bluuebunny/crossref_metadata_2025 huggingface dataset:
Synthesis of Chitosan Microparticles Encapsulating Bacterial Cell-Free Supernatants and Indole Acetic Acid, and Their Effects on Germination and Seedling Growth in Tomato (Solanum lycopersicum)
Authors: Gonzalez-Montfort Thania Soledad , Almaraz-Abarca Norma , Pérez-y-Terrón Rocío , Ocaranza-Sánchez Erik , Rojas-López Marlon
Published: November 2022

Encapsulation of biostimulant metabolites has gained popularity as it increases their shelf life and improves their absorption, being considered a good alternative for the manufacture of products that stimulate plant growth and fruit production. Cell-free supernatants (CFS) were obtained from nine indole-3-acetic acid (IAA) producing bacterial strains. Stenotrophomonas maltophilia (PT53T) produced the highest concentration of IAA (15.88 μg/mL) after 48 h of incubation. CFS from this strain, as well as an IAA standard were separately encapsulated in chitosan microparticles (CS-MP) using the ionic gelation method. The CS-MP were analyzed by Fourier transform infrared spectroscopy (FTIR), showing absorption bands at 1641, 1547, and 1218 cm−1, associated with the vibrations of the carbonyl C=O, the N-H amine, and the bond between chitosan (CHI) and sodium tripolyphosphate (TPP). The effects of unencapsulated CFS, encapsulated CFS (EN-CFS), and encapsulated IAA standard (EN-IAA) on germination and growth of seven-day-old tomato (Solanum lycopersicum) seedlings were studied. Results showed that both EN-CFS and EN-IAA significantly (

p
<
0.05

) increased seed germination rates by 77.5 and 80.8%, respectively. Both CFS and EN-IAA produced the greatest increase in aerial part length and fresh weight with respect to the treatment-free test. Therefore, it was concluded that the application of EN-CFS or EN-IAA could be a good option to improve the germination and growth of tomato seedlings.
Cytokines in Thyroid-Associated Ophthalmopathy
Authors: Zhang Pengbo , Zhu Huang
Published: November 2022

Thyroid-associated ophthalmopathy (TAO), also known as thyroid eye disease (TED) or Graves’ orbitopathy (GO), is a complex autoimmune condition causing visual impairment, disfigurement, and harm to patients’ physical and mental health. The pathogenesis of TAO has not been fully elucidated, and the mainstream view is that coantigens shared by the thyroid and orbit trigger remodeling of extraocular muscles and orbital connective tissues through an inflammatory response. In recent years, cytokines and the immune responses they mediate have been crucial in disease progression, and currently, common evidence has shown that drugs targeting cytokines, such as tocilizumab, infliximab, and adalimumab, may be novel targets for therapy. In this review, we summarize the research development of different cytokines in TAO pathogenesis in the hope of discovering new therapeutic targets.
Approaches for Delineating Groundwater Recharge Potential Zone Using Fuzzy Logic Model
Authors: Haile Mesfin Mamo
Published: November 2022

Groundwater recharge potential is an important spatial analysis to refer where the groundwater recharge is likely to happen. The present study discusses approaches for determining groundwater recharge potential zones in the Guder watershed, Ethiopia, using geospatial and fuzzy logic techniques on derived thematic layers based on significant influence of groundwater occurrence. A variety of thematic layers were created using satellite images and other ancillary data, including drainage density, slope, rainfall, geology, land use/land cover, lineament density, soil, and geomorphology. Researchers, expert opinion, and previous studies were used to estimate the membership value for each governing factors. According to the findings of the study, approximately 5.25% of the area has very high groundwater recharge potential, and 9.34% of the area has high groundwater recharge potential, while the areas with very low, low, and moderate groundwater recharge potential are approximately 20.04%, 34.21%, and 31.15%, respectively. The study’s findings were confirmed by looking at the cross-tabulation area between the contributing features and the final map, which demonstrated strong associations. The findings of the study will help decision-makers and policymakers plan to implement appropriate groundwater recharge procedures. Fuzzy logic approaches have been found to be a basic tool for estimating groundwater recharge locations when used with GIS.
The Use of Blended Teaching in Higher Medical Education during the Pandemic Era
Authors: Fu Xue-Tao , Hu Yi , Yan Bing-Chun , Jiao Yun-Gen , Zheng Shi-Jun , Wang Ying-Ge , Zhang Jiang-Yun , Wang Zheng-Bing
Published: November 2022

Objective. This study aims to compare the effect of blended teaching and traditional teaching in higher medical education during the pandemic era. Methods. Taking the teaching of neurology as an example, 293 Yangzhou University Clinical Medicine 2016 undergraduate students were selected as the research subjects, and were randomly divided into 2 groups a blended teaching group (n = 148) and a traditional teaching group (n = 145), and received blended teaching and traditional teaching, respectively. The blended teaching was based on a Massive Open Online Course, problem-based learning, and case-based learning and supplemented by Tencent video conferences, QQ messaging groups, and other auxiliary teaching tools. At the end of the course, the teaching effect and satisfaction rate were evaluated through theory assessment, practical skills assessment, and an anonymous questionnaire survey. Results. There were significant differences in theoretical achievements (81.83 ± 6.23 vs 76.79 ± 6.87,

P
<
0.001

) and practical skill achievements (84.74 ± 6.50 vs 78.48 ± 6.53,

P
<
0.001

). In addition, significant differences in all aspects of satisfaction rate were observed between the two groups (all

P
<
0.001

). Conclusion. Blended teaching is beneficial to students’ learning and stimulates their enthusiasm, cultivates clinical thinking ability, and improves teaching quality. Thus, it has played a positive role in the reform of higher medical teaching during the pandemic era.
Prognostic Nomogram for Gastrointestinal Stromal Tumors after Surgery Based on the SEER Database
Authors: Sun Qianhui , Chen Yunru , Li Tingting , Zhu Xiaoyu , Zhu Guanghui , Ni Baoyi , Gao Ruike , Li Jie
Published: November 2022

We aimed to determine prognostic factors and develop an effective and practical nomogram for predicting cancer‐specific survival in gastrointestinal stromal tumor (GIST) patients. Postoperative data were obtained from the SEER database (2000‐2018). Patients were divided into training and validation cohorts at random (7 : 3). Prognostic factors were screened, and a prognostic nomogram was established using log‐rank testing and Cox regression. We used DCA, ROC curves, C‐index, and calibration curves to evaluate our model’s predictive performance. The clinical value of the nomogram and the modified National Institute of Health (M‐NIH) classification were compared using the NRI and IDI. The Kaplan‐Meier method was applied to examine survival by risk group, and log‐rank tests were applied to compare variations in survival curves. Independent prognostic risk factors associated with cancer‐specific survival on multivariate Cox proportional hazards regression analysis were age, race, and tumor location, size, grade, and stage. Clinically relevant variables need to be considered in addition to statistically significant variables when developing prognostic models to aid clinical decision‐making. We included two additional variables (mitotic rate and chemotherapy) when constructing the prognostic model. The C‐index was 0.766 (95% confidence interval (CI): 0.737‐0.794) in the training cohort and 0.795 (95% CI: 0.754‐0.836) in the internal validation group suggesting robustness. The areas under the ROC curve for three‐year and five‐year survival were >0.700, indicating satisfactory discrimination. The calibration curves showed good agreement between the predictions of the nomogram and the actual results. The NRI (0.346 for 3‐year and 0.265 for 5‐year cancer‐specific survival for patients with GIST (GSS) prediction; validation cohort: 0.356 for 3‐year and 0.246 for 5‐year GSS prediction) and IDI values (0.047 for 3‐year and 0.060 for 5‐year GSS prediction; validation cohort: 0.071 for 3‐year and 0.084 for 5‐year GSS prediction) suggested that the established nomogram performed significantly better than the M‐NIH classification. The DCA indicated that the nomogram was clinically useful and had a high discriminative ability in identifying patients who were at high risk of poor outcomes. According to nomogram findings, patients were divided into three groups (high, moderate, and low risk), with significantly different prognoses in both cohorts. Our nomogram satisfactorily predicted survival in postsurgical GIST patients, which may assist clinicians to evaluate the postoperative status and guide subsequent treatments.
Exploring the Influencing Factors for Contraceptive Use among Women: A Meta-Analysis of Demographic and Health Survey Data from 18 Developing Countries
Authors: Islam Md. Akhtarul , Khan Md. Nafiul Alam , Raihan Hasin , Barna Sutapa Dey
Published: November 2022

Background. The primary objective of this research was to investigate how socioeconomic and demographic factors influence the usage of contraceptives by women in 18 developing countries. Methods. The study used the latest DHS data from 18 developing countries in order to acquire a broad perspective of contraceptive methods. We applied meta-analysis techniques for 18 developing countries to find out the summary results. Results. The overall summary effect showed that the variable respondent education (

OR
=
1.39

; 95% CI: 1.17 to 1.65), husband education (

OR
=
1.60

; 95% CI: 1.32 to 1.93), type of place of residence (

OR
=
0.88

; 95% CI 0.78 to 0.98), current working status (

OR
=
1.47

; 95% CI 1.30 to 1.66), age of the respondent (

OR
=
3.41

; 95% CI 2.35 to 4.93), breastfeeding status (

OR
=
1.34

; 95% CI 1.11 to 1.62), and desire for more children (

OR
=
0.53

; 95% CI 0.43 to 0.65) were the significant factors for contraceptive utilization in developing countries. Conclusions. According to the findings of this descriptive study, the respondent’s age, level of education, and work status were shown to be the most significant factors that influence the usage of contraceptives in developing countries. It is necessary to take reasonable steps in order to increase the rate of utilizing methods of contraception among women of reproductive age who are uneducated, living in rural areas, and unemployed.
Clinical Features and Risk Factors of Mortality in Patients with Posterior Reversible Encephalopathy Syndrome
Authors: Cui Hong-Wei , Lei Ru-Yi , Zhang Bo-Ai
Published: November 2022

Objective. Although the prognosis of posterior reversible encephalopathy syndrome (PRES) is usually favourable and most patients wholly recover, the disorder can result in death in some patients. To date, the data on clinical features and risk factors for death are still lacking; therefore, we aim to investigate the clinical features and long-term prognostic risk factors of PRES in the present study. Methods. The patients with PRES were identified from the First Affiliated Hospital of Zhengzhou University from June 2011 to June 2020. Clinical characteristics, laboratory tests, magnetic resonance imaging examinations, and treatment of all patients were analyzed retrospectively. All patients were followed up by telephone. Finally, the patients were divided into the survival group and death group for prognosis analysis. Results. A total of 92 patients with PRES were included; 84.8% of whom were female, with an average age of 25.4 (5–66) years at the onset of PRES. Epilepsy was the main clinical manifestation (72.8%). The in-hospital mortality rate was 2.17%. The 3-year all-cause survival rate for PRES patients was 86%. In univariate analysis, patients with systemic lupus erythematosus (

P
=
0.027

) and blood transfusion history within 1 month before onset (

P
=
0.027

), need for dialysis (

P
≤
0.001

), nephritis (

P
=
0.010

), stroke (

P
=
0.016

), and heart failure (

P
=
0.016

) were associated with death. In multivariate analysis, we found that heart failure (

OR
=
0.095

, 95% CI 0.020 to 0.441) and stroke (

OR
=
0.033

, 95% CI 0.002 to 0.467) were independent risk factors for death in PRES patients, while pregnancy was a protective factor for death in PRES patients (

OR
=
7.978

, 95% CI 1.446 to 44.006). Conclusions. Our results indicate that PRES could be considered as a sign of a very high-risk patient. We also demonstrated that heart failure and stroke were independent risk factors for death in patients with PRES; moreover, pregnancy was a protective factor.
Classification of Thoracic Diseases Based on Chest X-ray Images Using Kernel Support Vector Machine
Authors: Khan Rijah , Mehmood Tahir
Published: November 2022

Machine learning is the leading field of artificial intelligence that has achieved expert-level performance. Diagnosis and treatment of various medical diseases have led to advancements in medical imaging. Chest X-ray-based thoracic disease classification or identification is one of the potential applications in medical imaging based on machine learning. The study consists of 112,120 images of 30,804 individual patients with fourteen thoracic disease labels, which encapsulated the support vector machine (SVM). We have considered 04 kernels in SVM, namely, linear (L-SVM), polynomial (P-SVM), radial basis (R-SVM), and hyperbolic tangent (H-SVM) for classification of thoracic diseases based on X-ray images. To reduce the dimensionality and outliers from the SVM, variants are coupled with novel fast principal component analysis (FPCA). It appears that there is a significant

p
≤
0.05

difference between SVM variants where P-SVM and R-SVM next in order outperforms on most of the disease identification models with average validated classification accuracy ranging from 92% to 98%. The average calibrated accuracy ranges from 99.5% and reaches to 100% in most of the cases. The study is worth investigating as it is good for radiologists as they will be able to classify the diseases and it will help in improving and enhancing different medical techniques.
An Enhancer Demethylator Phenotype Converged to Immune Dysfunction and Resistance to Immune Checkpoint Inhibitors in Clear-Cell Renal Cell Carcinomas
Authors: Lu Xiaofan , Vano Yann , Helleux Alexandra , Su Xiaoping , Lindner Véronique , Davidson Guillaume , Mouawad Roger , Spano Jean-Philippe , Rouprêt Morgan , Elaidi Reza , Compérat Eva , Verkarre Virginie , Sun Chengming , Chevreau Christine , Bennamoun Mostefa , Lang Hervé , Tricard Thibault , Cheng Wenxuan , Xu Li , Davidson Irwin , Yan Fangrong , Fridman Wolf Herman , Sautes-Fridman Catherine , Oudard Stéphane , Malouf Gabriel G.
Published: November 2022

Purpose:

Immune checkpoint inhibitors (ICI) have revolutionized the treatment of patients with clear-cell renal cell carcinomas (ccRCC). Although analyses of transcriptome, genetic alterations, and the tumor microenvironment (TME) have shed light into mechanisms of response and resistance to these agents, the role of epigenetic alterations in this process remains fully unknown.

Experimental Design:

We investigated the methylome of six ccRCC cohorts as well as one cell line dataset. Of note, we took advantage of the BIONIKK trial aiming to tailor treatments according to Paris Descartes 4-gene expression subgroups, and performed Illumina EPIC profiling for 46 samples related to patients treated with ipilimumab plus nivolumab, and 17 samples related to patients treated with sunitinib.

Results:

A group of tumors associated with enhancer demethylation was discovered, namely TED. TED was associated with tumors with sarcomatoid differentiation and poor clinical outcome. TED harbored TET1 promoter demethylation, activated the gene expression signature of epithelial–mesenchymal transition and IL6/JAK/STAT3 pathways, and displayed a TME characterized by both immune activation and suppressive populations, fibroblast infiltration, and endothelial depletion. In addition, TED was a predictive factor of resistance to the combination of first-line ipilimumab-nivolumab in the BIONIKK clinical trial. Finally, TED was associated with activation of specific regulons, which we also found to be predictive of resistance to immunotherapy in an independent cohort.

Conclusions:

We report on the discovery of a novel epigenetic phenotype associated with resistance to ICIs that may pave the way to better personalizing patients’ treatments.

See related commentary by Zhou and Kim, p. 1170
Comparing the Working Memory Capacity with Cognitive Flexibility, Cognitive Emotion Regulation, and Learning Styles of University Students: A Domain General View
Authors: Guler Kahraman , Aydin Aylin
Published: November 2022

Introduction: The goal of this study was to investigate the relationship of working memory capacity with the use of cognitive emotion regulation strategies, cognitive flexibility level, and learning styles of university students. Methods: In the present study (N = 39), the participants completed the Emotion Regulation Questionnaire, Cognitive Flexibility Scale, Kolb Learning Styles Inventory, and Vermunt Learning Styles Inventory and three WM capacity (WMC) tasks that are Rotation Span Task, Operation Span Task, and Symmetry Span Task. Their WMCs were assessed, and the relationship of it was compared with cognitive emotion regulation, cognitive flexibility, and learning styles. Results: The results indicated that there is a significant difference and negative correlation (r= −0.341) between Operation Span Task and refocus on planning. Findings of the research indicated correlations between emotion regulation strategies and between cognitive flexibility and two emotion regulation strategies that are refocus on planning (r = 0.528) and positive reappraisal (r = 0.574). Only one learning style that is Processing Strategies in Vermunt Learning Style Inventory was found significantly different in terms of the cognitive flexibility level of the participants. The results also indicated a positive correlation between verbal and spatial WM tasks which support the domain general view for WMC. Conclusions: Further studies are advised to be conducted between cognitive emotion regulation strategies and working memory capacity as these findings may have significant implications for understanding the correlation between memory and emotion.
Development of Novel Tasks to Assess Outcome-Specific and General Pavlovian-to-Instrumental Transfer in Humans
Authors: Belanger Matthew J. , Chen Hao , Hentschel Angela , Garbusow Maria , Ebrahimi Claudia , Knorr Felix G. , Zech Hilmar G. , Pilhatsch Maximilian , Heinz Andreas , Smolka Michael N.
Published: November 2022

Introduction: The emergence of Pavlovian-to-instrumental transfer (PIT) research in the human neurobehavioral domain has been met with increased interest over the past two decades. A variety of PIT tasks were developed during this time; while successful in demonstrating transfer phenomena, existing tasks have limitations that should be addressed. Herein, we introduce two PIT paradigms designed to assess outcome-specific and general PIT within the context of addiction. Materials and Methods: The single-lever PIT task, based on an established paradigm, replaced button presses with joystick motion to better assess avoidance behavior. The full transfer task uses alcohol and nonalcohol rewards associated with Pavlovian cues and instrumental responses, along with other gustatory and monetary rewards. We constructed mixed-effects models with the addition of other statistical analyses as needed to interpret various behavioral measures. Results: Single-lever PIT: both versions were successful in eliciting a PIT effect (joystick: p < 0.001, ηp2 = 0.36, button-box: p < 0.001, ηp2 = 0.30). Full transfer task: it was determined that the alcohol and nonalcoholic reward cues selectively primed their respective reward-associated responses (gustatory version: p < 0.001, r = 0.59, and monetary version: p < 0.001, r = 0.84). The appetitive/aversive cues resulted in a general transfer effect (gustatory: p < 0.001, ηp2 = 0.09, and monetary: p < 0.001, ηp2 = 0.17). Discussion/Conclusion: Single-lever PIT: PIT was observed in both task versions. We posit that the use of a joystick is more advantageous for the analysis of avoidance behavior. It evenly distributes movement between approach and avoid trials, which is relevant to analyzing fMRI data. Full transfer task: While gustatory conditioning has been used in the past to elicit transfer effects, we present the first paradigm that successfully elicits both specific and general transfers in humans with gustatory alcohol rewards.
Audit of Asthma Exacerbation Management in a Swiss General Hospital
Authors: Schnyder Dominik , Lüthi-Corridori Giorgia , Leuppi-Taegtmeyer Anne Barbara , Boesing Maria , Geigy Nicolas , Leuppi Joerg Daniel
Published: November 2022

Background: Adequate management is crucial to reduce symptoms, hospitalization, and relapses in patients with asthma. Hospitals often struggle to meet treatment guidelines, and no recent data for Switzerland are available. Objectives: The aim of the study was to audit the asthma exacerbation management in the Cantonal Hospital of Baselland in order to evaluate the level of compliance with guidelines in a narrative discussion. Method: The study design is a retrospective observational cohort study. We evaluated all adult patients presenting to the hospital with a physician-diagnosed asthma exacerbation in 2018 and 2019. The asthma management patients received was compared to the Swiss guidelines and the international GINA guidelines. Results: 160 patients were included (mean age: 50 years old, 57.5% female). SpO2 and heart rate were assessed at presentation in nearly all patients. Peak expiratory flow (PEF) was measured in only 14%. Adequate management of asthma exacerbation with inhaled bronchodilator medication in a combination of short-acting beta-agonists and short-acting anticholinergics was administered to 96% of the patients. Patients with severe symptoms received systemic glucocorticosteroids within 6 h in 55%. At discharge, a reliever medication was prescribed for 64% of the patients and 55% received a new or increased controller therapy with inhaled glucocorticosteroid (ICS). 49% of the patients had no follow-up organized. Conclusion: To increase the guideline conformity and quality of asthma exacerbation management, the severity should be better assessed, especially by routinely performing PEF measurements. Treatment needs to be intensified; in particular, the ICS dose should be increased significantly and systemic glucocorticosteroids should be given with a lower threshold.
Leitlinie zu Evaluation und Management von pulmonalen Manifestationen bei Sjögren-Syndrom
Authors: Kahn Nicolas C.
Published: November 2022

Background: Pulmonary disease is a potentially serious yet underdiagnosed complication of Sjögren’s syndrome, the second most common autoimmune rheumatic disease. Approximately 16% of patients with Sjögren’s demonstrate pulmonary involvement with higher mortality and lower quality of life. Research question: Clinical practice guidelines for pulmonary manifestations of Sjögren’s were developed by the Sjögren’s Foundation after identifying a critical need for early diagnosis and improved quality and consistency of care. Study design: and methods: A rigorous and transparent method­ology was followed according to American College of Rheumatology guidelines. The Pulmonary Topic Review Group (TRG) developed clinical questions in the PICO (Patient, Intervention, Comparison, Outcome) format and selected literature search parameters. Each article was reviewed by a minimum of two TRG members for eligibility and assessment of quality of evidence and strength of recommendation. Guidelines were then drafted based on available evidence, expert opinion, and clinical importance. Draft recommendations with a clinical rationale and data extraction tables were submitted to a Consensus Expert Panel for consideration and approval, with at least 75% agreement required for individual recommendations to be included in the final version. Results: The literature search revealed 1,192 articles, of which 150 qualified for consideration in guideline development. Of the original 85 PICO questions posed by the TRG, 52 recommendations were generated. These were then reviewed by the Consensus Expert Panel and 52 recommendations were finalized, with a mean agreement of 97.71% (range, 79%-100%). The recommendations span topics of evaluating Sjögren’s patients for pulmonary manifestations and assessing, managing, and treating upper and lower airway disease, interstitial lung disease, and lymphoproliferative disease. Interpretation: Clinical practice guidelines for pulmonary manifestations in Sjögren’s will improve early identification, evaluation, and uniformity of care by primary care physicians, rheumatologists, and pulmonologists. Additionally, opportunities for future research are identified.
Autoimmune Liver Disease in Human Immunodeficiency Virus-Infected Patients: 3 Case Series
Authors: Rafael Maria Ana , Bordalo Ferreira Filipa , Theias Manso Rita , Peruzzu Francesca , Cardoso Mariana
Published: November 2022

We present 3 cases of autoimmune liver disease in human immunodeficiency virus (HIV)-infected patients and describe the different diagnostic and therapeutic approaches used in each case. The first patient was diagnosed with primary biliary cholangitis (PBC) with features of autoimmune hepatitis (AIH), requiring second-line therapy due to incomplete response to ursodeoxycholic acid. The second patient was diagnosed with AIH with features of PBC and had the particular challenges of presenting with advanced liver fibrosis and having a past history of disseminated cytomegalovirus infection. The last case concerns an AIH with acute liver injury, successfully treated with corticosteroids and azathioprine. Recently, the number of patients on antiretroviral therapy (ART) for HIV disease has increased significantly. Therefore, more patients with this chronic infection have been diagnosed with autoimmune diseases, leading to concerns regarding immunosuppressive therapies in this population. With these cases, we alert for these increasingly incident diseases and support the safety of immunosuppressive therapies, provided that HIV is suppressed with ART.
High-Frequency Deep Sclerotomy as Adjunctive Therapy in Open-Angle Glaucoma Patients
Authors: Kontic Marko , Todorovic Dusan , Zecevic Rada , Vulovic Tatjana Sarenac
Published: November 2022

Introduction: Glaucoma is a chronic, progressive disease of the optic nerve that can cause vision loss and blindness. High-frequency deep sclerotomy (HFDS) is a novel ab interno procedure used to lower intraocular pressure (IOP). This study aims at examining hypotensive effects of HFDS in patients with medically uncontrolled primary open-angle glaucoma (POAG). Methods: 23 patients (23 eyes) participated in this study. They were all affected by POAG and used maximum prescribed medical therapy. It is important to note that the target IOP was not detected in any study participant. The target IOP is the highest IOP value at which no new damages of the optic nerve occur. The procedure was performed with a custom-made, high-frequency dissection probe by applying bipolar current of 500 kHz. Six pockets (1 mm deep, 0.3 mm high, and 6 mm wide) were made ab interno in nasal sclera (through trabecular meshwork and Schlemm’s canal). Tobramycin/dexamethasone and pilocarpine eye drops were administered postoperatively for a month. Results: The mean value of the base IOP had been 25.6 mm Hg before the procedure. Significant complications were not recorded either during the surgery or in a postoperative follow-up period. The average IOP for our patients reduced by 8.6 mm Hg (33.6%) after a year. The mean value of the instilled anti-glaucoma eye drops had been 2.78 (SD = 0.45) before the HFDS and 0.61 (SD = 1.04) at the end of the research. The target IOP was not achieved in five cases (21.7% of our sample). Discussion/Conclusion: This study presents the data on our first surgical experience with HFDS that was conducted on 23 patients who had medically uncontrolled POAG. The results indicate that HFDS is safe and efficient in reducing IOP.
TBX20 Improves Contractility and Mitochondrial Function During Direct Human Cardiac Reprogramming
Authors: Tang Yawen , Aryal Sajesan , Geng Xiaoxiao , Zhou Xinyue , Fast Vladimir G. , Zhang Jianyi , Lu Rui , Zhou Yang
Published: November 2022

Background:

Direct cardiac reprogramming of fibroblasts into cardiomyocytes has emerged as a promising strategy to remuscularize injured myocardium. However, it is insufficient to generate functional induced cardiomyocytes from human fibroblasts using conventional reprogramming cocktails, and the underlying molecular mechanisms are not well studied.

Methods:

To discover potential missing factors for human direct reprogramming, we performed transcriptomic comparison between human induced cardiomyocytes and functional cardiomyocytes.

Results:

We identified
TBX20
(T-box transcription factor 20) as the top cardiac gene that is unable to be activated by the MGT133 reprogramming cocktail (
MEF2C
,
GATA4
,
TBX5
, and
miR-133
). TBX20 is required for normal heart development and cardiac function in adult cardiomyocytes, yet its role in cardiac reprogramming remains undefined. We show that the addition of TBX20 to the MGT133 cocktail (MGT+TBX20) promotes cardiac reprogramming and activates genes associated with cardiac contractility, maturation, and ventricular heart. Human induced cardiomyocytes produced with MGT+TBX20 demonstrated more frequent beating, calcium oscillation, and higher energy metabolism as evidenced by increased mitochondria numbers and mitochondrial respiration. Mechanistically, comprehensive transcriptomic, chromatin occupancy, and epigenomic studies revealed that TBX20 colocalizes with MGT reprogramming factors at cardiac gene enhancers associated with heart contraction, promotes chromatin binding and co-occupancy of MGT factors at these loci, and synergizes with MGT for more robust activation of target gene transcription.

Conclusions:

TBX20 consolidates MGT cardiac reprogramming factors to activate cardiac enhancers to promote cardiac cell fate conversion. Human induced cardiomyocytes generated with TBX20 showed enhanced cardiac function in contractility and mitochondrial respiration.
Abstract GS101: Myosin Binding Protein H-like Competes With Cardiac Myosin Binding Protein-c To Prolong Atrial Contraction
Authors: Araujo Kelly , Woulfe Kc , Wittenkeller Lucas , Barefield David Y
Published: November 2022

Myosin binding protein-H like (MyBP-HL) has recently been identified as a novel component of myofilaments in the cardiac atria. Loss of MyBP-HL is associated with dilated cardiomyopathy and arrhythmia in humans and mice. MyBP-HL is highly homologous to cardiac myosin binding protein-C (cMyBP-C), which has been studied in the context of genetic cardiomyopathy. Prior work using mouse atrial tissue lysates established MyBP-HL and cMyBP-C bind competitively and maintain a stoichiometric ratio compared to myosin heavy chain. We now confirm this competitive binding using neonatal rat ventricular cardiomyocytes (NRVMs) transfected with mouse
Mybphl
. Using confocal microscopy with internal controls, we found a linear, inverse relationshib between myosin binding protein levels with NRVMs with high levels of MyBP-HL expression showing a reduction in cMyBP-C levels compared to untransfected NRVMs that only express cMyBP-C. While we show that these two myosin binding proteins bind comparatively with each other, the functional consequences are unclear. We performed force-calcium measurements of permeabilized single atrial cardiomyocytes and found that loss of MyBP-HL does not significantly alter the calcium sensitivity of force development, or the specific force generated by isometric contraction. Because cMyBP-C is a known regulator of crossbridge sliding velocity, we evaluated contraction and relaxation kinetics of
Mybphl
single-myofibrils from WT and
Mybphl
homozygous null mice. Atrial myofibrils lacking
Mybphl
showed a significantly faster rate and shorter duration of their linear phase of relaxation, which is governed by the off-rate of myosin from actin. This suggests that extra cMyBP-C in the atria can hasten myofibril relaxation. In healthy atria with MyBP-HL replacing approximately half the cMyBP-C, the slow phase of relaxation is prolonged. While the physiological importance of this is not explored, it provides a mechanism by which loss of function mutations in
MYBPHL
cause contractile dysfunction.
Abstract GS106: Asporin, An Extracellular Matrix Protein, Is A Beneficial Regulator Of Cardiac Remodeling
Authors: Rai Deepika , Huang Chengqun , Thakur Reetu , Sharma Ankush , Gottlieb Roberta A , Piplani Honit
Published: November 2022

Heart failure is accompanied by adverse cardiac remodeling involving extracellular matrix (ECM). Cardiac ECM acts as a major reservoir for many proteins including growth factors, cytokines, collagens, and proteoglycans. Activated fibroblasts during cardiac injury can alter the composition and activity of these ECM proteins. Through unbiased analysis of a microarray dataset of human heart tissue comparing normal hearts (n=135) to hearts with ischemic cardiomyopathy (n=94), we identified Asporin (ASPN) as the top differentially regulated gene (DEG) in ischemic cardiomyopathy; its gene-ontology terms relate closely to fibrosis and cell death. ASPN is a Class I small leucine repeat protein member implicated in cancer, osteoarthritis, and periodontal ligament mineralization. However, its role in cardiac remodeling is still unknown. Here, we initially confirmed our big dataset analysis through cells, mice, and clinical atrial biopsy samples to demonstrate increased ASPN expression after pressure overload or cardiac ischemia/reperfusion injury. We tested the hypothesis that ASPN, being a TGFβ1 inhibitor, can attenuate fibrosis in mouse models of cardiac injury. We found that ASPN is released by cardiac fibroblasts and attenuates TGFβ signaling. Moreover, ASPN
-/-
mice displayed increased fibrosis and decreased cardiac function after pressure overload by transverse aortic constriction (TAC) in mice. In addition, ASPN protected cardiomyocytes from hypoxia/reoxygenation-induced cell death and regulated autophagy and mitochondrial bioenergetics in cardiomyocytes. Increased infarct size after ischemia/reperfusion injury in ASPN
-/-
mice confirmed ASPN’s contribution to cardiomyocyte viability. Echocardiography revealed a greater reduction in left ventricular systolic function post-I/R in the ASPN
-/-
animals compared to wild type. Furthermore, we developed an ASPN-mimic peptide using molecular modeling and docking which when administered to mice prevented TAC-induced fibrosis and preserved heart function. The peptide also reduced infarct size after I/R in mice, demonstrating the translational potential of ASPN-based therapy. Thus, we establish the role of ASPN as a critical ECM molecule that regulates cardiac remodeling to preserve heart function.
Abstract GS111: Inhibition Of Cardiac Glucose Transporter 1 Suppresses Early Glucose Dependency And Klf5 Activation And Treats Cardiomyopathy In Diabetes
Authors: Mia Sobuj , Kyriazis Ioannis D , Sidiropoulou Rafailia , Hill Daniel , Mantalaris Athanasios , Drosatos Konstantinos
Published: November 2022

Healthy hearts use more fatty acids (FA) than glucose for ATP synthesis but diabetic cardiomyopathy (DbCM) occurs with higher FA dependency. It remains controversial whether glucotoxicity or lipotoxicity or both account for DbCM. We recently discovered that insulin signaling inhibition and eventual FOXO1 activation stimulate cardiac KLF5 expression, which drives lipotoxicity and causes cardiac dysfunction. In the present study, we investigated the relative contribution of glucose in the activation of cardiac KLF5 and DbCM. We induced Type-1 diabetes (T1D) in C57BL/6 mice via intraperitoneal injections of streptozotocin (STZ). In contrast to late-stage diabetes (12 weeks post-STZ), cardiac KLF5 mRNA and protein levels were not increased in the early T1D stage (4 weeks post-STZ) although mice have mild cardiac dysfunction already. Seahorse analysis in adult cardiomyocytes isolated from mice with early T1D showed higher glucose and lower FA dependency compared to non-diabetic mice and mice in late T1D. To confirm whether hyperglycemia causes cardiac dysfunction, we treated diabetic mice with Dapagliflozin (DAPA, SGLT2 inhibitor) or STF-31, a GLUT1 inhibitor. These treatments restored normal dependency on fatty acids and prevented cardiac dysfunction. GC-MS analysis showed that the reversal of fuel dependency from glucose to fatty acids in late T1D is accompanied by increased glucose content opposite to the early T1D. Accordingly cardiac KLF5 is increased in late T1D, accompanied by severe cardiac dysfunction. The expression changes of KLF5 are mirrored by transcriptional activity of FOXO1 -shown by expression of FOXO1 targets- in early and late T1D. The changes in transcriptional activity are accompanied by differential FOXO1 acetylation, which is controlled by Sirtuin-1 and modulates its DNA affinity. Analysis of mouse cardiac tissue in early T1D and a human cardiomyocyte cell line (AC16) that was treated with high glucose showed higher Sirtuin-1 expression and stronger protein-protein interaction with FOXO1. To this end, mice that were subjected to treatment with either DAPA or STF31 for 12 Wks had improved cardiac function, lower cardiac KLF5 expression and decreased expression of cardiac KLF5 gene targets. Interestingly, GLUT1 mRNA levels were increased in late T1D compared to early T1D. Cardiomyocyte-specific KLF5 overexpression or adenovirus-mediated KLF5 overexpression in AC16 cells stimulated GLUT1 expression. Collectively, in early T1D, hearts rely more on glucose utilization in mitochondria. In late T1D, SIRT1-FOXO1-KLF5 axis causes lipotoxicity and subsequent induction of GLUT1 expression that contributes to glucotoxicity. Inhibition of GLUT1-dependent glucose uptake alleviates diabetic cardiomyopathy via inhibition of both early glucose dependency and late KLF5 activation.
Abstract OEC101: Cardiac Reprogramming Inducer ZNF281 Is Indispensable For Heart Development By Interacting With Key Cardiac Transcriptional Factors
Authors: Hashimoto Hisayuki , Komuro Jin , Nakamura Takahiro , Yuasa Shinsuke , Fukuda Keiichi
Published: November 2022

During direct cardiac reprogramming, cardiogenic transcription factors (TFs) cooperatively reshape the epigenomic landscape by activating cardiac enhancers, which describe a common epigenetic signature between
in vitro
cardiac reprogramming and
in vivo
heart development. We thus hypothesized that direct reprogramming could be a useful tool to study novel transcriptional regulators in heart development. To test this hypothesis, we focused on ZNF281/Zfp281(Zinc finger protein 281), a pluripotency regulator which showed up as an unexpected strong cardiac reprogramming inducer from our previous unbiased screen. Since Zfp281 is an essential factor for epiblast maturation, we established a Zfp281 floxed mouse line using CRISPR/Cas9 system to study the impact of Zfp281 after cardiac lineage commitment. We then generated multiple conditional Zfp281KO (cKO) mouse lines by crossing with different stage-specific Cre driver lines. Surprisingly, we found that mesodermal (Mesp1-Cre) Zfp281cKO mice were embryonic lethal as early as E9.5, associated with various cardiac anomalies. In contrast, cardiac progenitor (Nkx2-5-Cre) and cardiomyocyte (Myh6-Cre) Zfp281cKO mice survived to adulthood and appeared grossly normal, suggesting a crucial role of Zfp281 in mesodermal to cardiac differentiation. Next, we performed RNA-seq analysis to study the molecular mechanism of Zfp281 deficient cardiac anomalies. Interestingly, expression of cardiac TFs and sarcomeric genes were altered in Zfp281cKO embryos compared to control siblings. Consistently, co-immunoprecipitation analysis showed direct interaction of Zfp281 with Mesp1 and Gata4. These data indicate that Zfp281 directly interacts with cardiac TFs to cooperatively orchestrate the cardiogenic transcriptional network. In conclusion, by using multiple genetic mouse models, we discovered that cardiac reprogramming inducer Zfp281 plays a pivotal role in heart development. Although GWAS studies have identified various disease-causing mutations, the etiology for the majority of congenital heart disease remains largely unknown and there are still such missing pieces to be uncovered in the interdependent cardiogenic transcriptional network. Our study demonstrates and highlights the impact and possibility of direct reprogramming as a novel approach to study cardiovascular biology.
Abstract OEC102: Unbiased Discovery Of Allograft Inflammatory Factor 1 As A New And Critical Immunometabolic Regulatory Node During Cardiac Injury
Authors: DeBerge Matthew , Thorp Edward B
Published: November 2022

Introduction:
Myocardial infarction (MI) mobilizes macrophages, which are the central protagonists of tissue repair in the infarcted heart. While macrophages are necessary for repair after MI, they can also contribute to adverse remodeling and progression to heart failure. We sought to identify novel mediators of maladaptive macrophage responses by performing single cell RNA sequencing of a patient with ischemic cardiomyopathy. This led to the unbiased identification of
Allograft inflammatory factor 1
(
AIF1
) as the most significant differentially expressed gene in macrophages compared to other cardiac cells during human heart failure.

Hypothesis:
Since
AIF1
was specifically expressed in macrophages during ischemic heart failure, we next tested the hypothesis that AIF1 promoted maladaptive macrophage responses after MI.

Methods and Results:
Using autopsy samples from human acute MI patients or a murine model of permanent occlusion MI, we found that AIF1 was increased in macrophages, but not neutrophils or monocytes, after MI. To test the functional significance of AIF1 expression in macrophages after MI, we generated bone marrow chimeric mice where
Aif1
-deficiency was restricted to only cells of hematopoietic origin. Despite comparable levels of surgically induced injury, loss of
Aif1
was associated with increased proreparative macrophages, smaller infarct sizes, and preservation of cardiac function compared to wild-type controls. Therapeutic targeting of
Aif1
with anti-sense oligonucleotides also conferred cardioprotective responses after MI. Mechanistically, AIF1 promoted actin polymerization in bone marrow-derived macrophages after TLR4 stimulation leading to increased glycolytic metabolism and inflammatory IL-6 and TNF-α production. AIF1 also suppressed calcium uptake in the mitochondria to limit mitochondrial metabolism and anti-inflammatory IL-10 production.

Conclusion:
These data newly reveal a proinflammatory role for AIF1 in the infarcted heart through inflammatory glycolytic reprogramming of macrophages, which required both calcium mobilization and actin polymerization. Targeted knockdown of
AIF1
led to cardioprotection supporting further exploration of macrophage AIF1 as a therapeutic target after MI.
Abstract P1001: IGFBP2 Overcomes Cell Contact-mediated Proliferation Inhibition In Human IPSC-derived Cardiomyocytes
Authors: Lee Soah , Heinrich Paul , Lee Daniel , Goodyer William R , Galdos Francisco X , Zhu Han , Samad Tahmina , Xu Sidra , Lee Carissa , Duan Lauren , Wu Sean M
Published: November 2022

Introduction and Hypothesis:
Human induced pluripotent stem cells (hiPSCs) and their derivatives provide a promising cellular source for regenerative cardiac therapies. However, generating large-scale hiPSC-derived cardiomyocytes (hiPSC-CMs) remains a major hurdle, caused, in part, by cell contact-driven proliferation inhibition in dense culture conditions. We hypothesize that differentially-secreted factors in sparsely vs densely-cultured hiPSC-CMs are responsible for the increased proliferation rate in sparsely cultured hiPSC-CMs.

Methods:
We performed a high-throughput screening (HTS) assay to identify growth factors (GFs) released in the supernatant of sparsely- and densely-cultured hiPSC-CMs. Differential expression of candidate GFs in dense vs sparse culture conditions was confirmed by single cell RNA sequencing (scRNAseq), Western Blot (WB), and Immunocytochemistry (ICC) analyses. The confirmed GF hits were further validated by measuring the expression of the cell cycle marker Ki67 in hiPSC-CMs after GF treatment. The GF treatment effects on cell-cell contact was further explored mechanistically using siRNA-based N-cadherin (CDH2) knockdown (KD) model in vitro.

Results:
Our HTS assay identified IGFBP2, IGFBP6, PDGF-AA as top candidate GFs in the sparse culture supernatant. Subsequent WB, ICC, scRNAseq analyses confirmed 1.5, 3.5, 2.0-fold higher IGFBP2 expression in the sparse condition compared to the dense condition. Supplementation of recombinant IGFBP2 to densely cultured hiPSC-CMs showed a dosage-dependent increase in Ki67 expression level from 13.0 +/- 1.4% in control to 44.1 +/- 8.4% after 3nM IGFBP2 treatment, a level comparable to that achieved in sparse culture. Mechanistically, IGFBP2 treatment resulted in a shift of CDH2 localization from cell-cell junction to the cytoplasm. Reducing CDH2 expression by CDH2-KD to mimic loss of cell-cell contact promoted hiPSC-CM proliferation.

Conclusion:
We discovered an unexpected role of IGFBP2 to overcome cell contact-mediated inhibition of hiPSC-CM proliferation which provides a promising new approach to engineer in situ large-scale 3D cardiac tissues for regenerative applications.
Abstract P1002: Elucidating The Role Of Dyrk1a Kinase In Controlling Cardiomyocyte Proliferation
Authors: Rizzari Giorgia , Dirkx Ellen , Torrini Consuelo , Tomczyk Mateusz , Mano Miguel , Giacca Mauro
Published: November 2022

Despite the development of new pharmacological and device-based therapies, no approaches currently exist to compensate for the irreversible loss of cardiomyocytes occurring after myocardial infarction. In a search for novel treatments that induce cardiomyocyte proliferation, we performed a high-content, fluorescence-microscopy-based, high-throughput screening in neonatal rat cardiomyocytes, using a library of FDA/EMA-approved drug library compounds. The screening, which was based on EdU incorporation as an S-phase marker, identified harmine as the most powerful drug enhancing proliferation in combination with miR-199a-3p and miR-590-3p, the most effective microRNAs that the laboratory had previously identified. Harmine treatment of CD1 mice showed an improved cardiac outcome upon harmine treatment compared to controls. Harmine is a potent inhibitor of the Dyrk1 family of protein kinases, with high specificity for the Dyrk1a isoform. Dyrk1a downregulation experiments in neonatal rat ventricular cardiomyocytes showed increased proliferation, consistent with a negative role of this kinase. The simultaneous knockdown of YAP1 blunted the effect of Dyrk1a downregulation. Of note, the YAP total protein levels were unchanged upon either Dyrk1a silencing or overexpression, but there was a marked modification of YAP subcellular localisation, being YAP1 nuclear upon knockdown and mostly cytoplasmic upon overexpression. Dyrk1a downregulation and harmine treatment activated YAP-controlled gene expression. In the course of these experiments, we also noticed a reciprocal regulation of YAP onto DYRK1a, as YAP downregulation markedly decreased Dyrk1a protein levels. This finding was reinforced by the observations that, in cardiomyocytes, the YAP-TEAD inhibitor verteporfin also decreased Dyrk1a protein levels. Of note, this downregulation was not rescued by the proteasome inhibitor MG132, thus indicating that the level of regulation is most likely transcriptional. These results support the role of Dyrk1a as an anti-proliferative factor in cardiomyocytes and highlight a novel feedback connection between Dyrk1a and YAP.