ALKBH5 inhibitors as a potential treatment strategy in heart failure-inferences from gene expression profiling.

Heart Failure (HF) is a complex clinical syndrome in which the heart is unable to provide enough blood flow to meet metabolic needs and lacks efficient venous return. HF is a major risk factor for morbidity and mortality with cardiovascular diseases globally. Despite enormous research, the molecular markers relevant to disease prognosis and management remain not well understood. Here, we analyzed the whole transcriptomes of 18 failing hearts and 15 non-failing hearts (predominantly of Caucasian origin), by applying the standard in silico tools. The analyses revealed novel gene-markers including ALKBH5 of mRNA demethylation and KMT2E of histone modification processes, significantly over-expressed in the HF compared with the non-failing hearts (FDR < 0.05). To validate the over-expression of ALKBH5, we determined the global m6A level in hypoxic H9c2 cells using a dot blot assay. The global m6A level was found markedly lower in the hypoxic H9c2 cells than in the control cells. Additionally, the expression of ALKBH5 in the H9c2 cells was quantified by the qPCR and found to be 1.18 times higher at 12 h (p < 0.05), and 1.67 times higher at 24 h of hypoxia (p < 0.01) compared with the control cells, indicating a likely role of ALKBH5 in the failing cardiac cells. Furthermore, we identified several compounds through the virtual screening of 11,272 drug-like molecules of the ZINC15 database to inhibit the ALKBH5 in a molecular docking process. Collectively, the study revealed novel markers potentially involved in the pathophysiology of HF and suggested plausible therapeutic molecules for the management of the disease.

Epigenetic Regulation of Macrophage Polarization in Cardiovascular Diseases.

Cardiovascular diseases (CVDs) are the leading cause of hospitalization and death worldwide, especially in developing countries. The increased prevalence rate and mortality due to CVDs, despite the development of several approaches for prevention and treatment, are alarming trends in global health. Chronic inflammation and macrophage infiltration are key regulators of the initiation and progression of CVDs. Recent data suggest that epigenetic modifications, such as DNA methylation, posttranslational histone modifications, and RNA modifications, regulate cell development, DNA damage repair, apoptosis, immunity, calcium signaling, and aging in cardiomyocytes; and are involved in macrophage polarization and contribute significantly to cardiac disease development. Cardiac macrophages not only trigger damaging inflammatory responses during atherosclerotic plaque formation, myocardial injury, and heart failure but are also involved in tissue repair, remodeling, and regeneration. In this review, we summarize the key epigenetic modifications that influence macrophage polarization and contribute to the pathophysiology of CVDs, and highlight their potential for the development of advanced epigenetic therapies.

Potential therapeutic strategies for myocardial infarction: the role of Toll-like receptors.

Myocardial infarction (MI) is a life-threatening condition among patients with cardiovascular diseases. MI increases the risk of stroke and heart failure and is a leading cause of morbidity and mortality worldwide. Several genetic and epigenetic factors contribute to the development of MI, suggesting that further understanding of the pathomechanism of MI might help in the early management and treatment of this disease. Toll-like receptors (TLRs) are well-known members of the pattern recognition receptor (PRR) family and contribute to both adaptive and innate immunity. Collectively, studies suggest that TLRs have a cardioprotective effect. However, prolonged TLR activation in the response to signals generated by damage-associated molecular patterns (DAMPs) results in the release of inflammatory cytokines and contributes to the development and exacerbation of myocardial inflammation, MI, ischemia-reperfusion injury, myocarditis, and heart failure. The objective of this review is to discuss and summarize the association of TLRs with MI, highlighting their therapeutic potential for the development of advanced TLR-targeted therapies for MI.

GSK3ß mediates the spatiotemporal dynamics of NLRP3 inflammasome activation.

Subcellular machinery of NLRP3 is essential for inflammasome assembly and activation. However, the stepwise process and mechanistic basis of NLRP3 engagement with organelles remain unclear. Herein, we demonstrated glycogen synthase kinase 3ß (GSK3ß) as a molecular determinant for the spatiotemporal dynamics of NLRP3 inflammasome activation. Using live cell multispectral time-lapse tracking acquisition, we observed that upon stimuli NLRP3 was transiently associated with mitochondria and subsequently recruited to the Golgi network (TGN) where it was retained for inflammasome assembly. This occurred in relation to the temporal contact of mitochondria to Golgi apparatus. NLRP3 stimuli initiate GSK3ß activation with subsequent binding to NLRP3, facilitating NLRP3 recruitment to mitochondria and transition to TGN. GSK3ß activation also phosphorylates phosphatidylinositol 4-kinase 2 Α (PI4k2A) in TGN to promote sustained NLRP3 oligomerization. Our study has identified the interplay between GSK3ß signaling and the organelles dynamics of NLRP3 required for inflammasome activation and opens new avenues for therapeutic intervention.

The caspase-1 inhibitor VX765 upregulates connexin 43 expression and improves cell-cell communication after myocardial infarction via suppressing the IL-1ß/p38 MAPK pathway.

Connexin 43 (Cx43) is the most important protein in the gap junction channel between cardiomyocytes. Abnormalities of Cx43 change the conduction velocity and direction of cardiomyocytes, leading to reentry and conduction block of the myocardium, thereby causing arrhythmia. It has been shown that IL-1ß reduces the expression of Cx43 in astrocytes and cardiomyocytes in vitro. However, whether caspase-1 and IL-1ß affect connexin 43 after myocardial infarction (MI) is uncertain. In this study we investigated the effects of VX765, a caspase-1 inhibitor, on the expression of Cx43 and cell-to-cell communication after MI. Rats were treated with VX765 (16 mg/kg, i.v.) 1 h before the left anterior descending artery (LAD) ligation, and then once daily for 7 days. The ischemic heart was collected for histochemical analysis and Western blot analysis. We showed that VX765 treatment significantly decreased the infarct area, and alleviated cardiac dysfunction and remodeling by suppressing the NLRP3 inflammasome/caspase-1/IL-1ß expression in the heart after MI. In addition, VX765 treatment markedly raised Cx43 levels in the heart after MI. In vitro experiments were conducted in rat cardiac myocytes (RCMs) stimulated with the supernatant from LPS/ATP-treated rat cardiac fibroblasts (RCFs). Pretreatment of the RCFs with VX765 (25 µM) reversed the downregulation of Cx43 expression in RCMs and significantly improved intercellular communication detected using a scrape-loading/dye transfer assay. We revealed that VX765 suppressed the activation of p38 MAPK signaling in the heart tissue after MI as well as in RCMs stimulated with the supernatant from LPS/ATP-treated RCFs. Taken together, these data show that the caspase-1 inhibitor VX765 upregulates Cx43 expression and improves cell-to-cell communication in rat heart after MI via suppressing the IL-1ß/p38 MAPK pathway.

GSK-3ß-mediated activation of NLRP3 inflammasome leads to pyroptosis and apoptosis of rat cardiomyocytes and fibroblasts.

We previously demonstrated that GSK-3ß mediates NLRP3 inflammasome activation and IL-1ß production in cardiac fibroblasts (CFs) after myocardial infarction (MI). In this study, we show how GSK-3ß-mediated activation of the NLRP3 inflammasome/caspase-1/IL-1ß pathway leads to apoptosis and pyroptosis of cardiomyocytes (CMs) and CFs. Administration of lipopolysaccharide (LPS)/ATP to primary newborn rat cardiac fibroblasts (RCFs) led to increase in proteins of NLRP3, apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), caspase-1, IL-1ß, and IL-18. Additionally, the expression of caspase-3 and N-terminal fragments of gasdermin D (N-GSDMD) and the Bax/Bcl-2 ratio increased. Administration of the GSK-3ß inhibitor SB216763 reduced the levels of apoptosis- and pyroptosis-related proteins regulated by NLRP3 inflammasome activation in RCFs. Next, we transferred the culture supernatant of LPS/ATP-treated RCFs to in vitro primary newborn rat cardiomyocytes (RCMs). The results showed that SB216763 attenuate the upregulation of the ratios of Bax/Bcl-2 and the expression of caspase-3 and N-GSDMD in RCMs. Direct stimulation of RCMs and H9c2 cells with recombinant rat IL-1ß increased the p-GSK-3ß/GSK-3ß and Bax/Bcl-2 ratios and the expression of caspase-3 and N-GSDMD, while both SB216763 and TLR1 (an IL-1ß receptor inhibitor) markedly reduced these effects, as assessed using propidium iodide positive staining and the lactate dehydrogenase release assay. The caspase-11 inhibitor wedelolactone decreased the expression level of N-GSDMD but did not alter the p-GSK-3ß/GSK-3ß ratio. Lastly, we established a Sprague-Dawley rat MI model to confirm that SB216763 diminished the increase in caspase-3 and N-GSDMD expression and the Bax/Bcl-2 ratio in the ischemic area. These data demonstrate that GSK-3ß regulates apoptosis and pyroptosis of RCMs and RCFs due to NLRP3 inflammasome activation in RCFs.

Glycogen synthase kinase 3 beta inhibitor SB216763 improves Kir2.1 expression after myocardia infraction in rats.

BACKGROUND: Abnormal ion channel currents caused by myocardial electrical remodeling is one of the main causes of malignant arrhythmias. Glycogen synthase kinase 3ß (GSK-3ß) is the main therapeutic target following ischemia as it regulates nerve cell channels. However, few studies have investigated its role in myocardial electrical remodeling. The present study aimed to investigate the role of GSK-3ß in a rat myocardial infarction (MI)-induced electrical remodeling and potential effects on cardiac ionic channels including KCNJ2/Kir2.1/IK1. METHODS: Ligation of the left anterior descending artery in rats was performed to establish a MI model. The rats were randomly divided into three groups, the sham, MI, and MI + SB group. The animals in the latter group were administered SB216763 (GSK-3ß inhibitor) at a dose of 0.6 mg·kg-1·day-1. The ventricular function was assessed by echocardiography, electrocardiography, and histological analysis 7 days post-surgery. Serum was collected to measure lactate dehydrogenase and cardiac troponin I levels, and the mRNA and protein levels of the KCNJ2/Kir2.1/IK1 channel in the heart tissues were assessed. H9c2 cells were cultured to examine the effects of SB216763 on the protein expression of Kir2.1 channel under hypoxic conditions. RESULTS: The results revealed that SB216763 ameliorated acute cardiac injury and improved myocardial dysfunction. Moreover, SB216763 increased the mRNA and protein expression of Kir2.1 during MI. Furthermore, SB216763 treatment abrogated the decreased expression of Kir2.1 in H9c2 cells under hypoxic conditions. CONCLUSIONS: GSK-3ß inhibition upregulates Kir2.1 expression in a rat model of MI.

RNAi silencing of the 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR) gene inhibits vitellogenesis in Chinese mitten crab Eriocheir sinensis.

The sesquiterpenoid methyl farnesoate (MF), a de-epoxide form of insect juvenile hormone III (JH III), plays an essential role in regulating many crucial physiological processes in crustaceans including vitellogenesis and reproduction. 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR) is an important rate-limiting enzyme in the mevalonate pathway, which is critical for the synthesis of JH III and MF. In the present study, a full-length cDNA encoding HMGR (EsHMGR) in Eriocheir sinensis was isolated and characterised. Sequence analysis of EsHMGR revealed that it belongs to Class I HMGR family proteins with HMG-CoA-binding and NADPH-binding domains, both important for HMGR activity. In addition to its ubiquitous tissue expression, expression of EsHMGR was highly specific to the ovary, the main site of Vg synthesis. During ovarian development, EsHMGR expression in ovary displayed a stage-specific pattern, and was correlated with expression of vitellogenin (EsVg) in hepatopancreas, which suggests that EsHMGR possibly involved in vitellogenesis. To further investigate the functional role of EsHMGR in vitellogenin biosynthesis in E. sinensis, RNA interference-mediated gene silencing was carried out both in vitro and in vivo. Quantitative PCR results showed that injection of EsHMGR double-stranded RNA (dsRNA) led to a significant decrease in EsVg expression levels in ovary and hepatopancreas both in vitro and in vivo. Taken together, the results suggest that EsHMGR is involved in vitellogenin biosynthesis in female E. sinensis, which may provide a new resource for HMGR enzymes participating in reproduction in crustaceans.

Ticagrelor reduces doxorubicin-induced pyroptosis of rat cardiomyocytes by targeting GSK-3ß/caspase-1.

Doxorubicin (Dox) is a widely used clinical drug whose cardiotoxicity cannot be ignored. Pyroptosis (inflammatory cell death) has gradually gained attention in the context of Dox-induced cardiotoxicity. In addition to the inhibition of platelet activation by ticagrelor, little is known about its other pharmacological effects. Glycogen synthase kinase 3ß (GSK-3ß) has been shown to contribute to the pathological process of pyroptosis, but whether it is related to the potential role of ticagrelor is unclear. In this study, we investigated the effects of ticagrelor on Dox-induced pyroptosis in cardiomyocytes. Rats were treated with ticagrelor (7.5 mg/kg, i.g.) 1 h before intravenous injection of Dox (2.5 mg/kg), once every 3 days, six times in total. Hearts were collected for histochemical analysis and western blot detection 8 weeks after the last administration. Ticagrelor was shown to significantly improve cardiac function by inhibiting GSK-3ß/caspase-1/GSDMD activation. In vitro experiments were conducted using rat cardiac myocytes (RCMs) and rat embryonic cardiac-derived H9c2 cells. Pretreatment with ticagrelor (10 µm) significantly inhibited Dox (1 µm)-induced hypertrophy and reversed the upregulation of GSDMD-NT expression. We showed that ticagrelor suppressed the activation of Akt caused by Dox in the heart tissue as well as in RCMs/H9c2 cells caused by Dox. When GSK-3ß expression was absent in H9c2 cells, the inhibitory effect of ticagrelor on Dox-induced caspase-1/GSDMD activation was weakened. These data showed that ticagrelor reduced Dox-induced pyroptosis in rat cardiomyocytes by targeting GSK-3ß/caspase-1.

Molecular evidence for farnesoic acid O-methyltransferase (FAMeT) involved in the biosynthesis of vitellogenin in the Chinese mitten crab Eriocheir sinensis.

Sesquiterpenoid methyl farnesoate (MF), a crustacean equivalent of insect juvenile hormone (JH III), has essential functions in regulating physiological processes in crustaceans, including reproduction and vitellogenesis. Farnesoic acid O-methyltransferase (FAMeT) is a key rate-limiting enzyme catalyzing the conversion of farnesoic acid (FA) to JH/MF in insects and crustaceans. In this study, a full-length cDNA of EsFAMeT from Eriocheir sinensis was isolated and characterized. The deduced EsFAMeT amino acid sequence indicated there were two conserved Methyltransf-FA domains characteristic of FAMeT family proteins. With use of sequence alignment analysis procedures, there was an indication that FAMeT proteins are highly conserved among crustaceans and FAMeT is more closely related to crustacean FAMeT than to insect FAMeT. Results from quantitative real-time PCR analysis revealed there was ubiquitous EsFAMeT in all tissues examined, with greater abundances of mRNA transcripts in the ovary. The transcription of EsFAMeT indicated there were stage-specific patterns in the hepatopancreas and ovary during ovarian development, with the greatest abundance during ovarian development Stages II and III, respectively. To investigate functions of EsFAMeT in vitellogenin biosynthesis in E. sinensis, RNA interference-mediated gene knockdown was used in vitro and in vivo. Injection of EsFAMeT dsRNA resulted in a marked decrease in EsVg (encoding vitellogenin) transcripts in the ovary and hepatopancreas both in vitro and in vivo. Results from the present study indicated EsFAMeT is involved in vitellogenin biosynthesis in the ovary and hepatopancreas of E. sinensis, providing a new resource to study modulatory effects of the FAMeT family of enzymes in crustacean reproduction.

Targeting Glycogen Synthase Kinase 3 Beta Regulates CD47 Expression After Myocardial Infarction in Rats via the NF-κB Signaling Pathway.

The aim of this study was to investigate the effects of the GSK-3ß/NF-κB pathway on integrin-associated protein (CD47) expression after myocardial infarction (MI) in rats. An MI Sprague Dawley rat model was established by ligating the left anterior descending coronary artery. The rats were divided into three groups: Sham, MI, and SB + MI (SB216763) groups. Immunohistochemistry was used to observe the changes in cardiac morphology. A significant reduction in the sizes of fibrotic scars was observed in the SB + MI group compared to that in the MI group. SB216763 decreased the mRNA and protein expression of CD47 and NF-κB during MI. Primary rat cardiomyocytes (RCMs) and the H9c2 cell line were used to establish in vitro hypoxia models. Quantitative real-time PCR and western blotting analyses were conducted to detect mRNA and protein expression levels of CD47 and NF-κB and apoptosis-related proteins, respectively. Apoptosis of hypoxic cells was assessed using flow cytometry. SB216763 reduced the protein expression of CD47 and NF-κB in RCMs and H9c2 cells under hypoxic conditions for 12 h, and alleviated hypoxia-induced apoptosis. SN50 (an NF-κB inhibitor) also decreased CD47 protein expression in RCMs and H9c2 cells under hypoxic conditions for 12 h and protected cells from apoptosis. GSK-3ß upregulates CD47 expression in cardiac tissues after MI by activating NF-κB, which in turn leads to myocardial cell damage and apoptosis.

HSPBP1 facilitates cellular RLR-mediated antiviral response by inhibiting the K48-linked ubiquitination of RIG-I.

Retinoic acid-inducible gene I (RIG-I) plays a critical role in the recognition of intracytoplasmic viral RNA. Upon binding to the RNA of invading viruses, the activated RIG-I translocates to mitochondria, where it recruits adapter protein MAVS, causing a series of signaling cascades. In this study, we demonstrated that Hsp70 binding protein 1 (HSPBP1) promotes RIG-I-mediated signal transduction. The overexpression of HSPBP1 can increase the stability of RIG-I protein by inhibiting its K48-linked ubiquitination, and promote the activation of IRF3 and the production of IFN-ß induced by Sendai virus. Knockdown and knockout of HSPBP1 leads to down-regulation of virus-induced RIG-I expression, inhibits IRF3 activation, and reduces the production of IFNB1. These results indicate that HSPBP1 positively regulates the antiviral signal pathway induced by inhibiting the K48-linked ubiquitination of RIG-I.

Potential regulatory role of epigenetic RNA methylation in cardiovascular diseases.

Cardiovascular diseases (CVDs) are the leading cause of morbidity and mortality worldwide, especially in developing countries. To date, several approaches have been proposed for the prevention and treatment of CVDs. However, the increased risk of developing cardiovascular events that result in hospitalization has become a growing public health concern. The pathogenesis of CVDs has been analyzed from various perspectives. Recent data suggest that regulatory RNAs play a multidimensional role in the development of CVDs. Studies have identified several mRNA modifications that have contributed to the functional characterization of various cardiac diseases. RNA methylation, such as N6-methyladenosine, N1-methyladenosine, 5-methylcytosine, N7-methylguanosine, N4-acetylcytidine, and 2'-O-methylation are novel epigenetic modifications that affect the regulation of cell growth, immunity, DNA damage, calcium signaling, apoptosis, and aging in cardiomyocytes. In this review, we summarize the role of RNA methylation in the pathophysiology of CVDs and the potential of using epigenetics to treat such disorders.

Mitochondrial DUT-M potentiates RLR-mediated antiviral signaling by enhancing VISA and TRAF2 association.

Upon recognition of intracytoplasmic viral RNA, activated RIG-I is recruited to the mitochondrion-located adaptor protein VISA (also known as MAVS, CARDIF, and IPS-1). VISA then acts as a central signaling platform for linking RIG-I and downstream signaling components, such as TRAF2, 5, and 6, TBK1, and IKK, leading to activation of the kinases TBK1 and IKK. These activated kinases further phosphorylate the transcription factors IRF3/7 and NF-κB, leading to the induction of downstream antiviral genes. Here, we report a mitochondrial isoform, deoxyuridine triphosphate nucleotidohydrolase (dUTPase), DUT-M, as a positive regulator in RLR-VISA-mediated antiviral signaling. DUT-M interacts with VISA and RIG-I to facilitate the assembly of the VISA-TRAF2 complex and to augment the polyubiquitination of TRAF2, leading to potentiated activation of IRF3 dimerization and phosphorylation of P65, and enhanced VISA-mediated innate immune response. RLR-VISA-mediated IRF3 dimerization and P65 phosphorylation, were inhibited in DUT-knockdown and DUT-deficient 293 cells. Thus, DUT-M is a positive regulator of the RIG-I-VISA-mediated innate immune response to RNA viruses.

Peptidome characterization of ovarian cancer serum and the identification of tumor suppressive peptide ZYX36-58.

BACKGROUND: Several serum biomarkers, including miRNA, mRNA, protein and peptides in cancer patients are also important mediators of cancer progression. METHODS: The differentially expressed peptides between the serum of ovarian cancer patients and healthy controls were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The function of the peptides was analyzed by CCK8, transwell, wound healing, and flow cytometry analysis. And the mechanism of the peptides was analyzed by peptide pull down, and high-throughput RNA-sequencing. RESULTS: A total of 7 and 46 peptides were significantly up-regulated and down-regulated in the serum of ovarian cancer patients, respectively. The precursor proteins of the differentially expressed peptides mainly involved in the complement and coagulation cascades, platelet activation, phagosome and focal adhesion pathways. Interestingly, focal adhesion, platelet activation, platelet-cancer cell interaction, complement activation, coagulation cascades and phagosome formation are all critical factors for cancer initiation or progression, which indicated that the peptides may play a crucial role in cancer development. And we identified one peptide, ZYX36-58, which was down-regulated in the serum of ovarian cancer patients, significantly inhibited invasion and migration and promoted the apoptosis of ovarian cancer cells. Mechanistic study indicated that ZYX36-58 interacted with and increased the protein level of the antiangiogenic protein thrombospondin-1 (TSP1), which has a tumor suppressive effect on ovarian cancer. CONCLUSIONS: ZYX36-58, which was significantly down-regulated in the serum of ovarian cancer patients can significantly inhibit cell invasion, migration and promote apoptosis of ovarian cancer cells by binding and up-regulating TSP1 protein expression.

Cholesterol 25-hydroxylase protects against experimental colitis in mice by modulating epithelial gut barrier function.

Cholesterol 25-hydroxylase (CH25H) encodes the enzyme that converts cholesterol to 25-hydroxycholesterol (25-HC). 25-HC has been demonstrated to be involved in the pathogenesis of inflammatory bowel disease. However, the role of CH25H in experimental colitis remains unknown. Dextran sulfate sodium (DSS)-induced colitis was monitored in wild type and Ch25h-/- mice in 8-week-old male for 7 days by assessment of body weight, histology, inflammatory cellular infiltration, and colon length. The function of CH25H was investigated using loss-of-function and gain-of-function such as Ch25h-deficient mice, supplementation with exogenous 25-HC and treatment of 25-HC into Caco2 and HCT116 colonic epithelial cells. Ch25h-/- mice with DSS-induced colitis exhibited aggravated injury, including higher clinical colitis scores, severe injury of the epithelial barrier, lower tight junction protein levels and higher levels of IL-6. Supplementation with exogenous 25-HC ameliorated disease symptoms and reduced the extent of damage in DSS-induced colitis, which was characterized by lower colon damage, higher tight junction protein expression, significantly decreased local and systemic production of pro-inflammatory cytokines IL-6. In Caco2 and HCT116 cells, 25-HC induced tight junction genes expression in colon cancer epithelial cells. These effects of CH25H were obtained by promoting ATF3 expression. Taken together, our findings reveal a protective role for 25-HC in DSS-induced colitis and the ability of CH25H to maintain epithelial gut barrier function through ATF3 expression. Supplementation with exogenous 25-HC ameliorates disease symptoms, which provides a new therapeutic strategy for ulcerative colitis.

Synthesis of Aluminophosphate Molecular Sieves in Alkaline Media.

Unlike conventional aluminosilicate zeolites synthesized in alkaline media, aluminophosphate molecular sieves (AlPOs) have always been prepared under acidic conditions in the past three decades; this has been regarded as one of essential factors for synthesis, except for the case of silica-substituted analogues (SAPOs). For the first time, we demonstrate herein a simple and generalized route for synthesizing various types of aluminophosphate molecular sieves in alkaline media. A series of aluminophosphate sieves and their analogues have been prepared with different quaternary ammonium cations as structure-directing agents in this manner. The above successes have extended the systematic media from acidic or neutral to alkaline for the preparation of a series of aluminophosphate molecular sieves, which possibly open an alternative route for the synthesis of aluminophosphate molecular sieves.

Activation of NLRP3 inflammasome promotes the proliferation and migration of esophageal squamous cell carcinoma.

Inflammasomes can identify endogenous danger signals as an inflammatory immune response. As the most common inflammasome, the NLR pyrin family domain containing 3 (NLRP3) inflammasome is associated with the pathogenesis of different tumors. However, the function of the NLRP3 inflammasome in esophageal cancer (EC) has rarely been reported. Herein, the expression levels of the components of NLRP3 inflammasome and Ki­67 were analyzed by immunohistochemistry. Furthermore, correlations between the NLRP3 inflammasome and Ki­67 along with the clinicopathological features of EC patients were evaluated. The components of the NLRP3 inflammasome were also assessed by western blot analysis and quantitative PCR. NLRP3 was silenced or overexpressed in different esophageal squamous cell carcinoma (ESCC) cell lines, and cell viability, migration and invasion were assessed by CCK­8 and Transwell assays. The present results showed that high NLRP3 expression in the tumor specimens was significantly associated with TNM stage and T category. Spearman's correlation analysis revealed a positive correlation between NLRP3 and the Ki­67 proliferation index. The mRNA and protein levels of NLRP3, apoptosis­associated speck­like protein containing a CARD (ASC), cleaved caspase­1, and interleukin (IL)­1ß in tumor tissues were higher than those in non­cancerous tissues. The level of secreted IL­1ß in tumor tissues was also increased, as compared to that in normal tissues. Silencing of NLRP3 in KYSE­70 and TE13 cells strongly attenuated cell viability, decreased cell mobility in wound­healing assays and greatly diminished the ability of cell migration and invasion in the Transwell system. Overexpression of NLRP3 in KYSE­510 and EC9706 cells markedly promoted the proliferation, migration and invasion. Collectively, these results revealed that the the NLRP3 inflammasome is upregulated in human ESCC tissues and promotes ESCC progression. Hence, NLRP3 could be a promising new candidate diagnostic and prognostic target.