Growth of wildtype and mutant E. coli strains in minimal media for optimal production of nucleic acids for preparing labeled nucleotides

Since RNAs lie at the center of most cellular processes, there is a need for synthesizing large amounts of RNAs made from stable isotope-labeled nucleotides to advance the study of their structure and dynamics by nuclear magnetic resonance (NMR) spectroscopy. A particularly effective means of obtaining labeled nucleotides is to harvest these nucleotides from bacteria grown in defined minimal media supplemented with 15NH4Cl and various carbon sources. Given the high cost of carbon precursors required for labeling nucleic acids for NMR studies, it becomes important to evaluate the optimal growth for commonly used strains under standard minimal media conditions. Such information is lacking. In this study, we characterize the growth for Escherichia coli strains K12, K10zwf, and DL323 in three minimal media with isotopic-labeled carbon sources of acetate, glycerol, and glycerol combined with formate. Of the three media, the LeMaster-Richards and the Studier media outperform the commonly used M9 media and both support optimal growth of E. coli for the production of nucleotides. However, the growth of all three E. coli strains in acetate is reduced almost twofold compared to growth in glycerol. Analysis of the metabolic pathway and previous gene array studies help to explain this differential growth in glycerol and acetate. These studies should benefit efforts to make selective 13C-15N isotopic-labeled nucleotides for synthesizing biologically important RNAs

Older adults’ perspectives on using digital technology to maintain good mental health: interactive group study

Background: A growing number of apps to support good mental health and wellbeing are available on digital platforms. However, very few studies have examined older adults’ attitudes towards the use of these, despite increasing uptake of digital technologies by this demographic. Objective: The present study sought to explore older adults’ perspectives on technology to support good mental health. Methods: Fifteen older adults (>50 years old), in two groups, participated in sessions to explore the use of digital technologies to support mental health. Interactive activities were designed to capture participants’ immediate reactions to apps and websites designed to support mental health, and explore their experiences of using technology for these purposes in their own lives. Template analysis was used to analyse transcripts of the group discussions. Results: Older adults were motivated to turn to technology to improve mood through mechanisms of distraction, normalisation and facilitated expression of mental states, while aiming to reduce burden on others. Perceived barriers to use included fear of consequences, and the impact of low mood on readiness to engage with technology, as well as a lack of prior knowledge applicable to digital technologies. Participants were aware of websites available to support mental health, but awareness alone did not motivate use. Conclusions: Older adults are motivated to use digital technologies to improve their mental health, but barriers remain that developers need to address for this population to access them

Mitochondrial Dysfunction in Diaphragm Muscle Precedes the Cachectic Phenotype in LLC Tumor-Bearing Mice.

The defining feature of cancer cachexia is extensive weight loss and skeletal muscle atrophy. It is clinically important because cachexia reduces patient survival, results in functional impairment, and is estimated to be directly responsible for 20-40% of cancer deaths. Unfortunately, no clinical therapy exists and therefore, it is important to understand the molecular mechanisms responsible for rapid muscle wasting. Compared to limb muscles, the diaphragm is relatively understudied in cancer cachexia, but is likely to be adversely affected because cachexia is a systemic disease. Wasting of the primary inspiratory muscle may result in difficulty breathing and inability to adjust minute ventilation in response to a respiratory challenge. Based on emerging evidence, it is clear that oxidative stress is present in cachexia-induced wasting of the diaphragm; PURPOSE: we developed the hypothesis that mitochondrial dysfunction in the diaphragm precedes cachexia. METHODS: 1X106 Lewis Lung Carcinoma cells (LLC) or Phosphate-Buffered Saline (PBS, control) were implanted to the hind-flank of C57BL6/J mice at 8 wks of age. Tumors were allowed to develop for 1, 2, 3, or 4 wks. At designated time points diaphragms were collected and mitochondrial function was assessed by respiration and ROS production. RESULTS: Cancer cachexia was evident only at the 4 wk time point demonstrated by decrease in body mass and muscle atrophy in several limb muscles. Mitochondrial respiration, assessed by respiratory control ratio (state3/state 4 respiration), was significantly lower at 1 wk (pCONCLUSIONS:The molecular events that lead to muscle atrophy in cancer cachexia are unknown. We demonstrate that two hallmarks of mitochondrial dysfunction, altered respiration and ROS production, occur in the diaphragm well before the cancer cachexia phenotype is evident in the LLC model. These data suggest that the mitochondria are likely a suitable target to treat or prevent cancer cachexia-induced muscle wasting in the diaphragm

How The Mountain West States Voted In 2016: A Post-Election Analysis of Trends, Demographics, and Politics in America’s New Swing Region

Brookings Mountain West and the Greenspun College of Urban Affairs hosted a panel of experts in state and regional politics and history to examine election returns and exit polling and provide a first-read of the 2016 election. The Mountain West is now one of the nation’s most contested political regions. Its population growth and ever-shifting demographics make the region harder to predict and most susceptible to political swings. Five states in the Southern Mountain West – Arizona, Colorado, Nevada, New Mexico, and Utah – now hold more electoral votes than all individual states except Texas and California. In our current political climate these 37 electoral votes can determine the majority in the U.S. Senate and the presidency of the United States

Reversible shear thickening at low shear rates of electrorheological fluids under electric fields

Shear thickening is a phenomenon of significant viscosity increase of colloidal suspensions. While electrorheological (ER) fluids can be turned into a solid-like material by applying an electric field, their shear strength is widely represented by the attractive electrostatic interaction between ER particles. By shearing ER fluids between two concentric cylinders, we show a reversible shear thickening of ER fluids above a low critical shear rate (<1 s-1) and a high critical electric field strength (>100 V/mm), which could be characterized by a modified Mason number. Shear thickening and electrostatic particle interaction-induced inter-particle friction forces is considered to be the real origin of the high shear strength of ER fluids, while the applied electric field controls the extent of shear thickening. The electric field-controlled reversible shear thickening has implications for high-performance ER/magnetorheological (MR) fluid design, clutch fluids with high friction forces triggered by applying local electric field, other field-responsive materials and intelligent systems.Comment: 29pages, 9 figure

VarSight: prioritizing clinically reported variants with binary classification algorithms.

BackgroundWhen applying genomic medicine to a rare disease patient, the primary goal is to identify one or more genomic variants that may explain the patient's phenotypes. Typically, this is done through annotation, filtering, and then prioritization of variants for manual curation. However, prioritization of variants in rare disease patients remains a challenging task due to the high degree of variability in phenotype presentation and molecular source of disease. Thus, methods that can identify and/or prioritize variants to be clinically reported in the presence of such variability are of critical importance.MethodsWe tested the application of classification algorithms that ingest variant annotations along with phenotype information for predicting whether a variant will ultimately be clinically reported and returned to a patient. To test the classifiers, we performed a retrospective study on variants that were clinically reported to 237 patients in the Undiagnosed Diseases Network.ResultsWe treated the classifiers as variant prioritization systems and compared them to four variant prioritization algorithms and two single-measure controls. We showed that the trained classifiers outperformed all other tested methods with the best classifiers ranking 72% of all reported variants and 94% of reported pathogenic variants in the top 20.ConclusionsWe demonstrated how freely available binary classification algorithms can be used to prioritize variants even in the presence of real-world variability. Furthermore, these classifiers outperformed all other tested methods, suggesting that they may be well suited for working with real rare disease patient datasets

Disuse Atrophy Occurs Without a Change in Mitochondrial Respiratory Control Ratio During Hindlimb Unloading in Mice

Skeletal muscle atrophy commonly occurs during prolonged periods of inactivity, however, the precise mechanisms that cause muscle atrophy have yet to be determined. Specifically, a controversy exists on whether mitochondrial dysfunction is a cause or consequence of disuse muscle atrophy. PURPOSE: The purpose of this study was to determine if a change in the respiratory control ratio, which is a ratio of maximal O2 respiration to leak respiration, could be detected prior to muscle atrophy in a time-course study in mice. METHODS: Disuse atrophy was induced using hindlimb unloading (HU) in adult, C57BL/6J male mice for 0 (control), 1, 2, 3, or 7 days (n=~6-8/group). Following completion, gastrocnemius and soleus muscles were weighed and assessed for mitochondrial function in permeabilized muscle fibers. Here, we define mitochondrial function as the respiratory control ratio (RCR) determined by maximal ADP stimulated respiration (State 3) divided by leak or ATP synthase inhibited (state 4) respiration. A one-way ANOVA was used to determine differences between means. When significant F ratios were found, a Tukey post-hoc was used to compare differences between means. Values presented are mean ± standard error RESULTS: In both the gastrocnemius and soleus, muscle mass was not significantly different from control at day 1, but was significantly lower at 2, 3, and 7-day timepoints. In contrast, there was no significant difference in RCR in gastrocnemius (control 3.11±0.20, 1 day 2.96±0.60, 2 day 3.07±0.31, 3 day 3.08±0.25, 7 day 3.41± .29) or soleus (control 2.33±0.33, 1 day 2.77±0.33, 2 day 3.03±0.51, 3 day 2.93±0.30, 7 day 2.78±0.48). CONCLUSION: It is well established that HU causes rapid muscle atrophy. These data support mitochondrial RCR does decrease before muscle atrophy in either gastrocnemius or soleus muscle, and therefore may not be a primary cause of HU-induced muscle atrophy in mice

Fractional Synthetic Rate and Markers of Protein Turnover are Altered in the Diaphragms of Cachectic Mice

Cancer cachexia, a wasting syndrome characterized by rapid skeletal muscle wasting and fat loss, directly accounts for up to 20-40% of cancer-related deaths. All muscles, including respiratory muscles, are susceptible to atrophy because cancer cachexia is a systemic disease. Atrophy of the primary breathing muscle, the diaphragm, can lead to respiratory distress, which is commonly associated with a cachectic phenotype. Indeed, the diaphragm is more susceptible to atrophy in certain conditions, but little is known about the effects of cancer-cachexia on protein turnover in the diaphragm. Therefore, investigations into the alterations in protein turnover could provide insight to the molecular events and provide valuable information in the search for therapeutic targets. PURPOSE: The purpose of this study was to describe changes in diaphragmatic protein synthesis and molecular markers of synthesis and degradation during the progression of cancer cachexia. METHODS: C57BL6/J mice (8 wks old) were implanted with 1X106 Lewis Lung Carcinoma cells (LLC) or Phosphate-Buffered Saline (PBS, control). Tumors developed over a 1-4 wk time course and diaphragms were harvested at each time point (1, 2, 3, or 4 wks). Fractional synthetic rates (FSR) were determined using deuterium incorporation into muscle. Selected markers of protein synthesis and degradation pathways were analyzed by immunoblot analysis. One-Way ANOVA was used for statistical analyses, with significance set at pRESULTS: FSR trended downward over time, but did not reach significance. Similar to FSR, anabolic signaling markers (4EBP-1, ERK1/2, Deptor) did not demonstrate significant differences. p62, an autophagic degradation marker, was significantly less than PBS in 3 wk diaphragms (