bluuebunny/crossref_metadata_2025_split huggingface dataset:
Mutation in PDGFRbeta: a potential new pathogenic variant for mitral valve prolapse
Authors: Wu H W , Van Wijngaarden A L , Mousavi Gourabi B , Smits A M , Barge-Schaapveld D Q C M , De Vries A A F , Jukema J W , Kruithof B P T , Ajmone Marsan N 
Published: November 2023

Abstract

Background
Mitral valve prolapse (MVP) due to myxomatous degeneration is characterized by familial clustering, but few pathogenic genes have been identified. In a genetic screening program for MVP patients, we identified a platelet-derived growth factor receptor β (PDGFRβ)-E162K missense variant in a family with a bi-leaflet MVP phenotype in four affected family members. PDGFRβ is a receptor tyrosine kinase, which plays an important role in vascular development.

Purpose
In this study, we investigated whether the E162K substitution in PDGFRβ could lead to mitral valve abnormalities.

Methods
To assess the functional effects of the missense variant, proliferation and migration assays were performed using HeLa cells overexpressing wild-type (WT) or mutant PDGFRβ. Immunostainings were performed to determine PDGFRβ expression in the diseased valves from a family member undergoing surgical operation and from additional patients operated for MVP. Mouse models were created with the mouse equivalent of the human PDGFRβ-E162K variant (PDGFRβ-E161K) using CRISPR/CAS9. The variant was not lethal and a total of 12 homozygous PDGFRβ-E161K-/- mice, 11 heterozygous PDGFRβ-E161K+/- mice, and 7 WT mice were subjected to transthoracic echocardiography. In addition, histology was performed in all 30 mice to detect potential valvular defects.

Results
Stimulated HeLa cells expressing PDGFRβ-E162K showed reduced proliferation and migration, suggesting a loss of function of the mutant receptor. PDGFRβ was expressed in human diseased mitral valves. Echocardiographic evaluation revealed a significant larger mitral valve annulus diameter in both PDGFRβ-E161K+/- and PDGFRβ-E161K-/- compared to WT (WT: 1.97 mm; PDGFRβ-E161K+/-:2.13, p=0.031; PDGFRβ-E161K-/-: 2.42 mm, p&lt;0.0001). Histological data in PDGFRβ-E161K-/- mice showed significant leaflet thickening in the free edge (FE) of the posterior mitral valve leaflet (PMVL) compared to WT (WT: 72.34 ɥm; PDGFRβ-E161K-/-: 109.6 ɥm, p=0.040) and a larger leaflet area of the PMVL compared to WT (WT: 25280 ɥm²; PDGFRβ-E161K-/-: 44494 ɥm², p=0.020). Moreover, an altered organization of the extracellular matrix reminiscent of the myxomatous degeneration in MVP was observed.

Conclusions
The PDGFRβ-E162K variant is associated with familial MVP and alters the function of PDGFRβ. Mice harboring this mutation display mitral valve defects and can be used as a model for mitral valve disease.
Aortic valve calcification is promoted by clonal hematopoiesis associated with DNMT3A and TET2 gene driver mutations
Authors: Abplanalp W , Schuhmacher B , Mas-Peiro S , Zuriaga M A , Matesanz N , Fuster J J , John D , Muhly-Reinholz M , Vasa-Nicotera M , Merten M , Hoffmann J , Dimmeler S , Zeiher A M 
Published: November 2023

Abstract

Background
Clonal hematopoiesis (CH) due to DNMT3A or TET2-driver mutations is associated with coronary heart disease and worse prognosis among patients with aortic valve stenosis (AVS) and inflammation. However, it is unknown what role CH plays in the pathogenesis of AVS. AVS is the most common age-related heart valve disease, with no medical therapy to halt progression.

Methods and Results
Single-cell RNA-sequencing of immune cells of CAVD patients (n=13) with and without the most prevalent CH-mutations (DNMT3A and TET2), with mean CH-driver gene variant allele frequency was 6.5% for DNMT3A and 17.9% for TET2. Monocytes of patients harboring DNMT3A and TET2 mutations shared 836 upregulated genes, many associated with glycolytic, pro-inflammatory activation (CXCL10, IL38) and paracrine pro-calcific factors (oncostatin M (OSM), S100A9), suggesting that CH might be causally involved in AVS. Indeed, silencing of TET2 or DNMT3A in macrophages promotes M1-like polarization and increases release of pro-calcific mediators, particularly S100A9 and OSM. To assess whether CH-gene silenced macrophages may stimulate mesenchymal cells to secrete calcium, secreted factors from TET2 or DNMT3A-silenced macrophages were applied mesenchymal cells. Indeed, CH-gene silenced secretomes induced osteoblastic differentiation of mesenchymal cells, evidenced by increased Alizarin red calcium staining and elevated levels of ALP and RUNX2. This CH-mediated enhancement in osteoblastic conversion could be ablated by silencing of OSM. Finally, valves from atheroprone Ldlr-/- mice receiving TET2-/- bone marrow transplants demonstrated increased total calcified area (1.49-fold) along with increased numbers of calcification deposits (2.33-fold) as evidenced by von Kossa staining, along with enhanced OSM and S100A9 levels.

Conclusion
This is the first study to demonstrate that somatic CH-driver mutations in monocytes may accelerate AVS. Moreover, calcification was ablated via silencing of OSM in CH-gene silenced macrophages. These data suggest Oncostatin M may be a therapeutically promising target in halting the progression of AVS in patients harboring CH mutations.
The senescence-associated secretome for prediction of mortality in patients with aortic valve stenosis undergoing TAVR
Authors: Mas-Peiro S , Rasper T , Berkowitsch A , Dimmeler S , Zeiher A M 
Published: November 2023

Abstract

Background
Cellular senescence is an essential mechanism in the process of aging. The senescence-associated secretory phenotype (SASP) includes mainly pro-inflammatory cytokines, proteases, soluble signalling factors, and growth modulators. Degenerative aortic valve stenosis (AVS) is the prototypical inflammatory age-related cardiovascular disease.

Purpose
Our aim was to assess whether circulating concentrations of SASP factors are associated with prognosis in elderly patients with AVS undergoing transcatheter aortic valve replacement (TAVR), the current standard therapy for severe cases.

Methods
A total of 24 SASPs were measured using the Luminex Multiplex ELISA Assay (Biotechene, GmbH) in 120 AVS patients undergoing TAVR between February 2017 and February 2020 with a prospective follow-up of up to 4 years. Receiver operating characteristic (ROC) curves were created for each SASP protein in order to identify predictive markers. SASPs were considered positive if their baseline value was above the cut-off value derived from ROC-curves. The only 3 independent predictive factors were selected to develop an easy-to-calculate score based on the number of positive SASP proteins (0 to 3). Primary endpoint was long-term all-cause mortality.

Results
Median (IQR) age of the population was 83.2 (78.9-87.0) years, 56.7% were women. According to ROC curves, 9 out of 24 SASPs showed a significant area under the curve (AUC), with values above 0.623 (see Table). Only 3 of the markers reflecting potentially different underlying mechanisms remained independent predictors, after including all SASP factors in a Cox Regression analysis: GDF-15, ICAM-1 and Osteoprotegerin (all, p&lt;0.03). A score based on the number of baseline factors being above the AUC-derived threshold level showed an excellent discrimination ability to predict survival, with long-term all-cause mortality substantially increasing for each additional positive marker (see Kaplan-Meier curves).

Conclusion
In conclusion, the presence of elevated baseline levels for 0, 1, 2, or 3 of the independent SASP factors reflecting different underlying pathophysiological mechanisms predicts increased long-term all-cause mortality with profoundly improved discrimination power in AVS patients undergoing TAVR. If confirmed in further prospective studies, a combination of these 3 SASP markers could serve as a highly useful risk stratification tool in the elderly population with AVS.
Changes in the immune cell landscape in the infarcted mouse heart
Authors: Mitre L , Burkard A , Hansen S , Li X , Ehret F , Heitlinger S , Mwinyella T , Marchini T , Olawale T , Gissler M , Horstmann H , Wolf D , Westermann D 
Published: November 2023

Abstract

Background
Myocardial Infarction (MI) and Ischemic Heart Failure represent the leading causes of death globally. Myocardial remodeling after MI is accompanied and driven by a strong immune response with an accumulation of several, partially opposing immune cell types. The temporal and spatial dynamics of this cardiac specific immune cell infiltration, however, have not been systematically evaluated.

Purpose
We characterized the dynamics of the immune cell repertoire in murine hearts following myocardial infarction by multi-color flow cytometry using a novel 12-marker panel allowing us to characterize major principal leukocyte lineages with a total of 10 distinct leukocyte populations. In parallel, we studied the immune response after MI in secondary lymph organs such as heart´s draining lymph nodes and the spleen. In addition, we performed transcriptional analysis of the healthy and infarcted hearts by bulk RNA at selected time points.

Methods
Permanent ligation of the left descending coronary artery (LAD), ischemic/reperfusion injury (I/R) and sham surgery was performed on C57BL6/J, 10 weeks old male mice. Leukocytes dynamics and T cell responses were screened in infarcted and remote areas of the heart, as well as in blood, spleen, and heat´s draining lymph nodes after 1, 3, 7, 14 and 28 days by multi-color flow cytometry. Also, immune activation and exhaustion transcription analysis by bulk RNA of healthy and infarcted hearts was performed at day 3,7 and 28 days after MI.

Results
We describe a sequential shift in the resident heart leukocyte population, a change in T cell activation states and identified potential new targets such as NK and γ/δ T cells. While macrophages area the most frequent leukocytes under steady state, making up to 75% of the total relative leukocyte population, at day 28 Lymphocytes represent 50%, among which B cells represent the most of them, followed by T helper cells. At early stages, CD4+ T cells secret pro inflammatory cytokines in the heart and in the heart´s draining lymph nodes, however, as stable scar in the heart is formed at day 7 immunomodulatory cytokines secretion such as IL-10 was observed. This is consistent with the build-up of a relevant adaptive immune response involving antigen presenting DCs and effector T and B cells in the healing heart.

Conclusions
Our findings contribute to identify novel cellular and transcriptional targets in the fight against ischemic heart failure.
Quantitative assessment of myocardial fibrosis by digital image analysis: an adjunctive tool for pathologist ground truth
Authors: Maltes S , Abecasis J , Dias N C , Pinto D G , Santos R R , Lopes P M D , Madeira M , Cardim N , Ramos S , Felix A 
Published: November 2023

Abstract

Aims
Myocardial fibrosis (MF) is a common pathological process in a wide range of cardiovascular diseases. Its quantity renders both current myocardial condition and previous accumulated damage. We aimed to assess if the use of an automated artificial intelligence software for the quantification of MF on endomyocardial biopsies (EMB) might improve individual pathologist´s semi-quantification and agreement, in terms of precision.

Methods and Results
EMB samples were studied from intraoperative biopsies of the interventricular septum of 30 patients with severe aortic stenosis submitted to surgical aortic valve replacement. Tissue sections were stained with Masson´s trichrome for collagen/fibrosis and whole slide images (WSI) from the experimental glass slides were obtained at a resolution of 0.5mm using a digital microscopic scanner. Three experienced pathologists made a first quantification of MF excluding the subendocardium. An automatic algorithm, developed on QuPath software for Masson`s trichrome brightfield WSI, was applied and MF quantification ensued (Figure 1), being revealed to the pathologists. A new blinded quantification was performed by each of the pathologists after two weeks. The agreement of MF quantification and the impact of the automatic quantification were tested with Bland-Altman type methodology.
Median values of MF on EMB were 8.33% [IQR 5.00-12.08%] and 13.60% [IQR 7.32-21.2%], respectively for the first pathologist´s and automatic algorithm quantification, being highly correlated (R2: 0.79; p&lt;0.001) (Figure 2). Inter-observer discordance increased at first observation for higher percentages of MF, but the knowledge of the automatic quantification improved the overall pathologist´s agreement, which became unbiased for distinct grades of MF (Figure 3).

Conclusions
The use of an automatic digital pathology algorithm for the quantification of MF on EMB samples significantly improves the reproducibility of measurements by experienced pathologists. This may prove to be a useful adjunctive tool to both improve the reliability of the quantification of myocardial tissue components and imaging-pathology correlation studies.
Cardiac-specific deletion of myosin phosphatase targeting subunit 2 alleviates cardiac fibrosis in mineralocorticoid receptor-related hypertension
Authors: Ye Z , Okamoto R , Ito H , Moriwaki K , Ito R , Ida M , Ito M , Dohi K 
Published: November 2023

Abstract

Introduction
Myosin phosphatase targeting subunit 2 (MYPT2) is a crucial subunit of cardiac myosin light chain (MLC) phosphatase. The phosphorylation of MLC in the ventricle plays a pivotal role in cardiac hypertrophy and function. Our previous study demonstrated that overexpression of MYPT2 activated MLC phosphatase leading to cardiac dysfunction. Mineralocorticoid receptor (MR)-related hypertension is associated with extensive cardiac fibrosis. It remains unknown whether MYPT2 may play a role in the development of cardiac fibrosis in MR-related hypertension.

Purpose
Our purpose is to investigate the effect of MYPT2 on cardiac fibrosis in MR-related hypertension.

Methods
After the knockdown of MYPT2 using MYPT2-siRNA, HL-1 cells (murine cardiomyocytes) were incubated with aldosterone (ALDO) for 24h. Cardiac-­specific MYPT2 knockout (c-MYPT2-/-) mice were generated using the Cre-lox system (αMHC-­Cre; MYPT2f/f). MR-related hypertension mice were induced in c-MYPT2-/- and wild-type (MYPT2+/+) mice with the subcutanouse infusion of ALDO and 8% NaCl food for 4 weeks after uninephrectomy. Systolic blood pressure (SBP) was measured by tail cuffs every week. WB, qPCR, cardiac echo, and histological examinations were performed.

Results
In HL-1 cells, ALDO increased the expression of connective tissue growth factor (CTGF), MYPT2, and the catalytic subunit of the type-1 phosphatase δ isoform (PP1cδ) in a concentration-dependent manner. MYPT2 protein levels significantly decreased by 80% in HL-1 cells transfected with MYPT2 siRNA compared with cells transfected with unspecific siRNA. Knockdown of MYPT2 significantly decreased the protein level of CTGF. In the mice model, the MYPT2 expression in the heart from MYPT2-/- mice was decreased to less than 30% compared with MYPT2+/+ mice. Heart rate, blood pressure, and cardiac systolic function were normal in c-MYPT2-/- mice. PP1cδ was decreased in c-MYPT2-/- mice compared with MYPT2+/+ mice. The expression of cardiac MLC kinase was not changed. The level of phosphorylation of MLC in c-MYPT2-/- mice was significantly higher than MYPT2+/+. Blood pressure elevation and left ventricular hypertrophy were observed in both MR-related hypertension mice. No significant difference in heart size and the nuclear localization of MR in cardiomyocytes. However, ejection fraction and fractional shortening in echocardiography were higher in c-MYPT2-/- than in MYPT2+/+ in MR-related hypertension mice. Histopathological examinations revealed the degree of fibrosis in c-MYPT2-/- mice was lower than in MYPT2+/+ mice with the decreased expression of CTGF.

Conclusions
Cardiac-specific deletion of MYPT2 resulted in a decrease of MLC phosphatase and an increase of phosphorylation in MLC in vivo. MYPT2 deletion can inhibit cardiac fibrosis in a MR-related hypertension model.
High FABP4, associated with low-voltage areas, increases the epicardial fibrosis and its glucose metabolism
Authors: Vazquez-Abuin X , Fu X , Couselo-Seijas M , Lopez-Canoa J N , Martinez-Cereijo J M , Reija-Lopez L , Fernandez-Gonzalez A L , Gonzalez-Juanatey J R , Rodriguez-Manero M , Eiras S 
Published: November 2023

Abstract

Background
High plasma levels of FABP4 were associated with epicardial fat volume on persistent AF patients. FABP4 can be released by differentiated adipocytes and macrophages, which can be involved in fibrosis.

Purpose
Our main objective was to study the association between FABP4 and epicardial fibrosis, which can explain low-voltage areas (LVA) and atrial fibrillation (AF) recurrence.

Material and methods
Peripheral levels of FABP4 and Galectin-3 were analysed on AF patients undergoing pulmonary vein isolation (PVI) (n=299) and signed informed consent. During the intervention, LVA was evaluated by invasive electroanatomic voltage mapping (EAM). Epicardial (EAT) (n=6) and subcutaneous adipose tissue (SAT) (n=5) were obtained from undergoing open heart surgery patients. Stromal cells were isolated by collagenase digestion, cultured and treated with and without FABP4 100 ng/mL and/or galectin-3 30 ng/mL for 24 hours. The proliferation and migration rate were measured by wound healing assay; mitochondrial activity was studied by MTT assay; and glucose and lactate consumption were detected by colorimetric and fluorescence enzymatic assays, respectively. Gene expression levels were assessed by real-time PCR.

Results
Regression analysis determined that the main independent predictors of LVA were age, left atrial area and FABP4 levels. Preclinical "in vitro" studies have shown that FABP4 100 ng/mL induced fibrosis in epicardial stromal cells (p=0.0039) and subcutaneous stromal cells (p=0.0009), evidenced by wound healing assay and by the increment of associated fibroblast genes (COL1A2, VIM, TGF-β1). Moreover, the high FABP4 concentration modulates the protein levels and metabolic genes expression (CPT1, FASN, ATGL, related to transport, synthesis and degradation of fatty acids, respectively) on EAT stromal cells. Glucose metabolism without lactate production was also incremented after FABP4 treatment (p&lt;0.0001). Even, on SAT cells, released lactate was reduced after FABP4 treatment (p=0.0218).

Conclusions
Plasma FABP4 was a predictor of LVA. Our results suggest that this association might be explained by the fibrotic and metabolic role of FABP4 in epicardial cells. Future studies will shed light on the benefits on atrial remodelling with FABP4 modulators.Figure 1Figure 2
ERBB4 stimulation mitigates atrial fibrillation and remodeling in a porcine DOCA model
Authors: Tubeeckx M R L , Goovaerts B , Van Fraeyenhove J , Murphy S L , De Meyer G R Y , De Keulenaer G W , Heidbuchel H , Segers V F M 
Published: November 2023

Abstract

Background
Before patients have spontaneous AF, pathologic remodeling is often already extensively present in the atria, creating a substrate for AF. Currently, no therapies exist that target this atrial remodeling, that can be divided into electrical (ion channel alterations) and structural (fibrosis, dilatation) remodeling. JK07 is a long-acting neuregulin-fusion protein with preventive effects towards ventricular fibrosis in models of heart failure through selective stimulation of the ERBB4 receptor.

Purpose
To test the ability of JK07 to reduce atrial fibrosis and AF inducibility in the minipig model of deoxycorticosterone acetate (DOCA, an aldosterone agonist) induced hypertension.

Methods
18 Aachener minipigs were randomized into 3 groups: control (CTRL), DOCA + Vehicle (DOCA) and DOCA+JK07. The control group did not undergo disease induction or therapeutic intervention. To induce hypertension and atrial fibrosis, pellets releasing 100 mg/kg DOCA over a period of 60 days were implanted in minipigs in the DOCA+VEH and DOCA+JK07 groups. The DOCA-implanted animals underwent weekly treatment with JK07 (0.3 mg/kg; DOCA + JK07) or vehicle control (DOCA), starting on the day of implantation (total of 9 administrations). After 60 days, arterial blood pressure was measured invasively and a decapolar catheter was placed in the right atrium. AF inducibility was tested by performing 50 burst pacing episodes and quantified as the percentage of "successful" episodes where an AF run ≥ 5 sec could be induced after the burst, out of the total attempts per group. Atrial fibrosis was quantified using ImageJ software on Masson trichrome staining of atrial specimens.

Results
Mean arterial pressure was significantly higher in the DOCA+VEH (142 ± 10 mmHg) and DOCA+JK07 (132 ± 15 mmHg) groups than in the CTRL group (105 ± 8 mmHg, p&lt;0.01), without an effect of JK07.
AF inducibility in the DOCA+VEH group was significantly higher than in the CTRL group (66/250 vs. 9/300, p&lt;0.001), and the DOCA + JK07 group (66/250 vs. 8/300, p&lt;0.001).
The degree of fibrosis in the left atrial free wall was significantly higher in the DOCA group compared to CTRL (12.16 ± 5.06 vs. 5.14 ± 1.57, p=0.0089) and compared to DOCA+JK07 (12.16 ± 5.06 vs. 5.47 ± 3.69, p=0.026). Similar results were also found in the right atrial free wall and left and right atrial appendage.

Conclusions
ERBB4 stimulation prevents atrial fibrotic remodeling and AF inducibility in a porcine DOCA model. The effect of JK07 is unrelated to effects on blood pressure, but probably related to reduced atrial fibrogenesis.ERBB4 stimulation prevents AF inductionERBB4 stimulation prevents fibrosis
Interleukin 37 attenuates platelet activation and thrombosis formation
Authors: Gao W , Chen Y F , Luo X P 
Published: November 2023

Abstract

Aims
Interleukin 37 (IL-37) emerges as a natural suppressor of innate inflammatory and immune responses. Plasma IL-37 is abnormally increased in cardiovascular disease. Pleiotropic effects of IL-37 beyond anti-inflammation have been shown. However, the direct effects of IL-37 on platelet activation and thrombosis remain unclear. This study explored the direct effects of IL-37 on platelet activation and thrombosis.

Methods and results
A negative correlation between plasma IL-37 concentration and platelet aggregation was observed in patients with myocardial infarction. IL-37 directly inhibited ADP, thrombin and collagen induced platelet aggregation and dense granule release (Figure 1A, B). Platelet spreading in immobilized fibrinogen and clot retraction driven by integrin αIIbβ3 mediated out-side in signaling were inhibited by IL-37 concentration-dependently. In vivo thrombosis formation and stability in a FeCl3-injured mesenteric arteriole thrombosis mouse model was also severely suppressed by IL-37. Using co-immunoprecipitation, confocal microscopy and platelet-specific IL-1R8-deficient mice, we found that IL-37 bound to platelet IL-1R8 and IL-18Rα, and silencing of IL-1R8 impaired inhibitory effects of IL-37 on platelet activation. Mechanistic studies revealed that IL-37 combined with IL-1R8 to enhance PTEN activity and inhibited Akt, mitogen-activated protein kinase pathways. Platelet-specific IL-1R8-deficient mice presented impaired thrombosis and hemostasis but had improved cardiac function, reduced infarct size, decreased inflammatory response, and microthrombus after permanent ligation of the left anterior descending coronary. While control mice treated with IL-37 were protected from myocardial injury, IL-1R8-/- mice were not (Figure 1C, D).

Conclusions
IL-37 directly attenuated platelet activation and thrombosis formation by binding to platelet IL-18Rα and IL-1R8 and regulating the downstream signaling pathways, suggesting the potential role of exogenous IL-37 protein to prevent platelet hyperactivity thrombotic complications in patients with low plasma IL-37 levels.resultsmode picture