bluuebunny/crossref_metadata_2025 huggingface dataset:
Are Nonprofit Hospitals Addressing the Most Critical Community Health Needs That They Identify in Their Community Health Needs Assessments?
Authors: Franz Berkeley , Cronin Cory E. , Singh Simone
Published: August 2019

Context:

Virtually all nonprofit hospitals are in compliance with the Affordable Care Act's new Community Health Needs Assessments requirements.

Objective:

To assess what needs have emerged in the Community Health Needs Assessments hospitals complete nationally, the degree to which identified needs reflect the most pressing community health issues, and the extent to which hospitals address identified needs.

Design:

Using both bivariate and logistic regressions, we analyzed the Community Health Needs Assessments and implementation strategies of nonprofit hospitals to determine whether identified needs overlapped with county health-ranking indicators of need and whether institutional or community-level factors predicted hospital willingness to address identified needs.

Participants:

We included a 20% random sample of US nonprofit hospitals (n = 496).

Main Outcome Measures:

Our main outcome measures were whether nonprofit hospitals addressed each of the most common needs.

Results:

Mental health, access to care, obesity, substance abuse, diabetes, cancer, and the social determinants of health were the most commonly identified needs across the sample. The rate at which hospitals chose to address each of these needs in their implementation strategies, however, varied considerably, ranging from 56% (cancer) to 85% (obesity). We found that several institutional and community characteristics predicted hospital willingness to address each need; whether the community ranked a need as number 1 was a better predictor of hospital investment than the severity of the need, as measured by county health-rankings data.

Conclusions:

These findings may help inform local, state, and federal policy makers as they consider interventions aimed at encouraging hospitals to invest in improving the health of their communities.
Identifying Novel Predictors of State Legislative Action to Address Obesity
Authors: Arons Abigail , Pomeranz Jennifer , Hamad Rita
Published: August 2019

Objective:

There is wide variation in the number and types of obesity policies enacted across states, and prior studies suggest that partisan factors may not fully explain this variation. In this exploratory analysis, we examined the association of a broad array of state-level factors with the number and types of obesity policies across states.

Design:

We analyzed 32 predictor variables across 7 categories of state-level characteristics. We abstracted data from 1652 state obesity policies introduced during 2009-2014. We used multilevel regression models and principal component analysis to examine the association between state-level characteristics and policy outcomes.

Main Outcome Measures:

Our outcome measures included whether bills involved topics that were public health–oriented or business interest–oriented, whether bills were enacted into law, and the number of introduced bills and enacted laws per state.

Results:

Numerous state-level characteristics were associated with obesity-related bill introduction and law enactment, and different state characteristics were associated with public health–oriented versus business interest–oriented policies. For example, state-level demographics, economic factors, policy environment, public programs, and the prevalence of obesity's downstream consequences were associated with the number of public health laws whereas obesity prevalence and policy environment were associated with the number of business interest laws.

Conclusions:

Our results support the hypothesis that a variety of factors contribute to a complex state obesity policymaking environment, highlighting the need for future research to disentangle these key predictors.
Expert Interviews on Multilevel Barriers in Implementing Expedited Partner Therapy for Chlamydia
Authors: McCool-Myers Megan , Smith Akimi Dan-Yei , Kottke Melissa J.
Published: August 2019

Expedited partner therapy (EPT) is an evidence-based practice in which partners of patients with chlamydia are given antibiotics or a prescription for antibiotics without prior medical evaluation. This practice facilitates partner treatment and prevents reinfection of the patient. EPT remains underutilized due to multilevel implementation barriers. Barriers may exist at the level of legislation, policy, health care system, health care provider (HCP), pharmacist, patient, or partner. Qualitative interviews were performed with 11 EPT experts across the United States to uncover barriers in implementation. Thirty-four barriers were identified and grouped into 1 of 15 themes. The themes that surfaced most frequently were liability and adverse events (policy and HCP level), funding (policy level), electronic medical records (health care system level), and awareness (HCP and pharmacy level). Tailored implementation strategies are needed to increase awareness among HCPs and pharmacists, to establish funding and to streamline processes in health care systems.
Diurnal variation of inflammatory plasma proteins involved in pain
Authors: Jasim Hajer , Carlsson Anders , Gerdle Björn , Ernberg Malin , Ghafouri Bijar
Published: August 2019

Introduction:

Proteomics is a powerful approach for biochemical research because it directly studies the main functional components of biochemical systems. The understanding of the normal fluctuations of the proteome in health is essential to identify pain-specific biomarkers.

Objective:

To investigate fluctuations of the plasma proteome in healthy pain-free individuals.

Methods:

Blood samples were structurally collected in the early morning and evening from 10 clinically healthy individuals (26.3 ± 3.3 years). High abundant proteins were removed from plasma, and proteins were then analysed by nanoliquid chromatography combined with mass spectrometry. In addition, an assay of 71 cytokines/chemokines/growth factors was analysed.

Results:

Multivariate statistical analysis displayed that there were up to 64 proteins whose expression levels were significantly altered between the plasma samples collected during the morning and evening; no changes existed for the assay. The levels of 34 proteins were increased and 30 proteins were decreased during the evening compared with the morning sample. The increased proteins were involved in the biological processes such as protein activation cascade, complement activation, and stress response. The decreased proteins were involved in regulation of endopeptidase activity, inflammatory response, and protein metabolic processes.

Conclusion:

The circadian variations in the plasma proteome stress the need to collect blood samples of both patients and controls at a fixed time of the day. The results in this study might be useful for better understanding of the complexity of individual variation in the human plasma proteome over time and provide a baseline for improved pain biomarker discovery.
Characterization of Interstitial Lung Disease Associated With Anti-Ribonucleoprotein Antibodies
Authors: Lhote Raphael , Grenier Philippe , Haroche Julien , Miyara Makoto , Boussouar Samia , Mathian Alexis , Pha Micheline , Amoura Zahir , Cohen Aubart Fleur
Published: August 2019

Objectives

Interstitial lung disease (ILD) is a common feature of mixed connective tissue disease. However, many patients do not meet the criteria for mixed connective tissue disease and thus may be diagnosed as interstitial pneumonia with autoimmune features. The aim of this study was to characterize ILD associated with anti-ribonucleoprotein (RNP) antibodies.

Methods

Chest computed tomography scans of patients with anti-RNP antibody who were seen between January 2011 and October 2015 were reviewed. The underlying disease was classified with international criteria using clinical and serological features.

Results

Among 544 patients with anti-RNP antibodies, 188 had a chest computed tomography scan, and 48 (26%) of them had radiological features of ILD. The presence of ILD was significantly associated with dyspnea, crackles, arthritis, Raynaud phenomenon, myositis, and sicca syndrome. The most frequent pattern was nonspecific interstitial pneumonia in 39 patients (81%). Among patients with ILD, 17 (35%) had a radiological pattern consisting of cysts and ground-glass attenuation not fulfilling the lymphoid interstitial pneumonia criteria. In 3 patients, cysts were related to fibrosis; in 14 patients, cysts corresponded to an original ILD pattern.

Conclusions

Interstitial lung disease was found in 26% of patients with anti-RNP antibodies independently of the underlying disease. Anti-RNP–associated ILD mainly corresponds to nonspecific interstitial pneumonia or an original pattern consisting of cysts and ground-glass attenuation.
Patient-Reported Outcome Measures in Plastic Surgery
Authors: Sharma Kavita , Steele Kathryn , Birks Meg , Jones Georgina , Miller Gavin
Published: August 2019

Clinical care is often directed at improving patient's quality of life, the effectiveness of which can be measured by objective or subjective outcomes. Surgical intervention, especially in plastic and reconstructive surgery, can often provide hard objective measures, for example, breast size reduction after bilateral reduction mammaplasty, or improved joint position/movement after Dupuytren's surgery. These measurements do not describe how a particular intervention affects the patient's life, from their point of view.

Patient-reported outcome measures (PROMs) are validated questionnaires completed by patients about their health, functioning, health behaviors and quality of care. Patient-reported outcome measures reflect the patient's perspective and their use increases the meaningfulness of outcomes measured, for whatever purpose. There is therefore a growing interest in PROMs as part of routine clinical practice in the United Kingdom and worldwide.

This article aims to provide an introduction of PROMs to plastic surgeons, a description of how PROMs are developed and scored, along with the most commonly used tools currently most applicable to plastic surgery. The benefits of PROMs on a local, regional, and national level are discussed, and we aim to set out how these tools can be used in auditing individual and departmental performance against national standards. We will also provide suggestions for sustainable use of PROMs in the clinical environment, advice on choosing the right outcome measure, and our views on electronic data collection, analysis, and interpretation of the results.

Patient-reported outcome measures are highly relevant clinical practice and will undoubtedly with time become routine. We encourage plastic surgeons to explore the possibilities these tools can provide in helping improve the quality of care we deliver to our patients.
Autologous Fat Grafting in Young Patients
Authors: Maia Munique , Lukash Frederick N.
Published: August 2019

Goals/Purpose

Rhinoplasty is the most common procedure seen in the teenage population. Many of these patients have facial imbalance both recognized and unrecognized by the individual and family. Most often it involves chin or midface deficiency. When the discussion involves the possibility of additional surgery, such as an alloplastic chin implant or a surgical osteotomy, the conversation halts because the patient and family seek a simpler solution.

Autologous fat transfer is a useful adjunct to achieve facial balance in chin and cheek in the teen population. More recently, we have been using this technique to correct facial disharmony in rhinoplasty patients.

Methods/Technique

Fat grafting was performed at the time of rhinoplasty in 22 patients (age, 15–19 years). Presurgical planning involved cephalometric and computer-enhanced photographic analysis of the face. Midface retrusion and underprojected mentum were treated. Deficient sites were treated with small aliquots of fat, which were injected into the supraperiosteal plane. The average amounts of fat grafting injected per region were 2 to 3 mL for the malar region and 8 to 10 mL for the chin and geniomandibular borders.

Results/Complications

Twenty-two patients underwent augmentation of soft tissue and skeletal deficiencies. Analysis of postoperative results showed excellent outcomes with enhancement of facial profile and proportions. The mean follow-up was 3 years. The majority of patients reported high degree of satisfaction. No complications were observed.

Conclusions

The combination of rhinoplasty and autologous fat grafting offers very satisfactory esthetic outcomes and improvement of facial balance with minimal added time, cost, and risk.
Gallstone Pancreatitis
Authors: Mueck Krislynn M. , Wei Shuyan , Pedroza Claudia , Bernardi Karla , Jackson Margaret L. , Liang Mike K. , Ko Tien C. , Tyson Jon E. , Kao Lillian S.
Published: August 2019

Introduction:

Early cholecystectomy shortly after admission for mild gallstone pancreatitis has been proposed based on observational data. We hypothesized that cholecystectomy within 24 hours of admission versus after clinical resolution of gallstone pancreatitis that is predicted to be mild results in decreased length-of-stay (LOS) without an increase in complications.

Methods:

Adults with predicted mild gallstone pancreatitis were randomized to cholecystectomy with cholangiogram within 24 hours of presentation (early group) versus after clinical resolution (control) based on abdominal exam and normalized laboratory values. Primary outcome was 30-day LOS including readmissions. Secondary outcomes were time to surgery, endoscopic retrograde cholangiopancreatography (ERCP) rates, and postoperative complications. Frequentist and Bayesian intention-to-treat analyses were performed.

Results:

Baseline characteristics were similar in the early (n = 49) and control (n = 48) groups. Early group had fewer ERCPs (15% vs 29%, P = 0.038), faster time to surgery (16 h vs 43 h, P < 0.005), and shorter 30-day LOS (50 h vs 77 h, RR 0.68 95% CI 0.65 – 0.71, P < 0.005). Complication rates were 6% in early group versus 2% in controls (P = 0.613), which included recurrence/progression of pancreatitis (2 early, 1 control) and a cystic duct stump leak (early). On Bayesian analysis, early cholecystectomy has a 99% probability of reducing 30-day LOS, 93% probability of decreasing ERCP use, and 72% probability of increasing complications.

Conclusion:

In patients with predicted mild gallstone pancreatitis, cholecystectomy within 24 hours of admission reduced rate of ERCPs, time to surgery, and 30-day length-of-stay. Minor complications may be increased with early cholecystectomy. Identification of patients with predicted mild gallstone pancreatitis in whom early cholecystectomy is safe warrants further investigation.
Are results from randomized trials in anesthesiology robust or fragile? An analysis using the fragility index
Authors: Goerke Kale , Parke Matthew , Horn Jarryd , Meyer Chase , Dormire Kody , White Brad , Bright Trevor , Walters Corbin , Vassar Matt
Published: August 2019

Aim:

In anesthesiology, the findings from randomized controlled trials often underpin guidelines influencing clinical decision-making and therefore directly affect patient care. The aim of this study is to evaluate the fragility index and fragility quotient of randomized controlled trials published in the eight highest ranked anesthesiology journals. In addition, we assess the extent to which risk of bias scores, loss to follow-up, Web of Science Citation Index, and journal impact factor influence fragility index and fragility quotient.

Methods:

We included randomized trials published between 2014 and 2016 from the eight highest ranking anesthesiology journals based on Clarivate Analytics’ Science Citation Index and Google Scholar Metrics: Anesthesiology subcategory. We included journals that published general anesthesia topics and omitted specialty anesthesia journals. The fragility index and fragility quotient for all included trials were calculated. Risk of bias for each trial was evaluated using the Cochrane ‘risk of bias’ Tool 2.0.

Results:

One hundred and thirty one randomized control trials were included in this analysis. The median fragility index was 3 (interquartile range 1.0–5.5) with a fragility quotient of 0.03 (interquartile range 0.01–0.08). In 11% (14/131) of trials, the number of patients lost to follow-up was greater than the corresponding fragility index. Weak correlations were found between fragility index and total sample size (r = 0.13) and between fragility index and event frequency (r = 0.19). A near-negligible correlation was found between 5-year impact factor and fragility index (r = −0.03) and, similarly, between fragility index and Science Citation Index (r = −0.05). Ten trials were at high risk of bias with the randomization process found to be the domain at the highest risk of bias.

Conclusion:

In assessing the fragility of randomized controlled trials published in the top eight anesthesiology journals, our study suggests that statistically significant results in these journals are disconcertingly fragile. The median fragility index calculated from our 131 primary studies reveals that only three nonevents must be replaced with events to negate statistical significance. Although a current scale does not exist for fragility index ranges, many trials published by the top journals in anesthesiology are based on concerning methodology and highly fragile outcomes. With small median sample sizes and few patient events characterizing a large number of these trials, many of today's current guidelines and clinical practices may be founded on research containing statistical significance but lacking clinical significance.
Elucidating user behaviours in a digital health surveillance system to correct prevalence estimates
Authors: Liu Dennis , Mitchell Lewis , Cope Robert C. , Carlson Sandra J. , Ross Joshua V.
Published: August 2019

Estimating seasonal influenza prevalence is of undeniable public health importance, but remains challenging with traditional datasets due to cost and timeliness. Digital epidemiology has the potential to address this challenge, but can introduce sampling biases that are distinct to traditional systems. In online participatory health surveillance systems, the voluntary nature of the data generating process must be considered to address potential biases in estimates. Here we examine user behaviours in one such platform, FluTracking, from 2011 to 2017. We build a Bayesian model to estimate probabilities of an individual reporting in each week, given their past reporting behaviour, and to infer the weekly prevalence of influenza-like-illness (ILI) in Australia. We show that a model that corrects for user behaviour can substantially effect ILI estimates. The model examined here elucidates several factors, such as the status of having ILI and consistency of prior reporting, that are strongly associated with the likelihood of participating in online health surveillance systems. This framework could be applied to other digital participatory health systems where participation is inconsistent and sampling bias may be of concern.
A non-canonical role for p27Kip1in restricting proliferation of corneal endothelial cells during development
Authors: Defoe Dennis M. , Rao Huiying , Harris David J. , Moore Preston D. , Brocher Jan , Harrison Theresa A.
Published: August 2019

The cell cycle regulator p27Kip1is a critical factor controlling cell number in many lineages. While its anti-proliferative effects are well-established, the extent to which this is a result of its function as a cyclin-dependent kinase (CDK) inhibitor or through other known molecular interactions is not clear. To genetically dissect its role in the developing corneal endothelium, we examined mice harboring two loss-of-function alleles, a null allele (p27−) that abrogates all protein function and a knockin allele (p27CK−) that targets only its interaction with cyclins and CDKs. Whole-animal mutants, in which all cells are either homozygous knockout or knockin, exhibit identical proliferative increases (∼0.6-fold) compared with wild-type tissues. On the other hand, use of mosaic analysis with double markers (MADM) to produce infrequently-occurring clones of wild-type and mutant cells within the same tissue environment uncovers a roughly three- and six-fold expansion of individualp27CK−/CK−andp27−/−cells, respectively. Mosaicism also reveals distinct migration phenotypes, withp27−/−cells being highly restricted to their site of production andp27CK−/CK−cells more widely scattered within the endothelium. Using a density-based clustering algorithm to quantify dispersal of MADM-generated clones, a four-fold difference in aggregation is seen between the two types of mutant cells. Overall, our analysis reveals that, in developing mouse corneal endothelium, p27 regulates cell number by acting cell autonomously, both through its interactions with cyclins and CDKs and through a cyclin-CDK-independent mechanism(s). Combined with its parallel influence on cell motility, it constitutes a potent multi-functional effector mechanism with major impact on tissue organization.
Extended-spectrum beta-lactamase (ESBL)-producing and non-ESBL-producing Escherichia coli isolates causing bacteremia in the Netherlands (2014 – 2016) differ in clonal distribution, antimicrobial resistance gene and virulence gene content
Authors: van Hout Denise , Verschuuren Tess D. , Bruijning-Verhagen Patricia C.J. , Bosch Thijs , Schürch Anita C. , Willems Rob J.L. , Bonten Marc J.M. , Kluytmans Jan A.J.W.
Published: August 2019

Background

Knowledge on the molecular epidemiology of Escherichia coli causing E. coli bacteremia (ECB) in the Netherlands is mostly based on extended-spectrum beta-lactamase-producing E. coli (ESBL-Ec). We determined differences in clonality and resistance and virulence gene (VG) content between non-ESBL-producing E. coli (non-ESBL-Ec) and ESBL-Ec blood isolates with different epidemiological characteristics.

Materials/methods

A random selection of non-ESBL-Ec isolates as well as all available ESBL-Ec blood isolates was obtained from two Dutch hospitals between 2014 and 2016. Whole genome sequencing was performed to infer sequence types (STs), serotypes, acquired antibiotic resistance genes and VG scores, based on presence of 49 predefined putative pathogenic VG.

Results

ST73 was most prevalent among the 212 non-ESBL-Ec (N=26, 12.3%) and ST131 among the 69 ESBL-Ec (N=30, 43.5%). Prevalence of ST131 among non-ESBL-Ec was 10.4% (N=22, P value < 0.001 compared to ESBL-Ec). O25:H4 was the most common serotype in both non-ESBL-Ec and ESBL-Ec. Median acquired resistance gene counts were 1 (IQR 1 – 6) and 7 (IQR 4 – 9) for non-ESBL-Ec and ESBL-Ec, respectively (P value < 0.001). Among non-ESBL-Ec, acquired resistance gene count was highest among blood isolates from a primary gastro-intestinal focus (median 4, IQR 1 – 8). Median VG scores were 13 (IQR 9 – 20) and 12 (IQR 8 – 14) for non-ESBL-Ec and ESBL-Ec, respectively (P value = 0.002). VG scores among non-ESBL-Ec from a primary urinary focus (median 15, IQR 11 – 21) were higher compared to non-ESBL-Ec from a primary gastro-intestinal (median 10, IQR 6 – 13) or hepatic-biliary focus (median 11, IQR 5 – 18) (P values = 0.007 and 0.036, respectively). VG content varied between different E. coli STs.

Conclusions

Non-ESBL-Ec and ESBL-Ec blood isolates from two Dutch hospitals differed in clonal distribution, resistance gene and VG content. Also, resistance gene and VG content differed between non-ESBL-Ec from different primary foci of ECB.
Comparison of visceral adipose tissue DNA methylation and gene expression profiles in female adolescents with obesity
Authors: Barberio Matthew D. , Nadler Evan P. , Sevilla Samantha , Lu Rosemary , Harmon Brennan , Hubal Monica J.
Published: August 2019

Background

Epigenetic changes in visceral adipose tissue (VAT) with obesity and their effects on gene expression are poorly understood, especially during emergent obesity in youth. The current study tested the hypothesis that methylation and gene expression profiles of key growth factor and inflammatory pathways such as PI3K/AKT signaling are altered in VAT from obese compared to non-obese youth.

Methods

VAT samples from adolescent females grouped as Lean (L; n=15; age=15±3 yrs, BMI=21.9±3.0 kg/m2) or Obese (Ob; n=15, age=16±2 yrs, BMI=45.8±9.8 kg/m2) were collected. Global methylation (n=20) and gene expression (N=30) patterns were profiled via microarray and interrogated for differences between groups by ANCOVA (p<0.05), followed by biological pathway analysis.

Results

Overlapping differences in methylation and gene expression in 317 genes were found in VAT from obese compared to lean groups. PI3K/AKT Signaling (p=1.83×10−6; 10/121 molecules in dataset/pathway) was significantly overrepresented in Ob VAT according to pathway analysis. mRNA upregulations in the PI3K/AKT Signaling Pathway genes TFAM (p=0.03; Fold change=1.8) and PPP2R5C (p=0.03, FC=2.6) were confirmed via qRT-PCR.

Conclusion

Our analyses show obesity-related differences in DNA methylation and gene expression in visceral adipose tissue of adolescent females. Specifically, we identified methylation site/gene expression pairs differentially regulated and mapped these differences to PI3K/AKT signaling, suggesting that PI3K/AKT signaling pathway dysfunction in obesity may be driven in part by obesity-related changes in DNA methylation.
Leptin Resistance in the Ovary of Obese Mice Is Associated with Profound Changes in the Transcriptome of Cumulus Cells
Authors: Wołodko Karolina , Walewska Edyta , Adamowski Marek , Castillo-Fernandez Juan , Kelsey Gavin , Galvão António
Published: August 2019

Background/Aims

Obesity is associated with infertility, decreased ovarian performance and lipotoxicity. However, little is known about the aetiology of these reproductive impairments. Here, we hypothesise that the majority of changes in ovarian physiology in diet-induced obesity (DIO) are a consequence of transcriptional changes downstream of altered leptin signalling. Therefore, we investigated the extent to which leptin signalling is altered in the ovary upon obesity with particular emphasis on effects on cumulus cells (CCs), the intimate functional companions of the oocyte. Furthermore, we used the pharmacological hyperleptinemic (LEPT) mouse model to compare transcriptional profiles to DIO.

Methods

Mice were subjected to DIO for 4 and 16 weeks (wk) and leptin treatment for 16 days, to study effects in the ovary in components of leptin signalling at the transcript and protein levels, using Western blot, Real-time PCR and immunostaining. Furthermore, we used low-cell RNA sequencing to characterise changes in the transcriptome of CCs in these models.

Results

In the DIO model, obesity led to establishment of ovarian leptin resistance after 16 wk high fat diet (HFD), as evidenced by increases in the feedback regulator suppressor of cytokine signalling 3 (SOCS3) and decreases in the positive effectors phosphorylation of tyrosine 985 of leptin receptor (ObRb-pTyr985) and Janus kinase 2 (pJAK2). Transcriptome analysis of the CCs revealed a complex response to DIO, with large numbers and distinct sets of genes deregulated at early and late stages of obesity; in addition, there was a striking correlation between body weight and global transcriptome profile of CCs. Further analysis indicated that the transcriptome profile in 4 wk HFD CCs resembled that of LEPT CCs, in the upregulation of cellular trafficking and impairment in cytoskeleton organisation. Conversely, after 16 wk HFD CCs showed expression changes indicative of augmented inflammatory responses, cell morphogenesis, and decreased metabolism and transport, mainly as a consequence of the physiological changes of obesity.

Conclusion

Obesity leads to ovarian leptin resistance and major time-dependent changes in gene expression in CCs, which in early obesity may be caused by increased leptin signalling in the ovary, whereas in late obesity are likely to be a consequence of metabolic changes taking place in the obese mother.
ResQ, a release factor-dependent ribosome rescue factor in the Gram-positive bacterium Bacillus subtilis
Authors: Shimokawa-Chiba Naomi , Müller Claudia , Fujiwara Keigo , Beckert Bertrand , Ito Koreaki , Wilson Daniel N. , Chiba Shinobu
Published: August 2019

Rescue of the ribosomes from dead-end translation complexes, such as those on truncated (non-stop) mRNA, is essential for the cell. Whereas bacteria use trans-translation for ribosome rescue, some Gram-negative species possess alternative and release factor (RF)-dependent rescue factors, which enable an RF to catalyze stop codon-independent polypeptide release. We now discover that the Gram-positive Bacillus subtilis has an evolutionarily distinct ribosome rescue factor named ResQ. Genetic analysis shows that B. subtilis requires the function of either trans-translation or ResQ for growth, even in the absence of proteotoxic stresses. Biochemical and cryo-EM characterization demonstrates that ResQ binds to non-stop stalled ribosomes, recruits homologous RF2, but not RF1, and induces its transition into an open active conformation. Although ResQ is distinct from E. coli ArfA, they use convergent strategies in terms of mode of action and expression regulation, indicating that many bacteria may have evolved as yet unidentified ribosome rescue systems.
A Canal-Associated Neuron cAMP signalling pathway that regulatesC. eleganslarval development
Authors: Chien Jason , Wolf Fred W. , Grosche Sarah , Yosef Nebeyu , Garriga Gian , Mörck Catarina
Published: August 2019

Caenorhabditis eleganslarval development requires the function of the two Canal-Associated Neurons (CANs): killing the CANs by laser microsurgery or disrupting their development by mutating the geneceh-10results in early larval arrest. How these cells promote larval development, however, remains a mystery. In screens for mutations that bypass CAN function, we identified the genekin-29,which encodes a member of the Salt-Inducible Kinase (SIK) family and a component of a conserved pathway that regulates variousC. elegansphenotypes. Likekin-29loss, gain-of-function mutations in genes that may act upstream ofkin-29or growth in cyclic-AMP analogs bypassedceh-10larval arrest, suggesting that a conserved adenylyl cyclase/PKA pathway inhibits KIN-29 to promote larval development and that loss of CAN function results in dysregulation of KIN-29 and larval arrest. The adenylyl cyclase ACY-2 mediates CAN-dependent larval development:acy-2mutant larvae arrested development with a similar phenotype toceh-10mutants, and the arrest phenotype was suppressed by mutations inkin-29. ACY-2 is predominantly expressed in the CANs, and we provide evidence that theacy-2functions in the CANs to promote larval development. By contrast, cell-specific expression experiments suggest thatkin-29acts in both the hypodermis and neurons, but not in the CANs. Based on our findings, we propose that cAMP produced by ACY-2 in the CANs acts in neighboring neurons and hypodermal cells where it activates PKA and inhibits KIN-29 to promote larval development. We discuss how this conserved pathway could be partitioned between two cells.
Intercellular communication controls agonist-induced calcium oscillations independently of gap junctions in smooth muscle cells
Authors: Stasiak S.E. , Jamieson R.R. , Bouffard J. , Cram E.J. , Parameswaran H.
Published: August 2019

We report the existence of a unique mode of communication among human smooth muscle cells (SMCs) where they use force to frequency modulate long-range calcium waves. An important consequence of this mechanical signaling is that changes in stiffness of the underlying extracellular matrix can interfere with the frequency modulation of Ca2+ waves causing healthy SMCs to falsely perceive a much higher agonist dose than they actually received. This distorted sensing of contractile agonist dose on stiffer matrices is absent in isolated SMCs, even though the isolated cells can sense matrix rigidity. We show that intercellular communication that enables this collective Ca2+ response does not involve transport across gap junctions or extracellular diffusion of signaling molecules. The aberrant communication between cells that distorts the individual cell's perception of contractile stimulus can explain the sudden, exaggerated narrowing of the lumen when exposed to low dose of inhaled agonists in diseases like asthma.
Detection and quantification of GPCR mRNA: An assessment and implications of data from high-content methods
Authors: Sriram Krishna , Wiley Shu Z. , Moyung Kevin , Gorr Matthew W. , Salmerón Cristina , Marucut Jordin , French Randall P. , Lowy Andrew M. , Insel Paul A.
Published: August 2019

G protein-coupled receptors (GPCRs) are the largest family of membrane receptors and targets for approved drugs. Analysis of GPCR expression is thus important for drug discovery and typically involves mRNA-based methods. We compared transcriptomic cDNA [Affymetrix] microarrays, RNA-seq and qPCR-based TaqMan arrays for their ability to detect and quantify expression of endoGPCRs (non-chemosensory GPCRs with endogenous agonists). In human pancreatic cancer-associated fibroblasts, RNA-seq and TaqMan arrays yielded closely correlated values for GPCR number (~100) and expression levels, as validated by independent qPCR. By contrast, the microarrays failed to identify ~30 such GPCRs and generated data poorly correlated with results from those methods. RNA-seq and TaqMan arrays also yielded comparable results for GPCRs in human cardiac fibroblasts, pancreatic stellate cells, cancer cell lines and pulmonary arterial smooth muscle cells. The magnitude of mRNA expression for several Gq/11-coupled GPCRs predicted cytosolic calcium increase and cell migration by cognate agonists. RNA-seq also revealed splice variants for endoGPCRs. Thus, RNA-seq and qPCR-based arrays are better suited than microarrays for assessing GPCR expression and can yield results predictive of functional responses--findings that have implications for GPCR biology and drug discovery.

Abstract Graphic
BiofilmQ, a software tool for quantitative image analysis of microbial biofilm communities
Authors: Hartmann Raimo , Jeckel Hannah , Jelli Eric , Singh Praveen K. , Vaidya Sanika , Bayer Miriam , Vidakovic Lucia , Díaz-Pascual Francisco , Fong Jiunn C.N. , Dragoš Anna , Besharova Olga , Nadell Carey D. , Sourjik Victor , Kovács Ákos T. , Yildiz Fitnat H. , Drescher Knut
Published: August 2019

Biofilms are now considered to be the most abundant form of microbial life on Earth, playing critical roles in biogeochemical cycles, agriculture, and health care. Phenotypic and genotypic variations in biofilms generally occur in three-dimensional space and time, and biofilms are therefore often investigated using microscopy. However, the quantitative analysis of microscopy images presents a key obstacle in phenotyping biofilm communities and single-cell heterogeneity inside biofilms. Here, we present BiofilmQ, a comprehensive image cytometry software tool for the automated high-throughput quantification and visualization of 3D and 2D community properties in space and time. Using BiofilmQ does not require prior knowledge of programming or image processing and provides a user-friendly graphical user interface, resulting in editable publication-quality figures. BiofilmQ is designed for handling fluorescence images of any spatially structured microbial community and growth geometry, including microscopic, mesoscopic, macroscopic colonies and aggregates, as well as bacterial biofilms in the context of eukaryotic hosts.
Probe-Seq enables transcriptional profiling of specific cell types from heterogeneous tissue by RNA-based isolation
Authors: Amamoto Ryoji , Garcia Mauricio D. , West Emma R. , Choi Jiho , Lapan Sylvain W. , Lane Elizabeth A. , Perrimon Norbert , Cepko Constance L.
Published: August 2019

Recent transcriptional profiling technologies are uncovering previously-undefined cell populations and molecular markers at an unprecedented pace. While single cell RNA (scRNA) sequencing is an attractive approach for unbiased transcriptional profiling of all cell types, a complementary method to isolate and sequence specific cell populations from heterogeneous tissue remains challenging. Here, we developed Probe-Seq, which allows deep transcriptional profiling of specific cell types isolated using RNA as the defining feature. Dissociated cells are labelled using fluorescent in situ hybridization (FISH) for RNA, and then isolated by fluorescent activated cell sorting (FACS). We used Probe-Seq to purify and profile specific cell types from mouse, human, and chick retinas, as well as the Drosophila midgut. Probe-Seq is compatible with frozen nuclei, making cell types within archival tissue immediately accessible. As it can be multiplexed, combinations of markers can be used to create specificity. Multiplexing also allows for the isolation of multiple cell types from one cell preparation. Probe-Seq should enable RNA profiling of specific cell types from any organism.
Ctf4 organizes sister replisomes and Pol α into a replication factory
Authors: Yuan Zuanning , Georgescu Roxana , de Luna Almeida Santos Ruda , Zhang Daniel , Bai Lin , Yao Nina Y. , Zhao Gongpu , O’Donnell Michael E. , Li Huilin
Published: August 2019

The current view is that eukaryotic replisomes are independent. Here we show that Ctf4 tightly dimerizes CMG helicase, with an extensive interface involving Psf2, Cdc45, and Sld5. Interestingly, Ctf4 binds only one Pol α-primase. Thus, Ctf4 may have evolved as a trimer to organize two helicases and one Pol α-primase into a replication factory. In the 2CMG-Ctf43-1Pol α-primase factory model, the two CMGs nearly face each other, placing the two lagging strands toward the center and two leading strands out the sides. The single Pol α-primase is centrally located and may prime both sister replisomes. The Ctf4-coupled-sister replisome model is consistent with cellular microscopy studies revealing two sister forks of an origin remain attached and are pushed forward from a protein platform. The replication factory model may facilitate parental nucleosome transfer during replication.
Simulating metagenomic stable isotope probing datasets with MetaSIPSim
Authors: Barnett Samuel E. , Buckley Daniel H.
Published: August 2019

Background

DNA-stable isotope probing (DNA-SIP) links microorganisms to their in-situ function in diverse environmental samples. Combining DNA-SIP and metagenomics (metagenomic-SIP) allows us to link genomes from complex communities to their specific functions and improves the assembly and binning of these targeted genomes. However, empirical development of metagenomic-SIP methods is hindered by the complexity and cost of these studies. We developed a toolkit, ‘MetaSIPSim,’ to simulate sequencing read libraries for metagenomic-SIP experiments. MetaSIPSim is intended to generate datasets for method development and testing. To this end, we used MetaSIPSim generated data to demonstrate the advantages of metagenomic-SIP over a conventional shotgun metagenomic sequencing experiment.

Results

Through simulation we show that metagenomic-SIP improves the assembly and binning of isotopically labeled genomes relative to a conventional metagenomic approach. Improvements were dependent on experimental parameters and on sequencing depth. Community level G+C content impacted the assembly of labeled genomes and subsequent binning, where high community G+C generally reduced the benefits of metagenomic-SIP. Furthermore, when a high proportion of the community is isotopically labeled, the benefits of metagenomic-SIP decline. Finally, the choice of gradient fractions to sequence greatly influences method performance.

Conclusions

Metagenomic-SIP is a valuable method for recovering isotopically labeled genomes from complex communities. We show that metagenomic-SIP performance depends on optimization of experimental parameters. MetaSIPSim allows for simulation of metagenomic-SIP datasets which facilitates the optimization and development of metagenomic-SIP experiments and analytical approaches for dealing with these data.
KAT2-mediated acetylation switches the mode of PALB2 chromatin association to safeguard genome integrity
Authors: Fournier Marjorie , Bleuyard Jean-Yves , Couturier Anthony M. , Ellins Jessica , Hester Svenja , Smerdon Stephen J. , Tora László , Esashi Fumiko
Published: August 2019

The tumour suppressor PALB2 stimulates error-free repair of DNA breaks, whilst its steady-state chromatin association protects active genes from genotoxic stress. Here, we report that the lysine acetyltransferases 2A and 2B (KAT2A/B), commonly known to promote transcriptional activation, acetylate the PALB2 chromatin association motif (ChAM), providing a dynamic regulatory mechanism for PALB2. ChAM acetylation within a cluster of seven lysine residues (7K), detected in the chromatin-enriched fraction in undamaged cells, enhanced its association with nucleosomes while decreasing its non-specific binding to naked DNA. DNA damage triggered a rapid deacetylation of ChAM and a concomitant increase in PALB2 mobility. Significantly, a 7K-null mutation, which hindered ChAM binding to both nucleosomes and DNA, conferred deficiency in DNA repair and hypersensitivity to the anti-cancer drug olaparib. Thus, our study reveals a unique mechanism mediated by KAT2A/B-dependent acetylation of a non-histone protein, which fine-tunes the DNA damage response and hence promotes genome stability.
Modulation of sensory behavior and food choice by an enteric bacteria-produced neurotransmitter
Authors: O’Donnell Michael P. , Fox Bennett W. , Chao Pin-Hao , Schroeder Frank C. , Sengupta Piali
Published: August 2019

Animals coexist in commensal, pathogenic or mutualistic relationships with complex communities of diverse organisms including microbes1. Some bacteria produce bioactive neurotransmitters which have been proposed to modulate host nervous system activity and behaviors2. However, the mechanistic basis of this microbiota-brain modulation and its physiological relevance is largely unknown. Here we show that inC. elegans, the neuromodulator tyramine (TA) produced by gut-colonizing commensalProvidenciabacteria can bypass the requirement for host TA biosynthesis to manipulate a host sensory decision. Bacterially-produced TA is likely converted to octopamine (OA) by the host tyramine beta-hydroxylase enzyme. OA, in turn, targets the OCTR-1 receptor on the ASH/ASI sensory neurons to modulate an aversive olfactory response. We identify genes required for TA biosynthesis inProvidencia, and show that these genes are necessary for modulation of host behavior. We further find thatC. eleganscolonized byProvidenciapreferentially select these bacteria in food choice assays, and that this selection bias requires bacterially-produced TA. Our results demonstrate that a neurotransmitter produced by gut microbiota mimics the functions of the cognate host molecule to override host control of a sensory decision, thereby promoting fitness of both host and microbe.
Live-cell assays for cell stress responses reveal new patterns of cell signaling caused by mutations in rhodopsin, α-synuclein and TDP-43
Authors: Harlen Kevin M. , Roush Elizabeth C. , Clayton Joseph E. , Martinka Scott , Hughes Thomas E.
Published: August 2019

Many neurodegenerative diseases induce high levels of sustained cellular stress and alter a number of cellular processes. Genetically-encoded fluorescent biosensors are effective tools to examine neuronal activity and signaling in living cells. To examine how different mutations associated with neurodegenerative disease affect cell stress and signaling we created live-cell assays for ER-mediated cell stress and second messenger signaling. Analysis of the rhodopsin P23H mutation, the most common mutation in autosomal dominant Retinitis Pigmentosa, revealed increased cell stress levels compared to wild type rhodopsin. Moreover, this increase in cell stress correlated with blunted Ca2+signaling in a stress dependent manner. Analysis of single cell Ca2+signaling profiles revealed unique Ca2+signaling responses exist in cells expressing wild type or P23H mutants, further supporting the notion that second messenger signaling is affected by cell stress. To explore the use of the ER-stress biosensor in other neurodegenerative diseases we examined how various mutants of α-synuclein and TDP-43 affected ER-mediated cell stress. Mutants of both α-synuclein and TDP-43 associated with Parkinson’s Disease and ALS demonstrated increases in ER-mediated cell stress. This increased cell stress was accompanied by changes in phosphodiesterase activity. Both HEK293T and SH-SY5Y cells expressing these proteins displayed a shift towards increased cAMP degradation rates, likely due to increased phosphodiesterase activity. Together these data illustrate how biosensors can provide nuanced, new views of neurodegenerative disease processes.
Untangling the effects of cellular composition on coexpression analysis
Authors: Farahbod Marjan , Pavlidis Paul
Published: August 2019

Background

Coexpression analysis is one of the most widely used methods in genomics, with applications to inferring regulatory networks, predicting gene function, and interpretation of transcriptome profiling studies. Most studies use data collected from bulk tissue, where the effects of cellular composition present a potential confound. However, the impact of composition on coexpression analysis have not been studied in detail. Here we examine this issue for the case of human brain RNA analysis.

Results

We found that for most genes, differences in expression levels across cell types account for a large fraction of the variance of their measured RNA levels in brain (median R2= 0.64). We then show that genes that have similar expression patterns across cell types will have correlated RNA levels in bulk tissue, due to the effect of variation in cellular composition. We demonstrate that much of the coexpression in the bulk tissue can be attributed to this effect. We further show how this composition-induced coexpression masks underlying intra-cell-type coexpression observed in single-cell data. Attempt to correct for composition yielded mixed results.

Conclusions

The dominant coexpression signal in brain can be attributed to cellular compositional effects, rather than intra-cell-type regulatory relationships, and this is likely to be true for other tissues. These results have important implications for the relevance and interpretation of coexpression in many applications.