Exercise training decreases lactylation and prevents myocardial ischemia-reperfusion injury by inhibiting YTHDF2.

Exercise improves cardiac function and metabolism. Although long-term exercise leads to circulating and micro-environmental metabolic changes, the effect of exercise on protein post-translational lactylation modifications as well as its functional relevance is unclear. Here, we report that lactate can regulate cardiomyocyte changes by improving protein lactylation levels and elevating intracellular N6-methyladenosine RNA-binding protein YTHDF2. The intrinsic disorder region of YTHDF2 but not the RNA m6A-binding activity is indispensable for its regulatory function in influencing cardiomyocyte cell size changes and oxygen glucose deprivation/re-oxygenation (OGD/R)-stimulated apoptosis via upregulating Ras GTPase-activating protein-binding protein 1 (G3BP1). Downregulation of YTHDF2 is required for exercise-induced physiological cardiac hypertrophy. Moreover, myocardial YTHDF2 inhibition alleviated ischemia/reperfusion-induced acute injury and pathological remodeling. Our results here link lactate and lactylation modifications with RNA m6A reader YTHDF2 and highlight the physiological importance of this innovative post-transcriptional intrinsic regulation mechanism of cardiomyocyte responses to exercise. Decreasing lactylation or inhibiting YTHDF2/G3BP1 might represent a promising therapeutic strategy for cardiac diseases.

miR-486 attenuates cardiac ischemia/reperfusion injury and mediates the beneficial effect of exercise for myocardial protection.

Exercise and its regulated molecules have myocardial protective effects against cardiac ischemia/reperfusion (I/R) injury. The muscle-enriched miR-486 was previously identified to be upregulated in the exercised heart, which prompted us to investigate the functional roles of miR-486 in cardiac I/R injury and to further explore its potential in contributing to exercise-induced protection against I/R injury. Our data showed that miR-486 was significantly downregulated in the heart upon cardiac I/R injury. Both preventive and therapeutic interventions of adeno-associated virus 9 (AAV9)-mediated miR-486 overexpression could reduce cardiac I/R injury. Using AAV9 expressing miR-486 with a cTnT promoter, we further demonstrated that cardiac muscle cell-targeted miR-486 overexpression was also sufficient to protect against cardiac I/R injury. Consistently, miR-486 was downregulated in oxygen-glucose deprivation/reperfusion (OGDR)-stressed cardiomyocytes, while upregulating miR-486 inhibited cardiomyocyte apoptosis through PTEN and FoxO1 inhibition and AKT/mTOR activation. Finally, we observed that miR-486 was necessary for exercise-induced protection against cardiac I/R injury. In conclusion, miR-486 is protective against cardiac I/R injury and myocardial apoptosis through targeting of PTEN and FoxO1 and activation of the AKT/mTOR pathway, and mediates the beneficial effect of exercise for myocardial protection. Increasing miR-486 might be a promising therapeutic strategy for myocardial protection.

Extracellular Vesicles and Vascular Inflammation.

Vascular inflammation is the most common pathological feature in the pathogenesis of human disease. It is a complex immune process involved with many different types of cells including platelet, monocytes, macrophages, endothelial cells, and others. It is widely accepted that both innate and adaptive immune responses are important for the initiation and progression of vascular inflammation. The cell-cell interaction constitutes an important aspect of those immune responses in the vascular inflammation. Extracellular vesicles (EVs) are nanometer-sized double-layer lipid membrane vesicles released from most types of cells. They have been proved to play critical roles in intercellular communication in the occurrence and development of multisystem diseases. With the advancement of basal medical science, the biological roles of EVs in vascular inflammation have been clearer today. In this chapter, we will summarize the advance progress of extracellular vesicles in regulating vascular inflammation and its potential application in the clinical.

Correction to: Exercise-induced circulating extracellular vesicles protect against cardiac ischemia-reperfusion injury.

The original version of this article unfortunately contained a mistake.

Cardiac cell proliferation is not necessary for exercise-induced cardiac growth but required for its protection against ischaemia/reperfusion injury.

The adult heart retains a limited ability to regenerate in response to injury. Although exercise can reduce cardiac ischaemia/reperfusion (I/R) injury, the relative contribution of cardiac cell proliferation including newly formed cardiomyocytes remains unclear. A 4-week swimming murine model was utilized to induce cardiac physiological growth. Simultaneously, the antineoplastic agent 5-fluorouracil (5-FU), which acts during the S phase of the cell cycle, was given to mice via intraperitoneal injections. Using EdU and Ki-67 immunolabelling, we showed that exercise-induced cardiac cell proliferation was blunted by 5-FU. In addition, the growth of heart in size and weight upon exercise was unaltered, probably due to the fact that exercise-induced cardiomyocyte hypertrophy was not influenced by 5-FU as demonstrated by wheat germ agglutinin staining. Meanwhile, the markers for pathological hypertrophy, including ANP and BNP, were not changed by either exercise or 5-FU, indicating that physiological growth still developed in the presence of 5-FU. Furthermore, we showed that CITED4, a key regulator for cardiomyocyte proliferation, was blocked by 5-FU. Meanwhile, C/EBPß, a transcription factor responsible for both cellular proliferation and hypertrophy, was not altered by treatment with 5-FU. Importantly, the effects of exercise in reducing cardiac I/R injury could be abolished when cardiac cell proliferation was attenuated in mice treated with 5-FU. In conclusion, cardiac cell proliferation is not necessary for exercise-induced cardiac physiological growth, but it is required for exercise-associated protection against I/R injury.

Exercise-induced circulating extracellular vesicles protect against cardiac ischemia-reperfusion injury.

Extracellular vesicles (EVs) serve an important function as mediators of intercellular communication. Exercise is protective for the heart, although the signaling mechanisms that mediate this cardioprotection have not been fully elucidated. Here using nano-flow cytometry, we found a rapid increase in plasma EVs in human subjects undergoing exercise stress testing. We subsequently identified that serum EVs were increased by ~1.85-fold in mice after 3-week swimming. Intramyocardial injection of equivalent quantities of EVs from exercised mice and non-exercised controls provided similar protective effects against acute ischemia/reperfusion (I/R) injury in mice. However, injection of exercise-induced EVs in a quantity equivalent to the increase seen with exercise (1.85 swim group) significantly enhanced the protective effect. Similarly, treatment with exercise-induced increased EVs provided additional anti-apoptotic effect in H2O2-treated H9C2 cardiomyocytes mediated by the activation of ERK1/2 and HSP27 signaling. Finally, by treating H9C2 cells with insulin-like growth factor-1 to mimic exercise stimulus in vitro, we found an increased release of EVs from cardiomyocytes associated with ALIX and RAB35 activation. Collectively, our results show that exercise-induced increase in circulating EVs enhances the protective effects of endogenous EVs against cardiac I/R injury. Exercise-derived EVs might serve as a potent therapy for myocardial injury in the future.

Traditional Chinese Medication Qiliqiangxin Protects Against Cardiac Remodeling and Dysfunction in Spontaneously Hypertensive Rats.

Qiliqiangxin (QLQX), a traditional Chinese herbs medication, exerted protective effect in chronic heart failure patients in a multicenter randomized double-blind study. QLQX has also been found to improve cardiac function and reduce cardiac fibrosis in spontaneously hypertension animal model. However, the effect of longterm treatment with QLQX in such a condition and the related molecular mechanisms remain largely unknown. In the present study, thirteen-week-old spontaneously hypertensive rats (SHRs) were treated by daily intragastric administration of QLQX or saline for one year. Echocardiography, electron microscopy, and Masson's trichrome staining were used to determine cardiac function, mitochondria ultrastructure, and cardiac fibrosis, respectively. Quantitative reverse transcription polymerase chain reactions (qRT-PCRs) and Western blotting were used to determine gene expressions. We found that QLQX significantly improved cardiac function and reduced gene markers of pathological hypertrophy including ANP, BNP, and Myh7. QLQX also attenuated cardiac fibrosis and apoptosis in SHRs as evidenced by downregulation of α-SMA, collagen I, collagen III, and TGF-ß expressions and reduction of Bax to Bcl-2 ratio. Moreover, the damage of mitochondrial ultrastructure was greatly improved and the reduction of PPAR-α, PPAR-γ, and PGC-1α expression levels was significantly restored in SHRs by treatment with QLQX. In conclusion, longterm treatment with QLQX protects against cardiac remodeling and dysfunction in hypertension by increasing PPARs and PGC-1α.

Exosomes-Based Biomarkers for the Prognosis of Cardiovascular Diseases.

Cardiovascular diseases (CVDs) have a high prevalence and annually increasing incidence with high mortality and morbidity. Identification of biomarkers with high sensitivity and specificity for assessing the prognosis of CVDs is necessary for optimizing personalized treatment and reducing mortality. Exosomes have been proved to be accessible in nearly all body fluids and they can reflect disease stage or progression. Here we summarized exosomes-based biomarkers for the prognosis of coronary artery diseases, heart failure, stroke, hypertension, cardiac arrhythmia, cardiomyopathy, valvular heart diseases and pulmonary arterial hypertension. If exosome-based biomarkers can achieve additionally benefits as compared to the present prognostic biomarkers remains to be determined and multicenter studies with large cohorts of patients are highly needed.

Telocytes in exercise-induced cardiac growth.

Exercise can induce physiological cardiac growth, which is featured by enlarged cardiomyocyte cell size and formation of new cardiomyocytes. Telocytes (TCs) are a recently identified distinct interstitial cell type, existing in many tissues and organs including heart. TCs have been shown to form a tandem with cardiac stem/progenitor cells in cardiac stem cell niches, participating in cardiac regeneration and repair. Although exercise-induced cardiac growth has been confirmed as an important way to promote cardiac regeneration and repair, the response of cardiac TCs to exercise is still unclear. In this study, 4 weeks of swimming training was used to induce robust healthy cardiac growth. Exercise can induce an increase in cardiomyocyte cell size and formation of new cardiomyocytes as determined by Wheat Germ Lectin and EdU staining respectively. TCs were identified by three immunofluorescence stainings including double labelling for CD34/vimentin, CD34/platelet-derived growth factor (PDGF) receptor-α and CD34/PDGF receptor-ß. We found that cardiac TCs were significantly increased in exercised heart, suggesting that TCs might help control the activity of cardiac stem/progenitor cells, cardiomyocytes or endothelial cells. Adding cardiac TCs might help promote cardiac regeneration and renewal.

RNA m6A-Regulated circ-ZNF609 Suppression Ameliorates Doxorubicin-Induced Cardiotoxicity by Upregulating FTO.

Cardiac death is a major burden for cancer survivors, yet there is currently no effective treatment for doxorubicin (DOX)-induced cardiotoxicity. Here, we report that circ-ZNF609 knockdown knockdown had cardioprotective effects against DOX-induced cardiomyocyte toxicity. Mechanistically, circ-ZNF609 knockdown alleviated DOX-induced cardiotoxicity through attenuating cardiomyocyte apoptosis, reducing reactive oxygen species production, ameliorating mitochondrial nonheme iron overload. circ-ZNF609 inhibition blocked the elevation of RNA N6-methyladenosine (RNA m6A) methylation level in DOX-treated mice hearts, whereas m6A demethylase fat mass and obesity associated (FTO) acted as the downstream factor of circ-ZNF609. Moreover, the stability of circ-ZNF609 was regulated by RNA m6A methylation alteration, and suppression of RNA m6A methylation by methyltransferase like 14 (METTL14) modulated the function of circ-ZNF609. These data suggest that circ-ZNF609 inhibition represents a potential therapy for DOX-induced cardiotoxicity.

Diagnosis of acute myocardial infarction using a combination of circulating circular RNA cZNF292 and clinical information based on machine learning.

Circulating circular RNAs (circRNAs) are emerging as novel biomarkers for cardiovascular diseases (CVDs). Machine learning can provide optimal predictions on the diagnosis of diseases. Here we performed a proof-of-concept study to determine if combining circRNAs with an artificial intelligence approach works in diagnosing CVD. We used acute myocardial infarction (AMI) as a model setup to prove the claim. We determined the expression level of five hypoxia-induced circRNAs, including cZNF292, cAFF1, cDENND4C, cTHSD1, and cSRSF4, in the whole blood of coronary angiography positive AMI and negative non-AMI patients. Based on feature selection by using lasso with 10-fold cross validation, prediction model by logistic regression, and ROC curve analysis, we found that cZNF292 combined with clinical information (CM), including age, gender, body mass index, heart rate, and diastolic blood pressure, can predict AMI effectively. In a validation cohort, CM + cZNF292 can separate AMI and non-AMI patients, unstable angina and AMI patients, acute coronary syndromes (ACS), and non-ACS patients. RNA stability study demonstrated that cZNF292 was stable. Knockdown of cZNF292 in endothelial cells or cardiomyocytes showed anti-apoptosis effects in oxygen glucose deprivation/reoxygenation. Thus, we identify circulating cZNF292 as a potential biomarker for AMI and construct a prediction model "CM + cZNF292."

Exercise-induced circular RNA circUtrn is required for cardiac physiological hypertrophy and prevents myocardial ischaemia-reperfusion injury.

AIMS: Regular exercise training benefits cardiovascular health and effectively reduces the risk for cardiovascular disease. Circular RNAs (circRNAs) play important roles in cardiac pathophysiology. However, the role of circRNAs in response to exercise training and biological mechanisms responsible for exercise-induced cardiac protection remain largely unknown. METHODS AND RESULTS: RNA sequencing was used to profile circRNA expression in adult mouse cardiomyocytes that were isolated from mice with or without exercise training. Exercise-induced circRNA circUtrn was significantly increased in swimming-trained adult mouse cardiomyocytes. In vivo, circUtrn was found to be required for exercise-induced physiological cardiac hypertrophy. circUtrn inhibition abolished the protective effects of exercise on myocardial ischaemia-reperfusion remodelling. circUtrn overexpression prevented myocardial ischaemia-reperfusion-induced acute injury and pathological cardiac remodelling. In vitro, overexpression of circUtrn promoted H9 human embryonic stem cell-induced cardiomyocyte growth and survival via protein phosphatase 5 (PP5). Mechanistically, circUtrn directly bound to PP5 and regulated the stability of PP5 in a ubiquitin-proteasome-dependent manner. Hypoxia-inducible factor 1α-dependent splicing factor SF3B1 acted as an upstream regulator of circUtrn in cardiomyocytes. CONCLUSION: The circRNA circUtrn is upregulated upon exercise training in the heart. Overexpression of circUtrn can prevent myocardial I/R-induced injury and pathological cardiac remodelling.

Novel Analysis of Coronary Angiography in Predicting the Formation of Ventricular Aneurysm in Patients With Acute Myocardial Infarction After Percutaneous Coronary Intervention.

Background: Ventricular aneurysm (VA) is a serious complication of acute myocardial infarction (AMI), with a very poor prognosis. Early-stage prophylactic treatment is effective in preventing the formation of VAs. However, the existing predictive models for VA formation lack the sensitivity and specificity necessary for evaluating patients with MI. This study aimed to explore the potential use of coronary angiography and establish a more precise prediction model for VA in patients with MI. Methods: Patients with VA (n = 52) admitted to our medical center between June 2020 and July 2021 with previous emergency percutaneous coronary intervention for AMI were retrospectively included in this database study. Controls that matched 4:1 with the VA cases during the same period were enrolled. The baseline characteristics and coronary angiograms of the enrolled individuals were obtained from the electronic medical record system. The curve length of the distance from the main criminal lesion to its ostia (DLO) and distal (DLD) in the coronary artery were measured with ImageJ. Binary logistic regression analysis was used to identify the predictive factors. The model performance was evaluated by receiver operating characteristic curve analysis. Results: Binary analysis revealed maximum serum cardiac troponin I level (odds ratio [OR] = 1.046, 95% confidence interval [CI] = 1.027-1.066, P < 0.001), serum brain natriuretic peptide level (OR = 1.001, 95% CI = 1.000-1.002, P = 0.007), left anterior descending artery as the culprit lesion (OR = 5.091, 95% CI = 2.080-12.457, P < 0.001), and that single-vessel disease (OR = 1.809, 95% CI = 0.967-3.385, P < 0.001), stenosis in the main lesion (OR = 1.247, 95% CI = 1.173-1.327, P < 0.001), DLO (OR = 1.034, 95% CI = 1.019-1.049, P < 0.001), DLD (OR = 1.061, 95% CI = 1.043-1.079, P < 0.001), and DLD/DLD (OR = 0.033, 95% CI = 0.010-0.117, P < 0.001) were the independent variables for predicting VA formation in MI patients. Conclusion: Our study first used quantified information of coronary lesions to establish a predictive model and proved that a longer DLD had the greatest potential in predicting the incidence of VA. Its related parameters including DLO and DLO/DLD ratio were also correlated with the incidence of VA. These findings may provide a new reference for the early identification of high-risk MI patients and preventing VA.

Downregulation of circ-ZNF609 Promotes Heart Repair by Modulating RNA N6-Methyladenosine-Modified Yap Expression.

Circular RNAs take crucial roles in several pathophysiological processes. The regulatory role and its underlying mechanisms of circ-ZNF609 in the heart remains largely unknown. Here, we report that circ-ZNF609 is upregulated during myocardial ischemia/reperfusion (I/R) remodeling. Knockdown of circ-ZNF609 protects against acute I/R injury and attenuates left ventricle dysfunction after I/R remodeling in vivo. In vitro, circ-ZNF609 regulates cardiomyocyte survival and proliferation via modulating the crosstalk between Hippo-YAP and Akt signaling. Mechanically, N6-methyladenosine-modification is involved in the regulatory role of circ-ZNF609 on YAP. An in-depth study indicates that knockdown of circ-ZNF609 decreases the expression of YTHDF3 and further fine-tuned the accessibility of Yap mRNA to YTHDF1 and YTHDF2 to regulate YAP expression. circ-ZNF609 knockdown represents a promising therapeutic strategy to combat the pathological process of myocardial I/R injury.

METTL14 is required for exercise-induced cardiac hypertrophy and protects against myocardial ischemia-reperfusion injury.

RNA m6A modification is the most widely distributed RNA methylation and is closely related to various pathophysiological processes. Although the benefit of regular exercise on the heart has been well recognized, the role of RNA m6A in exercise training and exercise-induced physiological cardiac hypertrophy remains largely unknown. Here, we show that endurance exercise training leads to reduced cardiac mRNA m6A levels. METTL14 is downregulated by exercise, both at the level of RNA m6A and at the protein level. In vivo, wild-type METTL14 overexpression, but not MTase inactive mutant METTL14, blocks exercise-induced physiological cardiac hypertrophy. Cardiac-specific METTL14 knockdown attenuates acute ischemia-reperfusion injury as well as cardiac dysfunction in ischemia-reperfusion remodeling. Mechanistically, silencing METTL14 suppresses Phlpp2 mRNA m6A modifications and activates Akt-S473, in turn regulating cardiomyocyte growth and apoptosis. Our data indicates that METTL14 plays an important role in maintaining cardiac homeostasis. METTL14 downregulation represents a promising therapeutic strategy to attenuate cardiac remodeling.

Deregulated Cardiac Specific MicroRNAs in Postnatal Heart Growth.

The heart is recognized as an organ that is terminally differentiated by adulthood. However, during the process of human development, the heart is the first organ with function in the embryo and grows rapidly during the postnatal period. MicroRNAs (miRNAs, miRs), as regulators of gene expression, play important roles during the development of multiple systems. However, the role of miRNAs in postnatal heart growth is still unclear. In this study, by using qRT-PCR, we compared the expression of seven cardiac- or muscle-specific miRNAs that may be related to heart development in heart tissue from mice at postnatal days 0, 3, 8, and 14. Four miRNAs-miR-1a-3p, miR-133b-3p, miR-208b-3p, and miR-206-3p-were significantly decreased while miR-208a-3p was upregulated during the postnatal heart growth period. Based on these results, GeneSpring GX was used to predict potential downstream targets by performing a 3-way comparison of predictions from the miRWalk, PITA, and microRNAorg databases. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were used to identify potential functional annotations and signaling pathways related to postnatal heart growth. This study describes expression changes of cardiac- and muscle-specific miRNAs during postnatal heart growth and may provide new therapeutic targets for cardiovascular diseases.