Cardiomyocyte Injury Research Articles

The potential role of SARS-CoV-2 infection in acute coronary syndrome and type 2 myocardial infarction (T2MI): Intertwining spread.

Coronavirus disease 2019 (COVID-19) is a novel pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). It has been shown that SARS-CoV-2 infection-induced inflammatory and oxidative stress and associated endothelial dysfunction may lead to the development of acute coronary syndrome (ACS). Therefore, this review aimed to ascertain the link between severe SARS-CoV-2 infection and ACS. ACS is a spectrum of acute myocardial ischemia due to a sudden decrease in coronary blood flow, ranging from unstable angina to myocardial infarction (MI). Primary or type 1 MI (T1MI) is mainly caused by coronary plaque rupture and/or erosion with subsequent occlusive thrombosis. Secondary or type 2 MI (T2MI) is due to cardiac and systemic disorders without acute coronary atherothrombotic disruption. Acute SARS-CoV-2 infection is linked with the development of nonobstructive coronary disorders such as coronary vasospasm, dilated cardiomyopathy, myocardial fibrosis, and myocarditis. Furthermore, SARS-CoV-2 infection is associated with systemic inflammation that might affect coronary atherosclerotic plaque stability through augmentation of cardiac preload and afterload. Nevertheless, major coronary vessels with atherosclerotic plaques develop minor inflammation during COVID-19 since coronary arteries are not initially and primarily targeted by SARS-CoV-2 due to low expression of angiotensin-converting enzyme 2 in coronary vessels. In conclusion, SARS-CoV-2 infection through hypercytokinemia, direct cardiomyocyte injury, and dysregulation of the renin-angiotensin system may aggravate underlying ACS or cause new-onset T2MI. As well, arrhythmias induced by anti-COVID-19 medications could worsen underlying ACS.

MiR-16-5p Regulates Ferroptosis by Targeting SLC7A11 in Adriamycin-Induced Ferroptosis in Cardiomyocytes.

Adriamycin (ADR) is commonly used in tumor chemotherapy, but its nonreversible cardiotoxicity severely hampers its clinical application. Ferroptosis is an implicated cause of ADR-induced injury. However, the underlying molecular mechanisms remain poorly understood. This study explored whether ferroptosis is a pivotal pathogenic pathway underlying ADR-induced cardiotoxicity and the possible molecular mechanisms involved. In vivo and in vitro experimental models were used to study the mechanism of ADR-mediated ferroptosis. Ferroptosis levels were examined in mice and human/rat cardiomyocytes. Mechanistically, the expression levels of SLC7A11 and related miRNAs were examined. Bioinformatics prediction and luciferase reporter assays were used to verify the potential interaction between miR-16-5p and SLC7A11. The biological functions and interaction of SLC7A11 and miR-16-5p were investigated in vivo and in vitro. Our study observed that ADR treatment significantly increased ferroptosis levels in vivo and in vitro. Ferroptosis-related pharmacological interventions further confirmed these results. Our data displayed that the SLC7A11 level was significantly decreased in cardiac tissues and cells, while an increased expression level of miR-16-5p was observed. Moreover, upregulation of SLC7A1 and inhibition of miR-16-5p attenuated ADR-induced cardiomyocyte ferroptosis injury. Interactive rescue experiments showed that the protective effects of miR-16-5p inhibition on ADR-induced cardiomyocyte injury were blocked by SLC7A11 knockdown. Based on these findings, targeting miR-16-5p could partially reverse the ADR-induced cardiotoxicity by rescuing the SLC7A11 to attenuate ferroptosis. This study presents a pre-clinical basis to identify miR-16-5p/SLC7A11 as a potential treatment target for ADR-induced cardiotoxicity.

Association of high-sensitivity troponin T with outcomes in asymptomatic non-severe aortic stenosis: a post-hoc substudy of the SEAS trial

High-sensitivity Troponin T (hsTnT), a biomarker of cardiomyocyte overload and injury, relates to aortic valve replacement (AVR) and mortality in severe aortic stenosis (AS). However, its prognostic value remains unknown in asymptomatic patients with AS. We aimed to investigate if an hsTnT level >14pg/mL (above upper limit of normal 99th percentile) is associated with echocardiographic AS-severity, subsequent AVR, ischaemic coronary events (ICE), and mortality in asymptomatic patients with non-severe AS. In this post-hoc sub-analysis of the multicentre, randomised, double-blind, placebo-controlled SEAS trial (ClinicalTrials.gov, NCT00092677), we included asymptomatic patients with mild to moderate-severe AS. We ascertained baseline and 1-year hsTnT concentrations and examined the association between baseline levels and the risk of the primary composite endpoint, defined as the first event of all-cause mortality, isolated AVR (without coronary artery bypass grafting (CABG)), or ICE. Multivariable regressions and competing risk analyses examined associations of hsTnT level >14pg/mL with clinical correlates and 5-year risk of the primary endpoint. Between January 6, 2003, and March 4, 2004, a total of 1873 patients were enrolled in the SEAS trial, and 1739 patients were included in this post-hoc sub-analysis. Patients had a mean (SD) age of 67.5 (9.7) years, 61.0% (1061) were men, 17.4% (302) had moderate-severe AS, and 26.0% (453) had hsTnT level >14pg/mL. The median hsTnT difference from baseline to 1-year was 0.8pg/mL (IQR,-0.4 to 2.3). In adjusted linear regression, log(hsTnT) did not correlate with echocardiographic AS severity (p=0.36). In multivariable Cox regression, a hsTnT level >14pg/mL vs. hsTnT ≤14pg/mL was associated with an increased risk of the primary composite endpoint (HR, 1.41; 95% CI, 1.18-1.70; p=0.0002). In a competing risk model of first of the individual components of the primary endpoint, a hsTnT level >14pg/mL was associated with ICE risk (HR 1.71; 95% CI, 1.23-2.38; p=0.0013), but not with isolated AVR (p=0.064) or all-cause mortality (p=0.49) as the first event. hsTnT level is within the reference range (≤14pg/mL) in 3 out of 4 non-ischaemic patients with asymptomatic mild-to-moderate AS and remains stable during a 1-year follow-up regardless of AS-severity. An hsTnT level >14pg/mL was mainly associated with subsequent ICE, which suggest that hsTnT concentration is primarily a risk marker of subclinical coronary atherosclerotic disease. Merck & Co., Inc., the Schering-Plough Corporation, the Interreg IVA program, Roche Diagnostics Ltd., and Gangstedfonden. Open access publication fee funding provided by prof. Olav W. Nielsen and Department of Cardiology, Bispebjerg University Hospital, Denmark.

Plantamajoside alleviates hypoxia-reoxygenation injury through integrin-linked kinase/c-Src/Akt and the mitochondrial apoptosis signaling pathways in H9c2 myocardial cells

Myocardial ischemia–reperfusion injury(MIRI) is one of the common complications after myocardial infarction surgery, Oxidative stress is among the main mechanisms of myocardial ischemia–reperfusion injury. Plantamajoside (PMS), the main effective ingredient in the genus Plantain, has been reported to possess an antioxidation, anti-inflammatory and anti-apoptosis role. However, whether PMS can attenuate myocardial ischemia–reperfusion injury is not yet known. Herein, we explored the effects of PMS on hypoxia-reoxygenation (H/R) injury in H9c2 cardiomyocytes and the underling molecular mechanisms of the treatment. Network pharmacological analysis screened the top 31 key genes in the treatment of MIRI disease treated with PMS, and the result of molecular docking further illustrated the roles that the PMS play in the treatment of MIRI through its interference with integrin-linked kinase (ILK) target protein. PMS was not cytotoxic in the concentration range of 5–40 μM and increased cell survival after H/R injury in a concentration-dependent manner without affecting proliferation or growth. PMS significantly reduced the levels of lactate dehydrogenase, malonic dialdehyde, reactive oxygen species and cell apoptosis, and increased soperoxide dismutase activity compared with those of the H/R injury group. PMS promoted the protein and mRNA expression of ILK and Bcl-2, the protein expression of p-Akt, and reduced the protein and mRNA expression of Bax, Caspase-3, and Cytochrome c, the protein expression of p–c-Src. PMS has protective effects against H/R injury in H9c2 cells, and its protective mechanism may be related to reactive oxygen species clearance, activation of the ILK/c-Src/Akt pathway and inhibition of the mitochondrial apoptosis.

Tranilast reduces cardiomyocyte injury induced by ischemia‑reperfusion via Nrf2/HO‑1/NF‑κB signaling.

Tranilast, a synthetic derivative of a tryptophan metabolite, can be used to treat heart diseases. However, the specific mechanism underlying the effect of tranilast on ischemia-reperfusion (I/R) injury-induced cardiomyocyte apoptosis remains unclear. Therefore, the present study aimed to determine if tranilast could attenuate I/R-induced cardiomyocyte injury. A hypoxia/reoxygenation (H/R) model of H9c2 cardiomyocytes was established to simulate I/R-induced cardiomyocyte injury. The viability, apoptosis, inflammation and oxidative stress in H/R-induced H9c2 cells following treatment with tranilast were evaluated by Cell Counting Kit-8 and TUNEL assay. Commercially available kits were used to detect the levels of inflammatory markers and oxidative stress indicators. In addition, the expression levels of the apoptosis- and nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1)/NF-κB signalling pathway-associated proteins were detected by western blotting. The levels of reactive oxygen species were determined using 2',7'-dichlorofluorescin diacetate assay kit. The viability of H9c2 cells was decreased following induction with H/R. However, treatment with tranilast increased viability while decreasing apoptosis, oxidative stress and inflammatory response in H/R-induced H9c2 cells by activating Nrf2/HO-1/NF-κB signalling. Furthermore, treatment with ML-385, an Nrf2 inhibitor, reversed the effects of tranilast on H/R-induced H9c2 cells. In conclusion, the results of the present study suggested that tranilast could attenuate I/R-induced cardiomyocyte injury via the Nrf2/HO-1/NF-κB signalling pathway.

CircHSPG2 knockdown attenuates hypoxia-induced apoptosis, inflammation, and oxidative stress in human AC16 cardiomyocytes by regulating the miR-1184/MAP3K2 axis.

Circular RNAs (circRNAs) have been identified as vital regulators in cardiovascular diseases, including acute myocardial infarction (AMI). In this study, the function and mechanism of circRNA heparan sulfate proteoglycan 2 (circHSPG2) in hypoxia-induced injury in AC16 cardiomyocytes were investigated. AC16 cells were stimulated with hypoxia to establish an AMI cell model in vitro. Real-time quantitative PCR and western blot assays were performed to quantify the expression levels of circHSPG2, microRNA-1184 (miR-1184), and mitogen-activated protein kinase kinase kinase 2 (MAP3K2). Counting Kit-8 (CCK-8) assay was used to measure cell viability. Flow cytometry was performed to detect cell cycle and apoptosis. Enzyme-linked immunosorbent assay (ELISA) was used to determine the expression of inflammatory factors. Dual-luciferase reporter, RNA immunoprecipitation (RIP), and RNA pull-down assays were used to analyze the relationship between miR-1184 and circHSPG2 or MAP3K2. In AMI serum, circHSPG2 and MAP3K2 mRNA were highly expressed and miR-1184 was down-regulated. Hypoxia treatment elevated HIF1α expression and repressed cell growth and glycolysis. Moreover, hypoxia promoted cell apoptosis, inflammation, and oxidative stress in AC16 cells. Hypoxia-induced circHSPG2 expression in AC16 cells. CircHSPG2 knockdown alleviated hypoxia-induced AC16 cell injury. CircHSPG2 directly targeted miR-1184, and miR-1184 targeted and suppressed MAP3K2. Inhibition of miR-1184 or overexpression of MAP3K2 abolished the alleviated effect of circHSPG2 knockdown on hypoxia-induced AC16 cell injury. Overexpression of miR-1184 relieved hypoxia-induced impairment in AC16 cells by MAP3K2. CircHSPG2 could regulate MAP3K2 expression through miR-1184. CircHSPG2 knockdown protected AC16 cells from hypoxia-induced injury by regulating the miR-1184/MAP3K2 cascade.

Therapeutic Hypothermia Inhibits Hypoxia‐Induced Cardiomyocyte Apoptosis Via the MiR‐483‐3p/Cdk9 Axis

Background Therapeutic hypothermia has a beneficial effect on cardiac function after acute myocardial infarction, but the exact mechanism is still unclear. Recent research has suggested that microRNAs participate in acute myocardial infarction to regulate cardiomyocyte survival. This study aimed to explore the ability of hypothermia-regulated microRNA-483-3p (miR-483-3p) to inhibit hypoxia-induced myocardial infarction. Methods and Results Primary cardiomyocytes were cultured under hypoxia at 32 °C to mimic therapeutic hypothermia, and the differentially expressed microRNAs were determined by RNA sequencing. Therapeutic hypothermia recovered hypoxia-induced increases in apoptosis, decreases in ATP levels, and decreases in miR-483-3p expression. Overexpression of miR-483-3p exhibited effects similar to those of therapeutic hypothermia on hypoxia in the treatment of cardiomyocytes to associate with maintaining the mitochondrial membrane potential, and cyclin-dependent kinase 9 (Cdk9) was identified as a target gene with downregulated expression by miR-483-3p. Knockdown of Cdk9 also promoted cardiac survival, ATP production, and mitochondrial membrane potential stability under hypoxia. In vivo, the expression of miR-483-3p and Cdk9 was tested in the cardiac tissue of the mice with acute myocardial infarction, and the expression of miR-483-3p decreased and Cdk9 increased in the region of myocardial infarction. However, miR-483-3p was overexpressed with lentivirus, which suppressed apoptosis, infarct size (miR-483-3p,22.00±4.04% versus negative control,28.57±5.44%, P<0.05), and Cdk9 expression to improve cardiac contractility. Conclusions MiR-483-3p antagonizes hypoxia, leading to cardiomyocyte injury by targeting Cdk9, which is a new mechanism of therapeutic hypothermia.

Pro-fibrotic and apoptotic activities of circARAP1 in myocardial ischemia–reperfusion injury

Reperfusion modality can cause damage to cardiomyocytes, known as myocardial ischemia–reperfusion injury (MI/RI). Circular RNAs (circRNAs) are fundamental regulators associated with many cardiac diseases, including MI/RI. However, their functional impact on cardiomyocyte fibrosis and apoptosis remains elusive. Therefore, this study aimed to explore possible molecular mechanisms of circARPA1 in animal models and in hypoxia/reoxygenation (H/R)-treated cardiomyocytes. GEO dataset analysis showed that has_circ_0023461 (circARPA1) was differentially expressed in myocardial infarction samples. Real-time quantitative PCR further supported that circARPA1 was expressed at high levels in animal models and in H/R-triggered cardiomyocytes. Then, loss-of-function assays were performed to show that circARAP1 suppression effectively ameliorated cardiomyocyte fibrosis and apoptosis in MI/RI mice. Mechanistic experiments showed that miR-379-5p, KLF9 and Wnt signaling pathways were associated with circARPA1. circARPA1 can sponge miR-379-5p to regulate KLF9 expression, thereby activating the wnt/β-catenin pathway. Finally, gain-of-function assays revealed that circARAP1 aggravated MI/RI in mice and H/R-induced cardiomyocyte injury by regulating the miR-379-5p/KLF9 axis to activate Wnt/β-catenin signaling.

Downregulation of NUP93 aggravates hypoxia-induced death of cardiomyocytes in vitro through abnormal regulation of gene transcription.

Nuclear pore complex in the nuclear envelope plays an important role in controlling the transportation of RNAs, proteins and other macromolecules between the nucleus and cytoplasm. The relationship between abnormal expression of nucleoporins and cardiovascular diseases is unclear. In this study we investigated how myocardial infarction affected the expression and function of nucleoporins in cardiomyocytes. We separately knocked down 27 nucleoporins in rat primary myocardial cells. Among 27 nucleoporins, knockdown of Nup93, Nup210 and Nup214 markedly increased the expression of ANP and BNP, two molecular markers of cardiomyocyte function. We showed that Nup93 was significantly downregulated in hypoxic cardiomyocytes. Knockdown of Nup93 aggravated hypoxia-induced injury and cell death of cardiomyocytes, whereas overexpression of Nup93 led to the opposite effects. RNA-seq and bioinformatics analysis revealed that knockdown of Nup93 did not affect the overall transportation of mRNAs from the nucleus to the cytoplasm, but regulated the transcription of a large number of mRNAs in cardiomyocytes, which aremainly involved in oxidative phosphorylation and ribosome subunits. Most of the down-regulated genes by Nup93 knockdown overlapped with the genes whose promoters could be directly bound by Nup93. Among these genes, we demonstrated that Nup93 knockdown significantly down-regulated the expression of YAP1. Overexpression of YAP1 partially rescued the function of Nup93 knockdown and attenuated the effects of hypoxia on cell injury and cardiomyocyte death. We conclude that down-regulation of Nup93, at least partially, contributes to hypoxia-induced injury and cardiomyocyte death through abnormal interaction with the genome to dynamically regulate the transcription of YAP1 and other genes. These results reveal a new mechanism of Nup93 and might provide new therapeutic targets for the treatment of ischemia-induced heart failure.

Serum and glucocorticoid inducible kinase 1 modulates mitochondrial dysfunction and oxidative stress in doxorubicin-induced cardiomyocytes by regulating Hippo pathway via Neural precursor cell-expressed developmentally down-regulated 4 type 2.

Doxorubicin (Dox) was reported to cause mitochondrial dysfunction and oxidative stress in cardiomyocytes, leading to cardiomyocyte apoptosis and ultimately heart failure. Serum and glucocorticoid inducible kinase 1 (SGK1) participates in the progression of various cardiovascular diseases. Thus, we aimed to explore the role and regulatory mechanism of SGK1 in Dox-induced cardiomyocyte injury. The expression of SGK1 was evaluated in blood samples of heart failure children, and in myocardial tissues and blood samples of Dox-induced rats. Subsequently, we treated cardiomyocytes with Dox in vitro. A gain-of-function assay was performed to assess the effects of SGK1 on mitochondrial dysfunction and oxidative stress in Dox-induced cardiomyocytes. Furthermore, the modulation of SGK1 on Neural precursor cell-expressed developmentally down-regulated 4 type 2 (NEDD4-2) expression and the subsequent Hippo pathway was validated. In our study, we found that SGK1 was downregulated in blood samples of heart failure children, as well as myocardial tissues and blood samples of Dox-induced rats. SGK1 overexpression alleviated the decreases of mitochondrial complex activity, mitochondrial membrane potential, adenosine triphosphate (ATP) content and ATP synthetase activity stimulated by Dox. Besides, SGK1 overexpression reversed the promoting effects of Dox on oxidative stress and apoptosis. Mechanistically, SGK1 overexpression inhibited the expression of NEDD4-2 and blocked the subsequent activation of Hippo pathway. NEDD4-2 overexpression or activation of Hippo reversed the protective effects of SGK1 overexpression on Dox-induced cardiomyocyte injury. In conclusion, our results revealed that SGK1 modulated mitochondrial dysfunction and oxidative stress in Dox-induced cardiomyocytes by regulating Hippo pathway via NEDD4-2.

Metformin alleviates ethanol-induced cardiomyocyte injury by activating AKT/Nrf2 signaling in an ErbB2-dependent manner.

Metformin, a first-line oral anti-diabetic drug, has recently been reported to exert protective effect on various cardiovascular diseases. However, the potential role of metformin in ethanol-induced cardiomyocyte injury is still unknown. Therefore, this study was aimed to investigate the effect of metformin on ethanol-induced cardiomyocyte injury and its underlying mechanism. H9c2 cardiomyocytes were exposed to ethanol for 24h to establish an ethanol-induced cardiomyocyte injury model, and followed by treatment with metformin in the presence or absence of Lapatinib (an ErbB2 inhibition). CCK8 and LDH assays demonstrated that metformin improved cell viability in cardiomyocytes exposed to ethanol. Furthermore, metformin suppressed cardiomyocyte apoptosis and reduced the expressions of apoptosis-related proteins (Bax and C-CAS-3). In addition, our results showed that metformin activated the AKT/Nrf2 pathway, and then promoted Nrf2 nuclear translocation and the transcription of its downstream antioxidant genes (HO-1, CAT and SOD2), thereby inhibiting oxidative stress. Interestingly, we found that ErbB2 protein expression was significantly inhibited in ethanol-treated cardiomyocytes, which was markedly reversed by metformin. In contrast, Lapatinib largely abrogated the activation of AKT/Nrf2 signaling by metformin, accompanied by the increases in oxidative stress and cardiomyocyte apoptosis, indicating that metformin prevented ethanol-induced cardiomyocyte injury in an ErbB2-dependent manner. In summary, our study provides the first evidence that metformin protects cardiomyocyte against ethanol-induced oxidative stress and apoptosis by activating ErbB2-mediated AKT/Nrf2 signaling. Thus, metformin may be a potential novel treatment approach for alcoholic cardiomyopathy.

MicroRNA-494 regulates high glucose-induced cardiomyocyte apoptosis and autophagy by PI3K/AKT/mTOR signalling pathway.

Diabetic cardiomyopathy (DCM) is one of the major cardiovascular complications of diabetes. However, the mechanism of DCM is not fully understood. Studies have confirmed that certain microRNAs (miRNAs/miRs) are key regulators of DCM. The aim of this study was to investigate the role and mechanism of microRNA (miR)-494 in cardiomyocyte apoptosis and autophagy induced by high glucose (HG). By establishing a rat DCM model and an HG-treated H9c2 cells injury model, cardiac function was detected by echocardiography, myocardial tissue was stained by immunohistochemistry, and Cell Counting Kit-8 assay and lactate dehydrogenase assay were used to detect the cardiomyocyte injury. Cell apoptosis was detected by terminal deoxynucleotidyl transferase dUTP nick end labelling staining, and western blotting was used to detect death and autophagy. The results showed that the expression level of miR-494 was higher in the myocardial tissue of DCM rats and the myocardial cells of H9c2 treated with HG. Compared with the corresponding negative control groups, miR-494 mimics enhanced HG-induced apoptosis and autophagy, whereas miR-494 inhibitors showed the opposite effect, corresponding PI3K, AKT, and mTOR phosphorylation level has changed. These findings identify that miR-494 could regulate cell apoptosis and autophagy through PI3K/AKT/mTOR signalling pathway, participating in the regulation of cardiomyocyte cell damage after HG. These findings provide new insights for the further study of the molecular mechanism and treatment of DCM.

Qiangxin recipe improves doxorubicin-induced chronic heart failure by enhancing KLF5-mediated glucose metabolism

BackgroundQiangxin recipe (QXF) is a well-known Chinese herbal medicine commonly used in Asia for thousands of years to treat cardiovascular diseases, but its underlying mechanism remains unclear. PurposeThis study aimed to illustrate whether Qiangxin Recipe (QXF) induce glucose metabolism and inhibit cardiomyocyte apoptosis by promoting the activation of the transcription factor Krüppel like factor 5 (KLF5). Material and methodsIn vitro experiments, we constructed an H9C2 cardiomyocyte injury model using doxorubicin and used RNA-seq data analysis to detect the mechanism of QXF. In in vivo experiments, C57 BL/6 mice injected with doxorubicin (4 mg/kg every 6 days, for 30 days) to construct a CHF mouse model and randomly divided into to the normal control group, Dox group and Dox+QXF group (2.12 g/kg/day, 4.24 g/kg/day, for 30 days). Using Echocardiography, serum biochemical indices BNP, cTnl; and histopathological tests involving HE staining, Tunel staining and Immuno-dual fluorescence colocalization to analyze the therapeutic mechanism of QXF. ResultsWe verified that the Qiangxin recipe could reverse cardiomyocyte dying through enhancing glucose metabolism and reducing apoptosis to improve CHF. Mechanistically, we discovered that the Qiangxin recipe promoted the activation of transcription factor Krüppel-like factor 5 (KLF5) to induce glucose metabolism and inhibit apoptosis in cardiomyocytes. Further, we identified that KLF5 increased the promoter activity of hexokinase 2 (HK2) and B-cell CLL/lymphoma 2 (BCL2) genes, which further enhanced glucose metabolism and inhibited apoptosis of cardiomyocytes. ConclusionsWe highlighted the importance of KLF5-mediated signaling pathways in the treatment of CHF as shown by their participation in glucose metabolism and apoptosis in a doxorubicin-induced model of cardiomyocyte injury, as well as show that Qiangxin recipe can be used as a novel targeted therapy for the treatment of CHF. Compared with previous studies, we provide new ideas for the treatment of Doxorubicin-induced CHF from the perspective of energy metabolism.

Long noncoding RNA ACART knockdown decreases 3T3-L1 preadipocyte proliferation and differentiation.

Obesity is a main risk factor for diabetes and cardiovascular disorders and is closely linked to preadipocyte differentiation or adipogenesis. Peroxisome proliferator-activated receptor γ (PPARγ) is an indispensable transcription factor in adipogenesis. A newly identified long noncoding RNA, Acart, exerts a protective effect against cardiomyocyte injury by transactivating PPARγ signaling. However, the function of Acart in preadipocyte differentiation is unclear. To investigate the function of Acart in adipogenesis, a well-established preadipocyte, the 3T3-L1 cell line, was induced to differentiate, and Acart level was assessed during differentiation using quantitative real-time PCR. The biological role of Acart in adipogenesis was analyzed by assessing lipid droplet accumulation, PPARγ and CCAAT/enhancer-binding protein α (C/EBPα) expression, and 3T3-L1 cell proliferation and apoptosis after Acart silencing. We found that Acart level was promptly increased during preadipocyte differentiation in vitro. Acart was also significantly upregulated in obese mouse-derived subcutaneous, perirenal, and epididymal fat tissues compared with nonobese mouse-derived adipose tissues. Functionally, Acart depletion inhibited preadipocyte differentiation, as evidenced by a significant decrease in lipid accumulation and PPARγ and C/EBPα expression levels. Acart silencing also inhibited 3T3-L1 cell proliferation, whereas Acart overexpression accelerated 3T3-L1 cell proliferation and decreased cell apoptosis. Taken together, the current results reveal a novel function of Acart in regulating preadipocyte proliferation and differentiation.

The association of Hs-Troponin I with cardiac dysfunction and structural changes in chronic heart failure with reduced ejection fraction patients

Introduction: Cardiomyocyte death is the key pathological feature of heart failure with reduced ejection fraction (HFrEF). One of the circulating markers which represent myocardial cell death is cardiac troponin. High sensitivity-troponin I (hs-TnI) has been validated as an acute coronary syndrome diagnostic criteria. But its value and role in chronic heart failure, specifically HFrEF, has been variable in different studies. The aim of this study was to determine whether hs-TnI, an indicator of cardiomyocyte injury, is associated with functional and structural changes in HFrEF patients. Methods: A hospital-based cross-sectional analytic study was conducted on 72 patients with stable chronic HFrEF between April and October 2022. All subjects underwent echocardiography (Philips EPIQ7 and Affinity 70) and hs-troponin I serum measurement (Abbott Architect assay). Spearman’s test was used to evaluate the correlation between hs-troponin I and left ventricular ejection fraction (LVEF), E/A ratio, left ventricular end-diastolic volume (LVEDV), tricuspid annular plane systolic excursion (TAPSE) and left atrial volume index (LAVI). Results: hs-TnI concentrations were quantifiable in all samples ranging from 10 to 280.9 pg/mL. hs-TnI was associated significantly with LVEF (r=-0.39; p&lt;0.01), E/A ratio (r=0.328, p=0.01), and LVEDV (r=0.278, p=0.04). hs-TnI was not correlated with TAPSE and LAVI (r=0.241; p=0.07 and r=-0.13; p=0.429, respectively). The correlation between hs-troponin I with LVEF, E/A ratio and LVEDV proved that the degree of myocardial injury is directly associated with left ventricle (LV) systolic and diastolic dysfunction and also with LV dilatation. While it is not correlated with right ventricle (RV) systolic function. Hs-TnI may represent the injury in LV better than RV. Conclusion: These findings support the association of hs-TnI as a circulating marker of cardiomyocyte injury with LV dysfunction (systolic and diastolic) and dilatation in chronic HFrEF patients. This has expanded the role of hs-TnI in chronic heart disease.

Extracellular Histone-Induced Protein Kinase C Alpha Activation and Troponin Phosphorylation Is a Potential Mechanism of Cardiac Contractility Depression in Sepsis.

Reduction in cardiac contractility is common in severe sepsis. However, the pathological mechanism is still not fully understood. Recently it has been found that circulating histones released after extensive immune cell death play important roles in multiple organ injury and disfunction, particularly in cardiomyocyte injury and contractility reduction. How extracellular histones cause cardiac contractility depression is still not fully clear. In this work, using cultured cardiomyocytes and a histone infusion mouse model, we demonstrate that clinically relevant histone concentrations cause significant increases in intracellular calcium concentrations with subsequent activation and enriched localization of calcium-dependent protein kinase C (PKC) α and βII into the myofilament fraction of cardiomyocytes in vitro and in vivo. Furthermore, histones induced dose-dependent phosphorylation of cardiac troponin I (cTnI) at the PKC-regulated phosphorylation residues (S43 and T144) in cultured cardiomyocytes, which was also confirmed in murine cardiomyocytes following intravenous histone injection. Specific inhibitors against PKCα and PKCβII revealed that histone-induced cTnI phosphorylation was mainly mediated by PKCα activation, but not PKCβII. Blocking PKCα also significantly abrogated histone-induced deterioration in peak shortening, duration and the velocity of shortening, and re-lengthening of cardiomyocyte contractility. These in vitro and in vivo findings collectively indicate a potential mechanism of histone-induced cardiomyocyte dysfunction driven by PKCα activation with subsequent enhanced phosphorylation of cTnI. These findings also indicate a potential mechanism of clinical cardiac dysfunction in sepsis and other critical illnesses with high levels of circulating histones, which holds the potential translational benefit to these patients by targeting circulating histones and downstream pathways.

Kazinol B protects H9c2 cardiomyocytes from hypoxia/reoxygenation-induced cardiac injury by modulating the AKT/AMPK/Nrf2 signalling pathway

Context Kazinol B (KB), an isoprenylated flavan derived from Broussonetia kazinoki Sieb. (Moraceae) root, has long been used in folk medicine. Objective This study examines the protective effects of KB and its underlying mechanisms in hypoxia and reoxygenation (H/R)-induced cardiac injury in H9c2 rat cardiac myoblasts. Materials and methods H9c2 cells were incubated with various concentrations of KB (0, 0.3, 1, 3, 10 and 30 μM) for 2 h and then subjected to H/R insults. The protective effects of KB and its underlying mechanisms were explored. Results KB significantly elevated cell viability (1 μM, 1.21-fold; 3 μM, 1.36-fold, and 10 μM, 1.47-fold) and suppressed LDH release (1 μM, 0.77-fold; 3 μM, 0.68-fold, and 10 μM, 0.59-fold) in H/R-induced H9c2 cells. Further, 10 μM KB blocked apoptotic cascades, as shown by the Annexin-V/PI (0.41-fold), DNA fragmentation (0.51-fold), caspase-3 (0.52-fold), PARP activation (0.27-fold) and Bax/Bcl-2 expression (0.28-fold) assays. KB (10 μM) downregulated reactive oxygen species production (0.51-fold) and lipid peroxidation (0.48-fold); it upregulated the activities of GSH-Px (2.08-fold) and SOD (1.72-fold). KB (10 μM) induced Nrf2 nuclear accumulation (1.94-fold) and increased ARE promoter activity (2.15-fold), HO-1 expression (3.07-fold), AKT (3.07-fold) and AMPK (3.07-fold) phosphorylation. Nrf2 knockdown via using Nrf2 siRNA abrogated KB-mediated protective effects against H/R insults. Moreover, pharmacological inhibitors of AKT and AMPK also abrogated KB-induced Nrf2 activation and its protective function. Discussion and conclusions KB prevented H/R-induced cardiomyocyte injury via modulating the AKT and AMPK-mediated Nrf2 induction. KB might be a promising drug candidate for managing ischemic cardiac disorders.

Protective Effect of Sevoflurane Preconditioning on Cardiomyocytes Against Hypoxia/Reoxygenation Injury by Modulating Iron Homeostasis and Ferroptosis.

To investigate the mechanism whereby sevoflurane (Sev) protects cardiomyocytes from hypoxia/reoxygenation (H/R) injury. The rat cardiomyocyte line H9C2 was exposed to hypoxia (1% oxygen) for 24h, followed by reoxygenation for 2h to construct a model of H/R injury. H9C2 was exposed to 2.4% Sev for 45min before creating a hypoxic environment to observe the effect of Sev. MTT was taken to assess the viability of each group of cells, flow cytometry to detect cell apoptosis, and qRT-PCR or western blot to detect the expression of iron metabolism-related proteins and apoptosis-related proteins in the cells. And the kit determined the levels of total Fe and Fe2+ as well as factors related to oxidative stress in the cells. Administration of Sev significantly increased the cell viability of the H/R group while decreasing the expression of apoptosis-related proteins (Bax, cleaved caspase-3). Ferroportin 1 and mitochondrial ferritin, which are associated with iron metabolism, were considerably up-regulated by Sev, while iron regulatory protein 1, divalent metal transporter 1, and transferrin receptor 1 were significantly down-regulated in H/R cells. Additionally, Sev substantially reduced the levels of total Fe and Fe2+, reactive oxygen species, malondialdehyde, and 4-hydroxynonenal in H/R cells. In conclusion, Sev relieves H/R-induced cardiomyocyte injury by regulating iron homeostasis and ferroptosis.

PON2 ameliorates Ang II-induced cardiomyocyte injury by targeting the CANX/NOX4 signaling pathway.

The incidence of heart failure (HF) presents an escalating trend annually, second only to cancer. Few literatures are available regarding on the role of paraoxonase 2 (PON2) in HF so far despite the protective role of PON2 in cardiovascular diseases. PON2 expression in AC16 cells was examined with reverse transcriptase-quantitative polymerase chain reaction and western blot following angiotensin II (Ang II) challenging. After PON2 elevation, 2, 7-dichlorofluorescein diacetate assay estimated reactive oxygen species content, related kits appraised oxidative stress, enzyme-linked immunosorbent assayevaluated inflammatory levels, and Western blot was applied to the analysis of apoptosis levels. Research on cytoskeleton was conducted by immunofluorescence (IF), and Western blot analysis of the expressions of hypertrophy-related proteins was performed. BioGRID and GeneMania databases were used to analyze the relationship between PON2 and Calnexin(CANX), which was corroborated by co-immunoprecipitation experiment. Subsequently, PON2 and CANX were simultaneously overexpressed in AC16 cells induced by Ang II to further figure out the mechanism. PON2 expression was depleted in Ang II-induced AC16 cells. PON2 might mediate CANX/NOX4 signaling to inhibit oxidation, inflammatory, hypertrophy, and damage in Ang II-induced AC16 cells. PON2 can ease Ang II-induced cardiomyocyte injury via targeting CANX/NOX4 signaling.

LncRNA Fendrr: involvement in the protective role of nucleolin against H2O2-induced injury in cardiomyocytes

ABSTRACTBackground: Nucleolin is a multifunctional nucleolar protein with RNA-binding properties. Increased nucleolin expression protects cells from H2O2-induced damage, but the mechanism remains unknown. Long noncoding RNAs (lncRNAs) play crucial roles in cardiovascular diseases. However, the biological functions and underlying mechanisms of lncRNAs in myocardial injury remain unclear.Methods: In a nucleolin-overexpressing cardiac cell line, high-throughput technology was used to identify lncRNAs controlled by nucleolin. Cell counting kit-8 assay was used to determine cell viability, lactate dehydrogenase (LDH) assay to detect cell death, caspase activity assay and propidium iodide staining to confirm cell apoptosis, and RNA immunoprecipitation to examine the interaction between Fendrr and nucleolin.Results: We found that Fendrr expression was significantly downregulated in mouse hearts subjected to myocardial ischemia-reperfusion (MI/R) injury. High Fendrr expression abrogated H2O2-mediated injury in cardiomyocytes as evidenced by increased cell viability and decreased cell apoptosis. Conversely, Fendrr knockdown exacerbated the cardiomyocytes injury. Also, nucleolin overexpression inhibits Fendrr downregulation in H2O2-induced cardiomyocyte injury. Fendrr overexpression significantly reversed the role of the suppression of nucleolin expression in H2O2-induced cardiomyocytes.Conclusion: LncRNA Fendrr is involved in the cardioprotective effect of nucleolin against H2O2-induced injury and may be a potential therapeutic target for oxidative stress-induced myocardial injury.