bluuebunny/crossref_metadata_2025 huggingface dataset:
Impact of Oral Anticoagulation Therapy Versus Left Atrial Appendage Occlusion on Cognitive Function and Quality of Life in Patients With Atrial Fibrillation
Authors: Mohanty Sanghamitra , Mohanty Prasant , Trivedi Chintan , Assadourian Joanna , Mayedo Angel Quintero , MacDonald Bryan , Della Rocca Domenico G. , Gianni Carola , Horton Rodney , Al‐Ahmad Amin , Bassiouny Mohamed , Burkhardt John D. , Di Biase Luigi , Gurol M. Edip , Natale Andrea
Published: June 2021

Background

We compared the cognitive status and quality of life in patients with atrial fibrillation undergoing left atrial appendage occlusion (LAAO) or remaining on oral anticoagulation (OAC) after atrial fibrillation ablation.

Methods and Results

Cognition was assessed by the Montreal Cognitive Assessment (MoCA) survey at baseline and follow‐up. Consecutive patients receiving LAAO or OAC after atrial fibrillation ablation were screened, and patients with a score of ≤17 were excluded from the study. Quality of life was measured at baseline and 1 year using the Atrial Fibrillation Effect on Quality of Life survey. A total of 50 patients (CHA
2
DS
2
‐VASc [congestive heart failure, hypertension, age≥75 years, diabetes mellitus, stroke or transient ischemic attack, vascular disease, age 65–74 years, sex category] score: 3.30±1.43) in the LAAO group and 48 (CHA
2
DS
2
‐VASc score 2.73±1.25) in the OAC group were included in this prospective study. Mean baseline MoCA score was 26.18 and 26.08 in the LAAO and OAC groups, respectively (
P
=0.846). At 1 year, scores were 26.94 and 23.38 in the respective groups. MoCA score decreased by an estimated −2.74 (95% CI, −3.61 to −1.87;
P
<0.0001) points in the OAC group, whereas the change in the LAAO group was nonsignificant (0.79; (95% CI, −0.06 to 1.64;
P
=0.07). After adjusting for baseline clinical characteristics, remaining on OAC was an independent predictor of MoCA change at 1 year (regression coefficient, −3.38; 95% CI, −4.75 to −2.02;
P
<0.0001). Change in Atrial Fibrillation Effect on Quality of Life score did not differ significantly in achieving a clinically important difference between groups.

Conclusions

In this series, a significant difference in the postprocedure MoCA score was observed in postablation patients with atrial fibrillation receiving LAAO versus remaining on OAC with a substantial decline in the score in the OAC group. However, quality of life improved similarly across groups.

Registration

https://www.ClinicalTrials.gov
. Unique identifier: NCT01816308
Predictors of Use and Outcomes of Mechanical Valve Replacement in the United States (2008–2017)
Authors: Alkhouli Mohamad , Alqahtani Fahad , Simard Trevor , Pislaru Sorin , Schaff Hartzell V. , Nishimura Rich A.
Published: June 2021

Background

Contemporary nationwide data on the use, predictors, and outcomes of mechanical valve replacement in patients less than 70 years of age are limited.

Methods and Results

We identified hospitalizations for aortic valve replacement (AVR) or mitral valve replacement (MVR) in the Nationwide Inpatient Sample between January 1, 2008, and December 31, 2017. The study's end points included predictors of mechanical valve replacement and risk‐adjusted in‐hospital mortality. Among 253 100 hospitalizations for AVR, the use rate of mechanical prosthesis decreased from 45.3% in 2008 to 17.0% in 2017. Among 284 962 hospitalizations for MVR, mechanical prosthesis use decreased from 59.5% in 2008 to 29.2% in 2017 (
P
for trend<0.001). In multilogistic regression analyses, female sex, prior sternotomy, prior defibrillator, and South/West geographic location were predictive of mechanical valve use. The presence of bicuspid valve was a negative predictor of mechanical AVR (odds ratio [OR], 0.68; 95% CI, 0.66–0.69;
P
<0.001), whereas mitral stenosis was associated with higher mechanical MVR (OR, 1.28; 95% CI, 1.22–1.33;
P
<0.001). Unadjusted in‐hospital mortality decreased over time with AVR but not with MVR, regardless of prosthesis choice. Using years 2008 and 2009 as a reference, risk‐adjusted mortality also decreased over time with AVR but did not decrease after MVR.

Conclusions

There is a substantial decline in the use of mechanical valve replacement among patients aged ≤70 years in the United States. Long‐term durability data on bioprosthetic valve replacement are needed to better define the future role of mechanical valves in this age group.
Abstract 100: Discovery of Cardiac Fibroblast Enriched microRNAs and Their Pivotal Roles in the Regulation of Cardiac Remodeling and Fibrosis
Authors: Zhou Yue , Richards Arthur M , Wang Peipei
Published: June 2021

Cardiac fibroblast (cFB) responses to cardiac injury or overload directly contribute to deterioration of cardiac function in heart failure. MicroRNAs (miR) target multiple genes in cell signaling networks and are likely to have pivotal regulatory roles with respect to cFB function. To date cFB enriched miRs have not been reported. We have identified cFB enriched miRs which we hypothesize direct cFB proliferation and differentiation. Neonatal and adult rat cardiomyocytes (CM) and cFBs were isolated and cultured.
In vivo
and
in vitro
cardiac ischemic models comprised coronary artery ligation induced myocardial infarction (MI) in rats and cultured cFB exposed to hypoxia. RNA and protein were extracted for miR microarray, qPCR and Western Blot. Adult cFB transfected with miR mimics were tested for CCK-8 proliferation assay. Fifteen dysregulated miRs were selected from array profiles and qPCR validation. Among them miR-31, -199a, -214 and -222 were highly expressed in adult cFBs 10-90 folds vs. CM. CM specific miR-208a and 133a were undetectable in cFB. Neonatal cells showed directionally concordant but less pronounced differences. In early MI, cardiac miR-31 was up-regulated >30 fold vs. Sham (infarct), others increased 6-12 folds. All changes were ranked infarct>border>remote area. As a control, non-cFB enriched miR-125a remained unchanged. Hypoxia treatment of cFB
in vitro
up-regulated miR-31 but not the other miRs. Functional study by mimic transfection revealed differential roles of the miRs. MiR-31 increased cFB proliferation in CCK8 assay. MiR-199a and -222 had opposite effects. MiR-199a, but not miR-222, reversed the pro-fibrotic effects of TGF-β. MiR-199a reduced mRNA and protein expression of alpha smooth muscle actin (α-SMA), a myoFB differentiation marker and connective tissue growth factor (CTGF), a predicted target (miRDB). Conversely miR-31 increased α-SMA and CTGF. We provide the first report of 4 cFB enriched miRs and demonstrated their pro- vs. anti-fibrotic roles
in vitro
(miR-31 vs. miR-199 and -222 respectively). In early MI, the increase of pro-fibrotic miR-31 was predominant, whilst other miR dysregulation was secondary to cFB proliferation. cFB enriched miRs determine cFB fate and progression of cardiac fibrosis/remodeling.
Abstract 105: The Novel Heart-specific Ring-finger Protein 207 Regulates Energy Metabolism in Cardiomyocytes
Authors: Mizushima Wataru , Takahashi Hidehisa , Watanabe Masashi , Kinugawa Shintarou , Matsushima Shouji , Takada Shingo , Yokota Takashi , Furuhata Takaaki , Matsumoto Junichi , Tsuda Masaya , Katayama Takashi , Nakajima Takayuki , Hatakeyama Shigetsugu , Tsutsui Hiroyuki
Published: June 2021

Background:
Ring-finger proteins constitute a large protein family in the human genome and play essential roles in various biological processes. However, little is known about heart-specific Ring-finger proteins and those relations with cardiac functions. We performed the comprehensive analysis of the expression profiles of various kinds of Ring-finger proteins and found that Ring-finger protein 207 (RNF207) was largely expressed in the heart. The purpose of our study was to elucidate a role of RNF207 in the heart.

Methods and Results:
First, we confirmed that RNF207 was predominantly expressed in the heart at the mRNA and protein level. Next, we examined whether the expression of RNF207 changed in transverse aortic constriction (TAC) model mice and coronary ischemic heart failure model mice
.
4 weeks after TAC, mRNA level of RNF207 was significantly decreased to approximately 40% of that in sham mice. Moreover, we found that the protein level of RNF207 in the mice with heart failure was significantly reduced to around 70% of that in sham mice. Considering the well-known facts that the heart is a high-energy demanding organ and that the levels of ATP in cardiomyocytes are reduced in those model mice, we hypothesized that RNF207 got involved in cardiac energy metabolism. To investigate the hypothesis, we depleted RNF207 in rat neonatal cardiomyocytes (RNC) and performed metabolomic analysis. Metabolomic analysis revealed that ATP concentration and NADH/NAD
+
ratio were significantly lower and mitochondrial function was significantly reduced in RNF207 depleted RNC, compared to control NRC. Next, to elucidate the molecular mechanism by which RNF207 had effect on cardiac energy metabolism, we explored RNF207-associated proteins by mass spectrometric analysis. We identified voltage-dependent anion channel 1 (VDAC1) as a RNF207-associated protein. It has been shown that mitochondrial protein VDAC1 plays a crucial role in mitochondrial functions, such as energy metabolism. We confirmed that RNF207 directly interacted with VDAC1 by
in vitro
binding assay. Our results strongly indicate that RNF207 functions as a regulator of cardiac energy metabolism.

Conclusion:
RNF207 is a novel heart-specific protein and regulates energy metabolism in cardiomyocytes.
Abstract 110: The Estrogen-related Receptor Gamma Promotes Maturation of iPSC-derived Cardiac Myocytes
Authors: Sakamoto Tomoya , Leone Teresa C , Chen Huei-Sheng Vincent , Vega Rick B , Kelly Daniel P
Published: June 2021

The mammalian heart has a tremendous energy requirement to support normal contractile function. Recently, our group discovered that the estrogen-related receptor γ (ERRγ) coordinately controls oxidative energy metabolism and the type I fiber program in skeletal muscle. We hypothesized that this circuit is also operative in heart, serving to match capacity to produce ATP with energy utilizing processes such as contraction and ion transport. To identify downstream targets and pathways of ERRγ in cardiac myocytes, RNA-sequencing (RNA-seq) was conducted in neonatal rat ventricular myocytes (NRVMs) overexpressing ERRγ. ERRγ induced the expression of genes involved in mitochondrial oxidative phosphorylation, fatty acid oxidation, and the TCA cycle. Interestingly, ERRγ also induced expression of a broad program of adult cardiac contractile and structural genes including
Myh6
(αMHC) and Ca
2+
handling genes such as
Atp2a2
(SERCA2A). ERRγ-mediated activation of adult genes occurred concomitant with the suppression of fetal cardiac and embryonic/perinatal fast skeletal isoform contractile protein gene expression including
Myh7
(βMHC),
Myl1
and
Myh3/8
. These results prompted us to explore the role of ERRγ in differentiation and maturation of human iPS-derived cardiac myocytes (iPSC-CM). The expression level of ERRγ was markedly upregulated during the transition to cardiac lineage commitment in parallel with induction of cardiogenic transcription factors (e.g. NKX2.5 and GATA4). Forced expression of ERRγ in iPSC-CM activated the broad adult cardiac myocyte program including oxidative metabolism and adult contractile gene expression. Of note, ERRγ increased
MYH6
expression while decreasing expression of
MYH7
and the fetal gene markers
NPPA
(ANF) and
NPPB
(BNP). The expression of
TNNI3
, thought to be a marker of mature adult cardiomyocytes, was also induced by ERRγ. Conversely, CRISPR-Cas9n deletion of ERRγ in iPSC-CM resulted in diminished mitochondrial energy metabolic and adult cardiac contractile gene expression. These data provide evidence that ERRγ is a key nodal regulator of cardiac myocyte determination and terminal maturation. ERR signaling is a candidate target for driving iPSC-CM to the “adult” cardiac myocyte phenotype.
Abstract 115: Post-ischemic Estrogen Administration Protects the Myocardium Against Ischemia/Reperfusion Injury by Reducing Mitochondrial Protein Ubiquitination, Acetylation and Calpain10 Levels and Inhibition of the mPTP Opening via GPER1 Activation
Authors: Feng Yansheng , Madungwe Ngonidzashe Benson , Kabir Mohammad Enamul , Bopassa Jean C.
Published: June 2021

Introduction:
We recently found in isolated perfused hearts that acute pre-ischemic estrogen-induced cardioprotection against ischemia/reperfusion injury is mainly mediated via G protein-coupled estrogen receptor1 (GPER1) activation but not through classical estrogen receptors: alpha (ERα) and beta (ERβ).

Hypothesis:
We investigated whether acute post-ischemic estrogen (PI-E2) treatment can also induce cardioprotective effects via GPER1 activation in the intact animal subjected to ischemia/reperfusion injury.

Methods:
Male and ovariectomized female Sprague-Dawley rats were anesthetized with ketamine (80 mg/kg i.p.) and xylazine (8 mg/kg i.p.). Hearts were subjected to 35 min of the left anterior descending (LAD) artery occlusion, followed by 180 min reperfusion. An E2 bolus (50 mg/kg body weight) or PBS (same volume) was applied via the femoral vein 5 min before reperfusion and a GPER1 antagonist, G15, was given 10 min before E2. Area at risk (AAR) was identified using Evans Blue dye and myocardial infarct size assessed by TTC staining method. Mitochondria calcium retention capacity (CRC) required to induce mitochondrial permeability transition pore (mPTP) opening was assessed after 10 min reperfusion. Expression of ubiquitinated, acetylated, as well as calpains 1 and 10 proteins was measured by Western Blot in mitochondrial and cytosolic fractions.

Results:
We found that PI-E2 treatment reduced myocardial infarct size normalized to the AAR or the whole LV and improved mitochondrial CRC. PI-E2 treatment reduced mitochondrial protein acetylation, ubiquitination, and decreased calpain10 levels in mitochondrial but not in cytosolic fractions as compared to control, respectively. Interestingly, all these effects of E2 were abolished by addition of G15.

Conclusion:
Acute post-ischemic GPER1 activation by E2 induces cardioprotection against ischemia/reperfusion injury. PI-E2 effects through GPER1 involve the reduction of the levels of mitochondrial protein acetylation, ubiquitination, and calpain10. These PI-E2-GPER1 effects lead to inhibition of the mPTP opening.
Abstract 140: BMI- and Gender-specific Increase of MAP2K3/p38 Activity in Human Cardiac Hypertrophy
Authors: Newman Mackenzie , Infanto Aniello M , Watson Michael J , Hull Robert W , Yu Han-Gang
Published: June 2021

Introduction:
MAP2K3 is a stress-induced kinase and its role cardiac hypertrophy has been controversial: while
in vitro
studies have displayed a positive correlation in activity with hypertrophy,
in
vivo
studies have shown a negative association. Its direct downstream target, p38α MAPK, has been shown to be in activated in hypertrophic and failed hearts, but this has not been explored with regard to gender or BMI.

Hypothesis:
Early activation of p38 MAPK in hypertrophy is influenced by gender and BMI.

Methods:
Human heart samples were grouped into non-failed (NF), left ventricular hypertrophy (LVH), and NF without LVH (control). BMI < 25 was considered “lean”, 25 < BMI < 30 was considered overweight, and BMI > 30 was considered “obese”. RNA-Seq was used to generate cardiac gene expression profiles. Immunoblots were used to examine protein expression of MAP2K3, p38, and activated p38 (pp38; phospho-T180 and Y182).

Results:
We found that MAP2K3 mRNA levels were increased in obese males by 134% with LVH compared to non-LVH controls (FPKM: LVH = 19.2±3.5, n=3; non-LVH = 8.2±1.9, n=3; FPKM = Fragments Per Kilobase of transcript per Million mapped reads), without statistical significance (p>0.05). MAP2K3 protein expression was 5-fold higher in LVH (1.24±0.20, n=9) versus non-LVH hearts (0.25±0.03, n=15) (p<0.0001). Within all LVH hearts sampled, MAP2K3 protein expression is higher only in male overweight or obese (1.21±0.22, n=7) over lean (0.52±0.04, n=6) (p<0.05). No differences were detected in gene expression profiles of the four isoforms of p38 and total p38 protein expression levels between LVH and NF groups (p>0.05). Activated-p38 was detected in LVH without failure. Levels of p38 activation are higher in overweight/obese than in lean male group (pp38/p38 ratio: M_BMI>25 = 0.63 ± 0.03 (n=7), M_BMI<25 = 0.41 ± 0.12 (n=3), p<0.05). For BMI>25, levels of activated p38 are also higher in male (0.63 ± 0.03, n=7) than in female (0.08 ± 0.01, n=3) (p<0.0001), independent of LVH.

Conclusions:
We found increased protein expression of MAP2K3 and phosphorylation of p38 in cardiac hypertrophy positively associated with male obese human hearts.
Abstract 20: Novel Transfer of Low Viscosity Cyan Methacrylate on the Surfaces of the Sheep Heart Model and Paint Brush Technique
Authors: Arokiaraj Mark
Published: June 2021

Aim:
To develop a novel technique for the transfer of cyan methacrylate on the inner and outer surfaces of the sheep heart model.

Methods:
Low viscosity ultraviolet curable cyan methacrylate was mixed with commercially available dye to demonstrate the ability of transfer on the outer and inner surfaces of the in-vitro sheep heart model. 0.5 ml of cyan methacrylate was mixed with 0.2 ml of dye, and the material was injected over the surface of the heart in dry air, and it was allowed to fix for 2-3 seconds. Subsequently, the whole preparation was immersed in saline, and it was vigorously shaken to remove unbound compound. The similar experiment was performed without cyan methacrylate. By visual assessment, a significant quantity of the compound attaches to the surfaces of the heart compared to the dye alone, which was visualized after washing. After that, various techniques were investigated for efficient transfer of the compound on the heart surfaces soaked in saline. Of the various techniques paintbrush technique for the targeted transfer was effective (figure below). Also targeted areas like left atrial appendage, left ventricular inner surfaces at the origin of papillary muscle and left ventricular apex were some of the areas investigated successfully. The ultraviolet (UV) light source was a pen shaped device. It was observed that after ultraviolet treatment micro precipitates forms. The experiment was repeated in 3 different samples to observe the techniques.

Conclusion:
There is potential for transfer of low viscosity, UV curable cyan methacrylate on the surfaces of the heart. This could of potential to transfer growth factors or protein molecules attacked to cyan methacrylate.
Abstract 217: Long Non-coding RNA Myocardial Infarction-associated Transcript (MIAT) Protects Cardiac Progenitor Cells From Oxidative Stress-induced Cell Death
Authors: Yang Liu , Yu Yang , Arnone Baron , Boriboun Chan , Shi Jiawei , Qin Gangjian
Published: June 2021

Background:
Long non-coding RNAs (lncRNAs) are an emerging class of RNAs with no or limited protein-coding capacity; a few of which have recently been shown to regulate critical biological processes. Myocardial infarction-associated transcript (MIAT) is a conserved mammalian lncRNA, and single nucleotide polymorphisms (SNPs) in 6 loci of this gene have been identified to be strongly associated with the incidence and severity of human myocardial infarction (MI). However, whether and how MIAT impacts on the pathogenesis of MI is unknown.

Methods & Results:
Quantitative RT-PCR analyses revealed that MIAT is expressed in neonatal mouse heart and to a lesser extent in adult heart. After surgical induction of MI in adult mice, MIAT starts to increase in 2 hours, peaks at 6 hours in atria and 12 hours in ventricles, and decreases to baseline at 24 hours. Fluorescent
in situ
hybridization (FISH) revealed a slight increase in the number of MIAT-expressing cells in the infarct border zone at 12 hours post-MI. Moreover, qRT-PCR analyses of isolated cardiac cells revealed that MIAT is predominantly expressed in cardiosphere-derived cardiac progenitor cells (CPCs). Treatment of CPCs with H
2
O
2
led to a marked upregulation of MIAT, while knockdown (KD) of MIAT resulted in a significantly impaired cell survival
in vitro
with H
2
O
2
treatment and
in vivo
after administered in the ischemic/reperfused heart. Notably, bioinformatics prediction and RNA immunoprecipitation identified FUS (fused in sarcoma) as a novel MIAT-interacting protein. FUS-KD CPCs displayed reduced cell viability and increased apoptosis under oxidative stress. Furthermore, MIAT overexpression enhanced survival of WT CPCs but not FUS-KD CPCs, suggesting that the protective role of MIAT is mediated by FUS.

Conclusions:
MIAT interacts with FUS to protect CPCs from oxidative stress-induced cell death.
Abstract 222: Sigmar1 Mediates Mitochondrial Autophagy and Protects the Heart Against Ischemia/Reperfusion Injury
Authors: Bhuiyan Md. Shenuarin , Alam Shafiul , Green Jonette M. , Orr A. Wayne , James Jeanne , Osinska Hanna , Karch Jason , Molkentin Jeffery D. , Blaxall Burns C. , Lorenz John N. , Sadoshima Junichi , Robbins Jeffrey
Published: June 2021

Rationale:
Sigma 1 receptor (Sigmar1) is a highly expressed mitochondrion-associated ER membrane resident protein in different cell lines. We recently reported that Sigmar1 is highly expressed in cardiomyocytes but it’s molecular functions and role in the stress response still remains unknown.

Objective:
We investigated the functional role of Sigmar1 in mediating mitochondrial autophagy, mitochondrial fission and effects on stress resistance in the heart.

Methods and Results:
Subcellular fractionation and biochemical experiments confirmed Sigmar1 expression in the mitochondria, where it resides as an integral mitochondrial outer membrane protein. Sigmar1 overexpression induced mitochondrial fission, increased autophagosome formation and autophagic flux in cardiomyocytes. Similarly, cardiac specific Sigmar1 transgenic (Tg) mice showed increased levels of mitochondrial fission and mitochondrial autophagy without adverse effects. Conversely, Sigmar1 knockdown induced both mitochondrial elongation and accumulation of damaged mitochondria, whereas autophagosome formation and autophagic flux were reduced at baseline and in response to glucose deprivation in cardiomyocytes. Parallel studies using Sigmar1 knockout mice showed increased accumulation of abnormal mitochondria and significantly altered cardiac contractility. To define the functional significance of Sigmar1 in the cardiac stress response, we subjected the mice to ischemia/reperfusion (I/R) injury. Sigmar1 Tg mouse showed reduced infarct size, protected from I/R-injury induced adverse cardiac remodeling, and improved cardiac function associated with enhanced mitochondrial autophagy even 12 weeks after reperfusion injury. In contrary, knockdown of Sigmar1 evoked mitochondrial dysfunction, accumulation of abnormal mitochondria, enhanced adverse cardiac remodeling, aggravated cardiac dysfunction and increased susceptibility to I/R-injury.

Conclusions:
Our findings suggested that Sigmar1 is an integral mitochondrial outer membrane protein dispensable for constitutive mitochondrial quality control in normal hearts. Sigmar1 regulates mitochondrial autophagy to protect the heart against I/R injury-induced cardiac remodeling and dysfunction.
Abstract 224: LAMP-2 Deficiency Promotes Mitochondrial Damage in Danon Disease
Authors: Hashem Sherin , Gault Emily , Murphy Anne , Chi Neil , Adler Eric
Published: June 2021

Danon disease is a familial cardiomyopathy associated with impaired autophagic flux due to mutations in the gene encoding lysosomal associated membrane protein type 2 (LAMP-2). The majority of patients die from progressive heart failure in their twenties, yet to date the mechanisms by which LAMP-2 deficiency leads to cardiomyocyte dysfunction and death are not fully understood. To study the underlying molecular pathogenesis, we created induced pluripotent stem cell-derived cardiomyocytes (hiPSC- CMs) from patients with different
LAMP-2
mutations, and reported an increase in apoptosis and mitochondrial oxidative stress. In Danon hiPSC-CMs, undigested mitochondria within double-membrane autophagosomes were frequently observed suggesting impaired mitophagy. Danon hiPSC-CMs displayed increased translocation of PARKIN and p62 on mitochondria, suggesting increased mitochondrial damage. When mitochondrial bioenergetic profile was assessed, Danon hiPSC-CMs showed a severe impairment in mitochondrial respiratory capacity even when offered various oxidizable substrates, suggesting global mitochondrial damage. In addition, Danon hiPSC-CMs demonstrated excessive mitochondrial fragmentation and low mitochondrial membrane potential.
Lamp-2
knockout (KO) mice recapitulated key pathological features observed in Danon patients and their hiPSC-CMs.
Lamp-2
KO mouse hearts exhibited abnormal mitochondria, increased mitochondrial damage, and impaired mitophagy. Interestingly, altered mitochondrial respiration was observed in
Lamp-2
KO mouse hearts before the onset of significant impairment in contractile function. In summary, we have modeled Danon disease using hiPSC-CMs from patients with
LAMP-2
mutations as well as
Lamp-2
KO mice, allowing us to gain mechanistic insight into the pathogenesis of this disease. We demonstrate that LAMP-2 deficiency leads to impairment in mitophagic flux, accumulation of damaged mitochondria, and deterioration of mitochondrial function. Understanding the mechanisms underlying cardiomyocyte dysfunction will have important implications for the treatment of Danon disease as well as a various cardiovascular disorders associated with impaired autophagy.
Abstract 225: Assessing Sunitinib Cardiotoxicity Using Engineered Cardiac Microtissues
Authors: Truitt Rachel E , Ky Bonnie , Margulies Kenneth B
Published: June 2021

Introduction:
Sunitinib is an oral tyrosine kinase inhibitor that effectively treats a variety of solid tumors but carries a significant risk of hypertension and left ventricular (LV) dysfunction. Although AMP-K inhibition may be central to the development of cardiotoxicity, these mechanisms remain incompletely understood as does the pathogenic role of hypertension in the development of LV dysfunction. We hypothesized that sunitinib results in dose- and duration-dependent changes in cardiac energetics that are worsened by increased afterload.

Methods:
These studies employed the use of a cardiac microtissue (CMT) model where neonatal rat cardiomyocytes and fibroblasts self-assemble on microfabricated PDMS cantilevers. With these CMTs, afterload can be effectively modified (stiff vs. soft) through variations in the elastic modulus of the cantilevers. Caspase 3/7 activation and ATP levels were determined using luminescence assays. Spontaneous beating rate was assessed by counting the number of beats during a 1 min interval. Mitochondrial membrane potential (MMP) was measured by labeling cells with TMRM and analyzed via flow cytometry.

Results:
CMTs treated with as little as 1uM sunitinib for 4hrs demonstrated a nearly two-fold increase in activated caspase 3/7 compared to control (DMSO treated) CMTs. Moreover, there was a 2-fold increase in caspase activation when CMTs were cultured on stiff versus soft cantilevers (p=0.01). Dose - dependent decreases in spontaneous beating rate following treatment with 1uM sunitinib (p>0.05) or 10uM sunitinib (p<0.0001) were observed (p-values are compared to control). MMP decreased with 1uM sunitinib treatment after 4hr (28±2.7% vs. 22±2.1% for control, n= 2x10
4
, p<0.05) and 8hr (22±2.2% vs. 27±1.4% for control, n= 2x10
4
, p<0.05). ATP levels after 24hr of 1uM sunitinib treatment were also decreased relative to control (66.2± 2.4 pg vs. 53.4±3 pg; p=0.01).

Conclusions:
These findings demonstrate that the CMT model is a robust model to study sunitinib cardiotoxicity, and cardiotoxicity is a function of sunitinib dose, duration and afterload. Experiments with CMTs comprised of human iPS derived cardiomyocytes (iPS-CMs) are ongoing to understand the relevance of these findings in human cardiomyocytes.
Abstract 23: p53 Mediates
de novo
Cardiomyocte Differentiation From c-kit Positive Cardiac Progenitor Cells
Authors: Chen Zhongming , Zhu Wuqiang , Yellamilli Amritha , Bender Ingrid , van Berlo Jop H
Published: June 2021

The heart is arguably the least regenerative organ. Nonetheless, cardiac progenitor cells (CPCs) can readily be isolated from the adult heart. Moreover, we recently demonstrated that c-kit+ CPCs are able to generate de novo lineages of cardiomyocytes, endothelial cells and fibroblasts, albeit at low levels. In the present study, we assess if different pathophysiological stimuli could enhance cardiomyocyte lineage differentiation by CPCs. We first established that CPCs are heterogeneous in vivo and can be classified into three main populations based on the respective gene expression profiles: uncommitted, vascular and cardiogenic CPCs. Next, we show that transverse aortic constriction (TAC) surgery increased CPC derived cardiomyocytes by 3-fold, and enhanced non-cardiomyocyte lineages as well. Interestingly, anthracycline-induced cardiomyopathy increased CPC-derived cardiomyocytes by 35-fold. Immunostaining showed that doxorubicin stimulates p53 expression within CPCs and selectively enhanced CPC-derived cardiomyocyte lineage. The selective p53 inhibitor, pifithrin α, completely blocked the doxorubicin-mediated increase in de novo cardiomyocyte formation. Administration of p53 Activator III, RITA (reactivation of p53 and induction of tumor cell apoptosis), was sufficient to induce CPC-derived cardiomyocyte differentiation. These results demonstrate that anthracyclines can increase de novo cardiomyocyte differentiation from CPCs through activation of the p53 pathway. Ultimately, these findings might lead to new therapies for cardiac regeneration by inducing cardiomyocyte differentiation by endogenous CPCs.
Abstract 231: Improving Physical and Mental Health Through a Community-based, Multi-disciplinary Obesity Reduction Program Promoting Social Support and Adherence
Authors: Lemstra Mark
Published: June 2021

Background:
The Healthy Weights Initiative is a community-based, multi-disciplinary obesity reduction program that was developed after 153 community consultations and is delivered for free to obese adults. Social support was an important component and strongly promoted in the program. Each participant was asked to attend with “buddy” and to complete a social support contract.

Methods:
The initial 12 weeks of the program offered 60 group exercise sessions, 12 group cognitive therapy sessions, and 12 group dietary sessions with licensed professionals. During the second 12 week period, maintenance therapy includes 12 group exercise sessions.

Results:
Altogether, 234 people have completed the program (79.1%). Mean objective reductions included 12.6 lbs of body fat, 3.1 inches form the waist, 2.6 inches from the hips, blood cholesterol by 0.3 mmol/L, systolic blood pressure by 5.2 mmHg and diastolic by 2.5 mmHg. Depressed mood prevalence decreased from 45.7% to 11.7%; 84.5% of those who completed the program had improvements in health-related quality of life (HRQL), and significant increases in mean scores on eight dimensions of health were also observed (such as physical functioning, which increased by 15.5%). Using binary logistic regression, independent risk factors for not completing the program (no social support contract and lower education); not improving depressed mood after completion of the program (low general health); and not improving HRQL after completion of the program (smoking and no buddy in program) were determined.

Conclusion:
Comprehensive obesity reduction programs can be effective when there is extensive consultation at the community level and social support at the individual level. Successful obesity reduction programs can have important cardiovascular health implications by reducing blood pressure, blood cholesterol, and improving cardiovascular fitness.
Abstract 235: Integration of Patient-specific Computational Hemodynamics and Vessel Wall Shear Stress Into MRI Diagnosis of Vascular Diseases
Authors: Yu Huidan W , Chen Xi , Wang Zhiqiang , Chen Rou , Lin Chen , Kralik Stephen F , Patil Raveena
Published: June 2021

The research objective is to expand the capability of current MRI imaging technique in assessing the overall risk and predicted outcomes of atherosclerotic diseases through the quantification of individual patient-specific hemodynamics, including flow, pressure, and wall-shear stress. A unique computational modeling technique, named InVascular, is integrated directly into clinical MRI scanners as the extension of the image reconstruction and post-processing pipeline so that velocity, pressure, vorticity, and WSS can be available immediately with other diagnostic images. InVascular is a unified and GPU accelerated computation platform to model and simulate patient-specific hemodynamics and flow-vessel interaction based on MRI imaging data. In this study, we validate the efficiency and accuracy of InVascular through quantitative hemodynamics in vertebral and carotid arteries. A group of five volunteers participated in the scanning of high resolution time-of-flight (TOF) and low resolution electrocardiogram (ECG) gated phase contrast (PC) MR angiogram (MRA) images. For each case, InVascular successively processes the images to get vessel geometry from TOF MRA and velocity slices from PC MRA and solve the fluid dynamics inside the carotid arteries with PC MRA measured velocity at the inlet and outlet (Fig. 1 a-c). The velocity profiles from Invascular and PC MRA are compared at the same location (Fig. 1 d-g ). We conclude that integration of MRAs and InVascular can well captured the velocity fields as MRI measures. InVascular can provide quantitative pressure and WSS (Fig. 1h ) information as well.
Abstract 252: Chronic Relaxin Treatment Does Not Augment Angiotensin II Induced Aortic Pathologies in Mice
Authors: Dajee Maya , Johns Douglas G , Howatt Deborah A , Balakrishnan Anju , Seganish Michael , Da Cunha Valdeci , Moorleghen Jessica J , Lu Hong
Published: June 2021

Introduction:
Human relaxin is being tested clinically as an acute treatment for heart failure. The effects of relaxin on extracellular matrix remodeling, among several mechanisms, may provide myocardial benefit, however, the effects of relaxin on vascular extracellular matrix-related pathology has not been examined. Human aortic aneurysm is characterized by extracellular remodeling, as well as inflammation, thrombus formation and rupture. Studying relaxin in a model of aortic aneurysm might provide insight into mechanisms of the disease but also will assess whether such agents impact the incidence/severity of aortic aneurysms (AA).

Objectives:
The purpose of this study was to test the effects of relaxin, an anti-fibrotic agent, on angiotensin II (AngII)-induced AA, which is characterized by enhanced activation of matrix metalloproteinases (MMPs); additionally this model mimics human pathology.

Methods and Results:
Vehicle or relaxin was co-infused with AngII (1.44 mg/kg/d) in either C57BL/6 mice or apoE null mice through mini osmotic pumps for 28 days. Dose-dependent plasma relaxin concentrations were noted (12-60ng/ml). Relaxin did not augment AngII-induced aortic rupture rate, the most severe consequence of AA. Maximal ex vivo diameters of suprarenal aortas were measured for abdominal aortic dilation, and ascending aortic area was measured using an en face method to determine ascending aortic dilation. Maximal aortic diameter of suprarenal aortas did not show significant difference in relaxin and AngII infused mice with either mouse strain. Relaxin also had no effect on ascending aortic dilation in both mouse strains.

Conclusion:
Chronic relaxin infusion for 28days does not augment aortic aneurysms in AngII-infused mice, suggesting relaxin might not impact other matrix-dependent vascular pathologies, if used as a treatment for heart failure, although additional studies are warranted. Ongoing studies interrogate the potential benefits of molecular mechanisms of relaxin response in the AngII-induced aortic pathologies.
Abstract 254: A Novel Mechanism for Aberrant Aldosterone Production in Heart Failure
Authors: Endo Jin , Yamashita Kaoru , Yoshida Naohiro , Ito Kentaro , Ichihara Atsuhiro , Fukuda Keiichi , Sano Motoaki
Published: June 2021

Objective:
Aldosterone production by the adrenal gland is mainly regulated by the renin-angiotensin system (RAS), which is normally suppressed by a high-salt diet. However, plasma aldosterone levels are elevated in proportion to the severity of heart failure even if caused by high-salt intake. The inappropriate increase in plasma aldosterone level could not be diminished by pharmacological blockade of the RAS. We investigated the cellular mechanism underlying the aberrant aldosterone production in heart failure beyond the activation of RAS.

Methods and Results:
Dahl-salt sensitive rats fed high salt diet developed malignant hypertension, resulting in heart failure with increased plasma aldosterone level. The gene expression pattern of the adrenal gland from heart failure rats showed strikingly upregulated β3-adrenergic receptor (β3-AR) expression. Immunohistochemical staining confirmed that heart failure increased β3-AR expression in zona glomerulosa cells in adrenal gland of Dahl-salt sensitive rats. And the staining of pimonidazole, a hypoxia probe, revealed that the outer layer of adrenal cortex in the heart failure rat was under hypoxic condition. Hypoxia and DMOG, a chemical stabilizer of hypoxia-inducible factor, strongly induced β3-AR mRNA expression in H295R human adrenocortical carcinoma cells. Hypoxia-reoxygenation stimuli induced aldosterone production from H295R cells, which was blocked by SR59230A, a selective β3-AR antagonist, but not by losartan, an angiotensin II receptor blocker (ARB). Hypoxia endowed catecholamine responsiveness in H295R cells by increased β3-AR expression, wherein β3-AR agonist increased the phosphorylation levels of hormone-sensitive lipase (HSL) through activation of ERK pathway.

Conclusions:
In the present study, we demonstrated that the β3-AR induction in adrenal gland by hypoxia played an important role in aberrant aldosterone production in heart failure. The catecholamine stimuli through β3-AR in adrenocortical cells caused the ERK-mediated phosphorylation of HSL, suggesting that the HSL promotes lipolysis to supply substrates for the aldosterone biosynthesis.
Abstract 259: KChIP2 is a Key Transcriptional Regulator of Cardiac Excitability Under Normal and Pathogenic Conditions
Authors: Nassal Drew M , Wan Xiaoping , Liu Haiyan , Maleski Danielle , Ramirez-Navarro Angelina , Moravec Christine S , Ficker Eckhard , Laurita Kenneth R , Deschênes Isabelle
Published: June 2021

Introduction:
Arrhythmogenesis is the primary cause of death in patients with acquired heart disease, and is the consequence of profound dysregulation of both depolarizing and repolarizing currents. Reduction in expression of the auxiliary subunit K+ channel interacting protein 2 (KChIP2), which regulates the transient outward K+ current (Ito), is a common and early event in many cardiac pathologies. Notably, transcriptional changes observed in heart disease can be elicited through direct KChIP2 silencing without disease signaling, suggesting novel transcriptional capacity for KChIP2.

Methods and Results:
A miRNA microarray was conducted on neonatal rat ventricular myocytes (NRVM) following in vitro silencing of KChIP2, identifying the miR-34 family as a potential transcriptional target of KChIP2. qPCR confirmed reduction in miR-34b/c when over-expressing KChIP2 and increase following silencing. Luciferase assays conducted on the promoter for miR-34b/c reinforced KChIP2 repression on the promoter, while chromatin immunoprecipitation identified direct interaction of KChIP2 on the promoter. Previous studies show modified expression of KChIP2 can lead to changes in several ion channel subunits. Therefore, we investigated if this was the consequence of KChIP2 regulation via miR-34. Expression of miR-34b/c precursors reduced transcript levels of Nav1.5 and Navβ1, and reduced protein levels for Kv4.3, resulting in loss of INa and Ito. To determine the relevance in disease signaling, NRVMs were exposed to 100 μM phenylephrine for 48 hrs, significantly reducing KChIP2, Nav1.5, Navβ1, and Kv4.3, while elevating miR-34b/c. Maintaining KChIP2 expression by adenovirus or blocking miR-34 activity with antagomirs rescued the changes in channel expression. Consequently, miR-34 inhibition rescued the induction of sustained arrhythmias observed in a 2D culture of myocytes through the maintenance of cardiac excitability.

Conclusion:
Collectively, these observations identify dysregulation of the KChIP2/miR-34 axis as a nodal event in the development of electrical dysfunction that characterize cardiac pathologies, and further identifies miR-34 as a viable therapeutic target for managing arrhythmogenesis in patients with heart disease.
Abstract 262: A Rat Model of Heart Failure With Preserved Ejection Fraction Induced by Pressure Overload
Authors: Sun Shu-Yu , Campbell Barry , Da Cunha Valdeci , Hong Xuening , Huang Chin-hu , Kennan Richard , Ping Xiaoli , Shen Xiaolan , Small Kersten , Yang Liming , Zhao Huawei , Zheng Shuqin , Madwed Jeffrey B
Published: June 2021

The absence of animal models displaying clinical phenotypes of HFpEF represents a challenge to the pharmaceutical industry in the hope of identifying novel treatments that would benefit patients with this syndrome. We, at Merck, examined the supracoronary aortic banding (SAB) in rats as a potential model and set out to characterize its phenotypes. Juvenile Sprague-Dawley rats were used; a hemostatic clip with an opening of 0.7 mm was placed around the ascending aorta to elicit a pressure overload to the LV. Echocardiography was performed every 3 weeks and invasive hemodynamics was measured at week 9 to evaluate cardiac structural and functional effects; heart and lungs were harvested at the end for morphological and histological analysis. Nine weeks after SAB, LV hypertrophy and dysfunction were apparent, as manifested by increases in LVEDP, left atrial pressure, plasma BNP, LV and lung weights; LV relaxation time constant (Tau) and mitral early to late inflow ratio (E/A) were also increased; while EF and FS were preserved (see table, mean ± SEM). In addition, SAB rats had elevated mean pulmonary pressure (sham 14 ± 1.7 vs. SAB 37 ± 4.4 mm Hg, p<0.05) with an abnormal transpulmonary gradient (sham 12 ± 2.5 vs. SAB 23 ± 0.1 mm Hg, p<0.05), indicating comorbidity of pulmonary hypertension. Finally, piloerection, dyspnea and loss of muscle mass were evident and exercise tolerance measured by treadmill test was decreased in SAB rats. In summary, rats undergoing SAB exhibit many of the clinical phenotypes of HFpEF. Further work includes examining the effects of novel therapies in SAB rats on improving these clinical phenotypes of HFpEF and expanding the model to study morbidity and mortality.
Abstract 278: Domain-specific Roles for GRK2 in Cardiac Hypertrophy and Heart Failure
Authors: Schumacher Sarah M , Gao Erhe , Chuprun J. Kurt , Koch Walter J.
Published: June 2021

During heart failure (HF), cardiac levels and activity of the G protein-coupled receptor (GPCR) kinase (GRK) GRK2 are elevated and contribute to adverse remodeling and contractile dysfunction, while inhibition via a carboxyl-terminal peptide, βARKct, enhances heart function and can prevent HF. Mounting evidence supports the idea of a dynamic “interactome” in which GRK2 can uncouple GPCRs via novel protein-protein interactions. Several GRK2 interacting partners are important for adaptive and maladaptive myocyte growth; therefore, an understanding of domain-specific interactions with signaling and regulatory molecules could lead to novel targets for HF therapy. For instance, GRK2 contains a putative amino-terminal
R
egulator of
G
protein
S
ignaling (RGS) domain (βARKrgs) that directly interacts with Gαq and inhibits signaling. Previously, our lab investigated cardiac-specific transgenic expression of a fragment of this RGS domain (βARKnt), that did not reduce acute hypertrophy after pressure overload or demonstrate RGS activity
in vivo
against Gαq-mediated signaling. In contrast, βARKnt induced hypertrophy and elevated β-adrenergic receptor (βAR) density without altering agonist-induced contractility or adenylyl cyclase activity, due to a compensatory increase in GRK2 activity. Importantly, βAR downregulation in response to chronic agonist administration was attenuated by βARKnt expression, indicating a novel regulation of βAR receptor density. Herein, we investigated the effect of βARKnt expression during chronic pressure overload post trans-aortic constriction (TAC). Echocardiographic analysis revealed increased posterior wall thickness and left-ventricular mass 4 weeks post-TAC compared to non-transgenic littermate controls. Importantly, despite enhanced hypertrophy, the progression to HF was inhibited in βARKnt mice 14 weeks post-TAC. Histological analysis of interstitial fibrosis and cross-sectional area is underway to determine alterations in maladaptive remodeling. Further, cardiomyocyte signaling and βARKnt protein-binding partners are a focus, since our data indicate that βARKnt-mediated regulation of βAR density may provide a novel means of cardioprotection during pressure-overload induced HF.
Abstract 279: Stromal Cell-derived Factor-1 Prevents From Diabetic Cardiomyopathy via Cxcr7/ampk-mediated Nrf2 Activation in Type 2 Diabetic Mice
Authors: Wang Shudong , Gu Junlian , Yan Xiaoqing , Chen Jing , Chen Jun , Cai Lu , Liu Quan , Zheng Yang , Tan Yi
Published: June 2021

We have demonstrated that stromal cell-derived factor 1(SDF-1) protects against palmitate-induced cardiac cell death via CXCR7-mediated activation of AMPK signaling (Diabetes 62:2545-2558, 2013). Whether SDF-1 prevents diabetic cardiomyopathy (DCM) and what the underlying mechanism has not been addressed. Here we evaluated the prevention of SDF-1 from DCM in a high fat diet plus streptozotocin (HFD/STZ)-induced type 2 diabetic model in C57BL/6J mice. After 1 month on HFD, the HFD-fed mice were injected with one low dose STZ (100mg/kg body weight, ip). Five days after STZ injection, mice with blood glucose levels ≥250 mg/dl were defined as diabetic. In parallel, the age-matched normal diet-fed mice injected with a same volume of vehicle were used as control. After onset of diabetes, the mice were maintained on HFD or normal diet for another 4 months with or without SDF-1 treatment. Then cardiac function was assayed again, and the mice were sacrificed and cardiac tissue collected for cardiomyopathic index assay. We found that 1 month HFD feeding induced a significant insulin resistance without effect on cardiac function, but continued HFD feeding after STZ injection significantly increased insulin resistance and blood glucose, as well as blood insulin, triglyceride and cholesterol levels. Furthermore, HFD/STZT significantly impaired cardiac function, which were accompanied by increased cardiac inflammation, oxidative stress and fibrotic remodeling. Treatment with SDF-1 dose-dependently prevented diabetes-induced cardiac dysfunction, inflammation and remodeling, but without significant effects on the above mentioned other pathophysiological changes. Mechanistic study demonstrated that diabetes significantly inhibited the function of AMPK and Nrf2, and the expression of CXCR7, but not CXCR4; while treatment with SDF-1 significantly preserved AMPK and Nrf2 function and CXCR7 expression. Knockout CXCR4 did not affect SDF-1 preservation of AMPK and Nrf2 function, but SiRNA knockdown of AMPK resulted in blockade of SDF-1 preservation of Nrf2 function. These results indicate that SDF-1 prevents from DCM via CXCR7/AMPK-mediated Nrf2 activation in type 2 diabetic mice.