Apoptosis In H9c2 Cells Research Articles

Pinoresinol diglucoside ameliorates H/R-induced injury of cardiomyocytes by regulating miR-142-3p and HIF1AN.

This study is aimed to investigate the effect of pinoresinol diglucoside (PDG) in ameliorating myocardial ischemia-reperfusion injury (MIRI).Hypoxia/reperfusion (H/R)-induced H9c2 cardiomyocytes were used to establish an in-vitro ischemia-reperfusion injury model of cardiomyocytes. Cells were treated with 1 μmol/L of PDG. Reactive oxygen species (ROS) level was detected by a 2',7'-dichlorofluorescein-diacetateassay. The release of lactate dehydrogenase (LDH) and creatine kinase-MB (CK-MB) was examined by enzyme-linked immunosorbent assay. The viability and apoptosis of H9c2 cells were probed by MTT assay and flow cytometry. Besides this, Western blot and quantitative real-time PCR were used to detect microRNA-142-3p (miR-142-3p) and hypoxia-inducible factor 1 subunit alpha inhibitor (HIF1AN) expression levels. The binding sequence between miR-142-3p and HIF1AN 3'-untranslated region was validated by a dual-luciferase reporter gene assay.PDG treatment significantly reduced the level of ROS, LDH, and CK-MB, promoted viability, and inhibited the apoptosis of H9c2 cells. PDG treatment promoted miR-142-3p expression and inhibited HIF1AN expression in H9c2 cells. MiR-142-3p overexpression enhanced the effects of PDG on ROS, LDH, CK-MB levels, cell viability, and apoptosis in H9c2 cardiomyocytes, while overexpression of HIF1AN reversed the above effects.PDG ameliorates H/R-induced injury of cardiomyocytes by regulating miR-142-3p and HIF1AN.

Alpha-Lipoic Acid Protects Against Doxorubicin-Induced Cardiotoxicity by Regulating Pyruvate Dehydrogenase Kinase 4.

As a widely used anti-tumor anthracycline, the accumulation of Doxorubicin (DOX) in body causes irreparable cardiomyocyte damage and therefore is limited in clinical application. Strategies to prevent from DOX-associated cardiotoxicity are urgent for patients who undergo DOX-based chemotherapy. Since oxidative stress injury being the major reason for myocardial toxicity of DOX, here we demonstrated that, Alpha-lipoic acid (ALA), which is a reductive agent, plays a cardioprotective role in attenuating DOX-induced cardiotoxicity by inhibiting pyruvate dehydrogenase kinase 4 (PDK4) expression. In vivo, the beneficial effect of ALA was evidenced by increased survival rate, mechanical contraction, and oxidative phosphorylation, while decreased reactive oxidative species (ROS) and apoptosis. In vitro, PDK4 overexpression remarkably increased DOX-induced apoptosis and ROS production in H9C2 cells. Notably, the protective effect of ALA was abrogated by PDK4 overexpression. We further used PDK4 knockout mice to identify the role of PDK4 in DOX-induced cardiotoxicity. Results elicited that PDK4 deficiency showed a consistent effect in protecting DOX cardiotoxicity as ALA treatment, which was evidenced by restored redox homeostasis and mitochondrial metabolism, finally inhibited myocardial injury. In conclusion, the cardioprotective role of ALA against DOX cardiotoxicity was dependent on PDK4-mediated regulation of oxidative stress and mitochondria metabolism.

TIPE2 protects cardiomyocytes from ischemia-reperfusion-induced apoptosis by decreasing cell autophagy via the mTORC1 signaling pathway.

In cardiac ischemia-reperfusion (I/R), autophagy of hyperactivated cardiomyocytes degrades normal proteins and organelles, destroys cells and causes irreversible cell death. The present study aimed to determine the molecular mechanism through which TNF-α-induced protein 8-like protein 2 (TIPE2) regulates cardiomyocyte apoptosis via autophagy in I/R. The results revealed that the number of apoptotic cells and the protein expression levels of TIPE2 in the heart tissue of I/R model mice were significantly increased. In vitro, the overexpression of TIPE2 decreased oxygen glucose deprivation (OGD)-induced autophagy, apoptosis and activation of the mTOR complex 1 (mTORC1) signaling pathway in H9c2 cells. Treatment with the mTORC1 inhibitor not only inhibited the TIPE2-activated mTORC1 signaling pathway, but also increased OGD-induced autophagy and apoptosis of H9c2 cells. In conclusion, the results of the present study revealed that TIPE2 may protect cardiomyocytes from I/R-induced apoptosis by decreasing cell autophagy via the mTORC1 signaling pathway.

Mechanism of atractylenolide Ⅲ in alleviating H9c2 cell apoptosis through ROS/GRP78/caspase-12 signaling pathway based on molecular docking

This study aims to investigate the effect of atractylenolide Ⅲ(ATL-Ⅲ) on hydrogen peroxide(H_2O_2)-induced endoplasmic reticulum stress and apoptosis of H9 c2 cells via the ROS/GRP78/caspase-12 signaling pathway.The binding activity of ATL-Ⅲ to GRP78 was determined by molecular docking.The result showed that ATL-Ⅲ had a good binding activity to GRP78, and the binding activity of ATL-Ⅲ was stronger than that of its specific inhibitor.The endoplasmic reticulum stress model of H9 c2 was established by H_2O_2(100 μmol·L~(-1)) treatment.Five groups were designed: blank control group, model group, and ATL-Ⅲ(15, 30, and 60 μmol·L~(-1)) groups.Apoptosis was detected by Hoechst/PI double staining and flow cytometry.The levels of superoxide dismutase(SOD), malondialdehyde(MDA), and lactate dehydrogenase(LDH) were measured by colorimetry.The levels of reactive oxygen species(ROS) and calcium(Ca~(2+)) in cytoplasm were determined by the fluorescence probe DCFH-DA and the calcium fluorescence probe Flou-4, respectively.The protein levels of GRP78, caspase-12, and caspase-3 were determined by Western blot, and the mRNA levels of GRP78 and caspase-12 by RT-qPCR.N-acetyl-L-cysteine(NAC) and 4-phenylbutyric acid(4-PBA) were respectively used to inhibit ROS and GRP78, and then the mechanism of ATL-Ⅲ in protecting the cells from endoplasmic reticulum stress induced by H_2O_2 were deduced.ATL-Ⅲ(15, 30, and 60 μmol·L~(-1)) decreased the apoptosis rate and ROS, MDA, and LDH levels(P<0.01), increased the SOD activity(P<0.01), and down-regulated the protein levels of GRP78, caspase-12, and caspase-3 and the mRNA levels of GRP78 and caspase-12(P<0.05).The addition of NAC decreased the apoptosis rate and ROS, MDA, GRP78, caspase-12, and caspase-3 levels(P<0.01), while it elevated the SOD level(P<0.01).The addition of 4-PBA also decreased the apoptosis rate and the levels of GRP78, caspase-12, caspase-3, and Ca~(2+)(P<0.01).The effect of inhibitors were consistent with that of ATL-Ⅲ.In conclusion, ATL-Ⅲ can protect H9 c2 cardiomyocytes by regulating ROS/GRP78/caspase-12 signaling pathway to inhibit H_2O_2-induced endoplasmic reticulum stress and apoptosis.

Ononin alleviates endoplasmic reticulum stress in doxorubicin-induced cardiotoxicity by activating SIRT3

Doxorubicin (DOX) is a potent anthracycline antineoplastic drug. However, its dose-dependent cardiotoxicity limits its clinical application. Ononin is a natural isoflavone glycoside that is crucial in modulating apoptosis-related signaling pathways. In this study, we assessed the possible cardioprotective effects of ononin in DOX-induced cardiotoxicity and elucidated the underlying molecular mechanisms. In vitro and in vivo assessments were performed using DOX-treated H9C2 cells and rats, respectively. First, DOX was injected into the tail veins of Wistar rats to induce cardiomyopathy. Next, rats in the DOX + Ononin30 and DOX + Ononin60 groups were intragastrically administered ononin two weeks before DOX treatment. H9C2 cells were treated with vehicle or DOX with or without ononin. Next, 3-TYP was used to determine the relationship between endoplasmic reticulum (ER) stress and sirtuin 3 (SIRT3) expression. Ononin treatment ameliorated DOX-induced myocardial injury as determined by echocardiography. Furthermore, ononin partially restored DOX-induced cardiac dysfunction; the left ventricular ejection fraction (LVEF) and left ventricular systolic fractional shortening (LVFS) increased after pre-treatment with ononin. Further, ononin suppressed DOX-induced ER stress and apoptosis in rat cardiomyocytes and H9C2 cells. The Bax/Bcl-2 ratio and 78-kD glucose-regulated protein (GRP78) and CCAAT enhancer-binding protein (CHOP) expression levels were higher in the DOX-treated group than in the control group but ononin treatment improved these parameters. These effects are associated with SIRT3 activity. Moreover, 3-TYP blocked the ononin-mediated protective effects. Hence, ononin positively affected DOX-induced cardiotoxicity by inhibiting ER stress and apoptosis, possibly mediated by stimulation of the SIRT3 pathway.

A novel identified circular RNA, circSnap47, promotes heart failure progression via regulation of miR-223-3p/MAPK axis.

The aim of this study was to investigate the effect of circSnap47 on heart failure (HF) and its potential mechanisms. Quantitative real-time PCR (qRT-PCR) was performed to detect the mRNA expression levels of circSnap47 and miR-233-3p. The viability and apoptosis of H9C2 cells were assessed using CCK-8 and TUNEL assays. The expressions of interleukin (IL)-6, IL-1β, IL-18, and tumor necrosis factor-alpha were determined using ELISA and qRT-PCR. In addition, the expression of apoptosis-related proteins and mitogen-activated protein kinase (MAPK) signaling pathway-related proteins was analyzed using western blot. Moreover, HF-related circRNAs and miRNAs were predicted via bioinformatics analysis. The relationship between circSnap47 and miR-233-3p was further confirmed using a dual-luciferase reporter gene assay. In HF tissues and H9C2 cells treated with oxygen-glucose deprivation (OGD), circSnap47 was upregulated. Silencing circSnap47 increased cell viability and inhibited apoptosis. Besides, silencing circSnap47 alleviated OGD-induced inflammation in H9C2 cells. Moreover, we found that miR-233-3p was the downstream target gene of circSnap47. Our results also revealed that silencing circSnap47 relieved OGD-induced H9C2 cell damage by inactivating the miR-223-3p/MAPK axis. We confirmed that circSnap47 silencing inhibited HF progression via regulation of miR-223/MAPK axis, which will provide for a new therapeutic direction for the treatment of HF.

IFN-γ enhances the efficacy of mesenchymal stromal cell-derived exosomes via miR-21 in myocardial infarction rats

BackgroundMesenchymal stromal cells (MSCs) activated with IFN-γ elicit stronger physical effects. Exosomes (Exos) secreted from MSCs show protective effects against myocardial injury. This study aimed to determine whether Exos derived from IFN-γ-treated MSCs exhibit more potent cardioprotective function and the underlying mechanisms.MethodsH9c2 cells or human umbilical vein endothelial cells (HUVECs) were treated with Exos isolated from MSCs (Ctrl-Exo) or IFN-γ-primed MSCs (IFN-γ-Exo) under oxygen and glucose deprivation (OGD) conditions in vitro and in vivo in an infarcted rat heart. RNA sequencing was used to identify differentially expressed functional transcription factors (TFs). Quantitative reverse transcription-PCR (qPCR) was used to confirm the upregulated TFs and miRNA in IFN-γ-primed MSCs. Dual-luciferase reporter gene assay was used to analyze the transcriptional regulation of miRNAs by STAT1. The target of miR-21-5p (miR-21) was determined by luciferase reporter assays and qPCR. The function of BTG2 was verified in vitro under OGD conditions.ResultIFN-γ-Exo accelerated migration and tube-like structure formation and prevented OGD-induced apoptosis in H9c2. Similarly, IFN-γ-Exo treatment caused a decrease in fibrosis, reduced cardiomyocyte apoptosis, and improved cardiac function compared to Ctrl-Exo treatment. MiR-21 was significantly upregulated in IFN-γ-primed MSCs and IFN-γ-Exo. STAT1 transcriptionally induced miR-21 expression. Up-regulated miR-21 could inhibit BTG anti-proliferation factor 2 (BTG2) expressions. BTG2 promoted H9c2 cell apoptosis and reversed the protective effects of miR-21 under OGD conditions.ConclusionIFN-γ-Exo showed enhanced therapeutic efficacy against acute MI, possibly by promoting angiogenesis and reducing apoptosis by upregulating miR-21, which directly targeted BTG2.

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.

Activation of the Nrf2/GPX4 Signaling by Pratensein From Trifolium pretense Mitigates Ferroptosis in OGD/R-Insulted H9c2 Cardiomyocytes

Background: Pratensein (PTS) is a type of flavonoid that has been identified in various plants, such as Trifolium pretense L., with a considerable cytoprotective effect against exogenous stimuli. However, the biological function of PTS in cardiomyocytes in response to ischemia-reperfusion (I/R) conditions is unclear. Purpose: In our study, we examined the function of PTS in the progression of myocardial infarction (MI). Methods: In this study, we established an oxygen-glucose deprivation/reoxygenation (OGD/R) model in H9c2 cells. The Cell Counting Kit-8 assay was used to assess the viability of H9c2 cells. The TdT-mediated dUTP-biotin nick end labeling and flow cytometry assays confirmed apoptosis of H9c2 cells. Reactive oxygen species (ROS), malondialdehyde (MDA), glutathione (GSH) content, and Fe2+ level were evaluated. Western blotting was used to detect relative protein expression. Results: We firstly found that PTS reduced apoptosis of H9c2 cells in response to OGD/R stimulation. PTS attenuates the increase in ROS and MDA production and the decrease in GSH content caused by OGD/R. The increased Fe2+ level in OGD/R-treated H9c2 cells was also restrained by PTS. For mechanism studies, we found that the decreased expression levels of Nrf2 and GPX4 in OGD/R-treated H9c2 cells were significantly elevated after PTS treatment. Knockdown of Nrf2 in H9c2 cells reversed the protective effect of PTS on ferroptosis in H9c2 cells induced by OGD/R, indicated by reduced cell viability, increased apoptotic cells and oxidation markers, and increased Fe2+ level. Conclusion: Based on these findings, we speculated that PTS may protect H9c2 cells from OGD/R-caused ferroptosis by modulating the Nrf2/GPX4 signaling.

ASTRAGALOSIDE IV INHIBITS H2O2-INDUCED APOPTOSIS IN H9C2 CELLS BY ATTENUATING OXIDATIVE STRESS AND MITOCHONDRIAL DAMAGE

Objective: To investigate the effects and mechanisms of As-IV on H2O2-induced apoptosis in H9c2 cells. Design and method: H9c2 cells were divided into three groups, including normal control group (cultured without intervention), H2O2 group (cultured with 200umol/L H2O2 for 2 hours), As-IV group (cultured with 100umol/L As-IV for 1 hour), As-IV with H2O2 group (pretreated with 100umol/L As-IV for 1 hour, then incubated with H2O2 for 2 hours). FITC/PI and DCFH-DA are stained to examine levels of apoptosis and ROS receptively. Levels of SOD and MDA were measured. Stained with JC-1 probe to observe the change of mitochondrial membrane potential (MMP). Western-blot was detected to analyze the expression of Drp-1. Results: 1. The apoptosis rate of H2O2 group ((13.22 ± 3.17 vs 40.55 ± 13.74)%, P < 0.05) was significantly higher than that of control group, which was lower in As-IV with H2O2 group((23.43 ± 4.3 vs 40.55 ± 13.74) %, P < 0.05). 2. The ROS levels were higher in H2O2 group than control group((73.53 ± 5.33 vs 39.02 ± 7.90) %, P < 0.05), which were significant reduced in As-IV with H2O2 group((57.6 ± 4.63 vs 73.53 ± 5.33) %, P < 0.05) Meanwhile, the activity of MDA was increased and the activity of SOD was decreased in H2O2 group compared with that in control group((10.97 ± 0.56 vs 4.85 ± 0.55) nmol/mL, P < 0.05), ((1192.23 ± 341.59 vs 2710.95 ± 274.32)U/L, P < 0.05). Compared with H2O2 group, As-IV with H2O2 group significantly decrease the activity of MDA and increase the activity of SOD ((6.58 ± 0.79 vs 10.97 ± 0.56) nmol/mL, P < 0.05), ((1984.66 ± 188.81 vs 1192.23 ± 341.59) U/L, P < 0.05). 3.MMP was lower and the expression of Drp1 was higher in H2O2 group than that in control group ((0.31 ± 0.01 vs 1.52 ± 0.25) %, P < 0.05), ((0.86 ± 0.003 vs 0.45 ± 0.004) %, P < 0.05). Compared with H2O2 group, MMP was significantly increased and the expression of Drp1 was downregulated ((0.65 ± 0.38 vs 0.31 ± 0.01) %, P < 0.05), ((0.72 ± 0.005 vs 0.86 ± 0.003) %, P < 0.05). Conclusions: As-IV exerts protective effects on H2O2-induced apoptosis by attenuating oxidative stress and mitochondrial damage.

Exosomal circ-CACNG2 promotes cardiomyocyte apoptosis in multiple myeloma via modulating miR-197-3p/caspase3 axis

Multiple myeloma (MM) secreted exosomes are essential in MM-related complications such as osteolytic bone lesions and renal failure, but their role and underlying mechanism in cardiac complications has not yet been clarified. Here, we investigated the effects of U266 (a MM cell line) exosomes (U266-exo) on regulating the viability, cell cycle, oxidative stress and apoptosis of H9C2 cells and the role of circ-CACNG2 in these effects. We found that U266-exo coculture significantly inhibited viability and promoted apoptosis of H9C2 cells, and serum exosomes of MM patients harbored high level of circ-CACNG2. The clinical data analyses indicated that circ-CACNG2 was an independent prognostic and diagnostic indicator of MM-related cardiac complications. Also, in vitro experiments showed that circ-CACNG2 inhibited viability and promoted apoptosis of H9C2 cells. RIPA, pull-down assays, dual-luciferase reporter assays, and RNA FISH assays revealed that miR-197-3p could bind to circ-CACNG2 and caspase3 directly. Rescue experiments proved that circ-CACNG2 can increase the expression of caspase3 by binding to and decreasing the expression of miR-197-3p. In conclusion, MM-exosomes could inhibit cardiomyocyte viability and promote apoptosis partially through circ-CACNG2/miR-197-3p/caspase3 axis.

F-box and WD repeat-containing protein 7 ameliorates angiotensin II-induced myocardial hypertrophic injury via the mTOR-mediated autophagy pathway.

Myocardial hypertrophy is a common heart disease that is closely associated with heart failure. The expression of F-box and WD repeat-containing protein 7 (FBW7) is significantly downregulated in angiotensin (Ang) II-induced cardiac fibroblasts, suggesting that it may possess an important function in cardiac development. The present study attempted to explore the role of FBW7 in Ang II-induced myocardial hypertrophic injury and its associated mechanism of action. Reverse transcription-quantitative PCR and western blotting were used to determine the expression levels of FBW7 in Ang II-induced H9C2 cells. The expression levels of autophagy-related and mTOR signaling pathway-related proteins were detected using western blotting. Cell viability was assessed using the Cell Counting Kit-8 assay. The apoptosis rate of H9C2 cells was detected using TUNEL assay and western blotting. Cellular hypertrophy and fibrosis were assessed using phalloidin staining and western blotting. Levels of inflammatory factors were examined using ELISA and western blotting, whereas levels of oxidative stress-related markers were detected by corresponding kits. The results indicated that FBW7 expression was downregulated in Ang II-induced H9C2 cells. FBW7 upregulation enhanced the expression levels of autophagy-related proteins and activated mTOR-mediated cellular autophagy. FBW7 overexpression promoted the cell viability, inhibited Ang II-induced apoptosis, cellular hypertrophy and fibrosis in H9C2 cells via the autophagic pathway, as well as inflammation and oxidative stress. Overall, the data indicated that FBW7 overexpression ameliorated Ang II-induced hypertrophic myocardial injury via the mTOR-mediated autophagic pathway.

Design, synthesis, and in vitro protective effect evaluation of α-carboline derivatives against H2O2-induced cardiomyocyte injury

As one of the most important features of myocardial ischemia reperfusion (MI/R) injury, the overproduction of reactive oxygen species (ROS) overwhelms the intrinsic antioxidant and impairs the function of mitochondria and, finally, leads to cardiomyocyte death. To improve the damage of cardiomyocyte caused by oxidative stress, a series of α-carboline derivatives were designed and synthesized in this study. The biological studies revealed that most of the α-carbolines exhibited obvious protective activities against H2O2-induced cardiomyocyte injury. Among them, compound 14b significantly increased the cell viability in H2O2-induced oxidative stress in H9c2 cardiomyoblasts with a concentration-dependent manner, which was more potent than polydatin. Pretreatment of 14b obviously inhibited H2O2-induced lactate dehydrogenase (LDH) leakage, enhanced the capacity of endogenous antioxidant defenses, including superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px), and reduced the formation of the toxic product of lipid peroxidation (malondialdehyde, MDA). In addition, 14b effectively reduced the overproduction of ROS and restored the mitochondrial membrane potential ΔΨm, better than that of polydatin. Flow cytometry analysis demonstrated that 14b markedly reduced both necrosis and apoptosis in H9c2 cells after the exposure to H2O2. Further Western blot analysis revealed that 14b obviously decreased the ratio of Bax/Bcl-2 and reduced the expression of cytochrome c. Overall, these results revealed the potential of α-carboline 14b as a promising cardioprotective agent against H2O2-induced oxidative injury.

A possible new activator of PI3K-Huayu Qutan Recipe alleviates mitochondrial apoptosis in obesity rats with acute myocardial infarction.

Obesity, which has unknown pathogenesis, can increase the frequency and seriousness of acute myocardial infarction (AMI). This study evaluated effect of Huayu Qutan Recipe (HQR) pretreatment on myocardial apoptosis induced by AMI by regulating mitochondrial function via PI3K/Akt/Bad pathway in rats with obesity. For in vivo experiments, 60 male rats were randomly divided into 6 groups: sham group, AMI group, AMI (obese) group, 4.5, 9.0 and 18.0 g/kg/d HQR groups. The models fed on HQR with different concentrations for 2 weeks before AMI. For in vitro experiments, the cardiomyocytes line (H9c2) was used. Cells were pretreated with palmitic acid (PA) for 24 h, then to build hypoxia model followed by HQR‐containing serum for 24 h. Related indicators were also detected. In vivo, HQR can lessen pathohistological damage and apoptosis after AMI. In addition, HQR improves blood fat levels, cardiac function, inflammatory factor, the balance of oxidation and antioxidation, as well as lessen infarction in rats with obesity after AMI. Meanwhile, HQR can diminish myocardial cell death by improving mitochondrial function via PI3K/Akt/Bad pathway activation. In vitro, HQR inhibited H9c2 cells apoptosis, improved mitochondrial function and activated the PI3K/Akt/Bad pathway, but effects can be peripeteiad by LY294002. Myocardial mitochondrial dysfunction occurs following AMI and can lead to myocardial apoptosis, which can be aggravated by obesity. HQR can relieve myocardial apoptosis by improving mitochondrial function via the PI3K/Akt/Bad pathway in rats with obesity.

The role of empagliflozin in palmitate-induced ER stress and apoptosis in H9C2 cells

The role of empagliflozin in palmitate-induced ER stress and apoptosis in H9C2 cells

Endogenous Hydrogen Sulfide Persulfidates Caspase-3 at Cysteine 163 to Inhibit Doxorubicin-Induced Cardiomyocyte Apoptosis

Doxorubicin (DOX) is an efficient antitumor anthracycline drug, but its cardiotoxicity adversely affects the prognosis of the patients. In this study, we explored whether endogenous gasotransmitter hydrogen sulfide (H2S) could protect against DOX-induced cardiomyocyte apoptosis and its mechanisms. The results indicated that DOX significantly downregulated endogenous H2S production and endogenous synthetase cystathionine γ-lyase (CSE) expression and obviously stimulated the apoptosis in H9C2 cells. The supplement of H2S donor sodium hydrosulfide (NaHS) or overexpression of CSE inhibited DOX-induced H9C2 cell apoptosis. DOX enhanced the activities of caspase family members in cardiomyocytes, while NaHS attenuated DOX-enhanced caspase-3, caspase-2, and caspase-9 activities by 223.1%, 73.94%, and 52.29%, respectively. Therefore, taking caspase-3 as a main target, we demonstrated that NaHS or CSE overexpression alleviated the cleavage of caspase-3, suppressed caspase-3 activity, and inhibited the cleavage of poly ADP-ribose polymerase (PARP). Mechanistically, we found that H2S persulfidated caspase-3 in H9C2 cells and human recombinant caspase-3 protein, while the thiol-reducing agent dithiothreitol (DTT) abolished H2S-induced persulfidation of caspase-3 and thereby prevented the antiapoptotic effect of H2S on caspase-3 in H9C2 cells. The mutation of caspase-3 C148S and C170S failed to block caspase-3 persulfidation by H2S in H9C2 cells. However, caspase-3 C163S mutation successfully abolished the effect of H2S on caspase-3 persulfidation and the corresponding protection of H9C2 cells. Collectively, these findings indicate that endogenous H2S persulfidates caspase-3 at cysteine 163, inhibiting its activity and cardiomyocyte apoptosis. Sufficient endogenous H2S might be necessary for the protection against myocardial cell apoptosis induced by DOX. The results of the study might open new avenues with respect to the therapy of DOX-stimulated cardiomyopathy.

Role of sevoflurane in myocardial ischemia-reperfusion injury via the ubiquitin-specific protease 22/lysine-specific demethylase 3A axis

ABSTRACT Myocardial ischemia-reperfusion injury (MIRI) represents a coronary artery disease, accompanied by high morbidity and mortality. Sevoflurane post-conditioning (SPC) is importantly reported in myocardial disease. Accordingly, the current study sought to evaluate the role of Sevo in MI/RI. Firstly, MI/RI models were established and subjected to SPC. Subsequently, pathological injury in the myocardium, myocardial infarction areas, H9c2 cell viability, apoptosis, and levels of creatine kinase-MB (CK-MB), cardiac troponin I (cTnI), and lactate dehydrogenase (LDH) were all measured. Ubiquitin-specific peptidase (22USP22), lysine-specific demethylase 3A (KDM3A), and Yes1 associated transcriptional regulator (YAP1) were down-regulated in H9c2 cells using cell transfection to verify their roles. The interaction between USP22 and KDM3A and between KDM3A and YAP1 was further validated. USP 22, KDM3A, and YAP1 were found to be down-regulated in MI/RI and SPC protected MI/RI rats, as evidenced by up-regulated expressions of USP22, KDM3A, and YAP1, reduced pathological injury in the myocardium, myocardial infarction areas, apoptosis, and levels of CK-MB, cTnI, and LDH, and enhanced H9c2 cell viability; while the protective effects of Sevo were counteracted by silencing of USP22, KDM3A, and SPC upregulated USP22, which stabilized KDM3A protein levels via deubiquitination, and KDM3A inhibited histone 3 lysine 9 di-methylation (H3K9me2) levels in the YAP1 promoter to encourage YAP1 transcription, to reduce MI/RI.

MiR-223-3p affects myocardial inflammation and apoptosis following myocardial infarction via targeting FBXW7.

BackgroundMyocardial infarction (MI) is one of the main causes of disability and death in the world, leading to myocarditis and cardiomyocyte apoptosis. Studies have shown that microRNA (miRNA) is involved in myocarditis and apoptosis. The main purpose of this study was to explore the regulatory mechanism of miR-223-3p on myocarditis and apoptosis after MI.MethodsWe cultured H9c2 cells and detected the expression of miR-223-3p in cells treated with different concentrations of H2O2. Sprague Dawley (SD) rats were fed with normal diet, constructed an MI model and detect the expression of miR-223-3p in heart tissue. Overexpression or inhibition of miR-223-3p was conducted in MI model cells in vitro, and the contents of the inflammatory factors tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interleukin-10 (IL-10) in cell supernatant were detected by enzyme-linked immunosorbent assay (ELISA). Cell apoptosis was detected by flow cytometry. Luciferase activity assay was used to detect the targeting relationship between miR-223-3p and FBXW7, and the expression of FBXW7 in cells was also detected. Overexpression of miR-223-3p was performed in MI rats to detect the expression of inflammatory factors, FBXW7, and apoptosis in rat cardiac tissue.ResultsThe expression of miR-223-3p was down regulated in MI models established in vitro and in vivo. Overexpression of miR-223-3p can inhibit inflammatory response and apoptosis in H9c2 cells and cardiac tissues. It was revealed that miR-223-3p can inhibit the expression of FBXW7, reduce myocarditis and apoptosis after MI, and improve cardiac function.ConclusionsIt is possible that miR-223-3p reduces myocarditis and apoptosis after MI and improves cardiac function by targeted inhibition of FBXW7 expression.

Propofol protects cardiomyocytes from doxorubicin-induced toxic injury by activating the nuclear factor erythroid 2-related factor 2/glutathione peroxidase 4 signaling pathways

ABSTRACT Propofol offers important protective effects in ischemia/reperfusion-induced cardiomyocyte injury, but its specific mechanisms in doxorubicin (DOX)-induced cardiotoxicity have not been investigated. In this paper, we attempted to explore the effects of propofol on DOX-induced human cardiomyocyte injury and its related mechanisms. H9c2 cell viability was assessed by cell counting kit-8 and lactate dehydrogenase assay kit. Nuclear factor erythroid 2-related factor 2 (NRF2)/glutathione peroxidase 4 (GPx4) signaling pathway-related protein levels were measured by Western blot. Ferroptosis was evaluated by corresponding kits and Western blot and apoptosis was detected by CCK-8, terminal deoxynucleotidyl transferase dUTP nick-end labeling and Western blot. Oxidative stress was assessed by reactive oxygen species kit and the commercial kits, and inflammation response was analyzed by enzyme-linked immunosorbent assay and Western blot. The results showed that propofol attenuated DOX-induced cytotoxicity and activated Nrf2/GPx4 signaling pathways in H9c2 cells. In addition, propofol also alleviated DOX-induced ferroptosis, increased cell viability and inhibited apoptosis, oxidative stress and inflammatory responses in H9c2 cells through activation of Nrf2/GPx4 signaling pathways. In summary, propofol provides the protection against DOX-induced cardiomyocyte injury by activating Nrf2/GPx4 signaling, providing a new approach and theoretical basis for the repair of cardiomyocytes.

Dexmedetomidine promotes cell proliferation and inhibits cell apoptosis by regulating LINC00982 and activating the phosphoinositide-3-kinase (PI3K)/protein kinase B (AKT) signaling in hypoxia/reoxygenation-induced H9c2 cells

ABSTRACT Previous studies showed dexmedetomidine (DEX) could alleviate myocardial ischemia/reperfusion injury (MIRI). Nevertheless, the mechanisms by which DEX alleviated MIRI remain to be determined. Our results demonstrated that DEX reversed hypoxia/reoxygenation (H/R)-induced decreased proliferation, and enhanced LINC00982 level, apoptosis, and inflammation in H9c2 cells. Moreover, LINC00982 overexpression attenuated the DEX-mediated protective effect of H9c2 cells under H/R. In addition, DEX upregulated p-phosphoinositide-3-kinase (p-PI3K) and p-protein kinase B (p-AKT) levels, and the silencing of LINC00982 further enhanced this effect in H/R-induced H9c2 cells. Furthermore, LINC00982 deletion enhanced the protective effect of DEX on H9c2 cells under H/R condition, while PI3K inhibitor, LY294002, obviously reversed this phenomenon. In sum, our work determined that DEX could suppress cell apoptosis and inflammation in H/R-triggered H9c2 through downregulating LINC00982 and activating PI3K/AKT signaling.