Myocardial Cell Apoptosis Research Articles

Targeted activation of ERK1/2 reduces ischemia and reperfusion injury in hyperglycemic myocardium by improving mitochondrial function

Diabetes can increase the risk of coronary heart disease, and also increase the mortality rate of coronary heart disease in diabetic patients. Although reperfusion therapy can preserve the viable myocardium, fatal reperfusion injury can also occur. Studies have shown that diabetes can aggravate myocardial ischemia-reperfusion injury, ERK1/2 can reduce myocardial ischemia-reperfusion injury, but its mechanism in hyperglycemic myocardial ischemia-reperfusion injury is unclear. This study sought to explore the mechanism of extracellular signal-regulated kinase 1/2 (ERK1/2) in hyperglycemic myocardial ischemia reperfusion (I/R) injury. H9C2 cardiomyocytes were treated with high-glucose (HG) medium plus I/R stimulation to establish a hyperglycemia I/R model in vitro. The cells were treated with LM22B-10 (an ERK activator) or transfected with the constitutive activation of the mitogen-activated protein kinase 1 (CaMEK) gene. Myocardial cell apoptosis, mitochondria functional-related indicators, the oxidative stress indexes, and the expression levels of ERK1/2 protein were detected. The HG I/R injury intervention caused an increase in the ratio of apoptotic cardiomyocytes (P<0.05), but the phosphorylation level of the ERK1/2 protein did not increase further. Administering LM22B-10 or transfecting the CaMEK gene significantly activated the phosphorylation levels of ERK1/2 protein and reduced the proportion of cardiomyocyte apoptosis (P<0.05). HG I/R injury increased mitochondrial fission and reduced membrane potential. The intervention reduced the number of punctate mitochondria, increased the average network structure size and median branch length (P<0.01), increased the median network structure size and average branch length (P<0.05), and reduced the colocalization of Drp1 (Dynamin-Related protein1)/TOMM20 (Mitochondrial outer membrane translocation enzyme 20) (P<0.05) and Drp1 with serine 616 phosphorylation (Drp1s616) phosphorylation (P<0.01), thereby reducing mitochondrial fission, increasing membrane potential and mitochondrial function. HG I/R injury increased the level of oxidative stress, while administering LM22B-10 or transfecting the CaMEK gene reduced the level of oxidative stress (P<0.01). Targeting the activation of ERK1/2 protein phosphorylation reduced mitochondrial fission, increased membrane potential and mitochondrial function, reduced oxidative stress and myocardial cell apoptosis, and alleviated hyperglycemia myocardial I/R injury.

Expression of concern: The effects of tanshinone IIA on hypoxia/reoxygenation-induced myocardial microvascular endothelial cell apoptosis in rats via the JAK2/STAT3 signaling pathway [Biomedicine & Pharmacotherapy 83 (2016) 1116–1126

Expression of concern: The effects of tanshinone IIA on hypoxia/reoxygenation-induced myocardial microvascular endothelial cell apoptosis in rats via the JAK2/STAT3 signaling pathway [Biomedicine & Pharmacotherapy 83 (2016) 1116–1126

Visfatin Amplifies Cardiac Inflammation and Aggravates Cardiac Injury via the NF-κB p65 Signaling Pathway in LPS-Treated Mice.

Background Visfatin is an adipocytokine that has been demonstrated to be involved in cardiovascular diseases. This study aims at determining the role of visfatin in sepsis-induced cardiac injury and identify its possible mechanisms. Methods Dynamic changes in visfatin expression in mice with lipopolysaccharide- (LPS-) induced septicemia were measured. Additionally, mice were pretreated with visfatin and further administered LPS to observe the effects of visfatin on cardiac injury. Finally, septic mice were also pretreated with JSH-23 to investigate whether visfatin regulates cardiac injury via the NF-κB p65 pathway. Results Visfatin expression levels in both the heart and serum were increased in LPS-treated mice and peaked at 6 hours, and visfatin was derived from cardiac macrophages. In septic mice, pretreatment with visfatin reduced the survival rate, worsened cardiac dysfunction, and increased the expression of cardiac injury markers, including creatine kinase myocardial bound (CK-MB) and lactate dehydrogenase (LDH). Treatment with visfatin also increased the infiltration of CD3+ cells and F4/80+ cells, amplified the cardiac inflammatory response, and elevated myocardial cell apoptosis. Treatment with JSH-23 reversed the effects of visfatin in septic mice. Conclusions This study showed that visfatin amplifies the cardiac inflammatory response and aggravates cardiac injury through the p65 signaling pathway. Visfatin may be a clinical target for preventing cardiac injury in sepsis.

Research advance of the mechanism and treatment of septic cardiomyopathy

Sepsis is a life-threatening organ dysfunction caused by dysregulation of the body's response to infection. It is one of the common and serious complications in clinically critical patients with trauma, burn, shock, infection, etc., with high morbidity and mortality. Although the treatment of sepsis has made great achievements in clinical practice, the mortality of patients with sepsis is still increasing due to its secondary complications. Septic cardiomyopathy (SCM) is one of the major complications that threaten septic patient's life. SCM refers to myocardial dysfunction with the aggravation of the primary disease, which is manifested by biventricular dilatation accompanied by a decrease in left ventricular ejection fraction (LVEF). It is one of the major complications that threaten the life of patients with sepsis. The existing research shows that the mechanism of SCM includes myocardial mitochondrial dysfunction, myocardial cell apoptosis, calcium circulation disorder and its treatment including conventional treatment, β1 receptor blocker treatment and traditional Chinese medicine treatment,etc. This paper reviewed the pathogenesis of SCM and its related, in order to provide references for the rational diagnosis and treatment of SCM.

Polyphyllin I improves myocardial damage in coronary artery disease via modulating lipid metabolism and myocardial apoptosis.

Polyphyllin I (PPI) is a famous traditional medicine ingredient, which has been explored in wide range of areas. Nevertheless, whether PPI exerts any functions in coronary artery disease (CAD) is still uncertified. Herein, we probed the effect and mechanism of PPI on lipid metabolism and myocardial dysfunction in myocardial cells and CAD rat model. Hypoxia/reoxygenation (H/R)-treated H9c2 cells model was constructed for the in vitro experiments, and CAD model in vivo was established by high-fat feeding. After management with PPI, the correlated factors of lipid metabolism and myocardial function were investigated. The apoptosis of myocardial cells was assessed by Annexin V-FITC/PI kit and TUNEL staining. The apoptosis-associated factors (caspase 3, cleaved caspase 3, Bax, and Bcl-2) were tested by Western blot analysis. The MEK/ERK inhibitorwas applied and the functions of MEK/ERK pathway in myocardial damage were investigated. H/R-treated H9c2 cells model was constructed for the in vitro experiments, and CAD model in vivo was established by high-fat feeding. After management with PPI, the correlated factors of lipid metabolism and myocardial function were investigated. The apoptosis of myocardial cells was assessed by Annexin V-FITC/PI kit and TUNEL staining. The apoptosis-associated factors (caspase 3, cleaved caspase 3, Bax, and Bcl-2) were tested by Western blot analysis. The MEK/ERK inhibitorwas applied and the functions of MEK/ERK pathway in myocardial damage were investigated.PPI improved lipid metabolism disorder in H/R-induced H9c2 cells or in CAD rat model. Additionally, PPI attenuated myocardial dysfunction in CAD rats via enhancing left ventricular systolic pressure, maximum rate of change of left ventricular pressure (±dp/dtmax ), and arterial blood flow (CF). The apoptosis of myocardial cells was lessened by PPI management, which was further verified by reducing Bax and cleaved caspase 3 expression. Furthermore, PD0325901 (MEK/ERK inhibitor) weakened the effect of PPI on myocardial dysfunction, lipid metabolism, and myocardial cell apoptosis in CAD rats. The research confirmed the protective effect of PPI on myocardial damage in CAD, which was regulated by MEK/ERK pathway.

6-O-desulfated heparin attenuates myocardial ischemia/reperfusion injury in mice through the regulation of miR-199a-5p/klotho axis.

Heparin has been documented to reduce myocardial injury caused by ischemia/reperfusion (I/R), but its clinical application is limited due to its strong intrinsic anticoagulant property. Some desulfated derivatives of heparin display low anticoagulant activity and may have potential value as therapeutic agents for myocardial I/R injury. In this study, we observed that 6-O-desulfated heparin, a desulfated derivative of heparin, shortened the activated partial thromboplastin time and exhibited lower anticoagulant activity compared with heparin or 2-O-desulfated heparin (another desulfated derivative of heparin). Then, we explored whether 6-O-desulfated heparin could protect against myocardial I/R injury, and elucidated its possible mechanisms. Administration of 6-O-desulfated heparin significantly reduced creatine kinase activity, myocardial infarct size and cell apoptosis in mice subjected to 30min of myocardial ischemia following 2h of reperfusion, accompanied by a reverse in miR-199a-5p elevation, klotho downregulation and reactive oxygen species (ROS) accumulation. In cultured H9c2 cells, the mechanism of 6-O-desulfated heparin against myocardial I/R injury was further explored. Consistent with the results in vivo, 6-O-desulfated heparin significantly ameliorated hypoxia/reoxygenation-induced injury, upregulated klotho and decreased miR-199a-5p levels and ROS accumulation, and these effects were reversed by miR-199a-5p mimics. In conclusion, these results suggested that 6-O-desulfated heparin with lower anticoagulant activity attenuated myocardial I/R injury through miR-199a-5p/klotho and ROS signaling. Our study may also indicate that 6-O-desulfated heparin, as an excellent heparin derivative, is a potential therapeutic agent for myocardial I/R injury.

Dexmedetomidine reduces myocardial ischemia-reperfusion injury in young mice through MIF/AMPK/GLUT4 axis

BackgroundReperfusion of ischemic tissue has adverse impact on the myocardium. Dexmedetomidine (Dex) is a α2-adrenergic receptor (α2-AR) agonist with sedative and analgesic effects. Macrophage migration inhibition factor (MIF) is a pressure-regulating cytokine and is responsible for inflammatory and immune diseases. This study aims to reveal the consequences of Dex on myocardial ischemia-reperfusion injury (IRI) in young mice.MethodsFifty mice were raised and examined. At the end of the experiment, all mice were euthanized. The anterior descending department of the left coronary artery in mice was under ischemia for 60 min, then the ligation line was released and reperfused for 120 min to establish the IRI model. Mice were randomly divided into Sham, control, treatment using 4,5-dihydro-3-(4-hydroxyphenyl)-5-isoxazoleacetic acid (ISO-1), Dex treatment, and Dex combined ISO-1 treatment groups. Interleukin (IL)-6, IL-10 and tumor necrosis factor (TNF-α) were determined by enzyme-linked immunosorbent assay (ELISA). Reactive oxygen species (ROS) and ATP levels were recorded. The expressions of MIF, P-adenosine monophosphate-activated kinase α (AMPKα), glucose transporter (GLUT)4, Bax and Bcl-2 were detected by Western Blot (WB). Hematoxylin and Eosin (H&E) staining was used to study cell morphology. Apoptosis was detected by terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) assay. Echocardiography was carried out at the end of reperfusion, and the infarct size was calculated by Electron microscopy.ResultsI/R + Dex group showed significantly increased IL-6 and TNF-α levels and reduced myocardial cell necrosis and apoptosis. H&E staining showed alleviated myocardial disorder, myocardial cell swelling, myocardial fiber fracture, and inflammatory cell infiltration in I/R + Dex group. Myocardial cell necrosis and apoptosis were significantly reduced in I/R + Dex group. ATP level in myocardial tissue of mice in I/R group was substantially decreased, while that in Dex group was increased. WB results showed that MIF, P-AMPK α, GLUT4 and Bcl-2 levels were increased and Bax levels were decreased in I/R + Dex group.ConclusionDex may exert myocardial protection in young mice through MIF/AMPK/GLUT4 axis.

Experimental study of mitochondrion-targeted small molecule IR-61 ameliorated exhaustive exercise-induced cardiac injury in rats

To investigate the effects of mitochondrion-targeted cyanine fluorescent small molecule IR-61 on cardiac injury induced by exhaustive exercise in rats. Thirty-six adult male SD rats were randomly divided into 3 groups(n=12),control group (Ctrl), exhaustive exercise group (EE) and IR-61+ exhaustive exercise group (IR-61+EE). IR-61+EE group were intraperitoneally injected with 2 mg/kg IR-61 at the same time on day 1, 4 and 7. One hour after the end of the last drug administration, the two exhaustive exercise groups were subjected to exhaustive exercise modeling. The rats were placed on an animal treadmill with a slope of 0° at a speed of 10~15 m/min to coordinate their limbs running posture, and then ran at a speed of 25~30 m/min until exhaustion about 15 minutes later. After the animal models established, ECG was recorded by physiological recorder, myocardial injury was observed by light microscope, mitochondrial injury was observed by transmission electron microscope, myocardial cell apoptosis was detected by TUNEL method, markers of myocardial injury were detected by ELISA, and myocardial mitochondrial respiration rate was measured by high-resolution Oxygraph-2K mitochondrial instrument. ① Compared with Ctrl group, heart rate was increased, PR interval was shortened, QRS interval was prolonged, QTc was prolonged and ST segment was depressed significantly in EE group (P＜0.05). In EE group, myocardial fiber fracture and mitochondrial inner chamber swelling were obvious, mitochondrial crest was fuzzy, mitochondrial outer membrane was incomplete, and a large number of mitochondrial rupture and fusion were visible. In EE group, TUNEL staining cells were abundant, chromatin concentration and marginalization, nuclear membrane lysis, chromatin fragmentation into massive apoptotic bodies, apoptosis score increased (P＜0.05). The levels of creatine kinase isoenzyme-MB (CK-MB), cardiac troponin I(cTn-I) and N-terminal B-type natriuretic peptide (NT-proBNP) were increased in EE group (P＜0.05). Basal respiration rate, oxidative respiration rate of fatty acids and respiration rate of complex Ⅰ, Ⅱ and Ⅳ were all decreased (P＜ 0.05). ② Compared with EE group, the heart rate in IR-61+EE group was increased, PR interval was prolonged, QRS interval was shortened, QTc was shortened, ST segment was not significantly depressed (P＜0.05). In IR-61+EE group, myocardial fiber arrangement was loose, no obvious fracture was observed, mitochondrial inner ventricle was swelling, mitochondrial outer membrane was intact, TUNEL stained cells and unstained cells were observed, the overall morphology was more similar to Ctrl group. Apoptosis index was decreased (P＜0.05), the levels of CK-MB and cTn-I were decreased in IR-61+EE group (P＜0.05). The oxidative respiration rate of fatty acids and the respiration rate of complex Ⅱ and Ⅳ were increased (P＜0.05). Mitochondrion-targeted cyanine fluorescent small molecule IR-61 can improve cardiac electrical activity, reduce myocardial cell injury and mitochondrial injury, reduce myocardial cell apoptosis, and improve the myocardial mitochondrial energy metabolism condition in exhausted rats.

N-Acetylcysteine, an ROS Inhibitor, Alleviates the Pathophysiology of Hyperthyroidism-Induced Cardiomyopathy via the ROS/Ca2+ Pathway

Hyperthyroidism is common and can induce cardiomyopathy, but there is no effective therapeutic strategy. The purpose of this study was to investigate the molecular mechanism of hyperthyroidism-induced cardiomyopathy (HTC) and the effect of N-acetylcysteine (NAC), an ROS inhibitor, on the pathophysiology of HTC in vivo and in vitro. Compared with those in the control groups in vivo and in vitro, TT3 and TT4 were significantly increased, the structure of myocardial cells was enlarged and disordered, and interstitial fibrosis and the apoptosis of myocardial cells were markedly increased in the L-Thy group. The ROS and inflammatory response were increased in the hyperthyroidism group. In the NAC group, the contents of TT3 and TT4 were decreased, the myocardial cell structure was slightly disturbed, fibrosis and apoptosis were significantly reduced, and the ROS level and inflammatory response were significantly reduced. Interestingly, L-Thy decreased the viability of fibroblasts and H9c2 cells, suggesting that L-Thy-induced fibrosis was not caused by the proliferation of fibroblasts. The molecular mechanism of HTC could be explained by the fact that L-Thy could cause cardiac hypertrophy, inflammation, and fibrosis by regulating the Ca2+/calpain/Rcan1-dependent signalling pathway, the Ca2+/Rcan1/NF-κB/p65-dependent signalling pathway, and the Ca2+/ROS/Bcl-2/caspase-3-dependent signalling pathway. In conclusion, NAC can alleviate the pathophysiology of hyperthyroidism-induced cardiomyopathy, probably by regulating the ROS/Ca2+-dependent pathway.

Inhibition of TNF-α and JNK Signaling Pathway Can Reduce Paclitaxel-Induced Apoptosis of Mouse Cardiomyocytes.

Paclitaxel (PTX) is a widely used chemotherapeutic drug for treating tumors. However, studies have shown that it can cause cardiac problems such as arrhythmia, myocarditis, chronic cardiomyopathy, and heart failure. Therefore, it is essential to study the mechanism behind the cardiotoxicity of PTX in tumor treatment. In this study, we initially injected PTX into mice to establish a myocardial cell apoptosis model to observe the degree of damage to mouse myocardium caused by PTX. Upon determining the levels of mouse myocardial creatine phosphokinase (CK), myokinase isoenzyme (CK-MB), aspartate transaminase (AST), and lactate dehydrogenase (LDH), we found that all of these levels showed apparent increases in mice treated with PTX. Further analyses of the TNF-α level and the expression of Jun N-terminal kinase (JNK) and Bcl-2 family-related proteins in myocardial tissue were performed. It was found that PTX increased the protein levels of TNF-α, Bax, p-JNK, and JNK in myocardial tissue but decreased the protein level of Bcl-2. After 1 month of PTX treatment in mice, we inhibited the expression of TNF-α and JNK proteins, which reduced the effect of paclitaxel on the apoptosis of mouse cardiomyocytes. The protein levels of Bax, p-JNK, and TNF-α in cardiomyocytes were reduced, while there was a relative increase in the Bcl-2 protein level. The findings suggested that inhibition of the NK signaling pathway and TNF-α can lessen the effect of PTX on mouse cardiomyocytes.

HSP70 protects H9C2 cells from hypoxia and reoxygenation injury through STIM1/IP3R

Hypoxia/reoxygenation (H/R) is used as an in vivo model of ischemia/reperfusion injury, and myocardial ischemia can lead to heart disease. Calcium overload is an important factor in myocardial ischemia–reperfusion injury and can lead to apoptosis of myocardial cells. Therefore, it is of great clinical importance to find ways to regulate calcium overload and reduce apoptosis of myocardial cells, and thus alleviate myocardial ischemia–reperfusion injury. There is evidence that heat shock protein 70 (HSP70) has a protective effect on the myocardium, but the exact mechanism of this effect is not completely understood. Stromal interaction molecule 1 and inositol 1,4,5-triphosphate receptor (STIM/1IP3R) play an important role in myocardial ischemia–reperfusion injury. Therefore, this study aimed to investigate whether HSP70 plays an anti-apoptotic role in H9C2 cardiomyocytes by regulating the calcium overload pathway through STIM1/IP3R. Rat H9C2 cells were subjected to transient oxygen and glucose deprivation (incubated in glucose-free medium and hypoxia for 6 h) followed by re-exposure to glucose and reoxygenation (incubated in high glucose medium and reoxygenation for 4 h) to simulate myocardial ischemia reperfusion-induced cell injury. H9C2 cell viability was significantly decreased, and lactate dehydrogenase (LDH) release and apoptosis were significantly increased after oxygen and glucose deprivation. Transfection of HSP70 into H9C2 cells could reduce the corresponding effect, increase cell viability and anti-apoptotic signal pathway, and reduce the apoptotic rate and pro-apoptotic signal pathway. After hypoxia and reoxygenation, the expression of STIM1/IP3R and intracellular calcium concentration of HSP70-overexpressed H9C2 cells were significantly lower than those of hypoxia cells. Similarly, direct silencing of STIM1 by siRNA significantly increased cell viability and expression of anti-apoptotic protein Bcl-2 and decreased apoptosis rate and expression of pro-apoptotic protein BAX. These data are consistent with HSP70 overexpression. These results suggest that HSP70 abrogates intracellular calcium overload by inhibiting upregulation of STIM1/IP3R expression, thus reducing apoptosis in H9C2 cells and playing a protective role in cardiomyocytes.

MiR-199 Aggravates Doxorubicin-Induced Cardiotoxicity by Targeting TAF9b.

The clinical application of doxorubicin (DOX) is limited because of its cardiotoxicity. However, the pathogenic mechanism of DOX and the role of miRNA in DOX-induced cardiotoxicity remain to be further studied. This study aimed to investigate the role of miR-199 in DOX-mediated cardiotoxicity. A mouse model of myocardial cell injury induced by DOX was established. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the expression changes of miR-199 and TATA-binding protein associated factor 9B (TAF9b) in DOX-induced cardiac injury. Cell apoptosis was detected by TUNEL staining and flow cytometry. The expression levels of apoptosis-related proteins, namely, Bax and Bcl-2, were detected by qPCR. The expression of Beclin-1 and LC3b was detected by western blotting. The binding effect of miR-199 with TAF9b was verified by dual-luciferase reporter gene assay. In this study, overexpression of miR-199 could promote cardiotoxicity. Inhibition of miR-199 could alleviate DOX-mediated myocardial injury. Further studies showed that miR-199 targeted TAF9b. Moreover, miR-199 promoted apoptosis of myocardial cells and aggravated autophagy. Furthermore, we demonstrated that TAF9B knockdown reversed the myocardial protective effect of miR-199 inhibitors. Therefore, miR-199 promoted DOX-mediated cardiotoxicity by targeting TAF9b, thereby aggravating apoptosis and regulating autophagy.

Damage of uremic myocardium by p-cresyl sulfate and the ameliorative effect of Klotho by regulating SIRT6 ubiquitination

Uremic cardiomyopathy (UCM) is a common complication in patients with chronic kidney disease (CKD) and an important risk factor for death. P-Cresyl sulfate (PCS) is a damaging factor in UCM, and Klotho is a protective factor. However, the molecular mechanisms of Klotho and PCS in UCM and the relationship between PCS and Klotho are unclear. In vitro, Klotho treatment inhibited PCS-induced cardiomyocyte hypertrophy and apoptosis by blocking mTOR phosphorylation and inhibiting DNA double-strand breaks (DSBs), respectively. Moreover, PCS increased SIRT6 protein ubiquitination and downregulated SIRT6 protein expression, while Klotho inhibited SIRT6 protein ubiquitination and upregulated SIRT6 protein expression. In a mouse model of 5/6 nephrectomy (5/6Nx)-induced UCM, the expression of Klotho in the kidney and serum was decreased, and the expression of SIRT6 protein in myocardial tissues was lower. PCS further reduced Klotho and SIRT6 expression, aggravated heart structure and function abnormalities, and increased myocardial cell apoptosis in UCM mice. Administration of Klotho protein inhibited the downregulation of SIRT6 protein expression and improved cardiac structure and function. Furthermore, serum PCS level was associated with the left ventricular mass (LVM) and left ventricular mass index (LVMI) in hemodialysis patients. In conclusion, the uremic toxin PCS injures cardiomyocytes via mTOR phosphorylation and DSBs, and Klotho antagonizes the damaging effects of PCS. Moreover, the SIRT6 protein plays an important role in UCM, and Klotho suppresses SIRT6 ubiquitination induced by PCS, further improves cardiac structure and function in UCM and exerts protective effects.

Protective effect of ivabradine on mice with viral myocarditis and its mechanism

Viral myocarditis (VMC) is a type of cardiovascular disease caused by viral infection of myocardial cells characterized by myocardial interstitital inflammatory cell infiltration, myocardial fiber necrosis or figrinolysis. Ivabradine (IVA) is a commonly known drug used to control heart rate, resist inflammation and oxidative stress, particularly in VMC. Here, we have tried to evaluate the protective effects of IVA on mice with VMC and understand the possible mechanism behind this process. In order to complete the study, eighty male mice aged 6 weeks old were randomly divided into normal control, VMC model, low-dose IVA (L-IVA) and high-dose IVA (H-IVA) groups. Half an hour after modeling, IVA aqueous solution was administered intragastrically into L-IVA and H-IVA groups at 5 mg·kg-1·d-1 and 20 mg·kg-1·d-1, respectively for 14 consecutive days. Another 120 mice of the same batch were grouped and treated as described above. At 7 and 14 d, 6 mice in each group were sacrificed to obtain blood and heart samples. Body weight/heart weight (BW/HW) was calculated, hematoxylin-eosin staining was performed to observe the pathological changes of myocardium, and the level of cardiac troponin I (cTnI) was measured by ELISA. Related kits were employed to measure superoxide dismutase (SOD), malondialdehyde (MDA), glutathione peroxidase (GSH-Px) and catalase (CAT) activites in myocardial homogenate, and the levels of interleukin-6 (IL-6), IL-18, IL-1β and tumor necrosis factor-α (TNF-α) were determined by double-antibody sandwich ELISA. TUNEL assay was performed to detect the apoptosis of myocardial cells. The protein expressions of Bcl-2, Bax and Caspase-3 in myocardial cells were measured by Western blotting. Compared to the VMC model group, the heart/body weight ratio, myocardial pathological score, cTnI level, MDA activity, and levels of IL-6, IL-18, IL-1β and TNF-α were found decreased, while survival rate and activity of SOD, GSH-Px and CAT increased in L-IVA and H-IVA groups (P &lt;0.05). The apoptosis rate of myocardial cells declined in L-IVA and H-IVA groups (P &lt;0.05), especially in H-IVA group. IVA downregulated the protein expressions of Bax and Caspase-3 and upregulated that of Bcl-2 (P &lt;0.05). Thus, it has been found that IVA elevates the survival rate of VMC mice and relieves myocardial damage possibly by enhancing antioxidant capacity and modulating apoptosis-related proteins to suppress apoptosis.

Silencing lncRNA 93358 Inhibits the Apoptosis of Myocardial Cells in Myocardial Infarction Rats by Inducing the Expression of SLC8A1.

Objective To explore the inhibitor effects and mechanism of lncRNA 93358 against the apoptosis of myocardial cells in rats with myocardial infarction. Methods The myocardial infarction model was established in rats, which were identified by cardiac ultrasound. TTC staining was used to evaluate the degree of heart infarction, and HE staining was utilized to determine the pathological state in myocardial tissues. The apoptotic state in myocardial tissues was confirmed by TUNEL assay. lncRNA 93358 was screened out using a high-throughput sequencing which was confirmed by RT-qPCR. The interaction between miR-466c-3p and SLC8A1 was identified using the dual-luciferase reporter assay. The expression level of Bax, Bcl-2, and SLC8A1 was determined in lncRNA 93358 knockdown cells using RT-qPCR and Western blotting Results Massive myocardial necrosis was observed in model rats according to the results of TTC staining, HE staining, and TUNEL assay. lncRNA 93358 and Bax were found significantly upregulated, and Bcl-2 and SLC8A1 were greatly downregulated in model rats, which were dramatically reversed by the knockdown of lncRNA 93358, accompanied by the decline area of myocardial necrosis and decreased apoptotic myocardial cells. Conclusion Silencing lncRNA 93358 inhibits the apoptosis of myocardial cells in rats with myocardial infarction by inducing the expression of SLC8A1.

Ubiquitin-specific protease 7 regulates myocardial ischemia/reperfusion injury by stabilizing Keap1

Myocardial ischemia/reperfusion (I/R) injury is a complex pathological process that is still not fully understood. The oxidative stress response has a critical role in the occurrence and progression of myocardial ischemia/reperfusion injury. This study investigated the specific mechanism of ubiquitin-specific protease 7 (USP7) regulation of myocardial ischemia/reperfusion injury from the perspective of proteasome degradation and its relation with the Keap1 pathway, a vital regulator of cytoprotective responses to endogenous and exogenous stress induced by reactive oxygen species (ROS) and electrophiles. Our data indicated that USP7 expression is increased during myocardial ischemia/reperfusion injury in mice, while its inhibiting suppressed the generation of oxygen free radicals and myocardial cell apoptosis, reduced myocardial tissue damage, and improved heart function. Mechanistically, USP7 stabilizes Keap1 by regulating its ubiquitination. Taken together, these findings demonstrate the potential therapeutic effect of USP7 on myocardial ischemia/reperfusion injury.

Blocking the A2B adenosine receptor alleviates myocardial damage by inhibiting spleen-derived MDSC mobilisation after acute myocardial infarction

Background Myeloid-derived suppressor cell (MDSC) mobilisation is an important immune event in acute myocardial infarction (AMI). The A2B adenosine receptor (A2BAR) plays key role in regulating MDSC function, but its specific involvement in MDSC mobilisation in AMI remains unclear. Methods In AMI patients, the circulating MDSC ratio and A2BAR mRNA expression were measured. A mouse AMI model was established by left anterior descending coronary artery (LADCA) ligation. MDSCs were analysed by FACS and immunofluorescence staining (of heart tissue). A2BAR mRNA expression was assessed by qRT-PCR. Myocardial injury was detected by HE staining. Myocardial cell apoptosis was analysed by immunohistochemistry. Cardiac systolic function was evaluated by transthoracic echocardiography. Results In AMI patients, the circulating MDSC ratio was increased and positively correlated with A2BAR mRNA expression (r = 0.86, p < 0.01). In AMI model mice, the percentage of MDSCs was increased in the circulation and infarcted heart and decreased in the spleen. MRS-1754-mediated A2BAR inhibition decreased the MDSC ratio in the circulation and infarcted heart and prevented the decrease in MDSC number in the spleens of mice with AMI. A2BAR blockade inhibited myocardial cell apoptosis, alleviated myocardial inflammatory injury, and improved myocardial systolic function in the AMI mouse model. Similar results were found in mice after splenectomy. Additionally, spleen-derived MDSC injection increased the MDSC ratio in the infarcted heart, increased myocardial cell apoptosis, aggravated myocardial injury, and decreased cardiac systolic function in mice with AMI. Conclusion Blocking A2BAR alleviates myocardial damage and improves myocardial systolic function through inhibition of spleen-derived MDSC mobilisation after AMI. Key Messages Spleen-derived MDSC mobilisation aggravates myocardial inflammatory injury within 24 h of AMI. A2BAR promotes spleen-derived MDSC mobilisation within 24 h of AMI. Blocking A2BAR improves myocardial systolic function through inhibition of spleen-derived MDSC mobilisation.

Indole-3-Carbinol (I3C) Protects the Heart From Ischemia/Reperfusion Injury by Inhibiting Oxidative Stress, Inflammation, and Cellular Apoptosis in Mice.

Strategies for treating myocardial ischemia in the clinic usually include re-canalization of the coronary arteries to restore blood supply to the myocardium. However, myocardial reperfusion insult often leads to oxidative stress and inflammation, which in turn leads to apoptosis and necrosis of myocardial cells, for which there are no standard treatment methods. The aim of this study was to determine the pharmacological effect of indole-3-carbinol (I3C), a phytochemical found in most cruciferous vegetables, in a mouse model of myocardial ischemia/reperfusion injury (MIRI). Our results showed that I3C pretreatment (100 mg/kg, once daily, i. p.) prevented the MIRI-induced increase in infarct size and serum creatine kinase (CK) and lactate dehydrogenase (LDH) in mice. I3C pretreatment also suppressed cardiac apoptosis in MIRI mice by increasing the expression levels of the anti-apoptotic protein Bcl-2 and decreasing the expression levels of several apoptotic proteins, including Bax, caspase-3, and caspase-9. In addition, I3C pretreatment was found to reduce the levels of parameters reflecting oxidative stress, such as dihydroethidium (DHE), malondialdehyde (MDA), reactive oxygen species (ROS), and nitric oxide (NO), while increasing the levels of parameters reflecting anti-oxidation, such as total antioxidant capacity (T-AOC) and glutathione (GSH), in MIRI-induced ischemic heart tissue. I3C pretreatment was also able to remarkably decrease the expression of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6) mRNA in ischemic heart tissue. These results demonstrate that administration of I3C protects the heart from MIRI through its anti-apoptotic, antioxidant, and anti-inflammatory effects.

Stanniocalcin 1 alleviates myocardial ischemia-reperfusion injury through inhibiting inflammation and apoptosis of myocardial cells.

Myocardial ischemia-reperfusion injury (MIRI) is the main cause of death from ischemic heart diseases. Stanniocalcin 1 (STC1) has a potential therapeutic effect on MIRI. The purpose of this study is to investigate the effect of STC1 on inflammation and apoptosis of myocardium in MIRI. We used rats to make ischemia-reperfusion (I/R) models and determined the efficiency of modeling by 2, 3, 5-triphenyl tetrazolium chloride staining, echocardiography, and lactate dehydrogenase detection. We injected subcutaneously recombinant human STC1 (2.5 μg/kg, 5 μg/kg) into rats daily one week before modeling to detect the effect of STC1 pretreatment on inflammation and apoptosis of rat myocardial cells. In addition, we cultured rat myocardial cell lines (H9c2 cells) to investigate the effect of STC1 on myocardial cells. The cardiac function and structure of I/R rats were obviously destroyed. After treating rats with STC1, we found that the cardiac function and structure of the rats were significantly improved. In addition, STC1 reduced the expression of inflammatory factors and apoptosis levels in rat myocardium. Stimulation of STC1 also improved the viability of H9c2 cells in vitro. Therefore, STC1 can alleviate MIRI by inhibiting inflammation and apoptosis. It indicated that STC1 may have a potential therapeutic effect on MIRI.

Serum TP53 Protein Level as a Sensitive Biomarker for the Diagnosis of Myocardial Damage in Children.

BackgroundHigh levels of TP53 protein can lead to apoptosis of myocardial cells. However, TP53 protein influence of myocardial damage remains unclear. This prospective study investigated the involvement of TP53 protein in secondary myocardial damage in children up to 18 years of age.Material/MethodsSerum TP53 protein, N-terminal prohormone B-type natriuretic peptide (NT-ProBNP), cardiac troponin-I (cTnI), and creatine kinase isoenzyme MB (CK-MB) concentrations were measured in 50 hospitalized patients with secondary myocardial damage, 50 hospitalized patients without myocardial damage, and 50 healthy individuals (control). Cardiac damage was diagnosed based on cTnI, NT-ProBNP, and CK-MB levels, with electrocardiographic evidence as the reference. The appropriate cut-off value of TP53 protein for secondary myocardial damage was analyzed by receiver operating characteristic (ROC) curves.ResultsThe serum TP53 protein, NT-ProBNP, cTnI, and CK-MB concentrations of the patients with and without myocardial damage were 10.20±1.20 and 0.30±0.10 ng/L, 505.30 and 107.8 ng/L, 0.23±0.13 and 0.02±0.01 μg/L, and 28.30±5.13 and 12.24±4.29 IU/L, respectively. For the 50 patients with myocardial damage, the area under the ROC curve for serum TP53 protein, NT-ProBNP, cTnI, and CK-MB concentrations were 0.89 (95% CI: 0.81–0.95), 0.83 (95% CI: 0.77–0.91), 0.92 (95% CI: 0.84–0.97), and 0.85 (95% CI: 0.78–0.93), respectively, and the diagnostic cut-off values were 12.00 ng/L, 500.00 ng/L, 0.16 μg/L, and 27.00 IU/L, respectively, with positive likelihood ratios of 20.8, 13.2, 24.6, and 15.6.ConclusionsTP53 protein is a valid biomarker of secondary myocardial damage in pediatric patients and can be diagnostic.