bluuebunny/crossref_metadata_2025 huggingface dataset:
The FLIP-FIGNL1 complex regulates the dissociation of RAD51/DMC1 in homologous recombination and replication fork restart
Authors: Zhang Qianting , Fan Jiayi , Xu Wei , Cao Huiwen , Qiu Cheng , Xiong Yi , Zhao Huacun , Wang Yong , Huang Jun , Yu Chao
Published: July 2023

Recruitment of RAD51 and/or DMC1 recombinases to single-strand DNA is indispensable for homology search and strand invasion in homologous recombination (HR) and for protection of nascent DNA strands at stalled replication forks. Thereafter RAD51/DMC1 dissociate, actively or passively, from these joint molecules upon DNA repair or releasing from replication stress. However, the mechanism that regulates RAD51/DMC1 dissociation and its physiological importance remain elusive. Here, we show that a FLIP-FIGNL1 complex regulates RAD51 and DMC1 dissociation to promote meiotic recombination and replication fork restart in mammals. Mice lacking FLIP are embryonic lethal, while germline-specific deletion of FLIP leads to infertility in both males and females. FLIP-null meiocytes are arrested at a zygotene-like stage with massive RAD51 and DMC1 foci, which frequently co-localize with SHOC1 and TEX11. Furthermore, FLIP interacts with FIGNL1. Depletion of FLIP or FIGNL1 in cell lines destabilizes each other and impairs RAD51 dissociation. Thus, the active dissociation of RAD51/DMC1 by the FLIP-FIGNL1 complex is a crucial step required for HR and replication fork restart, and represents a conserved mechanism in somatic cells and germ cells.
Toward the confirmation of atmospheric neutrino oscillations
Authors: Oyama Yuichi
Published: July 2023

The atmospheric muon neutrino deficit, which was possible evidence of νμ↔ντ oscillation, was reported by the Kamiokande experiment from 1988. Many experimental efforts were made to examine the Kamiokande results. Experiments which contributed to the confirmation of νμ↔ντ oscillation are reviewed. Especially, long-baseline neutrino-oscillation experiments are described in detail.
A call for obesity prevention interventions for young children with intellectual and developmental disabilities
Authors: Schenkelberg Michaela A , Clarke Emily C , Wasser Heather , Ward Dianne S , Essenmacher Mikayla M , Thompson Kelsey L , Willis Erik A
Published: July 2023

Lay Summary

Health disparities among children with intellectual and developmental disabilities (IDD) are present in early childhood. Yet, this population is underrepresented in health behavior research. In this commentary the authors highlight the need for multi-level physical activity and nutrition research for obesity prevention with a specific focus on young children with Down syndrome, a population at greater risk of developing overweight and obesity compared to typically developing peers. This commentary describes the comorbidities and developmental challenges faced by many children with Down syndrome which may influence weight-related physical activity and nutrition behaviors. Additionally, the authors advocate for involving a multidisciplinary team of experts to inform the adaptation or development of multi-level, theory-driven behavioral interventions to prevent obesity among children with Down syndrome.
Longitudinal anellome dynamics in the upper respiratory tract of children with acute respiratory tract infections
Authors: Cao Le , Ma Yingying , Wan Zhenzhou , Li Bing , Tian Weimin , Zhang Chiyu , Li Yanpeng
Published: July 2023

Anelloviruses (AVs) are ubiquitous in humans and are the most abundant components of the commensal virome. Previous studies on the diversity, transmission, and persistence of AVs mainly focused on the blood or transplanted tissues from adults; however, the profile of the anellome in the respiratory tract in children are barely known. We investigated the anellome profile and their dynamics in the upper respiratory tract from a cohort of children with acute respiratory tract infections (ARTIs). Different to that in adult, betatorquevirus is the most abundant genus, followed by alphatorquevirus. We found that the relative abundance of betatorquevirus was higher in earlier time points, and in contrast, the abundance of alphatorquevirus was higher in later time points; these results might suggest that betatorquevirus decreased with age and alphatorquevirus increased with age in childhood. No difference regarding the diversity and abundance of anellome was found between single and multiple ARTIs, consistent with the idea that AV is not associated with certain disease. Most AVs are transient, and a small proportion (8 per cent) of them were found to be possibly persistent, with persistence time ranging from 1 month to as long as 56 months. Furthermore, the individual respiratory anellome appeared to be unique and dynamic, and the replacement of existing AVs with new ones are common over different time points. These findings demonstrate that betatorquevirus may be the early colonizer in children, and the individual respiratory anellome is unique, which are featured by both chronic infections and AV community replacement.
Interpretation of the Patient Health Questionnaire 9 in High-impact Chronic Pain
Authors: Aagaard Andrea , Ravn Sophie Lykkegaard , Andersen Tonny Elmose , Vaegter Henrik Bjarke
Published: July 2023

Objectives:

Depression is prevalent among patients with chronic pain and may impact pain management. Accurate assessment is, however, complicated by overlapping symptoms. This study investigated how patients with high-impact chronic pain interpreted and responded to the Patient Health Questionnaire 9 (PHQ-9) to identify problematic items and causes hereof.

Methods:

Cognitive interviews using the Three-Step Test-Interview procedure were conducted during completion of the PHQ-9 in 33 patients with high-impact chronic pain referred to interdisciplinary treatment. Responses were analyzed using four coding categories: i) “congruent” (response consistent with intention); ii) “incongruent” (response not consistent intention); iii) “ambiguous” (response both congruent and incongruent or insufficient to evaluate congruency); and iv) “confused” (response with confused or misunderstood statements). Next, content of responses problematic items was analyzed to identify causes for non-congruency and encountered response difficulties were identified across all items.

Results:

Three items (item 2, 6, and 9) performed as intended (>97% congruent responses), while seven items (item 1, 3, 4, 5, 7, 8, and 10) were identified as problematic (<50% congruent responses). Problematic items had one or more issues: Responses were based on i) pain related issues or ii) other (non-pain) factors unrelated to depression, or item structure caused response difficulties due to wordings, reversion, or having two questions in one.

Discussion:

Problematic items limit the construct validity of the PHQ-9, leaving increased risk of inflated depression scores in high-impact chronic pain. Identified problems should guide future revisions to enhance validity and screening accuracy for the benefit of both research and clinical practice.
Coronavirus Disease 2019 (COVID-19) Vaccination and Assisted Reproduction Outcomes
Authors: Chamani Isaac J. , Taylor Lauren L. , Dadoun Simon E. , McKenzie Laurie J. , Detti Laura , Ouellette Lara , McCulloh David H. , Licciardi Frederick L.
Published: July 2023

OBJECTIVE:

To assess the association between coronavirus disease 2019 (COVID-19) vaccination and female assisted reproduction outcomes through a systematic review and meta-analysis.

DATA SOURCES:

We searched Medline (OVID), EMBASE, Web of Science, Cochrane Library, and ClinicalTrials.gov on January 11, 2023, for original articles on assisted reproduction outcomes after COVID-19 vaccination. The primary outcome was rates of clinical pregnancy; secondary outcomes included number of oocytes retrieved, number of mature oocytes retrieved, fertilization rate, implantation rate, ongoing pregnancy rate, and live-birth rate.

METHODS OF STUDY SELECTION:

Two reviewers independently screened citations for relevance, extracted pertinent data, and rated study quality. Only peer-reviewed published studies were included.

TABULATION, INTEGRATION, AND RESULTS:

Our query retrieved 216 citations, of which 25 were studies with original, relevant data. Nineteen studies reported embryo transfer outcomes, with a total of 4,899 vaccinated and 13,491 unvaccinated patients. Eighteen studies reported data on ovarian stimulation outcomes, with a total of 1,878 vaccinated and 3,174 unvaccinated patients. There were no statistically significant results among our pooled data for any of the primary or secondary outcomes: clinical pregnancy rate (odds ratio [OR] 0.94, 95% CI 0.88–1.01, P=.10), number of oocytes retrieved (mean difference −0.26, 95% CI −0.68 to 0.15, P=.21), number of mature oocytes retrieved (mean difference 0.31, 95% CI −0.14 to 0.75, P=.18), fertilization rate (OR 0.99, 95% CI 0.87–1.11, P=.83), implantation rate (OR 0.92, 95% CI 0.84–1.00, P=.06), ongoing pregnancy rate (OR 0.95, 95% CI 0.86–1.06, P=.40), or live-birth rate (OR 0.95, 95% CI 0.78–1.17, P=.63). A subanalysis based on country of origin and vaccine type was also performed for the primary and secondary outcomes and did not change the study results.

CONCLUSION:

Vaccination against COVID-19 is not associated with different fertility outcomes in patients undergoing assisted reproductive technologies.

SYSTEMATIC REVIEW REGISTRATION:

PROSPERO, CRD42023400023.
Evaluation of the Recommended 30 cc/kg Fluid Dose for Patients With Septic Shock and Hypoperfusion With Lactate Greater Than 4 mmol/L
Authors: Yohannes Seife , Serafim Laura Piccolo , Slavinsky Victoria , O’Connor Timothy , Cabrera Mathew , Chin Meghan K. , Pratt Alexandra
Published: July 2023

OBJECTIVES:

The Surviving Sepsis Campaign Guidelines recommend fluid administration of 30 cc/kg ideal body weight (IBW) for patients with sepsis and lactate greater than 4 mmol/L within 3 hours of identification. In this study, we explore the impact of fluid dose on lactate normalization, treatment cost, length of stay, and mortality in patients with lactate greater than 4.

DESIGN:

Multicenter retrospective observational study.

SETTING:

Eight-hospital urban healthcare system in Northeastern United States.

PATIENTS:

Patients with sepsis, initial lactate value greater than 4 mmol/L, and received appropriate antibiotics within 3 hours.

INTERVENTIONS:

None.

MEASUREMENTS AND MAIN RESULTS:

We stratified patients into five groups based on the dose of fluid administered within 3 hours after sepsis identification. The groupings were less than 15 cc/kg IBW, 15.1–25 cc/kg IBW, 25.1–35 cc/kg IBW, 35.1–50 cc/kg IBW, and greater than 50 cc/kg IBW. We used the group that received a fluid dose of 25.1–35 cc/kg IBW, as a reference group. The mean age was 66 years, and 56% were male. Three hundred seventy-one (25%) received less than 15 cc/kg of IBW of crystalloid fluid, 278 (17%) received 15–25 cc/kg of IBW, 316 (21%) received 25.1–35 cc/kg of IBW, 319 (21%) received 35.1–50 cc/kg of IBW, and 207 (14%) received greater than 50 cc/kg of IBW. After multilinear regression, there was no significant difference in lactate normalization between the reference group and any of the other fluid groups. We also found no statistically significant difference in the observed/expected cost, or observed/expected length of stay, between the reference group and any of the other fluid groups. Mortality was higher among patients who received greater than 50 cc/kg IBW when compared to the recommended dose.

CONCLUSIONS:

In patients with sepsis and lactate value greater than 4 mmol/L, high or low fluid doses were not associated with better lactate clearance or patient outcomes. Greater than 50 cc/kg IBW dose of fluids within 3 hours is associated with higher mortality.
Developing a Machine Learning Risk-adjustment Method for Hospitalizations and Emergency Department Visits of Nursing Home Residents With Dementia
Authors: Xu Huiwen , Bowblis John R. , Becerra Adan Z. , Intrator Orna
Published: July 2023

Background:

Long-stay nursing home (NH) residents with Alzheimer disease and related dementias (ADRD) are at high risk of hospital transfers. Machine learning might improve risk-adjustment methods for NHs.

Objectives:

The objective of this study was to develop and compare NH risk-adjusted rates of hospitalizations and emergency department (ED) visits among long-stay residents with ADRD using Extreme Gradient Boosting (XGBoost) and logistic regression.

Research Design:

Secondary analysis of national Medicare claims and NH assessment data in 2012 Q3. Data were equally split into the training and test sets. Both XGBoost and logistic regression predicted any hospitalization and ED visit using 58 predictors. NH-level risk-adjusted rates from XGBoost and logistic regression were constructed and compared. Multivariate regressions examined NH and market factors associated with rates of hospitalization and ED visits.

Subjects:

Long-stay Medicare residents with ADRD (N=413,557) from 14,057 NHs.

Results:

A total of 8.1% and 8.9% residents experienced any hospitalization and ED visit in a quarter, respectively. XGBoost slightly outperformed logistic regression in area under the curve (0.88 vs. 0.86 for hospitalization; 0.85 vs. 0.83 for ED visit). NH-level risk-adjusted rates from XGBoost were slightly lower than logistic regression (hospitalization=8.3% and 8.4%; ED=8.9% and 9.0%, respectively), but were highly correlated. Facility and market factors associated with the XGBoost and logistic regression-adjusted hospitalization and ED rates were similar. NHs serving more residents with ADRD and having a higher registered nurse-to-total nursing staff ratio had lower rates.

Conclusions:

XGBoost and logistic regression provide comparable estimates of risk-adjusted hospitalization and ED rates.
Optimal Duration of High-Fidelity Simulator Training for Bronchoscope-Guided Intubation
Authors: Jiang Luyang , Yang Qingmei , Li Qingyue , Jiang Bailin , Laba Ciren , Feng Yi
Published: July 2023

Introduction

The optimal simulator training duration for flexible optical bronchoscopic (FOB) intubation is unknown. This study aimed to determine whether a learning curve-based training modality was noninferior to a fixed training time modality in terms of clinical FOB intubation time.

Methods

This multicenter, randomized, noninferiority study was conducted from May to August 2022. Anesthesiology residents or interns were enrolled. Eligible participants were randomized in a 1:1 ratio to receive new learning curve-based simulator training (individualized training time based on performance, group New) or reference fixed training time simulator training (1 hour, group Reference). The primary outcome was the time to complete FOB intubation in patients, which was defined as the time from the introduction of the FOB into the mouth until the first capnography visualization. The margin for detecting clinical significance was defined as 10 seconds.

Results

A total of 32 participants were included in the analysis (16 in each group). All trainees successfully intubated the patients. The mean intubation time (95% confidence interval [CI]) was 81.9 (65.7–98.1) seconds in group New and 97.0 (77.4–116.6) seconds in group Reference. The upper bound of the 1-sided 97.5% CI for the mean difference of clinical intubation time between groups was 9.3 seconds. Noninferiority was claimed. The mean duration of the training in group New was 28.4 (95% CI, 23.5–33.4) minutes. The total number of training procedures on simulators in group New was significantly less than that in group Reference (P < 0.01).

Conclusions

The clinical FOB intubation time in group New was noninferior to that in group Reference.
Epidemiology of Autism Spectrum Disorder in Buenos Aires, Argentina: Insights from Unique Disability ID Issuance Trends
Authors: Aragón-Daud Agustina , Vetere Giselle , Argañaraz Marina , Musich Francisco
Published: July 2023

Purpose

Autism Spectrum Disorder (ASD) prevalence rates vary greatly across regions and studies. Some countries, such as Argentina, have unknown rates. Due to this high heterogeneity in ASD prevalence worldwide, it is important to study its prevalence and characteristics in such countries to develop effective policies.

Methods

we analyzed official data from the Unique Disability IDs (UDIDs) issued for individuals with ASD in the Autonomous City of Buenos Aires from 2016 to 2021, examining age, gender, and diagnosis.

Results

most UDIDs were issued for males, for Pervasive Developmental Disorders, and almost half to individuals over 8 years old, with these trends remaining stable over these years. However, UDID issuance abruptly dropped in 2020.

Conclusions

our findings highlight epidemiological aspects of the autism population in Argentina, including challenges such as delayed diagnosis and high prevalence of broad diagnosis categories. Addressing these challenges requires further research and intervention to improve the quality of life of individuals with ASD.
Long-read Transcriptome Landscapes of Primary and Metastatic Liver Cancers at Transcript Resolution
Authors: Chen Zhiao , Shi Qili , Zhao Yiming , Xu Midie , Liu Yizhe , Li Xinrong , Liu Li , Sun Menghong , Wu Xiaohua , Shao Zhimin , Xu Ye , Wang Lu , He Xianghuo
Published: July 2023

Background

The liver is the sixth most common site of primary cancer in humans and is frequently colonized by metastases from cancers of other organs. Few studies have investigated the transcriptomic profiles of matched primary tumor and hepatic metastases of patients. Moreover, the read length of 100-200 bases in conventional short-read RNA sequencing is too short, which makes it difficult to directly infer the full-length transcript structure. To help develop effective treatments and improve survival, it is crucial to understand the complex and diverse molecular mechanisms of primary and metastatic liver cancers.

Methods

Ninety-five primary and secondary liver cancer patients who underwent hepatic resection were included with long-read sequencing isoform-sequencing and short-read RNA sequencing. We compared the transcriptome landscapes of primary and metastatic liver cancers and systematically investigated HCC, paired primary tumors and liver metastases, and matched non-tumor liver tissues.

Results

We defined the full-length isoform-level transcriptome of human primary and metastatic liver cancers and identified isoform-level diversity in HCC and metastasis-associated transcriptome variations in metastatic liver cancers. Specific RNA transcripts and isoform switching events with clinical implications were profoundly discovered in liver cancer. Metastasis-specific transcripts that can predict the metastatic risk and identify the primary sites of cancers of unknown primary liver metastasis patients were defined. Additionally, we found that adjacent paracancerous liver tissues are abnormal and characterized the premetastatic immunological and metabolic alterations in the liver that favor the spread of cancer metastases.

Conclusions

Our findings strongly highlight the powerfulness of full-length transcriptome profiling to yield novel biological insights into understanding the molecular basis of tumorigenesis and will further benefit the treatment of primary and metastatic liver cancers.
The Auxin and Cytokinin Balance Influence the in Vitro Regeneration ofPhalaenopsisShoots (Orchidaceae)
Authors: Cruz Kaliane Zaira Camacho Maximiano , Alencar Antonio André Silva , Santana Josefa Grasiela Silva , Oliveira Alves Laura Eliza
Published: July 2023

Orchids (Phalaenopsis) are considerably appreciated or their high and durable flowering rate and exotic appearance. The optimization of in vitro cultivation begins with an understanding of hormonal balance and its implications in production. This work aimed to regenerate shoots from seeds germination of thePhalaenopsisGolden Peoker ‘BL’ HCC / AOS’ under exogenous auxin and cytokinin influence. To do so, naphthalene acetic acid (NAA), 6-benzyladenine (BA), and Kinetin (Kin) were added to the culture medium to induce organogenesis, totaling 17 treatments with eight replications per treatment. The variables number of shoots (NS), shoot length (SL), and presence of root primordia (RP) were evaluated. A non-parametric test (the Friedman test), followed by the Bonferroni procedure, was used to compare the different groups. Hence, the MS medium provided 610 complete plants after 90 days of cultivation. The T6 treatment had the highest median NS (1.5 shoots per plant at 15 days) and SL. At 30 days, the highest NS was for the T16, with 6.15 shoots per plant; for SL, the one that stood out was the T9. Regarding RP in shoots, the best treatments were T2, T3, T9, T10, and T17, with the formation of RP at 15 days, remaining these treatments at 30 days. In conclusion, the T9 is the most suitable for an efficient protocol reproduction, as it showed the best plant development.

HIGHLIGHTS (MANDATORY)

The auxin and cytokinin balance promoted shoot regeneration.

The BA and NAA combined promoted the shoot formation ofPhalaenopsisGolden Peoker ‘BL’ HCC/AOS’.

The use of KIN isolated showed hyperhydricity and malformation in shoots ofPhalaenopsis.
The unique properties of Big tau in the visual system
Authors: Fischer Itzhak , Connors Theresa , Bouyer Julien , Jin Ying
Published: July 2023

Tau is a microtubule associated protein that plays important roles in regulating the properties of microtubules and axonal transport, as well as tauopathies associated with toxic aggregates leading to neurodegenerative diseases. It is encoded by the MAPT gene forming multiple isoforms by alternative splicing of exons 2/3 at the N-terminal and exon 10 which determines the numbers of microtubule binding repeats (3R or 4R). The high molecular weight (MW) tau isoform termed Big tau contains an additional large exon 4a generating a long projecting domain and expressed as a 110 kDa protein. Big tau was originally discovered in the peripheral nervous system but later found in selective CNS areas that project to the periphery as well as in the cerebellum and the visual system. However, there is a gap of knowledge in understanding the expression patterns and the role of Big tau during normal neuronal development and pathological conditions relative to the common low MW isoforms. Here we investigated the properties of Big tau in the retina and optic nerve and in particular its unique isoform structure as a middle MW of 90kDa and its distribution in retinal ganglion cells and axons of the optic nerve. We discovered that Big tau expresses the 4a exon as well as exons 6 and 10 (4R), lacking exons 2/3 but sharing the extensive phosphorylation characteristic of other tau isoforms. Importantly, early in development the visual system expresses only the low MW isoform (3R) switching to both the low and middle MW isoforms (4R) in adult retinal ganglion neurons and their corresponding axons. This is a unique structure and expression pattern of Big tau likely associated with different properties than what has been previously described, requiring more research to elucidate the detailed roles of Big tau in the visual system.
Distinguishing word identity and sequence context in DNA language models
Authors: Sanabria Melissa , Hirsch Jonas , Poetsch Anna R.
Published: July 2023

Transformer-based large language models (LLMs) are very suited for biological sequence data, because the structure of protein and nucleic acid sequences show many analogies to natural language. Complex relationships in biological sequence can be learned, although there may not be a clear concept of words, because they can be generated through tokenization. Training is subsequently performed for masked token prediction. With this strategy, the models learn both the token sequence identity and a larger sequence context. We developed a framework to interrogate what a model learns, which is both relevant for the interpretability of the model and to evaluate its potential for specific tasks.

We used a DNA language model, which is trained on the human reference genome with a Bidirectional Encoder Representations from Transformers (BERT) model. In this model, tokens are defined with overlapping k-mers. To gain insight into the model’s learning, we interrogated how the model performs predictions, extracted token embeddings, and defined a fine-tuning benchmarking task to predict the next tokens of different sizes without overlaps. This task is very suited to evaluate different pretrained DNA language models, also called foundation models, since it does not interrogate specific genome biology, does not depend on the tokenization strategy, the size of the vocabulary, the dictionary, or the number of parameters used to train the model. Lastly, the task performs without leakage of information from token identity into the prediction task, which makes it particularly useful to evaluate the learning of sequence context.

Through this assessment we discovered that the model with overlapping k-mers struggles to learn larger sequence context. Instead, the learned embeddings largely represent token sequence. Still, good performance is achieved for genome biology inspired fine-tuning tasks. Models with overlapping tokens may be used for tasks where a larger sequence context is of less relevance, but the token sequence directly represents the desired learning feature. This emphasizes the need to interrogate knowledge representation in biological large language models. Transparency is particularly important for biomedical use cases and an understanding of what the models are learning can be used to match the model to the desired task.
Faster action reprogramming, but not stopping, with proactive cues: Combining EMG and computational modelling in response-selective stop signal tasks
Authors: Salomoni Sauro E. , Gronau Quentin F. , Heathcote Andrew , Matzke Dora , Hinder Mark R.
Published: July 2023

The ability to stop simple ongoing actions has been extensively studied using the stop signal task, but less is known about inhibition in more complex scenarios. Here we used a task requiring bimanual responses to go stimuli, but selective inhibition of only one of those responses following a stop signal. We assessed how proactive cues affect the nature of both the responding and stopping processes, and the well-documented “stopping delay” in the continuing action following successful stopping. In this task, estimates of the speed of inhibition based on a simple-stopping model are inappropriate, and have produced inconsistent findings about the effects of proactive control on motor inhibition. We instead used a multi-modal approach, based on improved methods of detecting and interpreting partial electromyographical (EMG) responses and the recently proposed SIS (simultaneously inhibit and start) model of selective stopping behaviour. Our results provide clear and converging evidence that proactive cues reduce the stopping delay effect by slowing bimanual responses and speeding unimanual responses, with a negligible effect on the speed of the stopping process.
Increased collective migration correlates with germline stem cell competition in a basal chordate
Authors: Fentress Megan K. , De Tomaso Anthony W.
Published: July 2023

Cell competition is a process that compares the relative fitness of progenitor cells, resulting in ‘winners’, which contribute further to development, and ‘losers’, which are excluded, and is likely a universal quality control process that contributes to the fitness of an individual. Cell competition also has pathological consequences, and can create super-competitor cells responsible for tumor progression. We are studying cell competition during germline regeneration in the colonial ascidian,Botryllus schlosseri. Germline regeneration is due to the presence of germline stem cells (GSCs) which have a unique property: a competitive phenotype. When GSCs from one individual are transplanted into another, the donor and recipient cells compete for germline development. Often the donor GSCs win, and completely replace the gametes of the recipient-a process called germ cell parasitism (gcp). gcp is a heritable trait, and winner and loser genotypes can be found in nature and reared in the lab. However, the molecular and cellular mechanisms underlying gcp are unknown. Using an ex vivo migration assay, we show that GSCs isolated from winner genotypes migrate faster and in larger clusters than losers, and that cluster size correlates with expression of the Notch ligand, Jagged. Both cluster size and jagged expression can be manipulated simultaneously in a genotype dependent manner: treatment of loser GSCs with hepatocyte growth factor increases both jagged expression and cluster size, while inhibitors of the MAPK pathway decrease jagged expression and cluster size in winner GSCs. Live imaging in individuals transplanted with labeled winner and loser GSCs reveal that they migrate to the niche, some as small clusters, with the winners having a slight advantage in niche occupancy. Together, this suggests that the basis of GSC competition resides in a combination in homing ability and niche occupancy, and may be controlled by differential utilization of the Notch pathway.
Sensitivity and validation of porous membrane electrical cell substrate impedance spectroscopy (PM-ECIS) for measuring endothelial barrier properties
Authors: Ugodnikov Alisa , Chebotarev Oleg , Persson Henrik , Simmons Craig A.
Published: July 2023

Conventional trans-endothelial electrical resistance (TEER) setups are invasive and cannot directly measure monolayer integrity in co-culture. These limitations are addressed by porous membrane electrical cell-substrate impedance sensing (PM-ECIS), which measures barrier integrity in cell monolayers grown directly on permeable membranes patterned with electrodes. Here we advance the design and utility of PM-ECIS by investigating its sensitivity to working electrode size and correlation to TEER. Gold electrodes were fabricated on porous membrane inserts using hot embossing and UV lithography, with working electrode diameters of 250, 500, and 750 µm within the same insert. Frequency scans of confluent primary endothelial monolayers showed normalized resistance peaking at 4 kHz, consistent with traditional solid substrate ECIS. Sensitivity to resistance changes (4 kHz) during endothelial barrier formation was inversely proportional to electrode size, with the smallest electrodes being the most sensitive (p<0.001). Similarly, smaller electrodes were most sensitive to changes in impedance (40 kHz) corresponding to cell spreading and proliferation (p<0.001). Barrier disruption with EGTA was detectable by 250 and 750 µm (p<0.01), and 500 µm electrodes (p=0.058). Resistances measured by PM-ECIS vs. TEER for sodium chloride solutions were positively and significantly correlated for all electrode sizes (r>0.9; p<0.0001), but only with 750 µm electrodes for endothelial monolayers (r=0.71; p=0.058). These data inform design and selection of PM-ECIS electrodes for specific applications, and support PM-ECIS as a promising alternative to conventional TEER due to its capacity for direct, non-invasive, and real-time assessment of cells cultured on porous membranes.
Dynamic monitoring of viral gene expression reveals rapid antiviral effects of CD8 T cells recognizing the HCMV-pp65 antigen
Authors: Khan Fawad , Müller Thomas R. , Kasmapour Bahram , Ynga-Durand Mario Alberto , Eiz-Vesper Britta , von Einem Jens , Busch Dirk H. , Cicin-Sain Luka
Published: July 2023

Human Cytomegalovirus (HCMV) is a betaherpesvirus that causes severe disease in immunocompromised transplant recipients. Immunotherapy with CD8 T cells specific for HCMV antigens presented on HLA class-I molecules is explored as strategy for long-term relief to such patients, but the antiviral effectiveness of T cell preparations cannot be efficiently predicted by available methods. Therefore, we developed an Assay for Rapid Measurement of Antiviral T-cell Activity (ARMATA) by real-time automated fluorescent microscopy and used it to study the ability of CD8 T cells to neutralize HCMV and control its spread. As a proof of principle, we used TCR-transgenic T cells specific for the immunodominant HLA-A02-restricted tegumental phosphoprotein pp65. pp65 expression follows an early/late kinetic, but it is not clear at which stage of the virus cycle it acts as an antigen. We measured control of HCMV infection by T cells as early as 6 hours post infection (hpi). The timing of the antigen recognition indicated that it occurred before the late phase of the virus cycle, but also that virion-associated pp65 was not recognized during virus entry into cells. Monitoring of pp65 gene expression dynamics by reporter fluorescent genes revealed that pp65 was detectable as early as 6 hpi, and that a second and much larger bout of expression occurs in the late phase of the virus cycle by 48 hpi. Since transgenic (Tg)-pp65 specific CD8 T cells were activated even when DNA replication was blocked, our data argue that pp65 acts as an early virus gene for immunological purposes. Therefore, ARMATA does not only allow same-day identification of antiviral T-cell activity, but also provides a method to define the timing of antigen recognition in the context of HCMV infection.
Left Behind: Modelling the life expectancy disparities amongst people with disabilities in Low and Middle-Income Countries
Authors: Rotenberg Sara , Smythe Tracey , Kuper Hannah
Published: July 2023

Objective

To use life tables to model the differences in life expectancy for people with and without disabilities in low- and middle-income countries (LMICs).

Methods

We used data from a recent conducted a meta-analysis of analysis which gathered data from 70 studies to determine Hazard Ratios (HRs) for all-cause mortality by disability status for children 0-15, adults 15-59, and adults 60+, using the World Health Organisation’s definition of disability. To assess the disparities in life expectancy among people with disabilities across 136 low and middle-income countries (LMICs), we constructed life tables using these HRs.gap compared to the population average. These calculations were based on the 2020 United Nations population projections. The life expectancy gap was meta-analysed across countries to calculate the mean difference.

Findings

People with disabilities in LMIC had a mean life expectancy of 49.3 years (95%C.I. 47.4 – 51.2), compared to 68.5 years (95% C.I. 67.4 – 69.5) for the general population, giving a median gap of 19.2 years (95% C.I. 18.3 – 20.1). The disparity in life expectancy varied across countries, ranging from 9.6 years (95% C.I. 4.7 – 17.4) in Bosnia and Herzegovina to 30.6 years (95% C.I. 16.9 – 40.6) in Nigeria.

Conclusion

Life expectancy inequities represent an urgent threat to upholding the rights of persons with disabilities and achieving global goals. It is crucial to address the disparities in social determinants of health, and prioritise the inclusion of people with disabilities within health equity efforts to close this gap.
“I told myself, be bold and go and test”: Motivators and barriers to HIV testing among gays, bisexuals, and all other men who sex with men in Ghana – West Africa
Authors: Abu-Ba’are Gamji Rabiu , Aidoo-Frimpong Gloria , Stockton Melissa , Zigah Edem Yaw , Amuah Samuel , Amu-Adu Prince , Amoh-Otoo Richard Panix , Nyblade Laura , Torpey Kwasi , Nelson LaRon E.
Published: July 2023

Limited evidence from Ghana and SSA shows that despite consistently a disproportionately high burden of HIV, GBMSM infrequently often delay testing until the point of illness. We therefore used qualitative interviews to collect insights of experiences, motivators, and barriers to HIV testing among GBMSM. Two community-based organizations used snowball and convenience sampling to recruit 10 MSM for IDIs and 8 to 12 for FGDs. We transcribed, coded, identified and analyzed the relationship and commonalities between the participants’ responses. Under experiences with testing, 1) fear of HIV infection created a stressful HIV testing experience; and 2) friendly and supportive healthcare environment facilitated a positive experience in healthcare facilities. Motivators or facilitators of testing include: 1) the perception or belief that HIV testing is a HIV prevention strategy; 2) encouragement from friends and peers; 3) understanding risk associated with certain sexual behaviors such as transactional sex 4) education or information on HIV; 5) access to free testing and incentives; 6) early symptoms and provider recommendation. Barriers to HIV testing include: 1) negative community perceptions of HIV deter; 2) individual-level low risk perception or indifference about HIV infection; 3) location and cost; 4) inadequate testing availability; 5) Stigma at Healthcare facilities. The findings point to the need to address important issues around stigma, education, peer support and healthcare resources through interventions and research to improve HIV testing among GBMSM in the country.
A fast-slow trait continuum at the level of entire communities
Authors: Neyret Margot , Le Provost Gaëtane , Boesing Andrea Larissa , Schneider Florian D. , Baulechner Dennis , Bergmann Joana , Vries Franciska de , Fiore-Donno Anna Maria , Geisen Stefan , Goldmann Kezia , Merges Anna , Saifutdinov Ruslan A. , Simons Nadja K. , Tobias Joseph A. , Zaitsev Andrey S. , Gossner Martin M. , Jung Kirsten , Kandeler Ellen , Krauss Jochen , Penone Caterina , Schloter Michael , Schulz Stefanie , Staab Michael , Wolters Volkmar , Apostolakis Antonios , Birkhofer Klaus , Boch Steffen , Boeddinghaus Runa S. , Bolliger Ralph , Bonkowski Michael , Buscot Francois , Dumack Kenneth , Fischer Markus , Gan Huei Ying , Heinze Johannes , Hölzel Norbert , John Katharina , Klaus Valentin H. , Kleinebecker Till , Marhan Sven , Müller Jörg , Renner Swen C. , Rillig Matthias , Schenk Noëlle V. , Schöning Ingo , Schrumpf Marion , Seibold Sebastian , Socher Stephanie , Solly Emily F. , Teuscher Miriam , van Kleunen Mark , Wubet Tesfaye , Manning Pete
Published: July 2023

Across the tree of life, organismal functional strategies form a continuum from slow-to fast-growing organisms, in response to common drivers such as resource availability and disturbance. However, the synchronization of these strategies at the entire community level is untested. We combine trait data for >2800 above-and belowground taxa from 14 trophic guilds spanning a disturbance and resource availability gradient in German grasslands. Most guilds consistently respond to these drivers through both direct and trophically-mediated effects, resulting in a ‘slow-fast’ axis at the level of the entire community. Fast trait communities were also associated with faster rates of whole ecosystem functioning. These findings demonstrate that ‘slow’ and ‘fast’ strategies can be manifested at the level of whole ecosystems, opening new avenues of ecosystem-level functional classification.
Quantifying Amide-Aromatic Interactions at Molecular and Atomic Levels: Experimentally-determined Enthalpic and Entropic Contributions to Interactions of Amide sp2O, N, C and sp3C Unified Atoms with Naphthalene sp2C Atoms in Water
Authors: Zytkiewicz Emily , Shkel Irina A. , Cheng Xian , Rupanya Anuchit , McClure Kate , Karim Rezwana , Yang Sumin , Yang Felix , Record M. Thomas
Published: July 2023

In addition to amide hydrogen bonds and the hydrophobic effect, interactions involving π-bonded sp2atoms of amides, aromatics and other groups occur in protein self-assembly processes including folding, oligomerization and condensate formation. These interactions also occur in aqueous solutions of amide and aromatic compounds, where they can be quantified. Previous analysis of thermodynamic coefficients quantifying net-favorable interactions of amide compounds with other amides and aromatics revealed that interactions of amide sp2O with amide sp2N unified atoms (presumably C=O···H-N hydrogen bonds) and amide/aromatic sp2C (lone pair-π, n-π*) are particularly favorable. Sp3C-sp3C (hydrophobic), sp3C-sp2C (hydrophobic, CH-π), sp2C-sp2C (hydrophobic, π-π) and sp3C-sp2N interactions are favorable, sp2C-sp2N interactions are neutral, while sp2O-sp2O and sp2N-sp2N self-interactions and sp2O-sp3C interactions are unfavorable. Here, from determinations of favorable effects of fourteen amides on naphthalene solubility at 10, 25 and 45 °C, we dissect amide-aromatic interaction free energies into enthalpic and entropic contributions and find these vary systematically with amide composition. Analysis of these results yields enthalpic and entropic contributions to intrinsic strengths of interactions of amide sp2O, sp2N, sp2C and sp3C unified atoms with aromatic sp2C atoms. For each interaction, enthalpic and entropic contributions have the same sign and are much larger in magnitude than the interaction free energy itself. The amide sp2O-aromatic sp2C interaction is enthalpy-driven and entropically unfavorable, consistent with direct chemical interaction (e.g. lone pair-π) while amide sp3C- and sp2C-aromatic sp2C interactions are entropy-driven and enthalpically unfavorable, consistent with hydrophobic effects. These findings are relevant for interactions involving π-bonded sp2atoms in protein processes.

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Multivalent interactions of the disordered regions of XLF and XRCC4 foster robust cellular NHEJ and drive the formation of ligation-boosting condensatesin vitro
Authors: Vu Duc-Duy , Bonucci Alessio , Brenière Manon , Cisneros-Aguirre Metztli , Pelupessy Philippe , Wang Ziqing , Carlier Ludovic , Bouvignies Guillaume , Cortes Patricia , Aggarwal Aneel K. , Blackledge Martin , Gueroui Zoher , Belle Valérie , Stark Jeremy M. , Modesti Mauro , Ferrage Fabien
Published: July 2023

In mammalian cells, DNA double-strand breaks are predominantly repaired by non-homologous end joining (NHEJ). During repair, the Ku70/80 heterodimer (Ku), XRCC4 in complex with DNA Ligase 4 (X4L4), and XLF form a flexible scaffold that holds the broken DNA ends together. Insights into the architectural organization of the NHEJ scaffold and its regulation by the DNA-dependent protein kinase catalytic subunit (DNA-PKcs) have recently been obtained by single-particle cryo-electron microscopy analysis. However, several regions, especially the C-terminal regions (CTRs) of the XRCC4 and XLF scaffolding proteins, have largely remained unresolved in experimental structures, which hampers the understanding of their functions. Here, we used magnetic resonance techniques and biochemical assays to comprehensively characterize the interactions and dynamics of the XRCC4 and XLF CTRs at atomic resolution. We show that the CTRs of XRCC4 and XLF are intrinsically disordered and form a network of multivalent heterotypic and homotypic interactions that promotes robust cellular NHEJ activity. Importantly, we demonstrate that the multivalent interactions of these CTRs led to the formation of XLF and X4L4 condensatesin vitrowhich can recruit relevant effectors and critically stimulate DNA end ligation. Our work highlights the role of disordered regions in the mechanism and dynamics of NHEJ and lays the groundwork for the investigation of NHEJ protein disorder and its associated condensates inside cells with implications in cancer biology, immunology and the development of genome editing strategies.
Treatment of advanced atherosclerotic mice with the senolytic agent ABT-263 is associated with reduced indices of plaque stability and increased mortality
Authors: Karnewar Santosh , Karnewar Vaishnavi , Shankman Laura S. , Owens Gary K.
Published: July 2023

The use of senolytic agents to remove senescent cells from atherosclerotic lesions is controversial. A common limitation of previous studies is the failure to rigorously define the effects of senolytic agent ABT-263 (Navitoclax) on smooth muscle cells (SMC) despite studies claiming that they are the major source of senescent cells. Moreover, there are no studies of the effect of ABT-263 on endothelial cells (EC), which along with SMC comprise 90% of α-SMA+myofibroblast-like cells in the protective fibrous cap. Here we tested the hypothesis that treatment of advanced atherosclerotic mice with the ABT-263 will reduce lesion size and increase plaque stability. SMC (Myh11-CreERT2-eYFP) and EC (Cdh5-CreERT2-eYFP) lineage tracingApoe-/-mice were fed a WD for 18 weeks, followed by ABT-263 100mg/kg/bw for six weeks or 50mg/kg/bw for nine weeks. ABT-263 treatment did not change lesion size or lumen area of the brachiocephalic artery (BCA). However, ABT-263 treatment reduced SMC by 90% and increased EC-contributions to lesions via EC-to-mesenchymal transition (EndoMT) by 60%. ABT-263 treatment also reduced α-SMA+fibrous cap thickness by 60% and increased mortality by >50%. Contrary to expectations, treatment of WD-fedApoe-/-mice with the senolytic agent ABT-263 resulted in multiple detrimental changes including reduced indices of stability, and increased mortality.

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Highlights

Treatment of Apoe-/- mice with advanced atherosclerosis with the senolytic agent ABT-263 increased mortality by >50%.

ABT-263 showed a 90% reduction in SMC but a 60% increase in endothelial cell (EC) contributions to lesions via EC to mesenchymal transition (EndoMT) but prevented adaptive increases in investment of EC-derived cells into the fibrous cap via beneficial EndoMT to myofibroblast transitions that we have shown normally occur when SMC investment into fibrous cap of lesions is impaired.

Knock out (KO) of Klf4 in SMC, which results in smaller but more stable atherosclerotic lesions, was associated with reduced expression of pro-senescence markers, but preserved expression of the anti-senescence marker, telomerase reverse transcriptase although it is unclear if the latter is causal or an effect.
Oncostatin M is a Master Regulator of an Inflammatory Network inDnmt3a-Mutant Hematopoietic Stem Cells
Authors: Schwartz Logan S. , Young Kira A. , Stearns Timothy M. , Boyer Nathan , Mujica Kristina D. , Trowbridge Jennifer J.
Published: July 2023

Age-associated clonal hematopoiesis (CH) occurs due to somatic mutations accrued in hematopoietic stem cells (HSCs) that confer a selective advantage in the context of aging. The mechanisms by which CH-mutant HSCs gain this advantage with aging are not comprehensively understood. Using unbiased transcriptomic approaches, we identify Oncostatin M (OSM) signaling as a candidate contributor to aging-drivenDnmt3a-mutant CH. We find thatDnmt3a-mutant HSCs from young mice do not functionally respond to acute OSM stimulation with respect to proliferation, apoptosis, hematopoietic engraftment, or myeloid differentiation. However, youngDnmt3a-mutant HSCs transcriptionally upregulate an inflammatory cytokine network in response to acute OSM stimulation including genes encoding IL-6, IL-1β and TNFα. In addition, OSM-stimulatedDnmt3a-mutant HSCs upregulate the anti-inflammatory genesSocs3, Atf3andNr4a1, creating a negative feedback loop limiting sustained activation of the inflammatory network. In the context of an aged bone marrow (BM) microenvironment with chronically elevated levels of OSM,Dnmt3a-mutant HSCs upregulate pro-inflammatory genes but do not upregulateSocs3, Atf3andNr4a1. Together, our work suggests that chronic inflammation with aging exhausts the regulatory mechanisms in young CH-mutant HSCs that resolve inflammatory states, and that OSM is a master regulator of an inflammatory network that contributes to age-associated CH.