Platelet 5-HT receptor binding during depressive illness and tricyclic antidepressant treatment.
5-Hydroxytryptamine (5-HT) receptor binding was determined in intact human platelets using 125I-iodolysergic acid diethylamide (125I-iodoLSD). The assay revealed that platelets possess a single population of specific binding sites for 125I-iodoLSD which have a high affinity for the 5-HT2 receptor antagonists, ketanserin and spiperone. There was no difference in platelet 125I-iodoLSD receptor binding between depressed patients and controls. However, treatment with tricyclic antidepressants increased the number of 125I-iodoLSD receptor binding sites without significantly altering their affinity.


Novel insights into the function and dynamics of extracellular matrix in liver fibrosis.
Emerging evidence suggests that altered components and posttranslational modifications of proteins in the extracellular matrix (ECM) may both initiate and drive disease progression. The ECM is a complex grid consisting of multiple proteins, most of which play a vital role in containing the essential information needed for maintenance of a sophisticated structure anchoring the cells and sustaining normal function of tissues. Therefore, the matrix itself may be considered as a paracrine/endocrine entity, with more complex functions than previously appreciated. The aims of this review are to 1) explore key structural and functional components of the ECM as exemplified by monogenetic disorders leading to severe pathologies, 2) discuss selected pathological posttranslational modifications of ECM proteins resulting in altered functional (signaling) properties from the original structural proteins, and 3) discuss how these findings support the novel concept that an increasing number of components of the ECM harbor signaling functions that can modulate fibrotic liver disease. The ECM entails functions in addition to anchoring cells and modulating their migratory behavior. Key ECM components and their posttranslational modifications often harbor multiple domains with different signaling potential, in particular when modified during inflammation or wound healing. This signaling by the ECM should be considered a paracrine/endocrine function, as it affects cell phenotype, function, fate, and finally tissue homeostasis. These properties should be exploited to establish novel biochemical markers and antifibrotic treatment strategies for liver fibrosis as well as other fibrotic diseases.


Mesenchymal Stem Cells Isolated From Human Gliomas Increase Proliferation and Maintain Stemness of Glioma Stem Cells Through the IL-6/gp130/STAT3 Pathway.
Although mesenchymal stem cells (MSCs) have been implicated as stromal components of several cancers, their ultimate contribution to tumorigenesis and their potential to drive cancer stem cells, particularly in the unique microenvironment of human brain tumors, remain largely undefined. Consequently, using established criteria, we isolated glioma-associated-human MSCs (GA-hMSCs) from fresh human glioma surgical specimens for the first time. We show that these GA-hMSCs are nontumorigenic stromal cells that are phenotypically similar to prototypical bone marrow-MSCs. Low-passage genomic sequencing analyses comparing GA-hMSCs with matched tumor-initiating glioma stem cells (GSCs) suggest that most GA-hMSCs (60%) are normal cells recruited to the tumor (group 1 GA-hMSCs), although, rarely (10%), GA-hMSCs may differentiate directly from GSCs (group 2 GA-hMSCs) or display genetic patterns intermediate between these groups (group 3 GA-hMSCs). Importantly, GA-hMSCs increase proliferation and self-renewal of GSCs in vitro and enhance GSC tumorigenicity and mesenchymal features in vivo, confirming their functional significance within the GSC niche. These effects are mediated by GA-hMSC-secreted interleukin-6, which activates STAT3 in GSCs. Our results establish GA-hMSCs as a potentially new stromal component of gliomas that drives the aggressiveness of GSCs, and point to GA-hMSCs as a novel therapeutic target within gliomas.


Swimming motility, a virulence trait of Ralstonia solanacearum, is regulated by FlhDC and the plant host environment.
Swimming motility allows the bacterial wilt pathogen Ralstonia solanacearum to efficiently invade and colonize host plants. However, the bacteria are essentially nonmotile once inside plant xylem vessels. To determine how and when motility genes are expressed, we cloned and mutated flhDC, which encodes a major regulator of flagellar biosynthesis and bacterial motility. An flhDC mutant was nonmotile and less virulent than its wild-type parent on both tomato and Arabidopsis; on Arabidopsis, the flhDC mutant also was less virulent than a nonmotile fliC flagellin mutant. Genes in the R. solanacearum motility regulon had strikingly different expression patterns in culture and in the plant. In culture, as expected, flhDC expression depended on PehSR, a regulator of early virulence factors; and, in turn, FlhDC was required for fliC (flagellin) expression. However, when bacteria grew in tomato plants, flhDC was expressed in both wild-type and pehR mutant backgrounds, although PehSR is necessary for motility both in culture and in planta. Both flhDC and pehSR were significantly induced in planta relative to expression levels in culture. Unexpectedly, the fliC gene was expressed in planta at cell densities where motile bacteria were not observed, as well as in a nonmotile flhDC mutant. Thus, expression of flhDC and flagellin itself are uncoupled from bacterial motility in the host environment, indicating that additional signals and regulatory circuits repress motility during plant pathogenesis.


Not your regular high: cardiac dysrhythmias caused by loperamide.
Loperamide, a non-prescription anti-diarrheal agent, is a peripheral mu-opioid receptor agonist that is excluded from the blood-brain barrier by p-glycoprotein at therapeutic doses. Overdoses of loperamide penetrate the central nervous system (CNS), leading to abuse. We report cardiac conduction abnormalities and dysrhythmias after ingestion of a recreational supra-therapeutic dose of loperamide confirmed with an elevated blood loperamide concentration. A 48-year-old woman with a history of alcohol and benzodiazepine abuse presented to the emergency department (ED) with somnolence, weakness and slurred speech. She was taking 20 to 40 tablets of 2 mg loperamide 1-2 times/day for weeks along with clonazepam and whiskey. Vital signs were: blood pressure (BP), 124/90 mmHg; heart rate (HR), 88/min; respiratory rate(RR), 20/min; T, 36.9 °C; O2 saturation 100% on room air (RA). Glucose was 6.4 mmol/L. Electrocardiogram (ECG) had a ventricular rate of 58/min, QRS 164 ms, QT 582 ms with no discernable p-waves. Lactate was 3.5 mmol/L and potassium was 6.2 mEq/L. Labs were notable for an anion gap of 20 mEq/L, ethanol of 3.9 mmol/L, creatinine of 2.3 mg/dL and loperamide concentration of 210 ng/mL (average therapeutic plasma concentration 1.2 ng/mL). She became hypotensive, but responded to fluids. Following treatment for hyperkalemia with calcium, insulin, dextrose, and hypertonic sodium bicarbonate a repeat ECG had a ventricular rate of 66/min, QRS 156 ms, and QT 576 ms. Magnesium was given and pacer pads were placed. During the infusion of magnesium, her BP fell to 92/58 mmHg with a HR of 54/min, RR 14/min, O2 saturation of 97% on RA so the infusion was stopped. The ECG after the magnesium infusion had a ventricular rate of 51/min, QRS of 134 ms, and QT 614 ms. In the ICU she had multiple runs of non-sustained ventricular tachycardia that did not require therapy. Over the next 48 h she improved and was transferred to a floor bed. On day four of hospitalization the patient left against medical advice. At that time, her ECG showed sinus tachycardia with a heart rate 114/min, QRS 82 ms, QT 334 ms. Loperamide produces both QRS and QT prolongation at supra-therapeutic dosing. A blood loperamide concentration of 210 ng/mL is among the highest concentrations reported. Supra-therapeutic dosing of loperamide is promoted on multiple drug-use websites and online forums as a treatment for opioid withdrawal, as well as for euphoric effects. With the current epidemic of prescription opioid abuse, toxicity related to loperamide, an opioid agonist that is readily available without a prescription is occurring more frequently. It is important for clinicians to be aware of the potentially life-threatening toxicity related to loperamide abuse in order to provide proper diagnosis, management and patient education.


Comparative studies on the certification of reference materials by ICPMS and TIMS using isotope dilution procedures.
A comparison of different isotope dilution mass spectrometric (IDMS) procedures using inductively coupled plasma mass spectrometry (ICPMS) and thermal ionization mass spectrometry (TIMS) was carried out to examine the degree of equivalence between the used procedures in terms of requirements for reference material certification. The comparison was based on the measurement results and their uncertainties. The sample used in this study is a pure zinc metal to be certified by the Bureau Communie de Référence (BCR) for amount contents of different trace elements. This study focuses on cadmium and thallium. The TIMS values contributed to the certified values. To guarantee identical conditions as far as possible for the procedures under investigation, the samples were split into subsamples after spiking and digestion took place. Thus, every IDMS procedure started with an identical set of samples. In total, four different IDMS procedures and one external calibration procedure using internal standardization as an example of routine analysis were applied. The IDMS procedures divide in a group with and a group without trace/matrix separation. Multicollector TIMS (TI-MC-MS) and multicollector ICPMS (ICP-MC-MS) were used in combination with trace/matrix separation, whereas quadrupole ICPMS (ICP-QMS) and ICP-MC-MS were also applied to nonseparated samples. All IDMS results agree well within their combined uncertainties, while some results from the external calibration procedure do not. IDMS results obtained by ICPMS without separation are comparable to those obtained by TI-MC-MS with separation regarding precision and accuracy. The smallest uncertainties were achieved using ICP-MC-MS in combination with trace/matrix separation.


Virion-associated restriction endonucleases of chloroviruses.
Chloroviruses are large, double-stranded-DNA, plaque-forming viruses that infect certain eukaryotic chlorella-like green algae. The prototype of the genus is Paramecium bursaria chlorella virus 1 (PBCV-1). Chlorovirus genomes contain various amounts of methylated nucleotides due to virus-encoded DNA methyltransferases (MTases); about 25% of the MTases are associated with companion DNA site-specific (restriction) endonucleases (REases). These enzymes constitute virally encoded restriction-modification (R/M) systems. Although several of the chlorovirus R/M systems are characterized, their biological functions are unknown. The PBCV-1 proteome reveals that two virus-encoded REases, but not their companion MTases, are virion associated, suggesting that viral REases might help degrade the host DNA early in infection. To test this hypothesis, host chromosomal DNA from PBCV-1-infected cells was examined by pulsed-field gel electrophoresis. Initiation of host chromosomal DNA degradation occurred within 5 min postinfection (p.i.). The DNA degradation was insensitive to protein synthesis inhibitors or UV inactivation of virus particles, consistent with the agent being a small protein associated with the virion. Nuclease activities, including those of the two predicted REases and an uncharacterized general nuclease(s), were detected in disrupted PBCV-1 particles. The general nuclease(s) degraded both host and viral DNAs in vitro, although the viral DNA was not degraded in vivo, suggesting differential intracellular trafficking of the virion-associated nucleases. Infection with chloroviruses lacking an R/M system(s) resulted in either delayed host chromosomal DNA degradation or no detectable host chromatin changes. These immediate-early events associated with chlorovirus infections may facilitate rapid switching of the host transcriptional apparatus to viral transcription, which begins within 5 to 10 min p.i.


Application of clustering global optimization to thin film design problems.
Refinement techniques usually calculate an optimized local solution, which is strongly dependent on the initial formula used for the thin film design. In the present study, a clustering global optimization method is used which can iteratively change this initial formula, thereby progressing further than in the case of local optimization techniques. A wide panel of local solutions is found using this procedure, resulting in a large range of optical thicknesses. The efficiency of this technique is illustrated by two thin film design problems, in particular an infrared antireflection coating, and a solar-selective absorber coating.


Live cell imaging reveals actin-cytoskeleton-induced self-association of the actin-bundling protein WLIM1.
Crosslinking of actin filaments into bundles is essential for the assembly and stabilization of specific cytoskeletal structures. However, relatively little is known about the molecular mechanisms underlying actin bundle formation. The two LIM-domain-containing proteins define a novel and evolutionarily conserved family of actin-bundling proteins whose actin-binding and -crosslinking activities primarily rely on their LIM domains. Using TIRF microscopy, we describe real-time formation of actin bundles induced by tobacco NtWLIM1 in vitro. We show that NtWLIM1 binds to single filaments and subsequently promotes their interaction and zippering into tight bundles of mixed polarity. NtWLIM1-induced bundles grew by both elongation of internal filaments and addition of preformed fragments at their extremities. Importantly, these data are highly consistent with the modes of bundle formation and growth observed in transgenic Arabidopsis plants expressing a GFP-fused Arabidopsis AtWLIM1 protein. Using two complementary live cell imaging approaches, a close relationship between NtWLIM1 subcellular localization and self-association was established. Indeed, both BiFC and FLIM-FRET data revealed that, although unstable NtWLIM1 complexes can sporadically form in the cytosol, stable complexes concentrate along the actin cytoskeleton. Remarkably, disruption of the actin cytoskeleton significantly impaired self-association of NtWLIM1. In addition, biochemical analyses support the idea that F-actin facilitates the switch of purified recombinant NtWLIM1 from a monomeric to a di- or oligomeric state. On the basis of our data, we propose a model in which actin binding promotes the formation and stabilization of NtWLIM1 complexes, which in turn might drive the crosslinking of actin filaments.


Point mutations in the juxtamembrane domain of FLT3 define a new class of activating mutations in AML.
In acute myeloid leukemia (AML), two clusters of activating mutations are known in the FMS-like tyrosine kinase-3 (FLT3) gene: FLT3-internal tandem duplications (FLT3-ITDs) in the juxtamembrane (JM) domain in 20% to 25% of patients, and FLT3 point mutations in the tyrosine-kinase domain (FLT3-TKD) in 7% to 10% of patients, respectively. Here, we have characterized a new class of activating point mutations (PMs) that cluster in a 16-amino acid stretch of the juxtamembrane domain of FLT3 (FLT3-JM-PMs). Expression of 4 FLT3-JM-PMs in interleukin-3 (IL-3)-dependent Ba/F3 cells led to factor-independent growth, hyperresponsiveness to FLT3 ligand, and resistance to apoptotic cell death. FLT3-JM-PM receptors were autophosphorylated and showed a higher constitutive dimerization rate compared with the FLT3-wild-type (WT) receptor. As a molecular mechanism, we could show activation of STAT5 and up-regulation of Bcl-x(L) by all FLT3-JM-PMs. The FLT3 inhibitor PKC412 abrogated the factor-independent growth of FLT3-JM-PM-expressing cells. Compared with FLT3-ITD and FLT3-TKD mutants, the FLT3-JM-PMs showed a weaker transforming potential related to lower autophosphorylation of the receptor and its downstream target STAT5. Mapping of the FLT3-JM-PMs on the crystal structure of FLT3 showed that these mutations reduce the stability of the autoinhibitory JM domain, and provides a structural basis for the transforming capacity of this new class of gain-of-function mutations of FLT3.


Embryonic organizer specification in the mud snail Ilyanassa obsoleta depends on intercellular signaling.
In early embryos of the caenogastropod snail Ilyanassa obsoleta, cytoplasmic segregation of a polar lobe is required for establishment of the D quadrant founder cell, empowering its great-granddaughter macromere 3D to act as a single-celled organizer that induces ectodermal pattern along the secondary body axis of the embryo. We present evidence that polar lobe inheritance is not sufficient to specify 3D potential, but rather makes the D macromere lineage responsive to some intercellular signal(s) required for normal expression of 3D-specific phenotypes. Experimental removal of multiple micromeres resulted in loss of organizer-linked MAPK activation, complete and specific defects of organizer-dependent larval organs, and progressive cell cycle retardation, leading to equalization of the normally accelerated division schedule of 3D (relative to the third-order macromeres of the A, B and C quadrants). Ablation of the second-quartet micromere 2d greatly potentiated the effects of first micromere quartet ablation. Our findings link organizer activation in I. obsoleta to the putative ancestral spiralian mechanism in which a signal from micromeres leads to specification of 3D among four initially equivalent macromeres.


Overhauser effects in insulating solids.
We report magic angle spinning, dynamic nuclear polarization (DNP) experiments at magnetic fields of 9.4 T, 14.1 T, and 18.8 T using the narrow line polarizing agents 1,3-bisdiphenylene-2-phenylallyl (BDPA) dispersed in polystyrene, and sulfonated-BDPA (SA-BDPA) and trityl OX063 in glassy glycerol/water matrices. The (1)H DNP enhancement field profiles of the BDPA radicals exhibit a significant DNP Overhauser effect (OE) as well as a solid effect (SE) despite the fact that these samples are insulating solids. In contrast, trityl exhibits only a SE enhancement. Data suggest that the appearance of the OE is due to rather strong electron-nuclear hyperfine couplings present in BDPA and SA-BDPA, which are absent in trityl and perdeuterated BDPA (d21-BDPA). In addition, and in contrast to other DNP mechanisms such as the solid effect or cross effect, the experimental data suggest that the OE in non-conducting solids scales favorably with magnetic field, increasing in magnitude in going from 5 T, to 9.4 T, to 14.1 T, and to 18.8 T. Simulations using a model two spin system consisting of an electron hyperfine coupled to a (1)H reproduce the essential features of the field profiles and indicate that the OE in these samples originates from the zero and double quantum cross relaxation induced by fluctuating hyperfine interactions between the intramolecular delocalized unpaired electrons and their neighboring nuclei, and that the size of these hyperfine couplings is crucial to the magnitude of the enhancements. Microwave power dependent studies show that the OE saturates at considerably lower power levels than the solid effect in the same samples. Our results provide new insights into the mechanism of the Overhauser effect, and also provide a new approach to perform DNP experiments in chemical, biophysical, and physical systems at high magnetic fields.


Effect of California's proposition 187 on the use of primary care clinics.
California's ballot proposition 187, passed by the voters in November 1994, threatened to discontinue undocumented immigrants' eligibility for most health services while mandating that health care professionals report suspected undocumented patients to authorities. Although the proposition has not been put into practice, reports suggest that its passage was associated with a decline in the use of health services by some groups. To assess the effects of the passage of Proposition 187 on the use of primary care services, we surveyed a representative sample of California clinics serving low-income groups (n = 129). Using a mailed questionnaire and phone interviews with clinic directors, we obtained qualitative and quantitative data regarding the effects of the passage of the proposition on clinic use. Among primary care clinics statewide and clinics serving predominantly Latino patients, we detected no significant decline in total monthly visits following the election. Nevertheless, half of clinic directors (51%) thought that the number of clinic visits declined after the passage of Proposition 187, and many directors thought that the deterrent effects of the election persisted for weeks to months after the election. Whereas the number of visits probably declined at some clinics, only a small minority of patients at most primary care clinics could have been deterred from seeking care after the passage of Proposition 187.


Ability and Disability in Autism Spectrum Disorder: A Systematic Literature Review Employing the International Classification of Functioning, Disability and Health-Children and Youth Version.
This study is the first in a series of four empirical investigations to develop International Classification of Functioning, Disability and Health (ICF) Core Sets for Autism Spectrum Disorder (ASD). The objective was to use a systematic review approach to identify, number, and link functional ability and disability concepts used in the scientific ASD literature to the nomenclature of the ICF-CY (Children and Youth version of the ICF, covering the life span). Systematic searches on outcome studies of ASD were carried out in Medline/PubMed, PsycINFO, ERIC and Cinahl, and relevant functional ability and disability concepts extracted from the included studies. These concepts were then linked to the ICF-CY by two independent researchers using a standardized linking procedure. New concepts were extracted from the studies until saturation of identified ICF-CY categories was reached. Seventy-one studies were included in the final analysis and 2475 meaningful concepts contained in these studies were linked to 146 ICF-CY categories. Of these, 99 categories were considered most relevant to ASD (i.e., identified in at least 5% of the studies), of which 63 were related to Activities and Participation, 28 were related to Body functions, and 8 were related to Environmental factors. The five most frequently identified categories were basic interpersonal interactions (51%), emotional functions (49%), complex interpersonal interactions (48%), attention functions (44%), and mental functions of language (44%). The broad variety of ICF-CY categories identified in this study reflects the heterogeneity of functional differences found in ASD--both with respect to disability and exceptionality--and underlines the potential value of the ICF-CY as a framework to capture an individual's functioning in all dimensions of life. The current results in combination with three additional preparatory studies (expert survey, focus groups, and clinical study) will provide the scientific basis for defining the ICF Core Sets for ASD for multipurpose use in basic and applied research and every day clinical practice of ASD.


Aerobic catalytic systems inspired by copper amine oxidases: recent developments and synthetic applications.
Recent efforts to design synthetic quinone-based catalysts for the efficient aerobic oxidation of amines to imines have been inspired by copper amine oxidases (CuAOs), a family of metalloenzymes which selectively converts primary amines into aldehydes, using molecular oxygen through the cooperation of a quinone-based cofactor, 2,4,5-trihydroxyphenylalanine quinone (TPQ) and a copper ion. Two distinct classes of bioinspired quinone-based catalytic systems have been developed. The first class consists of catalytic systems, which mimic the activity of CuAOs by exhibiting exquisite selectivity for primary amines. The second class consists of catalytic systems, which allow the expansion of the substrate scope to the oxidation of α-branched primary amines and secondary amines including nitrogen heterocycles, two reaction types that natural CuAOs are not able to accomplish. These catalytic oxidative green processes can be applied to the C-H functionalization of primary amines and to the synthesis of several nitrogen-containing heterocycles.


Internal and external validity of attention-deficit hyperactivity disorder in a population-based sample of adults.
Follow-up studies of childhood ADHD have shown persistence of the disorder into adulthood, but no epidemiological data are yet available. ADHD DSM-IV symptoms were obtained by self-report in an adult population-based sample of 1813 adults (aged 18-75 years), that was drawn from an automated general practitioner system used in Nijmegen, The Netherlands. The structure of ADHD symptoms was analysed by means of confirmatory factor analyses. Other data used in this report are the General Health Questionnaire (GHQ-28), information about the presence of three core symptoms of ADHD in childhood, and about current psychosocial impairment. The three-factor model that allowed for cross-loadings provided the best fit in the entire sample. This result was replicated across gender and age subsamples. Inattentive and hyperactivity symptom scores were significantly associated with measures of impairment, even after controlling for the GHQ-28. Subjects with four or more inattentive or hyperactive-impulsive symptoms were significantly more impaired than subjects with two, one and no symptoms. The prevalence of ADHD in adults was 1.0% (95% CI 0.6-1.6) and 2.5% (1.9-3.4) using a cutoff of six and four current symptoms respectively, and requiring the presence of all three core symptoms in childhood. These results support the internal and external validity of ADHD in adults between 18 and 75 years. ADHD is not merely a child psychiatric disorder that persists into young adulthood, but an important and unique manifestation of psychopathology across the lifespan.


Altered hippocampal expression of glutamate receptors and transporters in GRM2 and GRM3 knockout mice.
Group II metabotropic glutamate receptors (mGluR2 and mGluR3, also called mGlu2 and mGlu3, encoded by GRM2 and GRM3, respectively) are therapeutic targets for several psychiatric disorders. GRM3 may also be a schizophrenia susceptibility gene. mGluR2-/- and mGluR3-/- mice provide the only unequivocal means to differentiate between these receptors, yet interpretation of in vivo findings may be complicated by secondary effects on expression of other genes. To address this issue, we examined the expression of NMDA receptor subunits (NR1, NR2A, NR2B) and glutamate transporters (EAAT1-3), as well as the remaining group II mGluR, in the hippocampus of mGluR2-/- and mGluR3-/- mice, compared with wild-type controls. mGluR2 mRNA was increased in mGluR3-/- mice, and vice versa. NR2A mRNA was increased in both knockout mice. EAAT1 (GLAST) mRNA and protein, and EAAT2 (GLT-1) protein, were reduced in mGluR3-/- mice, whereas EAAT3 (EAAC1) mRNA was decreased in mGluR2-/- mice. Transcripts for NR1 and NR2B were unchanged. The findings show a compensatory upregulation of the remaining group II metabotropic glutamate receptor in the knockout mice. Upregulation of NR2A expression suggests modified NMDA receptor signaling in mGluR2-/- and mGluR3-/- mice, and downregulation of glutamate transporter expression suggests a response to altered synaptic glutamate levels. The results show a mutual interplay between mGluR2 and mGluR3, and also provide a context in which to interpret behavioral and electrophysiological results in these mice.


Vascular supply of the distally based superficial sural artery flap: surgical safe zones based on component analysis using three-dimensional computed tomographic angiography.
Distal lower limb reconstruction remains challenging for surgeons. Since its description, the distally based superficial sural artery flap (sural flap) has gained popularity for these indications. The authors' aim was to determine which components were necessary to maintain this flap's arterial supply. Anatomical components were studied on 24 fresh adult cadaver legs and included the following: skin, adipose tissue, lesser saphenous vein, deep fascia, and sural nerve. Thirty-two flaps were harvested and divided into the following combination groups: cutaneous-venoneuroadipofascial (n = 5), venoneuroadipofascial (n = 3), neurofascial (n = 4), cutaneous-adipovenous (n = 2), cutaneous-venoadipofascial (n = 9), venoadipofascial (n = 5), and purely fascial (n = 4). Leg length, location of the sural nerve crossing the deep fascia, and location of peroneal and posterior tibial artery perforators were recorded. Twenty-eight of the combination flaps were injected with barium sulfate. Three-dimensional computed tomographic angiography was used to analyze the vascular territory of each flap. The cutaneous-venoneuroadipofascial and cutaneous-venoadipofascial flaps were perfused 86.5 percent and 80.2 percent, respectively, followed by cutaneous-adipovenous (75.7 percent), venoneuroadipofascial (87.1 percent), and venoadipofascial (74.8 percent) flaps. In contrast, the neurofascial and purely fascial flaps were merely perfused 40.8 percent and 44.1 percent, respectively, using only a perineural vascular network, with minimal fascial contribution. The lesser saphenous vein and the deep adipose tissue are necessary for the arterial supply of the sural flap. The deep fascia serves only as mechanical support. The sural nerve contributes to the vascular network, but its inclusion does not increase the vascular territory. Noninclusion of the sural nerve defines a new pivot point that should be identified before the harvest of the sural flap.


Do Interventions that Promote Awareness of Rights Increase Use of Maternity Care Services? A Systematic Review.
Twenty years after the rights of women to go through pregnancy and childbirth safely were recognized by governments, we assessed the effects of interventions that promote awareness of these rights to increase use of maternity care services. Using inclusion and exclusion criteria defined in a peer-reviewed protocol, we searched published and grey literature from one database of studies on maternal health, two search engines, an internet search and contact with experts. From the 707 unique documents found, 219 made reference to rights, with 22 detailing interventions promoting awareness of rights for maternal and newborn health. Only four of these evaluated effects on health outcomes. While all four interventions promoted awareness of rights, they did so in different ways. Interventions included highly-scripted dissemination meetings with educational materials and other visual aids, participatory approaches that combined raising awareness of rights with improving accountability of services, and broader multi-stakeholder efforts to improve maternal health. Study quality ranged from weak to strong. Measured health outcomes included increased antenatal care and facility birth. Improvements in human rights outcomes such as availability, acceptability, accessibility, quality of care, as well as the capacity of rights holders and duty bearers were also reported to varying extents. Very little information on costs and almost no information on harms or risks were described. Despite searching multiple sources of information, while some studies did report on activities to raise awareness of rights, few detailed how they did so and very few measured effects on health outcomes. Promoting awareness of rights is one element of increasing demand for and use of quality maternity care services for women during pregnancy, birth and after birth. To date efforts have not been well documented in the literature and the program theories, processes and costs, let alone health effects have not been well evaluated.


Optimizing time and resource allocation trade-offs for investment into morphological and behavioral defense.
Prey organisms are confronted with time and resource allocation trade-offs. Time allocation trade-offs partition time, for example, between foraging effort to acquire resources and behavioral defense. Resource allocation trade-offs partition the acquired resources between multiple traits, such as growth or morphological defense. We develop a mathematical model for prey organisms that comprise time and resource allocation trade-offs for multiple defense traits. Fitness is determined by growth and survival during ontogeny. We determine optimal defense strategies for environments that differ in their resource abundance, predation risk, and defense effectiveness. We compare the results with results of simplified models where single defense traits are optimized. Our results indicate that selection acts in favor of integrated traits. The selective advantage of expressing multiple defense traits is most pronounced at intermediate environmental conditions. Optimizing single traits generally leads to a more pronounced response of the defense traits, which implies that studying single traits leads to an overestimation of their response to predation. Behavioral defense and morphological defense compensate for and augment each other depending on predator densities and the effectiveness of the defense mechanisms. In the presence of time constraints, the model shows peak investment into morphological and behavioral defense at intermediate resource levels.


Evidence for a hyporesponsive limbic-hypothalamic-pituitary-adrenal axis following early-life repetitive hypoglycemia in adult male rats.
The developing limbic-hypothalamic-pituitary-adrenal (LHPA) axis is highly vulnerable to programming by early-life environmental factors, including exposure to synthetic glucocorticoids and nutrient deficiencies. Early-life repetitive hypoglycemia (RHG) is a common complication of insulin therapy for type-1 diabetes that may have long-term consequences in adulthood. Recent observations in a rat model of early RHG suggest persistent changes in LHPA axis function, including changes in relevant hormones and affective behaviors, which support a hyperresponsive LHPA axis. Thus, we hypothesized that early RHG would alter the expression of key genes regulating LHPA axis function in adulthood. The present study employed a rat model of insulin-induced RHG spanning postnatal days (P)24-28, a neurodevelopmental equivalent of early childhood in humans, to assess the long-term effects on mRNA levels for proteins relevant to the LHPA function and the corticosterone responses to ACTH stimulation of dispersed adrenocortical cells in vitro and restraint stress in vivo at adulthood. This early RHG model resulted in a hyporesponsive LHPA axis characterized by impaired corticosterone response, increased hippocampal glucocorticoid and mineralocorticoid receptor (GR and MR), decreased hypothalamic corticotropin-releasing hormone, increased adrenal steroidogenic-acute-regulatory protein and GR, and decreased adrenal MR, melanocortin-type-2 receptor and low-density lipoprotein receptor expression. Our findings highlight a complex environmental-gene interaction between RHG and LHPA axis during development that influences regulation of this axis in adulthood. The findings are consistent with the developmental origins of disease and underscore the influences of early-life events on the programming of a major regulatory system.


Setting standards for empirical bioethics research: a response to Carter and Cribb.
This paper responds to the commentaries from Stacy Carter and Alan Cribb. We pick up on two main themes in our response. First, we reflect on how the process of setting standards for empirical bioethics research entails drawing boundaries around what research counts as empirical bioethics research, and we discuss whether the standards agreed in the consensus process draw these boundaries correctly. Second, we expand on the discussion in the original paper of the role and significance of the concept of 'integrating' empirical methods and ethical argument as a standard for research practice within empirical bioethics.


Transient conformational remodeling of folding proteins by GroES-individually and in concert with GroEL.
The commonly accepted dogma of the bacterial GroE chaperonin system entails protein folding mediated by cycles of several ATP-dependent sequential steps where GroEL interacts with the folding client protein. In contrast, we herein report GroES-mediated dynamic remodeling (expansion and compression) of two different protein substrates during folding: the endogenous substrate MreB and carbonic anhydrase (HCAII), a well-characterized protein folding model. GroES was also found to influence GroEL binding induced unfolding and compression of the client protein underlining the synergistic activity of both chaperonins, even in the absence of ATP. This previously unidentified activity by GroES should have important implications for understanding the chaperonin mechanism and cellular stress response. Our findings necessitate a revision of the GroEL/ES mechanism.


Nonsteroidal anti-inflammatory drugs and the risk of hospitalization for acute renal failure.
Few studies have evaluated the association between use of nonsteroidal anti-inflammatory drugs (NSAIDs) and the risk of idiopathic acute renal failure (ARF) in the general population. Population-based case-control study among persons in the Canadian province of Saskatchewan who received at least 1 NSAID prescription between January 1, 1982, and December 31, 1986. Health department databases were used for case detection, as the sampling frame for selecting controls (n = 1997), and as the primary source of information on drug use and comorbidity. A total of 306 hospital records were reviewed. Twenty-eight patients who were hospitalized fulfilled the diagnostic criteria for ARF. The incidence rate of hospitalization for ARF among the general population not exposed to NSAIDs was 2 per 100000 person-years. Current exposure to NSAIDs, acetylsalicylic acid and other nephrotoxic drugs, male gender, increasing age, cardiovascular comorbidity, and recent hospitalization for disorders other than renal were found to be independent risk factors for ARF. Current NSAID users had an adjusted odds ratio for ARF of 4.1 (95% confidence interval, 1.5-10.8). The risk of ARF was especially high during the first month of use (odds ratio, 8.5). For prescribed dose, we found that users of high daily doses of NSAIDs experienced an odds ratio of 9.8 for ARF. In the general population, hospitalizations for ARF were found to be a rare condition. The 4-fold increase in risk associated with NSAID use was dose-dependent and occurred especially during the first month of therapy. Concurrent comedication with other potentially nephrotoxic agents should be prescribed with care, especially in the elderly.


Impact energy attenuation performance of football headgear.
Commercially available football head protectors were tested to determine their impact energy attenuation performance and ability to reduce the likelihood of concussion. Prospective study using standardised impact test methods with both rigid (magnesium) and Hybrid III headforms. Eight commercially available head protectors from six manufacturers were tested. The magnitude of the headform accelerations increased as the drop height was increased, ranging from a minimum of 64 g from a height of 0.2 m to a maximum of 1,132 g from a height of 0.6 m. The head injury criterion and maximum headform acceleration values followed a similar trend. A steep increase was noted in the magnitude of maximum headform acceleration and head injury criterion when the drop height was increased from 0.4 to 0.5 m. This indicates that the foam material was completely compressed at an impact energy above about 20 J and therefore offers little protection against impacts of greater severity. Repeated tests using a drop height of 0.3 m showed that some helmets exhibit a "memory" effect, whereby impact performance is reduced by up to 50% with repeated impacts. Laboratory tests indicate that current commercially available football headgear performance will not reduce the likelihood of concussion. The absence of internationally recognised standards for soft headgear designed to ameliorate concussion is a major deficiency in sports injury prevention.


Reinstitutionalization following psychiatric discharge among VA patients with serious mental illness: a national longitudinal study.
Patterns of reinstitutionalization following psychiatric hospitalization for individuals with serious mental illnesses (SMI) vary by medical and psychiatric health care settings. This report presents rates of reinstitutionalization across care settings for 35,527 patients following psychiatric discharge in the Department of Veterans Affairs (VA) health system, a national health care system. Over a 7-year follow-up period, 30,417 patients (86%) were reinstitutionalized. Among these patients, 73% were initially reinstitutionalized to inpatient psychiatric settings. Homelessness, medical morbidity, and substance use were associated with increased risks for reinstitutionalization. Despite the VA's increased emphasis on outpatient services delivery, the vast majority of patients experienced reinstitutionalization in the follow-up period. Study findings may inform efforts to refine psychiatric and medical assessment for service delivery for this vulnerable population.


The 2018 Global Research Expedition on Altitude Related Chronic Health (Global REACH) to Cerro de Pasco, Peru: an Experimental Overview.
What is the central question of this study? Herein, a methodological overview of our research team's (Global REACH) latest high altitude research expedition to Peru is provided. What is the main finding and its importance? The experimental objectives, expedition organization, measurements and key cohort data are discussed. The select data presented in this manuscript demonstrate the haematological differences between lowlanders and Andeans with and without excessive erythrocytosis. The data also demonstrate that exercise capacity was similar between study groups at high altitude. The forthcoming findings from our research expedition will contribute to our understanding of lowlander and indigenous highlander high altitude adaptation. In 2016, the international research team Global Research Expedition on Altitude Related Chronic Health (Global REACH) was established and executed a high altitude research expedition to Nepal. The team consists of ∼45 students, principal investigators and physicians with the common objective of conducting experiments focused on high altitude adaptation in lowlanders and in highlanders with lifelong exposure to high altitude. In 2018, Global REACH travelled to Peru, where we performed a series of experiments in the Andean highlanders. The experimental objectives, organization and characteristics, and key cohort data from Global REACH's latest research expedition are outlined herein. Fifteen major studies are described that aimed to elucidate the physiological differences in high altitude acclimatization between lowlanders (n = 30) and Andean-born highlanders with (n = 22) and without (n = 45) excessive erythrocytosis. After baseline testing in Kelowna, BC, Canada (344 m), Global REACH travelled to Lima, Peru (∼80 m) and then ascended by automobile to Cerro de Pasco, Peru (∼4300 m), where experiments were conducted over 25 days. The core studies focused on elucidating the mechanism(s) governing cerebral and peripheral vascular function, cardiopulmonary regulation, exercise performance and autonomic control. Despite encountering serious logistical challenges, each of the proposed studies was completed at both sea level and high altitude, amounting to ∼780 study sessions and >3000 h of experimental testing. Participant demographics and data relating to acid-base balance and exercise capacity are presented. The collective findings will contribute to our understanding of how lowlanders and Andean highlanders have adapted under high altitude stress.


A simple approach to fairer hospital benchmarking using patient experience data.
To determine the factors contributing most to variability in patient experience in order to present approaches for fairer benchmarking of hospitals and for quality improvement. Secondary analysis of data from a widely used survey on patient experience. Inpatients from all 24 acute hospitals in the Canton of Bern in Switzerland. Data collection followed the standardized and validated Picker Institute methodology for a period of 13 weeks in the fall/winter of 2005. Inpatients age 18 years and older (n = 14 089), discharged within the sampling period. 'Patient experience', measured by the total Picker Problem Score (PPS) and by six domain scores (care, communication, respect, cooperation, organization, discharge management). In regression analysis, the patient factors self-reported health, age and education explained the highest proportion of variability in the PPS (4.8, 2.2 and 0.7%, respectively). Multiple linear adjustment for factors associated with patients removed between 29 and 33% of variability between hospital categories. The domain score means varied from under 5% for 'respect towards the patient' to 34% for 'discharge management'. Ranking of hospitals by domain scores differed from the ranking based on the total PPS. Statistical adjustment for patient mix and additional stratification for some hospital factors make benchmarking using patient survey data fairer and more transparent. Use of our approach for presenting quality data may make interpretation easier for the different target groups and may enhance the relevance of such information for decision-making.


Using G6PD tests to enable the safe treatment of Plasmodium vivax infections with primaquine on the Thailand-Myanmar border: A cost-effectiveness analysis.
Primaquine is the only licensed antimalarial for the radical cure of Plasmodium vivax infections. Many countries, however, do not administer primaquine due to fear of hemolysis in those with glucose-6-phosphate dehydrogenase (G6PD) deficiency. In other settings, primaquine is given without G6PD testing, putting patients at risk of hemolysis. New rapid diagnostic tests (RDTs) offer the opportunity to screen for G6PD deficiency prior to treatment with primaquine. Here we assessed the cost-effectiveness of using G6PD RDTs on the Thailand-Myanmar border and provide the model as an online tool for use in other settings. Decision tree models for the management of P. vivax malaria evaluated the costs and disability-adjusted life-years (DALYs) associated with recurrences and primaquine-induced hemolysis from a health care provider perspective. Screening with G6PD RDTs before primaquine use was compared to (1) giving chloroquine alone and (2) giving primaquine without screening. Data were taken from a recent study on the impact of primaquine on P. vivax recurrences and a literature review. Compared to the use of chloroquine alone, the screening strategy had similar costs while averting 0.026 and 0.024 DALYs per primary infection in males and females respectively. Compared to primaquine administered without screening, the screening strategy provided modest cost savings while averting 0.011 and 0.004 DALYs in males and females respectively. The probabilistic sensitivity analyses resulted in a greater than 75% certainty that the screening strategy was cost-effective at a willingness to pay threshold of US$500, which is well below the common benchmark of per capita gross domestic product for Myanmar. In this setting G6PD RDTs could avert DALYs by reducing recurrences and reducing hemolytic risk in G6PD deficient patients at low costs or cost savings. The model results are limited by the paucity of data available in the literature for some parameter values, including the mortality rates for both primaquine-induced hemolysis and P. vivax. The online model provides an opportunity to use different parameter estimates to examine the validity of these findings in other settings.


Traumatic retinal detachment.
Seventy-seven patients developed retinal breaks following an episode of ocular contusion, and 65 (84.4%) of these developed rhegmatogenous retinal detachment. Surgical treatment successfully restored or maintained retinal apposition in 74 (96.1%) of the eyes. Thirty-six (46.8%) eyes recovered visual acuity of 6/9 or better. Of the retinal breaks recognised dialysis at the ora serrata was observed in 49 eyes, of which 28 were situated at the lower temporal quadrant. Seventeen eyes had irregular breaks arising within necrotic retina at the site of scleral impact. Twenty-four (31.2%) patients had retinal break or retinal detachment diagnosed within 24 hours of injury and 49 (63.6%) within six weeks. Immediate retinal detachment was a feature of necrotic retinal breaks, while inferior oral dialyses led to a slow accumulation of subretinal fluid. Delayed diagnosis of retinal detachment was due either to opaque media or to failure to examine the retina after injury. Visual prognosis was good when retinal break or detachment were diagnosed within six weeks of injury. However, those patients who escaped initial retinal examination and were lost to follow-up had a less favourable visual outcome.


Crystallographic and biochemical investigations of kumamolisin-As, a serine-carboxyl peptidase with collagenase activity.
Kumamolisin-As (previously called ScpA) is the first known example of a collagenase from the sedolisin family (MEROPS S53). This enzyme is active at low pH and in elevated temperatures. In this study that used x-ray crystallographic and biochemical methods, we investigated the structural basis of the preference of this enzyme for collagen and the importance of a glutamate residue in the unique catalytic triad (Ser(278)-Glu(78)-Asp(82)) for enzymatic activity. Crystal structures of the uninhibited enzyme and its complex with a covalently bound inhibitor, N-acetyl-isoleucyl-prolyl-phenylalaninal, showed the occurrence of a narrow S2 pocket and a groove that encompasses the active site and is rich in negative charges. Limited endoproteolysis studies of bovine type-I collagen as well as kinetic studies using peptide libraries randomized at P1 and P1', showed very strong preference for arginine at the P1 position, which correlated very well with the presence of a negatively charged residue in the S1 pocket of the enzyme. All of these features, together with those predicted through comparisons with fiddler crab collagenase, a serine peptidase, rationalize the enzyme's preference for collagen. A comparison of the Arrhenius plots of the activities of kumamolisin-As with either collagen or peptides as substrates suggests that collagen should be relaxed before proteolysis can occur. The E78H mutant, in which the catalytic triad was engineered to resemble that of subtilisin, showed only 0.01% activity of the wild-type enzyme, and its structure revealed that Ser(278), His(78), and Asp(82) do not interact with each other; thus, the canonical catalytic triad is disrupted.


Development of clinical immunity to malaria in highland areas of low and unstable transmission.
In highland areas of unstable, low malaria transmission, the extent to which immunity to uncomplicated malaria develops with age and intermittent parasite exposure has not been well characterized. We conducted active surveillance for clinical malaria during April 2003-March 2005 in two highland areas of western Kenya (Kapsisiywa and Kipsamoite). In both sites, annual malaria incidence was significantly lower in persons ≥ 15 years of age than in persons < 5 years of age (Kapsisiywa: incidence = 382.9 cases/1,000 persons among persons < 1-4 years of age versus 135.1 cases/1,000 persons among persons ≥ 15 years of age; Kipsamoite: incidence = 233.0 cases/1,000 persons in persons < 1-4 years of age versus 43.3 cases/1,000 persons in persons ≥ 15 years of age). In Kapsisiywa, among persons with malaria, parasite density and axillary body temperature were also significantly lower in persons ≥ 15 years of age than in persons < 5 years of age. Even in highland areas of unstable and low malaria transmission, age is associated with development of clinical immunity to malaria.


Recognizing an object from the sum of its parts: an intracranial study on alpha rhythms.
Little is known about the relation of alpha rhythms and object recognition. Alpha has been generally proposed to be associated with attention and memory and to be particularly important for the mediation of long-distance communication between neuronal populations. However, how these apply to object recognition is still unclear. This study aimed at describing the spatiotemporal dynamics of alpha rhythms while recognizing fragmented images of objects presented for the first time and presented again 24 hr later. Intracranial electroencephalography was performed in six epileptic patients undergoing presurgical evaluation. Time-frequency analysis revealed a strong alpha activity, mainly of the evoked type, propagating from posterior cerebral areas to anterior regions, which was similar whether the objects were recognized or not. Phase coherence analysis, however, showed clear phase synchronization specific for the moment of recognition. Twenty-four hr later, frontal regions displayed stronger alpha activity and more distributed phase synchronization than when images were presented for the first time. In conclusion, alpha amplitude seems to be related to nonspecific mechanism. Phase coherence analysis suggests a communicational role of alpha activity in object recognition, which may be important for the comparison between bottom-up representations and memory templates.


An Audit of UK Hospital Doctors' Knowledge and Experience of Myalgic Encephalomyelitis.
Background and Objectives: There is some evidence that knowledge and understanding of ME among doctors is limited. Consequently, an audit study was carried out on a group of hospital doctors attending a training event to establish how much they knew about ME and their attitudes towards it. Materials and Methods: Participants at the training event were asked to complete a questionnaire, enquiring about prior knowledge and experience of ME and their approaches to diagnosis and treatment. A total of 44 completed questionnaires were returned. Responses were tabulated, proportions selecting available options determined, 95% confidence limits calculated, and the significance of associations determined by Fisher's exact test. Results: Few respondents had any formal teaching on ME, though most had some experience of it. Few knew how to diagnose it and most lacked confidence in managing it. None of the respondents who had had teaching or prior experience of ME considered it a purely physical illness. Overall, 82% of participants believed ME was at least in part psychological. Most participants responded correctly to a series of propositions about the general epidemiology and chronicity of ME. There was little knowledge of definitions of ME, diagnosis, or of clinical manifestations. Understanding about appropriate management was very deficient. Similarly, there was little appreciation of the impact of the disease on daily living or quality of life. Where some doctors expressed confidence diagnosing or managing ME, this was misplaced as they were incorrect on the nature of ME, its diagnostic criteria and its treatment. Conclusion: This audit demonstrates that most doctors lack training and clinical expertise in ME. Nevertheless, participants recognised a need for further training and indicated a wish to participate in this. It is strongly recommended that factually correct and up-to-date medical education on ME be made a priority at undergraduate and postgraduate levels. It is also recommended that this audit be repeated following a period of medical education.


High leptin levels are associated with stroke.
Leptin, an important hormone for body weight regulation, may be involved in the pathogenesis of cardiovascular manifestations of obesity. We tested whether leptin may be an independent risk marker for stroke in a case-referent study. Definitive acute stroke events, defined by MONICA criteria, were identified from October 1, 1995 to April 30, 1999. Referents without known cardiovascular disease were randomly selected from a population census. Patient characteristics were taken from hospital files and leptin was analyzed in stored samples. Logistic regression analysis was used to determine possible differences in leptin levels between groups. One hundred and thirty-seven cases with ischemic stroke and 69 cases with hemorrhagic stroke were identified. In comparison with referents, male patients with stroke had significantly higher leptin levels. Both male and female stroke patients had increased blood pressure compared with the referents. In multivariate analyses, high leptin levels were associated with both ischemic (OR = 4.89; 95% CI: 1.89-12.62) and hemorrhagic (OR = 3.86; 95% CI: 1.13-13.16) stroke in men, and with ischemic stroke in women (OR = 4.10; 95% CI: 1.45-11.62). The combination of high leptin levels and increased blood pressure (systolic or diastolic) was associated with a strong positive interaction in males with hemorrhagic stroke. Leptin may be an important link for the development of cerebrovascular disease in the insulin resistance syndrome in men.


[Functional decline in older persons after acute hospitalization].
Many persons aged 65 years and older encounter new disabilities in activities of daily living after a period of acute hospitalization; this is defined as functional decline. A consequence of functional decline is they are not able to perform essential daily activities, such as bathing, dressing, and walking independently. Recovery from these new disabilities is difficult. Older persons who encounter functional decline report a lower health-related quality of life and are often in need of permanent home care. The systematic in-hospital recording of a patient's functional status prior to hospital stay, early ambulation, and adequate and effective rehabilitation or transitional care can contribute to the prevention of functional decline. However, these interventions are often not applied in clinical practice. Until now, the prevention of functional decline has only been possible in a limited number of older persons. Major research questions include: can all older persons recover from functional decline, which mechanisms contribute most to functional decline and how can these mechanisms be influenced?


Administration of probiotics influences F4 (K88)-positive enterotoxigenic Escherichia coli attachment and intestinal cytokine expression in weaned pigs.
This study evaluated the effect of the probiotics Pediococcus acidilactici and Saccharomyces cerevisiae boulardii on the intestinal colonization of O149 enterotoxigenic Escherichia coli harbouring the F4 (K88) fimbriae (ETEC F4) and on the expression of ileal cytokines in weaned pigs. At birth, different litters of pigs were randomly assigned to one of the following treatments: 1) control without antibiotics or probiotics (CTRL); 2) reference group in which chlortetracycline and tiamulin were added to weanling feed (ATB); 3) P. acidilactici; 4) S. cerevisiae boulardii; or 5) P. acidilactici + S. cerevisiae boulardii. Probiotics were administered daily (1 × 10(9) CFU per pig) during the lactation period and after weaning (day 21). At 28 days of age, all pigs were orally challenged with an ETEC F4 strain, and a necropsy was performed 24 h later. Intestinal segments were collected to evaluate bacterial colonization in the small intestine and ileal cytokine expressions. Attachment of ETEC F4 to the intestinal mucosa was significantly reduced in pigs treated with P. acidilactici or S. cerevisiae boulardii in comparison with the ATB group (P = 0.01 and P = 0.03, respectively). In addition, proinflammatory cytokines, such as IL-6, were upregulated in ETEC F4 challenged pigs treated with P. acidilactici alone or in combination with S. cerevisiae boulardii compared with the CTRL group. In conclusion, the administration of P. acidilactici or S. cerevisiae boulardii was effective in reducing ETEC F4 attachment to the ileal mucosa, whereas the presence of P. acidilactici was required to modulate the expression of intestinal inflammatory cytokines in pigs challenged with ETEC F4.


Optimisation of a reversed-phase high-performance liquid chromatographic method for the determination of flavone acetic acid and its major human metabolites in plasma and urine.
A high-performance liquid chromatographic method for the determination of flavone acetic acid (FAA) and its major human metabolites in plasma and urine is described. Two factors were identified as being the key to resolving the metabolites; pH and buffer ionic strength. Run at optimal conditions of 10 mM ammonium acetate, pH 5.5-propan-2-ol (80:20) and a column temperature of 40 degrees C on a muBondapak C18 10 microns particle column (30 cm X 3.8 mm I.D.), two major metabolites were identified [FAA, retention time (tR) 6.02 min +/- 0.5% coefficient of variation (C.V.); metabolite 1, tR 4.13 min +/- 1.1% C.V.; metabolite 2, tR 5.10 min +/- 0.5% C.V. and hesperidin, internal standard, tR 4.69 min +/- 1.6% C.V.]. A solid-phase technique using Bond Elut C2 40-microns particles is described which extracts FAA, metabolites and internal standard with efficiencies in excess of 90%. Considerable attention has to be paid to sample preparation: FAA has poor aqueous solubility at acidic pH and the metabolites degrade back to FAA via intermediates at alkaline pH. Both problems can be avoided by buffering and diluting samples with 10 mM ammonium acetate, pH 5.5.


High Risk of Infection During Triple Therapy with First-Generation Protease Inhibitors: A Nationwide Cohort Study.
Peginterferon (PegIFN) remains the backbone of therapy for chronic hepatitis C (CHC) in economically constrained regions. However, PegIFN may cause neutropenia and addition of a protease inhibitor can increase the likelihood of neutropenia. The aims of this study were to assess the occurrence of clinically relevant infections during first-generation protease inhibitor based therapy and its risk factors as well as the relation to treatment-induced neutropenia. This multicenter (n=45) retrospective cohort study included CHC patients treated in the Netherlands. Based on absolute neutrophil count, categories of neutropenia were defined as: severe (<500/μL), moderate (500-750/μL) and mild (750-1500/μL). Likewise, infections were classified as severe (intravenous antibiotics/hospitalization) and moderate (anti-infective treatment). We assessed risk factors for infections using multivariable regression analysis with correction for multiple measurements. We included 467 CHC patients, 319 (68%) were male and 111 (24%) had cirrhosis. A total of 185 clinically relevant infections (34 severe) occurred in 145 patients (31%). During treatment 310 patients experienced neutropenia (34 severe). Multivariable analysis identified female sex (OR 1.7, 95%CI 1.2-2.5), chronic obstructive pulmonary disease (COPD) (OR 2.7, 95%CI 1.6- 4.5) and diabetes mellitus (OR 1.7, 95%CI 1.0-3.0) as risk factors for infections. Neutropenia at the previous visit was not associated with infection (univariable analysis: OR 0.9, 95%CI 0.6-1.3). This study shows that therapy with first generation protease inhibitors was complicated by an infection in 31% of patients. Not neutropenia, but female sex, COPD and diabetes mellitus were independent risk factors for infection. These patients should be monitored carefully once a PegIFN regimen is initiated.


Simulation-based end-of-life care training during surgical clerkship: assessment of skills and perceptions.
Assessment of interpersonal and psychosocial competencies during end-of-life care training is essential. This study reports the relationship between simulation-based end-of-life care Objective Structured Clinical Examination ratings and communication skills, trust, and self-assessed empathy along with the perceptions of students regarding their training experiences. Medical students underwent simulation-based end-of-life care OSCE training that involved standardized patients who evaluated students' communication skills and physician trust with the Kalamazoo Essential Elements Communication Checklist and the Wake Forest Physician Trust Scale. Students also completed the Jefferson Scale of Physician Empathy. Pearson correlation was used to examine the relationship between OSCE performance grades and communication, trust, and empathy scores. Student comments were analyzed using the constant comparative method of analysis to identify dominant themes. The 389 students (mean age 26.6 ± 2.8 y; 54.5% female) had OSCE grades that were positively correlated with physician trust scores (r = 0.325, P < 0.01) and communication skills (r = 0.383, P < 0.01). However, OSCE grades and self-reported empathy were not related (r = 0.021, P = 0.68). Time of clerkship differed for OSCE grade and physician trust scores; however, there was no trend identified. No differences were noted between the time of clerkship and communication skills or empathy. Overall, students perceived simulation-based end-of-life care training to be a valuable learning experience and appreciated its placement early in clinical training. We found that simulation-based OSCE training in palliative and end-of-life care can be effectively conducted during a surgery clerkship. Moreover, the standardized patient encounters combined with the formal assessment of communication skills, physician trust, and empathy provide feedback to students at an early phase of their professional life. The positive and appreciative comments of students regarding the opportunity to practice difficult patient conversations suggest that attention to these professional characteristics and skills is a valued element of clinical training and conceivably a step toward better patient outcomes and satisfaction.


Clinical fluctuations in MuSK myasthenia gravis are related to antigen-specific IgG4 instead of IgG1.
We studied the longitudinal relation between disease severity and titers of antigen-specific IgG subclasses in sera of patients with myasthenia gravis and antibodies to Muscle Specific Kinase (MuSK MG). Six patients were included of whom 55 samples had been collected during 2.5-13.4 years. Anti-MuSK antibodies were determined by ELISA and with a cell-based immunofluorescence assay. Disease severity was scored on a semi continuous scale. Only antigen-specific IgG4, and not IgG1, titers were significantly associated with disease severity in a linear mixed effect model (p = 0.036). Levels of IgG4 antibodies were above IgG1 in all samples except in one patient who went into clinical remission while switching from IgG4 to IgG1. The results support an important role for IgG4 in the pathogenesis of MuSK MG, in contrast to MG with anti-acetylcholine receptor antibodies.


Environmental life cycle assessment of production of the high intensity sweetener steviol glycosides from Stevia rebaudiana leaf grown in Europe: The SWEET project.
There is an increasing interest in the use of non-nutritive sweeteners to replace added sugar in food and beverage products for reasons of improving consumer health. Much work has been done to understand safety of sweeteners, but very little on sustainability. To address that gap, this study presents the results of a life cycle assessment (LCA) of production of rebaudioside A 60%, 95% pure (RA60) steviol glycoside mix from Stevia rebaudiana leaf grown in Europe. An attributional cradle-to-factory-gate life cycle assessment was conducted on growing of stevia leaves and extraction of steviol glycosides in Europe. Primary data were used from a case study supply chain. Results are reported in impact categories from the ReCiPe 2016 (H) method, with focus given to global warming potential, freshwater eutrophication, water consumption, and land use. Impacts are expressed both in terms of production mass and sweetness equivalence, a common metric for understanding high intensity sweetener potency. Sweetness equivalence of RA60 is typically 200 to 300 times that of sugar. Comparison of environmental impact is made to sugar (sucrose) produced from both cane and beets. The research is part of the EU project SWEET (sweeteners and sweetness enhancers: impact on health, obesity, safety, and sustainability). Global warming potential for production of RA60 was found to be 20.25 kgCO2-eq/kgRA60 on a mass basis and 0.081 kgCO2-eq/kgSE on a sweetness equivalence basis. Field production of stevia leaves was found to be the main source of impact for most impact categories, and for all four focus categories. Extraction of the RA60 was the main source of impact for the others. Leaf processing and seedling propagation were minor contributors to life cycle impact. Removal of international transport from the supply chain reduced global warming potential by 18.8%. Compared with sugar on a sweetness equivalence basis, RA60 has approximately 5.7% to 10.2% the impact for global warming potential, 5.6% to 7.2% the impact for land use, and is lower across most other impact categories. This is the first LCA of steviol glycoside mix RA60 produced from leaf in Europe. The results indicate that RA60 can be used to reduce environmental impact of providing a sweet taste by replacing sugar across all impact categories. However, it is important to note that specific formulations in which RA60 is used will have a bearing on the final environmental impact of any food or beverage products. For solid foods, this requires further research. The online version contains supplementary material available at 10.1007/s11367-022-02127-9.


Functional ADA polymorphism increases sleep depth and reduces vigilant attention in humans.
Homeostatically regulated slow-wave oscillations in non-rapid eye movement (REM) sleep may reflect synaptic changes across the sleep-wake continuum and the restorative function of sleep. The nonsynonymous c.22G>A polymorphism (rs73598374) of adenosine deaminase (ADA) reduces the conversion of adenosine to inosine and predicts baseline differences in sleep slow-wave oscillations. We hypothesized that this polymorphism affects cognitive functions, and investigated whether it modulates electroencephalogram (EEG), behavioral, subjective, and biochemical responses to sleep deprivation. Attention, learning, memory, and executive functioning were quantified in healthy adults. Right-handed carriers of the variant allele (G/A genotype, n = 29) performed worse on the d2 attention task than G/G homozygotes (n = 191). To test whether this difference reflects elevated homeostatic sleep pressure, sleep and sleep EEG before and after sleep deprivation were studied in 2 prospectively matched groups of G/A and G/G genotype subjects. Deep sleep and EEG 0.75- to 1.5-Hz oscillations in non-REM sleep were significantly higher in G/A than in G/G genotype. Moreover, attention and vigor were reduced, whereas waking EEG alpha activity (8.5-12 Hz), sleepiness, fatigue, and α-amylase in saliva were enhanced. These convergent data demonstrate that genetic reduction of ADA activity elevates sleep pressure and plays a key role in sleep and waking quality in humans.


Half of breast cancer patients discontinue tamoxifen and any endocrine treatment before the end of the recommended treatment period of 5 years: a population-based analysis.
Observational studies on long-term endocrine treatment among breast cancer patients have presented discontinuation rates on tamoxifen, but lack information on the continuance of any endocrine treatment [both tamoxifen and aromatase inhibitors (AIs)] within the same cohort. In this study we determined switching rates from tamoxifen to AIs, discontinuation rates of tamoxifen only, discontinuation rates of any endocrine treatment and determinants of first treatment switch and treatment discontinuation. Patients with early stage breast cancer (stage I-IIIa) starting on tamoxifen were selected from the linked Eindhoven Cancer Registry-PHARMO RLS cohort in the period 1998-2006. Continuous use (allowing a 60 days gap between refills) of tamoxifen only and any endocrine treatment were determined after various follow-up periods: 1, 2, 3, 4, and 5 years. Time to first switch from tamoxifen to an AI was assessed. Cox regression was used to identify determinants of first treatment switch, discontinuation of tamoxifen, and discontinuation of any endocrine treatment. A total of 1,451 new early stage breast cancer patients started on tamoxifen. Of those, 380 had a treatment switch to an AI during follow-up. Of the patients followed for 5 years, 40% continuously used tamoxifen, which was 49% for any endocrine treatment. Older age (older than 70 versus 50-69 years) was independently associated with increased discontinuation of tamoxifen and any endocrine therapy. Patients with two or more concomitant diseases (versus no comorbidity) showed an increased likelihood to stop any endocrine treatment or switch treatment from tamoxifen to an AI. In conclusion, up to half of the breast cancer patients starting tamoxifen continued 5 years of endocrine treatment. Identification of patients at risk of discontinuation will assist in the development of interventions to improve treatment continuation comparable to that of patients included in clinical trials.


A catechol-O-methyltransferase that is essential for auditory function in mice and humans.
We have identified a previously unannotated catechol-O-methyltranferase (COMT), here designated COMT2, through positional cloning of a chemically induced mutation responsible for a neurobehavioral phenotype. Mice homozygous for a missense mutation in Comt2 show vestibular impairment, profound sensorineuronal deafness, and progressive degeneration of the organ of Corti. Consistent with this phenotype, COMT2 is highly expressed in sensory hair cells of the inner ear. COMT2 enzymatic activity is significantly reduced by the missense mutation, suggesting that a defect in catecholamine catabolism underlies the auditory and vestibular phenotypes. Based on the studies in mice, we have screened DNA from human families and identified a nonsense mutation in the human ortholog of the murine Comt2 gene that causes nonsyndromic deafness. Defects in catecholamine modification by COMT have been previously implicated in the development of schizophrenia. Our studies identify a previously undescribed COMT gene and indicate an unexpected role for catecholamines in the function of auditory and vestibular sense organs.


Wide-neck aneurysms: systematic review of the neurosurgical literature with a focus on definition and clinical implications.
Wide-necked aneurysms (WNAs) are a variably defined subset of cerebral aneurysms that require more advanced endovascular and microsurgical techniques than those required for narrow-necked aneurysms. The neurosurgical literature includes many definitions of WNAs, and a systematic review has not been performed to identify the most commonly used or optimal definition. The purpose of this systematic review was to highlight the most commonly used definition of WNAs. The authors searched PubMed for the years 1998-2017, using the terms "wide neck aneurysm" and "broad neck aneurysm" to identify relevant articles. All results were screened for having a minimum of 30 patients and for clearly stating a definition of WNA. Reference lists for all articles meeting the inclusion criteria were also screened for eligibility. The search of the neurosurgical literature identified 809 records, of which 686 were excluded (626 with < 30 patients; 60 for lack of a WNA definition), leaving 123 articles for analysis. Twenty-seven unique definitions were identified and condensed into 14 definitions. The most common definition was neck size ≥ 4 mm or dome-to-neck ratio < 2, which was used in 49 articles (39.8%). The second most commonly used definition was neck size ≥ 4 mm, which was used in 26 articles (21.1%). The rest of the definitions included similar parameters with variable thresholds. There was inconsistent reporting of the precise dome measurements used to determine the dome-to-neck ratio. Digital subtraction angiography was the only imaging modality used to study the aneurysm morphology in 87 of 122 articles (71.3%). The literature has great variability regarding the definition of a WNA. The most prevalent definition is a neck diameter of ≥ 4 mm or a dome-to-neck ratio of < 2. Whether this is the most appropriate and clinically useful definition is an area for future study.


Effect of exogenous estrogens and progestogens on the course of migraine during reproductive age: a consensus statement by the European Headache Federation (EHF) and the European Society of Contraception and Reproductive Health (ESCRH).
We systematically reviewed data about the effect of exogenous estrogens and progestogens on the course of migraine during reproductive age. Thereafter a consensus procedure among international experts was undertaken to develop statements to support clinical decision making, in terms of possible effects on migraine course of exogenous estrogens and progestogens and on possible treatment of headache associated with the use or with the withdrawal of hormones. Overall, quality of current evidence is low. Recommendations are provided for all the compounds with available evidence including the conventional 21/7 combined hormonal contraception, the desogestrel only oral pill, combined oral contraceptives with shortened pill-free interval, combined oral contraceptives with estradiol supplementation during the pill-free interval, extended regimen of combined hormonal contraceptive with pill or patch, combined hormonal contraceptive vaginal ring, transdermal estradiol supplementation with gel, transdermal estradiol supplementation with patch, subcutaneous estrogen implant with cyclical oral progestogen. As the quality of available data is poor, further research is needed on this topic to improve the knowledge about the use of estrogens and progestogens in women with migraine. There is a need for better management of headaches related to the use of hormones or their withdrawal.


The prevalence of psychiatric disorder among a community sample of crack cocaine users: an exploratory study with practical implications.
The Diagnostic Interview Schedule for DSM-IV was used to assess the lifetime prevalence of psychiatric disorder among not-in-treatment crack cocaine users (N = 313). The most common dependencies involved cocaine (59.7%), alcohol (37.7%), and cannabis (12.1%). The most common nondependency disorders were antisocial personality disorder (ASPD; 24%), depression (17.8%), and posttraumatic stress disorder (PTSD; 11.8%). Comorbidity was present in 36.4% of the sample. Proportionately more white than black users were dependent on cocaine, alcohol, amphetamine, and sedative-hypnotics. More white than black users experienced ASPD, depression, PTSD, and attention deficit disorder. Proportionately fewer black users suffered comorbid disorders. Proportionately more men had ASPD, whereas more women had PTSD and phobias. Multinominal modeling revealed that black users and users with a high school education were less likely to have comorbid disorders; married users were less likely to have nondependency disorders. White crack users were more likely to have comorbidity, complicating their treatment.


MS2 Coliphage as a Surrogate for 2009 Pandemic Influenza A (H1N1) Virus (pH1N1) in Surface Survival Studies on N95 Filtering Facepiece Respirators.
Research on influenza viruses regarding transmission and survival has surged in the recent years due to infectious emerging strains and outbreaks such as the 2009 Influenza A (H1N1) pandemic. MS2 coliphage has been applied as a surrogate for pathogenic respiratory viruses, such as influenza, as it's safe for personnel to handle and requires less time and labor to measure virus infectivity. However, direct comparisons to determine the effectiveness of coliphage as a surrogate for influenza virus regarding droplet persistence on personal protective equipment such as N95 filtering facepiece respirators (FFRs) are lacking. Persistence of viral droplets deposited on FFRs in healthcare settings is important to discern due to the potential risk of infection via indirect fomite transmission. The objective of this study was to determine if MS2 coliphage could be applied as a surrogate for influenza A viruses for studying persistence when applied to the FFRs as a droplet. The persistence of MS2 coliphage and 2009 Pandemic Influenza A (H1N1) Virus on FFR coupons in different matrices (viral media, 2% fetal bovine serum, and 5 mg ml-1 mucin) were compared over time (4, 12, 24, 48, 72, and 144 hours) in typical absolute humidity conditions (4.1 × 105 mPa [18°C/20% relative humidity (RH)]). Data revealed significant differences in viral infectivity over the 6-day period (H1N1- P <0.0001; MS2 - P <0.005), although a significant correlation of viral log10 reduction in 2% FBS (P <0.01) was illustrated. Overall, MS2 coliphage was not determined to be a sufficient surrogate for influenza A virus with respect to droplet persistence when applied to the N95 FFR as a droplet.


Living with inflammatory bowel disease: A Crohn's and Colitis Canada survey.
Despite improvements in therapies for inflammatory bowel diseases (IBDs), patient quality of life continues to be significantly impacted. To assess the impact of IBD on patients and families with regard to leisure, relationships, mental well-being and financial security, and to evaluate the quality and availability of IBD information. An online survey was advertised on the Crohn's and Colitis Canada website, and at gastroenterology clinics at the University of Alberta Hospital (Edmonton, Alberta) and University of Calgary Hospital (Calgary, Alberta). The survey was completed by 281 IBD patients and 32 family members. Among respondents with IBD, 64% reported a significant or major impact on leisure activities, 52% a significant or major impact on interpersonal relationships, 40% a significant or major impact on financial security, and 28% a significant or major impact on planning to start a family. Patient information needs emphasized understanding disease progression (84%) and extraintestinal symptoms (82%). There was a strong interest in support systems such as health care insurance (70%) and alternative therapies (66%). The most common source of information for patients was their gastroenterologist (70%); however, most (70%) patients preferred to obtain their information from the Crohn's and Colitis Canada website. The impact of IBD on interpersonal relationships and leisure activities was significant among IBD patients and their families. Understanding the disease, but also alternative treatment options, was of high interest. Currently, there is a discrepancy between interest in information topics and their availability. Respondents reported a strong desire to obtain information regarding disease progression, especially extraintestinal symptoms.


Caffeinated Gum Does Not Influence RPE-Regulated Cadence in Recreationally-Active College Females Regardless of Habitual Caffeine Consumption.
Caffeine (CAF) has been extensively studied for its ergogenic and analgesic effects during exercise. However, the majority of these studies have been conducted in male populations. This study investigated the effects of acute CAF chewing gum on self-selected exercise intensity during a rating of perceived exertion (RPE) production trial in active females (n = 16, 21.0 ± 2.8 y). Data were also analyzed based on habitual CAF consumption level. Participants completed a V̇O2peak trial, followed by a familiarization and two randomized, triple-blinded experimental RPE production trials on an arm ergometer [clamped resistance, blinded to self-selected cadence (CAD)] with either CAF gum (300 mg; 4.8 ± 0.7 mg/kg-1 body mass) or placebo (PLA), at a prescribed RPE of 4 and 7 (10 min each). Self-selected CAD did not statistically differ (p > 0.05) between CAF or PLA for an RPE4 (37.7 ± 1.6 vs. 37.6 ± 1.6 rev·min-1) or RPE7 (42.9 ± 1.6 vs. 41.2 ± 1.7 rev·min-1), respectively. There were no statistical differences between treatment groups for any other variables, except restlessness rating which was significantly higher (3.5 vs. 2.2; p = 0.03, d = 0.64) for the CAF group compared to PLA. Secondary analysis revealed no statistical differences for any variables between habitual consumers of low (23 ± 20 mg/day) or mod/high (195 ± 93 mg/day) CAF. Our data support previous studies examining CAF in women across different testing modalities and suggest that regardless of habitual CAF consumption, females might require higher doses of CAF to replicate subjective and physiological responses commonly observed using similar RPE production protocols in male participants. These findings support the need for additional investigations into female physiological and perceptual responses following CAF ingestion.


Homogeneous time resolved fluorescence resonance energy transfer using rare earth cryptates as a tool for probing molecular interactions in biology.
A homogeneous assay technology using time resolved fluorescence and fluorescence resonance energy transfer is described. A new class of fluorescent complexes, the cryptates, have been used as fluorescent donor with cross-linked allophycocyanin as acceptor. This new donor/acceptor shows an exceptionally high Förster distance R0 of 9 nm. This allows to build up a set of strategies to probe the interactions of biomolecules in biology, particularly for high throughput screening applications. In this article, we describe the basics of the technology and review applications developed for studying different key molecular interactions involved in cellular processes.


FTO genotype and weight loss in diet and lifestyle interventions: a systematic review and meta-analysis.
Studies have suggested that the fat mass and obesity-associated (FTO) genotype is associated with individual variability in weight loss in response to diet/lifestyle interventions, but results are inconsistent. We aimed to provide a summary of the literature evaluating the relation between the FTO genotype and weight loss in response to diet/lifestyle interventions. A search of English-language articles in the PubMed and Embase databases (through 30 April 2015) was performed. Eligible studies were diet/lifestyle weight-loss intervention studies conducted in adults that reported changes in body weight or body mass index (BMI) by the FTO variant rs9939609 (or its proxy). Differences in weight loss between FTO genotypes across studies were pooled with the use of fixed-effect models. A meta-analysis of 10 studies (comprising 6951 participants) that reported the results of additive genetic models showed that individuals with the FTO TA genotype and AA genotype (those with the obesity-predisposing A allele) had 0.18-kg (95% CI: -0.09-, 0.45-kg;P= 0.19; NS) and 0.44-kg (95% CI: 0.09-, 0.79-kg;P= 0.015) greater weight loss, respectively, than those with the TT genotype. A meta-analysis of 14 studies (comprising 7700 participants) that reported the results of dominant genetic models indicated a 0.20-kg (-0.43-, 0.04-kg) greater weight loss in the TA/AA genotype than in the TT genotype (P= 0.10). In addition, differences in weight loss between the AA genotype and TT genotype were significant in studies with a diet intervention only, adjustment for baseline BMI or body weight, and several other subgroups. However, the relatively small number of studies limited these stratified analyses, and there was no statistically significant difference between subgroups. This meta-analysis suggests that individuals carrying the homozygous FTO obesity-predisposing allele may lose more weight through diet/lifestyle interventions than noncarriers. Our data provide evidence for genetic variability in response to diet/lifestyle interventions on weight loss, although clinical applications of these findings need further investigations.


Significant correlation between visceral adiposity and high-sensitivity C-reactive protein (hs-CRP) in Japanese subjects.
High-sensitivity C-reactive protein (hs-CRP) is a marker for low-grade inflammation, as well as atherosclerosis, obesity, hyperglycemia and hypertension. Because the factor showing the strongest association with inflammation is currently unknown, we investigated the associations between hs-CRP and clinical and biochemical characteristics in Japanese subjects with mild obesity or impaired glucose tolerance. Subjects aged <65 years old, attending the Seirei Medical Examination Center, underwent complete physical and laboratory examinations. A total of 112 subjects (mean age 59.9±5.9 years old, males/females: n=50/62) with a waist circumference of >85 cm in males and >90 cm in females, homeostasis model assessment-insulin resistance (HOMA-IR) ≥1.7, or impaired glucose tolerance were eligible for this study. All subjects had normal albuminuria. Log-transformed hs-CRP concentrations were significantly correlated with BMI (r=0.278, p<0.01), HOMA-IR (r=0.296, p<0.005), 2-h post-challenge IRI during an oral glucose tolerance test (r=0.218, p<0.05), maximum intima-media thickness (r=0.240, p<0.05), visceral fat area evaluated by computed tomography (r=0.423, p<0.0001) and subcutaneous fat area (r=0.231, p<0.05). Multiple linear regression analysis showed that visceral fat was the most significantly correlated factor with hs-CRP. Visceral fat mass was a significant and independent predictor for serum hs-CRP levels in Japanese subjects with mild obesity and/or impaired glucose tolerance.


How viscoelastic is human blood plasma?
Blood plasma has been considered a Newtonian fluid for decades. Recent experiments (Brust et al., Phys. Rev. Lett., 2013, 110) revealed that blood plasma has a pronounced viscoelastic behavior. This claim was based on purely elastic effects observed in the collapse of a thin plasma filament and the fast flow of plasma inside a contraction-expansion microchannel. However, due to the fact that plasma is a solution with very low viscosity, conventional rotational rheometers are not able to stretch the proteins effectively and thus, provide information about the viscoelastic properties of plasma. Using computational rheology and a molecular-based constitutive model, we predict accurately the rheological response of human blood plasma in strong extensional and constriction complex flows. The complete rheological characterization of plasma yields the first quantitative estimation of its viscoelastic properties in shear and extensional flows. We find that although plasma is characterized by a spectrum of ultra-short relaxation times (on the order of 10-3-10-5 s), its elastic nature dominates in flows that feature high shear and extensional rates, such as blood flow in microvessels. We show that plasma exhibits intense strain hardening when exposed to extensional deformations due to the stretch of the proteins in its bulk. In addition, using simple theoretical considerations we propose fibrinogen as the main candidate that attributes elasticity to plasma. These findings confirm that human blood plasma features bulk viscoelasticity and indicate that this non-Newtonian response should be seriously taken into consideration when examining whole blood flow.


Repurposing the anticancer drug mitomycin C for the treatment of persistent Acinetobacter baumannii infections.
Acinetobacter baumannii is an emergent opportunistic bacterial pathogen responsible for recalcitrant infections owing to its high intrinsic tolerance to most antibiotics; therefore, suitable strategies to treat these infections are needed. One plausible approach is the repurposing of drugs that are already in use. Among them, anticancer drugs may be especially useful due their cytotoxic activities and ample similarities between bacterial infections and growing tumours. In this work, the effectiveness of four anticancer drugs on the growth of A. baumannii ATTC BAA-747 was evaluated, including the antimetabolite 5-fluorouracil and three DNA crosslinkers, namely cisplatin, mitomycin C (MMC) and merphalan. MMC was the most effective drug, having a minimum inhibitory concentration for 50% of growth in Luria-Bertani medium at ca. 7 µg/mL and completely inhibiting growth at 25 µg/mL. Hence, MMC was tested against a panel of 21 clinical isolates, including 18 multidrug-resistant (MDR) isolates, 3 of which were sensitive only to colistin. The minimum inhibitory concentrations and minimum bactericidal concentrations of MMC in all tested strains were found to be similar to those of A. baumannii ATCC BAA-747, and MMC also effectively killed stationary-phase, persister and biofilm cells. Moreover, MMC was able to increase survival of the insect larvae Galleria mellonella against an otherwise lethal A. baumannii infection from 0% to ≥53% for the antibiotic-sensitive A. baumannii ATCC BAA-747 strain and the MDR strains A560 and A578. Therefore, MMC is highly effective at killing the emergent opportunistic pathogen A. baumannii.


Human mature T cells that are anergic in vivo prevail in SCID mice reconstituted with human peripheral blood.
In these studies we have characterized the human cells that repopulate severe combined immunodeficient (SCID) mice after injection of adult peripheral blood or cord blood (hu-PBL-SCID mice). In all organs of the chimeras, and at any time point tested, single-positive (CD4+ or CD8+) T cells that expressed the alpha/beta T cell receptor (TCR) prevailed. All T cells were CD45RO+ and the majority were also HLA-DR+. Thus, the human T cells in the chimeras exhibited the phenotype of mature, memory cells that showed signs of recent activation. Cell cycle studies revealed a mitotically active human T cell population in the murine host. However, when freshly isolated from the chimeras, the human T cells were refractory to stimulation by anti-CD3 antibody but proliferated in response to exogenous interleukin 2. Chimera-derived human T cell lines retained this state of unresponsiveness to TCR-triggered proliferation for 4-6 wk in vitro. Subsequently, the T cell lines developed responses to anti-CD3 stimulation and 9 of 11 of the lines also proliferated in response to splenic stimulator cells of SCID mice. These data demonstrate that the human T cells are in a state of reversible anergy in the murine host and that xenoreactivity might play a critical role in hu-PBL-SCID mice. Mechanisms that may determine repopulation of SCID mice with human peripheral blood mononuclear cells are discussed.


Long-term in vivo impedance changes of subretinal microelectrodes implanted in dystrophic P23H rats.
Retinal prostheses are being developed to restore vision in blind patients with photoreceptor degeneration. Electrodes arrays were subretinally implanted in transgenic P23H rats with their photoreceptors degenerated. Electrical stability of the implants was evaluated by long-term monitoring of their impedance changes. Electrode impedances were found to increase by two log units over a three weeks period whereas no impedance increase was noted when the implants were located in the vitreous. In case of hemorrhage or major fibrous reactions, the impedance continued to increase steadily. After explantation, it recovered its initial value indicating no deterioration of the implant. Although the glial cell layer at the surface of the subretinal space was slightly larger, no major glial reaction was seen in direct contact to the implant. These results indicate that no functional testing should be considered before at least three weeks post implantation.


Medically Related Post-traumatic Stress in Children and Adolescents with Congenital Heart Defects.
Children and adolescents with a congenital heart defect (ConHD) frequently undergo painful or frightening medical procedures and hospitalizations. They often need multiple invasive procedures at a very young age and require regular checkups during their entire life. From other pediatric populations, it is known that these kinds of experiences can result in acute stress reactions and even in post-traumatic stress disorder (PTSD) in the long-term. PTSD and also subthreshold PTSD can lead to serious (psychosocial) impairment. However, limited information is available about PTSD in children with ConHD. Therefore, the aim of this review is to provide a summary of the current literature on post-traumatic stress (PTS) in children and adolescents with ConHD describing the prevalence of PTSD and its predictors/correlates. This review indicates that a range of 12-31% of children undergoing cardiac surgery develop PTSD. A range of 12-14% shows elevated post-traumatic stress symptoms (PTSS). These findings are comparable to those of hospitalized children without ConHD. Noteworthy, most studies used varying self-report questionnaires to measure PTSD and only one study used a semistructured interview. Although all studies point in the same direction of elevated PTSD and PTSS, systematic research is necessary to be able to draw firm conclusions. At present, as far as we know, in most clinics treating patients with ConHD, there is no regular screening for PTS in children with ConHD. In the reviewed literature, there is strong consensus that screening for PTSS and (preventive) psychological care for children and adolescents with ConHD is urgently needed.