bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abstract 250: Adult Cardiac Stem Cells regulation by microRNAs
Authors: Matsuda Alex J , Pinheiro Ana I , Amaral Andreia , Gama-Carvalho Margarida , Rosário Luís , Kajstura Jan , Anversa Piero 
Published: July 2021

In recent years stem and progenitor cells have been identified in the heart, changing our understanding and therapeutic approach of heart disease. These cells can proliferate and differentiate in vitro into beating cardiomyocytes, endothelial and vascular smooth muscle cells, providing the potential for tissue regeneration and repair.
microRNAs (miRs) have been identified as master switches controlling proliferation and differentiation, and in particular as key regulators of stem cells and cardiac development and function. Modulation of miR-regulated gene expression networks holds the potential to control cell fate and proliferation, with predictable biotechnological and therapeutic applications.
We have characterized the expression profile of a subset of 100 miRs with reported functions in stem cell and tissue differentiation in mouse Sca1+ cardiac progenitor cells (CPCs). CPC miR expression profiles were compared with bone marrow mesenchymal stem cells (MSCs) and mouse embryonic heart (E=9) to obtain insights into the origins and function of this rare cell population. Within the miR panel that was studied, a subset with relative high expression levels in CPCs was identified, which may act as negative regulators of differentiation and proliferation. Although the overall expression profile of Sca1+ CPCs is closer to BM-MSCs, the most highly expressed miRs in CPCs are distinctive and predicted to target key genes involved in the control of cell proliferation and adhesion, vascular function and cardiomyocyte differentiation.
Our results provide novel insights into the gene expression networks active in CPCs, which will now be explored in functional studies to identify key regulators of proliferation and differentiation.
Abstract 255: Modulation Of Cardiac Macrophages As A Strategy For Infarct Healing
Authors: Somasuntharam Inthirai , Carroll Sheridan , Brown Milton , Garcia Andres , Davis Michael 
Published: July 2021

Heart failure is the leading cause of death in the developed world and myocardial infarction (MI) is the most common cause. Macrophages are key cells that orchestrate the initial inflammatory as well as later stage wound healing responses following MI. These functions are carried out by pro-inflammatory (M1) and reparative (M2) macrophages respectively. Optimal healing response after MI requires a balancing act of the biphasic macrophage response, so as to not prolong inflammatory signals detrimental to wound healing. Taking advantage of the fact that interleukin-4 (IL-4) activates macrophages towards M2, we hypothesize that delivering IL-4 to the post-MI heart can alter the ratio of M2 to M1 macrophages in the infarct area and induce a better healing response. In this study, we validate our approach in vitro and perform in vitro optimization of a suitable delivery system.
RAW 264.7 macrophages were stimulated with IL-4 (10ng/uL) or LPS/IFN-γ (100ng/mL and 10ng/mL) for 24h and gene expression markers (qPCR) and Nitric Oxide (NO) levels (Griess assay) analyzed as indication of M1or M2 activation. Mouse aortic endothelial cells were treated with conditioned media from these cells for 24h and tube formation assessed on matrigel. A bioactive, protease-cleavable polyethylene glycol (PEG) hydrogel delivery system was evaluated for release of functional IL-4 to LPS-activated macrophages. Empty or IL-4 encapsulating hydrogel was placed on a trans-well above LPS-stimulated macrophages. Collagenase I at 0.1mg/mL was applied over 48h to degrade the gels and release IL-4 (n≥3 and p<0.05 considered significant by one-way ANOVA).
We demonstrate that IL-4 significantly upregulates M2 markers (MRC-1 and Arg-1) while IFN-γ and LPS upregulate M1 markers (NO and TNF-alpha). We observe enhanced tube density in endothelial cells treated with M2 media while M1 inhibited tube formation. Hydrogel release study shows a significant reduction in NO levels of LPS-stimulated macrophages when IL-4 is released, demonstrating that IL-4 is released from the gel in its bioactive form. In conclusion, we show that macrophages can indeed respond to changing stimuli and adopt distinct activation types and our PEG based hydrogel could be a potential delivery system for in vivo IL-4 delivery.
Abstract 267: Opposing Functions of Two Splice Variants of the Human Transcription Factor Grainyhead-like 3 in Endothelial Cells And in vivo
Authors: Altschmied Joachim , Büchner Nicole , Jakob Sascha , Farrokh Sabrina , Goy Christine , Schaal Heiner , Haendeler Judith 
Published: July 2021

Grainyhead-like 3 (GRHL3) is a member of the evolutionary conserved Grainyhead family of transcription factors. In humans, three isoforms are derived from differential first exon usage and alternative splicing, which differ only in their N-terminus. Isoform 2, the only variant also present in mouse, is required for endothelial cell (EC) migration and protects against apoptosis. The functions of the human specific isoforms 1 and 3, which are derived from an alternatively spliced pre-mRNA, have not yet been investigated, although all three isoforms are expressed in EC. Therefore, we have assessed their effects on EC migration and apoptosis.
Overexpression of the two proteins had opposite effects on EC migration, with isoform 1 acting pro-migratory. This protein also protected EC against apoptosis in an eNOS-dependent manner, whereas isoform 3 had no effect. These opposing outcomes with respect to apoptosis EC were corroborated by isoform-specific knockdowns.
With reporter assays using a GRHL3-specific luciferase reporter we demonstrated that both are active transcription factors. Microarray analyses revealed that they induce divergent target gene sets in EC. Two validated targets, Akt2 and Mxi1, which are upregulated by isoform1, are regulators of Akt1-, and thus eNOS-phosphorylation and apoptosis, which could explain the effects of this protein on these processes.
In vivo, overexpression of isoform 3 in zebrafish embryos resulted in increased lethality and severe deformations, while isoform 1 had no deleterious effect.
In conclusion, our data demonstrate that the splice variant derived isoforms 1 and 3 of the human transcription factor GRHL3 induce opposing effects in primary human endothelial cells and in a whole animal model, most likely through the induction of different target genes.
Abstract 304: Estrogen Plays a Protective Role in Advanced Heart Failure by Suppressing Cardiac Fibrosis Associated Genes via miR129 Induction
Authors: Sharma Salil , Iorga Andrea , Singh Harnek , Li Jingyaun , Eghbali Mansoureh 
Published: July 2021

We have previously shown that short term treatment of estrogen(E2) can rescue advance heart failure(HF) and decreases associated fibrosis. We hypothesized that E2 can reduce fibrosis by regulating the levels of specific microRNAs including miR129-5p(miR129) through ERβ mediated mechanism.
We used transaortic constriction to induce HF in male mice, and once the ejection fraction (EF) reached ~30%, one group of animals was sacrificed (HF), and the other group received 17b-estradiol via a subcutaneous pellet implant (0.012mg/pellet, n=16) (E2) for 10 days. Sham-operated mice served as CTRL. Serial echocardiography was performed to monitor cardiac structure and function. Short-term E2 treatment rescued pressure overload-induced decompensated HF in mice by restoring the EF from 33.17±1.12% to 53.05±1.29 (p <0.001, n=16). E2 decreased both interstitial and perivascular fibrosis in HF. Microarray analysis comparing HF with E2 revealed ~70 microRNAs including miR129 regulated by E2. qPCR validation revealed that E2 treatment upregulates miR129 by 2 folds compared to HF restoring it to CTRL levels. Treatment of HF with ERβ agonist (DPN), but not ERα agonist (PPT) resulted in the upregulation of miR129 indicating the E2 mediated induction of miR129 is mediated through ERβ. In vitro, angiotensin II treatment significantly downregulated miR129 expression in neonatal rat fibroblasts (NRVF) which was restored by E2 and DPN but not by E2+ERβ antagonist (PHPT) further confirming the role of ERβ in regulating miR129. In vitro, OE of miR129 in both neonatal and adult rat cardiac fibroblasts (ARVF) resulted in significant downregulation of transcripts of many in-silico predicted pro-fibrotic target genes including EGFR, RUNX, GREM1, COL2A, PDGFA, PDGFRA and the transcription factor SOX4. OE of miR129 in fibroblasts also resulted in downregulation of EGFR protein. Gain of miR129 prevented the transition of fibroblasts to myofibroblasts in both NRVF and ARVF and inhibited fibroblast proliferation in vitro.
In conclusion, E2 treatment during HF induces miR129 likely through ERβ. MiR129 represses fibrosis by targeting key genes associated with cardiac fibrosis, inhibits fibroblast proliferation and fibroblast to myofibroblast transition.
Abstract 356: Scl Represses Cardiogenesis via Distant Enhancers during Hemogenic Endothelium Specification
Authors: Org Tonis , Duan Dan , Ferrari Roberto , Montel-Hagen Amelie , Van Handel Ben , Sasidharan Rajkumar , Orkin Stuart H , Kurdistani Siavash , Mikkola Hanna K 
Published: July 2021

Understanding the mechanisms directing mesoderm specification holds a great potential to advance the development of cell-based therapies for cardiovascular and blood disorders. The bHLH transcription factor Scl is known as the master regulator of the hematopoietic fate. We recently discovered that, in addition to its critical function in promoting the establishment of hemogenic endothelium during hematopoietic stem/progenitor cell (HS/PC) development, Scl is also required to repress cardiomyogenesis in endothelium in hematopoietic tissues and endocardium in the heart. However, the mechanisms for the cardiac repression have remained unknown.
Using ChIP-sequencing and microarray analysis of Flk+ mesoderm differentiated from mouse ES cells, we show that Scl both directly activates a broad gene regulatory network required for hemogenic endothelium and HS/PC development (e.g. Runx1, cMyb, Lyl1, Mef2C, Sox7 etc.), and directly represses transcriptional regulators required for cardiogenesis (e.g. Gata4, Gata6, Myocd, etc.) and mesoderm development (Eomes, Mixl1, Etv2, etc.). Repression of cardiac and mesodermal programs occurs during a short developmental window through Scl binding to distant enhancers, while binding to hematopoietic regulators extends throughout HS/PC and red blood cell development and encompasses both distant and proximal binding sites. We also discovered that, surprisingly, Scl complex partners Gata 1 and 2 are dispensable for hematopoietic vs. cardiac specification and Scl binding to majority of its target genes. Nevertheless, Gata factors co-operate with Scl to activate selected transcription factors that facilitate HS/PC emergence from hemogenic endothelium. These results denote Scl as a true master regulator of hematopoietic vs. cardiac fate choice and suggest a mechanism by which lineage-specific bHLH factors direct the divergence of competing fates.
Abstract 109: Interleukin 17 A, Coronary Artery Disease And Gender of The Patients
Authors: Oliveira Dinaldo C , Heide Elayne , Rego Moacyr , Oliveira Danielle A , Sarinho Felipi W , Gomes Viviane R , Pita Maira G 
Published: July 2021

Introduction:
The effects of interleukins during development of coronary artery disease (CAD) have been studied by researchers and it may help to understand better this disease. The aim of this study was to evaluate if there are differences of concentrations of the IL 17 A between men and women with stable CAD.

Hypothesis:
There are no differences of concentrations of the IL 17 A according to gender of the patients.

Methods:
This is a cross-sectional, prospective, analytical study, conducted from August to December 2012. We included 40 patients (24 men (MP) and 16 women (WP) with stable CAD, CCS III or IV, ischemic myocardial scintigraphy, who had not been subjected to any kind of myocardial revascularization and with coronary stenosis ≥ 50% according to current coronary angiography. There were 20 healthy volunteers (12 men (MC) and 8 women (WC), to take up comparison of concentrations of interleukins (IL). Interleukins were evaluated in serum of patients and after 48 hours of cells in culture with and without stimulus. IL concentrations were expressed in pg / ml. Statistical analysis was performed using the Mann-Whitney or Student t test. P ≤ 0,05 was considered statistically significant.

Results:
The comparison between the groups showed:

IL 17 A: Serum: MP = 3.91 (3.91 - 3.91) vs MC = 3.91 (3.91 - 9.28), p = 0.31; culture 48 hours without stimulus: MP = 3.91 (3.91 - 3.91) vs MC = 3.91 (3.91 - 3.91), p = 0.45; culture 48 hours with stimulus: MP = 451.67 (99.02 - 892.58) vs MC = 135 (3.91 -- 285), p = 0.04.
IL 17 A: Serum: WP = 3.91 (3.91 - 3.91) vs WC = 3.91 (3.91 -- 6.25), p = 0.90; culture 48 hours without stimulus: WP = 3.91 (3.91 - 3.91) vs WC = 3.91 (3.91 -- 3.91), p = 0.20; culture 48 hours with stimulus: WP = 131.21 (3.91 - 231.97) vs WC = 173.78 (3.91 - 642), p = 0.24.
IL 17 A: Serum: MP = 3.91 (3.91 - 3.91) vs WP = 3.91 (3.91 - 3.91), p = 0.17; culture 48 hours without stimulus: MP = 3.91 (3.91 - 3.91) vs WP = 3.91 (3.91 - 3.91), p = 0.47; culture 48 hours with stimulus: MP = 451.67 (99.02 -- 892.58) vs WP = 131.21 (3.91 - 231.97), p = 0.02.

Conclusions:
There were no differences of concentrations of the IL 17 A between WP and WC. However, MP had higher concentrations of the IL 17 A than MC and WP. Therefore, it is possible that the IL 17 A take part in a pathway which determines different patterns of CAD according to gender.
Abstract 12: The Effect of Extracellular Matrix Stiffness on Human Bone Marrow-Derived Mesenchymal Stem Cell Differentiation
Authors: Müller Alison L , Li Yun , Hinz Boris , Freed Darren H 
Published: July 2021

Differentiation of human mesenchymal stem cells (hMSCs) has been shown to be influenced by the surrounding microenvironment. It is important to understand the physiological implications of the hMSC microenvironment regarding differentiation within the body, especially in patients with cardiovascular disease. We are interested in understanding the influence of differing extracellular matrix (ECM) stiffness found in the body that the MSC encounters during its journey from the bone marrow to the infarct scar in patients recovering from a myocardial infarction. As hMSCs must respond rapidly to their environment, we also investigated the influence of microRNA at different surface tensions.
HMSCs were isolated from the bone marrow of patients undergoing open heart surgery and cultured in standard DMEM/F12 with 20% FBS. We plated these cells on fibronectin-coated plates with surface tensions of 2kPa, simulating bone marrow; 15kPa, simulating left ventricle; and 100kPa, simulating a fibrotic environment. Protein and mRNA were collected for further analysis.
Our data have revealed that softer surface tensions, representing a bone marrow-undifferentiating environment, cause a decrease in the protein expression of EDA-fibronectin and alpha-smooth muscle actin. There were also increases in mRNA of myosin heavy chain-9 and 10, and collagen-1. Softer surface tensions also show a slight increase in miR-301a, although at 15kPa, miR-301a expression is increased even further. Our lab has previously shown that miR-301a is involved in maintaining a proliferative phenotype of hMSCs. Interestingly, Dicer1, responsible for processing microRNAs, is upregulated at softer surface tensions and attenuated at 15kPa. Dicer1 mRNA expression is attenuated at 15kPa. These results indicate that ECM stiffness influences hMSC differentiation and the increase in Dicer1 found with softer matrices could represent an umbrella miRNA inhibition effect to effectively suppress hMSC differentiation.
Abstract 135: Reprogramming of Pig Cardiac Fibroblasts to Cardiomyocyte Fate: Implications for Gene Therapy to Treat Myocardial Infarction
Authors: Singh Vivek P , Mathison Megumi , Xu Xueping , Patel Vivek K , Belaguli Narasimhaswamy S , Gibson Brian W , Cooney Austin J , Rosengart Todd K 
Published: July 2021

There is an urgent clinical need to develop new therapeutic approaches to treat heart failure, but the biology of cardiovascular regeneration is complex. A new generation of reprogramming technology involves trans-differentiating one adult somatic cell type directly into another. We reported previously that administration of gene transfer vectors encoding Gata4 (G), Mef 2c (M) and Tbx5 (T), reprograms rat cardiac fibroblasts into induced cardiomyocytes (iCMs) in vitro and improves cardiac function in myocardial infarction models. Previous cardiac reprogramming studies were restricted to rodent and human fibroblasts. However studies in large animal models relevant for pre-clinical studies is lacking.
The Aim of the present study was to determine the optimal combination of factors necessary for direct reprogramming of porcine fibroblasts towards a cardiomyocyte lineage for the first time. Here we have used human lentiviral system to express various cardiomyocyte enriched transcriptional regulators such as GMT, Hand2, myocardin, and two microRNAs, miR-590 and mir-199, in porcine cardiac fibroblasts. Reprogramming of fibroblasts into iCMs was determined 4 weeks post-virus transduction by FACS analysis for the activation of endogenous cardiac troponin T (cTnT).
GMT alone was not sufficient to reprogram porcine fibroblasts although this combination was shown previously to be sufficient to convert rodent fibroblasts into iCMs. Addition of mir-199 to GMT was also not sufficient to promote reprogramming. However, adding miR-590 or Hand2, and myocardin (HM) to GMT resulted in reprogramming, although the efficiency remained low. Together, our results show that combination of 5 distinct transcription regulators (GMTHM) or GMT plus mir-590 is necessary for reprogramming porcine fibroblast.
These findings demonstrate that pig fibroblasts can be directly reprogrammed toward the cardiomyocyte lineage, and represent a step toward possible therapeutic application of this reprogramming approach in a pre-clinical setting.
Abstract 168: Contribution of Vascular Endothelial Growth Factor Signaling to Fate Specification in Cardiomyocytes
Authors: Lu Taylor Y , Domigan Courtney K , Antanesian Vaspour , Nakashima Yasuhiro , Nakano Atsushi , Iruela-Arispe Luisa 
Published: July 2021

Vascular endothelial growth factor (VEGF) is one of the pivotal proangiogenic growth factors that has long contributed to our knowledge of blood vessel and circulatory maintenance as well as angiogenesis in both pathology and pathophysiology. However, the non-canonical functions of VEGF in cardiac morphogenesis have not been well characterized. Here, we examined how VEGF regulates cardiomyocyte cell fate.
Using chimeric embryos harboring both wild type and VEGF-null embryonic stem cells, we observed that derivatives of VEGF null cells were preferentially recruited to the atrium of the heart in comparison to the ventricles. To further provide physiologic context of this finding, we used reporter-LacZ staining and RT-PCR and found that endogenous VEGF was indeed expressed at much lower levels in the atrium but highly expressed in the ventricle early in cardiac morphogenesis. These data lead to our hypothesis that cell-autonomous expression of VEGF is a determinant of atrial vs. ventricular cardiomyocyte cell fate. To test this hypothesis, we used a VEGF knock-in mouse model of Sm22Cre x Rosa 26 VEGF. VEGF overexpression in cardiomyocytes (and smooth muscle) at E8.5 resulted in lethality by P1 and thickened atrial and ventricular walls in mutant embryos as characterized by histology (H&E, IF). We further explored the molecular changes underlying this phenotype via microarray and RT-PCR and find disruptions in molecular markers necessary for wall development, specifically: Notch-1, BMP10, Nrg-1.
Taken together, our data indicates that aberrant embryonic VEGF signaling disrupts several critical signaling pathways and that overexpression leads to disruption of cardiomyocyte proliferation and cardiac morphogenesis. These findings add to the foundation of better understanding heart development, laying the groundwork for future therapy of congenital and acquired cardiac disease.
Abstract 193: 18-HEPE, An EPA Metabolite Released By Macrophages, Prevents Against Pressure Overload-induced Cardiac Remodeling
Authors: Endo Jin , Sano Motoaki , Fukuda Keiichi , Arita Makoto 
Published: July 2021

N-3 polyunsaturated fatty acids (PUFAs) have potential cardiovascular benefit, although the mechanisms underlying this effect remain poorly understood. The present study investigated the impact of the enrichment of n-3 PUFAs in the setting of pressure overload induced cardiac hypertrophy using fat-1 mouse, a genetic model with elevated n-3 PUFA levels in tissues, as well as the possible cellular mechanism involved in such a role.
The fat-1 mice subjected to pressure overload by transverse aortic constriction showed less infiltration of macrophages and decreased interstitial fibrosis, leading to the preserved cardiac function as compared to wild type (WT) mice. Bone marrow (BM) transplantation experiments revealed that fat-1 transgenic BM cells, but not fat-1 transgenic cardiac cells, contributed the anti-remodeling effect, which was comparable between fat-1 and WT mice transplanted with fat-1 transgenic BM, but was absent in fat-1 mice transplanted with WT BM. In the co-culture assay with cardiac fibroblasts, the fat-1 transgenic macrophages showed anti-inflammatory and anti-fibrotic activities. Lipidomic analysis revealed selective enrichment of eicosapentaenoic acid (EPA) in fat-1 transgenic bone marrow (BM) cells and an EPA-metabolite 18-hydroxyeicosapentaenoic acid (18-HEPE) in fat-1 transgenic macrophages. 18-HEPE-rich milieu in the fat-1 transgenic heart was generated by BM-derived cells, most likely macrophages. 18-HEPE inhibited macrophage-mediated proinflammatory activation of cardiac fibroblasts in culture, and in vivo administration of 18-HEPE significantly attenuated the inflammation and fibrosis of hypertrophied hearts, resulting in improvement of cardiac function.
We found that the elevated n-3 PUFA level in BM-derived macrophages is responsible for cardiac protection, and 18-HEPE exhibited cardioprotective potential when administered in vivo.