A novel Alzheimer's disease drug candidate targeting inflammation and fatty acid metabolism.
CAD-31 is an Alzheimer's disease (AD) drug candidate that was selected on the basis of its ability to stimulate the replication of human embryonic stem cell-derived neural precursor cells as well as in APPswe/PS1ΔE9 AD mice. To move CAD-31 toward the clinic, experiments were undertaken to determine its neuroprotective and pharmacological properties, as well as to assay its therapeutic efficacy in a rigorous mouse model of AD. CAD-31 has potent neuroprotective properties in six distinct nerve cell assays that mimic toxicities observed in the old brain. Pharmacological and preliminary toxicological studies show that CAD-31 is brain-penetrant and likely safe. When fed to old, symptomatic APPswe/PS1ΔE9 AD mice starting at 10 months of age for 3 additional months in a therapeutic model of the disease, there was a reduction in the memory deficit and brain inflammation, as well as an increase in the expression of synaptic proteins. Small-molecule metabolic data from the brain and plasma showed that the major effect of CAD-31 is centered on fatty acid metabolism and inflammation. Pathway analysis of gene expression data showed that CAD-31 had major effects on synapse formation and AD energy metabolic pathways. All of the multiple physiological effects of CAD-31 were favorable in the context of preventing some of the toxic events in old age-associated neurodegenerative diseases.


Suppressing N-Myc downstream regulated gene 1 reactivates senescence signaling and inhibits tumor growth in hepatocellular carcinoma.
Hepatocellular carcinoma (HCC) is the fifth most lethal malignancy worldwide with no curative therapies. To discover potentially novel therapeutic targets for HCC, we previously studied the gene expression profiles of HCC patients and identified that significant upregulation of N-Myc downstream regulated gene 1 (NDRG1) is associated with more aggressive phenotypes and poorer overall survival of HCC patients. In this study, we further used a loss-of-function approach (RNA interference) to understand the role of NDRG1 in hepatocarcinogenesis. We found that suppression of NDRG1 significantly impaired HCC cell growth through inducing extensive cellular senescence of HCC cells both in vitro and in vivo, accompanied by cell cycle arrest at the G1 phase. The observed antitumor effects of NDRG1 suppression were correlated with activation of major senescence-associated signaling pathways, such as upregulation of tumor suppressors p53, p21 and p16, and decreased phosphorylated Rb. To obtain further insights into the clinical significance of NDRG1-modulated senescence in HCC patients, immunohistochemistry staining of 92 cases of HCC patients was done. We found that high NDRG1 expression (n = 66) is associated with low p21 (n = 82; P < 0.001) and low p16 (n = 86; P < 0.001) levels. In conclusion, this study demonstrated that NDRG1 is a potential therapeutic target for HCC because its suppression triggers senescence of HCC cells through activating glycogen synthase kinase-3β-p53 pathway, thereby inhibiting tumor progression.


Improving the stability of thermophilic anaerobic digesters treating SS-OFMSW through enrichment with compost and leachate seeds.
This paper examines the potential of improving the stability of thermophilic anaerobic digestion of source-sorted organic fraction of municipal solid waste (SS-OFMSW) by adding leachate and compost during inoculation. For this purpose, two stable thermophilic digesters, A (control) and B (with added leachate and compost), were subjected to a sustained substrate shock by doubling the organic loading rate for one week. Feeding was suspended then gradually resumed to reach the pre-shock loading rate (2 gVS/l/d). Digester A failed, exhibiting excessive increase in acetate and a corresponding decrease in pH and methane generation, and lower COD and solids removal efficiencies. In contrast, digester B was able to restore its functionality with 90% recovery of pre-shock methane generation rate at stable pH, lower hydrogen levels, and reduced VFAs and ammonia accumulation.


Toward a Neurobiological Basis for Understanding Learning in University Modeling Instruction Physics Courses.
Modeling Instruction (MI) for University Physics is a curricular and pedagogical approach to active learning in introductory physics. A basic tenet of science is that it is a model-driven endeavor that involves building models, then validating, deploying, and ultimately revising them in an iterative fashion. MI was developed to provide students a facsimile in the university classroom of this foundational scientific practice. As a curriculum, MI employs conceptual scientific models as the basis for the course content, and thus learning in a MI classroom involves students appropriating scientific models for their own use. Over the last 10 years, substantial evidence has accumulated supporting MI's efficacy, including gains in conceptual understanding, odds of success, attitudes toward learning, self-efficacy, and social networks centered around physics learning. However, we still do not fully understand the mechanisms of how students learn physics and develop mental models of physical phenomena. Herein, we explore the hypothesis that the MI curriculum and pedagogy promotes student engagement via conceptual model building. This emphasis on conceptual model building, in turn, leads to improved knowledge organization and problem solving abilities that manifest as quantifiable functional brain changes that can be assessed with functional magnetic resonance imaging (fMRI). We conducted a neuroeducation study wherein students completed a physics reasoning task while undergoing fMRI scanning before (pre) and after (post) completing a MI introductory physics course. Preliminary results indicated that performance of the physics reasoning task was linked with increased brain activity notably in lateral prefrontal and parietal cortices that previously have been associated with attention, working memory, and problem solving, and are collectively referred to as the central executive network. Critically, assessment of changes in brain activity during the physics reasoning task from pre- vs. post-instruction identified increased activity after the course notably in the posterior cingulate cortex (a brain region previously linked with episodic memory and self-referential thought) and in the frontal poles (regions linked with learning). These preliminary outcomes highlight brain regions linked with physics reasoning and, critically, suggest that brain activity during physics reasoning is modifiable by thoughtfully designed curriculum and pedagogy.


Assessment of efficacy of intervention in hearing impaired children with speech and language deficits.
The diagnosis of hearing loss in children with speech and language deficits by the otorhinolaryngologist involves a large allocation of sources. The ability to assess the efficacy of intervention in order to minimize speech and language deficits is an aspect in the care of these children which has not been generally considered as part of the child's ongoing medical care. These children have speech and language deficits predominantly from hearing impairment and occasionally from a primary language disorder. The effect of intervention (usually a hearing aid), the special education program for the hearing impaired child, and the language therapy for the child with a primary language disorder are seldom measured. The ability to assess and monitor the child's progress is essential for the care of these patients. The continual monitoring of the child's progress is accomplished by periodic assessment of the child's speech and language. An instrument to do this has been developed and used by the Department of Otorhinolaryngectomy at the Albert Einstein College of Medicine for a number of years. The utilization of this instrument, as demonstrated by a series of case reports, will be presented. These will include children in which intervention was successful and those for whom it was unsuccessful, with an analysis of the reasons underlying the effectiveness for each child. It is recommended that every child with a speech and language deficit should be monitored periodically so that the child's progress, or lack of progress, can be determined. If the child is not progressing, additional remediation can be instituted to attempt to prevent a permanent speech and language deficiency.


Success and failure of controlling the real-time functional magnetic resonance imaging neurofeedback signal are reflected in the striatum.
Over the last decades, neurofeedback has been applied in variety of research contexts and therapeutic interventions. Despite this extensive use, its neural mechanisms are still under debate. Several scientific advances have suggested that different networks become jointly active during neurofeedback, including regions generally involved in self-regulation, regions related to the specific mental task driving the neurofeedback and regions generally involved in feedback learning (Sitaram et al., 2017, Nature Reviews Neuroscience, 18, 86). To investigate the neural mechanisms specific to neurofeedback but independent from general effects of self-regulation, we compared brain activation as measured with functional magnetic resonance imaging (fMRI) across different mental tasks involving gradual self-regulation with and without providing neurofeedback. Ten participants freely chose one self-regulation task and underwent two training sessions during fMRI scanning, one with and one without receiving neurofeedback. During neurofeedback sessions, feedback signals were provided in real-time based on activity in task-related, individually defined target regions. In both sessions, participants aimed at reaching and holding low, medium, or high brain-activation levels in the target region. During gradual self-regulation with neurofeedback, a network of cortical control regions as well as regions implicated in reward and feedback processing were activated. Self-regulation with feedback was accompanied by stronger activation within the striatum across different mental tasks. Additional time-resolved single-trial analysis revealed that neurofeedback performance was positively correlated with a delayed brain response in the striatum that reflected the accuracy of self-regulation. Overall, these findings support that neurofeedback contributes to self-regulation through task-general regions involved in feedback and reward processing.


Dynamic coupling of voltage sensor and gate involved in closed-state inactivation of kv4.2 channels.
Voltage-gated potassium channels related to the Shal gene of Drosophila (Kv4 channels) mediate a subthreshold-activating current (I(SA)) that controls dendritic excitation and the backpropagation of action potentials in neurons. Kv4 channels also exhibit a prominent low voltage-induced closed-state inactivation, but the underlying molecular mechanism is poorly understood. Here, we examined a structural model in which dynamic coupling between the voltage sensors and the cytoplasmic gate underlies inactivation in Kv4.2 channels. We performed an alanine-scanning mutagenesis in the S4-S5 linker, the initial part of S5, and the distal part of S6 and functionally characterized the mutants under two-electrode voltage clamp in Xenopus oocytes. In a large fraction of the mutants (>80%) normal channel function was preserved, but the mutations influenced the likelihood of the channel to enter the closed-inactivated state. Depending on the site of mutation, low-voltage inactivation kinetics were slowed or accelerated, and the voltage dependence of steady-state inactivation was shifted positive or negative. Still, in some mutants these inactivation parameters remained unaffected. Double mutant cycle analysis based on kinetic and steady-state parameters of low-voltage inactivation revealed that residues known to be critical for voltage-dependent gate opening, including Glu 323 and Val 404, are also critical for Kv4.2 closed-state inactivation. Selective redox modulation of corresponding double-cysteine mutants supported the idea that these residues are involved in a dynamic coupling, which mediates both transient activation and closed-state inactivation in Kv4.2 channels.


Prediction of DNA binding motifs from 3D models of transcription factors; identifying TLX3 regulated genes.
Proper cell functioning depends on the precise spatio-temporal expression of its genetic material. Gene expression is controlled to a great extent by sequence-specific transcription factors (TFs). Our current knowledge on where and how TFs bind and associate to regulate gene expression is incomplete. A structure-based computational algorithm (TF2DNA) is developed to identify binding specificities of TFs. The method constructs homology models of TFs bound to DNA and assesses the relative binding affinity for all possible DNA sequences using a knowledge-based potential, after optimization in a molecular mechanics force field. TF2DNA predictions were benchmarked against experimentally determined binding motifs. Success rates range from 45% to 81% and primarily depend on the sequence identity of aligned target sequences and template structures, TF2DNA was used to predict 1321 motifs for 1825 putative human TF proteins, facilitating the reconstruction of most of the human gene regulatory network. As an illustration, the predicted DNA binding site for the poorly characterized T-cell leukemia homeobox 3 (TLX3) TF was confirmed with gel shift assay experiments. TLX3 motif searches in human promoter regions identified a group of genes enriched in functions relating to hematopoiesis, tissue morphology, endocrine system and connective tissue development and function.


Loss of BMPR2 leads to high bone mass due to increased osteoblast activity.
Imbalances in the ratio of bone morphogenetic protein (BMP) versus activin and TGFβ signaling are increasingly associated with human diseases yet the mechanisms mediating this relationship remain unclear. The type 2 receptors ACVR2A and ACVR2B bind BMPs and activins but the type 2 receptor BMPR2 only binds BMPs, suggesting that type 2 receptor utilization might play a role in mediating the interaction of these pathways. We tested this hypothesis in the mouse skeleton, where bone mass is reciprocally regulated by BMP signaling and activin and TGFβ signaling. We found that deleting Bmpr2 in mouse skeletal progenitor cells (Bmpr2-cKO mice) selectively impaired activin signaling but had no effect on BMP signaling, resulting in an increased bone formation rate and high bone mass. Additionally, activin sequestration had no effect on bone mass in Bmpr2-cKO mice but increased bone mass in wild-type mice. Our findings suggest a novel model whereby BMPR2 availability alleviates receptor-level competition between BMPs and activins and where utilization of ACVR2A and ACVR2B by BMPs comes at the expense of activins. As BMP and activin pathway modulation are of current therapeutic interest, our findings provide important mechanistic insight into the relationship between these pathways in human health.


Stakeholders' Decisions and Preferences for the Provision and Use of Medicinal Cannabis: A Scoping Review.
Background: The aim of this scoping review was to examine the extent that stakeholder's decisions about and preferences for the provision and use of medicinal cannabis (MC) had been investigated. We sought to identify which populations were examined, the methods used for eliciting preferences and exploring decisions, and the reported outcomes of studies. Methods: Electronic databases (PubMed, CINAHL, Embase, BSC and PsycINFO) and the reference lists of relevant articles were searched for studies published up to March 2022. Studies were included if stakeholder preferences for MC were (1) the primary focus of the research, or (2) an aspect of a larger preference focus. Studies that (3) described the decisions to use MC were also included. Results: Thirteen studies were reviewed. The population focus of these was primarily patient, with seven studies focused on general patient populations and five studies targeting specific patient populations such as cancer survivors, and people experiencing depression. Methods included health economics preference methods, qualitative interviews, and a single multicriteria decision-making study. Four categories of outcomes were defined and included comparisons of MC with a therapeutic alternative (n=5), preferences for MC attributes (n=5), administration preferences (n=4), and the decision process of users (n=2). Motivation differences in preference were found. Purely medicinal users and novice users place more importance on cannabidiol (CBD) than tetrahydrocannabinol. Overall, inhalation methods of administration were preferred due to quick onset of symptom relief. Price was the greatest influence on choice for recreational/medicinal users, whereas purely medicinal users were less price sensitive for products with higher CBD content. Conclusion: Studies examining public preferences for the provision and use of MC were absent. Revealed preference methods are a useful technique for understanding preferences for characteristics that are difficult to visibly assess such as cannabinoid or strain. The outcomes of symptom-specific multicriteria decision method studies that compare the benefit-safety profiles of commonly used treatments and MC may be a useful decision support tool for health practitioners. Studies with representative samples are needed to understand the impact of age, gender, and race on preferences for MC.


Salicylic acid-dependent restriction of Tomato ringspot virus spread in tobacco is accompanied by a hypersensitive response, local RNA silencing, and moderate systemic resistance.
Tomato ringspot virus (ToRSV, a Nepovirus sp.) systemically infects many herbaceous plants. Viral RNA accumulates in symptomatic leaves and in young, asymptomatic leaves that emerge late in infection. Here, we show that systemic infection by ToRSV is restricted in tobacco. After an initial hypersensitive response in inoculated leaves, only a few plants showed limited systemic symptoms. Viral RNA did not usually accumulate to detectable levels in asymptomatic leaves. ToRSV-derived small-interfering RNAs and PR1a transcripts were only detected in tissues that contained viral RNA, indicating local induction of RNA silencing and salicylic acid (SA)-dependent defense responses. Lesion size and viral systemic spread were reduced with SA pretreatment but enhanced in NahG transgenic lines deficient in SA accumulation, suggesting that SA-dependent mechanisms play a key role in limiting ToRSV spread in tobacco. Restriction of virus infection was enhanced in transgenic lines expressing the P1-HC-Pro suppressor of silencing. Knocking down the SA-inducible RNA-dependent RNA polymerase 1 exacerbated the necrotic reaction but did not affect viral systemic spread. ToRSV-infected tobacco plants were susceptible to reinoculation by ToRSV or Tobacco mosaic virus, although a small reduction in lesion size was observed. This moderate systemic resistance suggests inefficient induction or spread of RNA silencing and systemic acquired resistance signal molecules.


Insights into the molecular evolution of HslU ATPase through biochemical and mutational analyses.
The ATP-dependent HslVU complexes are found in all three biological kingdoms. A single HslV protease exists in each species of prokaryotes, archaea, and eukaryotes, but two HslUs (HslU1 and HslU2) are present in the mitochondria of eukaryotes. Previously, a tyrosine residue at the C-terminal tail of HslU2 has been identified as a key determinant of HslV activation in Trypanosoma brucei and a phenylalanine at the equivalent position to E. coli HslU is found in T. brucei HslU1. Unexpectedly, we found that an F441Y mutation in HslU enhanced the peptidase and caseinolytic activity of HslV in E. coli but it showed partially reduced ATPase and SulA degradation activity. Previously, only the C-terminal tail of HslU has been the focus of HslV activation studies. However, the Pro315 residue interacting with Phe441 in free HslU has also been found to be critical for HslV activation. Hence, our current biochemical analyses explore the importance of the loop region just before Pro315 for HslVU complex functionality. The proline and phenylalanine pair in prokaryotic HslU was replaced with the threonine and tyrosine pair from the functional eukaryotic HslU2. Sequence comparisons between multiple HslUs from three different biological kingdoms in combination with biochemical analysis of E. coli mutants have uncovered important new insights into the molecular evolutionary pathway of HslU.


Present and future association between obesity and hypogonadism in Italian males.
Obesity prevalence is increasing worldwide and it is nowadays considered a real public health problem. Obesity is associated with co-morbidities like cardiovascular diseases (CVD) and type 2 diabetes mellitus (T2DM), furthermore visceral obesity can be related to low testosterone (T) plasma levels. The link between obesity and hypogonadism (HG) is complex and not completely clarified. Current guidelines suggest that screening for HG should be done in subjects with obesity and T2DM. The aim of this evaluation is to assess the estimated actual and future prevalence of obesity and related co-morbidities, in particular HG, in the Italian general population. The Strategyst Consulting Inc. recently completed an epidemiology forecast model for several countries, looking at HG and CV/Metabolic Disease, based on National Health and Nutrition Examination Survey (NHANES) data collected between 1999-2010. Data from NHANES survey were used to evaluate the Italian estimated prevalence of obesity and HG. Results show that obesity estimated prevalence will increase in 2030 also in Italy. In addition, also the prevalence of obese CVD and T2DM subjects will increase too. Even Italian HG prevalence is estimated to increase in the next two decades, irrespective of T threshold considered (< 8, 10 and 12 nmol/L). In obese CVD subjects the relative risk (RR) of developing HG (T < 8 nmol/L) is four times greater than in not-CVD obese subjects (RR = 4.1, 3.1 and 1.9 accordingly to the aforementioned T thresholds for defining HG). Accordingly, the estimated percentage of hypogonadal obese CVD and T2DM subjects will rise in 2030. The Strategyst epidemiology forecast model has allowed to assess the current and future prevalence of obesity and its relative co-morbidities like HG in Italy. Data emerged from this evaluation suggest that obesity and HG prevalence will increase in Italian population and confirm the complex link between adipose tissue and male T levels.


Reptilian-transcriptome v1.0, a glimpse in the brain transcriptome of five divergent Sauropsida lineages and the phylogenetic position of turtles.
Reptiles are largely under-represented in comparative genomics despite the fact that they are substantially more diverse in many respects than mammals. Given the high divergence of reptiles from classical model species, next-generation sequencing of their transcriptomes is an approach of choice for gene identification and annotation. Here, we use 454 technology to sequence the brain transcriptome of four divergent reptilian and one reference avian species: the Nile crocodile, the corn snake, the bearded dragon, the red-eared turtle, and the chicken. Using an in-house pipeline for recursive similarity searches of >3,000,000 reads against multiple databases from 7 reference vertebrates, we compile a reptilian comparative transcriptomics dataset, with homology assignment for 20,000 to 31,000 transcripts per species and a cumulated non-redundant sequence length of 248.6 Mbases. Our approach identifies the majority (87%) of chicken brain transcripts and about 50% of de novo assembled reptilian transcripts. In addition to 57,502 microsatellite loci, we identify thousands of SNP and indel polymorphisms for population genetic and linkage analyses. We also build very large multiple alignments for Sauropsida and mammals (two million residues per species) and perform extensive phylogenetic analyses suggesting that turtles are not basal living reptiles but are rather associated with Archosaurians, hence, potentially answering a long-standing question in the phylogeny of Amniotes. The reptilian transcriptome (freely available at http://www.reptilian-transcriptomes.org) should prove a useful new resource as reptiles are becoming important new models for comparative genomics, ecology, and evolutionary developmental genetics.


Neurodegeneration caused by expression of human truncated tau leads to progressive neurobehavioural impairment in transgenic rats.
Human truncated tau protein is an active constituent of the neurofibrillary degeneration in sporadic Alzheimer's disease. We have shown that modified tau protein, when expressed as a transgene in rats, induced AD characteristic tau cascade consisting of tau hyperphosphorylation, formation of argyrophilic tangles and sarcosyl-insoluble tau complexes. These pathological changes led to the functional impairment characterized by a variety of neurobehavioural symptoms. In the present study we have focused on the behavioural alterations induced by transgenic expression of human truncated tau. Transgenic rats underwent a battery of behavioural tests involving cognitive- and sensorimotor-dependent tasks accompanied with neurological assessment at the age of 4.5, 6 and 9 months. Behavioural examination of these rats showed altered spatial navigation in Morris water maze resulting in less time spent in target quadrant (p<0.05) and fewer crossings over previous platform position (p<0.05) during probe trial. Spontaneous locomotor activity and anxiety in open field was not influenced by transgene expression. However beam walking test revealed that transgenic rats developed progressive sensorimotor disturbances related to the age of tested animals. The disturbances were most pronounced at the age of 9 months (p<0.01). Neurological alterations indicating impaired reflex responses were other added features of behavioural phenotype of this novel transgenic rat. These results allow us to suggest that neurodegeneration, caused by the non-mutated human truncated tau derived from sporadic human AD, result in the neuronal dysfunction consequently leading to the progressive neurobehavioural impairment.


Bradykinin receptor expression and bradykinin-mediated sensitization of human sensory neurons.
Bradykinin is a peptide implicated in inflammatory pain in both humans and rodents. In rodent sensory neurons, activation of B1 and B2 bradykinin receptors induces neuronal hyperexcitability. Recent evidence suggests that human and rodent dorsal root ganglia (DRG), which contain the cell bodies of sensory neurons, differ in the expression and function of key GPCRs and ion channels; whether BK receptor expression and function are conserved across species has not been studied in depth. In this study, we used human DRG tissue from organ donors to provide a detailed characterization of bradykinin receptor expression and bradykinin-induced changes in the excitability of human sensory neurons. We found that B2 and, to a lesser extent, B1 receptors are expressed by human DRG neurons and satellite glial cells. B2 receptors were enriched in the nociceptor subpopulation. Using patch-clamp electrophysiology, we found that acute bradykinin increases the excitability of human sensory neurons, while prolonged exposure to bradykinin decreases neuronal excitability in a subpopulation of human DRG neurons. Finally, our analyses suggest that donor's history of chronic pain and age may be predictors of higher B1 receptor expression in human DRG neurons. Together, these results indicate that acute BK-induced hyperexcitability, first identified in rodents, is conserved in humans and provide further evidence supporting BK signaling as a potential therapeutic target for treating pain in humans.


Benign and malignant soft-tissue tumors: posttreatment MR imaging.
Soft-tissue sarcoma requires aggressive treatment, often with a combination of radiation therapy, chemotherapy, and surgical resection. Even after multimodality treatment, local recurrence is common, and regular follow-up imaging at short intervals is required. Interpretation of posttreatment magnetic resonance (MR) images may be complicated by changes in the surgical bed or treatment field. The challenge of distinguishing posttreatment change from recurrent tumor may be minimized by using an organized, systematic approach to imaging, with emphasis on the patient's clinical and surgical history and a review of pretreatment images. Common changes that result from radiation therapy include soft-tissue trabeculation, increased fatty marrow, and focal marrow abnormalities. Rarely, radiation-induced malignancies may develop within the treatment field. Chemotherapy also influences posttreatment imaging appearance. Occasionally, it causes a substantial increase in tumor size that is a result of chemotherapy-induced hemorrhage. Although myocutaneous flaps used in reconstructive surgery may mimic a mass, they demonstrate time-dependent changes in size, signal intensity, and enhancement on MR images. Recurrent tumor is characterized by the presence of a discrete nodule or mass with signal characteristics that typically mirror those of the original tumor. MR imaging sequences such as unenhanced T1-weighted fat-suppressed and gradient-echo sequences may help differentiate posttreatment hemorrhage from local tumor recurrence. A consistent imaging approach combined with a detailed knowledge of the patient's history, familiarity with pretreatment images, and an understanding of the various posttreatment changes enables optimal monitoring of the treatment bed and maximizes accuracy in the detection of recurrence.


Stellar Shocks from Dark Matter Asteroid Impacts.
Macroscopic dark matter is almost unconstrained over a wide "asteroidlike" mass range, where it could scatter on baryonic matter with geometric cross section. We show that when such an object travels through a star, it produces shock waves that reach the stellar surface, leading to a distinctive transient optical, UV, and x-ray emission. This signature can be searched for on a variety of stellar types and locations. In a dense globular cluster, such events occur far more often than flare backgrounds, and an existing UV telescope could probe orders of magnitude in dark matter mass in one week of dedicated observation.


Lesion analysis of the brain areas involved in language comprehension.
The cortical regions of the brain traditionally associated with the comprehension of language are Wernicke's area and Broca's area. However, recent evidence suggests that other brain regions might also be involved in this complex process. This paper describes the opportunity to evaluate a large number of brain-injured patients to determine which lesioned brain areas might affect language comprehension. Sixty-four chronic left hemisphere stroke patients were evaluated on 11 subtests of the Curtiss-Yamada Comprehensive Language Evaluation - Receptive (CYCLE-R; Curtiss, S., & Yamada, J. (1988). Curtiss-Yamada Comprehensive Language Evaluation. Unpublished test, UCLA). Eight right hemisphere stroke patients and 15 neurologically normal older controls also participated. Patients were required to select a single line drawing from an array of three or four choices that best depicted the content of an auditorily-presented sentence. Patients' lesions obtained from structural neuroimaging were reconstructed onto templates and entered into a voxel-based lesion-symptom mapping (VLSM; Bates, E., Wilson, S., Saygin, A. P., Dick, F., Sereno, M., Knight, R. T., & Dronkers, N. F. (2003). Voxel-based lesion-symptom mapping. Nature Neuroscience, 6(5), 448-450.) analysis along with the behavioral data. VLSM is a brain-behavior mapping technique that evaluates the relationships between areas of injury and behavioral performance in all patients on a voxel-by-voxel basis, similar to the analysis of functional neuroimaging data. Results indicated that lesions to five left hemisphere brain regions affected performance on the CYCLE-R, including the posterior middle temporal gyrus and underlying white matter, the anterior superior temporal gyrus, the superior temporal sulcus and angular gyrus, mid-frontal cortex in Brodmann's area 46, and Brodmann's area 47 of the inferior frontal gyrus. Lesions to Broca's and Wernicke's areas were not found to significantly alter language comprehension on this particular measure. Further analysis suggested that the middle temporal gyrus may be more important for comprehension at the word level, while the other regions may play a greater role at the level of the sentence. These results are consistent with those seen in recent functional neuroimaging studies and offer complementary data in the effort to understand the brain areas underlying language comprehension.


[Possibilities of improving the effectiveness of therapy in patients with chronic cerebral ischemia against the background of COVID-19].
To study the possibility of improving the efficacy of treatment with mexidol in COVID-19 patients with chronic cerebral ischemia (CCI). Three hundred and four patients with CCI and COVID-19 were observed, group 1 (n=152) consisted of patients receiving basic therapy and mexidol, group 2 (n=152) received only basic therapy. Mexidol was administered intravenously for 14 days, 500 mg (10 ml) per 400 ml of saline solution, then Mexidol FORTE 250 was administered in a dose of 250 mg 3 times a day for 2 months. The state of cognitive functions (MoCA scale), sleep (Spiegel questionnaire), asthenia (MFI-20 scale), and quality of life (SIP questionnaire) were evaluated. Examinations were performed before treatment, 30 and 75 days after start of treatment. In group 1, there was a more complete and earlier recovery of the state of cognitive functions (an increase in indicators on the MoCA scale, p<0.01), a regression of asthenia (p<0.05), and normalization of sleep (p<0.01). By the end of the study, there were significantly more patients in group 1 with complete or significant recovery of all quality of life indicators. Long-term sequential therapy with mexidol provides a more complete recovery of impaired functions in patients with CCI and COVID-19.


Body height, body mass index, waist-hip ratio, fluctuating asymmetry and second to fourth digit ratio in subjects with latent toxoplasmosis.
Between 20% and 60% of the population of most countries are infected with the protozoan Toxoplasma gondii. Subjects with clinically asymptomatic life-long latent toxoplasmosis differ from those who are Toxoplasma free in several behavioural parameters. Case-control studies cannot decide whether these differences already existed before infection or whether they were induced by the presence of Toxoplasma in the brain of infected hosts. Here we searched for such morphological differences between Toxoplasma-infected and Toxoplasma-free subjects that could be induced by the parasite (body weight, body height, body mass index, waist-hip ratio), or could rather correlate with their natural resistance to parasitic infection (fluctuating asymmetry, 2D : 4D ratio). We found Toxoplasma-infected men to be taller and Toxoplasma-infected men and women to have lower 2D : 4D ratios previously reported to be associated with higher pre-natal testosterone levels. The 2D : 4D ratio negatively correlated with the level of specific anti-Toxoplasma antibodies in Toxoplasma-free subjects. These results suggest that some of the observed differences between infected and non-infected subjects may have existed before infection and could be caused by the lower natural resistance to Toxoplasma infection in subjects with higher pre-natal testosterone levels.


The Herts and Minds study: feasibility of a randomised controlled trial of Mentalization-Based Treatment versus usual care to support the wellbeing of children in foster care.
There is a lack of well-designed randomized controlled trials (RCTs) to investigate the efficacy of psychological therapies for children in foster care with emotional and behavioural difficulties. Mentalization-based therapy (MBT) focuses on supporting the carer-child relationship by promoting reflective capacity. This study examined the feasibility and acceptability of an RCT of MBT, delivered in a family-format, for children who are in foster care in the UK. Herts and Minds was a phase II, blinded feasibility RCT with follow-up of at 12 and 24 weeks post-randomisation. Participants were children (age 5-16) in foster care referred to a targeted mental health service, who had some level of difficulty as identified by the Strengths and Difficulties Questionnaire (SDQ). Aims were to assess: the feasibility of recruitment processes and study uptake; capacity to train mental health practitioners to deliver MBT to an acceptable level of treatment integrity; establish acceptability and credibility of MBT as an intervention for children in foster care; establish feasibility and acceptability to participants of conducting an RCT; and estimate the likely treatment efficacy effect size. Participants were randomly allocated to either MBT (n = 15) or Usual Clinical Care (UCC) (n = 21) individually or in sibling groups. A range of qualitative and quantitative data was gathered to assess feasibility. Feasibility was established with regard to: capacity to recruit participants to a study; capacity to train mental health practitioners to deliver MBT to an acceptable level of treatment integrity; acceptability and credibility of MBT; and feasibility and acceptability to participants of conducting an RCT. A number of issues made it difficult to estimate a likely treatment efficacy effect size. With modifications, it is feasible to run an RCT of MBT for children in foster care. Both the therapy and research design were acceptable to participants, but modifications may be needed regarding both the timing of assessments and the identification of appropriate primary outcome measures. Given the lack of evidenced based therapies for this population, such a trial would be a significant contribution to the field. Findings may be useful for other groups planning clinical trials of psychological therapies for children in foster care. ISRCTN 90349442 . The trial was retrospectively registered on 6 May 2016.


Technology evaluation: Stem-cell therapy, Aastrom Biosciences Inc.
The expansion of human stem cells and their genetic manipulation represent areas of increasing interest in the field of stem cell transplantation. Previously, stem cell transplantation has been accomplished by using cellular products obtained by large volume bone marrow or peripheral blood harvest. Difficulties with this approach include inadequate cell numbers and tumor cell contamination. Furthermore, for gene transfer modalities requiring proliferating progenitor cells, low gene expression would be expected in these products. To address these difficulties, the AastromReplicell System has been developed as a fully closed and automated system for expanding hematopoietic cells. Investigators at Aastrom have evaluated the conditions needed for optimal growth including the need for unpurified bone marrow or cord blood mononuclear cells, high cell densities, serum-containing medium and certain types of plastic surfaces. Studies have now been initiated to demonstrate the feasibility of generating enough cells to fully reconstitute hematopoiesis from small volumes of cellular progenitors. It has also been demonstrated that tumor cell contamination passively decreases during the culture period. It now remains to be shown in a direct comparison that this approach yields greater efficacy and a lower cost than transplantation with unmanipulated large volume marrow or peripheral blood stem cell products.


When is a little knowledge dangerous? Circumstances of recent heroin overdose and links to knowledge of overdose risk factors.
To describe the circumstances surrounding recent heroin overdose among a sample of heroin overdose survivors and the links to their knowledge of overdose risk. A cross-sectional survey of 257 recent non-fatal heroin overdose survivors was undertaken to examine self-reported knowledge of overdose risk reduction strategies, behaviour in the 12 h prior to overdose and attributions of overdose causation. Most of the overdoses occurred in public spaces as a result of heroin use within 5 min of purchasing the drug. A substantial number of overdoses occurred with no one else present and/or involved the concomitant use of other drugs. While knowledge of at least one overdose prevention strategy was reported by 90% of the sample, less then half of the sample knew any single strategy. Furthermore knowledge of the dangers of mixing benzodiazepines and/or alcohol with heroin was associated with an increased likelihood of such mixing being reported prior to overdose. While heroin users can articulate knowledge of key overdose risk reduction strategies, this knowledge was not generally associated with a reduction in risk behaviours but was in some cases associated with increased reports of overdose risk behaviours. Further research is required in order to better understand this paradoxical effect, focussing on risk reduction education amenable to the social contexts in which heroin use takes place.


Absolute lymphocyte count recovery following initial acute myelogenous leukemia therapy: Implications for adoptive cell therapy.
An adequate absolute lymphocyte count (ALC) is an essential first step in autologous chimeric antigen receptor (CAR) T-cell manufacturing. For patients with acute myelogenous leukemia (AML), the intensity of chemotherapy received may affect adequate ALC recovery required for CAR T-cell production. We sought to analyze ALC following each course of upfront therapy as one metric for CAR T-cell manufacturing feasibility in children and young adults with AML. ALC data were collected from an observational study of patients with newly diagnosed AML between the ages of 1 month and 21 years who received treatment between the years of 2006 and 2018 at one of three hospitals in the Leukemia Electronic Abstraction of Records Network (LEARN) consortium. Among 193 patients with sufficient ALC data for analysis, the median ALC following induction 1 was 1715 cells/μl (interquartile range: 1166-2388), with successive decreases in ALC with each subsequent course. Similarly, the proportion of patients achieving an ALC >400 cells/μl decreased following each course, ranging from 98.4% (190/193) after course 1 to 66.7% (22/33) for patients who received a fifth course of therapy. There is a successive decline of ALC recovery with subsequent courses of chemotherapy. Despite this decline, ALC values are likely sufficient to consider apheresis prior to the initiation of each course of upfront therapy for the majority of newly diagnosed pediatric AML patients, thereby providing a window of opportunity for T-cell collection for those patients identified at high risk of relapse or with refractory disease.


Kinetics of bone marrow repopulation in lethally irradiated rats after transplantation of vascularized bone marrow in syngeneic hind limb.
Grafting of the recipient with bone marrow cell suspension provides only few stromal cells and no autologous environment for cooperation between hemopoietic and stromal cells. Vascularized bone marrow grafts provide the recipient with bone marrow hemopoietic and stromal cells in their natural spatial relationship. It is expected that such a graft will resume its function soon after transplantation and almost immediately repopulate bone marrow cavities of the irradiated recipient as well as supply the sites of cytopoiesis with the necessary growth factors. The aim of the study was to investigate the process of repopulation of bone marrow cavities and lymphoid organs of irradiated recipient by bone marrow cells from syngeneic hind limb transplant. Lewis rats received total body irradiation of 8 Gy followed by orthotopic transplantation of syngeneic limb or i.v. infusion of equivalent amount of bone marrow cells. In control experiments the hind limb was shielded with lead plate during total body irradiation. Ten days after irradiation and hind limb transplantation the yield of nucleated cells from tibia was reaching values of normal animals. In rats receiving i.v. bone marrow cell infusion it was 40% of control values and in rats repopulated from shielded own hind limb it was 60% of controls. In all groups a higher percentage of early and immediate normoblasts and a reduced pool of juvenile and segmented neutrophils was observed. Thirty days after irradiation and repopulation procedures all parameters were returning to normal levels in each group. The results indicate that bone marrow cell transplantation in hind limb graft is highly effective in lethally irradiated animals in reconstituting bone marrow.


PAMAM dendrimers mediate siRNA delivery to target Hsp27 and produce potent antiproliferative effects on prostate cancer cells.
RNA interference (RNAi) holds great promise for the treatment of inherited and acquired diseases, provided that safe and efficient delivery systems are available. Herein we report that structurally flexible triethanolamine (TEA) core PAMAM dendrimers are able to deliver an Hsp27 siRNA effectively into prostate cancer (PC-3) cells by forming stable nanoparticles with siRNA, protecting the siRNA nanoparticles from enzymatic degradation, and enhancing cellular uptake of siRNA. The Hsp27 siRNA resulted in potent and specific gene silencing of heat-shock protein 27, an attractive therapeutic target in castrate-resistant prostate cancer. Silencing of the hsp27 gene led to induction of caspase-3/7-dependent apoptosis and inhibition of PC-3 cell growth in vitro. In addition, the siRNA-dendrimer complexes are non-cytotoxic under the conditions used for siRNA delivery. Altogether, TEA core PAMAM dendrimer-mediated siRNA delivery, in combination with RNAi that specifically targets Hsp27, may constitute a promising approach for combating castrate-resistant prostate cancer, for which there is no efficacious treatment.


Vaccine effectiveness in pandemic influenza - primary care reporting (VIPER): an observational study to assess the effectiveness of the pandemic influenza A (H1N1)v vaccine.
To determine influenza A (H1N1)v vaccine effectiveness (VE) in the Scottish population at an early stage of the 2009-10 H1N1v vaccination programme, using a sentinel surveillance network of 41 general practices contributing to the Practice Team Information (PTI) network. Retrospective cohort study using record linkage. Using the Community Health Index (CHI) number, general practice patient-level data were extracted and linked to the Scottish Morbidity Record (SMR) catalogue, containing information on all inpatient hospitalisations in Scotland. The Health Protection Scotland (HPS) data set was also used, consisting of laboratory-confirmed cases of influenza A (H1N1)v from the practices. The study involved a longitudinal evaluation of the aspect of the influenza A (H1N1)v vaccination programme implemented through general practice in autumn/winter 2009. At 25 December 2009, vaccine uptake estimates for the study population were 12.0% (95% CI 11.9 to 12.1). For those patients in an at-risk group (n = 59,721), the uptake rate was 37.5% (95% CI 37.1 to 37.9). Among the 1492 patients swabbed, 467 were positive for H1N1, giving a positivity rate of 31.3% [95% confidence interval (CI) 29.0 to 33.7]. Among those in a clinical risk group who were not vaccinated, 41.3% (95% CI 35.6 to 46.9) tested positive for influenza A (H1N1)v, a significant difference from the H1N1 positivity percentage among patients with no clinical risk (p < 0.01). Among those vaccinated and in a clinical risk group, only one patient (5%, 95% CI 0.3 to 23.6) tested after vaccination was positive for influenza A (H1N1)v. By comparing postvaccination swabs in those who were vaccinated with swabs taken in those who remained unvaccinated, the VE was found to be 95.0% (95% CI 76.0 to 100.0). In the study population there were 2739 admissions to hospital, of which 1241 were emergency admissions; all 48 emergency hospitalisations for influenza and pneumonia occurred in patients who did not receive the vaccine. VE for single or combined end points of influenza and pneumonia hospitalisation for all patients was estimated at 100.0% (95% CI infinity to 100.0). There were 132 hospitalisations in the unvaccinated group versus five in the vaccinated group for cardiovascular-related conditions. There were 193 hospitalisations in the unvaccinated group versus nine in those vaccinated in the group of patients admitted for influenza, pneumonia, chronic obstructive pulmonary disease (COPD) and cardiovascular-related conditions. VE for cardiovascular-related conditions alone, or in individuals with influenza, pneumonia COPD and cardiovascular-related conditions, was 71.1% (95% CI 11.3 to 90.6) and 64.7% (95% CI 12.0 to 85.8) respectively. Evidence from swabs submitted from patients in the cohort who presented in general practice with influenza-like illness suggests that the introduction of influenza A (H1N1)v vaccine in Scotland during 2009 was associated with a high degree of protection. Influenza A (H1N1)v immunisation in primary health-care settings appears to be both effective and widely acceptable, and should continue to be the mainstay of disease prevention for at-risk patients. A further analysis encompassing the whole influenza season is required to cover more days of vaccination exposure and increase precision.


A mass spectrometry-based unique fragment approach for the identification of microcystins.
Both qualitative and quantitative methods for the analysis of microcystin variants have been established. The qualitative method involves a unique fragment approach, specific for each variant in identifying microcystins, while the quantitative method involves the quantification of microcystins in cellular matrices of cyanobacteria and reservoir water samples. For the identification of fragments associated with their respective microcystin (MC) variants, theoretically obtained fragments of microcystins were compared with tandem mass spectrometric (MS/MS) fragments obtained from an ESI-Orbitrap mass spectrometer. Here, we present the first report of MC variants produced by the model algal strain M. aeruginosa NIES-843. A robust comparative study between the unique fragments and the conventional product ions for quantitative measurements of microcystin has also been carried out. The method with high robustness was further validated by determining the MC level changes in the intracellular matrices of M. aeruginosa samples, grown under high and low nitrogen conditions, and by testing the amount of MC in environmental water samples.


A macroepigenetic approach to identify factors responsible for the autism epidemic in the United States.
The number of children ages 6 to 21 in the United States receiving special education services under the autism disability category increased 91% between 2005 to 2010 while the number of children receiving special education services overall declined by 5%. The demand for special education services continues to rise in disability categories associated with pervasive developmental disorders. Neurodevelopment can be adversely impacted when gene expression is altered by dietary transcription factors, such as zinc insufficiency or deficiency, or by exposure to toxic substances found in our environment, such as mercury or organophosphate pesticides. Gene expression patterns differ geographically between populations and within populations. Gene variants of paraoxonase-1 are associated with autism in North America, but not in Italy, indicating regional specificity in gene-environment interactions. In the current review, we utilize a novel macroepigenetic approach to compare variations in diet and toxic substance exposure between these two geographical populations to determine the likely factors responsible for the autism epidemic in the United States.


Metabolic rate variation in Glossina pallidipes (Diptera: Glossinidae): gender, ageing and repeatability.
Despite the importance of metabolic rate in determining flight time of tsetse and in mediating the influence of abiotic variables on life history parameters (and hence abundance and distribution), metabolic rate measurements and their repeatability have not been widely assessed in these flies. We investigate age-related changes in standard metabolic rate (SMR) and its repeatability, using flow-through respirometry, for a variety of feeding, gender and pregnancy classes during early adult development in laboratory-reared individuals of the tsetse fly, Glossina pallidipes. Standard metabolic rate (144-635 microW) was generally within 22% of previous estimates, though lower than the values found using closed system respirometry. There was no significant difference between the genders, but metabolic rate increased consistently with age, probably owing to flight muscle development. Repeatability of metabolic rate was generally high (r=0.6-.09), but not in younger teneral adults and pregnant females (r approximately equal to 0.05-0.4). In these individuals, low repeatability values are a consequence of muscle or in utero larval development. Tsetse and other flies generally have a much higher metabolic rate, for a given size, than do other insect species investigated to date.


Antifungal activity of curcumin-silver nanoparticles against fluconazole-resistant clinical isolates of Candida species.
Candida species is the common form of opportunistic fungal infections, especially in immunosuppressed individuals. Fluconazole is the first-line therapy for candidiasis as it is affordable and readily available. However, the antifungal resistance pattern in high-risk patients is a major concern. The objective of the present study was to assess the anticandidal activity of curcumin-silver nanoparticles (C-Ag-NPs) against fluconazole-resistant Candida species isolated from HIV patients. Ten milliliters of 0.1 M silver nitrate (AgNO3) and 3 ml curcumin solution was heated in a water bath for 1 h at 60°C. The formation of the Ag-NPs was determined by color change from light yellow to brownish. The solution was centrifuged at 9000 rpm for 15 min and was washed with ethanol and later lyophilized for 24 h to obtain the purified curcumin-Ag-NPs (C-Ag-NPs). A stock of 1 mg/ml of C-Ag-NPs was prepared in deionized water. The agar diffusion test and broth dilution tests were conducted to determine the anticandidal activity of C-Ag-NPs. C-Ag-NPs showed a better antifungal activity compared to curcumin and AgNO3 solution. Candida glabrata and Candida albicans were the most inhibited and Candida tropicalis was the least inhibited species. The mean zone diameter was 22.2 ± 0.8 mm, 20.1 ± 0.8 mm, and 16.4 ± 0.7 mm against C. glabrata, C. albicans and C. tropicalis respectively. Other Candida species under the study were also inhibited. Inhibitory activity was dose dependent and it increased with concentration. The minimum inhibitory concentration values for different Candida species ranged from 31.2 μg/ml to 250 μg/ml. This is the first report on the antifungal activity of C-Ag-NPs against fluconazole-resistant Candida isolates. C-Ag-NPs can be explored further to identify a potential drug candidate that can be used for the treatment of candidiasis due to fluconazole-resistant strains of Candida species.


A stakeholder analysis of community-led collaboration to reduce health inequity in a deprived neighbourhood in South Korea.
Intersectoral collaboration amongst health and other sectors, as well as between government and non-governmental organisations, has been highlighted as a way to improve health equity. We used a mixed-methods approach to assess collaborative relationships between multiple government sectors and civil society and to suggest possible health promotion interventions and policy alternatives for the urban poor in deprived neighborhoods. A total of 18 participants involved in health promotion interventions and policy processes related to the inner-city area of Seoul were recruited using purposive sampling methods. Participants included stakeholders working for or engaging in governments (3), public health care institutions (5), social service providers (3), community-based organisations (CBOs) (4) and faith-based organisations (3). We conducted semi-structured, one-on-one interviews and then collected survey data. Quantitative data were analysed using social network analysis, and qualitative data were analysed through iterative and consensus processes. The social network analysis indicated that a CBO plays the most substantial role in sharing and controlling informational resources to promote health. A stakeholder analysis showed that the CBO neutrally and negatively viewed the possibility of collaboration with other stakeholders. Three themes related to challenges to intersectoral collaboration emerged: (1) lack of trust and communication, (2) need of a coalition with a committed leading actor for future collaboration and (3) organisational and political silos within and across public sectors. Increased understanding of the current status of and challenges to collaboration can inform the planning and implementation of complex intervening strategies and policies tailored to vulnerable people in deprived neighborhoods. Community-led collaborative actions empower people in marginalised communities to envision a healthier community.


Persistence and adherence with the new beta-3 receptor agonist, mirabegron, versus antimuscarinics in overactive bladder: Early experience in Canada.
Antimuscarinics are the principal pharmacological treatment for overactive bladder (OAB), but frequently give rise to anticholinergic side effects, such as dry mouth, a factor leading to poor persistence. The β3-adrenoceptor agonist mirabegron is devoid of significant anticholinergic activity, while being effective in OAB. We evaluated persistence and adherence with mirabegron versus antimuscarinics over 12 months. We obtained retrospective claims from a Canadian Private Drug Plan database for patients 18 years old and over, with a first claim for mirabegron or antimuscarinics during a 6-month index period (April-September 2013). A 6-month look-back identified those with no prior claims for OAB medication (treatment-naïve) or ≥1 prior OAB drug (treatment-experienced). Time to end of persistence (≥30 day therapy gap or switch of therapy) was evaluated over 12 months; adherence with medication (medication possession ratio) was also measured. Persistence data from 19 485 patients (74% female, 92% naïve, 19.9% aged ≥65 years) showed that for experienced patients the median number of days on mirabegron was 299 days, compared with a range of 96 to 242 days for the different antimuscarinics; for naïve patients, it was 196 versus 70 to 100 days, respectively. Persistence at 12 months was for mirabegron 39% versus 14% to 35% for antimuscarinics, (experienced) and 30% mirabegron versus 14% to 21% antimuscarinics, (naïve). Patients taking mirabegron demonstrated statistically significantly greater adherence than those taking antimuscarinics. Patients who received mirabegron remained longer on treatment than those treated with antimuscarinics, and had higher 12-month persistence and adherence rates.


Early feeding and risk of celiac disease in a prospective birth cohort.
Timing of gluten introduction has been associated with the risk of celiac disease (CD) in children, but the optimal time window is unknown. We aimed to study the effect of age of gluten introduction on the risk of CD, adjusting for continued breastfeeding. In The Norwegian Mother and Child Cohort Study, a prospective birth cohort including 107,000 children, CD was identified by questionnaires and by linkage to the Norwegian Patient Register. Gluten introduction was reported monthly from 0 to 6 months of age, and breastfeeding from 0 to 18 months. After exclusion of cases with insufficient information, 324 children with CD in a cohort of 82,167 were used in the analyses. Gluten was introduced before or at 4 months in 8.0%, 5 to 6 months in 45.3%, and after 6 months in 46.6%, whereas continued breastfeeding was stable at ≈ 78% at 6 months age. CD was diagnosed in 3.68/1000 of the infants with gluten introduction at 5 to 6 months compared with 4.15/1000 with late and 4.24/1000 with early gluten introduction. After adjustment for the child's age and gender, breastfeeding, and maternal CD, delayed gluten introduction was associated with an increased risk of CD (adjusted odds ratio, 1.27 [95% confidence interval, 1.01-1.65], P = .045). Breastfeeding >12 months was also associated with increased risk (adjusted odds ratio, 1.49 [95% confidence interval, 1.01-2.21], P = .046). We found an increased risk of CD in children introduced to gluten after 6 months and a higher risk in children breastfed after 12 months age.


Occludin-deficient embryonic stem cells can differentiate into polarized epithelial cells bearing tight junctions.
Occludin is the only known integral membrane protein of tight junctions (TJs), and is now believed to be directly involved in the barrier and fence functions of TJs. Occludin-deficient embryonic stem (ES) cells were generated by targeted disruption of both alleles of the occludin gene. When these cells were subjected to suspension culture, they aggregated to form simple, and then cystic embryoid bodies (EBs) with the same time course as EB formation from wild-type ES cells. Immunofluorescence microscopy and ultrathin section electron microscopy revealed that polarized epithelial (visceral endoderm-like) cells were differentiated to delineate EBs not only from wild-type but also from occludin-deficient ES cells. Freeze fracture analyses indicated no significant differences in number or morphology of TJ strands between wild-type and occludin-deficient epithelial cells. Furthermore, zonula occludens (ZO)-1, a TJ-associated peripheral membrane protein, was still exclusively concentrated at TJ in occludin-deficient epithelial cells. In good agreement with these morphological observations, TJ in occludin-deficient epithelial cells functioned as a primary barrier to the diffusion of a low molecular mass tracer through the paracellular pathway. These findings indicate that there are as yet unidentified TJ integral membrane protein(s) which can form strand structures, recruit ZO-1, and function as a barrier without occludin.


Genetic deletion of the A1 adenosine receptor limits myocardial ischemic tolerance.
Adenosine receptors may be important determinants of intrinsic ischemic tolerance. Genetically modified mice were used to examine effects of global A1 adenosine receptor (A1AR) knockout (KO) on function and ischemic tolerance in perfused mouse hearts. Baseline contractile function and heart rate were unaltered by A1AR KO, which was shown to abolish the negative chronotropic effects of 2-chloroadenosine (A1AR-mediated) without altering A2 adenosine receptor-mediated coronary dilation. Tolerance to 25 minutes global normothermic ischemia (followed by 45 minutes reperfusion) was significantly limited by A1AR KO, with impaired contractile recovery (reduced by 25%) and enhanced lactate dehydrogenase (LDH) efflux (increased by 100%). Functional effects of A1AR KO involved worsened systolic pressure development with little to no change in diastolic dysfunction. In contrast, cardiac specific A1AR overexpression enhanced ischemic tolerance with a primary action on diastolic dysfunction. Nonselective receptor agonism (10 micromol/L 2-chloroadenosine) protected wild-type and also A1AR KO hearts (albeit to a lesser extent), implicating protection via subtypes additional to A1ARs. However, A1AR KO abrogated effects of 2-chloroadenosine on ischemic contracture and diastolic dysfunction. These data are the first demonstrating global deletion of the A1AR limits intrinsic myocardial resistance to ischemia. Data indicate the function of intrinsically activated A1ARs appears primarily to be enhancement of postischemic contractility and limitation of cell death.


Video-based CBT-E improves eating patterns in obese patients with eating disorder: A single case multiple baseline study.
Cognitive Behavioral Therapy (CBT) is effective for treating eating disorders but it may be difficult to reach patients living far from urban centers. Mobile video-based psychotherapy may potentially improve service reach but has not yet been evaluated. The purpose of this study was to investigate the effects of mobile video-based CBT for eating disorder and to explore the feasibility to use this technology in clinical care. A controlled single case multiple baseline design was used which allowed for statistical analyses with randomization tests and non-overlap of all pairs (NAP). Five patients in the first stage of eating disorder treatment were included and the main outcome variable was daily meal frequency. Secondary outcome variables included eating disorder symptoms, psychological distress and treatment satisfaction. The treatment resulted in a significant (p < .01) increase in daily meal frequency with medium to large effect sizes (combined NAP = .89). Four participants reported reliable improvements in eating disorder symptoms and three reported improvements in mood. The participants reported high satisfaction with the treatment and with the mobile video-application despite some technical problems. Self-reported data on eating behavior is prone to be biased and the results of single case studies may have limited generalizability. CBT can be delivered effectively via a mobile video application and, despite some technological issues, can be well received by patients. All participants in this study had previous low access to mental health services and reported high satisfaction with the treatment format.


Routine defunctioning stoma after chemoradiation and total mesorectal excision: a single-surgeon experience.
To investigate the 10-year results of treating low rectal cancer by a single surgeon in one institution. From Oct 1998 to Feb 2009, we prospectively followed a total of 62 patients with cT2-4 low rectal cancer with lower tumor margins measuring at 3 to 6 cm above the anal verge. All patients received neoadjuvant chemoradiation (CRT) for 6 wk. Among them, 85% of the patients received 225 mg/m(2)/d 5-fluorouracil using a portable infusion pump. The whole pelvis received a total dose of 45 Gy of irradiation in 25 fractions over 5 wk. The interval from CRT completion to surgical intervention was planned to be approximately 6-8 wk. Total mesorectal excision (TME) and routine defunctioning stoma construction were performed by one surgeon. The distal resection margin, circumferential resection margin, tumor regression grade (TRG) and other parameters were recorded. We used TRG to evaluate the tumor response after neoadjuvant CRT. We evaluated anal function outcomes using the Memorial Sloan-Kettering Cancer Center anal function scores after closure of the defunctioning stoma. The median distance from the lower margin of rectal cancer to the anal verge was 5 cm: 6 cm in 9 patients, 5 cm in 32 patients, 4 cm in 10 patients, and 3 cm in 11 patients. Before receiving neoadjuvant CRT, 45 patients (72.6%) had a cT3-4 tumor, and 21 (33.9%) patients had a cN1-2 lymph node status. After CRT, 30 patients (48.4%) had a greater than 50% clinical reduction in tumor size. The final pathology reports revealed that 33 patients (53.2%) had a ypT3-4 tumor and 12 (19.4%) patients had ypN1-2 lymph node involvement. All patients completed the entire course of neoadjuvant CRT. Most patients developed only Grade 1-2 toxicities during CRT. Thirteen patients (21%) achieved a pathologic complete response. Few post-operative complications occurred. Nearly 90% of the defunctioning stomas were closed within 6 mo. The local recurrence rate was 3.2%. Pathologic lymph node involvement was the only prognostic factor predicting disease recurrence (36.5% vs 76.5%, P = 0.006). Nearly 90% of patients recovered sphincter function within 2 year after closure of the defunctioning stoma. Neoadjuvant CRT followed by TME, combined with routine defunctioning stoma construction and high-volume surgeon experience, can provide excellent surgical quality and good local disease control.


Estimation of Ki in a competitive enzyme-inhibition model: comparisons among three methods of data analysis.
There are a variety of methods available to calculate the inhibition constant (Ki) that characterizes substrate inhibition by a competitive inhibitor. Linearized versions of the Michaelis-Menten equation (e.g., Lineweaver-Burk, Dixon, etc.) are frequently used, but they often produce substantial errors in parameter estimation. This study was conducted to compare three methods of analysis for the estimation of Ki: simultaneous nonlinear regression (SNLR); nonsimultaneous, nonlinear regression, "KM,app" method; and the Dixon method. Metabolite formation rates were simulated for a competitive inhibition model with random error (corresponding to 10% coefficient of variation). These rates were generated for a control (i.e., no inhibitor) and five inhibitor concentrations with six substrate concentrations per inhibitor and control. The KM/Ki ratios ranged from less than 0.1 to greater than 600. A total of 3 data sets for each of three KM/Ki ratios were generated (i.e., 108 rates/data set per KM/Ki ratio). The mean inhibition and control data were fit simultaneously (SNLR method) using the full competitive enzyme-inhibition equation. In the KM,app method, the mean inhibition and control data were fit separately to the Michaelis-Menten equation. The SNLR approach was the most robust, fastest, and easiest to implement. The KM,app method gave good estimates of Ki but was more time consuming. Both methods gave good recoveries of KM and VMAX values. The Dixon method gave widely ranging and inaccurate estimates of Ki. For reliable estimation of Ki values, the SNLR method is preferred.


The Current Status of Cytomegalovirus (CMV) Prevalence in the MENA Region: A Systematic Review.
Human cytomegalovirus (CMV) is a highly prevalent herpesvirus worldwide. According to the Centers for Disease Control and Prevention (CDC) and the World Health Organization (WHO), CMV infects people of all ages, and by the age of five, approximately one-third of children in the United States are infected. Although the infection is generally asymptomatic, it can cause severe disease in immunocompromised patients, transplant and transfusion recipients, as well as newborn neonates. The objective of this study is to systematically review published literature on CMV in the MENA region to estimate its incidence in the region and describe its epidemiological and clinical significance. The literature was searched through four scientific databases: PubMed, Scopus, Science Direct, and Web of Science. A total of 72 studies from 11 countries satisfied the inclusion criteria, covering a period from 1988-2019. The CMV IgG seroprevalence ranged from 8.7%-99.2% (SD = 38.95%). CMV incidence in these countries ranged between 1.22% and 77% in transplant and transfusion recipients, with an increase in incidence with advanced age. However, the incidence rate was unclear for congenital CMV due to the variability of the reporting. This review highlights the need for more robust and well-designed studies to better estimate CMV incidence in the MENA region, standardize diagnostic criteria, and consider prophylactic and pre-emptive treatments to limit the morbidity and mortality of the disease.


Evidence of lifestyle modification in the management of hypercholesterolemia.
Coronary heart disease (CHD) is the leading cause of morbidity and mortality worldwide. The growth of ageing populations in developing countries with progressively urbanized lifestyles are major contributors. The key risk factors for CHD such as hypercholesterolemia, diabetes mellitus, and obesity are likely to increase in the future. These risk factors are modifiable through lifestyle. To review current literature on the potential benefit of cholesterol lowering in CHD risk reduction with a particular focus on the evidence of non-pharmacological/lifestyle management of hypercholesterolemia. Medline/PubMed systematic search was conducted using a two-tier approach limited to all recent English language papers. Primary search was conducted using key words and phrases and all abstracts were subsequently screened and relevant papers were selected. The next tier of searching was conducted by (1) reviewing the citation lists of the selected papers and (2) by using PubMed weblink for related papers. Over 3600 reports were reviewed. Target cholesterol levels set out in various guidelines could be achieved by lifestyle changes, including diet, weight reduction, and increased physical activity with the goal of reducing total cholesterol to <200 mg/dL and LDL-C<100 mg/dL. Various dietary constituents such as green tea, plant sterols, soy protein have important influences on total cholesterol. Medical intervention should be reserved for those patients who have not reached this goal after 3 months of non-pharmacological approach. CHD remains as a leading cause of death worldwide and hypercholesterolemia is an important cause of CHD. Non-pharmacological methods provide initial as well as long-term measures to address this issue.


Using Pleth Variability as a Triage Tool for Children With Obstructive Airway Disease in a Pediatric Emergency Department.
Patients with obstructive airway disease have varying degrees of pulsus paradoxus that correlate with illness severity. Pulsus paradoxus can be measured using plethysmography. We investigated whether plethysmograph (pleth) variability on admission to the pediatric emergency department (ED) could predict patient disposition. We hypothesized that patients with a larger pleth variability would have a higher likelihood of being admitted to a general pediatrics unit or the intensive care unit (ICU). We conducted a prospective single-center study of children aged 1 to 18 years who presented to a pediatric ED with a diagnosis of asthma or reactive airway disease. The pleth variability index (PVI) was calculated from their initial plethysmography tracing. Disposition from the ED was recorded as discharge, admission to the floor, or admission to the ICU. A total of 117 patients were included in our study. Forty-eight patients were discharged home, 61 were admitted to the floor, and 8 were admitted to the ICU. The median PVI for each of these groups was 0.27 (interquartile range [IQR], 0.19-0.39) for discharges, 0.29 (IQR, 0.20-0.44) for patients admitted to the floor, and 0.56 (IQR, 0.35-0.70) for patients admitted to the ICU. A Kruskal-Wallis test demonstrated a significant difference in the PVI between each of the groups (P = 0.0087). Our results suggest that PVI may be a useful tool in the triage of children who present to the ED with obstructive airway disease. Further studies should aim to assess the validity of PVI in predicting the response to bronchodilator therapy during the course of a patient's hospitalization.


Heparin binding domain in vitronectin is required for oligomerization and thus enhances integrin mediated cell adhesion and spreading.
Vitronectin is a multi-functional protein found predominantly as a monomer in blood and as an oligomer in the extracellular matrix. We have dissected the minimal regions of vitronectin protein needed for effective integrin dependent cell adhesion and spreading. A fragment of vitronectin containing the RGD integrin binding site showed similar binding affinity as that of full vitronectin protein to purified integrin alphavbeta3 but had diminished cell adhesion and spreading function in vivo. We demonstrate that the oligomeric state of the protein is responsible for this effect. We provide compelling evidence for the involvement of the heparin binding domain of vitronectin in the oligomerization process and show that such oligomerization reinforces the activity of vitronectin in cell adhesion and spreading.


The role of the hypoxia-inducible BH3-only proteins BNIP3 and BNIP3L in cancer.
In this review, we summarize current knowledge of the biological functions of the atypical BH3-only proteins BNIP3 and BNIP3L, focusing on the role of these proteins in cancer. Hypoxia increases the expression of BNIP3 through the transcription factor HIF-1, but despite a considerable number of investigations, it has proven difficult to establish a clear role for BNIP3 in the cellular hypoxic response. BNIP3 can induce a form of cell death that shows features of both necrosis and apoptosis, but unusually for a BH3-only protein, death occurs independently of the BH3 domain and is critically dependent on a C-terminal transmembrane domain, which also localizes the protein to the mitochondria. BNIP3 expression does not always result in cell death, suggesting that additional factors may suppress BNIP3 or cooperate with it to induce death. BNIP3 is highly expressed in some tumors, including those of the breast, lung and cervix. However, in colorectal and pancreatic cancers BNIP3 is frequently epigenetically silenced, possibly reflecting different functions for BNIP3 in different tissues. Recent reports have shown that BNIP3 can induce autophagy and there is some evidence to suggest this may represent an emerging role for BH3-only proteins in general. However, the mechanism through which BNIP3 induces autophagy and the cellular consequences of this are yet to be established.


Risk prediction error coding in orbitofrontal neurons.
Risk is a ubiquitous feature of life. It plays an important role in economic decisions by affecting subjective reward value. Informed decisions require accurate risk information for each choice option. However, risk is often not constant but changes dynamically in the environment. Therefore, risk information should be updated to the current risk level. Potential mechanisms involve error-driven updating, whereby differences between current and predicted risk levels (risk prediction errors) are used to obtain currently accurate risk predictions. As a major reward structure, the orbitofrontal cortex is involved in coding key reward parameters such as reward value and risk. In this study, monkeys viewed different visual stimuli indicating specific levels of risk that deviated from the overall risk predicted by a common earlier stimulus. A group of orbitofrontal neurons displayed a risk signal that tracked the discrepancy between current and predicted risk. Such neuronal signals may be involved in the updating of risk information.


Nma111p, the pro-apoptotic HtrA-like nuclear serine protease in Saccharomyces cerevisiae: a short survey.
The baker's yeast, Saccharomyces cerevisiae, is also capable of undergoing programmed cell death or apoptosis, for example in response to viral infection as well as during chronological and replicative aging. Intrinsically, programmed cell death in yeast can be induced by, for example, H2O2, acetic acid or the mating-type pheromone. A number of evolutionarily conserved apoptosis-regulatory proteins have been identified in yeast, one of which is the HtrA (high-temperature requirement A)-like serine protease Nma111p (Nma is nuclear mediator of apoptosis). Nma111p is a nuclear serine protease of the HtrA family, which targets Bir1p, the only known inhibitor-of-apoptosis protein in yeast. Nma111p mediates apoptosis in a serine-protease-dependent manner and exhibits its activity exclusively in the nucleus. How the activity of Nma111p is regulated has remained largely elusive, but some evidence points to a control by phosphorylation. Current knowledge of Nma111p's function in apoptosis will be discussed in the present review.


"The extension service is not an integration agency": the idea of race in the Cooperative Extension Service.
This article is an institutional history of the development of race policy within the federal Cooperative Extension Service. It demonstrates that the popular belief in African-American inferiority and pragmatic political compromises aimed at creating a bureaucracy serving the nation's agricultural constituency and ensuring its longevity, led to a conscious marginalization of African-American interests within the program. Federal extension officials not only tolerated, but actively supported, discrimination within the southern the southern branches of the service. African-American leadership protested against the adverse effects of racial policies in the Extension Service from the very beginning. While there were some limited positive gains in the number of staff and availability of services for rural African-Americans, these changes did not challenge the suzerainty of whites over the program. In the post-World War II era -- especially after 1950 -- African-Americans confronted white extension leadership. Extension officials hid behind a bureaucratic façade and a flawed interpretation of the federal-state cooperative agreement to delay institutional restructuring. Political appointees pushed the service toward policies of racial justice; however, extension leadership continued to move slowly on fundamental transformation. As a result, the adjustments did not lead to a fundamental re-thinking of race policy in the service and ultimately contributed to the disappearance of the African-American extension force to a significant degree.


The effect of diet and opponent size on aggressive interactions involving caribbean crazy ants (Nylanderia fulva).
Biotic interactions are often important in the establishment and spread of invasive species. In particular, competition between introduced and native species can strongly influence the distribution and spread of exotic species and in some cases competition among introduced species can be important. The Caribbean crazy ant, Nylanderia fulva, was recently introduced to the Gulf Coast of Texas, and appears to be spreading inland. It has been hypothesized that competition with the red imported fire ant, Solenopsis invicta, may be an important factor in the spread of crazy ants. We investigated the potential of interspecific competition among these two introduced ants by measuring interspecific aggression between Caribbean crazy ant workers and workers of Solenopsis invicta. Specifically, we examined the effect of body size and diet on individual-level aggressive interactions among crazy ant workers and fire ants. We found that differences in diet did not alter interactions between crazy ant workers from different nests, but carbohydrate level did play an important role in antagonistic interactions with fire ants: crazy ants on low sugar diets were more aggressive and less likely to be killed in aggressive encounters with fire ants. We found that large fire ants engaged in fewer fights with crazy ants than small fire ants, but fire ant size affected neither fire ant nor crazy ant mortality. Overall, crazy ants experienced higher mortality than fire ants after aggressive encounters. Our findings suggest that fire ant workers might outcompete crazy ant workers on an individual level, providing some biotic resistance to crazy ant range expansion. However, this resistance may be overcome by crazy ants that have a restricted sugar intake, which may occur when crazy ants are excluded from resources by fire ants.


Fatigability, Exercise Intolerance, and Abnormal Skeletal Muscle Energetics in Heart Failure.
Among central and peripheral factors contributing to exercise intolerance (EI) in heart failure (HF), the extent to which skeletal muscle (SM) energy metabolic abnormalities occur and contribute to EI and increased fatigability in HF patients with reduced or preserved ejection fraction (HFrEF and HFpEF, respectively) are not known. An energetic plantar flexion exercise fatigability test and magnetic resonance spectroscopy were used to probe the mechanistic in vivo relationships among SM high-energy phosphate concentrations, mitochondrial function, and EI in HFrEF and HFpEF patients and in healthy controls. Resting SM high-energy phosphate concentrations and ATP flux rates were normal in HFrEF and HFpEF patients. Fatigue occurred at similar SM energetic levels in all subjects, consistent with a common SM energetic limit. Importantly, HFrEF New York Heart Association class II-III patients with EI and high fatigability exhibited significantly faster rates of exercise-induced high-energy phosphate decline than did HFrEF patients with low fatigability (New York Heart Association class I), despite similar left ventricular ejection fractions. HFpEF patients exhibited severe EI, the most rapid rates of high-energy phosphate depletion during exercise, and impaired maximal oxidative capacity. Symptomatic fatigue during plantar flexion exercise occurs at a common energetic limit in all subjects. HFrEF and HFpEF patients with EI and increased fatigability manifest early, rapid exercise-induced declines in SM high-energy phosphates and reduced oxidative capacity compared with healthy and low-fatigability HF patients, suggesting that SM metabolism is a potentially important target for future HF treatment strategies.


Chemotherapy, biochemotherapy and anti-VEGF therapy in metastatic mucosal melanoma.
Mucosal melanomas arise from melanocytes located at various mucosal membranes, which demonstrate a clear demographic and ethnic disparity. This subtype of melanoma has more aggressive course and poorer prognosis than other subsets of melanomas. Due to their rarity, there is no well-established protocol of staging and treatment of mucosal melanomas. Significant advances have been achieved in novel immunotherapeutic and specific targeted agents for metastatic melanomas. For mucosal melanoma patients especially without known gene mutation, anti-angiogenic therapy combined with chemotherapy or other targeted drugs has shown promising efficacy, either as first- or second-line treatment. In some subset analysis, patients with mucosal melanoma, harboring wild type BRAF, might get more clinical and survival benefits than cutaneous ones. Therefore it is rational to further study these therapeutic strategies in the group of mucosal melanoma. Chemotherapy or biochemotherapy only showed limited efficacy without significant survival improvement, but larger sample size study is still required. Herein we presented a comprehensive review of chemotherapy/biochemotherapy and anti-vascular endothelial growth factor (anti-VEGF) therapy in metastatic mucosal melanoma.


Differentiating surface and bulk interactions using localized surface plasmon resonances of gold nanorods.
We demonstrate a novel localized surface-plasmon resonance sensor that can distinguish surface binding interactions from interfering bulk effects. This is accomplished by utilizing the longitudinal and transverse plasmon modes of gold nanorods. We have investigated, both numerically and experimentally, the effect of change in background refractive index and surface binding on the two resonances of a gold nanorod on an indium tin oxide coated glass substrate.


Bifunctional opioid/nociceptin hybrid KGNOP1 effectively attenuates pain-related behaviour in a rat model of neuropathy.
A bifunctional peptide containing an opioid and nociceptin receptor-binding pharmacophore, H-Dmt-D-Arg-Aba-β-Ala-Arg-Tyr-Tyr-Arg-Ile-Lys-NH2 (KGNOP1), was tested for its analgesic properties when administered intrathecally in naïve and chronic constriction injury (CCI)-exposed rats with neuropathy-like symptoms. KGNOP1 significantly increased the acute pain threshold, as measured by the tail-flick test, and also increased the threshold of a painful reaction to mechanical and thermal stimuli in CCI-exposed rats. Both of the effects could be blocked by pre-administration of [Nphe1]-Nociceptin (1-13)-NH2 (NPhe) or naloxone, antagonists for nociceptin and opioid receptors, respectively. This led us to conclude that KGNOP1 acts as a dual opioid and nociceptin receptor agonist in vivo. The analgesic effect of KGNOP1 proved to be more powerful than clinical drugs such as morphine and buprenorphine. Repeated daily intrathecal injections of KGNOP1 led to the development of analgesic tolerance, with the antiallodynic action being completely abolished on day 6. Nevertheless, the development of tolerance to the antihyperalgesic effect was delayed in comparison to morphine, which lost its efficacy as measured by the cold plate test after 3days of daily intrathecal administration, whereas KGNOP1 was efficient up to day 6. A single intrathecal injection of morphine to KGNOP1-tolerant rats did not raise the pain threshold in any of the behavioural tests; in contrast, a single intrathecal dose of KGNOP1 significantly suppressed allodynia and hyperalgesia in morphine-tolerant rats.


Recombination at the bar locus in an inverted attached-X system in Drosophila melanogaster.
Recombination at the Bar locus in Drosophila melanogaster was investigated in an inverted attached-X system which enhanced the frequency of homozygosis for the Bar region. Females among the progeny of homozygous B mothers were searched for changes of B to BB and to B(+). Marker genes were followed and exceptional half-tetrads were analyzed in regard to two hypotheses: that of exchange between obliquely synapsed members of the duplication, which is associated with exchange of outside markers, and that of intrachromosomal exchange, which does not involve recombination of markers.-Recombinant exceptions of B(+) /BB genotype, carrying the outside marker combinations predicted on the hypothesis of exchange between obliquely synapsed duplication members, were encountered repeatedly. It is established that B(+) and BB strands are reciprocal products of the same event.-Twelve nonrecombinant exceptional strands were isolated; ten of these were B(+) and two were BB. Only one of the nonrecombinant half-tetrads offered the opportunity to test the prediction of reciprocity of the intrachromosomal event. Analysis showed the exceptional female to be of the constitution BB/B, a type not expected on the hypothesis. While it could have arisen through some kind of copy error in the repair of a chromatid break, a valid test of the hypothesis of intrachromosomal exchange must rest on the isolation and analysis of more cases of the appropriate exceptional genotype.-In several cases Bar changes were found to be associated with aberrations; all but one of these involved spontaneous, cytologically identifiable deletions.


Sugar import and phytopathogenicity of Spiroplasma citri: glucose and fructose play distinct roles.
We have shown previously that the glucose PTS (phosphotransferase system) permease enzyme II of Spiroplasma citri is split into two distinct polypeptides, which are encoded by two separate genes, crr and ptsG. A S. citri mutant was obtained by disruption of ptsG through homologous recombination and was proved unable to import glucose. The ptsG mutant (GII3-glc1) was transmitted to periwinkle (Catharanthus roseus) plants through injection to the leaf-hopper vector. In contrast to the previously characterized fructose operon mutant GMT 553, which was found virtually nonpathogenic, the ptsG mutant GII3-glc1 induced severe symptoms similar to those induced by the wild-type strain GII-3. These results, indicating that fructose and glucose utilization were not equally involved in pathogenicity, were consistent with biochemical data showing that, in the presence of both sugars, S. citri used fructose preferentially. Proton nuclear magnetic resonance analyses of carbohydrates in plant extracts revealed the accumulation of soluble sugars, particularly glucose, in plants infected by S. citri GII-3 or GII3-glc1 but not in those infected by GMT 553. From these data, a hypothetical model was proposed to establish the relationship between fructose utilization by the spiroplasmas present in the phloem sieve tubes and glucose accumulation in the leaves of S. citri infected plants.


Inhibition of SHP-1 Expands the Repertoire of Antitumor T Cells Available to Respond to Immune Checkpoint Blockade.
The presence and activity of CD8+ T cells within the tumor microenvironment are essential for the control of tumor growth. Utilizing B16-F10 melanoma tumors that express altered peptide ligands of chicken ovalbumin, OVA257-264, we measured high- and low-affinity OVA-specific responses following adoptive transfer of OT-I CD8+ T cell into mice subsequently challenged with tumors. T-cell receptor (TCR) affinity positively correlated with the frequency of OT-I tumor-infiltrating lymphocytes (TIL). Differences in TCR affinity inversely corresponded to in vivo tumor growth rate. Blockade of the PD-1 and CTLA-4 checkpoints preferentially increased the frequency and antitumor function of TIL responding to high-affinity antigens, while failing to enhance the antitumor activity of low-affinity T cells. To determine whether lowering the TCR activation threshold could enhance the breadth and magnitude of the antitumor T-cell response, we inhibited Src homology region 2 domain-containing phosphatase 1 (SHP-1) in OT-I T cells prior to tumor antigen exposure. SHP-1 knockdown increased the cytokine-producing potential of high- and low-affinity T cells but failed to enhance control of tumor growth. In contrast, when SHP-1 knockdown of OT-I T cells was combined with immunotherapy, we observed a significant and long-lasting suppression of tumor growth mediated by low-affinity T cells. We conclude that lowering the TCR activation threshold by targeting SHP-1 expands the repertoire of T cells available to respond to conventional checkpoint blockade, leading to enhanced control of tumor growth.


Radiation exposure to personnel during examination of limbs of horses with a portable hand-held fluoroscopic unit.
To determine radiation exposure to personnel during fluoroscopic imaging of limbs of horses with a portable unit and to determine distance from the c-arm at which radioprotective clothing is not required. Repeated-measures cohort study. Part 1, 1 forelimb and 1 hind limb from each of 5 equine cadavers; parts 2 and 3, personnel involved during imaging of limbs of 5 and 9 horses, respectively. Radiation exposure rates were mapped around the suspended c-arm of a portable fluoroscopy unit during imaging of various joints of equine cadaver limbs. During similar examinations in live horses, exposure rates to the fluoroscopist and assistant were measured. Mean duration for fluoroscopy of various joints was determined by observing an experienced fluoroscopist. Exposure to fluoroscopists and assistants per examination and per annum was estimated. Radiation exposure rates were dependent on distance and direction relative to the c-arm and consistently highest on the tube side of the unit. Exposure was significantly greater than background amounts until approximately 4.7 m from the c-arm. During examination of live horses, exposure was highest to the fluoroscopist's hand nearest the tube. Typically, exposure to the fluoroscopist and assistant during carpal examination was 25 to 40 times greater than that for comparable radiographic examination. Annual exposure for fluoroscopists was more than twice the recommended maximum permissible dose. Fluoroscopic imaging of limbs of horses represents a major source of radiation exposure. Annual maximum permissible doses of radiation will be rapidly exceeded if required radioprotective clothing is not worn.


Knowledge of measures to safeguard harvested rainwater quality in rural domestic households.
Given the possibility of waterborne diseases caused by inappropriate rainwater harvesting systems, a survey was conducted in Uganda to assess existing knowledge of both physical and non-physical measures that safeguard harvested rainwater. Households who had received rainwater tanks were assessed on issues related to harvested rainwater quality. The study shows that 84% of respondents were aware of various sources of rainwater contamination, but only 5% were aware that they needed to adjust use of rainwater, depending on whether they cleaned the tank or not. Most of the respondents were not aware that gutter cleaning was necessary to improve water quality. Indeed, as the water from the collection surface is channelled through gutters, a number of measures need to be taken to control the entry of contaminations and subsequent growth of pathogens in the tank, e.g. first flush diverts, installation of filters, chemical use and mesh cleaning. The majority, however, did not take adequate care of the gutters and this impacts on health and social livelihood. Overall, the findings emphasize the need to provide more information to households when installing water harvesting tanks to ensure that the harvested rainwater is of high quality.