tbuckley/cabot-search huggingface dataset:
Abstract Tu067: ROCK2 Specific Inhibition Prevents Isoproterenol Induced Takotsubo Cardiomyopathy in Mice
Authors: Daniel Fehrenbach, Jean Wassenaar, Charles Smart, Lin Zhong, Francis Miller, Meena Madhur
Journal: Circulation Research (2024)

Takotsubo Cardiomyopathy (TCM) is an acute, life-threatening condition characterized by a drop in ejection fraction (EF) and apical ballooning of the left ventricle (LV). TCM is most frequently observed in women and is associated with an emotional or physical stressor. Recent work has demonstrated that the Rho kinase ROCK2 contributes to heart failure phenotypes in mice in other models of heart failure. The role of ROCK2 in TCM is unknown. We tested the hypothesis that ROCK2 inhibition attenuates isoproterenol (ISO)-induced TCM in mice. To induce TCM, male and female mice were given a single dose of 200mg/kg ISO, i.p. and cardiac function was assessed by conscious echocardiography. The effect of ISO overdose on acute heart failure over a 7-day time course was determined followed by an intervention study in which mice were treated with the ROCK2 specific inhibitor KD025/belumosudil (50mg/kg, i.p.) concurrent with heart failure induction. Though ISO induced a drop in EF in both sexes after 24 hours, females exhibited lower EF (46.64±3.11%) than males (68.36±2.62, p&lt;0.0001) mirroring the increased prevalence of clinically defined TCM in females in humans. Female mice were used for the remaining experiments. Invasive hemodynamics revealed reduced contraction and relaxation (dp/dt max and dp/dt min) as well as an increased relaxation time constant Tau 24 hours after ISO injection. A time course study demonstrated that EF begins to normalize by Day 3 and is restored to baseline levels by Day 7. Importantly, ROCK2-specific inhibition prevented ISO-induced drop in EF, thinning of the LV wall, and LV volume expansion. Here, we demonstrate that female mice are more susceptible to ISO-induced cardiac dysfunction. Furthermore, we show that ROCK2 activation is necessary for the development of TCM in mice. These results suggest that FDA-approved KD025/belumosudil could potentially be repurposed for treatment in TCM.
Abstract Tu140: Evaluation of the effects of mitoquinone on doxorubicin-induced acute cardiac damage
Authors: Meagan Lyons, Juliet Melnik, Lloyd Yan, Kelly Annie Mercado, Sasha Ghofrani, Nelma Malik, Kimberly Dawes, Gabby Ghasb, Jonathan Amora, Lindon Young, Robert Barsotti, Qian CHEN
Journal: Circulation Research (2024)

Background: Doxorubicin (DOX)-induced cardiotoxicity remains a high priority to overcome due to irreversibility and lack of abundant preventions. DOX damages the heart tissue principally by overproduction of mitochondrial superoxide and intercalation with the topoisomerase II in the nucleus. Ultimately, cardiac cells become apoptotic, and heart function declines. Our lab has shown that Mitoquinone (MQ), a mitochondria-targeted antioxidant, improves H9c2 cell viability in the presence of DOX. We hypothesize that MQ would counteract DOX to preserve cardiac cells and function in isolated rat hearts, possibly by reducing mitochondrial superoxide production and intracellular accumulation. Methods: H9c2 cells (passages &lt; 18) were incubated with DOX (40 µM) with or without MQ (0.005 μM - 5 μM) for 24 hours before measuring mitochondrial superoxide anion levels and DOX intracellular accumulation. Furthermore, heart rate (HR) and left ventricular end-systolic pressure (LVESP) were recorded before and after a 60-minute infusion of DOX or DOX with MQ in isolated rat hearts. TUNEL staining was performed on heart tissue slides to assess apoptosis. All the data was expressed as a ratio between treatment and DOX alone or between final to initial values. Statistical significance was evaluated using ANOVA with Student Newman Keuls post-hoc analysis. Results: Hearts perfused with DOX (25 µM, n = 5) displayed lower final LVESP with 39 ± 5% of the initial baseline. By contrast, hearts treated with 0.25 µM MQ (n = 4) or 1 µM MQ (n = 5) manifested exhibited a higher final LVESP to 44 ± 8% and 90 ± 6% of initial baselines, respectively. Additionally, MQ administration significantly reduced cardiac cell apoptosis to 8 ± 1% (0.25 µM, n = 2) and 6 ± 1% (1 µM, n = 5, p &lt; 0.05) when compared to DOX (88 ± 2%, n = 4). In contrast to DOX, MQ (1 µM, n = 4) showed 27 ± 9% (n = 4) reduction in mitochondrial superoxide levels and 54 ± 6% (n = 6, p &lt; 0.05) less intracellular DOX deposition in H9c2 cells. Conclusion: This study suggests acute DOX administration to the heart severely compromises cardiac systolic function with higher induction of cell apoptosis. MQ exerts cardio-protection with higher systolic function and cell viability by attenuating mitochondrial superoxide levels and intracellular DOX accumulation.
Abstract Tu020: Contrasting effect of Ox-PAPC and PC-KLH in atherosclerotic plaques or peripheral blood T cell, macropahges and dendritic cell activation
Authors: Johan Frostegard, Sabbir Ahmed, Johnny Steuer, Peter Gillgren, Mizanur Rahman
Journal: Circulation Research (2024)

Background: Atherosclerosis is a chronic inflammation characterized by accumulation of immune cells, oxidized low density lipoprotein (OxLDL) and dead cells in the lesions, both of the latter have the hapten phosphorylcholine (PC) as an important antigen. PC is also exposed on microorganisms and other compounds and needs a carrier to be antigenic. Antibodies against PC (anti-PC) ameliorates deleterious effects of both OxLDL and dead cells. Oxidized-1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphatidylcholine (Ox-PAPC) is a major compound in the lipid moiety of OxLDL exposing PC. Methods: We studied effect of Ox-PAPC and PC with keyhole limpet cyanid (KLH) on T cells, dendritic cells (DC) and macrophages (MQ). We examined the influence of Ox-PAPC and PC conjugated to KLH (PC-KLH) on T cells, DC, and MQ from peripheral blood or atherosclerotic plaques. We investigated phospholipid and glutathione (GSH) levels in the blood plasma of healthy controls (n=30) and patients (n=30). Results: Ox-PAPC, in contrast to PAPC and PC-KLH, independently of DC, induced proinflammatory T cell activation, which was inhibited by mito tempo, inhibitor of reactive oxygen species (ROS). Ox-PAPC triggered inflammatory activities while reducing metabolic activities in T cells. PC-KLH, on the other hand, reduced CD86 and CD11C, proinflammatory cytokines, and induced anti-inflammatory IL-10 in DC. Ox-PAPC induced ROS, GSH, and proinflammatory MQ activation, inhibited by Mito tempo or GSH inhibitor. Additionally, Ox-PAPC, in the presence of PCSK9, elevated CD86 expression and lipid peroxidation. Patients exhibited higher GSH levels comapre to control groups, associated with phospholipid levels. Conclusions: The study reveals the differential effects of Ox-PAPC on various immune cell types, showcasing T cell activation independent of DC and heightened macrophage activation in the presence of PCSK9. Ox-PAPC-induced redox imbalance regulates T cell activation, apoptosis, and MQ activation, potentially rendering plaques more vulnerable and contributing to lesion formation and thus plaque rupture. In contrast, PC-KLH had anti-inflammatory effects, inducing anti-inflammatory cytokines from T cells, DC and MQ. PC-effects thus vary from proinflammatory to anti-inflammatory depending on presentation. Inhibition of OxPAPC effects by agents such as anti-PC could thus be anti-atherogenic, while on the other hand PC presented on a protein as KLH could be anti-atherogenic and anti-inflammatory itself.
Abstract Or107: MyBP-HL nonsense mutations fail in sarcomere incorporation leads to modifications in myocyte contraction trough calcium modifications.
Authors: Alejandro Alvarez-Arce, Geena Fritzmann, Kelly Araujo, Alexandra Pena, Lucas Wittenkeller, David Barefield
Journal: Circulation Research (2024)

Introduction: Heart function depends on the cardiomyocyte contractile apparatus and proper sarcomere protein expression. Mutations in sarcomere genes cause inherited forms of cardiomyopathy and arrhythmias, including atrial fibrillation. AF is the most common cardiac arrhythmia and is associated with ischemic stroke, heart failure, and substantial morbidity and mortality. Recently, our group described a novel sarcomere component, myosin binding protein-H like (MyBP-HL), that is mainly expressed in the cardiac atria. MyBP-HL is composed of two immunoglobulin domains and one fibronectin domain, with homology to last three C-terminal domains of cardiac myosin binding protein-C (cMyBP-C). cMyBP-C nonsense mutations prevent his incorporation into the sarcomere. C-MyBP-C truncated peptides are identified by cells as poisoning peptides, and they are rapidly degraded. cMyBP-C nonsense mutations are causative of hypertrophic cardiomyopathy (HCM). The MyBP-HL Arg255stop variant has been described in humans and has been linked to ventricular conduction system abnormalities, atrial enlargement, dilated cardiomyopathy, and atrial and ventricular arrhythmias. The gnomAD database reports 9 nonsense mutations for MYBPHL in humans (Gln29, Trp54, Arg113, Tyr123, Trp158, Trp192, Lys250, Arg255, and Tyr307). Hypothesis: MYBPHL nonsense mutations fail to incorporate into the sarcomere will lead to contractile disfunction. Results: We demonstrated that MyBP-HL overexpression showed to be able to incorporate in a similar pattern to cMyBP-C C-zone doublets. Nevertheless, nonsense mutations disrupt normal sarcomere localization. MyBP-HL nonsense mutations promote an increase in contraction amplitude and slower relaxation kinetics. Also, we observed an increase in sarcomere length. Similarly, MyBP-HL nonsense mutations lead to increase in amplitude peak of calcium transients, and reduction in velocity to maximal calcium peak. In accordance with calcium modifications, we observed through mass spectrometry an increase in calpain 6 abundance promoted by MyBP-HL nonsense mutations. Conclusion: Based on these data, we demonstrated that MYBPHL nonsense mutations prevent MyBP-HL incorporation into the thick filament. The failure of MyBP-HL truncating peptides to incorporate into the sarcomere leads to a deficiencies in myocytes contraction patterns, changes in calcium patterns and increase in degradation proteins abundance, which are related to AF prevalence.
Abstract Tu062: FLNC Deficiency Triggers Unfolded Protein Response and Leads to Dilated Cardiomyopathy
Authors: He Xuan, Jiacheng Li, Hong Lian, Yu Nie
Journal: Circulation Research (2024)

Background: The FLNC gene encodes filamin C, which is highly expressed in cardiomyocytes. Dysfunction of filamin C, due to different types of FLNC mutations, has been reported to be associated with various types of cardiomyopathy. Truncating mutations in FLNC often result in insufficient expression of filamin C, resulting in dilated cardiomyopathy (DCM). However, the specific processes via which the deficiency of filamin C causes DCM remain incompletely comprehended. Aims: To investigate the pathogenic mechanisms of DCM caused by the deficiency of filamin C in cardiomyocytes using a mouse model with cardiac-specific knockout of Flnc and to explore potential therapeutic approaches. Methods and Results: We used tamoxifen to induce cardiac-specific Flnc gene knockout in cardiomyocytes and found that mice with cardiac-specific Flnc knockout exhibited left ventricular dilation, heart failure, increased heart weight, cardiomyocyte enlargement and fibrosis, suggesting that Flnc deficiency in cardiomyocytes causes DCM. Transcriptional profiling of primary cardiomyocytes isolated on the seventh day after tamoxifen injections showed the activation of ER stress and the unfolded protein response (UPR) upon Flnc deletion. Then we measured the expression levels of proteins in UPR pathway in cardiac tissue and found that PDI and pIRE1a proteins were significantly upregulated, while there were no changes in the other two branches of UPR, including the PERK and ATF6 branches. Intraperitoneal injection of a PDI inhibitor E64FC26 significantly improved heart function and reduced fibrosis in cardiac-specific Flnc deletion mice. Conclusion: Cardiac-specific knockout of Flnc leads to DCM in mice, during which the UPR pathway is activated. Treatment with the PDI inhibitor E64FC26 improves the heart function and the fibrosis in Flnc -deficient DCM mice, offering novel perspectives on the pathological mechanisms of DCM and providing new insights into therapeutic strategies in treating DCM caused by FLNC dysfunction.
Abstract Mo119: PI3Kgamma regulates CamKII mediated phosphorylation of phospholamban and SR calcium cycling
Authors: Maradumane Mohan, Conner Witherow, Robert Papay, Yu Sun, Kate Stenson, Anushruti Ashok, Sathyamangla Naga Prasad
Journal: Circulation Research (2024)

Although phosphoinositide 3-kinase (PI3Kγ) null mice (PI3Kγ -/- ) show increased ventricular rate/contraction, it is unknown whether PI3Kγ regulates calcium recycling machinery underlying this phenotype. Primary adult cardiomyocytes from PI3Kγ -/- mice show altered sarcoendoplasmic reticulum (SR) calcium cycling following caffeine. Unexpectedly, PI3Kγ -/- cardiomyocytes showed significant reduction in phosphorylation of phospholamban (PLN) at Thr17, a key regulator of SR calcium re-uptake without changes in phosphorylation at Ser16. Furthermore, loss in PLN phosphorylation in PI3Kγ -/- cardiomyocytes was associated with augmented interaction with SR calcium ATPase (SERCA). Surprisingly, cardiomyocyte-specific overexpression of kinase-dead PI3Kγ (PI3Kγ inact ) in global PI3Kγ -/- mice (PI3Kγ inact /PI3Kγ -/- ) normalized caffeine-induced calcium re-uptake, PLN phosphorylation at Thr17 and decreased PLN-SERCA interaction. These data suggested kinase-independent function of PI3Kγ in regulation of SR calcium load and PLN phosphorylation. Since phosphorylation of Thr17 of PLN is carried out by Ca 2+ /Calmodulin dependent protein kinase (CamKII), we probed for the role of PI3Kγ in regulation of CamKII in PLN phosphorylation. Mechanistically, PI3Kγ exhibits scaffolding function in recruitment of CamKII to PLN at SR to mediate PLN phosphorylation. Furthermore, we showed that PI3Kγ directly interacts with CamKII. The study unravels a yet to be recognized kinase-independent role of PI3Kγ in regulating PLN with implications in cardiac function.
Abstract Mo111: Comparison of Pre-Clinical Models of Heart Failure with Preserved Ejection Fraction Using Multimodal Approaches
Authors: Jean Wassenaar, Pranoy Sangowdar, Charles Smart, Daniel Fehrenbach, Lin Zhong, Daniel Colvin, Francis Miller, Jeffrey Dendy, Amanda Doran, Meena Madhur
Journal: Circulation Research (2024)

There are various pre-clinical models of heart failure with preserved ejection fraction (HFpEF) designed to model different clinical phenotypes of HFpEF. However, detailed assessment and head to head comparison of diastolic function by invasive and non-invasive techniques in these models is lacking. Our goal was to comprehensively evaluate cardiac structure and function in three HFpEF mouse models – deoxycortisterone acetate (DOCA)-salt, Nω-nitro-l-arginine methyl ester high fat diet (LNAME+HFD), and aged female mice fed HFD with angiotensin infusion (Ang+HFD) – using both non-invasive and invasive methods. We employed pulse wave and tissue Doppler echocardiography, cardiac magnetic resonance imaging (CMR), and left ventricular (LV) catheterization in the commonly used C57BL/6J mouse, enabling direct comparisons between models. Cardiac hypertrophy was assessed by heart weight (HW) to tibia length (TL) ratio. The Mann-Whitney test was used for statistical testing within each model. We found that on echocardiography, all models demonstrated diastolic dysfunction with significant differences (p &lt; 0.05) or trends for significance (p &lt; 0.1) on tissue doppler peak mitral annular velocity during early filling (e’), ratio of peak early mitral flow velocity (E) to e’ (E/e’), ratio of peak early mitral flow velocity (E) to e’ (E/e’), isovolumetric relaxation time (IVRT), and E pressure half time (E PHT). CMR was notable for findings of reduced right ventricular ejection function in the LNAME+HFD group (36.1±2.7% vs 49.8±1.4% in chow controls, p&lt;0.01). On invasive hemodynamics, only DOCA-salt had a higher LV diastolic time constant (t, 11.7±1.4 vs 6.9±0.8 ms in sham controls, p=0.02) and only Ang+HFD had a higher minimum rate of pressure change in the ventricle (dP/dt min , -3506±342 vs -5268±227 in chow fed, vehicle infusion controls, p&lt;0.01). No model showed a difference in LV end diastolic pressure. For cardiac hypertrophy, we found that both DOCA-salt (8.1±0.2 vs 10.3±0.5, p&lt;0.01) and Ang+HFD (9.2±0.3 vs 11.5±0.4, p&lt;0.01) models demonstrated increased HW to TL ratio, while LNAME+HFD did not. These results highlight differences in cardiac structure and function in HFpEF mouse models and the need for multi-modal assessment in pre-clinical studies.
Abstract Tu028: NADPH Oxidase Inhibition Antagonizes Endothelial Pro-inflammatory and Pro-oxidant Signaling Resulting in Enhanced Coronary Vasorelaxation in Diabetic Models
Authors: Debolina Banerjee, Hang Xing, Ju-Woo Nho, Janelle Li, Hanisah Iddrisu, Meghamsh Kanuparthy, frank sellke, Jun Feng
Journal: Circulation Research (2024)

Introduction: NADPH oxidase (NOX) overactivation and reactive oxygen species amplification may underlie worse cardiac outcomes in patients with diabetes. Our group has shown impaired endothelium-dependent coronary vasodilation in diabetes, with restored endothelial function following NOX inhibition. Persistently high levels of oxidative stress may be driving this impaired coronary microvascular reactivity by altering small-conductance potassium channel activity. We hypothesized that NADPH oxidase inhibition would be vasoprotective through reductions in pro-oxidant and pro-inflammatory signaling. Methods: Human coronary arterial endothelial cells (HCAECs) were obtained from diabetic (D) and nondiabetic (C) patients (N = 4 per group). HCAECs were cultured in normo- or hyperglycemic conditions, with one hyperglycemic group receiving NADPH oxidase inhibitor apocynin (DT). Molecular signaling was assessed using immunoblotting. Results: Diabetes resulted in significantly increased pro-oxidant markers, including NOX 4 (p = 0.005), phosphorylated endothelial nitric oxide synthase (p-eNOS) (p = 0.007), and eNOS (p = 0.003). Pro-inflammatory Nrf2 was also increased (p = 0.0003), with a trend toward increased NFkB (p = 0.09) in D compared to C. Antioxidant peroxiredoxin (PRDX) 1 was significantly reduced in D compared to C (p = 0.02). NOX inhibition rescued these effects, with reversed trends in NOX 4 (p = 0.008), p-eNOS (p = 0.03), and PRDX 1 (p = 0.02) following apocynin treatment. NOX inhibition resulted in de novo decrease in NOX 1 in DT when compared to D (p = 0.008) and C (p = 0.02), and decrease in phosphorylated NFkB (p-NFkB) when compared to D (p = 0.02). There was a trend toward decreased total NFkB (p = 0.07) and p-NFkB:NFkB ratio (p = 0.06) and increased antioxidant thioredoxin (TXN) 2 (p = 0.05) in diabetic cells following treatment. Antioxidant superoxide dismutase (SOD) 2 (p = 0.01) and TXN 1 (p = 0.02) were increased in D when compared to C, while apocynin treatment reversed these changes (SOD 2, p = 0.0006; TXN 1, p = 0.04). There were no significant differences in NOX 2, SOD 1, catalase, PRDX 2, or PRDX 3 between the groups. Conclusions: Apocynin overall suppressed pro-oxidant and pro-inflammatory signaling in human diabetic coronary endothelial cells, resulting in improved coronary endothelium-dependent vasorelaxation. Cardioprotective strategies that incorporate NADPH oxidase inhibition in the setting of diabetes warrant further investigation.
Abstract Tu144: DIAPH1 regulates stress induced senescence in Human cardiomyocytes
Authors: Gautham Yepuri, Kaamashri Mangar, Ann Marie Schmidt, Ravichandran Ramasamy
Journal: Circulation Research (2024)

Introduction: Stress induced senescence (SIS), defined as the loss of cell physiology/function associated with increased senescence associated secretory phenotypes (SASPs), is a potential risk factor for cardiac dysfunction and failure. Human Diaphanous 1(DIAPH1), a formin molecule is known to regulate actin-cytoskeleton dynamics, trafficking of organelles and signal transduction. Our recent work demonstrated the importance of DIAPH1-MFN2 interactions in regulating actin-cytoskeleton dynamics and the mitochondria-sarcoplasmic reticulum(Mito-SR) distance thereby regulating cardiomyocyte function and pathological responses to ischemia reperfusion injury. We tested the hypothesis that DIAPH1 regulated stress induced senescence in cardiomyocytes under aging and hypoxia/reoxygenation (H/R) conditions. Methods: Human cardiomyocytes (HiPSC-CMs) from two different sources of HiPSCs which are double positive for TNT2 and TNNI3 were used. To understand the role of DIAPH1 in SIS, we have used two different approaches. HiPSC-CMs were either kept for 120days in culture or at day25 were treated with mitomycin(200nM/72hrs). DIAPH1 was silenced using shRNA approach ( shDIAPH1 ). Results: We observed increased DIAPH1 gene expression at 120 days of culture vs day 30. Interestingly, shDIAPH1 HiPSC-CMs, attenuated the number of SA-β-galactosidase positive cells, reduced expression of senescence markers SERPINE, CDKN1A, CDKN2A, TP53 , COL1A2 along with improved mitochondrial function when compared to shScr controls. In experiments where HiPSC-CMs at day 25 were treated with mitomycin to induced senescence, we observed that silencing DIAPH1 attenuated the number of SA-β-galactosidase positive cells, reduced F/G actin ratio and robustly decreased DIAPH1-MFN2 interactions as measured by DUOLINK proximity ligation assay. We probed the RNA sequencing datasets from our recent publication (Yepuri et.al., Nat. Comm. 2023) and observed that senescence markers, SASPs and cytokines including CDKN1A, H2AX, LMNA, IL1A, IL6, SERPINE were significantly down regulated in shDIAPH1 HiPSC-CMs exposed to H/R conditions. Conclusion: Taken together our data demonstrates that DIAPH1 mediates stress induced senescence in cardiomyocytes and is linked to actin-cytoskeleton dynamics and DIAPH1-MFN2 interactions. Importantly, our data indicates a causative role of DIAPH1 in SIS and sets the stage for probing DIAPH1 as a therapeutic target for protection of hearts from stress induced senescence.