bluuebunny/crossref_metadata_2025 huggingface dataset:
KLF7 promotes adipocyte inflammation and glucose metabolism disorder by activating the PKCζ/NF‐κB pathway
Authors: Yang Xin , Liang Maodi , Tang Yihan , Ma Dingling , Li Menghuan , Yuan Chenggang , Hou Yanting , Sun Chaoyue , Liu Jie , Wei Qianqian , Chang Yongsheng , Wang Cuizhe , Zhang Jun
Published: June 2023

In the obesity context, inflammatory cytokines secreted by adipocytes lead to insulin resistance and are key to metabolic syndrome development. In our previous study, we found that the transcription factor KLF7 promoted the expression of p‐p65 and IL‐6 in adipocytes. However, the specific molecular mechanism remained unclear. In the present study, we found that the expression of KLF7, PKCζ, p‐IκB, p‐p65, and IL‐6 in epididymal white adipose tissue (Epi WAT) in mice fed a high‐fat diet (HFD) was significantly increased. In contrast, the expression of PKCζ, p‐IκB, p‐p65, and IL‐6 was significantly decreased in Epi WAT of KLF7 fat conditional knockout mice. In 3T3‐L1 adipocytes, KLF7 promoted the expression of IL‐6 via the PKCζ/NF‐κB pathway. In addition, we performed luciferase reporter and chromatin immunoprecipitation assays, which confirmed that KLF7 upregulated the expression of PKCζ transcripts in HEK‐293T cells. Collectively, our results show that KLF7 promotes the expression of IL‐6 by upregulating PKCζ expression and activating the NF‐κB signaling pathway in adipocytes.
Near‐infrared fluorophoreIR‐61 delays postovulatory aging of mouse oocytes through suppressing oxidative stress mediated by mitochondrial protection
Authors: Qu Jiadan , Luo Yunyao , Qin Lifeng , Guo Jing , Zhu Ling , Li Chong , Xie Juan , Shi Chunmeng , Huang Guoning , Li Jingyu
Published: June 2023

Postovulatory aging can trigger deterioration of oocyte quality and subsequent embryonic development, and thus reduce the success rates of assisted reproductive technology (ART). The molecular mechanisms underlying postovulatory aging, and preventative strategies, remain to be explored. The near‐infrared fluorophore IR‐61, a novel heptamethine cyanine dye, has the potential for mitochondrial targeting and cell protection. In this study, we found that IR‐61 accumulated in oocyte mitochondria and reduced the postovulatory aging‐induced decline in mitochondrial function, including mitochondrial distribution, membrane potential, mtDNA number, ATP levels, and mitochondrial ultrastructure. In addition, IR‐61 rescued postovulatory aging‐caused oocyte fragmentation, defects in spindle structure, and embryonic developmental potential. RNA sequencing analysis indicated that the postovulatory aging‐induced oxidative stress pathway might be inhibited by IR‐61. We then confirmed that IR‐61 decreased the levels of reactive oxygen species and MitoSOX, and increased GSH content in aged oocytes. Collectively, the results indicate that IR‐61 may prevent postovulatory aging by rescuing oocyte quality, promoting successful rate in ART procedure.
Numeracy Education for Health Care Providers: A Scoping Review
Authors: Goldstein Casey , Woods Nicole , MacKinnon Rebecca , Fazelzad Rouhi , Gill Bhajan , Giuliani Meredith Elana , Papadakos Tina , Wei Qinge , Papadakos Janet
Published: June 2023

Introduction:

Health numeracy (numerical literacy) refers to an individual's ability to use numerical information to make effective health decisions. Numeracy is fundamental in the role of a health care provider, forming the basis of evidence-based medicine and effective patient-provider communication. Despite a high level of education, many health care providers struggle with numeracy. Numeracy is often integrated into training curricula; however, teaching modality, competencies covered, learner satisfaction, and effectiveness of these educational interventions varies.

Method:

A scoping review was conducted to explore and summarize what is known about numeracy skills education programs for health care providers. A comprehensive literature search was conducted from January 2010 to April 2021 in 10 databases. Controlled vocabulary terms and text words were used. The search was restricted to human studies, adults, and the English language. Articles were included if they were related to numeracy education for health care providers or trainees and provided details regarding methods, evaluation, and results.

Results:

The literature search retrieved 31,611 results and 71 met the inclusion criteria. Most interventions were conducted in a university setting, and targeted nursing students, medical students, resident physicians, and pharmacy students. Common numeracy concepts included statistics/biostatistics, medication calculations, evidence-based medicine, research methodology, and epidemiology. A variety of teaching modalities were used, which most often combined active approaches (eg, workshops, laboratories, small-group exercises, and discussion boards) with passive approaches (eg, traditional lectures and didactic teaching). Measured outcomes included knowledge and skills, self-efficacy, attitudes, and engagement.

Discussion:

Although efforts have been made to incorporate numeracy into training curricula, greater emphasis should be placed on developing strong numeracy skills in health care providers, particularly given the role numerical information plays in clinical decision making, evidence-based practices, and patient-provider communication.
Basaloid Squamous Cell Carcinomas of the Thymus With Prominent B-Cell Lymphoid Hyperplasia
Authors: Weissferdt Annikka , Moran Cesar A.
Published: June 2023

Ten cases of basaloid squamous cell carcinomas of the thymus are presented. The patients are 6 women and 4 men ranging in ages between 51 and 72 years (average: 61.5 y), who presented with nonspecific symptoms of cough, dyspnea, and chest pain with no history of malignancy, myasthenia gravis, or other autoimmune disease. Surgical resection of the mediastinal masses via thoracotomy or sternotomy was performed in all patients. Grossly, the tumors varied in size from 2 to 8 cm, were light tan in color, solid and slightly hemorrhagic, and had infiltrative borders. Histologically, scanning magnification showed elongated interanastomosing ribbons of tumors cells embedded in a lymphoid stroma containing germinal centers. At higher magnification, the tumors cells were round to oval with moderate amounts of lightly eosinophilic cytoplasm, oval nuclei, moderate cellular atypia, and mitotic activity ranging from 3 to 5 mitotic figures per 10 HPFs. In 8 cases, the tumor invaded perithymic adipose tissue, in 1 case the tumor infiltrated pericardium, and in 1 case, the tumor involved the pleura. Immunohistochemical stains showed positive staining in the epithelial component for pancytokeratin, p63, keratin 5/6, and p40, while CD20 and CD79a characterized the lymphoid component. Clinical follow-up was obtained in 7 patients. Two patients died within 24 months and 5 patients remained alive between 12 and 60 months. The current cases highlight the unusual feature of B-cell lymphoid hyperplasia in these tumors and their potential aggressive behavior.
OPTIMAL PLASMA CONCENTRATION OF THROMBOMODULIN ALFA TO TREAT SEPSIS-INDUCED DISSEMINATED INTRAVASCULAR COAGULATION
Authors: Imaura Masaharu , Katagiri Fumihiko , Nagase Satoshi , Hatoyama-Tanaka Saori , Takahashi Hiroyuki , Takayanagi Risa , Kanno Hiroshi , Yamada Yasuhiko
Published: June 2023

Thrombomodulin alfa (TM alfa) has been shown effective for treatment of disseminated intravascular coagulation (DIC) associated with sepsis, although the optimal therapeutic plasma concentration has not been clarified. In the present study, the plasma trough concentration of TM alfa in septic patients with DIC was determined, then the cutoff value for that concentration showing influence on treatment outcome was calculated using a receiver operating characteristic curve. With a cutoff value of 1,010, the area under the curve of the receiver operating characteristic was 0.669 (95% confidence interval, 0.530–0.808), with sensitivity of 0.458 and specificity of 0.882. To evaluate its accuracy, patients were divided into those above or below the cutoff value, and 90-day survival rates were compared. The above-cutoff group showed a significantly higher 90-day survival rate (91.7%) as compared with the below-cutoff group (63.4%) (P = 0.017), with a hazard ratio of 0.199 (95% confidence interval, 0.045–0.871). Interestingly, the incidence of hemorrhagic adverse effects was not significantly different between the groups. Based on these results, the recommended plasma trough concentration of TM alfa for treatment of septic DIC is 1,010 ng/mL, which should minimize the risk of severe bleeding while maximizing the therapeutic effect.
Evaluation of a portable retinal imaging device: towards a comparative quantitative analysis for morphological measurements of retinal blood vessels
Authors: Martinez-Perez M. Elena , Hughes Alun D. , Thom Simon A. McG. , Parker Kim H. , Witt Nicholas W.
Published: June 2023

This study investigated the possibility of using low-cost, handheld, retinal imaging devices for the automatic extraction of quantifiable measures of retinal blood vessels. Initially, the available handheld devices were compared using a Zeiss model eye incorporating a USAF resolution test chart to assess their optical properties. The only suitable camera of the five evaluated was the Horus DEC 200. This device was then subjected to a detailed evaluation in which images in human eyes taken from the handheld camera were compared in a quantitative analysis with those of the same eye from a Canon CR-DGi retinal desktop camera. We found that the Horus DEC 200 exhibited shortcomings in capturing images of human eyes by comparison with the Canon. More images were rejected as being unevaluable or suffering failures in automatic segmentation than with the Canon, and even after exclusion of affected images, the Horus yielded lower measurements of vessel density than the Canon. A number of issues affecting handheld cameras in general and some features of the Horus in particular have been identified that might contribute to the observed differences in performance. Some potential mitigations are discussed which might yield improvements in performance, thus potentially facilitating use of handheld retinal imaging devices for quantitative retinal microvascular measurements.
Signal detection models as contextual bandits
Authors: Sherratt Thomas N. , O'Neill Erica
Published: June 2023

Signal detection theory (SDT) has been widely applied to identify the optimal discriminative decisions of receivers under uncertainty. However, the approach assumes that decision-makers immediately adopt the appropriate acceptance threshold, even though the optimal response must often be learned. Here we recast the classical normal–normal (and power-law) signal detection model as a contextual multi-armed bandit (CMAB). Thus, rather than starting with complete information, decision-makers must infer how the magnitude of a continuous cue is related to the probability that a signaller is desirable, while simultaneously seeking to exploit the information they acquire. We explain how various CMAB heuristics resolve the trade-off between better estimating the underlying relationship and exploiting it. Next, we determined how naive human volunteers resolve signal detection problems with a continuous cue. As anticipated, a model of choice (accept/reject) that assumed volunteers immediately adopted the SDT-predicted acceptance threshold did not predict volunteer behaviour well. The Softmax rule for solving CMABs, with choices based on a logistic function of the expected payoffs, best explained the decisions of our volunteers but a simple midpoint algorithm also predicted decisions well under some conditions. CMABs offer principled parametric solutions to solving many classical SDT problems when decision-makers start with incomplete information.
A surface plasmon resonance nanostructure containing graphene and BaTiO
3
layers for sensitive defection of organic compounds
Authors: Taya Sofyan A. , Daher Malek G. , Almawgani Abdulkarem H. M. , Hindi Ayman Taher , Colak Ilhami
Published: June 2023

Organic compound-based sensors are used in a variety of significant fields, including medical research, azeotropic calibration, vegetable oil extraction, the shoe industry and geothermal power plants. Here, a high-performance, two-dimensional material-based organic compound sensor has been proposed using a surface plasmon resonance (SPR) nanostructure consisting of a BK7 glass prism, Ag, BaTiO
3
, Ag, graphene and sensing layer. The reflectivity curves of the SPR device have been investigated when the sensing media are Pentane, n-Hexane, n-Heptane and n-Octane. The thickness of the BaTiO
3
layer and the number of graphene sheets have been optimized to maximize the sensitivity. The highest sensitivity attained is 220.83 deg/RIU for n-Octane with 45 nm silver/10 nm BaTiO
3
/8 nm silver and four layers of graphene. We believe that the SPR-based sensors are simple and can replace the spectrometry, chromatography and electrochemical based sensors. The proposed design is extremely effective for diverse applications in biological, industrial and chemical detection because of its simple structure and great performance.
Complex relationships between climate and reproduction in a resident montane bird
Authors: Whitenack Lauren E. , Welklin Joseph F. , Branch Carrie L. , Sonnenberg Benjamin R. , Pitera Angela M. , Kozlovsky Dovid Y. , Benedict Lauren M. , Heinen Virginia K. , Pravosudov Vladimir V.
Published: June 2023

Animals use climate-related environmental cues to fine-tune breeding timing and investment to match peak food availability. In birds, spring temperature is a commonly documented cue used to initiate breeding, but with global climate change, organisms are experiencing both directional changes in ambient temperatures and extreme year-to-year precipitation fluctuations. Montane environments exhibit complex climate patterns where temperatures and precipitation change along elevational gradients, and where exacerbated annual variation in precipitation has resulted in extreme swings between heavy snow and drought. We used 10 years of data to investigate how annual variation in climatic conditions is associated with differences in breeding phenology and reproductive performance in resident mountain chickadees (Poecile gambeli) at two elevations in the northern Sierra Nevada mountains, USA. Variation in spring temperature was not associated with differences in breeding phenology across elevations in our system. Greater snow accumulation was associated with later breeding initiation at high, but not low, elevation. Brood size was reduced under drought, but only at low elevation. Our data suggest complex relationships between climate and avian reproduction and point to autumn climate as important for reproductive performance, likely via its effect on phenology and abundance of invertebrates.
Carrier frequency estimation of pathogenic variants of autosomal recessive and X-linked recessive Mendelian disorders using exome sequencing data in 1,642 Thais
Authors: Chetruengchai Wanna , Phowthongkum Prasit , Shotelersuk Vorasuk
Published: June 2023

People with autosomal recessive disorders often were born without awareness of the carrier status of their parents. The American College of Medical Genetics and Genomics (ACMG) recommends screening 113 genes known to cause autosomal recessive and X-linked conditions in couples seeking to learn about their risk of having children with these disorders to have an appropriate reproductive plan. Here, we analyzed the exome sequencing data of 1,642 unrelated Thai individuals to identify the pathogenic variant (PV) frequencies in genes recommended by ACMG. The ascertainment bias was controlled by excluding the carriers of the PV in the genes for the conditions that are attributed to their offspring disorders. In the 113 ACMG-recommended genes, 165 PV and likely PVs in 60 genes of 559 exomes (34%, 559/1642) were identified. The carrier rate was increased to 39% when glucose-6-phosphate dehydrogenase (G6PD) was added. The carrier rate was still as high as 14.7% when thalassemia and hemoglobinopathies were excluded. In addition to thalassemia, hemoglobinopathies, and G6PD deficiency, carrier frequencies of >1% were found for Gaucher disease, primary hyperoxaluria, Pendred syndrome, and Wilson disease. Nearly 2% of the couples were at risk of having offsprings with the tested autosomal recessive conditions. The expanded carrier screening focused on common autosomal recessive conditions in Thai seems to be benefit among the study samples.
A Novel Bleeding Risk Stratification Scheme in Japanese Patients with Non-valvular Atrial Fibrillation: The J-RISK AF study
Authors: Akao Masaharu , Tomita Hirofumi , Nakai Michikazu , Kodani Eitaro , Suzuki Shinya , Hayashi Kenshi , Sawano Mitsuaki , Goya Masahiko , Yamashita Takeshi , Fukuda Keiichi , Tsuda Toyonobu , Isobe Mitsuaki , Toyoda Kazunori , Miyamoto Yoshihiro , Okamura Tomonori , Sasahara Yusuke , Okumura Ken
Published: June 2023

Background

Oral anticoagulants (OAC) reduce the risk of ischemic stroke, but may increase the risk of major bleeding in non-valvular atrial fibrillation (NVAF) patients. Various risk scores, such as HAS-BLED, ATRIA, and ORBIT, have been proposed to assess the risk of major bleeding in NVAF patients receiving OAC. However, limited data are available on bleeding risk stratification in Japanese NVAF patients.

Methods

Of the 16,098 NVAF patients from the J-RISK AF study, the combined data of the five major AF registries in Japan (J-RHYTHM Registry, Fushimi AF Registry, Shinken Database, Keio interhospital Cardiovascular Studies, and Hokuriku-Plus AF Registry), we analyzed 11,539 patients receiving OAC (median age, 71 years; female, 39.6%; median CHADS2score, 2). Multivariable Cox-hazard proportional analysis was performed to explore significant risk factors for major bleeding. Using those factors, we developed a novel bleeding risk stratification scheme and compared its predictive performance with previously reported risk scores.

Results

During the 2-year follow-up period, major bleeding occurred in 274 patients (1.3% per patient-year). On multivariable analysis, advanced age, uncontrolled hypertension, history of bleeding, anemia, thrombocytopenia, and concomitant antiplatelet agents were significantly associated with higher incidence of major bleeding. We developed a novel risk stratification system, J-RISK bleeding score, that had good predictive performance (C-statistics 0.67) for major bleeding. The predictive performance of our score was better than previous scores.

Conclusion

Our findings suggest that our novel risk stratification system, the J-RISK bleeding score, is more useful than previous score systems for Japanese NVAF patients receiving OAC.
Trends and landscape of cardiovascular disease studies characteristics on ClinicalTrials.gov from 2012 to 2021
Authors: Rawlley Bharat , Bansal Kannu , Dayal Utkarsh , Julka Dhawani , Salooja Ishita , Sanchez Andres Cordova , Gupta Kartik , Kumar Sandeep , Chaudhuri Debanik
Published: June 2023

Introduction

To analyze cardiovascular disease (CVD) studies from the United States registered onClinicalTrials.govfocusing on characteristics associated with their external validity; the presence of Data Monitoring Committee/Data Safety Monitoring Board (DMC/DSMB), randomization, blinding, and gender of the principal investigators (PIs).

Methods

We queried the Application Programming Interface (API) ofClinicalTrials.govto extract data on characteristics of the identified studies, most notably, DMC/DSMB status, Design Allocation, Design masking, and gender of PIs.

Results

We identified 536 studies pertinent to CVD for this analysis. Out of 536, 471 (88%) were interventional studies (Clinical trials) and 65 (12%) were observational studies with interventions. Amongst these, 261 (49%) reported having a DMC, 312 (66%) reported randomization, 224 (48%) reported masking and 122 (19%) of the PIs were women. No trend was seen in the annual proportion of studies with DMC, randomization, masking, and women as PIs (P-trend > 0.05 for all). Multivariable logistic regression analyses were notable for higher odds of DMC (aOR, 5.28; 95% CI, 2.70 – 10.90; P < 0.05) and blinding (aOR, 2.42; 95% CI, 1.29 – 4.64; P < 0.05) in NIH-funded studies and higher odds of being terminated/suspended or withdrawn in basic science studies (aOR, 2.83; 95% CI, 1.07 - 6.90; P < 0.05). No relation was seen between any characteristics and the study being completed.

Conclusions

We report on the absence of DMCs, randomization, blinding, women as PIs, and lack of cross-gender collaboration in the leadership of CVD studies without any favorable trend over the past decade. This calls for comprehensive efforts to improve these trends and ultimately, the external validity of studies. We also call for an overhaul of the definition of the phase of a clinical trial that centers around a drug being the intervention.
Evidence for protein leverage on Total Energy Intake, but not Body Mass Index, in a large cohort of older adults
Authors: Honfo Sèwanou H. , Senior Alistair M. , Legault Véronique , Presse Nancy , Turcot Valérie , Gaudreau Pierrette , Simpson Stephen J. , Raubenheimer David , Cohen Alan A.
Published: June 2023

BACKGROUND

Protein leverage (PL), the phenomenon of food consuming until absolute intake of protein meets a target value, regardless of shortfall or overconsuming for other nutrients in the diet and total energy intake (TEI). Evidence for PL was observed in humans, recently in a cohort of youth with obesity. This study aimed to test for PL and the protein leverage hypothesis (PLH) in a cohort of older adults.

METHODS

We conducted a retrospective analysis of dietary intake in a cohort of 1699 community-dwelling older adults aged 67-84 years from the NuAge cohort. We computed TEI and the energy contribution (EC) from each macronutrient. The strength of leverage of macronutrients was assessed through power functions (TEI=µ * ECL). Body mass index (BMI) was calculated, and mixture models were fitted to predict TEI and BMI from macronutrient ECs.

RESULTS

The mean TEI was 7,673 kJ and macronutrient ECs were 50.4 %, 33.2 % and 16.4 %, respectively for carbohydrates, fat, and protein. High carbohydrate intake was associated with low fat intake. There was a strong negative association (L= -0.37; p < 0.001) between the protein EC and TEI. Each percent of energy intake from protein reduced TEI by 77 kJ on average,ceteris paribus. BMI was unassociated with TEI in this cohort, so the PLH could not be tested here.

CONCLUSIONS

Findings indicate clear evidence for PL on TEI, but not on BMI, likely because TEI and BMI become increasingly uncoupled during aging.
Prognostic Impact of Preoperative Atrial Fibrillation in Patients Undergoing Heart Surgery in Cardiogenic Shock
Authors: Kowalewski Mariusz , Raffa Giuseppe M. , Pasierski Michał , Pasierski Michał , Kołodziejczak Michalina , Litwinowicz Radosław , Wańha Wojciech , Wojakowski Wojciech , Rogowski Jan , Jasiński Marek , Widenka Kazimierz , Hirnle Tomasz , Deja Marek , Bartus Krzysztof , Lorusso Roberto , Tobota Zdzisław , Maruszewski Bohdan , Suwalski Piotr
Published: June 2023

Background

Surgical intervention in the setting of cardiogenic shock (CS) is burdened with high mortality. Due to acute condition, detailed diagnoses and risk assessment is often precluded. Atrial fibrillation (AF) is a risk factor for perioperative complications and worse survival but little is known about AF patients operated in CS. Current analysis aimed to determine prognostic impact of preoperative AF in patients undergoing heart surgery in CS.

Methods

We analyzed data from the Polish National Registry of Cardiac Surgery (KROK) Procedures. Between 2012 and 2021, 4,852 patients presented with CS and were referred for cardiac surgery in 37 centers. Total of 624 (13%) patients had AF history. Cox proportional hazards models were used for computations. Propensity score (nearest neighbor) matching for the comparison of patients with and without AF was performed.

Results

Median follow-up was 4.6 years (max.10.0), mean age was 62 (±15) years and 68% patients were men. Thirty-day mortality was 36%. The origin of CS included acute myocardial infarction (36%), acute aortic dissection (22%) and valvular dysfunction (13%). In an unadjusted analysis, patients with underlying AF had almost 20% higher mortality risk (HR 1.19, 95% CIs 1.06-1.34; P=0.004). Propensity score matching returned 597 pairs with similar baseline characteristics; AF remained a significant prognostic factor for worse survival (HR 1.19, 95% CI 1.00-1.40; P=0.045).

Conclusions

Among patients with CS referred for cardiac surgery, history of AF was a significant risk factor for mortality. Role of concomitant AF ablation and/or left atrial appendage occlusion should be addressed in the future.
High rates of SARS-CoV-2 infection in pregnant Ugandan women and association with stunting in infancy
Authors: Jacobson Karen B. , Röltgen Katharina , Lam Brandon , Nayebare Patience , Kakuru Abel , Kizza Jimmy , Aguti Miriam , Nankya Felistas , Briggs Jessica , Takahashi Saki , Greenhouse Bryan , Rodriguez-Barraquer Isabel , Ploeg Kattria van der , Wohlstadter Jacob N. , Sigal George B. , Roh Michelle E , Nankabirwa Joaniter I. , Cuu Gloria , Gaw Stephanie L. , Rosenthal Philip J. , Kamya Moses R. , Ssewanyana Isaac , Dorsey Grant , Boyd Scott D. , Jagannathan Prasanna
Published: June 2023

Background

SARS-CoV-2 has been well studied in resource-rich areas but many questions remain about effects of infection in African populations, particularly in vulnerable groups such as pregnant women.

Methods

We describe SARS-CoV-2 immunoglobulin (Ig) G and IgM antibody responses and clinical outcomes in mother-infant dyads enrolled in malaria chemoprevention trials in Uganda.

Results

From December 2020 to February 2022, among 400 unvaccinated pregnant women, serologic assessments revealed that 128 (32%) were seronegative for anti-SARS-CoV-2 IgG and IgM at enrollment and delivery, 80 (20%) were infected either prior to or early in pregnancy, and 192 (48%) were infected or re-infected with SARS-CoV-2 during pregnancy. We observed preferential binding of plasma IgG to Wuhan-Hu-1-like antigens in individuals seroconverting up to early 2021, and to Delta variant antigens in a subset of individuals in mid-2021. Breadth of IgG binding to all variants improved over time. No participants experienced severe respiratory illness during the study. SARS-CoV-2 infection in early pregnancy was associated with lower median length-for-age Z-score at age 3 months compared with no infection or late pregnancy infection (- 1.54 versus −0.37 and −0.51, p=0.009).

Conclusion

Pregnant Ugandan women experienced high levels of SARS-CoV-2 infection without severe respiratory illness. Variant-specific serology testing demonstrated evidence of antibody affinity maturation at the population level. Early gestational SARS-CoV-2 infection was associated with shorter stature in early infancy.

Funding

This work was supported by: Stanford MCHRI/Stephen Bechtel Endowed Fellowship in Pediatric Translational Medicine (KJ), Swiss National Science Foundation PRIMA grant PR00P3_208580 (KR), the Bill and Melinda Gates Foundation, and NIAID (T32-AI052073, U01- AI141308, U01-AI155325).
The Benchtop mesoSPIM: a next-generation open-source light-sheet microscope for large cleared samples
Authors: Vladimirov Nikita , Voigt Fabian F. , Naert Thomas , Araujo Gabriela R. , Cai Ruiyao , Reuss Anna Maria , Zhao Shan , Schmid Patricia , Hildebrand Sven , Schaettin Martina , Groos Dominik , Mateos José María , Bethge Philipp , Yamamoto Taiyo , Aerne Valentino , Roebroeck Alard , Ertürk Ali , Aguzzi Adriano , Ziegler Urs , Stoeckli Esther , Baudis Laura , Lienkamp Soeren S. , Helmchen Fritjof
Published: June 2023

In 2015, we launched the mesoSPIM initiative (www.mesospim.org), an open-source project for making light-sheet microscopy of large cleared tissues more accessible. Meanwhile, the demand for imaging larger samples at higher speed and resolution has increased, requiring major improvements in the capabilities of light-sheet microscopy. Here, we introduce the next-generation mesoSPIM (“Benchtop”) with significantly increased field of view, improved resolution, higher throughput, more affordable cost and simpler assembly compared to the original version. We developed a new method for testing objectives, enabling us to select detection objectives optimal for light-sheet imaging with large-sensor sCMOS cameras. The new mesoSPIM achieves high spatial resolution (1.5 µm laterally, 3.3 µm axially) across the entire field of view, a magnification up to 20x, and supports sample sizes ranging from sub-mm up to several centimetres, while being compatible with multiple clearing techniques. The new microscope serves a broad range of applications in neuroscience, developmental biology, and even physics.
Comparison of Transcatheter Edge-to-Edge Mitral Valve Repair for Primary Mitral Regurgitation Outcomes to Hospital Volumes of Surgical Mitral Valve Repair
Authors: Grayburn Paul A. , Mack Michael J. , Manandhar Pratik , Kosinski Andrzej S. , Sannino Anna , Smith Robert L. , Szerlip Molly , Vemulapalli Sreekanth
Published: June 2023

Background

Transcatheter edge-to-edge mitral valve (MV) repair (TEER) is an effective treatment for patients with primary mitral regurgitation (MR) at prohibitive risk for surgical MV repair (MVr). High volume MVr centers and high volume TEER centers have better outcomes than low volume centers, respectively. However little is known about whether MVr volume, and specifically complex MVr volume, predicts TEER outcomes. We hypothesized that high volume MV surgical centers would have superior risk-adjusted outcomes for TEER than tlow volume centers.

Methods

We combined data from the ACC/STS TVT registry and the STS adult cardiac surgery database. Complex MVr, defined as leaflet resection or artificial chords with or without annuloplasty was evaluated as a continuous variable and as pre-defined categories (<25, 25-49 and ≥ 50 MV repairs/year). A generalized linear mixed model was used to evaluate risk-adjusted in-hospital/30-day mortality, 30 day HF readmission and TEER success (MR ≤ 2+ and gradient < 5 mmHg).

Results

The study comprised 41,834 patients from 500 sites. TEER mortality at 30-days was 3.5% with no significant difference across MVr volume on unadjusted (p=0.141) or adjusted (p=0.071) analysis of volume as a continuous variable. One-year mortality was 15.0% and was lower for higher MVr volume centers when adjusted for clinical and demographic variables (p=0.027). HF readmission at one year was 9.4% and was statistically significantly lower in high volume centers on both unadjusted (p=0.017) or adjusted (p-0.015) analysis. TEER success was 54.6% and was not statistically significantly different across MV surgical site volumes (p=0.4271).

Conclusions

TEER can be safely performed in centers with low volumes of complex MV repair. However, one-year mortality and HF readmission are superior at centers with higher MVr volume.
Click-Based Determination of Accumulation of Molecules inEscherichia coli
Authors: Ongwae George M. , Liu Zichen , Feng Shasha , Chordia Mahendra D. , Sharifian Gh. Mohammad , Dash Rachita , Dalesandro Brianna E. , Guo Taijie , Sharpless Karl Barry , Dong Jiajia , Siegrist M. Sloan , Im Wonpil , Pires Marcos M.
Published: June 2023

Gram-negative bacterial pathogens pose a significant challenge in drug development due to their outer membranes, which impede the permeation of small molecules. The lack of widely adoptable methods to measure the cytosolic accumulation of compounds in bacterial cells has hindered drug discovery efforts. To address this challenge, we developed the CHloroalkane Azide Membrane Permeability (CHAMP) assay, specifically designed to assess molecule accumulation in the cytosol of Gram-negative bacteria. The CHAMP analysis utilizes biorthogonal epitopes anchored within HaloTag-expressing bacteria and measures the cytosolic arrival of azide-bearing test molecules through strain-promoted azide-alkyne cycloaddition. This workflow allows for robust and rapid accumulation measurements of thousands of azide-tagged small molecules. Our approach consistently yields a large number of accumulation profiles, significantly exceeding the scale of previous measurements inEscherichia coli(E. coli). We have validated the CHAMP assay across various chemical and biological contexts, including hyperporinated cells, membrane-permeabilized cells, and E. coli strains with impaired TolC function, a key component of the efflux pump. The CHAMP platform provides a simple, high-throughput, and accessible method that enables the analysis of over 1,000 molecules within hours. This technique addresses a critical gap in antimicrobial research, potentially accelerating the development of effective agents against Gram-negative pathogens.
Cell arrangement impacts metabolic activity and antibiotic tolerance inPseudomonas aeruginosabiofilms
Authors: Dayton Hannah , Kiss Julie , Wei Mian , Chauhan Shradha , LaMarre Emily , Cornell William Cole , Morgan Chase J. , Janakiraman Anuradha , Min Wei , Tomer Raju , Price-Whelan Alexa , Nirody Jasmine A , Dietrich Lars E.P.
Published: June 2023

Cells must access resources to survive, and the anatomy of multicellular structures influences this access. In diverse multicellular eukaryotes, resources are provided by internal conduits that allow substances to travel more readily through tissue than they would via diffusion. Microbes growing in multicellular structures, called biofilms, are also affected by differential access to resources and we hypothesized that this is influenced by the physical arrangement of the cells. In this study, we examined the microanatomy of biofilms formed by the pathogenic bacteriumPseudomonas aeruginosaand discovered that clonal cells form striations that are packed lengthwise across most of a mature biofilm’s depth. We identified mutants, including those defective in pilus function and in O-antigen attachment, that show alterations to this lengthwise packing phenotype. Consistent with the notion that cellular arrangement affects access to resources within the biofilm, we found that while the wild type shows even distribution of tested substrates across depth, the mutants show accumulation of substrates at the biofilm boundaries. Furthermore, we found that altered cellular arrangement within biofilms affects the localization of metabolic activity, the survival of resident cells, and the susceptibility of subpopulations to antibiotic treatment. Our observations provide insight into cellular features that determine biofilm microanatomy, with consequences for physiological differentiation and drug sensitivity.
Astrocyte interferon-gamma signaling dampens inflammation during chronic central nervous system autoimmunity via PD-L1
Authors: Smith Brandon C. , Tinkey Rachel A. , Mariam Arshiya , Habean Maria L. , Dutta Ranjan , Williams Jessica L.
Published: June 2023

Multiple sclerosis (MS) is an inflammatory and neurodegenerative disease of the central nervous system (CNS). Infiltrating inflammatory immune cells perpetuate demyelination and axonal damage in the CNS and significantly contribute to pathology and clinical deficits. While the cytokine interferon (IFN)γ is classically described as deleterious in acute CNS autoimmunity, we and others have shown astrocytic IFNγ signaling also has a neuroprotective role. Here, we performed RNA sequencing and ingenuity pathway analysis on IFNγ-treated astrocytes and found that PD-L1 was prominently expressed. Using a PD-1/PD-L1 antagonist, we determined that apoptosis was reduced in leukocytes exposed to IFNγ-treated astrocytesin vitro. To further elucidate the role of astrocytic IFNγ signaling on the PD-1/PD-L1 axisin vivo, we induced the experimental autoimmune encephalomyelitis (EAE) model of MS inAldh1l1-CreERT2,Ifngr1fl/flmice. Mice with conditional astrocytic deletion of IFNγ receptor exhibited a reduction in PD-L1 expression which corresponded to increased infiltrating leukocytes, particularly from the myeloid lineage, and exacerbated clinical disease. PD-1 agonism reduced EAE severity and CNS-infiltrating leukocytes. Importantly, PD-1 is expressed by myeloid cells surrounding MS lesions. These data support that IFNγ signaling in astrocytes diminishes inflammation during chronic autoimmunity via upregulation of PD-L1, suggesting potential therapeutic benefit for MS patients.
N-terminal targeting sequences and coding sequences act in concert to determine the localization and trafficking pathway of apicoplast proteins inToxoplasma gondii
Authors: Anjum Sofia , Prasad Aparna , Mastud Pragati , Patankar Swati
Published: June 2023

Toxoplasma gondiihas a relict plastid, the apicoplast, to which proteins are targeted after synthesis in the cytosol. Proteins exclusively found in the apicoplast use a Golgi-independent route for trafficking, while dually targeted proteins found in both the apicoplast and the mitochondrion use a Golgi-dependent route. For apicoplast targeting, N-terminal signal sequences have been shown to direct the localization of different reporters. In this study, we use chimeric proteins to dissect out the roles of N-terminal sequences and coding sequences in apicoplast localization and the choice of the trafficking route. We show that when the N-termini of a dually targeted protein,TgTPx1/2, or of an apicoplast protein,TgACP, are fused with the reporter protein, enhanced Green Fluorescent Protein (eGFP) or endogenous proteins,TgSOD2,TgSOD3,TgACP orTgTPx1/2, the chimeric proteins exhibit flexibility in apicoplast targeting depending on the coding sequences. Further, the chimeras that are localized to the apicoplast use different trafficking pathways depending on the combination of the N-terminal signals and the coding sequences. This report shows, for the first time, that in addition to the N-terminal signal sequences, targeting and trafficking signals also reside within the coding sequences of apicoplast proteins.
Effect of a retinoic acid analogue on BMP-driven pluripotent stem cell chondrogenesis
Authors: Mancini Fabrizio E. , Humphreys Paul E.A. , Woods Steven , Bates Nicola , Cuvertino Sara , O’Flaherty Julieta , Biant Leela , Domingos Marco A.N. , Kimber Susan J.
Published: June 2023

Osteoarthritis is the most common degenerative joint condition, leading to articular cartilage (AC) degradation, chronic pain and immobility. The lack of appropriate therapies that provide tissue restoration combined with the limited lifespan of joint-replacement implants indicate the need for alternative AC regeneration strategies. Differentiation of human pluripotent stem cells (hPSCs) into AC progenitors may provide a long-term regenerative solution but are still limited due to the continued reliance upon growth factors to recapitulate developmental signalling processes. Recently, TTNPB, a small molecule activator of retinoic acid receptors (RARs), has been shown to be sufficient to guide mesodermal specification and early chondrogenesis of hPSCs. Here, we modified our previous differentiation protocol, by supplementing cells with TTNPB and administering BMP2 at specific times to enhance early development. Transcriptomic analyses indicated that activation of RAR signalling significantly upregulated genes related to limb and embryonic skeletal development in the early stages of the protocol and upregulated genes related to AC development in later stages. Chondroprogenitors obtained from RAPID-E could generate cartilaginous pellets that expressed AC-related matrix proteins such as Lubricin, Aggrecan, and Collagen II. This protocol could lay the foundations for cell therapy strategies for osteoarthritis and improve the understanding of AC development in humans.
Drug Resistance without a cost? Common and uncommon routes to fosfomycin resistance in UropathogenicEscherichia coli
Authors: Bermudez Tomas A. , Brannon John R. , Dudipala Neha , Reasoner Seth , Morales Grace , Wiebe Michelle , Cecala Mia , Beebout Connor , Amir Omar , Hadjifrangiskou Maria
Published: June 2023

Fosfomycin kills bacteria by blocking the binding of phosphoenolpyruvate (PEP) to the bacterial enzyme MurA and halting peptidoglycan synthesis. While its use has increased with the emergence of antibiotic resistance, the mechanisms leading to fosfomycin resistance remain relatively unexplored. In uropathogenicEscherichia coli(UPEC) that accounts for >75% of urinary tract infections (UTIs), fosfomycin enters the cell primarily through UhpT, which transports glucose-6-phosphate (G6P) glycolysis intermediate into the cell. Mutations inuhpT lead to fosfomycin resistance and have been identified during antimicrobial susceptibility testing (AST) in non-susceptible inner colonies that form within the zone of inhibition. However, EUCAST and CLSI guidelines differ in how to read fosfomycin AST when such resistant colonies arise. Work from our lab and others demonstrated that glycolysis is dispensable during acute UTI. Moreover, G6P is scarce in urine, prompting us to test the hypothesis thatuhpmutations may not impart a fitness cost to the pathogen. We report that loss ofuhpindeed does not impair UPEC pathogenesis and that clinical isolates exist that lack theuhplocus altogether. Analysis of non-susceptible inner colonies revealed a suite of novel genes involved in fosfomycin resistance. One of them is PykF that converts PEP to pyruvate during glycolysis. Single deletions ofpykFor its anaerobic homologpykAdo not attenuate UPEC. Based on our data, we raise the alarm that multiple routes lead to fosfomycin resistance and do not affect pathogenesis and propose that the current EUCAST and CLSI guidelines unify into how they evaluate fosfomycin AST.

IMPORTANCE

While fosfomycin resistance is rare, the observation of non-susceptible subpopulations among clinicalEscherichia coliisolates is a common phenomenon during antimicrobial susceptibility testing (AST) in American and European clinical labs. Previous evidence suggests that mutations eliciting this phenotype are of high biological cost to the pathogen during infection, leading to current recommendations of neglecting non-susceptible colonies during AST. Here we report that the most common route to fosfomycin resistance, as well as novel routes described in this work do not impair virulence in uropathogenicE. coli, the major cause of urinary tract infections, suggesting a re-evaluation of current susceptibility guidelines is warranted.
Defining the impact of flavivirus envelope protein glycosylation on sensitivity to broadly neutralizing antibodies
Authors: Contreras Maya , Stuart Jackson B. , Levoir Lisa M. , Belmont Laura , Goo Leslie
Published: June 2023

Antibodies targeting the so-called envelope dimer epitope (EDE) cross-neutralize Zika virus (ZIKV) and all four dengue virus (DENV) serotypes and have thus inspired an epitope-focused vaccine design against these flaviviruses. There are two EDE antibody subclasses (EDE1, EDE2) distinguished by their dependence on viral envelope (E) proteinN-linked glycosylation at position N153 (DENV) or N154 (ZIKV) for binding. Here, we determined how E glycosylation affectsneutralizationby EDE and other broadly neutralizing antibodies. Consistent with structural studies, mutations abolishing the N153/N154 glycosylation site increased DENV and ZIKV sensitivity to neutralization by EDE1 antibodies. Surprisingly, these mutations also increased sensitivity to EDE2 antibodies although they occurred at predicted contact sites. Despite preserving the glycosylation site motif (N-X-S/T), substituting the threonine at ZIKV E residue 156 with a serine resulted in loss of glycan occupancy accompanied with increased neutralization sensitivity to EDE antibodies. For DENV, the presence of a serine instead of a threonine at E residue 155 retained glycan occupancy, but nonetheless increased sensitivity to EDE antibodies, in some cases to a similar extent as mutation at N153, which abolishes glycosylation. E glycosylation site mutations also increased ZIKV and DENV sensitivity to other broadly neutralizing antibodies, but had limited effects on ZIKV-or DENV-specific antibodies. Thus, E protein glycosylation is context-dependent and modulates the potency of broadly neutralizing antibodies in a manner not predicted by existing structures. Manipulating E protein glycosylation could be a novel strategy for engineering vaccine antigens to elicit antibodies that broadly neutralize ZIKV and DENV.

IMPORTANCE

Antibodies that can potently cross-neutralize Zika (ZIKV) and dengue (DENV) viruses are attractive to induce via vaccination to protect against these co-circulating flaviviruses. Structural studies have shown that viral envelope protein glycosylation is important for binding by one class of these so-called broadly neutralizing antibodies, but less is known about the determinants of neutralization. Here, we investigated how envelope protein glycosylation impacts broadly neutralizing antibody potency. By characterizing a panel of ZIKV and DENV variants encoding envelope protein glycosylation site mutations, we found that glycan occupancy was not always predicted by an intact N-X-S/T sequence motif. Moreover, envelope protein glycosylation status alters the neutralization potency of broadly neutralizing antibodies in a manner unexpected from their predicted binding mechanism as determined by existing structures. We highlight the complex role and determinants of envelope protein glycosylation that should be considered in the design of vaccine antigens to elicit broadly neutralizing antibodies.
GRAS1 non-coding RNA protects against DNA damage and cell death by binding and stabilizing NKAP
Authors: Su Tong , Trang Nhu , Zhu Jonathan , Kong Lingbo , Cheung Darin , Chou Vita , Ellis Lauren , Huang Calvin , Camden Nichelle , McHugh Colleen A.
Published: June 2023

Non-coding RNA (ncRNA) gene products are involved in diverse biological processes including splicing, epigenetic regulation, gene expression, proliferation, and metabolism. The biological mechanisms by which ncRNAs contribute to cell survival remain poorly understood. We found that the Growth Regulator Antisense 1 (GRAS1) long non-coding RNA (lncRNA) transcript promotes growth in multiple human cell types by protecting against DNA damage. Knockdown of GRAS1 induced DNA damage and cell death, along with significant expression changes in DNA damage response, intrinsic apoptotic signaling, and cellular response to environmental stimulus genes. Extensive DNA damage occurred after GRAS1 knockdown, with numerous double strand breaks occurring in each cell. The number of cells undergoing apoptosis and with fragmented nuclei increased significantly after GRAS1 knockdown. We used RNA antisense purification and mass spectrometry (RAP-MS) to identify the NF-κB activating protein (NKAP) as a direct protein interaction partner of GRAS1 lncRNA. NKAP protein was degraded after GRAS1 knockdown, in a proteasome-dependent manner. Overexpression of GRAS1 or NKAP mitigated the DNA damage effects of GRAS1 knockdown. In summary, GRAS1 and NKAP directly interact to protect against DNA damage and cell death in multiple human cell lines.