Hypertrophic Markers Research Articles

Establishment of a HFpEF model using female Dahl salt-sensitive rats: a valuable tool for elucidating the pathophysiology of HFpEF in women.

The pathogenesis of heart failure with preserved ejection fraction (HFpEF) remains unclear, and effective treatments are limited. HFpEF is more prevalent in females, indicating potential gender differences in its pathogenesis. However, no female HFpEF model animals have been established. Hypertension is a major contributor to HFpEF, and sympathetic activation is thought to play a role in both conditions. This study aimed to establish a female HFpEF model using hypertensive Dahl salt-sensitive rats and to assess the presence of sympathetic activation. Seven-week-old female Dahl salt-sensitive rats were fed an 8% high-salt diet (HS group, n = 6), while a low-salt diet group (LS group, n = 9) served as controls. The HS group exhibited increased systolic blood pressure and heart rate. Echocardiography revealed an increased left ventricular (LV) wall thickness, a decreased E/A ratio, and an increased E/e' ratio, all indicative of diastolic dysfunction without reduced LV ejection fraction. Additionally, the HS group showed elevated LV end-diastolic pressure, LV weight, and lung weight, along with histological cardiomyocyte hypertrophy and interstitial fibrosis. Gene expression markers for cardiac hypertrophy and fibrosis were also increased. Renal function was significantly impaired, and plasma norepinephrine levels were elevated, consistent with heightened pre-sympathetic neuronal activity in the brain. In conclusion, high salt loading from 7 weeks of age in female Dahl salt-sensitive rats induced hypertensive HFpEF phenotypes with LV hypertrophy and fibrosis, and sympathetic activation by 16 to 19 weeks of age. This model provides a valuable tool for studying HFpEF pathophysiology in women.

Matrigel-Encapsulated Articular cartilage derived fibronectin adhesion assay derived Chondroprogenitors for Enhanced Chondrogenic Differentiation: An In Vitro Evaluation

In cartilage research, three-dimensional (3D) culture models are pivotal for assessing chondrogenic differentiation potential. Standard pellet cultures, despite their utility, pose challenges like uneven differentiation and handling difficulties. This study explores the use of Matrigel, an extracellular matrix-based hydrogel, to encapsulate fibronectin adhesion assay-derived chondroprogenitors (FAA-CPs) and evaluate their chondrogenic differentiation potential. FAA-CPs, isolated from human articular cartilage and expanded to passage 2, were either polymerized in Matrigel or cultured as standard pellets. Both groups underwent chondrogenic differentiation for 28 days and osteogenic differentiation for 21 days. Comprehensive analyses included histological staining, gene expression (SOX-9, ACAN, COL2A1 for chondrogenesis; COL1A1, RUNX2, COL10A1 for osteogenesis), and biochemical assays for glycosaminoglycans (GAG) and Collagen type II. The results demonstrated that Matrigel-encapsulated FAA-CPs achieved greater GAG accumulation, as evidenced by enhanced Alcian Blue and Safranin O staining, compared to standard pellets. However, the Collagen type II deposition, both histologically and quantitatively, was reduced in Matrigel constructs. Gene expression analysis showed no significant differences in key chondrogenic and osteogenic markers between the two groups. Despite improved handling and GAG deposition, Matrigel did not enhance uniform chondrogenic differentiation nor offer significant benefits for osteogenic differentiation, showing comparable hypertrophic markers to the standard method. While Matrigel encapsulation offers advantages in handling and enhances GAG accumulation quantitatively, these benefits were not reflected in staining results. Furthermore, Matrigel did not significantly outperform standard pellet cultures in chondrogenic or osteogenic differentiation. These findings suggest a need for further refinement and in vivo validation.

Protective effect of maltol on pathological response of cardiomyocyte in dystrophic mice.

Heart diseases are a significant contributor to global morbidity and mortality, and despite their diverse and complex mechanisms, treatment options remain limited. Maltol, a natural compound with antioxidant and anti-inflammatory activities, exhibits potential for addressing this need. This study evaluates the cardioprotective effects of maltol in isoproterenol (ISO)-induced cardiac stress models and Duchenne muscular dystrophy (DMD). Maltol's cardiac cytotoxicity was assessed in rodent (H9c2) and human (AC16) cells and compared with that of dapagliflozin to illustrate its cardiac safety. In ISO-induced stress models, maltol significantly reduced hypertrophic markers and inflammation while enhancing autophagy and antioxidant pathways. In the mdx mice, a DMD model, maltol treatment improved cardiac contractility and reduced pathogenic remodeling. Enhanced phosphorylation of phospholamban and trends toward higher SERCA2a expression indicated enhanced Ca2+ handling, which is crucial in DMD cardiomyopathy. This study demonstrated that maltol has the potential to provide therapeutic benefits for DMD and other cardiac conditions characterized by hypertrophy and inflammation, as evidenced by its well-known antioxidant properties, low cytotoxicity, and capacity to enhance cardiac function and Ca2+ handling.

Abstract 4134695: The Effect and Mechanism of Histone Lactylation In Pathological Cardiac Hypertrophy Induced by Pressure Overload

Introduction: Cardiac hypertrophy is characterized by significant metabolic changes, notably an increase in glycolysis. Histone lactylation, a post-translational modification influenced by the glycolytic state of cells, plays a crucial role in regulating gene transcription. However, the relationship between histone lactylation and cardiac hypertrophy remains unclear. Research Questions: This study aims to elucidate the effects and underlying mechanisms of histone lactylation in the progression of cardiac hypertrophy induced by pressure overload. Methods and Results: We observed an increase in histone lactylation levels in hypertrophic hearts from both humans and mice. To investigate further, male mice underwent transverse aortic constriction (TAC) to induce cardiac hypertrophy, followed by either an intraperitoneal injection of Oxamate (an LDHA inhibitor) to reduce histone lactylation or sodium lactate to increase it. Reducing histone lactylation protected the hearts from TAC-induced hypertrophy and fibrosis, preserving cardiac function, while increased histone lactylation exacerbated cardiac hypertrophy and fibrosis, impairing cardiac function. Cardiomyocyte-specific deletion of LDHA also led to a reduction in cardiac hypertrophy. In vitro experiments showed that inhibiting histone lactylation reduced the expression of hypertrophic markers and the hypertrophic growth of cardiomyocytes stimulated by phenylephrine. Using co-immunoprecipitation, we confirmed that P300 and GCN5 are the transferases mediating histone lactylation in cardiomyocytes. Mechanistic studies using the CUT&Tag technique revealed lactate-dependent histone modification was enriched at the promoter of TGFB2, activating its transcription. Inhibiting cardiac-specific TGFB2 expression through adeno-associated viral delivery of shRNA reduced TAC-induced cardiac hypertrophy. Furthermore, TGFB2 overexpression induced a hypertrophic phenotype in cardiomyocytes and activated the PI3K/AKT/mTOR pathways. This hypertrophic phenotype, induced by TGFB2 overexpression, was suppressed by inhibitors of PI3K or AKT. Conclusion: Our findings reveal a critical role for histone lactylation in the development of pathological cardiac hypertrophy by regulating TGFB2 expression and the PI3K/AKT/mTOR pathway.

Efficient Production of Chondrocyte Particles from Human iPSC-Derived Chondroprogenitors Using a Plate-Based Cell Self-Aggregation Technique.

The limited capacity of articular cartilage for self-repair is a critical challenge in orthopedic medicine. Here, we aimed to develop a simplified method of generating chondrocyte particles from human-induced pluripotent stem cell-derived expandable limb-bud mesenchymal cells (ExpLBM) using a cell self-aggregation technique (CAT). ExpLBM cells were induced to form chondrocyte particles through a stepwise differentiation protocol performed on a CAT plate (prevelex-CAT®), which enables efficient and consistent production of an arbitrary number of uniformly sized particles. Histological and immunohistochemical analyses confirmed that the generated chondrocyte particles expressed key cartilage markers, such as type II collagen and aggrecan, but not hypertrophic markers, such as type X collagen. Additionally, when these particles were transplanted into osteochondral defects in rats with X-linked severe combined immunodeficiency, they demonstrated successful engraftment and extracellular matrix production, as evidenced by Safranin O and Toluidine Blue staining. These data suggest that the plate-based CAT system offers a robust and scalable approach to produce a large number of chondrocyte particles in a simplified and efficient manner, with potential application to cartilage regeneration. Future studies will focus on refining the system and exploring its clinical applications to the treatment of cartilage defects.

Sex-Specific Changes in Cardiac Function and Electrophysiology During Progression of Adenine-Induced Chronic Kidney Disease in Mice.

Chronic kidney disease (CKD) and cardiovascular disease (CVD) often co-exist, with notable sex-dependent differences in manifestation and progression despite both sexes sharing similar risk factors. Identifying sex-specific diagnostic markers in CKD-induced CVD could elucidate why the development and progression of these diseases differ by sex. Adult, C57BL/6J male and female mice were fed a high-adenine diet for 12 weeks to induce CKD, while control mice were given a normal diet. Adenine-treated males showed more severe CKD than females. Cardiac physiology was evaluated using electrocardiogram (ECG) and echocardiogram markers. Only adenine-treated male mice showed markers of left ventricular (LV) hypertrophy. Adenine males showed markers of LV systolic and diastolic dysfunction throughout regimen duration, worsening as the disease progressed. Adenine males had prolonged QTc interval compared to adenine females and control males. We identified a new ECG marker, Speak-J duration, which increased with disease progression and appeared earlier in adenine-treated males than in females. We identified sex-dependent differences in cardiac structure, function, and electrophysiology in a CKD-induced CVD mouse model, with adenine-treated males displaying markers of LV hypertrophy, dysfunction, and electrophysiological changes. This study demonstrates the feasibility of using this model to investigate sex-dependent cardiac differences resulting from CKD.

A Multicenter Evaluation of the Impact of Therapies on Deep Learning-based Electrocardiographic Hypertrophic Cardiomyopathy Markers

Artificial intelligence-enhanced electrocardiography (AI-ECG) can identify hypertrophic cardiomyopathy (HCM) on 12-lead ECGs and offers a novel way to monitor treatment response. While the surgical or percutaneous reduction of the interventricular septum (SRT) represented initial HCM therapies, mavacamten offers an oral alternative. We aimed to assess the use of AI-ECG as a strategy to evaluate biological response to SRT and mavacamten. We applied an AI-ECG model for HCM detection to ECG images from patients who underwent SRT across 3 sites: Yale New Haven Health System (YNHHS), Cleveland Clinic Foundation (CCF), and Atlantic Health System (AHS); and to ECG images from patients receiving mavacamten at YNHHS. A total of 70 patients underwent SRT at YNHHS, 100 at CCF, and 145 at AHS. At YNHHS, there was no significant change in the AI-ECG HCM score before versus after SRT (pre-SRT: median 0.55 [IQR 0.24-0.77] vs post-SRT: 0.59 [0.40-0.75]). The AI-ECG HCM scores also did not improve post SRT at CCF (0.61 [0.32-0.79] vs 0.69 [0.52-0.79]) and AHS (0.52 [0.35-0.69] vs 0.61 [0.49-0.70]). Among 36 YNHHS patients on mavacamten therapy, the median AI-ECG score before starting mavacamten was 0.41 (0.22-0.77), which decreased significantly to 0.28 (0.11-0.50, p <0.001 by Wilcoxon signed-rank test) at the end of a median follow-up period of 237 days. In conclusion, we observed a lack of improvement in AI-based HCM score with SRT, in contrast to a significant decrease with mavacamten. Our approach suggests the potential role of AI-ECG for serial point-of-care monitoring of pathophysiological improvement following medical therapy in HCM using ECG images.

MiR-421-mediated suppression of FGF13 as a novel mechanism ameliorates cardiac hypertrophy by inhibiting endoplasmic reticulum stress

Pathological cardiac hypertrophy is an independent risk factor for heart failure. Currently, clinical treatments offer limited effectiveness, and both mortality and morbidity from cardiac hypertrophy and heart failure continue to be significant. Therefore, it is extremely urgent to find new intervention targets to prevent and alleviate pathological cardiac hypertrophy. In this study, we explored FGF13 expression and its upstream regulators in hypertrophic hearts. Firstly, we observed an increase in FGF13 expression levels in human hypertrophic myocardium tissues, as well as in mouse models of TAC-induced hypertrophy and in neonatal rat cardiomyocyte (NRCM) models induced by isoproterenol (ISO). Moreover, these elevated levels of FGF13 were shown to positively correlate with hypertrophic markers, including ANP and BNP. By using both gain-of-function and loss-of-function approaches in an in vitro hypertrophy model, we demonstrated that FGF13 knockdown could inhibit endoplasmic reticulum stress (ERS), thereby ameliorating cardiomyocyte hypertrophy. Meanwhile, we investigated the upstream regulators of FGF13 in hypertrophic hearts, and a dual-luciferase reporter assay confirmed that FGF13 is a direct target of miR-421. Overexpression of miR-421 decreased the protein level of FGF13 and ameliorated ISO-induced cardiomyocyte hypertrophy via modulating ER stress. In contrast, overexpression of FGF13 attenuated the ameliorative effect of miR-421 on ISO-induced cardiomyocyte hypertrophy. Taken together, the present results suggested that miR-421 ameliorated ISO-induced cardiomyocyte hypertrophy by negatively regulating FGF13 expression. This finding may offer a novel approach for the treatment of cardiac hypertrophy.

Dapagliflozin mitigates cellular stress and inflammation through PI3K/AKT pathway modulation in cardiomyocytes, aortic endothelial cells, and stem cell-derived β cells

Dapagliflozin (DAPA), a sodium-glucose cotransporter 2 (SGLT2) inhibitor, is well-recognized for its therapeutic benefits in type 2 diabetes (T2D) and cardiovascular diseases. In this comprehensive in vitro study, we investigated DAPA’s effects on cardiomyocytes, aortic endothelial cells (AECs), and stem cell-derived beta cells (SC-β), focusing on its impact on hypertrophy, inflammation, and cellular stress. Our results demonstrate that DAPA effectively attenuates isoproterenol (ISO)-induced hypertrophy in cardiomyocytes, reducing cell size and improving cellular structure. Mechanistically, DAPA mitigates reactive oxygen species (ROS) production and inflammation by activating the AKT pathway, which influences downstream markers of fibrosis, hypertrophy, and inflammation. Additionally, DAPA’s modulation of SGLT2, the Na+/H + exchanger 1 (NHE1), and glucose transporter (GLUT 1) type 1 highlights its critical role in maintaining cellular ion balance and glucose metabolism, providing insights into its cardioprotective mechanisms. In aortic endothelial cells (AECs), DAPA exhibited notable anti-inflammatory properties by restoring AKT and phosphoinositide 3-kinase (PI3K) expression, enhancing mitogen-activated protein kinase (MAPK) activation, and downregulating inflammatory cytokines at both the gene and protein levels. Furthermore, DAPA alleviated tumor necrosis factor (TNFα)-induced inflammation and stress responses while enhancing endothelial nitric oxide synthase (eNOS) expression, suggesting its potential to preserve vascular function and improve endothelial health. Investigating SC-β cells, we found that DAPA enhances insulin functionality without altering cell identity, indicating potential benefits for diabetes management. DAPA also upregulated MAFA, PI3K, and NRF2 expression, positively influencing β-cell function and stress response. Additionally, it attenuated NLRP3 activation in inflammation and reduced NHE1 and glucose-regulated protein GRP78 expression, offering novel insights into its anti-inflammatory and stress-modulating effects. Overall, our findings elucidate the multifaceted therapeutic potential of DAPA across various cellular models, emphasizing its role in mitigating hypertrophy, inflammation, and cellular stress through the activation of the AKT pathway and other signaling cascades. These mechanisms may not only contribute to enhanced cardiac and endothelial function but also underscore DAPA’s potential to address metabolic dysregulation in T2D.Graphical abstract

Cardiac secreted-HSP90alpha in cardiomyocyte hypertrophy by N-Cadherin mediated signaling under pressure overload

Abstract Background Left ventricular hypertrophy (LVH) in response to pressure overload is strongly associated with adverse cardiovascular outcomes. Heat shock protein 90 (HSP90) has been reported to be involved in cardiac remodeling under stress. Aims Here, we evaluate whether serum HSP90α (an isoform of HSP90) increases and associates with cardiac hypertrophy in patients with hypertension or severe aortic stenosis (AS), and explore the potential mechanisms in experimental pressure overload mouse model. Methods Serum HSP90α levels in patients with hypertension or aortic stenosis (AS) were determined by ELISA. Pressure overload mouse model was built by a transverse aortic constriction (TAC). Cardiac function was assessed by echocardiography. Cardiac hypertrophy was examined by histology, western blot and RNA analysis. Results Serum levels of HSP90α were higher in patients with hypertension or AS than in the control group, respectively, and the levels positively correlated with LVH. HSP90α levels also increased in heart tissues of patients with obstructive hypertrophic cardiomyopathy (HCM), and mice after TAC, in parallel with the hypertrophic markers. TAC induced the enhanced expression and secretion of HSP90α from cardiomyocytes and cardiac fibroblasts. Knockdown of HSP90α or blockade of extracellular HSP90α (eHSP90α) prevent the development of LVH under pressure overload in vivo and in vitro. By Nano-HPLC-MS/MS analysis, we identified eHSP90α interacted with N-cadherin in cardiomyocyte surface, which induced the activation of β-catenin and promoted β-catenin to bind TCF7 (a member of TCF/Lef family) in nucleus, enhanced the transcription of hypertrophic genes, contributing to cardiac hypertrophy and dysfunction under pressure overload. Conclusions Our data demonstrated that cardiac secreted-HSP90α promoted myocardial hypertrophy by N-Cadherin mediated β-catenin/TCF7 signaling under pressure overload. It indicates the therapeutic potential of targeting HSP90α-initiated signaling against cardiac hypertrophy and dysfunction under pressure overload.

Serum exosomal long non-coding RNA H19 as a theragnostic target for atrial fibrillation

Abstract Background/Introduction Exosomes are membrane vesicles of 30- to 150-nm secreted by most cell types that can mediate intercellular communication by transmitting various forms of RNAs. Long non-coding RNAs (lncRNAs, ≥ 200 nucleotides length) have been reported to play important roles in the pathophysiological processes that promote the development and progression of many diseases. Interestingly, recent studies have shown that exosomal lncRNAs exhibit different expression profiles in various diseases and are being extensively studied in the medical field as an ideal target for the diagnosis and treatment. However, there is still little known about their role in atrial fibrillation (AF). Purpose The aim of this study was to identify a novel theragnostic target exosomal lncRNA for AF. Methods First, serum exosomes from controls and patients with persistent AF were isolated and characterized by transmission electron microscopy, nanoparticle tracking analysis, and western blot. Next, serum exosomal lncRNA expression profiles were explored using RNA-sequencing analysis. In addition, the expression levels of candidate exosomal lncRNAs were confirmed using qRT-PCR (n = 50 per group). Finally, AF-related pathophysiological mechanisms of exosomal lncRNA were investigated in angiotensin II (Ang II)-treated human induced pluripotent stem cell-derived atrial cardiomyocytes (iPSC-atrial CMs). Results After RNA-sequencing analysis, we identified 27 differentially expressed lncRNAs (i.e., 4 upregulated and 23 downregulated lncRNAs with a |fold change| ≥ 2 and p &amp;lt; 0.05) in serum exosomes from patients with persistent AF compared with the controls. In fact, qRT-PCR reveled that exosomal lncRNA H19 was consistently downregulated in the serum of patients with persistent AF compared with the controls (p &amp;lt; 0.01). In addition, exosomal lncRNA H19 exhibited significant diagnostic validity for AF. Notably, we confirmed that exosomal lncRNA H19 was involved in AF-related pathophysiological mechanisms. In Ang II-treated iPSC-atrial CMs, inhibition of lncRNA H19 significantly aggravated Ang II-induced increases in levels of hypertrophic markers (ANP, BNP and β-MHC), cell surface area and inflammation (p &amp;lt; 0.05). Mechanistically, we found that loss of lncRNA H19 weakens the inhibition of its direct target microRNAs, miR-141-3p and miR-200a-3p, leading to downregulation of their target PTEN, thereby promoting the atrial remodeling and the consequent AF. Conclusion In conclusion, serum-derived exosomal lncRNA H19 could serve as a potential target for the diagnosis and treatment of AF.

Intact DNA repair in differentiated cardiomyocytes is essential for maintaining cardiac function in response to pressure overload in mice

Abstract Introduction DNA in every cell is continuously damaged and DNA repair mechanisms are essential for protection against DNA damage-induced aging-related diseases. For example, deficient repair of endogenously generated DNA damage in mice with cardiomyocyte-restricted inactivation of Xpg, is associated with progressive heart failure. Purpose Here we tested the hypothesis that unrepaired DNA damage in differentiated cardiomyocytes increases cardiac vulnerability in response to hemodynamic overload. Methods To increase the burden of spontaneous DNA damage, we generated mice with cardiomyocyte-restricted inactivation of DNA repair endonuclease XPG. At 8 weeks of age, αMHC-Xpgc/- and control (Ctrl) mice were subjected to pressure overload by transverse aortic constriction (TAC). Eight weeks after TAC, left ventricular (LV) function was assessed using echocardiography and hemodynamic measurements, followed by molecular and histological analyses. Results Cardiomyocyte-restricted inactivation of Xpg resulted in systolic as well as diastolic LV dysfunction (Table). TAC-induced LV hypertrophy is similar in both groups (Ctrl 38%; αMHC-Xpgc/- 34%). In Ctrl mice, LV hypertrophy was accompanied by minimal LV dilation and modest changes in systolic and diastolic LV function. Conversely, TAC in αMHC-Xpgc/- produced severe LV dysfunction and resulted in overt congestive heart failure, demonstrated by aggravation of LV dilation, and marked increases in LV end-diastolic pressure, left atrial weight and lung fluid weight, which was accompanied by further increases in the expression levels of the hypertrophic marker genes atrial natriuretic peptide and beta-myosin heavy chain (Table). Moreover, lectin staining revealed a decrease in capillary density and TUNEL staining revealed further elevated levels of myocardial apoptosis. In addition, a significant increase of myocardial collagen content was observed. Conclusion Cardiomyocyte-restricted loss of DNA repair protein XPG increases cardiac vulnerability to develop heart failure in response to pressure overload. These findings underscore the importance of genomic stability for maintenance of cardiac function, not only during basal conditions, but also in response to a pathological stimulus.

The Therapeutic Potential of Adipose-Derived Mesenchymal Stem Cell Secretome in Osteoarthritis: A Comprehensive Study

Osteoarthritis (OA) is a degenerative joint disease characterized by cartilage degradation and inflammation. This study investigates the therapeutic potential of secretome derived from adipose tissue mesenchymal stem cells (ASCs) in mitigating inflammation and promoting cartilage repair in an in vitro model of OA. Our in vitro model comprised chondrocytes inflamed with TNF. To assess the therapeutic potential of secretome, inflamed chondrocytes were treated with it and concentrations of pro-inflammatory cytokines, metalloproteinases (MMPs) and extracellular matrix markers were measured. In addition, secretome-treated chondrocytes were subject to a microarray analysis to determine which genes were upregulated and which were downregulated. Treating TNF-inflamed chondrocytes with secretome in vitro inhibits the NF-κB pathway, thereby mediating anti-inflammatory and anti-catabolic effects. Additional protective effects of secretome on cartilage are revealed in the inhibition of hypertrophy markers such as RUNX2 and COL10A1, increased production of COL2A1 and ACAN and upregulation of SOX9. These findings suggest that ASC-derived secretome can effectively reduce inflammation, promote cartilage repair, and maintain chondrocyte phenotype. This study highlights the potential of ASC-derived secretome as a novel, non-cell-based therapeutic approach for OA, offering a promising alternative to current treatments by targeting inflammation and cartilage repair mechanisms.

Protective Role of (-)-Epicatechin on Trimethylamine-N-Oxide (TMAO)-Induced Cardiac Hypertrophy via SP1/SIRT1/SUMO1 Signaling Pathway

(-)-Epicatechin (EPI) is beneficial for cardiovascular health. Trimethylamine N-oxide (TMAO), a gut microbe-derived food metabolite, is strongly associated with the risk of cardiovascular diseases. However, the effects and underlying mechanisms of EPI on TMAO-induced cardiac hypertrophy remain unclear. This study aimed to determine whether EPI inhibits TMAO-induced cardiac hypertrophy. Plasma levels of TMAO in control participants and patients with cardiac hypertrophy were measured and analyzed. Male C57BL/6 mice were randomly divided into control group, TMAO group, EPI group and TMAO + EPI group. According to the groups assignments, mice received intraperitoneal (i.p.) injection of normal saline or i.p. injection of TMAO (150 mg/kg/day) for 14 days. The EPI group was given intragastric (i.g.) administration of EPI alone (1 mg/kg/day) for 21 days, and TMAO + EPI group received i.g. administration of EPI for 7 days before starting i.p. injection of TMAO, continuing until the end of the TMAO treatment. Histological analyses of the mice’s hearts was accessed by H&E and Masson staining. In vitro, H9c2 cells were induced to hypertrophy by TMAO (10 µM) for 24 h and were pre-treated with or without EPI (10 µM) for 1 h. Protein level of cardiac hypertrophy markers and Sp1/SIRT1/SUMO1 pathway were determined by western blot. The plasma level of TMAO was 2.66 ± 1.59 μmol/L in patients with cardiac hypertrophy and 0.62 ± 0.30 μmol/L in control participants. EPI attenuated TMAO-induced hypertrophy in H9c2 cells. In vivo, TMAO induced cardiac hypertrophy and impaired the cardiac function of mice. Pathological staining showed that TMAO induced cardiac hypertrophy and collagen deposition in mice. EPI treatment improved the cardiac function, inhibited the myocardial hypertrophy induced by TMAO. EPI significantly attenuated the TMAO-induced upregulation of ANP and BNP and the downregulation of SP1, SIRT1 and SUMO1 in vivo and in vitro. EPI may suppress TMAO-induced cardiac hypertrophy by activating the Sp1/SIRT1/SUMO1 signaling pathway.

8519 Investigating the Role of Protein Inhibitor of Activated STAT1 (Pias1) in Growth Plate Chondrocyte Maturation

Abstract Disclosure: J. Baronas: None. U. Ahmed: None. J.N. Hirschhorn: None. N.E. Renthal: None. Development of growth plate chondrocytes is a complex process that involves a wide array of regulatory elements. Recently our laboratory conducted a genome wide CRISPR knockout (KO) screen of growth plate chondrocyte maturation to discover novel genes and gene networks in the proliferation and maturation of chondrocytes. Among targets uncovered is Protein Inhibitor of Activated STAT 1 (Pias1), an E3 SUMO ligase that plays a multi-faceted role in genetic regulation, driving post-translational modifications, directly binding to transcription factors, and functioning as a DNA-binding protein. While Pias1 has been shown to be involved in the regulation of many cellular processes across multiple tissues, its function in growth plate development and chondrocyte maturation has been previously unstudied. In our genome-wide screen, our laboratory identified that loss of Pias1 led to early maturation of chondrocytes. This project seeks to investigate the role of Pias1 in chondrocyte maturation and differentiation. We first developed a Pias1-KO chondrocyte cell line, observing upregulation of hypertrophic markers Cd200 and Col10a1 in KOs relative to control via qPCR, in addition to proliferative regulators IHH and Pth1R. Through Bulk-RNAseq of the same cell line, we identified additional chondrocyte markers as upregulated in KO, as well as osteoblast marker Sp7. Members of the HoxD family, regulators of limb development, were downregulated. Cell culture data suggests that loss of Pias1 leads to increased cell proliferation and increased matrix production, indicating dysregulation of multiple chondrocyte processes. Immunohistochemistry of micromass culture sections also suggests morphological differences between KO and control, with KO cells significantly larger in diameter. Early SUMO-1 profiling yields limited evidence of decreased global SUMOylation in KO cells, one possible mechanistic explanation for the observed dysregulation. Ongoing studies are focused on identifying specific SUMOylation targets of Pias1 in chondrocytes, analyzing proteomic differences in KO and control, seeking putative DNA-binding sites of Pias1, and observing in vivo murine Pias1-KO growth plates. Our findings suggest that PIAS1 plays a crucial role in the growth plate by prolonging growth and delaying terminal hypertrophy. Our study aims to provide a better understanding of the regulatory pathways involved in growth plate development and related phenotypes such as height, and contribute to the development of novel therapies for growth-related disorders, such as achondroplasia and hypochondroplasia. Presentation: 6/1/2024

Fat Embolism Does Not Alter Cardiac Structure or Induce Pathological Changes in a Rat Model

IntroductionFat embolism (FE) encompasses conditions in which fatty substance becomes embedded in a tissue/organ. Fat emboli most commonly affect the lungs in a trauma setting. This can lead to both significant pathology locally and systemically including changes in structure, inflammatory response, activation of the renin-angiotensin system, and subsequent hypoxia. In fact, changes in skin, brain, lungs, and kidneys have been noted in FE syndrome. Because there is an extensive record of pathology reports on this condition without evidence of direct cardiac involvement, as well as our studies showing apparent complete recovery after the acute embolism, we hypothesized that structural changes similar to the lung and at the same time course would not be observed in the heart. MethodsWe used a rat model of FE previously described by our group where we have documented significant lung pathology. In this study, we analyzed both pulmonary and cardiac structure, histology, and gene expression at 48 h and 10 wks post fat injection to mimic FE. ResultsDespite severe inflammatory evidence and structural changes to the lung and vasculature up to 10 wks after FE, we found no significant alterations to cardiovascular morphometry including lumen patency ratio, adventitia/media ratio, fibrosis content, and heart chamber/wall dimensions in stained histological sections. Additionally, genetic markers of cardiac pathological hypertrophy were not significantly elevated 48 h or 10 wks after fat treatment. Oil Red O staining showed increased fat droplet content within lung and aorta tissue, but not in the myocardium. ConclusionsOur study suggests that, in contrast to the lungs, the heart is more resistant to the inflammatory and remodeling responses that result from FE, possibly due to the organ-specific differences in fat retention.

Apelin-13's Actions in Controlling Hypertension-Related Cardiac Hypertrophy and the Expressions of Inflammatory Cytokines.

As a key molecule for improving cardiovascular diseases, Apelin-13 was surveyed in this work to explain its actions in controlling inflammation, pyroptosis, and myocardial hypertrophy. First, mouse models with myocardial hypertrophy were established. Then, assessments were made on the pathological variation in the heart of mouse, on the cardiac functions, as well as on the expressions of cardiac hypertrophy markers (β-MHC, ANP, and BNP), inflammatory factors (TNF-α, COX2, IL-6, ICAM-1, and VCAM-1), myocardial cell pyroptosis markers (NLRP3, ASC, c-caspase-1, and GSDMD-N), and Hippo pathway proteins (p-YAP, YAP, LATS1, and p-LATS1) by HE staining, echocardiography scanning, and western blot tests separately. The expressions of such inflammatory factors as in myocardial tissue were acquired by ELISA. After inducing the phenotype of H9c2 cell hypertrophy by noradrenaline, we used CCK-8 kits to know about the activity of H9c2 cells treated with Apelin-13, and performed ɑ-actinin staining to measure the changes in volumes of such cells. As unraveled through this work, Apelin-13 refrained the activation of the Hippo pathway, which in turn attenuated the hypertrophy, inflammation, and pyroptosis of myocardial tissue and H9c2 cells. Hence, Apelin-13 can be considered as a target for hypertension treatment.

Aerobic Exercise Improved Diacylglycerol/Protein Kinase C and Phosphatidylinositol 4,5-Biphosphate Pathway in Cardiac Tissue of Obese Rats

Background: Obesity, characterized by abnormal accumulation or excess lipids, is a major cause of increased incidence of cardiovascular diseases. Objectives: The purpose of this experiment is to investigate the effect of aerobic exercise on the expression of sn-1,2-diacylglycerol (DAG), Protein kinase C (PKC), and Phosphatidylinositol 4,5-biphosphate (PIP2) genes in experimental rat models of diet-induced obesity. Methods: Twenty-four Wistar rats were randomly divided into four groups: (1) control, (2) obesity, (3) exercise, and (4) obesity + exercise. At the end of the experiment, cardiac hypertrophy markers and real-time PCR were used to evaluate and compare the gene expression levels of DAG, PKC, and PIP2 in cardiac tissue across all groups. Results: A high-fat diet containing fructose significantly increased the gene expression levels of DAG, PKC, and PIP2 in the cardiac tissue of obese rats compared to the control group. However, 6 weeks of aerobic exercise led to a significant decrease in DAG and PKC gene expression compared to the obese rats. Although there was no significant change in PIP2 gene expression in the obesity + exercise group compared to obese rats. Conclusions: Based on the results of this study, it appears that aerobic exercise can modulate the expression of DAG and PKC genes, which play a critical role in cardiac remodeling.

TongGuanWan Alleviates Doxorubicin- and Isoproterenol-Induced Cardiac Hypertrophy and Fibrosis by Modulating Apoptotic and Fibrotic Pathways.

Heart failure, a major public health issue, often stems from prolonged stress or damage to the heart muscle, leading to cardiac hypertrophy. This can progress to heart failure and other cardiovascular problems. Doxorubicin (DOX), a common chemotherapy drug, and isoproterenol (ISO), a β-adrenergic agonist, both induce cardiac hypertrophy through different mechanisms. This study investigates TongGuanWan (TGW,), a traditional herbal remedy, for its effects on cardiac hypertrophy and fibrosis in DOX-induced H9c2 cells and ISO-induced mouse models. TGW was found to counteract DOX-induced increases in H9c2 cell surface area (n = 8, p < 0.01) and improve biomarkers like ANP (n = 3, p < 0.01)) and BNP (n = 3, p < 0.01). It inhibited the MAPK pathway (n = 4, p < 0.01) and GATA-4/calcineurin/NFAT-3 signaling, reduced inflammation by decreasing NF-κB p65 translocation, and enhanced apoptosis-related factors such as caspase-3 (n = 3, p < 0.01), caspase-9 (n = 3, p < 0.01), Bax (n = 3, p < 0.01), and Bcl-2 (n = 3, p < 0.01). Flow cytometry showed TGW reduced apoptotic cell populations. In vivo, TGW reduced heart (n = 8~10, p < 0.01), and left ventricle weights (n = 6~7), cardiac hypertrophy markers (n = 3, p < 0.01), and perivascular fibrosis in ISO-induced mice, with Western blot analysis confirming decreased levels of fibrosis-related factors like fibronectin, α-SMA (n = 3, p < 0.05), and collagen type I (n = 3, p < 0.05). These findings suggest TGW has potential as a therapeutic option for cardiac hypertrophy and fibrosis.

Effect of Cannabistilbene I in Attenuating Angiotensin II-Induced Cardiac Hypertrophy: Insights into Cytochrome P450s and Arachidonic Acid Metabolites Modulation.

Introduction: This research investigated the impact of Cannabistilbene I on Angiotensin II (Ang II)-induced cardiac hypertrophy and its potential role in cytochrome P450 (CYP) enzymes and arachidonic acid (AA) metabolic pathways. Cardiac hypertrophy, a response to increased stress on the heart, can lead to severe cardiovascular diseases if not managed effectively. CYP enzymes and AA metabolites play critical roles in cardiac function and hypertrophy, making them important targets for therapeutic intervention. Methods: Adult human ventricular cardiomyocyte cell line (AC16) was cultured and treated with Cannabistilbene I in the presence and absence of Ang II. The effects on mRNA expression related to cardiac hypertrophic markers and CYP were analyzed using real-time polymerase chain reaction, while CYP protein levels were measured by Western blot analysis. AA metabolites were quantified using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Results: Results showed that Ang II triggered hypertrophy, as evidenced by the increase in hypertrophic marker expression, and enlarged the cell surface area, effects that were alleviated by Cannabistilbene I. Gene expression analysis indicated that Cannabistilbene I upregulated CYP1A1, leading to increased enzymatic activity, as evidenced by 7-ethoxyresorufin-O-deethylase assay. Furthermore, LC-MS/MS analysis of AA metabolites revealed that Ang II elevated midchain (R/S)-hydroxyeicosatetraenoic acid (HETE) concentrations, which were reduced by Cannabistilbene I. Notably, Cannabistilbene I selectively increased 19(S)-HETE concentration and reversed the Ang II-induced decline in 19(S)-HETE, suggesting a unique protective role. Conclusion: This study provides new insights into the potential of Cannabistilbene I in modulating AA metabolites and reducing Ang II-induced cardiac hypertrophy, revealing a new candidate as a therapeutic agent for cardiac hypertrophy.