psychopathologies mentioned were virtually identical for male and female suicides (1.22 and 1.21) for whom comorbidity was registered. Mean mentions of physical disease were lower for male suicides: 1.64 vs 1.77. Multiple medical conditions were registered for 4% of male suicides and 6% of female suicides. Suicides manifested excess comorbidity for depression and mood disorders (adjusted odds ratio (AOR) = 34.6, 95% CI = 32.41 to 36.92), schizophrenia (AOR = 2.5, 95% CI = 2.16 to 2.88), and cancer (AOR = 2.1, 95% CI = 1.93 to 2.19), but unexpectedly no excess comorbidity for HIV. The US system of death certification appears to be less fastidious than the Australian system. Although comorbidity patterns were very similar, only half as much psychopathology was reported for US suicides as for Australian suicides. A questionable deficit of comorbid physical disease was also documented for US suicides. Findings have important implications for medical training, as well as for suicide surveillance, policy, and prevention.


Serotonin selectively inhibits growth cone motility and synaptogenesis of specific identified neurons.
The motile activity of growth cones of specific identified neurons is inhibited by the neurotransmitter serotonin, although other identified neurons are unaffected. As a consequence, affected neurons are unable to form electrical synapses, whereas other neurons whose growth is unaffected can still interconnect. This result demonstrates that neurotransmitters can play a prominent role in regulating neuronal architecture and connectivity in addition to their classical role in neurotransmission.


Combined Face-Brain Morphology and Associated Neurocognitive Correlates in Fetal Alcohol Spectrum Disorders.
Since the 1970s, a range of facial, neurostructural, and neurocognitive adverse effects have been shown to be associated with prenatal alcohol exposure. Typically, these effects are studied individually and not in combination. Our objective is to improve the understanding of the teratogenic effects of prenatal alcohol exposure by simultaneously considering face-brain morphology and neurocognitive measures. Participants were categorized as control (n = 47), fetal alcohol syndrome (FAS, n = 22), or heavily exposed (HE) prenatally, but not eligible for a FAS diagnosis (HE, n = 50). Structural brain MRI images and high-resolution 3D facial images were analyzed using dense surface models of features of the face and surface shape of the corpus callosum (CC) and caudate nucleus (CN). Asymmetry of the CN was evaluated for correlations with neurocognitive measures. (i) Facial growth delineations for FAS, HE, and controls are replicated for the CN and the CC. (ii) Concordance of clinical diagnosis and face-based control-FAS discrimination improves when the latter is combined with specific brain regions. In particular, midline facial regions discriminate better when combined with a midsagittal profile of the CC. (iii) A subset of HE individuals was identified with FAS-like CN dysmorphism. The average of this HE subset was FAS-like in its facial dysmorphism. (iv) Right-left asymmetry found in the CNs of controls is not apparent for FAS, is diminished for HE, and correlates with neurocognitive measures in the combined FAS and HE population. Shape analysis which combines facial regions with the CN, and with the CC, better identify those with FAS. CN asymmetry was reduced for FAS compared to controls and is strongly associated with general cognitive ability, verbal learning, and recall in those with prenatal alcohol exposure. This study further extends the brain-behavior relationships known to be vulnerable to alcohol teratogenesis.


Early multisensory integration of self and source motion in the auditory system.
Discriminating external from self-produced sensory inputs is a major challenge for brains. In the auditory system, sound localization must account for movements of the head and ears, a computation likely to involve multimodal integration. Principal neurons (PNs) of the dorsal cochlear nucleus (DCN) are known to be spatially selective and to receive multimodal sensory information. We studied the responses of PNs to body rotation with or without sound stimulation, as well as to sound source rotation with stationary body. We demonstrated that PNs are sensitive to head direction, and, in the presence of sound, they differentiate between body and sound source movement. Thus, the output of the DCN provides the brain with enough information to disambiguate the movement of a sound source from an acoustically identical relative movement produced by motion of the animal.


Vulnerability as practice in diagnosing multiple conditions.
The paper contributes to contemporary understandings of vulnerability by expanding their scope with an understanding of vulnerability as generated through institutionalised practices. The argument draws on experiential accounts of navigating the practices of diagnosis by people living with multiple conditions of ill-health and disability. Vulnerability as a concept is used widely across different domains and conveys a multitude of meanings. Contemporary biomedicine, and its associated health systems and services, understands vulnerability mostly as inherent to particular physical and mental bodily conditions that put people at risk of ill-health or emotionally fragility. This may combine with a more epidemiological understanding of vulnerability as the experience of certain population groups subject to entrenched structural inequalities. Philosophers and feminists have argued that vulnerability is a universal experience of being human while political commentators have explored its potential as a resource for resistance and action. Diagnosis within medicine and psychiatry has been the subject of extensive social analysis, critique and activism. The paper draws on first-hand experiential accounts collected through face-to-face interviews with people living with multiple conditions about their experiences of diagnosis, mostly at the primary care level. We identify five aspects to diagnostic practice that are harmful and exacerbate the experience of vulnerability: temporal sequencing; diagnostic authority; medical specialisation; strategic symptom selection; medical isolation. However, these diagnostic practices are not best understood only in terms of the power asymmetries inherent to the medical consultation, but are embedded into the very institution of diagnosis. The paper thus proposes a combined approach to vulnerability that recognises it as a universal condition of humanity but one that becomes animated or amplified for some bodies, through their own inherent incapacities or the external structures of inequality, and through the practices of medicine as situated in particular times and places.


Circulating Tumor Cell Phenotyping via High-Throughput Acoustic Separation.
The study of circulating tumor cells (CTCs) offers pathways to develop new diagnostic and prognostic biomarkers that benefit cancer treatments. In order to fully exploit and interpret the information provided by CTCs, the development of a platform is reported that integrates acoustics and microfluidics to isolate rare CTCs from peripheral blood in high throughput while preserving their structural, biological, and functional integrity. Cancer cells are first isolated from leukocytes with a throughput of 7.5 mL h-1 , achieving a recovery rate of at least 86% while maintaining the cells' ability to proliferate. High-throughput acoustic separation enables statistical analysis of isolated CTCs from prostate cancer patients to be performed to determine their size distribution and phenotypic heterogeneity for a range of biomarkers, including the visualization of CTCs with a loss of expression for the prostate specific membrane antigen. The method also enables the isolation of even rarer, but clinically important, CTC clusters.


The critical-care nurse practitioner curriculum at the University of Pennsylvania: update and revision.
This paper describes the philosophical, conceptual, and programmatic characteristics of the Adult Critical Care Nurse Practitioner (CCNP) Program, which began at the University of Pennsylvania in 1993. It outlines the rationale and strategies for curriculum revisions during the 3rd year of program implementation. The basic content and sequencing of courses in the program did not change. Curricular revision in theoretical and field work courses consisted of modifying the approach to teaching with emphasis on critical thinking, diagnostic reasoning, and clinical decision making. Finally, it discusses the future challenges for educating CCNPs, with an emphasis on market demands and research challenges.


Neural substrates of the development of cognitive control in children ages 5-10 years.
Cognitive conflict detection and resolution develops with age across childhood and likely supports age-related increases in other aspects of cognitive and emotional development. Little is known about the neural correlates of conflict detection and resolution in early childhood. In the current study, we investigated age-related change in neural recruitment during a blocked spatial-incompatibility task (Simon task) in children ages 5-10 years using fMRI. Cortical thickness was measured using structural MRI. Across all children, there was greater activation in right prefrontal and bilateral parietal cortices for incompatible than compatible conditions. In older children, compared with younger children, there was decreased activation and decreased gray matter thickness in the medial PFC. Thickness and activation changes across age were associated within participants, such that thinner cortex was associated with less activation in the rostral ACC. These findings suggest that developmental change in medial PFC activation supports performance on cognitive control tasks in early childhood.


The pharmacokinetics and the pharmacodynamics of cannabinoids.
There is increasing interest in the use of cannabinoids for disease and symptom management, but limited information available regarding their pharmacokinetics and pharmacodynamics to guide prescribers. Cannabis medicines contain a wide variety of chemical compounds, including the cannabinoids delta-9-tetrahydrocannabinol (THC), which is psychoactive, and the nonpsychoactive cannabidiol (CBD). Cannabis use is associated with both pathological and behavioural toxicity and, accordingly, is contraindicated in the context of significant psychiatric, cardiovascular, renal or hepatic illness. The pharmacokinetics of cannabinoids and the effects observed depend on the formulation and route of administration, which should be tailored to individual patient requirements. As both THC and CBD are hepatically metabolized, the potential exists for pharmacokinetic drug interactions via inhibition or induction of enzymes or transporters. An important example is the CBD-mediated inhibition of clobazam metabolism. Pharmacodynamic interactions may occur if cannabis is administered with other central nervous system depressant drugs, and cardiac toxicity may occur via additive hypertension and tachycardia with sympathomimetic agents. More vulnerable populations, such as older patients, may benefit from the potential symptomatic and palliative benefits of cannabinoids but are at increased risk of adverse effects. The limited availability of applicable pharmacokinetic and pharmacodynamic information highlights the need to initiate prescribing cannabis medicines using a 'start low and go slow' approach, carefully observing the patient for desired and adverse effects. Further clinical studies in the actual patient populations for whom prescribing may be considered are needed, to derive a better understanding of these drugs and enhance safe and optimal prescribing.


Epidemiology and outcomes of clostridium difficile-associated disease among patients on prolonged acute mechanical ventilation.
Patients receiving prolonged acute mechanical ventilation (PAMV), although comprising a third of all mechanical ventilation (MV) patients, consume two-thirds of all the resources allocated to MV, and their numbers are projected to double by 2020. By virtue of their prolonged hospital length of stay (median LOS, 17 days), they are subject to such nosocomial infections as Clostridium difficile-associated disease (CDAD), the incidence and age-adjusted case fatality rate of which doubled between 2000 and 2005. We examined the rates and outcomes of CDAD among adult PAMV patients. We analyzed 2005 data from the Health Care Utilization Project/Nationwide Inpatient Sample from the Agency for Healthcare Research and Quality. PAMV and CDAD were identified using the International Classification of Diseases, ninth revision, clinical modification codes 96.72 and 008.45, respectively. Among 64,910 adult PAMV patients who were discharged in 2005, 3,468 patients (5.34%) had a concurrent diagnosis of CDAD (PAMV patients who were discharged with concomitant diagnosis of CDAD [CDAD+]). CDAD+ patients who were discharged were older (mean [+/- SD] age, 66.7 +/- 15.9 vs 63.7 +/- 16.9 years, respectively; p < 0.001) and were more likely to have been admitted to the hospital from a long-term care facility (5.7% vs 2.9%, respectively; p < 0.001) than PAMV patients who were discharged without CDAD (CDAD-). Although crude hospital mortality rates did not differ among PAMV patients who were discharged from the hospital by CDAD status (CDAD+, 32.6%; CDAD-, 33.0%; p = 0.598), both unadjusted calculations and propensity-score adjustment showed a substantial increase in LOS (6.1 days; 95% confidence interval [CI], 4.9 to 7.4) and total costs ($10,355; 95% CI, $7,540 to $13,170) among CDAD+ patients. PAMV patients have an order of magnitude higher risk of having CDAD than other hospitalized patients. Concurrent CDAD infection is associated with increased hospital LOS and costs. The PAMV population is an attractive target for aggressive measures aimed at CDAD prevention.


Effect of Helicobacter pylori Infection on GATA-5 and TFF1 Regulation, Comparison Between Pediatric and Adult Patients.
GATA factors, which constitute a family of transcription regulatory proteins, participate in gastrointestinal development. Trefoil factor 1 (TFF1) plays a crucial role in mucosal defense and healing, and evidence suggests that GATA-5 mediated its regulation. Gastric cancer is a multiple-step process triggered by Helicobacter pylori and is characterized by accumulation of molecular and epigenetic alteration. The aim of this study was to evaluate the effect of H. pylori infection on the regulation of GATA-5 and TFF1 in vitro and in vivo. Infected cells exhibited upregulation of GATA-5 and TFF1 after 48 h. An increase in GATA-5 and TFF1 mRNA levels was also found in mice samples after 6 and 12 months of infection, respectively. In human samples, we found an association between H. pylori infection and GATA-5 upregulation. In fact, among H. pylori-infected patients, hypermethylation was observed in 45.5% of pediatric samples, in 62.6% of chronic gastritis samples, and in 63% of gastric cancer samples. Regarding TFF1, the expression levels were similar in pediatrics and adults patients, and were independent of H. pylori infection, and the expression of these factors was downregulated in gastric cancer samples. GATA-5 promoter methylation was associated with a decrease in TFF1 mRNA levels. Our results suggest that the upregulation of GATA-5 and TFF1 observed in vitro and in vivo may be correlated with a protective effect of the mucosa in response to infection. The epigenetic inactivation of GATA-5 observed in human biopsies from infected patients may suggest that this alteration is an early event occurring in association with H. pylori infection.


Real-time monitoring of neural differentiation of human mesenchymal stem cells by electric cell-substrate impedance sensing.
Stem cells are useful for cell replacement therapy. Stem cell differentiation must be monitored thoroughly and precisely prior to transplantation. In this study we evaluated the usefulness of electric cell-substrate impedance sensing (ECIS) for in vitro real-time monitoring of neural differentiation of human mesenchymal stem cells (hMSCs). We cultured hMSCs in neural differentiation media (NDM) for 6 days and examined the time-course of impedance changes with an ECIS array. We also monitored the expression of markers for neural differentiation, total cell count, and cell cycle profiles. Cellular expression of neuron and oligodendrocyte markers increased. The resistance value of cells cultured in NDM was automatically measured in real-time and found to increase much more slowly over time compared to cells cultured in non-differentiation media. The relatively slow resistance changes observed in differentiating MSCs were determined to be due to their lower growth capacity achieved by induction of cell cycle arrest in G0/G1. Overall results suggest that the relatively slow change in resistance values measured by ECIS method can be used as a parameter for slowly growing neural-differentiating cells. However, to enhance the competence of ECIS for in vitro real-time monitoring of neural differentiation of MSCs, more elaborate studies are needed.


Improved coalescence stability of monodisperse phospholipid-coated microbubbles formed by flow-focusing at elevated temperatures.
Monodisperse phospholipid-coated ultrasound contrast agent (UCA) microbubbles can be directly synthesized in a lab-on-a-chip flow-focusing device. However, high total lipid concentrations are required to minimize on-chip bubble coalescence. Here, we characterize the coalescence probability and the long-term size stability of microbubbles formed using DPPC and DSPC based lipid mixtures as a function of temperature. We show that the coalescence probability can be dramatically reduced by increasing the temperature during bubble formation. Moreover, it is shown that the increased coalescence stability can be explained from an exponential increase of the relative viscosity in the thin liquid film between the colliding bubbles. Furthermore, it was found that the relative viscosity of a DPPC lipid mixture is 7.6 times higher than that of a DSPC mixture and that it can be explained solely from the higher DPPC liposome concentration. Regarding long-term bubble stability, the ratio of the initial on-chip bubble size to the final stable bubble size was always found to be 2.2 for DPPC and DSPC coated bubbles with 10 mol% DPPE-PEG5000, independent of the temperature. Moreover, it was demonstrated that the microbubble suspensions formed at elevated temperatures are highly stable over a time window of 2 to 4 days when collected in a vial. All in all, this work shows that, by increasing the temperature during bubble formation from room temperature to 70 °C, the efficiency of the use of phospholipids in microbubble formation by flow-focusing can be increased by 5 times.


Best behaviour? Ontologies and the formal description of animal behaviour.
The development of ontologies for describing animal behaviour has proved to be one of the most difficult of all scientific knowledge domains. Ranging from neurological processes to human emotions, the range and scope needed for such ontologies is highly challenging, but if data integration and computational tools such as automated reasoning are to be fully applied in this important area the underlying principles of these ontologies need to be better established and development needs detailed coordination. Whilst the state of scientific knowledge is always paramount in ontology and formal description framework design, this is a particular problem with neurobehavioural ontologies where our understanding of the relationship between behaviour and its underlying biophysical basis is currently in its infancy. In this commentary, we discuss some of the fundamental problems in designing and using behaviour ontologies, and present some of the best developed tools in this domain.