Directional specificity in the regeneration of lamprey spinal axons.
After spinal transection in ammocoetes (lamprey larvae) 4 to 5 years old, functional recovery is accompanied by a limited regeneration in which axons grow as far as 5 millimeters beyond the scar. In axotomized giant interneurons labeled intracellularly with horseradish peroxidase 16 to 120 days after transection, 74 percent of regenerating neurites grew in their normal projection pattern, rostal and contralateral to the cell body. One third of the neurites originated anomalously from posterior dendrites. Despite their initial abnormal orientation, 80 percent of these neurites looped contralaterally and rostrally to assume the normal projection path. The directional specificity persisted when giant interneurons were located in islands formed by double simultaneous cord transection. This limited regeneration seems to be characterized by directional selectivity that cannot be attributed to nonspecific influences, such as a tendency of neurites to grow in an already established direction or a trophic effect of the zone of injury.


On the other hand am I rational? Hemispheric activation and the framing effect.
In recent decades the investigation of framing effects has become the foremost studied phenomenon of rational/irrational decision making. Two experiments were conducted to determine whether the functional specializations of the left and the right hemispheres would produce different responses to a traditional framing task. In Experiment 1, a behavioral task of finger tapping was used to induce asymmetrical activation of the respective hemispheres. In Experiment 2, a monaural listening procedure was used. In both experiments, the predicted results were found. Framing effects were found when the right hemisphere was selectively activated whereas they were not observed when the left hemisphere was selectively activated. The results are interpreted in light of a dual-processing approach of decision-making.


MARCO, TLR2, and CD14 are required for macrophage cytokine responses to mycobacterial trehalose dimycolate and Mycobacterium tuberculosis.
Virtually all of the elements of Mycobacterium tuberculosis (Mtb) pathogenesis, including pro-inflammatory cytokine production, granuloma formation, cachexia, and mortality, can be induced by its predominant cell wall glycolipid, trehalose 6,6'-dimycolate (TDM/cord factor). TDM mediates these potent inflammatory responses via interactions with macrophages both in vitro and in vivo in a myeloid differentiation factor 88 (MyD88)-dependent manner via phosphorylation of the mitogen activated protein kinases (MAPKs), implying involvement of toll-like receptors (TLRs). However, specific TLRs or binding receptors for TDM have yet to be identified. Herein, we demonstrate that the macrophage receptor with collagenous structure (MARCO), a class A scavenger receptor, is utilized preferentially to "tether" TDM to the macrophage and to activate the TLR2 signaling pathway. TDM-induced signaling, as measured by a nuclear factor-kappa B (NF-kappaB)-luciferase reporter assay, required MARCO in addition to TLR2 and CD14. MARCO was used preferentially over the highly homologous scavenger receptor class A (SRA), which required TLR2 and TLR4, as well as their respective accessory molecules, in order for a slight increase in NF-kappaB signaling to occur. Consistent with these observations, macrophages from MARCO(-/-) or MARCO(-/-)SRA(-/-) mice are defective in activation of extracellular signal-related kinase 1/2 (ERK1/2) and subsequent pro-inflammatory cytokine production in response to TDM. These results show that MARCO-expressing macrophages secrete pro-inflammatory cytokines in response to TDM by cooperation between MARCO and TLR2/CD14, whereas other macrophage subtypes (e.g. bone marrow-derived) may rely somewhat less effectively on SRA, TLR2/CD14, and TLR4/MD2. Macrophages from MARCO(-/-) mice also produce markedly lower levels of pro-inflammatory cytokines in response to infection with virulent Mtb. These observations identify the scavenger receptors as essential binding receptors for TDM, explain the differential response to TDM of various macrophage populations, which differ in their expression of the scavenger receptors, and identify MARCO as a novel component required for TLR signaling.


Fate of plasma membrane during endocytosis. III. Evidence for incomplete breakdown of immunoglobulin in lysosomes of cultured fibroblasts.
Rat embryo fibroblasts, when cultured in the presence of control rabbit immunoglobulins (C IgG), doubly labeled by (3)H-acetylation (A) and then conjugated with flourescein (F), take up FAC IgG continuously for at least 72 h. They return the major part of their intake back to the medium in the form of breakdown products of very low molecular weight. Gel filtration and immunological analyses of cells and medium at various times indicate that essentially all the FAC IgG molecules taken up undergo digestion of their Fc part, but that the Fab part of only about three-fourths of the molecules is degraded. The rest remains stored intracellularly in the form of F(ab')2-type fragments that slowly dissociate into Fab'-type fragments. When FAC IgG was incubated in vitro in the presence of a hepatic lysosomal extract, complete digestion of the Fc part likewise occurred, but the Fab' part of most if not all the molecules proved resistant to breakdown, and remained as Fab'-type fragments. Cell fractionation experiments have demonstrated that the storage compartment of the FAC IgG and of its digestion residues: (a) shows a density distribution pattern in a sucrose gradient identical to that of the lysosomal marker N-acetyl-beta-glucosaminidase and clearly dissociated from that of the Golgi marker galactosyltransferase, and (b) accompanies the lysosomal marker in its density shift induced by exposure of the cells to chloroquine. It is concluded that storage and processing of FAC IgG by rat fibroblasts occur in a single, digestively active compartment of lysosomal nature, and that resistance to digestion of certain Fab'-type fragments accounts largely for the inability of the lysososmal enzymes to completely digest the FAC IgG taken up. This conclusion implies that the intracellular storage compartment through which, in earlier work, plasma membrane patches were found to transit after endocytosis and before recycling to the cell surface consists of authentic lysosomes.


A highly active catalyst for Huisgen 1,3-dipolar cycloadditions based on the tris(triazolyl)methanol-Cu(I) structure.
A new tris(1-benzyl-1H-1,2,3-triazol-4-yl)methanol ligand 3 has been prepared by a triple Cu(I)-catalyzed alkyne-azide 1,3-dipolar cycloaddition (CuAAC). Ligand 3 forms a stable complex with CuCl, which catalyzes the Huisgen 1,3-dipolar cycloaddition on water or under neat conditions. Low catalyst loadings, short reaction times at room temperature, and compatibility with free amino groups make 3.CuCl an outstanding catalyst for CuAAC.


Prevention of posttraumatic alterations in lymphocyte subpopulations in mice by immunomodulating drugs.
Infection is a major cause of morbidity and mortality in traumatized victims, as well as an unfortunate complication following surgical procedures. The incidence of such infections is probably related to immune suppression occurring after severe trauma and surgery. Cell-mediated immunity is an arm of immunity that is probably important in defense against bacterial as well as fungal and viral infections. The status of cell-mediated immunity appears to be reflected by the helper/inducer to suppressor/cytotoxic lymphocyte (Ly1+/Ly2+ in the mouse model) ratio. A depressed ratio has been shown to correlate with other measures of suppressed immunity. Using monoclonal antibodies and immunofluorescent analysis, splenic lymphocyte subpopulations were enumerated at different intervals after injury in mice receiving various injuries (laparotomy, 40% liver resection, burn injury covering 20% of total body surface area, and crush injury-amputation of hind limb). An immediate fall in the Ly1+/Ly2+ ratio was observed following the most severe injuries, namely, liver resection, burn, and crush injury-amputation, but not following laparotomy. Changes in the ratio were reflected by both decreases in helper populations (Ly1+) and increases in suppressor populations (Ly2+). Prostaglandins and histamines have been implicated as possible mediators in producing down-regulation of the immune system. Posttraumatic lymphocyte alterations were prevented in crush injury-amputation group mice when drugs (ibuprofen and cimetidine) to block such mediators were administered prior to trauma; changes were also prevented by administering cyclophosphamide, a drug which in low dose might inhibit proliferation of suppressor lymphocyte populations.


Identification of open reading frames unique to a select agent: Ralstonia solanacearum race 3 biovar 2.
An 8x draft genome was obtained and annotated for Ralstonia solanacearum race 3 biovar 2 (R3B2) strain UW551, a United States Department of Agriculture Select Agent isolated from geranium. The draft UW551 genome consisted of 80,169 reads resulting in 582 contigs containing 5,925,491 base pairs, with an average 64.5% GC content. Annotation revealed a predicted 4,454 protein coding open reading frames (ORFs), 43 tRNAs, and 5 rRNAs; 2,793 (or 62%) of the ORFs had a functional assignment. The UW551 genome was compared with the published genome of R. solanacearum race 1 biovar 3 tropical tomato strain GMI1000. The two phylogenetically distinct strains were at least 71% syntenic in gene organization. Most genes encoding known pathogenicity determinants, including predicted type III secreted effectors, appeared to be common to both strains. A total of 402 unique UW551 ORFs were identified, none of which had a best hit or >45% amino acid sequence identity with any R. solanacearum predicted protein; 16 had strong (E < 10(-13)) best hits to ORFs found in other bacterial plant pathogens. Many of the 402 unique genes were clustered, including 5 found in the hrp region and 38 contiguous, potential prophage genes. Conservation of some UW551 unique genes among R3B2 strains was examined by polymerase chain reaction among a group of 58 strains from different races and biovars, resulting in the identification of genes that may be potentially useful for diagnostic detection and identification of R3B2 strains. One 22-kb region that appears to be present in GMI1000 as a result of horizontal gene transfer is absent from UW551 and encodes enzymes that likely are essential for utilization of the three sugar alcohols that distinguish biovars 3 and 4 from biovars 1 and 2.


Regulation of tRNA suppressor activity by an intron-encoded polyadenylation signal.
A 26-nt sequence from the 3' UTR of the yeast GAL7 mRNA directs accurate and efficient cleavage and polyadenylation to form the 3' end of the GAL7 mRNA in vivo and in vitro. Here we asked whether this polyadenylation signal can function within the context of a tRNA. Insertion of the GAL7 signal into the intron of the dominant SUP4 nonsense suppressor allowed us to judge the effect of the insert on SUP4 function by observation of nonsense suppression efficiency in vivo. The GAL7 signal impairs the function of SUP4 in an orientation-dependent manner in vivo, consistent with its ability to specify cleavage and polyadenylation in this context in vitro. Mutation of a UA repeat within the GAL7 signal restores SUP4 function partially, consistent with the role of this repeat as an efficiency element in polyadenylation. Mutations that impair the mRNA 3' end-processing factors Rna14p and Rna15p restore suppressor function partially. Northern blot analysis, PCR amplification, and DNA sequence analysis show that the GAL7 signal directs polyadenylation within the body of pre-SUP4 and within the terminator, suggesting that polyadenylation inhibits 5' and 3' end processing, as well as removal of the pre-tRNA intron. These findings indicate that the GAL7 polyadenylation signal is capable of targeting a pre-tRNA to the mRNA processing pathway.


Inhaled nitric oxide and intravenous magnesium sulphate for the treatment of persistent pulmonary hypertension of the newborn.
The aim of this study was to compare the response and survival rates of term infants with persistent pulmonary hypertension of the newborn (PPHN) on high frequency oscillatory ventilation (HFOV) treated with either inhaled nitric oxide (iNO) or intravenous magnesium sulphate (MgSO4). This was a randomised controlled study. The inclusion criteria were infants with respiratory distress, oxygen index equal to or greater than 25 despite HFOV support, and echocardiographic evidence of PPHN. Infants in the MgSO4 group (n is 13) were loaded with MgSO4 200 mg/kg infused over half an hour, followed by continuous infusion at 50-150 mg/kg/hour to attain a serum magnesium level of 5.0-7.0 mmol/L. Infants in the iNO group (n is 12) were administered nitric oxide at an initial concentration of 20 ppm. Analysis was done on an intention-to-treat basis. There was no significant difference in the median age when the vasodilators were commenced (MgSO4 group: 14.0 hours, interquartile range [IQR]: 7.5, 27.0; iNO group: 14.8 hours, IQR: 12.5, 35.3, p is 0.8). There was no significant difference in the proportion of infants who responded primarily to either vasodilator (MgSO4: 23.3%, iNO: 33.3%, p is 1.0) . After switching over to iNO following a failed MgSO4 therapy, a significantly higher proportion (9 out of 10) of the non-respondents in the MgSO4 group recovered from PPHN and survived compared to the non-respondents in the iNO group (1 out of 8) who switched over to intravenous MgSO4 (p is less than 0.03). Infants who were administered iNO following a failed MgSO4 therapy were associated with a better outcome than those who were administered MgSO4 following a failed iNO therapy.


Synthesis of bis(trithio)phosphines by oxidative transfer of phosphorus(i).
The synthesis of novel bis(trithio)phosphines is achieved by oxidative addition of tetrathiocins to the phosphorus(i) reagent [PIdppe][Br] in good yields under ambient conditions. These bis(trithio)phosphines and the related intermediate diphosphine species are characterized by X-ray diffraction and multinuclear NMR and a mechanism is proposed for the formation of these molecules. In contrast, the related reaction with diphenyldisulfide produces a mononuclear tris(phenylthio)phosphine.


Intergenic RNA mainly derives from nascent transcripts of known genes.
Eukaryotic genomes undergo pervasive transcription, leading to the production of many types of stable and unstable RNAs. Transcription is not restricted to regions with annotated gene features but includes almost any genomic context. Currently, the source and function of most RNAs originating from intergenic regions in the human genome remain unclear. We hypothesize that many intergenic RNAs can be ascribed to the presence of as-yet unannotated genes or the "fuzzy" transcription of known genes that extends beyond the annotated boundaries. To elucidate the contributions of these two sources, we assemble a dataset of more than 2.5 billion publicly available RNA-seq reads across 5 human cell lines and multiple cellular compartments to annotate transcriptional units in the human genome. About 80% of transcripts from unannotated intergenic regions can be attributed to the fuzzy transcription of existing genes; the remaining transcripts originate mainly from putative long non-coding RNA loci that are rarely spliced. We validate the transcriptional activity of these intergenic RNAs using independent measurements, including transcriptional start sites, chromatin signatures, and genomic occupancies of RNA polymerase II in various phosphorylation states. We also analyze the nuclear localization and sensitivities of intergenic transcripts to nucleases to illustrate that they tend to be rapidly degraded either on-chromatin by XRN2 or off-chromatin by the exosome. We provide a curated atlas of intergenic RNAs that distinguishes between alternative processing of well-annotated genes from independent transcriptional units based on the combined analysis of chromatin signatures, nuclear RNA localization, and degradation pathways.


Factors influencing progress toward sympatric speciation.
Many factors could influence progress towards sympatric speciation. Some of the potentially important ones include competition, mate choice and the degree to which alternative sympatric environments (resources) are discrete. What is not well understood is the relative importance of these different factors, as well as interactions among them. We use an individual-based numerical model to investigate the possibilities. Mate choice was modelled as the degree to which male foraging traits influence female mate choice. Competition was modelled as the degree to which individuals with different phenotypes compete for portions of the resource distribution. Discreteness of the environment was modelled as the degree of bimodality of the underlying resource distribution. We find that strong mate choice was necessary, but not sufficient, to cause sympatric speciation. In addition, sympatric speciation was most likely when the resource distribution was strongly bimodal and when competition among different phenotypes was intermediate. Even under these ideal conditions, however, sympatric speciation occurred only a fraction of the time. Sympatric speciation owing to competition on unimodal resource distributions was also possible, but much less common. In all cases, stochasticity played an important role in determining progress towards sympatric speciation, as evidenced by variation in outcomes among replicate simulations for a given set of parameter values. Overall, we conclude that the nature of competition is much less important for sympatric speciation than is the nature of mate choice and the underlying resource distribution. We argue that an increased understanding of the promoters and inhibitors of sympatric speciation is best achieved through models that simultaneously evaluate multiple potential factors.


Acute coronary insufficiency. An urgent surgical condition.
In 41 of 220 consecutive patients who had a coronary artery bypass operation between July 1973 and March 1974 the operation was for acute coronary insufficiency (recurrent chest pain with transient electrocardiographic changes persisting after admission to hospital). Their mean age was 54 (range 33-70 years). Eleven patients had had angina before, 14 had had at least one myocardial infarction, and 16 presented de novo. Eight of the latter 16 patients required only a single graft, usually to the left anterior descending artery, a significantly greater number than the two of the other 25 patients (P less than 0.01). Fourteen of these 16 patients had normal ventricular contraction, a significantly higher proportion than the 13 of the remaining 25 (p less than 0.05). No collaterals were seen in any of the 10 with single-vessel disease, which was significantly fewer than five out of 18 with double- and nine out of 13 with triple-vessel disease (P less than 0.005). Patients with rapidly developing obstruction, especially in the proximal left anterior descending artery, may not have time to develop collaterals, present acutely with good ventricular function, and may be particularly at risk. There was no operative mortality. The patients had a perioperative myocardial infarction, and there was one late death. At follow-up averaging 9-7 months (range 5-14 months) 32 (80%) patients were angina-free, no myocardial infarctions had occurred, and 85% were fully employed. Urgent coronary artery bypass grafting is a safe and effective treatment for acute coronary insufficiency.


Are community midwives addressing the inequities in access to skilled birth attendance in Punjab, Pakistan? Gender, class and social exclusion.
Pakistan is one of the six countries estimated to contribute to over half of all maternal deaths worldwide. To address its high maternal mortality rate, in particular the inequities in access to maternal health care services, the government of Pakistan created a new cadre of community-based midwives (CMW). A key expectation is that the CMWs will improve access to skilled antenatal and intra-partum care for the poor and disadvantaged women. A critical gap in our knowledge is whether this cadre of workers, operating in the private health care context, will meet the expectation to provide care to the poorest and most marginalized women. There is an inherent paradox between the notions of fee-for-service and increasing access to health care for the poorest who, by definition, are unable to pay. Data will be collected in three interlinked modules. Module 1 will consist of a population-based survey in the catchment areas of the CMW's in districts Jhelum and Layyah in Punjab. Proportions of socially excluded women who are served by CMWs and their satisfaction levels with their maternity care provider will be assessed. Module 2 will explore, using an institutional ethnographic approach, the challenges (organizational, social, financial) that CMWs face in providing care to the poor and socially marginalized women. Module 3 will identify the social, financial, geographical and other barriers to uncover the hidden forces and power relations that shape the choices and opportunities of poor and marginalized women in accessing CMW services. An extensive knowledge dissemination plan will facilitate uptake of research findings to inform positive developments in maternal health policy, service design and care delivery in Pakistan. The findings of this study will enhance understanding of the power dynamics of gender and class that may underlie poor women's marginalization from health care systems, including community midwifery care. One key outcome will be an increased sensitization of the special needs of socially excluded women, an otherwise invisible group. Another expectation is that the poor, socially excluded women will be targeted for provision of maternity care. The research will support the achievement of the 5th Millennium Development Goal in Pakistan.


Viral infection potentiates the increase in airway blood flow produced by substance P.
We examined the effect of respiratory tract infection with Sendai virus on the responsiveness of airway blood flow to substance P (SP) in rats. Pathogen-free rats were inoculated with either Sendai virus suspension or sterile viral growth medium into each nostril. Five days later, we measured airway and esophageal blood flows before and immediately after injection of SP or histamine into the left ventricle of rats in both groups using a modification of the reference-sample microsphere technique. Viral infection potentiated the increase in airway blood flow evoked by SP but not by histamine. We also examined the effect of neutral endopeptidase (NEP) and angiotensin-converting enzyme (ACE) on the SP-induced increase in airway blood flow. Both phosphoramidon (NEP inhibitor) and captopril (ACE inhibitor) potentiated the increase in airway blood flow produced by SP in pathogen-free rats. In the presence of both peptidase inhibitors, a submaximal dose of SP increased blood flow to a similar level in infected and pathogen-free rats. Thus decreased activity of both ACE and NEP may be involved in the exaggerated increase in airway blood flow evoked by SP in virus-infected rats.


Use of 2-azido-3-[125I]iodo-7,8-dibromodibenzo-p-dioxin as a probe to determine the relative ligand affinity of human versus mouse aryl hydrocarbon receptor in cultured cells.
The aryl hydrocarbon receptor (AhR) is a ligand-induced transcription factor that is activated by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and other related compounds, leading to toxicity. There is considerable variation in the response to TCDD among different species, and this may be correlated to differences in the AhR. Variations in the structure of the AhR could result in altered biochemical properties of the receptor, such as ligand affinity or transactivation potential. The difference between the mouse AhR b-1 allele (mAhR(b-1)) and human AhR (hAhR), in terms of their relative affinity for a photoaffinity ligand (2-azido-3-[(125)I]iodo-7,8-dibromodibenzo-p-dioxin), was assessed using both in vitro assays and assays performed directly in cell culture. Results revealed that the hAhR has a lower affinity for the photoaffinity ligand compared with mAhR(b-1). In contrast with a previous study, we found that a single amino acid (valine 381) in hAhR is responsible for the lower ligand affinity, and mutating this residue to alanine results in restoration of high ligand affinity in hAhR. In vitro ligand binding assays are limited by the low concentrations of protein in the assays, and it is not appropriate to compare ligand affinities of different receptors using this method without performing a competition assay or increasing the protein concentration in the assay. Because of the limitation of the in vitro assay, the relative ligand occupancy of mAhR(b-1) and hAhR was compared most effectively within cells, revealing that mAhR(b-1) has a 10-fold higher relative ligand affinity in cells, whereas mAhR(d) has a 2-fold higher relative ligand affinity than hAhR.


Trauma-Informed HIV Prevention and Treatment.
The high prevalence of trauma and its negative impact on health and health-promoting behaviors underscore the need for multi-level interventions to address trauma and its associated sequelae to improve physical and mental well-being in both HIV-infected and HIV-uninfected populations. Growing global awareness of the intersection of trauma and HIV has resulted in development and testing of interventions to address trauma in the context of HIV treatment and HIV prevention in the USA and globally. Despite increasing recognition of the widespread nature of trauma and the importance of trauma to HIV transmission around the globe, several gaps remain. Through a survey of the literature, we identified eight studies (published in the past 5 years) describing interventions to address the effects of trauma on HIV-related outcomes. In particular, this study focused on the levels of intervention, populations the interventions were designed to benefit, and types of trauma addressed in the interventions in the context of both HIV prevention and treatment. Remarkably absent from the HIV prevention, interventions reviewed were interventions designed to address violence experienced by men or transgender individuals, in the USA or globally. Given the pervasive nature of trauma experienced generally, but especially among individuals at heightened risk for HIV, future HIV prevention interventions universally should consider becoming trauma-informed. Widespread acknowledgement of the pervasive impact of gender-based violence on HIV outcomes among women has led to multiple calls for trauma-informed care (TIC) approaches to improve the effectiveness of HIV services for HIV-infected women. TIC approaches may be relevant for and should also be tested among men and all groups with high co-occurring epidemics of HIV and trauma (e.g., men who have sex with men (MSM), transgendered populations, injection drug users, sex workers), regardless of type of trauma experience.


The generation and trapping of solitary waves over topography
Observations of tidal flow over a sill show that upstream solitary wave generation occurred in conjunction with the downstream movement of a streamline bifurcation over the obstacle crest. Solitary waves appeared upstream of the bifurcation, riding on the surface stratification. Shear flow instabilities on the plunging interface were also evident and may contribute to wave generation through subharmonic interaction. Nonlinear waves generated downstream of an internal control became trapped in the supercritical flow.


Familial, structural, and environmental correlates of MRI-defined bone marrow lesions: a sibpair study.
The aim of this study was to estimate the heritability and describe the correlates of bone marrow lesions in knee subchondral bone. A sibpair design was used. T2- and T1-weighted MRI scans were performed on the right knee to assess bone marrow lesions at lateral tibia and femora and medial tibia and femora, as well as chondral defects. A radiograph was taken on the same knee and scored for individual features of osteoarthritis (radiographic osteoarthritis; ROA) and alignment. Other variables measured included height, weight, knee pain, and lower-limb muscle strength. Heritability was estimated with the program SOLAR (Sequential Oligogenetic Linkage Analysis Routines). A total of 115 siblings (60 females and 55 males) from 48 families, representing 95 sib pairs, took part. The adjusted heritability estimates were 53 +/- 28% (mean +/- SEM; p = 0.03) and 65 +/- 32% (p = 0.03) for severity of bone marrow lesions at lateral and medial compartments, respectively. The estimates were reduced by 8 to 9% after adjustment for chondral defects and ROA (but not alignment). The adjusted heritability estimate was 99% for prevalent bone marrow lesions at both lateral and medial compartments. Both lateral and medial bone marrow lesions were significantly correlated with age, chondral defects, and ROA of the knee (all p < 0.05). Medial bone marrow lesions were also more common in males and were correlated with body mass index (BMI). Thus, bone marrow lesions have a significant genetic component. They commonly coexist with chondral defects and ROA but only share common genetic mechanisms to a limited degree. They are also more common with increasing age, male sex, and increasing BMI.


The efficacy and safety of multiple sessions of multisite transcranial random noise stimulation in treating chronic tinnitus.
Random noise stimulation was reported as the more effective and safer type of electrical stimulation techniques in relieving tinnitus symptoms. The multisite protocol of transcranial random noise stimulation has shown additional favorable effects. Here we will discuss the role of applying eight sessions of multisite transcranial random noise stimulation in decreasing tinnitus loudness and annoyance without exerting additional adverse effects. Twenty-nine subjects (8 female), the mean age of (45.34±9.57) with chronic tinnitus received transcranial random noise stimulation in the multisite protocol, 10min of auditory-transcranial random noise stimulation applied over the T3, T4 preceded by 10min of prefrontal-transcranial random noise stimulation applied over F4, FP1. In the first group, only one session was applied and the multiple-sessions group contained eight repeated sessions. Visual analog scale scores for loudness and distress were recorded before and immediately after the treatment. Multivariate repeated measure ANOVA test was used and minimal detectable change calculated. There was a statistically and clinically significant reduction in tinnitus loudness and annoyance in both groups (p<0.05, effect size (η2)>0.8), while the amount of annoyance suppression in the multiple-sessions group was significantly greater than the single-session group. The patients of the multiple-sessions transcranial random noise stimulation group reported an improvement in their sleep and lower tinnitus handicap inventory scores without experiencing any additional adverse effects of the intervention. The results of this study showed a substantial improvement in tinnitus symptoms by using the multiple sessions of transcranial random noise stimulation in the multisite protocol without producing any additional side effects. We suggest further clinical trials with long-term follow-up be investigated.


Lactate transport in mammalian ventricle. General properties and relation to K+ fluxes.
Net cellular L-lactate efflux associated with accelerated anaerobic glycolysis has been implicated as a potential cause of the marked cellular K+ loss contributing to lethal cardiac arrhythmias in ischemic heart and to impaired function of fatigued skeletal muscle. To examine the mechanisms of transsarcolemmal L-lactate movement in the heart, isolated guinea pig ventricular myocytes were loaded with the fluorescent H+ or K+ indicators, carboxy SNARF-1 or PBFI, respectively, under whole-cell patch-clamp conditions. With H+ as the only permeable monovalent cation, a rapid increase in extracellular L-lactate concentration ([L-]o) from 0 to 30 mmol/L at constant pHo (7.35) caused an intracellular acidification averaging 0.18 +/- 0.02 pH units in 60 seconds (n = 7), reflecting L-lactate influx in association with H+ influx (or OH- efflux). Under voltage-clamp conditions, no significant electrogenic current was associated with H(+)-coupled L-lactate influx, and membrane potential (-75 to +75 mV) had no effect on the degree of acidification produced by 30 mmol/L [L-]o, indicating that L-lactate influx was predominantly nonelectrogenic. Acidification in response to increased [L-]o was saturable (Km, approximately 5 mmol/L), partially stereospecific for L-lactate over D-lactate, and inhibited by 55 +/- 7% and 82 +/- 7% by the monocarboxylate carrier inhibitors alpha-cyano-4-hydroxycinnamate and mersalyl acid, respectively, consistent with a carrier-mediated transport mechanism. Extracellular K+ inhibited H(+)-coupled L-lactate influx by 36 +/- 2%, suggesting that K+ either inhibited or substituted for H+ in cotransport with L-lactate. However, in myocytes loaded with PBFI, no significant increase in [K+]i was detected during exposure to 30 mmol/L [L-]o, suggesting that only a minor component, if any, of L-lactate influx was cotransported or codiffused with K+.


Cardiotoxicity of anthracycline in young breast cancer female patients: the possibility of detection of early cardiotoxicity by TDI.
Tissue Doppler imaging (TDI) was investigated for its applicability for detecting cardiac function and early cardiotoxicity in breast cancer patients treated with anthracyclines. A total of 40 women (age range 18 to 65 years) were enrolled, who had not received anthracyclines previously and had normal systolic and diastolic cardiac function. All healthy patients in the control group were of the same age. Each patient underwent not only standard echocardiographic measurements (ventricular dimensions and wall thickness, ejection fraction, E-wave deceleration time (DT), E/A ratio), but also specific imagings (E-septum separation, pulmonary venous flow), and in addition the myocardial velocity of many segments of mitral anulus obtained with pulsed wave tissue Doppler imaging were performed during the one year of observation period. Based on the results we found that systolic left ventricular function did not change significantly - neither in the study nor in the control group. Diastolic left ventricular function was impaired in 39 patients (97.5%), and 30 (75%) of these showed clear changes by means of both the traditional E/A ratio and TDI. Diastolic dysfunction in 9 patients (22.5%), however, could be detected only with TDI. The analysis of myocardial velocity in different segments showed that diastolic dysfunction does not develop in a homogeneous way but in a different way in segments. Diastolic function was intact in the control group during the study. The detectable myocardial damage occurred in the study group of young female patients without risk factors as a result of one year anthracycline therapy was so severe that the possible outcome might be serious congestive heart failure or death. Our results confirmed our assumptions that TDI is a more precise and useful examination method than traditional ones (E/A ratio or deceleration time) to demonstrate isolated diastolic dysfunction as a result of chemotherapy. Relevant extra information might be given by TDI compared to parameters describing diastolic functions depending on several changing values. TDI may become a regularly and more widely used noninvasive method to detect subclinical cardiotoxicity emerging after chemotherapy.


Cannabidiol Modulates Cytokine Storm in Acute Respiratory Distress Syndrome Induced by Simulated Viral Infection Using Synthetic RNA.
Introduction: In the absence of effective antivirals and vaccination, the pandemic of COVID-19 remains the most significant challenge to our health care system in decades. There is an urgent need for definitive therapeutic intervention. Clinical reports indicate that the cytokine storm associated with acute respiratory distress syndrome (ARDS) is the leading cause of mortality in severe cases of some respiratory viral infections, including COVID-19. In recent years, cannabinoids have been investigated extensively due to their potential effects on the human body. Among all cannabinoids, cannabidiol (CBD) has demonstrated potent anti-inflammatory effects in a variety of pathological conditions. Therefore, it is logical to explore whether CBD can reduce the cytokine storm and treat ARDS. Materials and Methods: In this study, we show that intranasal application of Poly(I:C), a synthetic analogue of viral double-stranded RNA, simulated symptoms of severe viral infections inducing signs of ARDS and cytokine storm. Discussion: The administration of CBD downregulated the level of proinflammatory cytokines and ameliorated the clinical symptoms of Poly I:C-induced ARDS. Conclusion: Our results suggest a potential protective role for CBD during ARDS that may extend CBD as part of the treatment of COVID-19 by reducing the cytokine storm, protecting pulmonary tissues, and re-establishing inflammatory homeostasis.


The many landscapes of recombination in Drosophila melanogaster.
Recombination is a fundamental biological process with profound evolutionary implications. Theory predicts that recombination increases the effectiveness of selection in natural populations. Yet, direct tests of this prediction have been restricted to qualitative trends due to the lack of detailed characterization of recombination rate variation across genomes and within species. The use of imprecise recombination rates can also skew population genetic analyses designed to assess the presence and mode of selection across genomes. Here we report the first integrated high-resolution description of genomic and population variation in recombination, which also distinguishes between the two outcomes of meiotic recombination: crossing over (CO) and gene conversion (GC). We characterized the products of 5,860 female meioses in Drosophila melanogaster by genotyping a total of 139 million informative SNPs and mapped 106,964 recombination events at a resolution down to 2 kilobases. This approach allowed us to generate whole-genome CO and GC maps as well as a detailed description of variation in recombination among individuals of this species. We describe many levels of variation in recombination rates. At a large-scale (100 kb), CO rates exhibit extreme and highly punctuated variation along chromosomes, with hot and coldspots. We also show extensive intra-specific variation in CO landscapes that is associated with hotspots at low frequency in our sample. GC rates are more uniformly distributed across the genome than CO rates and detectable in regions with reduced or absent CO. At a local scale, recombination events are associated with numerous sequence motifs and tend to occur within transcript regions, thus suggesting that chromatin accessibility favors double-strand breaks. All these non-independent layers of variation in recombination across genomes and among individuals need to be taken into account in order to obtain relevant estimates of recombination rates, and should be included in a new generation of population genetic models of the interaction between selection and linkage.


Visual Tracking via Dynamic Memory Networks.
Template-matching methods for visual tracking have gained popularity recently due to their good performance and fast speed. However, they lack effective ways to adapt to changes in the target object's appearance, making their tracking accuracy still far from state-of-the-art. In this paper, we propose a dynamic memory network to adapt the template to the target's appearance variations during tracking. The reading and writing process of the external memory is controlled by an LSTM network with the search feature map as input. A spatial attention mechanism is applied to concentrate the LSTM input on the potential target as the location of the target is at first unknown. To prevent aggressive model adaptivity, we apply gated residual template learning to control the amount of retrieved memory that is used to combine with the initial template. In order to alleviate the drift problem, we also design a "negative" memory unit that stores templates for distractors, which are used to cancel out wrong responses from the object template. To further boost the tracking performance, an auxiliary classification loss is added after the feature extractor part. Unlike tracking-by-detection methods where the object's information is maintained by the weight parameters of neural networks, which requires expensive online fine-tuning to be adaptable, our tracker runs completely feed-forward and adapts to the target's appearance changes by updating the external memory. Moreover, the capacity of our model is not determined by the network size as with other trackers - the capacity can be easily enlarged as the memory requirements of a task increase, which is favorable for memorizing long-term object information. Extensive experiments on the OTB and VOT datasets demonstrate that our trackers perform favorably against state-of-the-art tracking methods while retaining real-time speed.


Open-label placebo treatment does not enhance cognitive abilities in healthy volunteers.
The use of so-called 'smart drugs' such as modafinil to improve cognitive performance has recently attracted considerable attention. However, their side effects have limited user enthusiasm. Open-label placebo (OLP) treatment, i.e., inert treatments that are openly disclosed to individuals as having no active pharmacological ingredient, has been shown to improve various medical symptoms and conditions, including those related to cognitive performance. OLP treatment could therefore be an exciting alternative to pharmacological cognitive enhancers. Here, we used a randomized-controlled design to investigate the effect of a 21-day OLP treatment on several sub-domains of cognitive performance in N = 78 healthy volunteers. Subjective and objective measures of cognitive performance as well as different measures of well-being were obtained before and after the treatment period. Using a combination of classic Frequentist and Bayesian analysis approaches showed no additional benefit from OLP treatment in any of the subjective or objective measures of cognitive performance. Our study thus highlights possible limitations of OLP treatment in boosting cognitive performance in healthy volunteers. These findings are discussed in the light of expectancy-value considerations that may determine OLP efficacy.


Cyclic AMP-dependent phosphodiesterase isozyme-specific potentiation by protein kinase C in hypertrophic cardiomyopathic hamster hearts.
We recently reported that protein kinase C (PKC) potentiates cAMP-dependent phosphodiesterase (PDE) activity in Syrian hamster hearts with hypertrophic cardiomyopathy (HCM) but not in control hamster hearts. In this study, we examined the mechanism of this PKC/PDE interaction by identifying the PDE isozyme that is the target of PKC modulation. Using Mono-Q high performance liquid chromatography, both control and HCM hamster cardiac PDE could be partially purified into the calcium/calmodulin-dependent (I), the cGMP-stimulated (II), and the low KM (III) isozyme fractions. The elution profiles of PDE isozyme fractions were similar to those in isolated hamster cardiac myocytes. The percentages of PDE isozymes activities (I/II/III) were 68.8:22.4:8.8% and 51.1:38.4:10.5% for HCM and control hearts, respectively (n = 4), suggesting a change in the quantitative expression of isozymes activities in HCM hearts with a significant increase in the calcium/calmodulin-dependent PDE isozyme activities (p < 0.05 compared with control). The addition of exogenous PKC (100 munits of rat brain) produced a 60% stimulation in the calcium/calmodulin-dependent PDE isozyme fraction but not in other PDE isozymes of HCM and in none of the isozymes in control hearts. This PKC-mediated potentiation of the calcium/calmodulin-dependent PDE activity was completely blocked by the PKC-specific peptide inhibitor PKC(19-31). Analysis of enzymatic kinetics showed that PKC enhanced the calcium/calmodulin-dependent PDE isozyme activity in HCM by increasing its Vmax (from 350 pmol/mg/min at baseline to 758 pmol/min/mg with PKC) without changing its KM (0.69 microM at baseline versus 0.89 microM with PKC). These results suggest that there are both quantitative and qualitative abnormalities in the expression of the calcium/ calmodulin-dependent PDE isozyme in HCM hearts and that the PKC modulation of PDE activity in the HCM heart is isozyme specific.


Characterization of GABAA receptor ligands with automated patch-clamp using human neurons derived from pluripotent stem cells.
Automated patch clamp is a recent but widely used technology to assess pre-clinical drug safety. With the availability of human neurons derived from pluripotent stem cells, this technology can be extended to determine CNS effects of drug candidates, especially those acting on the GABAA receptor. iCell Neurons (Cellular Dynamics International, A Fujifilm Company) were cultured for ten days and analyzed by patch clamp in the presence of agonist GABA or in combination with positive allosteric GABAA receptor modulators. Both efficacy and affinity were determined. In addition, mRNA of GABAA receptor subunits were quantified by qRT-PCR. We have shown that iCell Neurons are compatible with the IonFlux microfluidic system of the automated patch clamp instrument. Resistance ranging from 15 to 25MΩ was achieved for each trap channel of patch clamped cells in a 96-well plate format. GABA induced a robust change of current with an EC50 of 0.43μM. Positive GABAA receptor modulators diazepam, HZ-166, and CW-04-020 exhibited EC50 values of 0.42μM, 1.56μM, and 0.23μM, respectively. The α2/α3/α5 selective compound HZ-166-induced the highest potentiation (efficacy) of 810% of the current induced by 100nM GABA. Quantification of GABAA receptor mRNA in iCell Neurons revealed high levels of α5 and β3 subunits and low levels of α1, which is similar to the configuration in human neonatal brain. iCell Neurons represent a new cellular model to characterize GABAergic compounds using automated patch clamp. These cells have excellent representation of cellular GABAA receptor distribution that enable determination of total small molecule efficacy and affinity as measured by cell membrane current change.


Candidate lung tumor susceptibility genes identified through whole-genome association analyses in inbred mice.
We performed a whole-genome association analysis of lung tumor susceptibility using dense SNP maps ( approximately 1 SNP per 20 kb) in inbred mice. We reproduced the pulmonary adenoma susceptibility 1 (Pas1) locus identified in previous linkage studies and further narrowed this quantitative trait locus (QTL) to a region of less than 0.5 Mb in which at least two genes, Kras2 (Kirsten rat sarcoma oncogene 2) and Casc1 (cancer susceptibility candidate 1; also known as Las1), are strong candidates. Casc1 knockout mouse tumor bioassays showed that Casc1-deficient mice were susceptible to chemical induction of lung tumors. We also found three more genetic loci for lung adenoma development. Analysis of one of these candidate loci identified a previously uncharacterized gene Lasc1, bearing a nonsynonymous substitution (D102E). We found that the Lasc1 Glu102 allele preferentially promotes lung tumor cell growth. Our findings demonstrate the prospects for using dense SNP maps in laboratory mice to refine previous QTL regions and identify genetic determinants of complex traits.


Models of maternity care: evidence for midwifery continuity of care.
There has been substantial reform in the past decade in the provision of maternal and child health services, and specifically regarding models of maternity care. Increasingly, midwives are working together in small groups to provide midwife-led continuity of care. This article reviews the current evidence for models of maternity care that provide midwifery continuity of care, in terms of their impact on clinical outcomes, the views of midwives and childbearing women, and health service costs. A systematic review of midwife-led continuity of care models identified benefits for women and babies, with no adverse effects. Non-randomised studies have shown benefits of midwifery continuity of care for specific groups, such as Aboriginal and Torres Strait Islander women. There are also benefits for midwives, including high levels of job satisfaction and less occupational burnout. Implementing midwifery continuity of care in public and private settings in Australia has been challenging, despite the evidence in its favour and government policy documents that support it. A reorganisation of the way maternity services are provided in Australia is required to ensure that women across the country can access this model of care. Critical to such reform is collaboration with obstetricians, general practitioners, paediatricians and other medical professionals involved in the care of pregnant women, as well as professional respect for the central role of midwives in the provision of maternity care. More research is needed into ways to ensure that all childbearing women can access midwifery continuity of care.


Transcriptomic profiling to identify genes involved in Fusarium mycotoxin Deoxynivalenol and Zearalenone tolerance in the mycoparasitic fungus Clonostachys rosea.
Clonostachys rosea strain IK726 is a mycoparasitic fungus capable of controlling mycotoxin-producing Fusarium species, including F. graminearum and F. culmorum, known to produce Zearalenone (ZEA) and Deoxynivalenol (DON). DON is a type B trichothecene known to interfere with protein synthesis in eukaryotes. ZEA is a estrogenic-mimicing mycotoxin that exhibits antifungal growth. C. rosea produces the enzyme zearalenone hydrolase (ZHD101), which degrades ZEA. However, the molecular basis of resistance to DON in C. rosea is not understood. We have exploited a genome-wide transcriptomic approach to identify genes induced by DON and ZEA in order to investigate the molecular basis of mycotoxin resistance C. rosea. We generated DON- and ZEA-induced cDNA libraries based on suppression subtractive hybridization. A total of 443 and 446 sequenced clones (corresponding to 58 and 65 genes) from the DON- and ZEA-induced library, respectively, were analysed. DON-induced transcripts represented genes encoding metabolic enzymes such as cytochrome P450, cytochrome c oxidase and stress response proteins. In contrast, transcripts encoding the ZEA-detoxifying enzyme ZHD101 and those encoding a number of ATP-Binding Cassette (ABC) transporter transcripts were highly frequent in the ZEA-induced library. Subsequent bioinformatics analysis predicted that all transcripts with similarity to ABC transporters could be ascribed to only 2 ABC transporters genes, and phylogenetic analysis of the predicted ABC transporters suggested that they belong to group G (pleiotropic drug transporters) of the fungal ABC transporter gene family. This is the first report suggesting involvement of ABC transporters in ZEA tolerance. Expression patterns of a selected set of DON- and ZEA-induced genes were validated by the use of quantitative RT-PCR after exposure to the toxins. The qRT-PCR results obtained confirm the expression patterns suggested from the EST redundancy data. The present study identifies a number of transcripts encoding proteins that are potentially involved in conferring resistance to DON and ZEA in the mycoparasitic fungus C. rosea. Whilst metabolic readjustment is potentially the key to withstanding DON, the fungus produces ZHD101 to detoxify ZEA and ABC transporters to transport ZEA or its degradation products out from the fungal cell.


Cloning, pharmacological characterization and distribution of a novel galanin receptor.
The neuropeptide galanin mediates a diverse spectrum of biological activities by interacting with specific G-protein-coupled receptors. Through expression cloning, human and rat GALR1 receptor cDNA clones have previously been isolated and characterized. In this study, we have used homology screening to isolate a rat brain cDNA clone encoding a second galanin receptor subtype, the GALR2 receptor. The isolated cDNA encodes a 372-amino-acid G-protein-coupled receptor that shares 38% overall amino-acid identity with the rat GALR1 receptor. The pharmacological profile of the rat GALR2 receptor is similar to that of the rat GALR1 receptor. The rat GALR2 receptor binds galanin, N-terminal galanin fragments, and the putative galanin receptor antagonists galantide, C7, M35 and M40 with high affinity but it does not bind C-terminal galanin fragments. Galanin increases intracellular inositol phosphate levels in HEK 293 cells expressing the rat GALR2 receptor via a pertussis toxin-insensitive G-protein. The rat GALR2 receptor mRNA is highly expressed in several brain regions, including hypothalamus and hippocampus as well as the anterior pituitary, with lower levels of expression detected in amygdala, and regions of cortex. It is also highly expressed in the GH3 pituitary cell line and in gut tissues, and to a lower extent in spleen, lung, skeletal muscle, heart, kidney, liver and testis. These results suggest that GALR2 receptor mediates galanin's regulation of pituitary hormone secretion and possibly food intake.


Heart-lung interactions during mechanical ventilation: the basics.
The hemodynamic effects of mechanical ventilation can be grouped into three clinically relevant concepts. First, since spontaneous ventilation is exercise. In patients increased work of breathing, initiation of mechanical ventilatory support may improve O2 delivery because the work of breathing is reduced. Second, changes in lung volume alter autonomic tone, pulmonary vascular resistance, and at high lung volumes compress the heart in the cardiac fossa similarly to cardiac tamponade. As lung volume increases so does the pressure difference between airway and pleural pressure. When this pressure difference exceeds pulmonary artery pressure, pulmonary vessels collapse as they pass form the pulmonary arteries into the alveolar space increasing pulmonary vascular resistance. Hyperinflation increases pulmonary vascular resistance impeding right ventricular ejection. Anything that over distends lung units will increase their vascular resistance, and if occurring globally throughout the lung, increase pulmonary vascular resistance. Decreases in end-expiratory lung volume cause alveolar collapse increases pulmonary vasomotor tone by the process of hypoxic pulmonary vasoconstriction. Recruitment maneuvers that restore alveolar oxygenation without over distention will reduce pulmonary artery pressure. Third, positive-pressure ventilation increases intrathoracic pressure. Since diaphragmatic descent increases intra-abdominal pressure, the decrease in the pressure gradient for venous return is less than would otherwise occur if the only change were an increase in right atrial pressure. However, in hypovolemic states, it can induce profound decreases in venous return. Increases in intrathoracic pressure decreases left ventricular afterload and will augment left ventricular ejection. In patients with hypervolemic heart failure, this afterload reducing effect can result in improved left ventricular ejection, increased cardiac output and reduced myocardial O2 demand. This brief review will focus primarily on mechanical ventilation and intrathoracic pressure as they affect right and left ventricular function and cardiac output.


Torsion-vibration coupling in S1 toluene: Implications for IVR, the torsional barrier height, and rotational constants.
We have examined the S1←S0 transition of toluene in the region from the 0(0)(0) band to ∼210 cm(-1) above it. The spectrum reveals methyl rotor levels of 0(0) toluene up to m = 6 and of the lowest frequency vibration, 20(1), up to m = 4. The rotor levels of both 20(1) and 0(0) are perturbed by torsion-vibration coupling. The inclusion of torsion-vibration coupling leads to the S1 torsional barrier, V6, being revised from -26.376 cm(-1) to -5.59 cm(-1). The torsion-vibration coupling constant is determined to be 21.1 cm(-1). This situation is the S1 analogue of that recently reported for S0 toluene [Gascooke et al., J. Chem. Phys. 142, 024315 (2015)]. Torsion-vibration coupling alters both the rotor band positions and the rotational contours, which particularly affects the rotational constants associated with motion around the a-axis, about which the methyl group rotates. Every vibrational state (indicated generically by X) will be involved in the corresponding X - X20(1) torsion-vibration coupling; so, this interaction permeates the vib-rotor manifold, providing a mechanism to enhance intramolecular vibrational energy redistribution.


orf4 of the Bacillus cereus sigB gene cluster encodes a general stress-inducible Dps-like bacterioferritin.
The function of orf4 in the sigB cluster in Bacillus cereus ATCC 14579 remains to be explored. Amino-acid sequence analysis has revealed that Orf4 is homologous with bacterioferritins and Dps. In this study, we generated an orf4-null mutant and produced recombinant protein rOrf4 to establish the role of orf4. In vitro, the purified rOrf4 was found to exist in two distinct forms, a dimeric form and a polymer form, through size exclusion analysis. The latter form exhibited a unique filament structure, in contrast to the typical spherical tetracosamer structure of bacterioferritins; the former can be induced to form rOrf4 polymers immediately after the addition of FeCl(2). Catalysis of the oxidation of ferrous irons by ferroxidase activity was detected with rOrf4, and the mineralized irons were subsequently sequestered only in the rOrf4 polymer. Moreover, rOrf4 exerted DNA-protective activity against oxidative damage via DNA binding in a nonspecific manner, as is seen with Dps. In vivo, deletion of orf4 had no effect on activation of the alternative sigma factor sigma(B), and therefore, orf4 is not associated with sigma(B) regulation; however, orf4 can be significantly upregulated upon environmental stress but not H(2)O(2) treatment. B. cereus strains with constitutive Orf4 expression exhibited a viability higher than that of the orf4-null mutant, under specific oxidative stress or heat shock. Taken together, these results suggest that Orf4 functions as a Dps-like bacterioferritin in response to environmental stress and can provide cell protection from oxidative damage through iron sequestration and DNA binding.


Serial changes of elevated serum IgG4 levels in IgG4-related systemic disease.
Autoimmune pancreatitis (AIP) and Mikulicz's disease have recently been recognized as pancreatic or salivary gland lesions of IgG4-related systemic disease. These are frequently associated with elevated serum IgG4 levels. This study aimed to clarify clinical implications of serial changes of elevated serum IgG4 levels in IgG4-related systemic diseases. Serial changes of elevated serum IgG4 levels were examined in patients with IgG4-related systemic diseases. Patients Serial changes of elevated serum IgG4 levels were examined in 44 patients: AIP (n=24), Mikulicz's disease (n=8), pancreatic cancer (n=5), bile duct cancer (n=1), sclerosing cholangitis (n=1), hypereosinophilic syndrome (n=1), chronic thyroiditis (n=1), hypophysitis (n=1), idiopathic pancreatitis (n=1), and Behcet's disease (n=1). The serum IgG4 levels decreased in all patients with AIP and Mikulicz's disease after steroid therapy. The serum IgG4 levels were normalized in 46% of AIP patients and 38% of Mikulicz's disease patients. The serum IgG4 levels were not normalized at remission in 3 of 4 relapsed AIP patients, and re-elevation of serum IgG4 levels was detected in all relapsed patients. Elevated serum IgG4 levels decreased in 3 patients with pancreatic cancer after resection or chemotherapy, and decreased in patients with hypereosinophilic syndrome, sclerosing cholangitis, and hypophysitis after steroid therapy. Measurement of serial serum IgG4 levels is useful to determine the disease activity of IgG4-related systemic diseases.


Targeted amplicon deep sequencing of ama1 and mdr1 to track within-host P. falciparum diversity throughout treatment in a clinical drug trial.
Antimalarial therapeutic efficacy studies are routinely conducted in malaria-endemic countries to assess the effectiveness of antimalarial treatment strategies. Targeted amplicon sequencing (AmpSeq) uniquely identifies and quantifies genetically distinct parasites within an infection. In this study, AmpSeq of Plasmodium falciparum apical membrane antigen 1 ( ama1), and multidrug resistance gene 1 ( mdr1), were used to characterise the complexity of infection (COI) and drug-resistance genotypes, respectively. P. falciparum-positive samples were obtained from a triple artemisinin combination therapy clinical trial conducted in 30 children under 13 years of age between 2018 and 2019 in Kilifi, Kenya. Nine of the 30 participants presented with recurrent parasitemia from day 26 (624h) onwards. The ama1 and mdr1 genes were amplified and sequenced, while msp1, msp2 and glurp data were obtained from the original clinical study. The COI was comparable between ama1 and msp1, msp2 and glurp; overall, ama1 detected more microhaplotypes. Based on ama1, a stable number of microhaplotypes were detected throughout treatment until day 3. Additionally, a recrudescent infection was identified with an ama1 microhaplotype initially observed at 30h and later in an unscheduled follow-up visit. Using the relative frequencies of ama1 microhaplotypes and parasitemia, we identified a fast (<1h) and slow (>5h) clearing microhaplotype. As expected, only two mdr1 microhaplotypes (NF and NY) were identified based on the combination of amino acid polymorphisms at codons 86 and 184. This study highlights AmpSeq as a tool for highly-resolution tracking of parasite microhaplotypes throughout treatment and can detect variation in microhaplotype clearance estimates. AmpSeq can also identify slow-clearing microhaplotypes, a potential early sign of selection during treatment. Consequently, AmpSeq has the capability of improving the discriminatory power to distinguish recrudescences from reinfections accurately.


Modeling the relative dynamics of DNA-coated colloids.
We construct a theoretical model for the dynamics of a microscale colloidal particle, modeled as an interval, moving horizontally on a DNA-coated surface, modelled as a line coated with springs that can stick to the interval. Averaging over the fast DNA dynamics leads to an evolution equation for the particle in isolation, which contains both friction and diffusion. The DNA-induced friction coefficient depends on the physical properties of the DNA, and substituting parameter values typical of a 1 μm colloid coated densely with weakly interacting DNA gives a coefficient about 100 times larger than the corresponding coefficient of hydrodynamic friction. We use a mean-field extension of the model to higher dimensions to estimate the friction tensor for a disc rotating and translating horizontally along a line. When the DNA strands are very stiff and short, the friction coefficient for the disc rolling approaches zero while the friction for the disc sliding remains large. Together, these results could have significant implications for the dynamics of DNA-coated colloids or other ligand-receptor systems, implying that DNA-induced friction between colloids can be stronger than hydrodynamic friction and should be incorporated into simulations, and that it depends nontrivially on the type of relative motion, possibly causing the particles to assemble into out-of-equilibrium metastable states governed by the pathways with the least friction.


A CTP-dependent archaeal riboflavin kinase forms a bridge in the evolution of cradle-loop barrels.
Proteins of the cradle-loop barrel metafold are formed by duplication of a conserved betaalphabeta-element, suggesting a common evolutionary origin from an ancestral group of nucleic acid-binding proteins. The basal fold within this metafold, the RIFT barrel, is also found in a wide range of enzymes, whose homologous relationship with the nucleic acid-binding group is unclear. We have characterized a protein family that is intermediate in sequence and structure between the basal group of cradle-loop barrels and one family of RIFT-barrel enzymes, the riboflavin kinases. We report the structure, substrate-binding mode, and catalytic activity for one of these proteins, Methanocaldococcus jannaschii Mj0056, which is an archaeal riboflavin kinase. Mj0056 is unusual in utilizing CTP rather than ATP as the donor nucleotide, and sequence conservation in the relevant residues suggests that this is a general feature of archaeal riboflavin kinases.


Gene coexpression network analysis of fruit transcriptomes uncovers a possible mechanistically distinct class of sugar/acid ratio-associated genes in sweet orange.
The ratio of sugars to organic acids, two of the major metabolites in fleshy fruits, has been considered the most important contributor to fruit sweetness. Although accumulation of sugars and acids have been extensively studied, whether plants evolve a mechanism to maintain, sense or respond to the fruit sugar/acid ratio remains a mystery. In a prior study, we used an integrated systems biology tool to identify a group of 39 acid-associated genes from the fruit transcriptomes in four sweet orange varieties (Citrus sinensis L. Osbeck) with varying fruit acidity, Succari (acidless), Bingtang (low acid), and Newhall and Xinhui (normal acid). We reanalyzed the prior sweet orange fruit transcriptome data, leading to the identification of 72 genes highly correlated with the fruit sugar/acid ratio. The majority of these sugar/acid ratio-related genes are predicted to be involved in regulatory functions such as transport, signaling and transcription or encode enzymes involved in metabolism. Surprisingly, only three of these sugar/acid ratio-correlated genes are weakly correlated with sugar level and none of them overlaps with the acid-associated genes. Weighted Gene Coexpression Network Analysis (WGCNA) has revealed that these genes belong to four modules, Blue, Grey, Brown and Turquoise, with the former two modules being unique to the sugar/acid ratio control. Our results indicate that orange fruits contain a possible mechanistically distinct class of genes that may potentially be involved in maintaining fruit sugar/acid ratios and/or responding to the cellular sugar/acid ratio status. Therefore, our analysis of orange transcriptomes provides an intriguing insight into the potentially novel genetic or molecular mechanisms controlling the sugar/acid ratio in fruits.


Dectin-1 regulates IL-10 production via a MSK1/2 and CREB dependent pathway and promotes the induction of regulatory macrophage markers.
In response to infection by fungal pathogens, the innate immune system recognises specific fungal pathogen associated molecular patterns (PAMPs) via pattern recognition receptors including the C-type lectin dectin-1 and members of the Toll Like Receptor (TLR) family. Stimulation of these receptors leads to the induction of both pro- and anti-inflammatory cytokines. The protein kinases MSK1 and 2 are known to be important in limiting inflammatory cytokine production by macrophages in response to the TLR4 agonist LPS. In this study we show that MSKs are also activated in macrophages by the fungal derived ligand zymosan, as well as the dectin-1 specific agonists curdlan and depleted zymosan, via the ERK1/2 and p38α MAPK pathways. Furthermore, we show that MSKs regulate dectin-1 induced IL-10 production, and that this regulation is dependent on the ability of MSKs to phosphorylate the transcription factor CREB. IL-10 secreted in response to zymosan was able to promote STAT3 phosphorylation via an autocrine feedback loop. Consistent with the decreased IL-10 secretion in MSK1/2 knockout macrophages, these cells also had decreased STAT3 tyrosine phosphorylation relative to wild type controls after stimulation with zymosan. We further show that the reduction in IL-10 production in the MSK1/2 macrophages results in increased secretion of IL-12p40 in response to zymosan relative to wild type controls. The production of high levels of IL-10 but low levels of IL-12 has previously been associated with an M2b or 'regulatory' macrophage phenotype, which was initially described in macrophages stimulated with a combination of immune complexes and LPS. We found that zymosan, via dectin-1 activation, also leads to the expression of SphK1 and LIGHT, markers of a regulatory like phenotype in mouse macrophages. The expression of these makers was further reinforced by the high level of IL-10 secreted in response to zymosan stimulation.


Modulation of forskolin binding to rat brain membranes.
High affinity binding sites for [3H]forskolin have been identified in rat brain membranes. These sites have a Kd of 15 nM and a Bmax of about 200 fmol/mg protein. The binding of [3H]forskolin to those high affinity sites in rat brain membranes is increased about two-fold by addition of MgCl2 or MnCl2. Smaller increases are observed in the presence of calcium, sodium, or potassium. The binding of [3H]forskolin is also increased in the presence of NaF or GppNHp, agents that are known to activate adenylate cyclase through the stimulatory guanine nucleotide regulatory protein (Ns). The increase in [3H]forskolin binding in the presence of NaF or GppNHp is due to an increase in the number of binding sites with no change in the apparent Kd for the binding sites. The NaF- and GppNHp-stimulated binding requires the presence of magnesium or manganese. The binding of [3H]forskolin to rat brain membranes is reduced in membranes that are heated or pretreated with chymotrypsin, trypsin, or N-ethylmaleimide. NaF stabilizes the binding sites to thermal denaturation. The data demonstrate that the number of high affinity forskolin binding sites are increased under conditions that promote the activation of the catalytic protein of adenylate cyclase by the Ns protein. It is suggested that the high affinity forskolin binding sites are associated with a complex of the catalytic protein and the activated Ns protein.


Endothelial and platelet microparticles in vasculitis of the young.
Microparticles are released from endothelial cells in response to a variety of injurious stimuli and recently have been shown to be increased in a number of diseases associated with endothelial dysfunction. This study examined endothelial microparticle (EMP) and platelet microparticle (PMP) profiles in children with systemic vasculitis to test the hypothesis that EMPs may provide a noninvasive means of examining endothelial activation or injury. The study cohort comprised 39 children with systemic vasculitis at various stages of disease activity, 24 control children with febrile disease, and a control group of 43 healthy subjects. Plasma was ultracentrifuged at 17,000g for 60 minutes, and the microparticle pellets were examined using flow cytometry. Plasma from patients with active systemic vasculitis contained significantly higher numbers of E-selectin-positive EMPs compared with that from patients in remission, healthy controls, or febrile disease controls (P = 0.000 for each). A similar result was obtained for the numbers of EMPs expressing the marker CD105. There was also a significant increase in PMPs expressing CD42a in the active vasculitis group as compared with the other groups, but this difference was not significant for PMPs expressing P-selectin. The EMP counts correlated with the Birmingham Vasculitis Activity Score and the acute-phase reactant levels in the patients with systemic vasculitis, but there was a poor correlation overall between EMP counts and the acute-phase reactant levels in the febrile disease controls. EMPs may provide a window to the activated endothelium and could provide important pathophysiologic insights into the vascular injury associated with vasculitis of the young.


Attitudes and opinions of faculty tutors about problem-based learning.
There has been little systematic attention to the opinions of faculty in evaluating problem-based learning (PBL). The purpose of this report is to describe the attitudes and opinions of tutors in PBL programs about the relative merits of PBL and traditional medical education, and to examine the influences of selected variables on these attitudes. Questionnaires containing both Likert-type and open-ended questions were sent to 1,287 faculty members who had served as PBL tutors at 22 U.S. and Canadian medical schools during the 1992-93 academic year. All schools with identifiable PBL programs (broadly defined) were included in the survey. The overall response rate was 69% (882 of 1,287). As a group, the respondents were experienced in both PBL and traditional curricula, with an average of 3.75 years of experience in the former and 11.41 years of experience in the latter. Regarding both their overall attitudes and their opinions about seven of nine specific areas, the respondents evaluated PBL more positively than traditional methods. This was especially evident in the ratings of student interest and enthusiasm, faculty interest and enthusiasm, the respondents' personal satisfaction, student reasoning, and preparation for clinical rotations. The two methods were judged to be approximately equally efficient for learning. Traditional methods were judged to be superior for teaching factual knowledge of basic sciences. PBL was particularly popular with faculty in PBL-track programs, with faculty in both the newest and the oldest PBL programs, and with faculty in either primary care or "nontraditional" specialties. The findings confirm and extend the picture of PBL strengths and weaknesses that can be derived from prior anecdotal program descriptions and small sample studies. Experienced faculty seem to prefer PBL in most respects, although they have some serious reservations. The present findings also agree with prior outcome studies of PBL, suggesting that the most strongly held faculty opinions may have a factual basis.


Elaborations on Corallopyronin A as a Novel Treatment Strategy Against Genital Chlamydial Infections.
Ascending Chlamydia trachomatis infection causes functional damage to the fallopian tubes, which may lead to ectopic pregnancy and infertility in women. Treatment failures using the standard regimens of doxycycline and azithromycin have been observed. We tested the polyketide-derived α-pyrone antibiotic Corallopyronin A (CorA) that inhibits the bacterial DNA dependent RNA polymerase and has strong activity against various extracellular and some intracellular bacteria. Extensive testing in cell culture infection models and in an ex vivo human fallopian tube model under different oxygen concentrations was performed to assess the anti-chlamydial efficacy of CorA at physiological conditions. CorA showed high efficacy against C. trachomatis (MICN/H: 0.5 μg/mL for serovar D and L2), C. muridarum (MICN/H: 0.5 μg/mL), and C. pneumoniae (MICN/H: 1 μg/mL) under normoxic (N) and hypoxic (H) conditions. Recoverable inclusion forming units were significantly lower already at 0.25 μg/mL for all tested chlamydiae. CorA at a concentration of 1 μg/mL was also effective against already established C. trachomatis and C. pneumoniae infections (up to 24 h.p.i.) in epithelial cells, while efficacy against C. muridarum was limited to earlier time points. A preliminary study using a C. muridarum genital infection model revealed corresponding limitations in the efficacy. Importantly, in an ex vivo human fallopian tube model, the growth of C. trachomatis was significantly inhibited by CorA at concentrations of 1-2 μg/mL under normoxic and hypoxic conditions. The overall high efficacies of CorA against C. trachomatis in cell culture and an ex vivo human fallopian tube model under physiological oxygen concentrations qualifies this drug as a candidate that should be further investigated.


Targeting Macrophages Sensitizes Chronic Lymphocytic Leukemia to Apoptosis and Inhibits Disease Progression.
The role of monocytes/macrophages in the development and progression of chronic lymphocytic leukemia (CLL) is poorly understood. Transcriptomic analyses show that monocytes/macrophages and leukemic cells cross talk during CLL progression. Macrophage depletion impairs CLL engraftment, drastically reduces leukemic growth, and favorably impacts mouse survival. Targeting of macrophages by either CSF1R signaling blockade or clodrolip-mediated cell killing has marked inhibitory effects on established leukemia also. Macrophage killing induces leukemic cell death mainly via the TNF pathway and reprograms the tumor microenvironment toward an antitumoral phenotype. CSF1R inhibition reduces leukemic cell load, especially in the bone marrow, and increases circulating CD20(+) leukemic cells. Accordingly, co-targeting TAMs and CD20-expressing leukemic cells provides a survival benefit in the mice. These results establish the important role of macrophages in CLL and suggest therapeutic strategies based on interfering with leukemia-macrophage interactions.


Chronic myelogenous leukemia: a multivariate analysis of the associations of patient characteristics and therapy with survival.
The prognostic importance of patient pretreatment clinical and laboratory features was investigated in a group of 303 patients with Philadelphia chromosome-positive benign-phase chronic myelogenous leukemia. Intensive chemotherapy was given to 97 patients, and 78 underwent an early elective splenectomy. The overall median survival time, dated from hospital admission, was 39 months. Patient characteristics associated with shortened survival were age 60 years or older, black race, the presence of hepatomegaly, splenomegaly, symptoms, weight loss, and poor performance status. Adverse blood and bone marrow parameters were anemia, thrombocytosis or thrombocytopenia, a high proportion of peripheral blasts plus promyelocytes or of basophils, a high proportion of marrow blasts or basophils, decreased marrow megakaryocytes, and cytogenetic abnormalities in addition to the Philadelphia chromosome. Several of these factors were interrelated. A multivariate regression analysis demonstrated that the combination blood basophilia, race, additional cytogenetic abnormalities, age and marrow basophilia had the strongest predictive relationship to survival time. This resulted in a model segregating patients into low-, intermediate-, and high-risk groups, with median survivals of 53, 39, and 25 months, respectively. Another model was derived that did not include the marrow features and identified splenomegaly and platelet counts as adding to the prognosis prediction by blood basophilia, race, and age. Evaluation of the effect of therapy, after adjusting for differences in prognostic characteristics, showed that intensive chemotherapy was associated with survival prolongation among patients at intermediate and high risk of death. We conclude that a combination of pretreatment factors identifies different risk subcategories in patients with chronic myelogenous leukemia and is helpful in assessing overall prognosis and treatment effect.


Flow-through anterior thigh flaps with a short pedicle for reconstruction of lower leg and foot defects.
New flow-through perforator flaps with a large, short vascular pedicle are proposed because of their clinical significance and a high success rate for reconstruction of the lower legs. Of 13 consecutive cases, the authors describe two cases of successful transfer of a new short-pedicle anterolateral or anteromedial thigh flow-through flap for coverage of soft-tissue defects in the legs. This new flap has a thin fatty layer and a small fascial component, and is vascularized with a perforator originating from a short segment of the descending branch of the lateral circumflex femoral system. The advantages of this flap are as follows: flow-through anastomosis ensures a high success rate for free flaps and preserves the recipient arterial flow; there is no need for dissecting throughout the lateral circumflex femoral system as the pedicle vessel; minimal time is required for flap elevation; there is minimal donor-site morbidity; and the flap is obtained from a thin portion of the thigh. Even in obese patients, thinning of the flap with primary defatting is possible, and the donor scar is concealed. This flap is suitable for coverage of defects in legs where a single arterial flow remains. It is also suitable for chronic lower leg ulcers, osteomyelitis, and plantar coverage.


Degradation paths of manganese-based MOF materials in a model oxidative environment: a computational study.
Stability is the key property of functional materials. In this work we investigate computationally the degradative potential of a model Mn-BTC (BTC = benzene-1,3,5-tricarboxylate) metal-organic framework (MOF) building block in aqueous solutions under oxidative conditions. Model density functional theory calculations have shown that the direct hydrolysis of the Mn-containing moieties is more difficult than their decomposition via oxidation-induced paths. While the interaction with H2O2 species is of non-covalent nature and requires O-O-bond breaking to initiate Mn-center oxidation, open-shell O2 species readily oxidize radical Mn-centers and form bonds of σ-, π-, or δ-symmetry with the metal. The oxidative transformations of di-Mn paddle-wheel carboxylate structure-forming units are accompanied with substantial distortions of the coordination polyhedra that, together with the increased Lewis acidity of the oxidized metal centers, facilitates the hydrolysis leading to the degradation of the structure at a larger scale. Whereas such a mechanism is expected to hamper the catalytic applications of such Mn-MOFs, the associated structural response to oxidizing and radical species can create a basis for the construction of Mn-MOF-based drug delivery systems with increased bio-compatibility.


Purification and 1H NMR spectroscopic characterization of phase II metabolites of tolfenamic acid.
Tolfenamic acid, an anti-inflammatory drug (NSAID), is metabolized in vivo to form several oxidative metabolites which are all conjugated with beta-D-glucuronic acid. In this study, the metabolites of tolfenamic acid were identified by 1H nuclear magnetic resonance (NMR) spectroscopy in urine samples obtained on days 7 to 10 from a human volunteer after oral administration of 200 mg of the drug three times per day (steady-state plasma concentration). The metabolites of tolfenamic acid were initially concentrated by preparative solid phase extraction (PSPE) chromatography, thereby removing the endogenous polar compounds that are present in the urine. The individual metabolites were purified by preparative high performance liquid chromatography (HPLC) and then identified using 1H NMR. Both one- and two-dimensional NMR experiments were performed to identify the phase II metabolites of tolfenamic acid; the study shows the applicability of 1H NMR for the identification of drug metabolites in biological fluids. In addition to NMR analysis, two metabolites were also identified by mass spectrometry (MS). The glucuronides of the following parent compounds, N-(2-methyl-3-chlorophenyl)-anthranilic acid (T), N-(2-hydroxymethyl-3-chlorophenyl)-anthranilic acid (1), N-(2-hydroxymethyl-3-chloro-4-hydroxyphenyl)-anthranilic acid (2), N-(2-formyl-3-chlorophenyl) anthranilic acid (3), N-(2-methyl-3-chloro-4-hydroxyphenyl)-anthranilic acid (4), N-(2-methyl-3-chloro-5-hydroxyphenyl)-anthranilic acid (5), N-(2-carboxy-3-chlorophenyl)-anthranilic acid (6), N-(2-hydroxymethyl-3-chlorophenyl)-4-hydroxy-anthranilic acid (7), N-(2-methyl-3-chlorophenyl)-5-hydroxy-anthranilic acid (8), N-(2-methyl-3-chloro-4-metoxyphenyl)-anthranilic acid (9), N-(2-methyl-3-chlorophenyl)-4-hydroxy-anthranilic acid (10), and N-(2-methyl-4-hydroxyphenyl)-anthranilic acid (11) were identified. The phase II metabolites (5-11) had not previously been identified in urine from humans administered tolfenamic acid. The phase I metabolites of the glucuronides 7, 8, 10, and 11 were identified here for the first time. An HPLC method was developed that simultaneously separates all the phase II metabolites identified as well as some phase I metabolites in urine samples obtained after intake of tolfenamic acid.


The effects of psilocybin on cognitive and emotional functions in healthy participants: Results from a phase 1, randomised, placebo-controlled trial involving simultaneous psilocybin administration and preparation.
Psilocybin, a psychoactive serotonin receptor partial agonist, has been reported to acutely reduce clinical symptoms of depressive disorders. Psilocybin's effects on cognitive function have not been widely or systematically studied. The aim of this study was to explore the safety of simultaneous administration of psilocybin to healthy participants in the largest randomised controlled trial of psilocybin to date. Primary and secondary endpoints assessed the short- and longer-term change in cognitive functioning, as assessed by a Cambridge Neuropsychological Test Automated Battery (CANTAB) Panel, and emotional processing scales. Safety was assessed via endpoints which included cognitive function, assessed by CANTAB global composite score, and treatment-emergent adverse event (TEAE) monitoring. In this phase 1, randomised, double-blind, placebo-controlled study, healthy participants (n = 89; mean age 36.1 years; 41 females, 48 males) were randomised to receive a single oral dose of 10 or 25 mg psilocybin, or placebo, administered simultaneously to up to six participants, with one-to-one psychological support - each participant having an assigned, dedicated therapist available throughout the session. In total, 511 TEAEs were reported, with a median duration of 1.0 day; 67% of all TEAEs started and resolved on the day of administration. There were no serious TEAEs, and none led to study withdrawal. There were no clinically relevant between-group differences in CANTAB global composite score, CANTAB cognitive domain scores, or emotional processing scale scores. These results indicate that 10 mg and 25 mg doses of psilocybin were generally well tolerated when given to up to six participants simultaneously and did not have any detrimental short- or long-term effects on cognitive functioning or emotional processing. EudraCT (https://www.clinicaltrialsregister.eu/) number: 2018-000978-30.


Certification of caffeine reference material purity by ultraviolet/visible spectrophotometry and high-performance liquid chromatography with diode-array detection as two independent analytical methods.
Caffeine reference material certified for purity is produced worldwide, but no research work on the details of the certification process has been published in the literature. In this paper, we report the scientific details of the preparation and certification of pure caffeine reference materials. Caffeine was prepared by extraction from roasted and ground coffee by dichloromethane after heating in deionized water mixed with magnesium oxide. The extract was purified, dried, and bottled in dark glass vials. Stratified random selection was applied to select a number of vials for homogeneity and stability studies, which revealed that the prepared reference material is homogeneous and sufficiently stable. Quantification of caffeine purity % was carried out using a calibrated UV/visible spectrophotometer and a calibrated high-performance liquid chromatography with diode-array detection method. The results obtained from both methods were combined to drive the certified value and its associated uncertainty. The certified value of the reference material purity was found to be 99.86% and its associated uncertainty was ±0.65%, which makes the candidate reference material a very useful calibrant in food and drug chemical analysis.


Association Between Alcohol Use Disorders and Outcomes of Patients Hospitalized With Community-Acquired Pneumonia.
Patients with alcohol use disorder (AUD) are at elevated risk of developing pneumonia, but few studies have assessed the outcomes of pneumonia in patients with AUD. To compare the causes, treatment, and outcomes of pneumonia in patients with and without AUD and to understand the associations of comorbid illnesses, alcohol withdrawal, and any residual effects due to alcohol itself with patient outcomes. A retrospective cohort study was conducted of 137 496 patients 18 years or older with pneumonia who were admitted to 177 US hospitals participating in the Premier Healthcare Database from July 1, 2010, to June 30, 2015. Statistical analysis was conducted from October 27, 2017, to August 20, 2018. Alcohol use disorders identified from International Classification of Diseases, Ninth Revision, Clinical Modification codes. Pneumonia cause, antibiotic treatment, inpatient mortality, clinical deterioration, length of stay, and cost. Associations of AUD with these variables were studied using generalized linear mixed models. Of 137 496 patients with community-acquired pneumonia (70 358 women and 67 138 men; mean [SD] age, 69.5 [16.2] years), 3.5% had an AUD. Patients with an AUD were younger than those without an AUD (median age, 58.0 vs 73.0 years; P < .001), more often male (77.3% vs 47.8%; P < .001), and more often had principal diagnoses of aspiration pneumonia (10.9% vs 9.8%; P < .001), sepsis (38.6% vs 30.7%; P < .001), or respiratory failure (9.3% vs 5.5%; P < .001). Their cultures more often grew Streptococcus pneumoniae (43.7% vs 25.5%; P < .001) and less frequently grew organisms resistant to guideline-recommended antibiotics (25.0% vs 43.7%; P < .001). Patients with an AUD were treated more often with piperacillin-tazobactam (26.2% vs 22.5%; P < .001) but equally as often with anti-methicillin-resistant Staphylococcus aureus agents (32.9% vs 31.8%; P = .11) compared with patients without AUDs. When adjusted for demographic characteristics and insurance, AUD was associated with higher mortality (odds ratio, 1.40; 95% CI, 1.25-1.56), length of stay (risk-adjusted geometric mean ratio, 1.24; 95% CI, 1.20-1.27), and costs (risk-adjusted geometric mean ratio, 1.33; 95% CI, 1.28-1.38). After additional adjustment for differences in comorbidities and risk factors for resistant organisms, AUD was no longer associated with mortality but remained associated with late mechanical ventilation (odds ratio, 1.28; 95% CI, 1.12-1.46), length of stay (risk-adjusted geometric mean ratio, 1.04; 95% CI, 1.01-1.06), and costs (risk-adjusted geometric mean ratio, 1.06; 95% CI, 1.03-1.09). Models segregating patients undergoing alcohol withdrawal showed that poorer outcomes among patients with AUD were confined to the subgroup undergoing alcohol withdrawal. This study suggests that, compared with hospitalized patients with community-acquired pneumonia but without AUD, those with AUD less often harbor resistant organisms. The higher age-adjusted risk of death among patients with AUD appears to be largely attributable to differences in comorbidities, whereas greater use of health care resources may be attributable to alcohol withdrawal.


Effect of tailoring anticoagulant treatment duration by applying a recurrence risk prediction model in patients with venous thromboembolism compared to usual care: A randomized controlled trial.
Patients with unprovoked (i.e., without the presence of apparent transient risk factors such as recent surgery) venous thromboembolism (VTE) are at risk of recurrence if anticoagulants are stopped after 3-6 months, yet their risk remains heterogeneous. Thus, prolonging anticoagulant treatment should be considered in high-risk patients, whereas stopping is likely preferred in those with a low predicted risk. The Vienna Prediction Model (VPM) could aid clinicians in estimating this risk, yet its clinical effects and external validity are currently unknown. The aim of this study was to investigate the clinical impact of this model on reducing recurrence risk in patients with unprovoked VTE, compared to usual care. In a randomized controlled trial, the decision to prolong or stop anticoagulant treatment was guided by predicted recurrence risk using the VPM (n = 441), which was compared with usual care (n = 442). Patients with unprovoked VTE were recruited from local thrombosis services in the Netherlands (in Utrecht, Harderwijk, Ede, Amersfoort, Zwolle, Hilversum, Rotterdam, Deventer, and Enschede) between 22 July 2011 and 30 November 2015, with 24-month follow-up complete for all patients by early 2018. The primary outcome was recurrent VTE during 24 months of follow-up. Secondary outcomes included major bleeding and clinically relevant non-major (CRNM) bleeding. In the total study population of 883 patients, mean age was 55 years, and 507 (57.4%) were men. A total of 96 recurrent VTE events (10.9%) were observed, 46 in the intervention arm and 50 in the control arm (risk ratio 0.92, 95% CI 0.63-1.35, p = 0.67). Major bleeding occurred in 4 patients, 2 in each treatment arm, whereas CRNM bleeding occurred in 20 patients (12 in intervention arm versus 8 in control arm). The VPM showed good discriminative power (c-statistic 0.76, 95% CI 0.69-0.83) and moderate to good calibration, notably at the lower spectrum of predicted risk. For instance, in 284 patients with a predicted risk of >2% to 4%, the observed rate of recurrence was 2.5% (95% CI 0.7% to 4.3%). The main limitation of this study is that it did not enroll the preplanned number of 750 patients in each study arm due to declining recruitment rate. Our results show that application of the VPM in all patients with unprovoked VTE is unlikely to reduce overall recurrence risk. Yet, in those with a low predicted risk of recurrence, the observed rate was also low, suggesting that it might be safe to stop anticoagulant treatment in these patients. Netherlands Trial Register NTR2680.


Using serological measures to monitor changes in malaria transmission in Vanuatu.
With renewed interest in malaria elimination, island environments present unique opportunities to achieve this goal. However, as transmission decreases, monitoring and evaluation programmes need increasingly sensitive tools to assess Plasmodium falciparum and Plasmodium vivax exposure. In 2009, to assess the role of serological markers in evaluating malaria transmission, a cross-sectional seroprevalence study was carried out in Tanna and Aneityum, two of the southernmost islands of the Vanuatu archipelago, areas where malaria transmission has been variably reduced over the past few decades. Malaria transmission was assessed using serological markers for exposure to P. falciparum and P. vivax. Filter blood spot papers were collected from 1,249 people from Tanna, and 517 people from Aneityum to assess the prevalence of antibodies to two P. falciparum antigens (MSP-119 and AMA-1) and two P. vivax antigens (MSP-119 and AMA-1). Age-specific prevalence was modelled using a simple catalytic conversion model based on maximum likelihood to generate a community seroconversion rate (SCR). Overall seropositivity in Tanna was 9.4%, 12.4% and 16.6% to P. falciparum MSP-119, AMA-1 and Schizont Extract respectively and 12.6% and 15.0% to P. vivax MSP-119 and AMA-1 respectively. Serological results distinguished between areas of differential dominance of either P. vivax or P. falciparum and analysis of age-stratified results showed a step in seroprevalence occurring approximately 30 years ago on both islands, indicative of a change in transmission intensity at this time. Results from Aneityum suggest that several children may have been exposed to malaria since the 2002 P. vivax epidemic. Seroepidemiology can provide key information on malaria transmission for control programmes, when parasite rates are low. As Vanuatu moves closer to malaria elimination, monitoring changes in transmission intensity and identification of residual malaria foci is paramount in order to concentrate intervention efforts.


The relative efficacy of 3 different freehand frontal ventriculostomy trajectories: a prospective neuronavigation-assisted simulation study.
OBJECTIVE Ventriculostomy is a relatively common neurosurgical procedure, often performed in the setting of acute hydrocephalus. Accurate positioning of the catheter is vital to minimize morbidity and mortality, and several anatomical landmarks are currently used. The aim of this study was to prospectively evaluate the relative performance of 3 recognized trajectories for frontal ventriculostomy using imaging-derived metrics: perpendicular to skull (PTS), contralateral medial canthus/external auditory meatus (CMC/EAM), and ipsilateral medial canthus/external auditory meatus (IMC/EAM). METHODS Participants completed 9 simulated ventriculostomy attempts (3 of each trajectory) on a model head with Medtronic StealthStation coregistered imaging. Performance measures were distance of the ventricular catheter tip to the foramen of Monro (FoM) and presence of the catheter tip in a lateral ventricle. RESULTS Thirty-one individuals of varying seniority and prior ventriculostomy experience performed a total of 279 simulated freehand frontal ventriculostomies. The PTS and CMC/EAM trajectories were found to be significantly more likely to result in both the catheter tip being closer to the FoM and in a lateral ventricle compared with the IMC/EAM trajectory. These findings were not influenced by the prior ventriculostomy experience of the participant, corroborating the significance of these results. CONCLUSIONS The PTS and CMC/EAM trajectories were superior to the IMC/EAM trajectories during freehand frontal ventriculostomy in this study, and further data from studies incorporating varying ventricular sizes and bur hole locations are required to facilitate a change in clinical practice. In addition, neuronavigation and other guidance techniques for ventriculostomy are becoming increasingly popular and may be superior to freehand techniques, necessitating further prospective data evaluating their safety, efficacy, and feasibility for routine clinical use.