MiR-181a protects the heart against myocardial infarction by regulating mitochondrial fission via targeting programmed cell death protein 4.

Worldwide, myocardial infarction (MI) is the leading cause of death and disability-adjusted life years lost. Recent researches explored new methods of detecting biomarkers that can predict the risk of developing myocardial infarction, which includes identifying genetic markers associated with increased risk. We induced myocardial infarction in mice by occluding the left anterior descending coronary artery and performed TTC staining to assess cell death. Next, we performed ChIP assays to measure the enrichment of histone modifications at the promoter regions of key genes involved in mitochondrial fission. We used qPCR and western blot to measure expression levels of relative apoptotic indicators. We report that miR-181a inhibits myocardial ischemia-induced apoptosis and preserves left ventricular function after MI. We show that programmed cell death protein 4 (PDCD4) is the target gene involved in miR-181a-mediated anti-ischemic injury, which enhanced BID recruitment to the mitochondria. In addition, we discovered that p53 inhibits the expression of miR-181a via transcriptional regulation. Here, we discovered for the first time a mitochondrial fission and apoptosis pathway which is controlled by miR-181a and involves PDCD4 and BID. This pathway may be controlled by p53 transcriptionally, and we presume that miR-181a may lead to the discovery of new therapeutic and preventive targets for ischemic heart diseases.

18F-FDG PET imaging-monitored anti-inflammatory therapy for acute myocardial infarction: Exploring the role of MCC950 in murine model.

BACKGROUND: MCC950 is a novel NLRP3 inflammasome inhibitor that possesses potent anti-inflammatory properties in acute myocardial infarction (AMI). However, the lack of noninvasive monitoring methods limits its potential clinical translation. Thus, we sought to investigate whether 18F-FDG PET imaging can monitor the therapeutic effects of MCC950 in an AMI murine model. METHODS: C57BL/6 mice were used to generate an AMI model. MCC950 or sterile saline was intraperitoneally administered 1 hour after surgery and then daily for 7 consecutive days. 18F-FDG PET (inflammation) imaging was used to monitor inflammatory changes on days 3 and 5. Immunohistochemistry and Western blot were used to detect inflammatory markers and to confirm the PET imaging results. 18F-FDG PET (viability) imaging was used to quantitate the viability defect expansion on days 7 and 28. Cardiac ultrasound and survival analyses were performed to evaluate the cardiac function and survival rate. Adverse remodeling was determined by Wheat Germ Agglutinin (WGA) and Masson trichrome staining. RESULTS: The FDG-PET (inflammation) imaging revealed that MCC950 treatment led to lower 18F-FDG inflammatory uptakes, at the infarct region, on days 3 and 5 when compared to the MI group. The decreased M1 macrophages and neutrophils infiltration and the remission of the NLRP3/IL-1ß pathway, confirmed the FDG-PET (inflammation) imaging results. The FDG-PET (viability) imaging revealed that MCC950 significantly decreased the expansion of the viability defect, demonstrating its myocardial preservation effects. The acute FDG-PET (inflammation) signal positively correlated with the late viability defect and with the reduction in the left ventricular ejection fraction (LVEF). Additionally, the alleviated adverse remodeling and the improved survival rate further support the anti-inflammatory efficiency of MCC950 in AMI. CONCLUSION: Using 18F-FDG PET imaging, we noninvasively demonstrated the therapeutic effects of MCC950 in AMI and showed that 18F-FDG PET imaging holds promising application potentials in MCC950's clinical translation.

Parkin overexpression alleviates cardiac aging through facilitating K63-polyubiquitination of TBK1 to facilitate mitophagy.

Cumulative clinical and experimental evidence has revealed a cardinal role for mitochondrial integrity in cardiac aging. Parkin-mediated mitophagy is essential to ensure mitochondrial quality control in myocardium. This study was designed to examine the impact of Parkin overexpression on aging-induced myocardial anomalies and the underlying mechanisms with a focus on Parkin-regulated mitophagy. Cardiac function, myocardial apoptosis, mitochondrial ultrastructure and mitophagy were examined in young (3 mo) and old (24-26 mo) wild-type (WT) and Parkin transgenic mice. Our data revealed compromised myocardial function and mitochondrial morphology along with overtly apoptosis with advanced aging, the effects of which were attenuated by Parkin overexpression. Advanced aging dampened mitophagy as evidenced by decreased levels of Parkin, LC3II, phosphorylation of p62 and TBK1 in isolated mitochondria as well as reduced mitochondria autophagosomes, the effects of which were mitigated by restoration of mitophagy via Parkin overexpression. Using the low-dose doxorubicin (DOX) in vitro model of cell senescence, we noted that Parkin-offered beneficial effect against senescence was abolished by the TBK1 kinase inhibitor BX795. With TBK1 overexpression in cardiomyocytes, we uncovered the interaction of Parkin with TBK1 using a Co-immunoprecipitation (Co-IP) assay. The interaction of Parkin with TBK1 contributed to K63-linked polyubiquitination of TBK1. Our study also noted that DOX disturbed K63-linked polyubiquitination of TBK1 with downregulation of Parkin. Parkin overexpression promoted K63-linked polyubiquitination of TBK1 through Lys30 and Lys401 residues to foster TBK1 phosphorylation to facilitate efficient mitophagy. In summary, these findings suggested that Parkin effectively rescued cardiac aging through promoting K63-linked polyubiquitination of TBK1 to facilitate mitophagy.

Predictive model for acute abdominal pain after transarterial chemoembolization for liver cancer.

BACKGROUND: Transarterial chemoembolization (TACE) is the first-line treatment for patients with unresectable liver cancer; however, TACE is associated with postembolization pain. AIM: To analyze the risk factors for acute abdominal pain after TACE and establish a predictive model for postembolization pain. METHODS: From January 2018 to September 2018, all patients with liver cancer who underwent TACE at our hospital were included. General characteristics; clinical, imaging, and procedural data; and postembolization pain were analyzed. Postembolization pain was defined as acute moderate-to-severe abdominal pain within 24 h after TACE. Logistic regression and a classification and regression tree were used to develop a predictive model. Receiver operating characteristic curve analysis was used to examine the efficacy of the predictive model. RESULTS: We analyzed 522 patients who underwent a total of 582 TACE procedures. Ninety-seven (16.70%) episodes of severe pain occurred. A predictive model built based on the dataset from classification and regression tree analysis identified known invasion of blood vessels as the strongest predictor of subsequent performance, followed by history of TACE, method of TACE, and history of abdominal pain after TACE. The area under the receiver operating characteristic curve was 0.736 [95% confidence interval (CI): 0.682-0.789], the sensitivity was 73.2%, the specificity was 65.6%, and the negative predictive value was 92.4%. Logistic regression produced similar results by identifying age [odds ratio (OR) = 0.971; 95%CI: 0.951-0.992; P = 0.007), history of TACE (OR = 0.378; 95%CI: 0.189-0.757; P = 0.007), history of abdominal pain after TACE (OR = 6.288; 95%CI: 2.963-13.342; P < 0.001), tumor size (OR = 1.978; 95%CI: 1.175-3.330; P = 0.01), multiple tumors (OR = 2.164; 95%CI: 1.243-3.769; P = 0.006), invasion of blood vessels (OR = 1.756; 95%CI: 1.045-2.950; P = 0.034), and TACE with drug-eluting beads (DEB-TACE) (OR = 2.05; 95%CI: 1.260-3.334; P = 0.004) as independent predictive factors for postembolization pain. CONCLUSION: Blood vessel invasion, TACE history, TACE with drug-eluting beads, and history of abdominal pain after TACE are predictors of acute moderate-to-severe pain. The predictive model may help medical staff to manage pain.

Sirt3 deficiency exacerbates diabetic cardiac dysfunction: Role of Foxo3A-Parkin-mediated mitophagy.

Diabetic cardiomyopathy (DCM) is often associated with suppressed cardiac autophagy, mitochondrial structural and functional impairment. Sirtuin-3 (Sirt3) has been reported to play a crucial role in mitochondrial homeostasis and confers a protective role against the onset and development of DCM although the precise mechanism(s) remains elusive. Here we hypothesized that Sirt3 exerts cardioprotection against DCM by activating Parkin-mediated mitophagy, en route to preserved mitochondrial homeostasis and suppressed cardiomyocyte apoptosis. Adult male wild-type (WT) and Sirt3 knockout (Sirt3KO) mice were treated with streptozotocin (STZ) or vehicle for 3months prior to assessment of echocardiographic property, interstitial fibrosis, cardiomyocyte apoptosis, mitochondrial morphology, cardiac autophagy and cell signaling molecules. Our findings revealed that STZ-induced diabetes mellitus prompted cardiac dysfunction, interstitial fibrosis, cardiomyocyte apoptosis and mitochondrial injury, accompanied with suppressed autophagy and mitophagy, the effects of which were aggravated by Sirt3KO. To the contrary, Sirt3 overexpression in vitro activated autophagy and mitophagy, inhibited mitochondrial injury and cardiomyocyte apoptosis, the effects of which were attenuated by autophagy inhibition using 3-MA. Moreover, deacetylation of Foxo3A and expression of Parkin were decreased by Sirt3KO, while these effects were facilitated by Sirt3OE in diabetic and high glucose settings. Taken together, our data suggested that suppressed Sirt3-Foxo3A-Parkin signaling mediated downregulation of mitophagy may play a vital role in the development of diabetic cardiomyopathy. This article is part of a Special Issue entitled: Genetic and epigenetic control of heart failure edited by Dr. Jun Ren & Yingmei Zhang.

Helix B surface peptide attenuates diabetic cardiomyopathy via AMPK-dependent autophagy.

BACKGROUND: Erythropoietin (EPO) has been reported to exert protective effects on a host of damaged tissues. However, the erythropoietic effect of this hormone can result in high risks of thrombosis, stroke, and hypertension, remarkably limiting the clinical use of EPO. Helix B surface peptide (HBSP) is a small peptide derived from the helix-B domain of EPO. Surprisingly, HBSP retains the tissue protective properties of EPO without altering the hematocrit. Thus, we evaluated the possible role of HBSP on diabetic cardiomyopathy. METHODS: Diabetes was induced in mice by intraperitoneal injections of streptozocin (STZ). Mice were randomly treated with normal saline or HBSP. Cardiac function, fibrosis, apoptosis, and myocardial mitochondrial morphology were examined. For in vitro experiments, H9C2 myoblast cells were randomly grouped as normal glucose (NG, 5 mM), NG+HBSP (100 nM), high glucose (HG, 33 mM), HG+HBSP (100 nM), HG+HBSP+3-methyladenine (3-MA, 10 mM), HG+rapamycin (Rapa, 100 nM), and HG+HBSP+Compound C (CC, 10 mM). Autophagosomes, LC3 dots, apoptosis and mitochondria membrane potential (MMP) of H9C2 cells were examined.The expressions of LC3, p62, p-AMPK (Thr172) and p-mTOR (Ser2448) were examined by Western blot. RESULTS: HBSP markedly improved cardiac function, attenuated cardiac interstitial fibrosis, inhibited myocardial apoptosis, and ameliorated mitochondrial ultrastructure in mice with diabetic cardiomyopathy. HG reduced autophagy in H9C2 cells. HBSP enhanced autophagy in HG-treated H9C2 cells. HBSP reduced the apoptosis index of HG-treated H9C2 cells. HBSP increased the MMP of HG-treated H9C2 cells. HBSP increased the levels of p-AMPK (Thr172), and reduced p-mTOR (Ser2448) in HG-treated H9C2 cells, and the increase of p-AMPK (Thr172) was accompanied by the stimulation of autophagy. Autophagy inhibitor 3-MA and AMPK inhibitor CC mitigated HBSP-induced beneficial effect, whereas autophagy inducer Rapa alleviated the HG-induced cell apoptosis. CONCLUSIONS: HBSP attenuates diabetic cardiomyopathy via autophagy mediated by AMPK-dependent pathway. HBSP may be a potential therapeutic intervention for diabetic cardiomyopathy.

Activation of Cannabinoid Receptor Type II by AM1241 Ameliorates Myocardial Fibrosis via Nrf2-Mediated Inhibition of TGF-ß1/Smad3 Pathway in Myocardial Infarction Mice.

AIMS: Myocardial interstitial fibrosis is a major histologic landmark resulting in cardiac dysfunction after myocardial infarction (MI). Activation of cannabinoid receptor type II (CB2 receptor) have been demonstrated to reduce fibrosis in hepatic cirrhotic rat. However, the anti-fibrotic effect of CB2 receptor activation in infarcted hearts was still unclear. In this study, we aimed to investigate the effects of a CB2 receptor selective agonist AM1241 on myocardial fibrosis post MI in mice. METHODS: Echocardiograph was conducted to assess cardiac function. Fibrosis markers such as type I and type III collagen, fibronectin, Plasminogen activator inhibitor(PAI)-1 and tissue inhibitor of metalloprotease(TIMP)-1 were examined by Western blot, while collagens were directly observed by Sirius-red staining. Primary cultured cardiac fibroblasts(CFs) were subjected to hypoxia/serum deprivation (H/SD) injury to simulate ischemic conditions in vivo. Nrf2 siRNA were applied to explore the role of Nrf2 and TGF-ß1/Smad3 pathway in this process. RESULTS: Echocardiography showed that AM1241 significantly improved cardiac function, suppressed the expression of fibrosis markers such as collagen I and collagen III, fibronectin, PAI-1 and TIMP-1 in mice with MI. In cardiac fibroblasts subjected to H/SD injury, AM1241 reduced the elevated levels of α-SMA, collagen I and collagen III, which were partially abrogated by the Nrf2 siRNA transfection. Furthermore, AM1241 not only activated and accelerated the translocation of Nrf2 to nucleus, but also inhibited TGF-ß1/ Smad3 pathway in an Nrf2 dependent manner. CONCLUSION: CB2 receptor agonist AM1241 alleviated myocardial interstitial fibrosis via Nrf2 -mediated down-regulation of TGF-ß1/Smad3 pathway, which suggested that CB2 receptor activation might represent a promising target for retarding cardiac fibrosis after MI.

Mst1 participates in the atherosclerosis progression through macrophage autophagy inhibition and macrophage apoptosis enhancement.

Emerging evidence favors the notion that macrophage autophagy plays a prominent role in the pathogenesis of vulnerable plaque, suggesting the therapeutic potential of targeting autophagy in atherosclerosis. Here ApoE(-/-) mice were crossed with Mst1 knockout or Mst1 Tg mice to generate ApoE(-/-):Mst1(-/-) and ApoE(-/-):Mst1Tg mice. All animals were fed high-fat-diet for 4months to induce arterial atherosclerosis. Murine macrophage RAW264.7 cells were subjected to ox-LDL (50µg/mL) in an effort to examine the cellular mechanisms. A significant increase in the levels of Mst1 and p-Mst1 was observed in the aorta of ApoE(-/-) mice. Mst1 knockout significantly reduced atherosclerotic area, decreased lipid core area and macrophage accumulation as compared with ApoE(-/-) mice. Along the same line, Mst1 overexpression increased plaque area, lipid core and macrophage accumulation as compared with ApoE(-/-) mice. Mst1 deficiency significantly increased levels of Beclin1 and LC3II, while decreased that of p62 in aortic atherosclerosis. Moreover, in vitro data indicated that Mst1 knockdown prompted more typical autophagosomes upon ox-LDL challenge. Mst1 knockdown also enhanced autophagic flux as evidenced by GFP-mRFP-LC3 staining, increased LC3-II expression and decreased p62 expression in the presence of bafilomycin A1. Mst1 knockdown decreased, while Mst1 overexpression increased macrophage apoptosis upon ox-LDL exposure. In conclusion, Mst1 deficiency diminishes atherosclerosis and stabilizes atherosclerotic plaques in ApoE(-/-) mice. Mst1 may participate in atherosclerosis progression through inhibition of macrophage autophagy and promotion of macrophage apoptosis.

Cardiac-Specific Knockout of ETA Receptor Mitigates Paraquat-Induced Cardiac Contractile Dysfunction.

Paraquat (1,1'-dim ethyl-4-4'-bipyridinium dichloride), a highly toxic quaternary ammonium herbicide widely used in agriculture, exerts potent toxic prooxidant effects resulting in multi-organ failure including the lung and heart although the underlying mechanism remains elusive. Recent evidence suggests possible involvement of endothelin system in paraquat-induced acute lung injury. This study was designed to examine the role of endothelin receptor A (ETA) in paraquat-induced cardiac contractile and mitochondrial injury. Wild-type (WT) and cardiac-specific ETA receptor knockout mice were challenged to paraquat (45 mg/kg, i.p.) for 48 h prior to the assessment of echocardiographic, cardiomyocyte contractile and intracellular Ca(2+) properties, as well as apoptosis and mitochondrial damage. Levels of the mitochondrial proteins for biogenesis and oxidative phosphorylation including UCP2, HSP90 and PGC1α were evaluated. Our results revealed that paraquat elicited cardiac enlargement, mechanical anomalies including compromised echocardiographic parameters (elevated left ventricular end-systolic and end-diastolic diameters as well as reduced factional shortening), suppressed cardiomyocyte contractile function, intracellular Ca(2+) handling, overt apoptosis and mitochondrial damage. ETA receptor knockout itself failed to affect myocardial function, apoptosis, mitochondrial integrity and mitochondrial protein expression. However, ETA receptor knockout ablated or significantly attenuated paraquat-induced cardiac contractile and intracellular Ca(2+) defect, apoptosis and mitochondrial damage. Taken together, these findings revealed that endothelin system in particular the ETA receptor may be involved in paraquat-induced toxic myocardial contractile anomalies possibly related to apoptosis and mitochondrial damage.