Luteolin induces hippocampal neurogenesis in the Ts65Dn mouse model of Down syndrome.
Studies have shown that the natural flavonoid luteolin has neurotrophic activity. In this study, we investigated the effect of luteolin in a mouse model of Down syndrome. Ts65Dn mice, which are frequently used as a model of Down syndrome, were intraperitoneally injected with 10 mg/kg luteolin for 4 consecutive weeks starting at 12 weeks of age. The Morris water maze test was used to evaluate learning and memory abilities, and the novel object recognition test was used to assess recognition memory. Immunohistochemistry was performed for the neural stem cell marker nestin, the astrocyte marker glial fibrillary acidic protein, the immature neuron marker DCX, the mature neuron marker NeuN, and the cell proliferation marker Ki67 in the hippocampal dentate gyrus. Nissl staining was used to observe changes in morphology and to quantify cells in the dentate gyrus. Western blot assay was used to analyze the protein levels of brain-derived neurotrophic factor (BDNF) and phospho-extracellular signal-regulated kinase 1/2 (p-ERK1/2) in the hippocampus. Luteolin improved learning and memory abilities as well as novel object recognition ability, and enhanced the proliferation of neurons in the hippocampal dentate gyrus. Furthermore, luteolin increased expression of nestin and glial fibrillary acidic protein, increased the number of DCX+ neurons in the granular layer and NeuN+ neurons in the subgranular region of the dentate gyrus, and increased the protein levels of BDNF and p-ERK1/2 in the hippocampus. Our findings show that luteolin improves behavioral performance and promotes hippocampal neurogenesis in Ts65Dn mice. Moreover, these effects might be associated with the activation of the BDNF/ERK1/2 pathway.


The drug target genes show higher evolutionary conservation than non-target genes.
Although evidence indicates that drug target genes share some common evolutionary features, there have been few studies analyzing evolutionary features of drug targets from an overall level. Therefore, we conducted an analysis which aimed to investigate the evolutionary characteristics of drug target genes. We compared the evolutionary conservation between human drug target genes and non-target genes by combining both the evolutionary features and network topological properties in human protein-protein interaction network. The evolution rate, conservation score and the percentage of orthologous genes of 21 species were included in our study. Meanwhile, four topological features including the average shortest path length, betweenness centrality, clustering coefficient and degree were considered for comparison analysis. Then we got four results as following: compared with non-drug target genes, 1) drug target genes had lower evolutionary rates; 2) drug target genes had higher conservation scores; 3) drug target genes had higher percentages of orthologous genes and 4) drug target genes had a tighter network structure including higher degrees, betweenness centrality, clustering coefficients and lower average shortest path lengths. These results demonstrate that drug target genes are more evolutionarily conserved than non-drug target genes. We hope that our study will provide valuable information for other researchers who are interested in evolutionary conservation of drug targets.


The peripheral sensory nervous system in the vertebrate head: a gene regulatory perspective.
In the vertebrate head, crucial parts of the sense organs and sensory ganglia develop from special regions, the cranial placodes. Despite their cellular and functional diversity, they arise from a common field of multipotent progenitors and acquire distinct identity later under the influence of local signalling. Here we present the gene regulatory network that summarises our current understanding of how sensory cells are specified, how they become different from other ectodermal derivatives and how they begin to diversify to generate placodes with different identities. This analysis reveals how sequential activation of sets of transcription factors subdivides the ectoderm over time into smaller domains of progenitors for the central nervous system, neural crest, epidermis and sensory placodes. Within this hierarchy the timing of signalling and developmental history of each cell population is of critical importance to determine the ultimate outcome. A reoccurring theme is that local signals set up broad gene expression domains, which are further refined by mutual repression between different transcription factors. The Six and Eya network lies at the heart of sensory progenitor specification. In a positive feedback loop these factors perpetuate their own expression thus stabilising pre-placodal fate, while simultaneously repressing neural and neural crest specific factors. Downstream of the Six and Eya cassette, Pax genes in combination with other factors begin to impart regional identity to placode progenitors. While our review highlights the wealth of information available, it also points to the lack information on the cis-regulatory mechanisms that control placode specification and of how the repeated use of signalling input is integrated.


Psychiatric aspects of phosphodiesterases: An overview.
Phosphodiesterases (PDE) are exciting new targets in medical sciences. These enzymes are some of the key mediators of cellular functions in the body and hence are attractive sites for drug-induced modulations. With the finding that Tofisopam, a new anxiolytic, inhibits PDEs, the authors were inspired to look into the role of PDE and drugs acting on them in psychiatry. Hence, the review was undertaken. We found several research materials available highlighting the role of PDE in cellular functions and the possible newer etiological mechanisms of neuropsychiatric illnesses such as schizophrenia, depression/anxiety disorders, and cognitive dysfunction involving PDEs. We also found that there are many molecules acting on PDEs, which have the potential to alter the way we treat mental illnesses today. This article is intended to provide an in-depth look at these enzymes so that more cost-effective therapeutic molecules may be synthesized and marketed in India for managing mental illnesses.


The phenotype of spontaneous preterm birth: application of a clinical phenotyping tool.
Spontaneous preterm birth (SPTB) is a complex condition that is likely a final common pathway with multiple possible causes. We hypothesized that a comprehensive classification system appropriately could group women with similar STPB causes and could provide an explanation, at least in part, for the disparities in SPTB that are associated with race and gestational age at delivery. This was a planned analysis of a multicenter, prospective study of singleton SPTBs. Women with SPTB at <34 weeks' gestation were included. We defined 9 potential SPTB phenotypes based on clinical data: infection/inflammation, maternal stress, decidual hemorrhage, uterine distention, cervical insufficiency, placental dysfunction, premature rupture of the membranes, maternal comorbidities, and familial factors. Each woman's condition was evaluated for each phenotype. Delivery gestational age was compared between those with and without each phenotype. Phenotype profiles were also compared between women with very early (20.0-27.9 weeks' gestation) SPTB vs those with early SPTB (28.0-34.0 weeks' gestation) and between African American and white women. Statistical analysis was by t test and χ(2) test, as appropriate. The phenotyping tool was applied to 1025 women with SPTBs who delivered at a mean 30.0 ± 3.2 (SD) weeks' gestation. Of these, 800 women (78%) had ≥2 phenotypes. Only 43 women (4.2%) had no phenotypes. The 281 women with early SPTBs were more likely to have infection/inflammation, decidual hemorrhage, and cervical insufficiency phenotypes (all P ≤ .001). African American women had more maternal stress and cervical insufficiency but less decidual hemorrhage and placental dysfunction compared with white women (all P < .05). Gestational age at delivery decreased as the number of phenotypes that were present increased. Precise SPTB phenotyping classifies women with SPTBs and identifies specific differences between very early and early SPTB and between African American and white women.


Injury Risk and Noise Exposure in Firefighter Training Operations.
Firefighters have high rate of injuries and illnesses, as well as exposures to high levels of noise. This study explored the relationship between noise exposure and injury among firefighters. We recruited firefighters undergoing vehicle extrication and structural collapse emergency response training at a highly realistic training facility. Demographics, health status, body mass index (BMI), and history of serious injuries (i.e. injuries requiring first aid treatment, treatment in a medical clinic or office, or treatment at a hospital) were assessed at baseline, and daily activities, injury events, and near misses were assessed daily via surveys. Participants' noise exposures were monitored for one 24-h period using noise dosimeters. We used a mixed-effects logistic regression model to estimate the odds of injury events and near misses associated with noise exposure as an independent variable. Of 56 subjects, 20 (36%) reported that they had ever suffered a serious injury during firefighting activities, and 9 (16%) reported a serious injury within the past year. We estimated rates of 6.6 lifetime serious injuries per 100 FTE 16.1 serious injuries per 100 FTE within the past year. Our models indicated a significant increase in injury events and near misses among those with higher BMI, and as well as a dose-response relationship between near misses/injuries and increasing noise levels. Noise levels >90 dBA in the 30 min prior to time of injury or near miss were associated with substantially increased odds ratios for injury or near miss. Our models further indicated that perceived job demands were significantly associated with increased risk of injury or near miss. Our results suggest that noise exposures may need to be incorporated into injury prevention programs for firefighters to reduce injuries among this high-risk occupational group.


[Popular medicine: benefits and drawbacks of medicinal plants].
Descriptive study which aims to verify if the herbalist offer the information correct for the use of the medicinal plants; if they give for the customers concerning the possible poisonings or interaction with the allopathic guide and also if there is a criteria for the commercialization of the phytotherapics. A questionnaire was used containing open and closed questions involving aspects of the performance of the herbalists with the phytotherapics. The herbalists know the majority of the medicinal plants, however, there are some gaps concerning the correct indication of these products, of the collateral effect and toxicities. The herbalist lack to better know the principles of the grass, the therapeutically indications active, to guide the users concerning the possible pharmacological interactions or medicaments poisonings and regarding the cleanness, storage, time of useful life and contraindications of the product.


Functionally Conserved Noncoding Regulators of Cardiomyocyte Proliferation and Regeneration in Mouse and Human.
The adult mammalian heart has little regenerative capacity after myocardial infarction (MI), whereas neonatal mouse heart regenerates without scarring or dysfunction. However, the underlying pathways are poorly defined. We sought to derive insights into the pathways regulating neonatal development of the mouse heart and cardiac regeneration post-MI. Total RNA-seq of mouse heart through the first 10 days of postnatal life (referred to as P3, P5, P10) revealed a previously unobserved transition in microRNA (miRNA) expression between P3 and P5 associated specifically with altered expression of protein-coding genes on the focal adhesion pathway and cessation of cardiomyocyte cell division. We found profound changes in the coding and noncoding transcriptome after neonatal MI, with evidence of essentially complete healing by P10. Over two-thirds of each of the messenger RNAs, long noncoding RNAs, and miRNAs that were differentially expressed in the post-MI heart were differentially expressed during normal postnatal development, suggesting a common regulatory pathway for normal cardiac development and post-MI cardiac regeneration. We selected exemplars of miRNAs implicated in our data set as regulators of cardiomyocyte proliferation. Several of these showed evidence of a functional influence on mouse cardiomyocyte cell division. In addition, a subset of these miRNAs, miR-144-3p, miR-195a-5p, miR-451a, and miR-6240 showed evidence of functional conservation in human cardiomyocytes. The sets of messenger RNAs, miRNAs, and long noncoding RNAs that we report here merit further investigation as gatekeepers of cell division in the postnatal heart and as targets for extension of the period of cardiac regeneration beyond the neonatal period.


Androgen Suppresses Hyperosmolarity-Induced Inflammatory Mediators in Human Corneal Epithelial Cells.
To investigative the effects of sex steroids on hyperosmolar stress-induced proinflammatory cytokine expression of tumor necrosis factor-alpha (TNF-α), interleukin (IL)-8, and IL-6, and on the mitogen-activated protein kinase pathway in immortalized human corneal epithelial cells (hCECs). Immortalized hCECs were cultured with keratinocyte-free medium until reaching 80% confluency with either 10 M dihidrotestosteron (DHT) or 10 M 17-β-estradiol, and then, the medium was changed to hyperosmolar for various time points. After hyperosmolar treatment, a real-time polymerase chain reaction was performed to show the TNF-α, IL-8, and IL-6 gene expression levels in hCECs. In addition, the treated cells were lysed, and Western blot analysis was applied for phosphorylated and nonphosphorylated forms of extracellular signal-regulated kinase 1/2 (ERK1/2), p38 kinase, and c-Jun N-terminal kinase 1/2 (JNK1/2). hCECs viability was measured with Annexin V/propidium iodide. Pretreatment with 10 M DHT or 17-β-estradiol inhibited the high osmolarity-induced expression of TNF-α, IL-8, and IL-6. The upregulation of p-ERK, p-JNK, and p-p38 with high osmolarity was inhibited partially by DHT, but 17-β-estradiol pretreatment only affected p-p38 for a short time interval. In addition, DHT increased cell viability of hCECs under hyperosmolar conditions. Our results demonstrated that DHT and 17-β-estradiol decreased the proinflammatory cytokine gene expression levels which were stimulated by high osmolarity in immortalized hCECs. The mitogen-activated protein kinase signaling pathway is partially involved in the regulatory effects of DHT on hCECs. These findings may contribute to the etiologic role and therapeutic implications of sex steroids in certain ocular surface diseases.


Resolvin E1 reduces proinflammatory markers in human pancreatic islets in vitro.
In clinical islet transplantation, inflammatory responses initiated by the transplanted islets and by the host immune system cause acute and chronic graft loss. The resolution of acute inflammation is an active process mediated by specific signals and mediators such as resolvin E1 (RvE1). We investigated the effect of RvE1 on i) the inflammatory status of human pancreatic islets, ii) islet viability and apoptosis, and iii) the instant blood-mediated inflammatory reaction (IBMIR) IN VITRO. Pro-inflammatory cytokines and tissue factor (TF) in isolated human islets were determined by real-time RT-qPCR (mRNA levels), CBA and Gyrolab bioaffy (protein levels) after lipopolysaccaride (LPS) stimulation. Islet viability was measured using insulin secretion in a dynamic model, ADP/ATP ratio and total ATP content. Apoptosis was measured using commercial kits after stimulation with proinflammatory cytokines. To assess effect on IBMIR, human islets were mixed with non-anticoagulated, RvE1 or vehicle pretreated ABO-compatible blood in heparin-coated tubing loops. Treatment of human islets with RvE1 (500 nM) for 24 h reduced LPS-induced increase in mRNA and protein levels of selected pro-inflammatory markers (IL-8, MCP-1, and TF). RvE1 lowered the ADP/ATP ratio, but had no effect on insulin secretion. RvE1 reduced the apoptotic effect of proinflammatory cytokines. Additionally, RvE1 reduced platelet consumption and TAT complex formation during the first 5 min after islet-blood contact. RvE1 suppresses proinflammatory markers and lowers the ADP/ATP ratio in human islets IN VITRO. RvE1 demonstrates anti-apoptotic effects in a proinflammatory milieu. Additionally, RvE1 has modest dampening effects on IBMIR. We conclude that RvE1 may have potential in clinical islet transplantation.


Family Presence at the PICU Bedside: A Single-Center Retrospective Cohort Study.
To determine factors associated with bedside family presence in the PICU and to understand how individual factors interact as barriers to family presence. Mixed methods study. Tertiary children's hospital PICU. Five hundred twenty-three children of less than 18 years enrolled in the Seattle Children's Hospital Outcomes Assessment Program from 2011 to 2017. None. Quantitative: Family was documented every 2 hours. Exposures included patient and illness characteristics and family demographic and socioeconomic characteristics. We used multivariable logistic regression to identify factors associated with presence of less than 80% and stratified results by self-reported race. Longer PICU length of stay (LOS), public insurance, and complex chronic conditions (C-CD) were associated with family presence of less than 80%. Self-reported race modified these associations; no factors were associated with lower bedside presence for White families, in contrast with multiple associations for non-White families including public insurance, C-CD, and longer LOS. Qualitative: Thematic analysis of social work notes for the 48 patients with family presence of less than 80% matched on age, LOS, and diagnosis to 48 patients with greater than or equal to 95% family presence. Three themes emerged: the primary caregiver's prior experiences with the hospital, relationships outside of the hospital, and additional stressors during the hospitalization affected bedside presence. We identified sociodemographic and illness factors associated with family bedside presence in the PICU. Self-reported race modified these associations, representing racism within healthcare. Family presence at the bedside may help identify families facing greater disparities in healthcare access.


George Chance and Frank Holdsworth: Understanding Spinal Instability and the Evolution of Modern Spine Injury Classification Systems.
The concept of spinal cord injury has existed since the earliest human civilizations, with the earliest documented cases dating back to 3000 BC under the Egyptian Empire. Howevr, an understanding of this field developed slowly, with real advancements not emerging until the 20th century. Technological advancements including the dawn of modern warfare producing mass human casualties instigated revolutionary advancement in the field of spine injury and its management. Spine surgeons today encounter "Chance" and "Holdsworth" fractures commonly; however, neurosurgical literature has not explored the history of these physicians and their groundbreaking contributions to the modern understanding of spine injury. A literature search using a historical database, Cochrane, Google Scholar, and PubMed was performed. As needed, hospitals and native universities were contacted to add their original contributions to the literature. George Quentin Chance, a Manchester-based British physician, is well known to many as an eminent radiologist of his time who described the eponymous fracture in 1948. Sir Frank Wild Holdsworth (1904-1969), a renowned British orthopedic surgeon who laid a solid foundation for rehabilitation of spinal injuries under the aegis of the Miners' Welfare Commission, described in detail the management of thoraco-lumbar junctional rotational fracture. The work of these 2 men laid the foundation for today's understanding of spinal instability, which is central to modern spine injury classification and management algorithms. This historical vignette will explore the academic legacies of Sir Frank Wild Holdsworth and George Quentin Chance, and the evolution of spinal instability and spine injury classification systems that ensued from their work.


Glucose metabolism measured by positron emission tomography is reduced in patients with white matter presumably ischemic lesions.
The severity and progression of white matter ischemic lesion (WMIL) are closely linked to vascular dementia. The function of neural tissue is closely linked to glucose consumption as the most important energy-supplying metabolic process. At present, [18]fluorine-fluorodeoxy glucose ([18]FDG) positron emission tomography (PET) can provide regional and 3-dimensional quantification of glucose metabolism in the human brain. Although MMSE and MoCA are commonly used screens in cognitive impairment, no research team has yet validated their performance in WMIL. The purpose of our study was to compare MMSE and MoCA in screening for cognitive impairment and to explore the correlations between CMRglu values and executive function. All the participants underwent comprehensive clinical, MoCA, MMSE, MRI, and PET examinations. Patients in the WMIL group were subdivided into 3 severity subgroups according to the Fazekas scale. The MoCA scores were lower in the WMIL group. Our research indicates that MoCA is a more sensitive screening tool than the commonly used MMSE in detecting cognitive impairment in patients with WMIL. CMRglu values of gray matter were decreased in the WMIL group. Reductions of CMRglu in parietal lobe, frontal lobe, and white matter centrum semiovale were observed to different degrees in the WMIL groups according to the modified Fazekas scale. A significant negative correlation was found between executive function and CMRglu in the frontal lobe. MoCA appears to be a more sensitive screening tool than the commonly used MMSE in detecting cognitive impairment in patients with WMIL. CMRglu can potentially be used as a biomarker for predicting the severity of WMIL.


Phosphorylation of insulin receptors solubilized from rat skeletal muscle.
A method has been developed to solubilize insulin receptors from skeletal muscles. Rat hindlimb muscles were rapidly frozen in liquid nitrogen, powdered, extracted with buffered Triton X-100, and partially purified by differential centrifugation followed by wheat germ agglutinin affinity chromatography. The solubilized receptors exhibit typical curvilinear Scatchard plots in insulin binding assays: rapid, Mn2+-dependent autophosphorylation of the beta-subunit on exposure to insulin as well as insulin-stimulated kinase activity toward histone H2B. Furthermore, when intact soleus muscles were incubated in phosphate-depleted medium containing Na2H[32P]PO4, addition of insulin stimulated the in situ phosphorylation of the beta-subunit of the insulin receptor. The ability to rapidly and efficiently isolate insulin receptors from skeletal muscle may permit investigation of factors that modulate insulin action in this tissue.


Vectors for P element-mediated gene transfer in Drosophila.
We have constructed and tested several new vectors for P element-mediated gene transfer. These vectors contain restriction sites for cloning a wide variety of DNA fragments within a small, non-autonomous P element and can be used to efficiently transduce microinjected DNA sequences into the germ line chromosomes of D. melanogaster. The P element in one vector also carries the rosy gene which serves as an easily scored marker to facilitate the transfer of DNA fragments that do not themselves confer a recognizable phenotype. The failure of certain P element constructs to function as vectors suggests that P element sequences, in addition to the 31 bp inverse terminal repeats, are required in cis for transposition. Moreover, removal of the first 38 bp of the autonomous 2.9 kb P element appears to destroy its ability to provide a trans-acting factor (s) required for the transposition of non-autonomous P elements. Finally, we describe a genomic sequence arrangement that apparently arose by the transposition of a 54 kb composite P element from a tetramer plasmid.


Submillisecond rotational dynamics of spin-labeled myosin heads in myofibrils.
The rotational motion of crossbridges, formed when myosin heads bind to actin, is an essential element of most molecular models of muscle contraction. To obtain direct information about this molecular motion, we have performed saturation transfer EPR experiments in which spin labels were selectively and rigidly attached to myosin heads in purified myosin and in glycerinated myofibrils. In synthetic myosin filaments, in the absence of actin, the spectra indicated rapid rotational motion of heads characterized by an effective correlation time of 10 microseconds. By contrast, little or no submillisecond rotational motion was observed when isolated myosin heads (subfragment-1) were attached to glass beads or to F-actin, indicating that the bond between the myosin head and actin is quite rigid on this time scale. A similar immobilization of heads was observed in spin-labeled myofibrils in rigor. Therefore, we conclude that virtually all of the myosin heads in a rigor myofibril are immobilized, apparently owing to attachment of heads to actin. Addition of ATP to myofibrils, either in the presence or absence of 0.1 mM Ca2+, produced spectra similar to those observed for myosin filaments in the absence of actin, indicating rapid submillisecond rotational motion. These results indicate that either (a) most of the myosin heads are detached at any instant in relaxed or activated myofibrils or (b) attached heads bearing the products of ATP hydrolysis rotate as rapidly as detached heads.


Modulation of macrophage phenotype and protein secretion via heparin-IL-4 functionalized supramolecular elastomers.
Hallmark of the in situ tissue engineering approach is the use of bioresorbable, synthetic, acellular scaffolds, which are designed to modulate the inflammatory response and actively trigger tissue regeneration by the body itself at the site of implantation. Much research is devoted to the design of synthetic materials modulating the polarization of macrophages, which are essential mediators of the early stages of the inflammatory response. Here, we present a novel method for the functionalization of elastomers based on synthetic peptide chemistry, supramolecular self-assembly, and immobilization of heparin and interleukin 4 (IL-4), which is known to skew the polarization of macrophages into the wound healing "M2" phenotype. Ureido-pyrimidinone (UPy)-modified chain extended polycaprolactone (CE-UPy-PCL) was mixed with a UPy-modified heparin binding peptide (UPy-HBP) to allow for immobilization of heparin, and further functionalization with IL-4 via its heparin binding domain. As a first proof of principle, CE-UPy-PCL and UPy-HBP were premixed in solution, dropcast and exposed to primary human monocyte-derived macrophages, in the presence or absence of IL-4-heparin functionalization. It was demonstrated that the supramolecular IL-4-heparin functionalization effectively promoted macrophage polarization into an anti-inflammatory phenotype, in terms of morphology, immunohistochemistry and cytokine secretion. Moreover, the supramolecular functionalization approach used was successfully translated to 3D electrospun scaffolds for in situ tissue engineering purposes, where UPy-HBP retention, and heparin and IL-4 attachment to the supramolecular scaffolds were proven over 7 days. Lastly, human monocyte-derived macrophages were cultured on 3D scaffolds, which, in case of IL-4-heparin functionalization, were proven to promote of an anti-inflammatory environment on protein level. This study presents a novel method in designing a versatile class of functionalized elastomers that effectively harness the anti-inflammatory behavior of macrophages in vitro, and as such, may be instrumental for the development of a new class of synthetic materials for in situ tissue engineering purposes. Macrophages and their phenotypic and functional plasticity play a pivotal role in metabolic homeostasis and tissue repair. Based on this notion, bioactivated materials modulating macrophage polarization were extensively investigated in the past. Here, we designed immunomodulating, synthetic materials based on supramolecular immobilization of a heparin binding peptide, and further bioactivation with heparin and IL-4, an anti-inflammatory cytokine responsible for M2 activation and polarization. Human monocyte-derived macrophages cultured on heparin-IL-4 bioactivated materials displayed an elongated morphology and an anti-inflammatory phenotype, with downregulation of pro-inflammatory cytokines and promotion of anti-inflammatory cytokines over time. This study represents the first step in designing a novel class of synthetic, bioactivated materials that harness the regenerative behavior of host macrophages towards in situ tissue regeneration.


Feeding Stimulates Sphingosine-1-Phosphate Mobilization in Mouse Hypothalamus.
Previous studies have shown that the sphingolipid-derived mediator sphingosine-1-phosphate (S1P) reduces food intake by activating G protein-coupled S1P receptor-1 (S1PR1) in the hypothalamus. Here, we examined whether feeding regulates hypothalamic mobilization of S1P and other sphingolipid-derived messengers. We prepared lipid extracts from the hypothalamus of C57Bl6/J male mice subjected to one of four conditions: free feeding, 12 h fasting, and 1 h or 6 h refeeding. Liquid chromatography/tandem mass spectrometry was used to quantify various sphingolipid species, including sphinganine (SA), sphingosine (SO), and their bioactive derivatives SA-1-phosphate (SA1P) and S1P. In parallel experiments, transcription of S1PR1 (encoded in mice by the S1pr1 gene) and of key genes of sphingolipid metabolism (Sptlc2, Lass1, Sphk1, Sphk2) was measured by RT-PCR. Feeding increased levels of S1P (in pmol-mg-1 of wet tissue) and SA1P. This response was accompanied by parallel changes in SA and dihydroceramide (d18:0/18:0), and was partially (SA1P) or completely (S1P) reversed by fasting. No such effects were observed with other sphingolipid species targeted by our analysis. Feeding also increased transcription of Sptlc2, Lass1, Sphk2, and S1pr1. Feeding stimulates mobilization of endogenous S1PR1 agonists S1P and SA1P in mouse hypothalamus, via a mechanism that involves transcriptional up-regulation of de novo sphingolipid biosynthesis. The results support a role for sphingolipid-mediated signaling in the central control of energy balance.


Ortner's syndrome in association with mitral valve prolapse.
The case of an 83-year-old woman with a history of hypertension, valvular heart disease, atrial fibrillation, and cardiomegaly is presented. The patient also had progressive hoarseness of her voice and intermittent dysphagia. Ear, nose, and throat examination revealed left vocal cord paralysis. Echocardiography revealed severely dilated left (LA) and right atria (RA), moderate mitral regurgitation, severe tricuspid regurgitation, and prolapse of both these valves. A review of literature of Ortner's or cardiovocal syndrome is presented. Ortner's syndrome due to mitral valve prolapse has not been reported previously.


Slight Variations in the Sequence Downstream of the Polyadenylation Signal Significantly Increase Transgene Expression in HEK293T and CHO Cells.
Compared to transcription initiation, much less is known about transcription termination. In particular, large-scale mutagenesis studies have, so far, primarily concentrated on promoter and enhancer, but not terminator sequences. Here, we used a massively parallel reporter assay (MPRA) to systematically analyze the influence of short (8 bp) sequence variants (mutations) located downstream of the polyadenylation signal (PAS) on the steady-state mRNA level of the upstream gene, employing an eGFP reporter and human HEK293T cells as a model system. In total, we evaluated 227,755 mutations located at different overlapping positions within +17..+56 bp downstream of the PAS for their ability to regulate the reporter gene expression. We found that the positions +17..+44 bp downstream of the PAS are more essential for gene upregulation than those located more distal to the PAS, and that the mutation sequences ensuring high levels of eGFP mRNA expression are extremely T-rich. Next, we validated the positive effect of a couple of mutations identified in the MPRA screening on the eGFP and luciferase protein expression. The most promising mutation increased the expression of the reporter proteins 13-fold and sevenfold on average in HEK293T and CHO cells, respectively. Overall, these findings might be useful for further improving the efficiency of production of therapeutic products, e.g., recombinant antibodies.


Transplantation of Flk-1+ human bone marrow-derived mesenchymal stem cells promotes behavioral recovery and anti-inflammatory and angiogenesis effects in an intracerebral hemorrhage rat model.
Mesenchymal stem cells (MSCs) have been successfully used for the treatment of experimental intracerebral hemorrhage (ICH). However, the neuroprotective mechanisms through which MSCs improve neurological functional recovery are not fully understood. In the present study, we tested the hypothesis that treatment with MSCs inhibits inflammation after ICH and reduces subsequent brain injury. Adult rats subjected to stereotaxic injection of collagenase VII were transplanted with a subpopulation of human bone marrow-derived MSCs (hBMSCs), termed fetal liver kinase (Flk)-1(+) hBMSCs, or saline into the ipsilateral brain parenchyma 1 day after ICH. Significant recovery of behavior was noted in the Flk-1(+) hBMSC-treated rats beginning 3 days after ICH compared with the control group. Brain water content was significantly decreased in the ipsilateral hemispheres of the Flk-1(+) hBMSC-treated rats when compared with the controls 3 days after ICH. The relative hemorrhage volume was reduced 55 days after Flk-1(+) hBMSC treatment. However, this change was not statistically significant. Flk-1(+) hBMSCs significantly inhibited the proliferation of rat peripheral blood mononuclear cells (rPBMCs) induced in a mixed lymphocyte reaction. Consistently, we found a significant anti-inflammatory effect of Flk-1(+) hBMSCs on the ICH brain, including a decrease in neutrophil infiltration and microglial activation in the peri-ICH area, and downregulation of inflammatory mediators, such as interleukin (IL)-1β, IL-2, IL-4, IL-6, and tumor necrosis factor (TNF)-α. In addition, Flk-1+ hBMSC treatment significantly increased vascular density in the peri-ICH area, and transplanted Flk-1(+) hBMSCs were found to be incorporated into the cerebral vasculature 55 days after transplantation. Overall, these data suggest an essential role for Flk-1(+) hBMSCs in reducing inflammatory infiltration, promoting angiogenesis, and improving functional recovery after ICH in rats.


Symptom clusters in traumatic spinal cord injury: an exploratory literature review.
Chronic pain, spasticity, and depression are three common secondary health problems experienced by persons living with traumatic spinal cord injury. Limited research exists related to the interaction of these symptoms and their cumulative effect on patient outcomes and quality of life. This article is a report of an exploratory literature review that uses the conceptual approach of symptom clustering to classify secondary health problems in spinal cord injury. CINAHL, MEDLINE, and PsycINFO databases were searched for studies published in English between 1990 and 2008. Eleven articles met the inclusion criteria for review. Studies were categorized by levels of evidence defined by the American Association of Neuroscience Nurses Clinical Practice Guidelines. Research on the interaction of the symptom pair chronic pain and depression was evident. Several surveys reported the occurrence of multiple symptoms, but none specifically addressed the management of the interactive effects of the symptom cluster chronic pain, spasticity, and depression. Increased disciplinary knowledge of symptom clusters in the spinal cord injury population is needed for research and practice. Furthermore, determining symptom clusters in this population may assist clinicians in providing targeted interventions to simultaneously treat multiple symptoms.


Molecular cloning and characterization of the pseudorabies virus US1 gene.
Using polymerase chain reaction, a 1050-bp sequence of the US1 gene was amplified from the pseudorabies virus (PRV) Becker strain genome; identification of the US1 gene was confirmed by further cloning and sequencing. Bioinformatics analysis indicated that the PRV US1 gene encodes a putative polypeptide with 349 amino acids. The encoded protein, designated PICP22, had a conserved Herpes_IE68 domain, which was found to be closely related with the herpes virus immediate early regulatory protein family and is highly conserved among the counterparts encoded by Herpes_IE68 genes. Multiple nucleic acid sequence and amino acid sequence alignments suggested that the product of PRV US1 has a relatively higher homology with ICP22-like proteins of genus Varicellovirus than with those of other genera of Alphaherpesvirinae. In addition, phylogenetic analysis showed that PRV US1 has a close evolutionary relationship with members of the genus Varicellovirus, especially Equid herpes virus 1 (EHV-1), EHV-4 and EHV-9. Antigen prediction indicated that several potential B-cell epitopes are located in PICP22. Also, subcellular localization analysis demonstrated that PICP22 is predominantly located in the cytoplasm, suggesting that it might function as a cytoplasmic-targeted protein.


Reliability of the OptoJump Next System for Measuring Temporal Values in Elite Racewalking.
Hanley, B and Tucker, CB. Reliability of the OptoJump Next system for measuring temporal values in elite racewalking. J Strength Cond Res 33(12): 3438-3443, 2019-Racewalking is an Olympic event where athletes are not permitted a visible loss of contact, with the result that competitors try to minimize flight times. The accuracy of measurements taken during testing is dependent on valid and reliable systems to determine temporal values. The aim of the study was to compare different methodologies used to measure contact and flight times in overground and treadmill racewalking. Eighteen racewalkers completed overground and instrumented treadmill trials at 5 speeds, during which flight and contact times were measured using the OptoJump Next photocell system (1,000 Hz), high-speed videography (500 Hz), and force plates (1,000 Hz). Results from OptoJump Next were extracted using 5 settings based on the number of light-emitting diodes (LEDs) activated (GaitIn_GaitOut) and annotated as 0_0, 1_1, 2_2, 3_3, and 4_4. Regarding flight time measurements for the overground condition, the 2_2 LED setting had the best 95% confidence interval (95% CI) for intraclass correlation coefficient (ICC) (0.978-0.988), the least bias (0.000 seconds), and the lowest random error (RE) (0.008 seconds). For the treadmill condition, the 0_0 LED setting had the best 95% CI for ICC (0.890-0.957), the least bias (0.004 seconds), and the lowest RE (0.017 seconds). Although high-speed videography also provided highly reliable results, the equally reliable and quicker availability of results using OptoJump Next is beneficial in laboratory-based testing. Coaches and researches are advised to alter the system's LED settings as appropriate and to report these settings with their findings.


Urine Proteome Biomarkers in Kidney Diseases. I. Limits, Perspectives, and First Focus on Normal Urine.
Urine proteome is a potential source of information in renal diseases, and it is considered a natural area of investigation for biomarkers. Technology developments have markedly increased the power analysis on urinary proteins, and it is time to confront methodologies and results of major studies on the topics. This is a first part of a series of reviews that will focus on the urine proteome as a site for detecting biomarkers of renal diseases; the theme of the first review concerns methodological aspects applied to normal urine. Main issues are techniques for urine pretreatment, separation of exosomes, use of combinatorial peptide ligand libraries, mass spectrometry approaches, and analysis of data sets. Available studies show important differences, suggesting a major confounding effect of the technologies utilized for analysis. The objective is to obtain consensus about which approaches should be utilized for studying urine proteome in renal diseases.


Isolation and characterization of a 14500 molecular weight protein from brains and tissue cultures persistently infected with borna disease virus.
A protein with an apparent molecular weight of 14500 (14.5K) was extractable from homogenates of Borna disease virus-infected brains and tissue cultures using high concentrations of detergent and salt and by differential centrifugation procedures. The protein, present in an aggregated form, was remarkably resistant to proteinase K. Specific antibodies prepared in the homologous system (rat) recognized the 14.5K protein from various sources (infected brain of rat, mouse or chicken, and tissue cultures), but did not neutralize infectivity nor stain Borna disease virus-specific antigens from in vitro or in vivo preparations. Post-infection immune sera from different animal species did not detect the protein. This 14.5K protein was infection-specific but not disease-specific, and is inferred to be part of an internal virion component.


DNA from resin-embedded organisms: Past, present and future.
Past claims have been made for fossil DNA recovery from various organisms (bacteria, plants, insects and mammals, including humans) dating back in time from thousands to several million years BP. However, many of these recoveries, especially those described from million-year-old amber (fossil resin), have faced criticism as being the result of modern environmental contamination and for lack of reproducibility. Using modern genomic techniques, DNA can be obtained with confidence from a variety of substrates (e.g. bones, teeth, gum, museum specimens and fossil insects) of different ages, albeit always less than one million years BP, and results can also be obtained from much older materials using palaeoproteomics. Nevertheless, new attempts to determine if ancient DNA (aDNA) is present in insects preserved in 40 000-year old sub-fossilised resin, the precursor of amber, have been unsuccessful or not well documented. Resin-embedded specimens are therefore regarded as unsuitable for genetic studies. However, we demonstrate here, for the first time, that although a labile molecule, DNA is still present in platypodine beetles (Coleoptera: Curculionidae) embedded in six-year-old and two-year-old resin pieces from Hymenaea verrucosa (Angiospermae: Fabaceae) collected in Madagascar. We describe an optimised method which meets all the requirements and precautions for aDNA experiments for our purpose: to explore the DNA preservation limits in resin. Our objective is far from starting an uncontrolled search for aDNA in amber as it was in the past, but to start resolving basic aspects from the DNA preservation in resin and search from the most modern samples to the ancient ones, step by step. We conclude that it is therefore possible to study genomics from resin-embedded organisms, although the time limits remain to be determined.


How to write a manuscript.
Writing a scientific manuscript is an important component of academic medicine. For most novice writers, this task may seem quite formidable. This article will review the basic processes of writing a manuscript, with emphasis on the proper structure and organization of the article. The likelihood of a manuscript being accepted is highly dependent on its content and structure. A structured outline is the cornerstone of a successful manuscript. The current article presents the tools necessary for writing a successful manuscript.


Scoping the impact of COVID-19 on the nexus of statelessness and health in Council of Europe member states.
Stateless communities in Europe include ethnic Russians in the Baltic States, recent migrants, refugees, Roma, and other members of minority groups. Increases in COVID-19 infection have been observed in many European countries, including reported outbreaks in groups that include people and communities affected by statelessness, who often live in congested and sub-standard unhygienic conditions, work in informal sectors which hampers their adherence to public health measures (self-isolation/physical distancing/hand sanitation), or who are detained in immigration detention centres. The impact of COVID-19 on stateless people in Europe (estimated to be at least 600,000) is currently under researched, and there is an imperative to understand their experiences and situation, in order to generate evidence based measures, responses and actions to protect those most at risk. In order to better understand their unique position during the COVID-19 pandemic, we conducted a scoping review to explore and assess the nexus between statelessness and health during COVID-19 in Europe. Literature was found representing ten Council of Europe countries (Bulgaria, Denmark, Greece, Italy, Romania, Russia, Slovakia, Ireland, Ukraine and the United Kingdom), with 15 publications representing multiple countries. Four publications specifically focused on stateless people. The remainder focused on populations which include people disproportionately affected by statelessness both in the migratory context and those in situ (minority groups including Roma and ethnic Russians, and refugees and migrants). Three themes emerged from the analysis (Environmental determinants of health; access to healthcare services; and racism and vilification), with higher level abstraction centring on the nexus between existing adverse environmental determinants of health, compounded barriers to access healthcare during COVID-19; and the concerning rise in hate crime and scapegoating of minority populations during the COVID-19 emergency. Whilst the right to healthcare is a fundamental human right, with universal application and with access to healthcare services ensured to every human being without regards to race, religion or other criteria, including nationality status, this appears not to be the case for populations affected by statelessness during the COVID-19 health and state emergency. The right to a nationality (and realisation of the right to health and access to healthcare/public services) in the current pandemic times is crucial in a targeted effective and culturally sensitive public health response. The hidden nature of statelessness, coupled with the marginalisation of stateless people, exacerbates the structural underpinning and interplay between statelessness, human rights, health rights and right to nationality during the COVID-19 pandemic. The review further highlights the need to protect stateless people. We further cannot underestimate the need for sensitive legal, health and social response measures to tackle disease transmission in vulnerable groups, continued statelessness of people in Europe, and hate crime, xenophobia and discrimination of those perceived to be at risk of contagion.


Empathy at a distance: a qualitative study on the impact of publically-displayed art on observers.
While there is some evidence in the literature on the impact of art therapy for consumers, there is comparatively little written on how art that has been created by consumers impacts on those observing the art. This paper reports on a qualitative research study that sought to determine if publically-displayed art created by young consumers impacted on stigma reduction and self-help-seeking behaviours of the observers. The findings derived from the thematic analysis of qualitative interviews suggested that publically-displayed art is a safe medium, through which empathy and understanding towards young people with mental illness can be enhanced, and that the art generates discussion and self-help behaviours for mental illness. These findings highlight how mental health nurses can promote social inclusion and reduce stigma through public mental health initiatives that are an important inclusion in the scope of mental health nursing practice.


Molecular weight distribution evaluation of polysaccharides and glycoconjugates using analytical ultracentrifugation.
We review here the advances that have been made in methodology using the analytical ultracentrifuge for characterising polydisperse polymer systems with a quasi-continuous distribution of molecular weight. These advances include improved ways of defining the weight and z-average molecular weights of a distribution from sedimentation equilibrium experiments and hence the ratio Mz/Mw, off-line calibration of size exclusion chromatography by sedimentation equilibrium and conversion of a sedimentation coefficient distribution into a molecular weight distribution. Although these methods are applicable to other polymeric systems, we focus on polysaccharides, mucins and other glycoconjugates, systems which particularly benefit from the availability of recently available long optical path length cells for the minimisation of complications through thermodynamic non-ideality.


[Identification and differentiation of Campylobacter fetus subspecies by PCR].
The species Campylobacter (C.) fetus is divided into the subspecies venerealis and fetus, which differ in epidemiology and clinical importance. The differences between these subspecies make an accurate distinction essential. Differentiation of C. fetus by traditional microbiological methods is only based on two reactions (tolerance to glycin, Na selenite reduction), in which C. fetus ssp. venerealis reacts negatively. However, the value of both reactions is limited. We used a specific PCR-based assay for identifying and differentiating the two C. fetus subspecies, which was recently developed by HUM et al. (1997). In this assay, a 764 bp amplicon is produced using primers MG3F and MG4R for both subspecies of C. fetus. In contrast to HUM et al. (1997), this amplicon was approximately 200 bp smaller. This discrepancy can't be explained. Afterwards, the primers VenSF and VenSR are used for differentiation. The identification of the sub-species venerealis is based on the presence of a 142 bp amplicon, which is not formed with subspecies fetus. The type strains of both C. fetus subspecies were used as positive controls. Non-specific reactions were not observed. In this PCR assay, 73 field strains were investigated (among them 24 C. fetus ssp. veneralis, 26 C. fetus ssp. fetus). In these investigations, the method has proved its diagnostic suitability. The results of the traditional microbiological differentiation of the C. fetus field strains could be confirmed by the PCR assay. In future, the traditional phenotypic characterization of C. fetus subspecies remains indispensable, but this PCR assay constitutes a valuable method for the confirmation of these results.


What can we learn from global sensitivity analysis of biochemical systems?
Most biological models of intermediate size, and probably all large models, need to cope with the fact that many of their parameter values are unknown. In addition, it may not be possible to identify these values unambiguously on the basis of experimental data. This poses the question how reliable predictions made using such models are. Sensitivity analysis is commonly used to measure the impact of each model parameter on its variables. However, the results of such analyses can be dependent on an exact set of parameter values due to nonlinearity. To mitigate this problem, global sensitivity analysis techniques are used to calculate parameter sensitivities in a wider parameter space. We applied global sensitivity analysis to a selection of five signalling and metabolic models, several of which incorporate experimentally well-determined parameters. Assuming these models represent physiological reality, we explored how the results could change under increasing amounts of parameter uncertainty. Our results show that parameter sensitivities calculated with the physiological parameter values are not necessarily the most frequently observed under random sampling, even in a small interval around the physiological values. Often multimodal distributions were observed. Unsurprisingly, the range of possible sensitivity coefficient values increased with the level of parameter uncertainty, though the amount of parameter uncertainty at which the pattern of control was able to change differed among the models analysed. We suggest that this level of uncertainty can be used as a global measure of model robustness. Finally a comparison of different global sensitivity analysis techniques shows that, if high-throughput computing resources are available, then random sampling may actually be the most suitable technique.


[Primary hyperparathyroidism with severe skeletal involvement in one patient with parathyroid carcinoma].
Parathyroid carcinoma is an uncommon cause of primary hyperparathyroidism; however, when this condition is severe, cancer must be suspected. We report on a 28-year-old male with severe hypercalcemia, cachexia, acute pancreatitis, urolithiasis, anemia and a severe skeletal involvement with multiple fractures. The patient had a 4-cm parathyroid tumor, that was surgically excised, along with the ipsilateral thyroid lobe. During the postoperative period, he had a severe and prolonged hungry bone syndrome, with a slow recovery of fractures, with functional and anatomical sequelae in the extremities. PTH levels were adequate for the serum calcium during the 16 months of follow-up.


Evidence of the involvement of caspase-1 under physiologic and pathologic cellular stress during human pregnancy: a link between the inflammasome and parturition.
Caspase-1 is a component of the NALP3 inflammasome, a cytosolic multiprotein complex that mediates the processing of pro-inflammatory caspases and cytokines. The inflammasome represents the first line of defense against cellular stress and is a crucial component of innate immunity. Caspase-1 is the enzyme responsible for the cleavage and activation of interleukin (IL)-1 beta, which is a potent pro-inflammatory cytokine, and plays a central role in the mechanisms leading to labor (preterm and term) particularly in the context of intrauterine infection/inflammation. In addition, caspase-1 cleaves IL-18 and IL-33. The objectives of this study were to determine whether there is a relationship between amniotic fluid concentrations of caspase-1 and gestational age, parturition (term and preterm), and intra-amniotic infection/inflammation (IAI). A cross-sectional study was conducted including 143 pregnant women in the following groups: (1) mid-trimester of pregnancy (n = 18); (2) term not in labor (n = 25); (3) term in labor (n = 28); (4) preterm labor (PTL) who delivered at term (n = 23); (5) PTL without IAI who delivered preterm (n = 32); (6) PTL with IAI who delivered preterm neonates (n = 17). Caspase-1 concentrations in amniotic fluid were determined by a specific and sensitive immunoassay. Non-parametric statistics were used for analysis. (1) Caspase-1 was detected in amniotic fluid of women at term, but in none of the mid-trimester samples. (2) Patients in labor at term had a significantly higher median amniotic fluid concentration of caspase-1 than women at term not in labor (term in labor: 10.5 pg/mL, range 0.0-666.0 vs. term not in labor: 5.99 pg/mL, range 0.0-237.4; p < 0.05). (3) Among patients with spontaneous PTL, those with IAI (median 41.4 pg/mL, range 0.0-515.0) had a significantly higher median amniotic fluid caspase-1 concentration than those without IAI who delivered preterm (median 0.0 pg/mL, range 0.0-78.4) and than those who delivered at term (median 0.0 pg/mL, range 0.0-199.5); p < 0.001 for both comparisons. (1) The presence and concentration of caspase-1 in the amniotic fluid varies as a function of gestational age. (2) Women with spontaneous labor at term had a higher median caspase-1 amniotic fluid concentration than women at term without labor. This suggests that the inflammasome may be activated in spontaneous parturition at term. Since most women with labor do not have intra-amniotic infection, we propose that cellular stress during labor accounts for activation of the inflammasome. (3) Preterm labor associated with infection/inflammation was also associated with a high concentration of caspase-1, suggesting that infection may induce caspase-1 production and activation of the inflammasome. (4) The sequential activation of the inflammasome and caspase-1, leading to interleukin-1 beta processing and secretion, is a candidate pathway leading to the activation of the common pathway of parturition.


Acquired initiating mutations in early hematopoietic cells of CLL patients.
Appropriate cancer care requires a thorough understanding of the natural history of the disease, including the cell of origin, the pattern of clonal evolution, and the functional consequences of the mutations. Using deep sequencing of flow-sorted cell populations from patients with chronic lymphocytic leukemia (CLL), we established the presence of acquired mutations in multipotent hematopoietic progenitors. Mutations affected known lymphoid oncogenes, including BRAF, NOTCH1, and SF3B1. NFKBIE and EGR2 mutations were observed at unexpectedly high frequencies, 10.7% and 8.3% of 168 advanced-stage patients, respectively. EGR2 mutations were associated with a shorter time to treatment and poor overall survival. Analyses of BRAF and EGR2 mutations suggest that they result in deregulation of B-cell receptor (BCR) intracellular signaling. Our data propose disruption of hematopoietic and early B-cell differentiation through the deregulation of pre-BCR signaling as a phenotypic outcome of CLL mutations and show that CLL develops from a pre-leukemic phase. The origin and pathogenic mechanisms of CLL are not fully understood. The current work indicates that CLL develops from pre-leukemic multipotent hematopoietic progenitors carrying somatic mutations. It advocates for abnormalities in early B-cell differentiation as a phenotypic convergence of the diverse acquired mutations observed in CLL.


Seipin traps triacylglycerols to facilitate their nanoscale clustering in the endoplasmic reticulum membrane.
Seipin is a disk-like oligomeric endoplasmic reticulum (ER) protein important for lipid droplet (LD) biogenesis and triacylglycerol (TAG) delivery to growing LDs. Here we show through biomolecular simulations bridged to experiments that seipin can trap TAGs in the ER bilayer via the luminal hydrophobic helices of the protomers delineating the inner opening of the seipin disk. This promotes the nanoscale sequestration of TAGs at a concentration that by itself is insufficient to induce TAG clustering in a lipid membrane. We identify Ser166 in the α3 helix as a favored TAG occupancy site and show that mutating it compromises the ability of seipin complexes to sequester TAG in silico and to promote TAG transfer to LDs in cells. While the S166D-seipin mutant colocalizes poorly with promethin, the association of nascent wild-type seipin complexes with promethin is promoted by TAGs. Together, these results suggest that seipin traps TAGs via its luminal hydrophobic helices, serving as a catalyst for seeding the TAG cluster from dissolved monomers inside the seipin ring, thereby generating a favorable promethin binding interface.


Differential control of human Treg and effector T cells in tumor immunity by Fc-engineered anti-CTLA-4 antibody.
Anti-CTLA-4 mAb is efficacious in enhancing tumor immunity in humans. CTLA-4 is expressed by conventional T cells upon activation and by naturally occurring FOXP3+CD4+ Treg cells constitutively, raising a question of how anti-CTLA-4 mAb can differentially control these functionally opposing T cell populations in tumor immunity. Here we show that FOXP3high potently suppressive effector Treg cells were abundant in melanoma tissues, expressing CTLA-4 at higher levels than tumor-infiltrating CD8+ T cells. Upon in vitro tumor-antigen stimulation of peripheral blood mononuclear cells from healthy individuals or melanoma patients, Fc-region-modified anti-CTLA-4 mAb with high antibody-dependent cell-mediated cytotoxicity (ADCC) and cellular phagocytosis (ADCP) activity selectively depleted CTLA-4+FOXP3+ Treg cells and consequently expanded tumor-antigen-specific CD8+T cells. Importantly, the expansion occurred only when antigen stimulation was delayed several days from the antibody treatment to spare CTLA-4+ activated effector CD8+T cells from mAb-mediated killing. Similarly, in tumor-bearing mice, high-ADCC/ADCP anti-CTLA-4 mAb treatment with delayed tumor-antigen vaccination significantly prolonged their survival and markedly elevated cytokine production by tumor-infiltrating CD8+ T cells, whereas antibody treatment concurrent with vaccination did not. Anti-CTLA-4 mAb modified to exhibit a lesser or no Fc-binding activity failed to show such timing-dependent in vitro and in vivo immune enhancement. Thus, high ADCC anti-CTLA-4 mAb is able to selectively deplete effector Treg cells and evoke tumor immunity depending on the CTLA-4-expressing status of effector CD8+ T cells. These findings are instrumental in designing cancer immunotherapy with mAbs targeting the molecules commonly expressed by FOXP3+ Treg cells and tumor-reactive effector T cells.


Prevalence of Anxiety as a Variable in Treatment Outcomes for Individuals With Chronic Refractory Cough.
Anxiety is a mental state characterized by an intense sense of tension, worry, or apprehension relative to something adverse that might happen in the future. Anxiety is a known comorbidity in cough patients, yet its prevalence among those with chronic refractory cough (CRC) is unknown. Anxiety is not typically assessed during evaluation for CRC, but treatments for CRC such as neuromodulators and behavioral cough suppression therapy (BCST) may potentially attenuate anxiety. This preliminary study investigates the potential prevalence of anxiety in CRC and its possible role in treatment outcomes. CRC patients seen in a specialty clinic at the University of Utah or the University of Montana completed the Leicester Cough Questionnaire (LCQ) pre- and post-BCST treatment. Participants were dichotomized into positive anxiety screen (PAS) and negative anxiety screen (NAS) groups based on presence or absence of documented anxiety within electronic medical records at the University of Utah and based on a Generalized Anxiety Disorder-7 score > 5 at the University of Montana. Of the 86 total participants, 37 (43%) were in the PAS group (29 females, Mage = 56 ± 13) and 49 (57%) were in the NAS group (36 females, Mage = 64 ± 14). Eighty-nine percent of CRC participants with a PAS reported a clinically meaningful improvement in LCQ total score following treatment compared to 65% of NAS participants. Furthermore, mean pre- to posttreatment change scores on the LCQ were significantly greater within the PAS group (p = .002, Cohen's d = 0.7, indicating a moderate to large effect size). This preliminary study suggests that (a) anxiety may be prevalent among those with CRC and (b) those patients who screen positive for anxiety report greater benefit from BCST.


Assessing the ecological status in the context of the European Water Framework Directive: where do we go now?
The Water Framework Directive (WFD) is now well established as the key management imperative in river basins across Europe. However, there remain significant concerns with the way WFD is implemented and there is now a need for water managers and scientists to communicate better in order to find solutions to these concerns. To address this, a Science-Policy Interface (SPI) activity was launched in 2010 led by Directorate-General for Research and Innovation and Onema (the French national agency for water and aquatic ecosystems), which provided an interactive forum to connect scientists and WFD end-users. One major aim of the SPI activity was to establish a list of the most crucial research and development needs for enhancing WFD implementation. This paper synthesises the recommendations from this event highlighting 10 priority issues relating to ecological status. For lakes, temporary streams and transitional and coastal waters, WFD implementation still suffers from a lack of WFD-compliant bioassessment methods. For rivers, special attention is required to assess the ecological impacts of hydromorphological alterations on biological communities, notably those affecting river continuity and riparian covering. Spatial extrapolation tools are needed in order to evaluate ecological status for water bodies for which no data are available. The need for more functional bioassessment tools as complements to usual WFD-compliant tools, and to connect clearly good ecological state, biodiversity and ecosystem services when implementing WFD were also identified as crucial issues.


Structure of a pilin monomer from Pseudomonas aeruginosa: implications for the assembly of pili.
Type IV pilin monomers assemble to form fibers called pili that are required for a variety of bacterial functions. Pilin monomers oligomerize due to the interaction of part of their hydrophobic N-terminal alpha-helix. Engineering of a truncated pilin from Pseudomonas aeruginosa strain K122-4, where the first 28 residues are removed from the N terminus, yields a soluble, monomeric protein. This truncated pilin is shown to bind to its receptor and to decrease morbidity and mortality in mice upon administration 15 min before challenge with a heterologous strain of Pseudomonas. The structure of this truncated pilin reveals an alpha-helix at the N terminus that lies across a 4-stranded antiparallel beta-sheet. A model for a pilus is proposed that takes into account both electrostatic and hydrophobic interactions of pilin subunits as well as previously published x-ray fiber diffraction data. Our model indicates that DNA or RNA cannot pass through the center of the pilus, however, the possibility exists for small organic molecules to pass through indicating a potential mechanism for signal transduction.


Purification, characterization, and milk coagulating properties of ginger proteases.
Ginger proteases are used as milk coagulants in making a Chinese traditional milk product (Jiangzhinai or Jiangzhuangnai), suggesting their potential as a source of rennet substitute that might be applicable in the modern dairy industry. In this study, ginger proteases were extracted from fresh ginger rhizome by using phosphate buffer and subsequently purified by ion exchange chromatography. Ginger proteases, all with a molecular weight around 31 kDa, were found to exist in 3 forms with isoelectric point values around 5.58, 5.40, and 5.22, respectively. These enzymes had very similar biochemical behavior, exhibiting optimal proteolytic activity from 40 to 60 °C and maximum milk clotting activity at 70 °C. They were capable of hydrolyzing isolated α(S1)-, β-, and κ-casein, of which α(S1)-casein was most susceptible to the enzyme; κ-casein was hydrolyzed with a higher specificity than α(S1)- and β-casein. In addition, the ginger proteases exhibited a similar affinity for κ-casein and higher specificity with increasing temperature. Gel electrophoresis and mass spectra indicated that Ala90-Glu91 and His102-Leu103 of κ-casein were the preferred target bonds of ginger proteases. The milk clotting activity, affinity, and specificity toward κ-casein showed that ginger protease is a promising rennet-like protease that could be used in manufacturing cheese and oriental-style dairy foods.


Ankyrin-based cardiac arrhythmias: a new class of channelopathies due to loss of cellular targeting.
This review addresses a new mechanism for arrhythmia due to abnormal cellular localization of membrane ion channels and transporters. The focus is on ankyrins, a family of proteins that localize diverse membrane ion channels and transporters, and recent evidence that mutations affecting functions of ankyrins result in cardiac arrhythmia. A loss-of-function mutation of ankyrin-B in humans and a null mutation in mice result in a dominantly-inherited fatal cardiac arrhythmia initially classified as type 4 long QT syndrome. Characterization of additional probands suggests ankyrin-B mutations cause a new cardiac arrhythmia syndrome associated with sinus node dysfunction that is distinct from long QT syndrome. Ankyrin-B mutation results in elevated calcium transients in cardiomyocytes accompanied by loss of cellular targeting of Na/K ATPase, Na/Ca exchanger, and InsP3 receptor (all ankyrin-binding proteins) to cardiomyocyte membrane domains. The principal voltage-gated Na channel in heart, Nav1.5, is directly associated with ankyrin-G, which is encoded by a distinct gene from ankyrin-B. Mutation of Nav1.5 causing loss of binding to ankyrin-G results in Brugada syndrome and loss of targeting of Nav1.5 to the cell surface of cardiomyocytes. Ankyrin-B and ankyrin-G are recently recognized constituents of the heart that target diverse ion channels/pumps/transporters to physiologic sites of action in cardiomyocytes. Mutations of ankyrin-B cause a newly defined cardiac arrhythmia syndrome associated with abnormal calcium homeostasis in a mouse model. Ankyrin-G associates with the principal voltage-gated Na channel in the heart, and loss of this interaction due to mutation of Nav1.5 results in Brugada syndrome.


Massive haematemesis due to strangulated gangrenous gastric herniation as the delayed presentation of post-traumatic diaphragmatic rupture.
Diaphragmatic injuries are quite uncommon and often result from either blunt or penetrating trauma. Diaphragmatic ruptures are usually associated with abdominal trauma; however, it can occur in isolation. Acute traumatic rupture of the diaphragm may go unnoticed and there is often a delay between the injury and the diagnosis. Patients present with non-specific symptoms and may complain of chest pain, abdominal pain, dyspnoea, tachypnoea and cough, heartburn and symptoms of gastro-oesophageal reflux. Respiratory distress and faeco-pneumothorax have been reported. We present an interesting case of traumatic diaphragmatic hernia presenting 5 years after a road traffic accident as acute abdomen and massive haematemesis due to strangulated gangrenous gastric hernia.


The relevance of European Biota Quality Standards on the ecological water quality as determined by the multimetric macro-invertebrate index: A Flemish case study.
European Biota Quality Standards (EQSbiota), for compounds with low water solubility and high biomagnification, were created to sustain water quality and protect top predators and humans from secondary poisoning. In reality, for multiple compounds, an exceedance of these standards is often reported in literature without a decrease in ecological water quality determined by biotic indices. In the present study, threshold concentrations were defined in biota (from 44 sampling locations throughout Flanders (Belgium)), above which a good ecological water quality, assessed by the Multimetric Macroinvertebrate Index Flanders (MMIF), was never reached. Threshold values were compared to current EQSbiota. Accumulated perfluoroctane sulfonate (PFOS), mercury (Hg), hexabromocyclododecane (HBCD), polybrominated diphenyl ethers (PBDEs), dioxins and polychlorinated biphenyls (PCBs) concentrations were measured in muscle tissue of European yellow eel (Anguilla anguilla) and perch (Perca fluviatilis). Fluoranthene and benzo(a)pyrene were also analyzed in translocated mussels (Dreissena bugensis, D. polymorpha and Corbicula fluminea). Threshold values could only be calculated using a 90th quantile regression model for PFOS (in perch; 12 µg/kg ww), PCBs (in eel; 328 µg/kg ww) and benzo(a)pyrene (in mussels: 4.35 µg/kg ww). The lack of a significant regression model for the other compounds indicated an effective threshold value higher than the concentrations measured in the present study. Alternatively, the 95th percentile of concentrations measured in locations with a good ecological quality (MMIF≥0.7), was calculated for all compounds as an additional threshold value. Finally, fish concentrations were standardized for 5% lipid content (or 26% dry weight content for PFOS and Hg). Threshold values for PFOS and benzo(a)pyrene and the 95th percentiles for dioxins and fluoranthene were comparable to the existing standards. For all other compounds, the 95th percentile was higher than the current EQSbiota, while for HBCD, it was lower. These results strongly advise revising and fine-tuning the current EQSbiota, especially for ∑PBDE and HBCD.


The prevalence of sickle cell disease in Abu-AL-Khasib district of southern Iraq.
At attempt was made to estimate the prevalence of sickle cell disease among the primary school children in Abul-al-Khasib district of southern Iraq which is known for a relatively high prevalence. Examination of 706 children in the age group of 10 to 12 years from 10 out of 20 schools randomly chosen in four villages of Abu-al-Khasib showed an overall prevalence rate of 16% as compared to 2.5% seen in a control population of 525 children belonging to five urban schools in Basrah. Sickle cell trait was evident in 13.3% of the cases. Sickledex was used for initial testing and those showing positive sickling phenomenon were confirmed by cellulose acetate haemoglobin electrophoresis. The need for introducing early screening measures as well as marriage/genetic counselling as part of the overall health and welfare services in stressed.


Effects of tryptophan on depression and aggression in STZ-D mice.
Streptozocin-induced diabetic (STZ-D) mice have reduced brain concentrations of tryptophan, a precursor substance for 5-hydroxytryptamine, and show lengthened immobility in Porsolt's swim test, a putative animal model of depression. This study investigated whether tryptophan affects behavior in Porsolt's swim test in STZ-administered male National Institutes of Health Swiss mice. In addition, the effect of tryptophan on behavior in the resident-intruder test of aggression was studied. Tryptophan is effective in the treatment of mild depression and may reduce aggressive behavior. Diabetes was induced with injection of 200 mg/kg body wt i.p. STZ. Two weeks after STZ treatment, the mice received 0, 50, and 100 mg/kg i.p. tryptophan 60 min before the swim test. The STZ-administered mice exhibited lengthened immobility in the swim test, and tryptophan caused a dose-related shortening in their immobility times. The control and STZ mice, which were isolated for 1 wk before the resident-intruder test, did not show any difference in the time spent in social investigation or aggressive or defensive behaviors. However, 100 mg/kg i.p. tryptophan 60 min before the test reduced the social interaction and aggressive behavior of the STZ-D mice but increased these behaviors in controls. Results indicate that tryptophan shortens the increased immobility time and reduces social and aggressive behavior in STZ-D mice. Therefore, the reported reductions in the brain-tryptophan concentrations in STZ-D mice may participate in regulating their behavior.


Relationships among oncostatin M, insulin resistance, and chronic inflammation: a pilot study.
Objective Activated macrophages (M1-type macrophages) in adipose tissue secrete many proinflammatory cytokines that induce insulin resistance (IR). Oncostatin M (OSM), a member of the interleukin-6 (IL-6) family of Gp130 cytokines, plays an important role in a variety of biological functions, including the regulation of inflammatory responses. Proinflammatory cytokines released in patients with IR trigger a chronic, low-grade inflammatory reaction in blood vessel walls. This inflammator response leads to endothelial damage, which is the main mechanism for atherosclerosis and many cardiovascular diseases. Animal studies have reported a relationship between OSM and IR. To the best of our knowledge, however, few clinical studies have examined this topic. Therefore, we studied the relationship between serum levels of OSM and IR. Subjects and methods This prospective cross-sectional case-control study enrolled 50 people with IR (according to the HOMA-IR and QUICKI indices) and 34 healthy controls. The fasting blood concentrations of insulin, glucose, blood urea nitrogen (BUN), creatinine, aspartate aminotransferase (AST), alanine aminotransferase (ALT), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), triglyceride, total cholesterol, C-reactive protein (CRP), and OSM were determined. Results There were no significant differences between the two groups in age, sex, and HbA1c levels. Univariate analyses showed that waist circumference (WC) and levels of fasting glucose, insulin, CRP, HDL-C, OSM, HOMA-IR, and QUICKI differed between the two study groups. In multivariate analyses, both IR indices (QUICKI and HOMA) and OSM differed between the two groups. Conclusion OSM was correlated with the IR indices (QUICKI and HOMA). For simplicity, it might replace the other IR indices in the future. Further detailed studies are needed to confirm this.


Plasma concentrations of pregnancy-associated glycoproteins measured using anti-bovine PAG-2 antibodies on day 120 of gestation predict abortion in dairy cows naturally infected with Neospora caninum.
The present study sought to determine: (i) the effects of Neospora caninum infection and twin pregnancy on plasma pregnancy-associated glycoprotein-2 (PAG-2) concentrations throughout pregnancy and (ii) whether plasma PAG-2 concentrations could predict abortion in N. caninum-infected cows. The study was performed on a commercial Holstein-Friesian dairy herd in northeastern Spain and the final data included those recorded in 53 non-aborting and 19 aborting animals. Blood samples were collected immediately before pregnancy diagnosis (on Days 40, 90, 120, 150, 180 and 210 post-insemination) in non-aborting cows or until the time of abortion detection in aborting cows. General lineal models (GLM) repeated measures anova revealed the different behaviour of PAG-1 and PAG-2, and significant effects of Neospora seropositivity, cool season and twin pregnancy on plasma PAG-2 concentrations throughout gestation (between-subject effects). In addition, based on the odds ratios, the likelihood of abortion increased in Neospora-seropositive cows (by a factor of 7.0) compared to seronegative animals and decreased in cows with a high plasma PAG-2 concentration (>4.5 ng/ml) on Day 120 of pregnancy (by a factor of 0.24), compared to the remaining cows. In conclusion, there is a relationship between plasma PAG-2 concentrations and the risk of abortion in Neospora-infected dairy cows. Thus, plasma PAG concentrations measured using anti-boPAG-2 antiserum on Day 120 of gestation could serve as an indicator of the abortion risk in N. caninum infected animals; values <4.5 ng/ml indicating a high risk of abortion in chronically infected animals.


Inhibiting angiogenesis: interview with Dr Jean Plouët. Interview by Emma Quigley.
Dr Jean Plouët (Centre de Recherche Cardiovasculaire, INSERM U 689/Institut des Vaisseaux et du Sang) was interviewed by Emma Quigley (Commissioning Editor, Expert Opinion) on 20th June 2006. Born in Brittany, France, Dr Jean Plouët received his MD degree from Nantes University in 1977 and his PhD in Molecular Biology from Paris VII University in 1981. His first research topic, under the supervision of Dr Courtois and Dr Barritault was devoted to the purification and the description of the mechanisms of action of eye-derived growth factor, which turned out to be the retinal fibroblast growth factor (FGF)-2. He was appointed as Charge de Recherche by the Centre National de la Recherche Scientifique in 1981. He later conducted his research on the effect of FGF-2 on light transduction and demonstrated the role of opsin phosphorylation on FGF-2 release from retinal discs in Dr Pouliquen's laboratory. From 1987 to 1989, Dr Plouët worked in the Cancer Research Institute at the University of California, San Francisco, in Dr Gospodarowicz's laboratory where he co-discovered vascular endothelial growth factor (VEGF) with Dr Napoleone Ferrara. He established his own group in Toulouse where he worked on the mechanisms of action of VEGF and vascular targeting by anti-idiotypoc antibodies against VEGF and FGF. In 2005, Dr Plouët moved to Paris where he now works mainly on endogenous inhibitors of angiogenesis acting downstream of VEGF.


Physical Activity Reduces Risk of Premature Mortality in Patients With Type 1 Diabetes With and Without Kidney Disease.
The aims of the study were to assess how baseline leisure-time physical activity (LTPA) and its exercise components intensity, duration, and frequency are associated with all-cause and cardiovascular mortality in patients with type 1 diabetes 1) overall, 2) stratified by presence or absence of chronic kidney disease (CKD), and 3) stratified by sex. The study design was prospective and observational and included 2,639 patients with type 1 diabetes from the ongoing nationwide multicenter Finnish Diabetic Nephropathy (FinnDiane) Study. Mean follow-up time was 11.4 ± 3.5 years. LTPA was assessed by using a validated self-report questionnaire. Three hundred ten patients (11.7%) had CKD defined as an estimated glomerular filtration rate of ≤60 mL/min/1.73 m2. During follow-up, 270 deaths occurred. LTPA and all its components were associated with all-cause mortality, even after adjustment for the potential confounders sex, diabetic nephropathy, duration of diabetes, age at onset of diabetes, systolic blood pressure, triglycerides, BMI, and HbA1c. Only exercise intensity was associated with cardiovascular mortality after adjustment for the confounders. Of the patients with CKD, 127 died during follow-up. The total amount of LTPA and exercise frequency were independently associated with lower risk of all-cause mortality when adjusted for covariates. Exercise is associated with a lower risk of premature all-cause and cardiovascular mortality in patients with type 1 diabetes. This study also demonstrates that physical activity is associated with a lower risk of mortality in patients with type 1 diabetes and CKD.


[Ultrasonography and rheumatology].
Muskuloskeletal ultrasound has been incorporated by rheumatologist to the clinical practice over the past decade. The technical improvements of the devices allowed the production of high quality images contributing to better identification of joint inflammation and structural damage. In this review, we highlight the applications of ultrasound in the study of different rheumatic conditions.


Experiences of HIV-related stigma among HIV-positive older persons in Uganda--a mixed methods analysis.
There is limited data on stigma among older HIV-infected adults in sub-Saharan Africa. We describe the experiences of stigma and disclosure in a cohort of HIV-positive older people in Uganda. Using data from the Wellbeing of Older Peoples' Study of Kalungu (rural site) and Wakiso district (peri-urban site) residents, we measured self-reported stigma levels for 183 respondents (94 on antiretroviral therapy (ART); 88, not on ART) using a stigma score generated using three questions on stigma perceptions where 0 meant no stigma at all and 100 was maximum stigma. Based on two questions on disclosure, an overall score was computed. High disclosure was assigned to those who often or very often disclosed to the family and were never or seldom afraid to disclose elsewhere. We examined the experiences of HIV stigma of 25 adults (52% females) using semi-structured, open-ended interviews and monthly oral diaries over one year. Mean age of the respondents was 70 years (range 60-80 years) and 80% of all respondents were enrolled in ART. Interview transcripts were analysed using thematic content analysis. Overall, 55% of respondents had a high disclosure score, meaning they disclosed easily, and 47% had a high stigma score. The stigma scores were similar among those with high and low disclosure scores. In multivariate analyses with disclosure and stigma scores as dependent variables none of the respondents' characteristics had a significant effect at the 5% level. Qualitative data revealed that stigma ranges from: (1) perceptions (relatively passive, but leading to behaviour such as gossip, especially if not intended maliciously); to (2) discriminatory behaviour (active or enacted stigma; from malicious gossip to outright discrimination). Despite the relatively high levels of disclosure, older people suffer from high levels of stigma of various forms apart from HIV-related stigma. Efforts to assess for different forms of stigma at an individual level deserve greater attention from service providers and researchers, and must be context specific.


c-kit expression identifies cardiovascular precursors in the neonatal heart.
Directed differentiation of embryonic stem cells indicates that mesodermal lineages in the mammalian heart (cardiac, endothelial, and smooth muscle cells) develop from a common, multipotent cardiovascular precursor. To isolate and characterize the lineage potential of a resident pool of cardiovascular progenitor cells (CPcs), we developed BAC transgenic mice in which enhanced green fluorescent protein (EGFP) is placed under control of the c-kit locus (c-kit(BAC)-EGFP mice). Discrete c-kit-EGFP(+) cells were observed at different stages of differentiation in embryonic hearts, increasing in number to a maximum at about postnatal day (PN) 2; thereafter, EGFP(+) cells declined and were rarely observed in the adult heart. EGFP(+) cells purified from PN 0-5 hearts were nestin(+) and expanded in culture; 67% of cells were fluorescent after 9 days. Purified cells differentiated into endothelial, cardiac, and smooth muscle cells, and differentiation could be directed by specific growth factors. CPc-derived cardiac myocytes displayed rhythmic beating and action potentials characteristic of multiple cardiac cell types, similar to ES cell-derived cardiomyocytes. Single-cell dilution studies confirmed the potential of individual CPcs to form all 3 cardiovascular lineages. In adult hearts, cryoablation resulted in c-kit-EGFP(+) expression, peaking 7 days postcryolesion. Expression occurred in endothelial and smooth muscle cells in the revascularizing infarct, and in terminally differentiated cardiomyocytes in the border zone surrounding the infarct. Thus, c-kit expression marks CPc in the neonatal heart that are capable of directed differentiation in vitro; however, c-kit expression in cardiomyocytes in the adult heart after injury does not identify cardiac myogenesis.


Uterine plasminogen activator activity: modulation by steroid hormones.
We have used a sensitive and quantitative assay to investigate the hormonal regulation of plasminogen activator (PA) activity in the rat uterus. PA activity is increased 5-fold (per U protein or DNA) by low physiological (0.1 micrograms) doses of estradiol, with increases in activity first observed at approximately 12 h. The stimulation of PA activity shows strict specificity among the steroid hormones, being stimulated by estrogens only or by high doses of dihydrotestosterone, which are known to affect the estrogen receptor system, and this stimulation is suppressed markedly by triphenylethylene antiestrogens. Comparative dose-response studies with a variety of estrogens of different uterotropic potencies indicate a good correlation between the potencies of different estrogens in stimulating PA activity and uterine growth (diethylstilbestrol = 17 beta-estradiol greater than estrone = 17 alpha-estradiol greater than estriol), with the exception of the zearalanol estrogen P-1496, which was consistently a potent stimulator of PA activity while being a very weak uterotropic agent. These studies suggest that increases in uterine PA levels may serve as a good marker of estrogen action in the uterus. Although the role of PA in uterine function remains unknown at present, its relatively large increase (up to 25-fold increase in content per uterus) may play a role in tissue remodeling during uterine growth.


Models of suicide risk: nice persons.
The hypotheses are advanced that: (1) certain types of persons at risk for suicide represent identifiable clinical models, and (2) that such models provide relatively homogeneous samples from which specific high-risk indicators can be derived for clinical application to others who represent that model. Nine-hundred and eighty-six psychiatric inpatients representing the "Nice Persons" model, including 35 suicides, were randomly divided into an index and a validation set. Statistical analysis of the index set (N = 579), using a screening procedure followed by linear discriminant and linear logistic procedures, identified 11 high-risk indicators from 184 prospectively determined variables. When applied to the independent validation set (N = 407), the 11 indicators identified the suicides in that set with a sensitivity of 46% and specificity of 88%. Though not statistically significant, this level of efficiency is clinically of considerable potential value. Most importantly, the hypothesis is testable and is shown to deserve further development.


Cochlear implants in children: a study of parental stress and adjustment.
A contextual model of stress was employed to examine the impact of cochlear implantation on parents' levels of stress and psychological adjustment. This model provided a framework for identifying stressors tied directly to the situation, and pointed to important life roles that may be altered. Twenty-four mothers of children implanted with cochlear implants completed a series of questionnaires assessing stressors related to daily parenting tasks, time demands, and childhood deafness. Standardized measures of depression, anxiety, and somatic complaints were also administered. The results provided preliminary evidence that parents of children receiving cochlear implants experience higher levels of stress and poorer psychological adjustment than parents of normally hearing children. Greater parenting stress and emotional distress were reported by parents of children using multichannel as opposed to single-channel devices, with little evidence that this was related strongly to length of time using the devices. Finally, in analyses controlling for type of device and length of use, parenting stress as measured in the current study, accounted for substantial proportions of the variance in psychological distress. The implications of these findings, as well as directions for future research, are discussed.


Modified house tympanoplasty for successful closure of near-total tympanic membrane perforations.
To demonstrate the use of a modified House technique in the successful closure of near-total tympanic membrane perforations. This is a retrospective analysis over a 25-year span of the senior author's experience with a particular technique of closing large tympanic membrane perforations. The charts of patients were reviewed and the data were analyzed. The modified House technique combines an underlay graft placement with an advancement ear canal skin flap. It provides vascularization and subsequent early epithelialization of a large perforation. In this series, the senior author operated on 46 patients. Forty-three patients had one ear operated on and three had bilateral surgery. The closure rate was 98% (48 of 49 ears). The one failure had a subsequent successful closure at a later date. This is a simple endaural technique that guarantees successful closure of near-total tympanic membrane perforations using locally available tragal perichondrium.


Effects of topical mitomycin-C on the tracheal epithelia of rabbits following tracheostomy
We aimed to investigate the topical application of mitomycin-C (MMC) after the conventional tracheostomy in a rabbit model. Twenty-four male New Zealand White rabbits were randomly divided among 3 equal groups (n: 8). Trache- ostomies were performed on 16 subjects. Group 1 which served as a control for all tracheal measurements. After tracheostomy, we applied sterile saline (group 2) or MMC at 0.8 mg/mL (group 3) around the tracheotomy site for 5 min. At the 3rd week after surgery, all tracheas were subjected to morphometric and histopathological examinations, including tracheal lumen diameter (LD), number of capillary vessels (CV), subepithelial tissue thickness (SETT), fibroblasts, and inflammatory cells (IC). There was a statistically significant difference between the two tracheostomy groups themselves and the control group for LD (p = 0.035), CV (p = 0.006), SETT, fibroblasts, and IC (p < 0.001). Histopathological analysis showed the decreased LD, CV, SETT, IC, and fibroblasts compared to MMC with tracheostomy groups. MMC was more effective than saline for LD, CV, SETT, IC, and fibroblasts. Wound healing modulation may prevent scar formation. Fibrosis decreased following tracheostomy in the group treated with MMC. Fibroblasts appear to be key cells mediating these effects.


Transcriptome analysis suggests a role for the differential expression of cerebral aquaporins and the MAPK signalling pathway in human temporal lobe epilepsy.
Epilepsies are common disorders of the central nervous system (CNS), affecting up to 2% of the global population. Pharmaco-resistance is a major clinical challenge affecting about 30% of temporal lobe epilepsy (TLE) patients. Water homeostasis has been shown crucial for regulation of neuronal excitability. The control of water movement is achieved through a family of small integral membrane channel proteins called aquaporins (AQPs). Despite the fact that changes in water homeostasis occur in sclerotic hippocampi of people with TLE, the expression of AQPs in the epileptic brain is not fully characterised. This study uses microarray and ELISA methods to analyse the mRNA and protein expression of the human cerebral AQPs in sclerotic hippocampi (TLE-HS) and adjacent neocortex tissue (TLE-NC) of TLE patients. The expression of AQP1 and AQP4 transcripts was significantly increased, while that of the AQP9 transcript was significantly reduced in TLE-HS compared to TLE-NC. AQP4 protein expression was also increased while expression of AQP1 protein remained unchanged, and AQP9 was undetected. Microarray data analysis identified 3333 differentially regulated genes and suggested the involvement of the MAPK signalling pathway in TLE pathogenesis. Proteome array data validated the translational profile for 26 genes and within the MAPK pathway (e.g. p38, JNK) that were identified as differentially expressed from microarray analysis. ELISA data showed that p38 and JNK inhibitors decrease AQP4 protein levels in cultured human primary cortical astrocytes. Elucidating the mechanism of selective regulation of different AQPs and associated regulatory proteins may provide a new therapeutic approach to epilepsy treatment.


Common Patterns of Congenital Lower Extremity Shortening: Diagnosis, Classification, and Follow-up.
Congenital lower limb shortening is a group of relatively rare, heterogeneous disorders. Proximal focal femoral deficiency (PFFD) and fibular hemimelia (FH) are the most common pathologic entities in this disease spectrum. PFFD is characterized by variable degrees of shortening or absence of the femoral head, with associated dysplasia of the acetabulum and femoral shaft. FH ranges from mild hypoplasia to complete absence of the fibula with variable shortening of the tibia. The development of the lower limb requires complex and precise gene interactions. Although the etiologies of PFFD and FH remain unknown, there is a strong association between the two disorders. Associated congenital defects in the lower extremity are found in more than 50% of patients with PFFD, ipsilateral FH being the most common. FH also has a strong association with shortening and bowing of the tibia and with foot deformities such as absence of the lateral rays of the foot. Early diagnosis and radiologic classification of these abnormalities are imperative for appropriate management and surgical planning. Plain radiography remains the main diagnostic imaging modality for both PFFD and FH, and appropriate description of the osseous abnormalities seen on radiographs allows accurate classification, prognostic evaluation, and surgical planning. Minor malformations may commonly be misdiagnosed.


Assessment of antibiotic self-medication practice among public in the northwestern region of Pakistan.
Self-medication with antibiotics is a common practice, which may lead to the development of antimicrobial resistance (AMR)-a major health concern worldwide. The most common reason for the development of AMR is a lack of education and regulatory policies and the lack of community pharmacists. To assess various factors that lead to self-medication with antibiotics, which might cause AMR and hinder effective healthcare. A cross-sectional study was carried out using a predesigned questionnaire to collect data from 800 respondents. The respondents were selected by simple random sampling during November 2014 to January 2015 from different regions of Khyber Pakhtunkhwa (KPK), Pakistan. Only properly completed questionnaires were assessed for different variables. The collected data were analysed using SPSS V.16. 527 people completed and returned the questionnaire-a response rate of 66%. Self-medication with antibiotics was reported by 135 participants (26%), with a higher prevalence of men than women (48% vs 38%, respectively). The main reason for self-medication was previous experience with the same antibiotic (68%). The most commonly used antibiotics were amoxicillin-clavulanate (40%) and major indications for self-medication were sore throat (29%) and flu (24%). Of the 527 respondents, only 104 (20%) were aware of AMR. This study is the first to evaluate self-medication with antibiotics in KPK, Pakistan. In view of the high prevalence of self-medication, introduction of a public health policy through drug regulatory authorities, public awareness programmes/campaigns, patient education about AMR and appropriate use of antibiotics are critically required. The role of community pharmacists needs to be strengthened.