Signaling from the RNA sensor RIG-I is regulated by ufmylation.

The RNA-binding protein RIG-I is a key initiator of the antiviral innate immune response. The signaling that mediates the antiviral response downstream of RIG-I is transduced through the adaptor protein MAVS and results in the induction of type I and III interferons (IFNs). This signal transduction occurs at endoplasmic reticulum (ER)­mitochondrial contact sites, to which RIG-I and other signaling proteins are recruited following their activation. RIG-I signaling is highly regulated to prevent aberrant activation of this pathway and dysregulated induction of IFN. Previously, we identified UFL1, the E3 ligase of the ubiquitin-like modifier conjugation system called ufmylation, as one of the proteins recruited to membranes at ER­mitochondrial contact sites in response to RIG-I activation. Here, we show that UFL1, as well as the process of ufmylation, promote IFN induction in response to RIG-I activation. We found that following RNA virus infection, UFL1 is recruited to the membrane-targeting protein 14­3-3ε and that this complex is then recruited to activated RIG-I to promote downstream innate immune signaling. Importantly, we found that 14­3-3ε has an increase in UFM1 conjugation following RIG-I activation. Additionally, loss of cellular ufmylation prevents the interaction of 14­3-3ε with RIG-I, which abrogates the interaction of RIG-I with MAVS and thus the downstream signal transduction that induces IFN. Our results define ufmylation as an integral regulatory component of the RIG-I signaling pathway and as a posttranslational control for IFN induction.

FTO Suppresses STAT3 Activation and Modulates Proinflammatory Interferon-Stimulated Gene Expression.

Signaling initiated by type I interferon (IFN) results in the induction of hundreds of IFN-stimulated genes (ISGs). The type I IFN response is important for antiviral restriction, but aberrant activation of this response can lead to inflammation and autoimmunity. Regulation of this response is incompletely understood. We previously reported that the mRNA modification m6A and its deposition enzymes, METTL3 and METTL14 (METTL3/14), promote the type I IFN response by directly modifying the mRNA of a subset of ISGs to enhance their translation. Here, we determined the role of the RNA demethylase fat mass and obesity-associated protein (FTO) in the type I IFN response. FTO, which can remove either m6A or cap-adjacent m6Am RNA modifications, has previously been associated with obesity and body mass index, type 2 diabetes, cardiovascular disease, and inflammation. We found that FTO suppresses the transcription of a distinct set of ISGs, including many known pro-inflammatory genes, and that this regulation requires its catalytic activity but is not through the actions of FTO on m6Am. Interestingly, depletion of FTO led to activation of the transcription factor STAT3, whose role in the type I IFN response is not well understood. This activation of STAT3 increased the expression of a subset of ISGs. Importantly, this increased ISG induction resulting from FTO depletion was partially ablated by depletion of STAT3. Together, these results reveal that FTO negatively regulates STAT3-mediated signaling that induces proinflammatory ISGs during the IFN response, highlighting an important role for FTO in suppression of inflammatory genes.

Pediatric Simulation-Based Prehospital Training Course in Botswana.

Audience: This simulation-based training focuses on the most common and high risk pediatric prehospital scenarios in low- and middle-income countries (LMIC). The curriculum was developed based on a needs assessment to train Ministry of Health and Wellness (MOHW) prehospital providers in Botswana specifically for pediatric resuscitation and could be used for emergency medical services (EMS) providers in other LMIC. After participating in this curriculum, providers should enhance their assessment and interventions in acutely ill pediatric prehospital patients. Length of Curriculum: The entire course was designed to be presented over two days with 6-8 hours of instruction each day. Introduction: In recent years, prehospital medicine has shown continued growth in LMICs, specifically in Sub-Saharan Africa. As these programs develop focused training for the pediatric population, equipping the workforce with pediatric resuscitation skills is essential. A few years after its inception, the Botswana MOHW identified deficiencies in their current training program and sought external expertise and educational training. We partnered with the MOHW to create and implement a novel, prehospital simulation curriculum to teach pediatric resuscitation to prehospital providers. Our aim was to create a curriculum based on the needs of the community that could also be implemented in other similar resource-limited settings. This course included didactic sessions, five simulation scenarios using low fidelity mannequins and three pediatric-focused skill sessions. This program was found to be effective based on statistically significant improvement in written and simulation post-test scores. Educational Goals: The objective of this educational project was to design, implement, and evaluate a curriculum relevant to an EMS system based in a LMIC, so that it could be a basis for curricula for use in similar contexts. The educational goal is to improve prehospital providers performance in common pediatric resuscitations. Educational Methods: The educational methods used in this curriculum included simulation using rapid cycle deliberate practice (RCDP), didactic lectures, and hands on skills training for common pediatric scenarios. Outcomes were measured by comparing performance on written and simulation-based pre-and post-tests. Research Methods: Participants completed written and simulation-based pre- and post-tests covering the concepts taught in the curriculum. Continuous variables (written and simulation test scores) were compared between two dependent groups (pre- and post-trainings) using paired t-tests. Results: Mean written test scores increased by 11%, from 75% to 86% (p<0.0001), while mean simulated test scores increased by 22% (from 56% to 78 % (p<0.0001). Discussion: The curriculum we developed focused on high-yield pediatric skills based on the needs of the Botswana MOHW EMS program. We believe simulation training was an excellent and effective method for this type of training. We specifically designed RCDP scenarios for the training, due to the limited experience of the prehospital providers at that time. RCDP offers ample opportunities for feedback with immediate practice and improvement. Trainees demonstrated retention of knowledge and improved performance in simulation-based testing. The overall satisfaction level of the trainees was high and suggests additional training would be beneficial and desired. Additionally, as the results of our needs assessment mirrored common chief complaints in other LMIC countries in Sub-Saharan Africa1,2 we feel that this curriculum can be utilized and adopted with minor modifications in other LMIC settings, particularly where EMS programs are developing and in circumstances where few EMS providers have had extensive field experience. Topics: Respiratory distress, asthma, dehydration, hypovolemic shock, hypoglycemia, seizure, toxic ingestion, newborn resuscitation, precipitous delivery, traumatic injury, EMS, Botswana, global health, collaboration, rapid cycle deliberate practice (RCDP), medical simulation.

Haptics-Based Apps for Middle School Students with Visual Impairments.

Twenty haptics-based computer applications (apps) have been created to utilize a low-cost, force feedback haptic device, the Novint Falcon, to provide students with tactile and kinesthetic sensations while learning about math and science. These low-cost apps, developed specifically for students with visual impairments (yet practical for all students), add to the accessible resources available for math and science. This article outlines the motivation, development, and testing of these PC-based applications that incorporate computer haptics, auditory cues, and high-contrast visuals. Included is a brief overview of two of the apps, one with science content and one with math content, in order to provide the reader with some insight into the student experience. The results of testing six of the apps in classroom settings show that the device and software are feasible for teachers to implement and significant learning gains can be achieved for students who use them. Student attitudes toward the apps were positive, implying that not only are the apps useful in the classroom, but engaging as well.

Staging laparoscopy in pancreatic cancer: a potential role for advanced laparoscopic techniques.

BACKGROUND: The role of staging laparoscopy in pancreatic cancer in the age of high-resolution CT scans is under debate. This study's aim is to evaluate the efficacy of staging laparoscopy in this disease. STUDY DESIGN: A retrospective cohort study was conducted evaluating patients who underwent operative treatment for radiographic stage I to III pancreatic cancer between July 2003 and October 2012. Radiographic follow-up was 94% at 6 months. RESULTS: Of 274 patients who met inclusion criteria, 136 underwent staging laparoscopy, which identified radiographic occult distant metastases in 2% (3 of 136). However, subsequent laparotomy identified an additional 9% (12 of 136) harboring distant metastases in regions not visualized on standard staging laparoscopy; specifically, the posterior liver surface, paraduodenal retroperitoneum, proximal jejunal mesentery, and lesser sac. The remaining 138 patients underwent initial staging laparotomy, which showed similar results identifying radiographic occult distant disease in 11% (15 of 138). Within 6 months after the operation, peritoneal or subcapsular liver metastases developed in an additional 6% (15 of 257)-disease that potentially could have been diagnosed at the time of operation-providing a false-negative rate of 88% for staging laparoscopy compared with 36% for staging laparotomy. CONCLUSIONS: Despite the availability of high-resolution CT scans, occult distant metastases can still be found in 11% of patients during the operation. In the absence of reliable risk factors to predict distant metastases, staging laparoscopy should be offered to all patients with radiographic localized disease. However, the results favor extended laparoscopic staging with evaluation of the posterior liver surface, mobilization of the duodenum, evaluation of the proximal jejunal mesentery, and visualization of the lesser sac.

Improving fall risk assessment in home care: interdisciplinary use of the Timed Up and Go (TUG).

The Timed Up and Go (TUG) is a popular, effective, and valid test of functional mobility and fall risk that is often completed by registered nurses (RNs) and physical therapists (PTs) throughout the course of a home care episode. As reimbursement becomes tied to outcomes, it is essential that all disciplines are consistent in their methods when administering the TUG. Results of this study confirm the hypothesis that test-specific training will significantly improve reliability of the TUG when completed by 2 different disciplines. The purpose of this article is to describe an initiative that provided tool-specific training to all clinical staff at our home care agency. The inter-rater reliability between PTs and RNs improved significantly from 0.77 to 0.86 (p = 0.001) after standardized training on administration of the TUG.

Template directed formation of nanoparticle decorated multi-walled carbon nanotube bundles with uniform diameter.

Bundles of multi-walled carbon nanotubes of uniform diameter decorated with Ni nanoparticles were synthesized using mesoporous silicates as templates. The ordered morphology and the narrow pore size distribution of mesoporous silicates provide an ideal platform to synthesize uniformly sized carbon nanotubes. In addition, homogeneous sub-10 nm pore sizes of the templates allow in situ formation of catalytic nanoparticles with uniform diameters which end up decorating the carbon nanotubes. The resulting carbon nanotubes are multi-walled with a uniform diameter corresponding to the pore diameter of the template used during the synthesis that are decorated with the catalysts used to synthesize them. They have a narrow size distribution which can be used in many energy related fields of research.

Folate and homocysteine phenotypes: Comparative findings using research and clinical laboratory data.

OBJECTIVES: A low folate/high homocysteine phenotype is associated with several pathologies, including spina bifida and cardiovascular disease. Folate and total homocysteine (tHcy) measurements are used clinically to assess risk and the need for folic acid supplementation and in research to investigate the metabolic basis of disease. Red blood cell (RBC) folate, the best indicator of long-term folate status, is usually measured as "total" folate. However, different folate derivatives support distinct biochemical functions, suggesting a need to develop more precise methods. This study was designed to evaluate a method based on stable isotope dilution liquid chromatography-multiple reaction monitoring/mass spectrometry (LC-MRM/MS). DESIGN AND METHODS: We used LC-MRM/MS to quantify the RBC folate derivatives 5-methyltetrahydrofolate (5-CH(3)-THF), tetrahydrofolate (THF), and 5,10-methenyltetrahydrofolate (5,10-methenylTHF) in pre-menopausal women. The concentration of each folate derivative was assessed for utility in predicting tHcy levels, and compared to folate and tHcy measurements derived by routine clinical laboratory methods. RESULTS: LC-MRM/MS was qualitatively and quantitatively superior to routine clinical laboratory methods for determining folate and tHcy concentrations. RBC 5-CH(3)-THF had a reciprocal relationship with tHcy (p=0.0003), whereas RBC THF and RBC 5,10-methenylTHF had direct relationships (p=0.01, 0.04 respectively). In combination, these three variables accounted for 42% of the variation in tHcy. CONCLUSIONS: Robust methods for measuring RBC 5-CH(3)-THF would improve the utility of folate/homocysteine phenotyping in patient management. The use of LC-MRM/MS would allow studies of hyperhomocysteinemia and diseases associated with a low folate/high homocysteine phenotype to be performed with less measurement error and greater statistical power to generate data with the potential to elucidate the etiologic mechanisms of complex diseases and traits.

Genetic and biochemical determinants of serum concentrations of monocyte chemoattractant protein-1, a potential neural tube defect risk factor.

BACKGROUND: Women with the AA genotype at the (-2518)A>G promoter polymorphism of CCL-2, which encodes the potent pro-inflammatory chemokine monocyte chemoattractant protein 1 (MCP-1), may be at increased risk for having offspring affected by spina bifida. As the A allele at this locus has been associated with decreased transcription of MCP-1 mRNA relative to the G allele, the observed genetic association suggests that the risk of spina bifida may be increased in the offspring of women with low MCP-1 levels. The present study was undertaken to identify potential determinants of MCP-1 levels in women of reproductive age. METHODS: A small cohort of Caucasian and African-American women of reproductive age was recruited to participate in an exploratory investigation of the determinants of several disease-related, biochemical phenotypes, including MCP-1. Subjects completed a brief questionnaire and provided a fasting blood sample for biochemical and genetic studies. Potential biochemical, genetic, and lifestyle factors were assessed for their association with MCP-1 levels using linear regression analyses. RESULTS: In this cohort, MCP-1 levels were significantly higher in Caucasians as compared to African-Americans. Further, among women of both races, there was evidence that MCP-1 levels were associated with smoking status, MTHFR 677C>T genotype, and red blood cell tetrahydrofolate levels. CONCLUSIONS: The results of these analyses indicate that, if maternal CCL-2 genotype is related to the risk of spina bifida, this relationship is likely to be more complex than initially hypothesized, perhaps depending upon folate intake, MTHFR 677C>T genotype, the distribution of folate derivatives, and immune/inflammatory activity.

The surprisingly elusive crystal structure of sodium metabisulfite.

The crystal structure of Na2S2O5, a simple and common ionic compound, is reported here for the first time. The crystals form non-merohedral twins, with the twin domains related by a twofold axis which bisects the angle between the a and c axes of each unit cell. The structure was determined from a single-crystal fragment of a twinned crystal that had undergone cleavage along the twin boundary. In addition to the problems associated with twinning, space-group determination proved difficult as well, with the structure initially determined in the P2(1) space group appearing to be disordered with two rotational conformers of the metabisulfite ion occupying equivalent sites in the lattice. An analysis at low temperature provided new weak reflections which were inconsistent with the original unit cell, but indexed to the correct unit cell, allowing for space group and crystal structure determination. The apparent structure, which appeared disordered in P2(1), seems to have resulted from an apparently fortuitous superposition of two conformationally inequivalent S2O5(2-) anions in the asymmetric unit of the correct structure in the P2(1)/n space group. The metabisulfite ions in this structure do not adopt the C(s) geometry observed in previously determined crystal structures containing S2O5(2-). The structures of both ions in the asymmetric unit are effectively conformational mirror images of one another with two of the O atoms on each S atom in the ion approaching an eclipsed geometry. This observation provides further evidence that the structures of sulfur-oxy anions in the solid state are dictated mainly by interionic coulombic forces rather than by intraionic bonding interactions