Cardiomyocyte Pyroptosis Research Articles

Inhibition of circDGKZ ameliorates myocardial ischemia/reperfusion injury by targeting miR-345-5p/TLR4.

This study aims to explore the molecular mechanism of circular RNAs' (circRNAs) potential involvement in myocardial ischaemia-reperfusion injury (MIRI). Differently expressed genes in myocardial infarction (MI) were identified by screening the GEO database. Serum was collected from MI patients and healthy volunteers (n=5 for each group). AC16 cells were cultured and exposed to hypoxia/reperfusion (H/R) treatment for the cell experiments. Then candidate genes were validated in human serum and the H/R model. Quantitative real-time PCR and western blot were used to detect expression of key molecules such as circDGKZ, miR-345-5p, and Toll-like receptor 4 (TLR4), as well as pyroptosis markers such as NOD-like receptor thermal protein domain-associated protein 3 (NLRP3), ASC, C-caspase1, interleukin (IL)-1β, and IL-18. CircDGKZ was positively correlated in human serum (P<0.05) and in AC16 cells (P<0.01). Knockdown of circDGKZ inhibited cardiomyocyte pyroptosis and the TLR4/nuclear factor kappa B (NF-κB) signalling pathway (all P<0.05). A luciferase assay was used to detect the molecule interaction. MiR-345-5p was regulated by circDGKZ and regulated TLR4 in cardiomyocytes both through direct interaction (P<0.01). The stability and distribution of circRNA or linear RNA were examined by subcellular localization and RNA decay assays. CircDGKZ was stably expressed in cardiomyocytes and mainly distributed in the cytoplasm (P<0.01). Knockdown of circDGKZ also promoted the degradation of NLRP3 by inducing autophagy (P<0.05). MIRI rat models were constructed (n=5 for each group), and the cellular results were further confirmed in rat models (P<0.05). Knockdown of circDGKZ interrupted pyroptosis and induced autophagy of cardiomyocytes via regulating miR-345-5p/TLR4/NF-κB.

Growth differentiation factor 11 regulates high glucose-induced cardiomyocyte pyroptosis and diabetic cardiomyopathy by inhibiting inflammasome activation

BackgroundDiabetic cardiomyopathy (DCM) is a crucial complication of long-term chronic diabetes that can lead to myocardial hypertrophy, myocardial fibrosis, and heart failure. There is increasing evidence that DCM is associated with pyroptosis, a form of inflammation-related programmed cell death. Growth differentiation factor 11 (GDF11) is a member of the transforming growth factor β superfamily, which regulates oxidative stress, inflammation, and cell survival to mitigate myocardial hypertrophy, myocardial infarction, and vascular injury. However, the role of GDF11 in regulating pyroptosis in DCM remains to be elucidated. This research aims to investigate the role of GDF11 in regulating pyroptosis in DCM and the related mechanism.Methods and resultsMice were injected with streptozotocin (STZ) to induce a diabetes model. H9c2 cardiomyocytes were cultured in high glucose (50 mM) to establish an in vitro model of diabetes. C57BL/6J mice were preinjected with adeno-associated virus 9 (AAV9) intravenously via the tail vein to specifically overexpress myocardial GDF11. GDF11 attenuated pyroptosis in H9c2 cardiomyocytes after high-glucose treatment. In diabetic mice, GDF11 alleviated cardiomyocyte pyroptosis, reduced myocardial fibrosis, and improved cardiac function. Mechanistically, GDF11 inhibited pyroptosis by preventing inflammasome activation. GDF11 achieved this by specifically binding to apoptosis-associated speck-like protein containing a CARD (ASC) and preventing the assembly and activation of the inflammasome. Additionally, the expression of GDF11 during pyroptosis was regulated by peroxisome proliferator-activated receptor α (PPARα).ConclusionThese findings demonstrate that GDF11 can treat diabetic cardiomyopathy by alleviating pyroptosis and reveal the role of the PPARα-GDF11-ASC pathway in DCM, providing ideas for new strategies for cardioprotection.

Malus toringoides (Rehd.) Hughes decoction alleviates isoproterenol-induced cardiac fibrosis by inhibiting cardiomyocyte inflammation and pyroptosis via the HK1/NLRP3 signaling pathway.

Malus toringoides (Rehd.) Hughes, called "Eseye (Ese)," is a traditional medicinal plant from the Tibet province of China that has proven effective in treating cardiac conditions due to its anti-inflammatory, antioxidative, and antiapoptotic properties. In this study, we explored the underlying protective mechanisms of Ese decoction in isoproterenol (ISO)-induced cardiac fibrosis (CF) and established the fact that treatment with an Ese decoction attenuated tissue injury, decreased the release of IL-1β, IL-18, TNF-α, and caspase-3, and elevated the Bax/Bcl-2 ratio in CF mice. We also found that with Ese treatment damage to the mitochondrial ultrastructure of myocardium was alleviated, and the level of reactive oxygen species was markedly diminished. Ese inhibited the expression of proteins associated with pyroptosis by the HK1/NLRP3 signaling pathway and also improved CF. Due to the anti-inflammatory, antioxidative, and antiapoptotic characteristics of Ese decoction, we found that Ese protected against ISO-induced CF, by inhibiting inflammation and pyroptosis as mediated by the HK1/NLRP3 signaling pathway.

N6-Methyladenosine Demethylase ALKBH5 Promotes Pyroptosis by Modulating PTBP1 mRNA Stability in LPS-Induced Myocardial Dysfunction.

This study aims to investigate the mechanism by which alkB homolog 5 (ALKBH5) regulates polypyrimidine tract-binding protein 1 (PTBP1) to mediate cardiomyocyte pyroptosis in sepsis-induced myocardial injury. Lipopolysaccharide (LPS)-exposed H9C2 cell and rat models were established to mimic septic myocardial injury both in vitro and in vivo. The mRNA and protein levels of ALKBH5 and PTBP1 in the LPS-induced cell and septic rat models were detected. CCK-8 and flow cytometry were applied to detect cell viability and pyroptosis. H&E staining was used to observe myocardial tissue damage in rats, and immunohistochemistry to analyze the expression of pyroptosis and inflammation-related proteins in rat tissues. Elevated expressions of both ALKBH5 and PTBP1 were found in the myocardial tissues of LPS-induced septic rats. ALKBH5 knockdown could restore the cell viability and cell pyroptosis inhibited by LPS, while ALKBH5 promoted PTBP1 mRNA stability by affecting its N6-methyladenosine (m6A) modification. In vivo experiments showed that PTBP1 knockdown could largely reverse the antiproliferative and pro-pyroptosis effects of ALKBH5 in LPS-exposed H9C2 cells. ALKBH5 knockdown in in vivo experiments was found to suppress the expressions of pyroptosis biomarkers and attenuate myocardial injury in septic rats. ALKBH5 promoted mRNA stability and the expression of PTBP1 through m6A modification to induce pyroptosis in cardiomyocytes and ultimately aggravate sepsis-induced myocardial dysfunction.

Peptides Are Cardioprotective Drugs of the Future: The Receptor and Signaling Mechanisms of the Cardioprotective Effect of Glucagon-like Peptide-1 Receptor Agonists.

The high mortality rate among patients with acute myocardial infarction (AMI) is one of the main problems of modern cardiology. It is quite obvious that there is an urgent need to create more effective drugs for the treatment of AMI than those currently used in the clinic. Such drugs could be enzyme-resistant peptide analogs of glucagon-like peptide-1 (GLP-1). GLP-1 receptor (GLP1R) agonists can prevent ischemia/reperfusion (I/R) cardiac injury. In addition, chronic administration of GLP1R agonists can alleviate the development of adverse cardiac remodeling in myocardial infarction, hypertension, and diabetes mellitus. GLP1R agonists can protect the heart against oxidative stress and reduce proinflammatory cytokine (IL-1β, TNF-α, IL-6, and MCP-1) expression in the myocardium. GLP1R stimulation inhibits apoptosis, necroptosis, pyroptosis, and ferroptosis of cardiomyocytes. The activation of the GLP1R augments autophagy and mitophagy in the myocardium. GLP1R agonists downregulate reactive species generation through the activation of Epac and the GLP1R/PI3K/Akt/survivin pathway. The GLP1R, kinases (PKCε, PKA, Akt, AMPK, PI3K, ERK1/2, mTOR, GSK-3β, PKG, MEK1/2, and MKK3), enzymes (HO-1 and eNOS), transcription factors (STAT3, CREB, Nrf2, and FoxO3), KATP channel opening, and MPT pore closing are involved in the cardioprotective effect of GLP1R agonists.

Protective role of hydrogen sulfide against diabetic cardiomyopathy by inhibiting pyroptosis and myocardial fibrosis

Diabetic cardiomyopathy (DCM) contributes significantly to the heightened mortality rate observed among diabetic patients, with myocardial fibrosis (MF) being a pivotal element in the disease's progression. Hydrogen sulfide (H2S) has been shown to mitigate MF, but the specific underlying mechanisms have yet to be thoroughly understood. A connection has been established between the evolution of DCM and the incidence of cardiomyocyte pyroptosis. Our research offers insights into H2S protective impact and its probable mode of action against DCM, analyzed through the lens of MF. In this study, a diabetic rat model was developed using intraperitoneal injections of streptozotocin (STZ), and hyperglycemia-stimulated cardiomyocytes were employed to replicate the cellular environment of DCM. There was a marked decline in the expression of cystathionine γ-lyase (CSE), a catalyst for H2S synthesis, in both the STZ-induced diabetic rats and hyperglycemia-stimulated cardiomyocytes. Experimental results in vivo indicated that H2S ameliorates MF and enhances cardiac functionality in diabetic rats by mitigating cardiomyocyte pyroptosis. In vitro assessments highlighted the induction of cardiomyocyte pyroptosis and the subsequent decline in cell viability under hyperglycemic conditions. However, the administration of sodium hydrosulfide (NaHS) curtailed cardiomyocyte pyroptosis and augmented cell viability. In contrast, propargylglycine (PAG), a CSE inhibitor, reversed the effects rendered by NaHS administration. Additional exploration indicated that the mitigating effect of H2S on cardiomyocyte pyroptosis is modulated through the ROS/NLRP3 pathway. In essence, our findings corroborate the potential of H2S in alleviating MF in diabetic subjects. This therapeutic effect is likely attributable to the regulation of cardiomyocyte pyroptosis via the ROS/NLRP3 pathway. This discovery furnishes a prospective therapeutic target for the amelioration and management of MF associated with diabetes.

MiR-29b-3p regulates cardiomyocytes pyroptosis in CVB3-induced myocarditis through targeting DNMT3A.

Viral myocarditis (VMC) is a disease resulting from viral infection, which manifests as inflammation of myocardial cells. Until now, the treatment of VMC is still a great challenge for clinicians. Increasing studies indicate the participation of miR-29b-3p in various diseases. According to the transcriptome sequencing analysis, miR-29b-3p was markedly upregulated in the viral myocarditis model. The purpose of this study was to investigate the role of miR-29b-3p in the progression of VMC. We used CVB3 to induce primary cardiomyocytes and mice to establish a model of viral myocarditis. The purity of primary cardiomyocytes was identified by immunofluorescence. The cardiac function of mice was detected by Vevo770 imaging system. The area of inflammatory infiltration in heart tissue was shown by hematoxylin and eosin (H&E) staining. The expression of miR-29b-3p and DNMT3A was detected by quantitative real time polymerase chain reaction (qRT-PCR). The expression of a series of pyroptosis-related proteins was detected by western blot. The role of miR-29b-3p/DNMT3A in CVB3-induced pyroptosis of cardiomyocytes was studied in this research. Our data showed that the expression of miR-29b-3p was upregulated in CVB3-induced cardiomyocytes and heart tissues in mice. To explore the function of miR-29b-3p in CVB3-induced VMC, we conducted in vivo experiments by knocking down the expression of miR-29b-3p using antagomir. We then assessed the effects on mice body weight, histopathology changes, myocardial function, and cell pyroptosis in heart tissues. Additionally, we performed gain/loss-of-function experiments in vitro to measure the levels of pyroptosis in primary cardiomyocytes. Through bioinformatic analysis, we identified DNA methyltransferases 3A (DNMT3A) as a potential target gene of miR-29b-3p. Furthermore, we found that the expression of DNMT3A can be modulated by miR-29b-3p during CVB3 infection. Our results demonstrate a correlation between the expression of DNMT3A and CVB3-induced pyroptosis in cardiomyocytes. These findings unveil a previously unidentified mechanism by which CVB3 induces cardiac injury through the regulation of miR-29b-3p/DNMT3A-mediated pyroptosis.

PLD2 deletion ameliorates sepsis-induced cardiomyopathy by suppressing cardiomyocyte pyroptosis via the NLRP3/caspase 1/GSDMD pathway

ObjectiveSepsis-induced cardiomyopathy (SICM) is a life-threatening complication. Phospholipase D2 (PLD2) is crucial in mediating inflammatory reactions and is associated with the prognosis of patients with sepsis. Whether PLD2 is involved in the pathophysiology of SICM remains unknown. This study aimed to investigate the effect of PLD2 knockout on SICM and to explore potential mechanisms.MethodsThe SICM model was established using cecal ligation and puncture in wild-type and PLD2-knockout mice and lipopolysaccharide (LPS)-induced H9C2 cardiomyocytes. Transfection with PLD2-shRNA lentivirus and a PLD2 overexpression plasmid were used to interfere with PLD2 expression in H9C2 cells. Cardiac pathological alterations, cardiac function, markers of myocardial injury, and inflammatory factors were used to evaluate the SICM model. The expression of pyroptosis-related proteins (NLRP3, cleaved caspase 1, and GSDMD-N) was assessed using western blotting, immunofluorescence, and immunohistochemistry.ResultsSICM mice had myocardial tissue damage, increased inflammatory response, and impaired heart function, accompanied by elevated PLD2 expression. PLD2 deletion improved cardiac histological changes, mitigated cTNI production, and enhanced the survival of the SICM mice. Compared with controls, PLD2-knockdown H9C2 exhibits a decrease in inflammatory markers and lactate dehydrogenase production, and scanning electron microscopy results suggest that pyroptosis may be involved. The overexpression of PLD2 increased the expression of NLRP3 in cardiomyocytes. In addition, PLD2 deletion decreased the expression of pyroptosis-related proteins in SICM mice and LPS-induced H9C2 cells.ConclusionPLD2 deletion is involved in SICM pathogenesis and is associated with the inhibition of the myocardial inflammatory response and pyroptosis through the NLRP3/caspase 1/GSDMD pathway.

STUB1 is acetylated by KAT5 and alleviates myocardial ischemia-reperfusion injury through LATS2-YAP-β-catenin axis

Myocardial ischemia-reperfusion injury (MIRI) is involved in the pathogenesis of multiple cardiovascular diseases. This study elucidated the biological function of lysine acetyltransferase 5 (KAT5) in cardiomyocyte pyroptosis during MIRI. Oxygen-glucose deprivation/reoxygenation and left anterior descending coronary artery ligation were used to establish MIRI models. Here we show, KAT5 and STIP1 homology and U-box-containing protein 1 (STUB1) were downregulated, while large tumor suppressor kinase 2 (LATS2) was upregulated in MIRI models. KAT5/STUB1 overexpression or LATS2 silencing repressed cardiomyocyte pyroptosis. Mechanistically, KAT5 promoted STUB1 transcription via acetylation modulation, and subsequently caused ubiquitination and degradation of LATS2, which activated YAP/β-catenin pathway. Notably, the inhibitory effect of STUB1 overexpression on cardiomyocyte pyroptosis was abolished by LATS2 overexpression or KAT5 depletion. Our findings suggest that KAT5 overexpression inhibits NLRP3-mediated cardiomyocyte pyroptosis to relieve MIRI through modulation of STUB1/LATS2/YAP/β-catenin axis, providing a potential therapeutic target for MIRI.

Phospholipid transfer protein ameliorates sepsis-induced cardiac dysfunction through NLRP3 inflammasome inhibition.

Cardiomyocyte pyroptosis is a primary contributor to sepsis-induced cardiac dysfunction (SICD). Recombinant phospholipid transfer protein (PLTP) have been demonstrated to possess anti-inflammatory and antiseptic properties. However, the effect of PLTP on SICD remains unknown. In this study, we established the in vivo and in vitro sepsis model with the recombinant PLTP treatment. The survival rates of mice, mouse cardiac function, cell viability, the protein level of proinflammatory cytokine, and lactate dehydrogenase level were evaluated. The cardiomyocyte pyroptotic changes were observed. The distribution of PLTP and NOD-like receptor thermal protein domain associated protein 3 (NLRP3) in mouse myocardial tissue and expression of PLTP, apoptosis associated speck like protein containing a CARD (ASC), NLRP3, caspase-1, interleukin (IL)-1β, and Gasdermin D (GSDMD) were detected. PLTP ameliorated the cecal ligation and puncture-induced mouse survival rate decrease and cardiac dysfunction, inhibited the IL-1β, IL-18, and tumor necrosis factor (TNF)-α release, and blocked the NLRP3 inflammasome/GSDMD signaling pathway in septic mice. In vitro, PLTP reversed the lipopolysaccharide-induced cardiomyocyte pyroptosis, expression of IL-1β, IL-6, TNF-α, and activation of the NLRP3 inflammasome/GSDMD signal pathway. Moreover, PLTP could bind to NLRP3 and negatively regulate the activity of the NLRP3 inflammasome/GSDMD signal pathway. This study demonstrated that PLTP can ameliorate SICD by inhibiting inflammatory responses and cardiomyocyte pyroptosis by blocking the activation of the NLRP3 inflammasome/GSDMD signaling pathway.

MG53 protects against Coxsackievirus B3-induced acute viral myocarditis in mice by inhibiting NLRP3 inflammasome-mediated pyroptosis via the NF-κB signaling pathway

Pyroptosis, a novel programmed cell death mediated by NOD-like receptor protein 3 (NLRP3) inflammasome, is a critical pathogenic process in acute viral myocarditis (AVMC). Mitsugumin 53 (MG53) is predominantly expressed in myocardial tissues and has been reported to exert cardioprotective effects through multiple pathways. Herein, we aimed to investigate the biological function of MG53 in AVMC and its underlying regulatory mechanism in pyroptosis. BALB/c mice and HL-1 cells were infected with Coxsackievirus B3 (CVB3) to establish animal and cellular models of AVMC. As inflammation progressed in the myocardium, we found a progressive decrease in myocardial MG53 expression, accompanied by a significant enhancement of cardiomyocyte pyroptosis. MG53 overexpression significantly alleviated myocardial inflammation, apoptosis, fibrosis, and mitochondrial damage, thereby improving cardiac dysfunction in AVMC mice. Moreover, MG53 overexpression inhibited NLRP3 inflammasome-mediated pyroptosis, reduced pro-inflammatory cytokines (IL-1β/18) release, and suppressed NF-κB signaling pathway activation both in vivo and in vitro. Conversely, MG53 knockdown reduced cell viability, facilitated cell pyroptosis, and increased pro-inflammatory cytokines release in CVB3-infected HL-1 cells by promoting NF-κB activation. These effects were partially reversed by applying the NF-κB inhibitor BAY 11–7082. In conclusion, our results suggest that MG53 acts as a negative regulator of NLRP3 inflammasome-mediated pyroptosis in CVB3-induced AVMC, partially by inhibiting the NF-κB signaling pathway. MG53 is a promising candidate for clinical applications in AVMC treatment.

PLCE1 enhances mitochondrial dysfunction to promote GSDME-mediated pyroptosis in doxorubicin-induced cardiotoxicity

BackgroundThe therapeutic value and long-term application of doxorubicin (DOX) were hampered by its severe irreversible cardiotoxicity. Phospholipase C epsilon 1 (PLCE 1) was reported as a new member of the phospholipase C (PLC) family which controls the level of phosphoinositides in cells. Pyroptosis is a newly discovered inflammatory type of regulated cell death. Recent studies have consolidated that chemotherapeutic drugs lead to pyroptosis. Additionally, the phosphoinositide signaling system has remarkable effects on the execution of cell death. We aim to investigate the role of PLCE1 and the mechanism of pyroptosis from the context of DOX-induced cardiotoxicity. MethodsIn the current study, in vitro and in vivo experiments were performed to dissect the underlying mechanism of cardiomyocyte pyroptosis during DOX-induced cardiac injury. The molecular mechanism of PLCE1 was identified by the human cardiomyocyte AC16 cell line and C57BL/6 mouse model. ResultsThe results here indicated that PLCE1 high expressed and pyroptotic cell death presented in cardiomyocytes after DOX application, which was negatively correlated to heart function. DOX-induced cell model disclosed pyroptosis mediated by Gasdermin E (GSDME) protein and involved in mitochondrial damage. Conversely, the deletion of PLCE1 ameliorated mitochondrial dysfunction by suppressing ROS accumulation and reversing mitochondrial membrane potential, and then increased cell viability effectively. More importantly, the in vivo experiment demonstrated that inhibition of PLCE1 reduced pyroptotic cell death and improved heart effect. ConclusionsWe discovered firstly that PLCE1 inhibition protected cardiomyocytes from DOX-induced pyroptotic injury and promoted cardiac function. This information offers a theoretical basis for promising therapy.

The E3 ubiquitin ligase MARCH2 protects against myocardial ischemia-reperfusion injury through inhibiting pyroptosis via negative regulation of PGAM5/MAVS/NLRP3 axis

Inflammasome activation and pyroptotic cell death are known to contribute to the pathogenesis of cardiovascular diseases, such as myocardial ischemia-reperfusion (I/R) injury, although the underlying regulatory mechanisms remain poorly understood. Here we report that expression levels of the E3 ubiquitin ligase membrane-associated RING finger protein 2 (MARCH2) were elevated in ischemic human hearts or mouse hearts upon I/R injury. Genetic ablation of MARCH2 aggravated myocardial infarction and cardiac dysfunction upon myocardial I/R injury. Single-cell RNA-seq analysis suggested that loss of MARCH2 prompted activation of NLRP3 inflammasome in cardiomyocytes. Mechanistically, phosphoglycerate mutase 5 (PGAM5) was found to act as a novel regulator of MAVS-NLRP3 signaling by forming liquid-liquid phase separation condensates with MAVS and fostering the recruitment of NLRP3. MARCH2 directly interacts with PGAM5 to promote its K48-linked polyubiquitination and proteasomal degradation, resulting in reduced PGAM5–MAVS co-condensation, and consequently inhibition of NLRP3 inflammasome activation and cardiomyocyte pyroptosis. AAV-based re-introduction of MARCH2 significantly ameliorated I/R-induced mouse heart dysfunction. Altogether, our findings reveal a novel mechanism where MARCH2-mediated ubiquitination negatively regulates the PGAM5/MAVS/NLRP3 axis to protect against cardiomyocyte pyroptosis and myocardial I/R injury.

IP3R2-mediated Ca2+ release promotes LPS-induced cardiomyocyte pyroptosis via the activation of NLRP3/Caspase-1/GSDMD pathway

Pyroptosis plays a crucial role in sepsis, and the abnormal handling of myocyte calcium (Ca2+) has been associated with cardiomyocyte pyroptosis. Specifically, the inositol 1,4,5-trisphosphate receptor type 2 (IP3R2) is a Ca2+ release channel in the endoplasmic reticulum (ER). However, the specific role of IP3R2 in sepsis-induced cardiomyopathy (SIC) has not yet been determined. Thus, this study aimed to investigate the underlying mechanism by which IP3R2 channel-mediated Ca2+ signaling contributes to lipopolysaccharide (LPS)—induced cardiac pyroptosis. The SIC model was established in rats by intraperitoneal injection of LPS (10 mg/kg). Cardiac dysfunction was assessed using echocardiography, and the protein expression of relevant signaling pathways was analyzed using ELISA, RT-qPCR, and western blot. Small interfering RNAs (siRNA) and an inhibitor were used to explore the role of IP3R2 in neonatal rat cardiomyocytes (NRCMs) stimulated by LPS in vitro. LPS-induced NLRP3 overexpression and GSDMD-mediated pyroptosis in the rats’ heart. Treatment with the NLRP3 inhibitor MCC950 alleviated LPS-induced cardiomyocyte pyroptosis. Furthermore, LPS increased ATP-induced intracellular Ca2+ release and IP3R2 expression in NRCMs. Inhibiting IP3R activity with xestospongin C (XeC) or knocking down IP3R2 reversed LPS-induced intracellular Ca2+ release. Additionally, inhibiting IP3R2 reversed LPS-induced pyroptosis by suppressing the NLRP3/Caspase-1/GSDMD pathway. We also found that ER stress and IP3R2-mediated Ca2+ release mutually regulated each other, contributing to cardiomyocyte pyroptosis. IP3R2 promotes NLRP3-mediated pyroptosis by regulating ER Ca2+ release, and the mutual regulation of IP3R2 and ER stress further promotes LPS-induced pyroptosis in cardiomyocytes.

NOX4 aggravates doxorubicin-induced cardiomyocyte pyroptosis by increasing reactive oxygen species content and activating the NLRP3 inflammasome.

Nicotinamide adenine dinucleotide phosphate oxidase 4 (NOX4)-mediated reactive oxygen species (ROS) has been reported to induce cardiomyocyte apoptosis, but its effect on pyroptosis of cardiomyocytes has been rarely reported. This paper aimed to explore the effects of NOX4-mediated ROS production on doxorubicin (DOX)-induced myocardial injury and pyroptosis through nucleotide-binding and oligomerization domain-like receptor protein 3 (NLRP3) inflammasome. HL-1 cells were treated with DOX or mice (30 mice were divided into five groups with six mice/group) underwent intraperitoneal injection with DOX (5 mg/kg, once a week, five times) to induce myocardial injury, followed by assessment of NOX4 and NLRP3 expression in cell supernatant and myocardial tissues. In cardiomyocyte HL-1 cells, cell proliferation was tested by MTT assay and the activity of ROS by probes. The superoxide dismutase (SOD) activity, malondialdehyde (MDA) content, and glutathione (GSH) activity were evaluated by kits. The expression of pyroptosis proteins was assessed by western blotting. Subsequently, the expression of NOX4 or NLRP3 was altered to determine the effect of NOX4 or NLRP3 expression on cardiomyocyte injury and pyroptosis. The animal models were utilized to evaluate the changes in the cardiac function of mice using an echocardiographic system, with these parameters measured including left ventricular ejection fraction (LVEF), left ventricular fractional shortening (LVFS), and left ventricular end-diastolic diameter (LVEDD). Furthermore, the content of myocardial injury markers and the protein expression of pyroptosis proteins were determined to evaluate myocardial injury in the mice. DOX treatment led to cardiomyocyte injury and pyroptosis, as evidenced by weakened LVEF, LVFS, and cell proliferation (P<0.05), elevated LVEDD, ROS, and MDA (P<0.05), increased expression of pyroptosis proteins (P<0.05), and decreased SOD and GSH (P<0.05). Additionally, NOX4 and NLRP3 were highly-expressed (P<0.05) in cell supernatant and myocardial tissues. In DOX-induced HL-1 cells, the overexpression of NOX4 intensified ROS levels to aggravate cardiomyocyte injury and pyroptosis, which was reversed by treatment of the ROS scavenger N-acetyl-cysteine. Furthermore, it was revealed that the combination of short hairpin RNA (sh)-NOX4 and overexpressed (oe)-NLRP3 reversed the cardioprotective effects of sh-NOX4 and increased myocardial tissue or cell injury and pyroptosis in vitro and in vivo. No mice died during the animal experiments, and only two were ruled out due to a weight loss greater than 20%. NOX4-mediated ROS production activated NLRP3 inflammasome, thereby aggravating DOX-induced myocardial injury in vitro and in vivo.

LncRNA SNHG1 alleviates myocardial ischaemia-reperfusion injury by regulating the miR-137-3p/KLF4/TRPV1 axis.

Myocardial ischaemia-reperfusion injury (MIRI) contributes to serious myocardial injury and even death. Long non-coding RNAs (lncRNAs) have been reported to play pivotal roles in the occurrence and development of MIRI. Here, the detailed molecular mechanism of lncRNA SNHG1 in MIRI was explored. A cell model of MIRI was established through hypoxia/reoxygenation (H/R) stimulation. Cell viability and pyroptosis were evaluated utilizing MTT, PI staining, and flow cytometry. Interleukin (IL)-1β and IL-18 secretion levels were examined by ELISA. The gene and protein expression were detected by RT-qPCR and western blot, respectively. Dual luciferase reporter gene, RIP and ChIP assays were performed to analyse the molecular interactions. The results showed that lncRNA SNHG1 overexpression alleviated H/R-induced HL-1 cell pyroptosis (all P<0.05). LncRNA SNHG1 promoted KLF4 expression by sponging miR-137-3p. miR-137-3p silencing alleviated H/R-induced pyroptosis in HL-1 cells (all P<0.05), which was abolished by KLF4 knockdown (all P<0.05). KLF4 activated the AKT pathway by transcriptionally activating TRPV1 in HL-1 cells (all P<0.05). TRPV1 knockdown reversed the alleviation of SNHG1 upregulation on H/R-induced pyroptosis in HL-1 cells (all P<0.05). These results showed that lncRNA SNHG1 assuaged cardiomyocyte pyroptosis during MIRI progression by regulating the KLF4/TRPV1/AKT axis through sponging miR-137-3p. Our findings may provide novel therapeutic targets for MIRI.

Pharmacological inhibition of ICOS attenuates the protective effect of exercise on cardiac fibrosis induced by isoproterenol

AimsTo investigate the cardioprotective mechanism of exercise or exercise combined with inducible costimulatory molecules (ICOS) monoclonal antibody (mAb) therapy against isoproterenol (ISO)-induced cardiac remodeling. Main methodsTotally 24 male C57BL/6J mice were randomly divided into four groups: the control group (normal saline treatment), ISO group (subcutaneous injection of isoproterenol, 10 mg/kg/day, once daily for 5 consecutive days), the exercise with subcutaneous ISO injection group (EPI), and the exercise with injected with ISO and ICOS mAb group (EPII). The mice in EPI and EPII group were trained on a small animal treadmill for 4 weeks (13 m/min, 0% grade, 60min/day). Key findingsExercise significantly attenuated CD45+, Mac-2 inflammatory cell infiltration, cardiac fibrosis and inhibited the RIPK1/RIPK3/MLKL/CaMKII and cardiomyocyte pyroptosis pathways to counter ISO-induced severe cardiac injury. The administration of the ICOS mAb may inhibit the cardioprotection of exercise against ISO-induced heart damage. Compared to those in EPI, our data showed that the increasing levels of myocardial fibrosis, the leukocyte infiltration of cardiac tissue and proteins expression of cardiac myocyte necrosis and pyroptosis signaling pathways in the EPII group. SignificanceOur results demonstrated that exercise decreased leukocyte infiltration in heart, inhibited the cardiomyocyte pyroptosis and necroptosis signaling pathways, and attenuated inflammatory responses to alleviate ISO-induced cardiac fibrosis. However, the antifibrotic effects of combined treatment with exercise and ICOS mAb intervention did not exhibit synergistic enhancement.

Atrazine exposure promotes cardiomyocyte pyroptosis to exacerbate cardiotoxicity by activating NF-κB pathway

Atrazine is a ubiquitous herbicide with persistent environmental presence and accumulation in the food chain, posing potential health hazards to organisms. Increasing evidence suggests that atrazine may have detrimental effects on various organ systems, including the nervous, digestive, and immune systems. However, the specific toxicity and underlying mechanism of atrazine-induced cardiac injury remain obscure. In this study, 4-week-old male C57BL/6 mice were administered atrazine via intragastric administration at doses of 50 and 200 mg/kg for 4 and 8 weeks, respectively. Our findings showed that atrazine exposure led to cardiac fibrosis, as evidenced by elevated heart index and histopathological scores, extensive myofiber damage, and interstitial collagen deposition. Moreover, atrazine induced cardiomyocyte apoptosis, macrophage infiltration, and excessive production of inflammatory factors. Importantly, atrazine upregulated the expressions of crucial pyroptosis proteins, including NLRP3, ASC, CASPASE1, and GSDMD, via the activation of NF-κB pathway, thus promoting cardiomyocyte pyroptosis. Collectively, our findings provide novel evidence demonstrating that atrazine may exacerbate myocardial fibrosis by inducing cardiomyocyte pyroptosis, highlighting its potential role in the development of cardiac fibrosis.

Colchicine ameliorates myocardial injury induced by coronary microembolization through suppressing pyroptosis via the AMPK/SIRT1/NLRP3 signaling pathway

BackgroundCoronary microembolization(CME)is a common complication in acute coronary syndrome and percutaneous coronary intervention, which is closely related to poor prognosis. Pyroptosis, as an inflammatory programmed cell death, has been found to be associated with CME-induced myocardial injury. Colchicine (COL) has potential benefits in coronary artery disease due to its anti-inflammatory effect. However, the role of colchicine in pyroptosis-related CME-induced cardiomyocyte injury is unclear. This study was carried out to explore the effects and mechanisms of colchicine on myocardial pyroptosis induced by CME.MethodsThe CME animal model was constructed by injecting microspheres into the left ventricle with Sprague-Dawley rats, and colchicine (0.3 mg/kg) pretreatment seven days before and on the day of modeling or compound C(CC)co-treatment was given half an hour before modeling. The study was divided into 4 groups: Sham group, CME group, CME + COL group, and CME + COL + CC group (10 rats for each group). Cardiac function, serum myocardial injury markers, myocardial histopathology, and pyroptosis-related indicators were used to evaluate the effects of colchicine.ResultsColchicine pretreatment improved cardiac dysfunction and reduced myocardial injury induced by CME. The main manifestations were the improvement of left ventricular systolic function, the decrease of microinfarction area, and the decrease of mRNA and protein indexes related to pyroptosis. Mechanistically, colchicine increased the phosphorylation level of adenosine monophosphate-activated protein kinase (AMPK), promoted the expression of silent information regulation T1 (SIRT1), and inhibited the expression of NOD-like receptor pyrin containing 3 (NLRP3) to reduce myocardial pyroptosis. However, after CC co-treatment with COL, the effect of colchicine was partially reversed.ConclusionColchicine improves CME-induced cardiac dysfunction and myocardial injury by inhibiting cardiomyocyte pyroptosis through the AMPK/SIRT1/NLRP3 signaling pathway.

SIAH1 Promotes the Pyroptosis of Cardiomyocytes in Diabetic Cardiomyopathy via Regulating IκB-α/NF-κВ Signaling.

Inflammation-mediated dysfunction of cardiomyocytes is the main cause of diabetic cardiomyopathy (DCM). The present study aimed to investigate the roles of siah E3 ubiquitin protein ligase 1 (SIAH1) in DCM. The online dataset GSE4172 was used to analyze the differentially expressed genes in myocardial inflammation of DCM patients. RT-qPCR was conducted to detect mRNA levels. Enzyme-Linked Immunosorbent Assay (ELISA) was performed to detect cytokine release. Western blot was used to detect protein expression. Lactate dehydrogenase (LDH) assay was used to determine cytotoxicity. In vitro ubiquitination assay was applied to determine the ubiquitination of nuclear factor kappa B inhibitor alpha (1κВ-α). Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay was used to detect the death of cardiomyocytes. Flow cytometry was applied for determining cardiomyocyte pyroptosis. The results showed that SIAH1 was overexpressed in human inflammatory cardiomyopathy. High expression of SIAH1 was associated with inflammatory response. SIAH1 was also overexpressed lipopolysaccharide (LPS)-induced inflammatory cardiomyopathy model in vitro. However, SIAH1 knockdown suppressed the inflammatory-related pyroptosis of cardiomyocytes. SIAH1 promoted the ubiquitination of 1κВ-α and activated nuclear factor kappa В (NF-κВ) signaling, which promoted the pyroptosis of cardiomyocytes. In conclusion, SIAH1 exacerbated the progression of human inflammatory cardiomyopathy via inducing the ubiquitination of 1κВ-α and activation of NF-κВ signaling. Therefore, SIAHI/IκB-α/NF-κB signaling may be a potential target for human inflammatory cardiomyopathy.