Enhancer Of Zeste Homolog 2 Downregulation Research Articles

Lysophosphatidic Acid Induces Podocyte Pyroptosis in Diabetic Nephropathy by an Increase of Egr1 Expression via Downregulation of EzH2.

Podocyte damage and renal inflammation are the main features and pathogenesis of diabetic nephropathy (DN). Inhibition of lysophosphatidic acid (LPA) receptor 1 (LPAR1) suppresses glomerular inflammation and improves DN. Herein, we investigated LPA-induced podocyte damage and its underlying mechanisms in DN. We investigated the effects of AM095, a specific LPAR1 inhibitor, on podocytes from streptozotocin (STZ)-induced diabetic mice. E11 cells were treated with LPA in the presence or absence of AM095, and the expression of NLRP3 inflammasome factors and pyroptosis were measured. A chromatin immunoprecipitation assay and Western blotting were performed to elucidate underlying molecular mechanisms. Gene knockdown by transfecting small interfering RNA was used to determine the role of the transcription factor Egr1 (early growth response protein 1) and histone methyltransferase EzH2 (Enhancer of Zeste Homolog 2) in LPA-induced podocyte injury. AM095 administration inhibited podocyte loss, NLRP3 inflammasome factor expression, and cell death in STZ-induced diabetic mice. In E11 cells, LPA increased NLRP3 inflammasome activation and pyroptosis via LPAR1. Egr1 mediated NLRP3 inflammasome activation and pyroptosis in LPA-treated E11 cells. LPA decreased H3K27me3 enrichment at the Egr1 promoter in E11 cells by downregulating EzH2 expression. EzH2 knockdown further increased LPA-induced Egr1 expression. In podocytes from STZ-induced diabetic mice, AM095 suppressed Egr1 expression increase and EzH2/H3K27me3 expression reduction. Collectively, these results demonstrate that LPA induces NLRP3 inflammasome activation by downregulating EzH2/H3K27me3 and upregulating Egr1 expression, resulting in podocyte damage and pyroptosis, which may be a potential mechanism of DN progression.

Expression of Enhancer of Zeste Homolog 2 (EZH2) Gene in Acute Myeloid Leukemia.

Enhancer of Zeste Homolog 2 (EZH2) is the catalytic subunit of the chromatin modifying enzyme polycomb repressive complex 2 (PRC2). As a complex, these proteins selectively silence target genes through trimethylation of histone 3 at lysine 27. EZH2 is strongly oncogenic. It has been observed in various malignancies which makes it an interesting therapeutic target. Whether it functions as a tumor suppressor or oncogene in acute leukemia is not settled. Aim of this work: This study aimed at determining the expression levels of EZH2 gene in a cohort of adult Egyptian patients newly diagnosed with acute myeloid leukemia (AML). The present study included 45 de novo AML patients and 40 healthy subjects of matched age and sex as a control group. All study participants were subjected to complete blood count (CBC), bone marrow examination, immunophenotyping, conventional cytogenetic studies and Detection of EZH2 gene expression levels by real time quantitative polymerase chain reaction (RQ-PCR). EZH2 was significantly downregulated in AML patients compared to controls (p<0.001). There was no significant difference in EZH2 level when considering age, sex, bone marrow blasts count, cytogenetic studies, type or site of infection. Low EZH2 expression was associated with higher mortality (31 patients, 68.9%). Low EZH2 expression is prevalent in Egyptian AML patients subsequently; it is suggested to function as tumor suppressor gene rather than an oncogene. Moreover, EZH2 downregulation is associated with resistance to chemotherapy and high mortality rate.

Mechanical stretch aggravates vascular smooth muscle cell apoptosis and vascular remodeling by downregulating EZH2

Enhancer of zeste homolog 2 (EZH2) was recently found to play an important role in cardiovascular disease. However, the role of EZH2 in vascular remodeling induced by mechanical stretch is poorly understood. The aim of the present work was to investigate the role of EZH2 in regulating smooth muscle cell function through mechanical stretch assays and to explore the underlying mechanisms. WT C57BL/6J mice underwent sham surgery or abdominal aortic constriction. The level of EZH2 expression was determined by Western blotting and immunohistochemical staining. We demonstrated the thickness of vascular remodeling by HE staining. JASPAR was used to predict transcription factors that could affect EZH2. Chromatin immunoprecipitation was used to substantiate the DNAprotein interactions. Promoter luciferase assays were performed to demonstrate the activity of the transcription factors. We found that in vivo, AAC significantly reduced EZH2 protein levels in the thoracic aorta. Smooth muscle-specific overexpression of EZH2 was sufficient to attenuate the AAC-induced reduction in trimethylation of Lys-27 in histone 3 and thickening of the arterial media. Administration of GSK-J4 (an inhibitor of H3K27me3 demethylase) induced the same effects. In addition, we found that mechanical stretch regulated the expression of EZH2 through the Yes-associated protein (YAP)- transcriptional factor TEA domain 1 (TEAD) pathway. TEAD1 bound directly to the promoter of EZH2, and blocking the YAP-TEAD1 interaction inhibited EZH2 downregulation due to mechanical stretch. This study reveals that mechanical stretch downregulates EZH2 through the YAP-TEAD1 pathway, thereby aggravating smooth muscle cell apoptosis and vascular remodeling.

LncRNA colorectal neoplasia differentially expressed promotes glycolysis of liver cancer cells by regulating hypoxia-inducible factor 1α.

LncRNAs are associated with tumorigenesis of liver cancer. LncRNA Colorectal Neoplasia Differentially Expressed (CRNDE) was identified as an oncogenic lncRNA and involved in tumor growth and metastasis. The role of CRNDE in liver cancer was investigated. CRNDE was elevated in liver cancer cells. Knockdown of CRNDE decreased cell viability and inhibited proliferation of liver cancer. Moreover, knockdown of CRNDE reduced levels of extracellular acidification rate, glucose consumption, and lactate production to repress glycolysis of liver cancer. Silence of CRNDE enhanced the expression of miR-142 and reduced enhancer of zeste homolog 2 (EZH2) and hypoxia-inducible factor 1α (HIF-1α). Over-expression of HIF-1α attenuated CRNDE silence-induced decrease of glucose consumption and lactate production. Injection with sh-CRNDE virus reduced in vivo tumor growth of liver cancer through up-regulation of miR-142 and down-regulation of EZH2 and HIF-1α. In conclusion, knockdown of CRNDE suppressed cell proliferation, glycolysis, and tumor growth of liver cancer through EZH2/miR-142/HIF-1α.

Dextran Sulfate Inhibits Cell Proliferation and Induces Apoptosis by Regulating EZH2 in Gastric Carcinoma.

Gastric cancer (GC) is one of the most common gastrointestinal malignancies. According to reports, the enhancer of zeste homolog 2 (EZH2) exhibits carcinogenic function in a variety of cancers. Therefore, EZH2 may be a potential therapeutic target for the treatment of human cancer. Macromolecular Dextran Sulfate (DS) has been displayed to play a critical role in tumor inhibition. However, the molecular mechanism by which DS mediates this effect is unclear. In this study, we explored the effects of DS on the proliferation and apoptosis of gastric cancer and the related mechanisms. Cell proliferation and counting assays, as well as cell colony formation assays, revealed that DS inhibited the proliferation and tumorigenesis of GC cells. Additionally, flow cytometry analysis displayed that DS blocked the cell cycle of GC cells in the G1/S phase and promoted their apoptosis. Bioinformatics analyses, enzyme-linked immunosorbent assays, immunohistochemistry, and other methods were applied to measure the expression of EZH2 in human GC cells and tissues. Further studies have shown that DS treatment can reduce the expression of proliferating cell nuclear antigen (PCNA) and increase the level of the ratio of Bax: Bcl-2 protein in GC cells. In addition, DS reduced EZH2 levels and increased CXXC finger protein 4 levels both in vitro and in vivo. In addition, down-regulation of EZH2 with EZH2 inhibitors reversed the inhibitory effect of DS on gastric cancer cells. Collectively, our work demonstrates that DS suppresses proliferation and promotes apoptosis of GC cells by regulating EZH2. Our study suggests that DS is a promising therapeutic compound for the treatment of GC.

Downregulation of EZH2 in Trophoblasts Induces Decidual M1 Macrophage Polarization: a Potential Cause of Recurrent Spontaneous Abortion

Macrophages are known to be pivotal for ensuring the establishment of the immune tolerance microenvironment at the maternal–fetal interface. In particular, trophoblasts stay in close contact with decidual macrophages (DMs), which have been reported to play an active role in the modulation of the polarization of DMs. Thus, any dysfunction of trophoblasts might be associated with certain pregnancy‐related complications, such as recurrent spontaneous abortion (RSA). Enhancer of zeste homolog 2 (EZH2) is an important epigenetic regulatory gene that has been previously shown to be related to immune regulation. The present study assessed the expression of EZH2 in villi tissue obtained from healthy controls and RSA patients. Trophoblasts conditioned medium was collected to incubate macrophages differentiated from the THP‐1 cell line. The expression and function of EZH2 in trophoblasts were knocked down either by the use of siRNA or GSK126 as an inhibitor. Our results show a significant decrease in the expression of EZH2 in villi tissue from RSA patients as compared to healthy controls. Further, the inhibition of expression or function of EZH2 in trophoblasts promoted M1 macrophage polarization, which might be involved in the pathogenesis of RSA. Moreover, the suppression of EZH2 was found to affect the secretion of immune and inflammatory cytokines in trophoblasts. Altogether, these results indicated the importance of EZH2 in the regulation of immune functions of trophoblasts and thus highlighted its potential to be explored as a therapeutic target to prevent and treat pregnancy loss.

FLT3 Inhibition Downregulates EZH2 in AML and Promotes Myeloid Differentiation

Internal tandem duplication mutations in the Fms-like tyrosine kinase 3 (FLT3-ITD) are frequently recurring in AML and confer a poor prognosis. FLT3 inhibitors (FLT3i) such as gilteritinib are efficacious in relapsed AML. Clinical responses to FLT3i include myeloid differentiation of the FLT3-ITD clone in about 50% of patients. How FLT3i induce this response in a subset of patients is unknown.The FLT3i-induced differentiation response seen in clinical trials has not previously been demonstrated in animal models. We modeled FLT3i-induced differentiation in murine Flt3 ITD/ITDDnmt3a -/- AML model (Meyer et al., Cancer Discovery, 2016). Treatment with FLT3i in vitro accelerated differentiation of cKIT+ leukemic splenocytes as assessed by colony morphology in serial re-plating assays. To characterize the differentiation response in vivo, we transplanted CD45.2+ leukemic splenocytes from moribund mice into sub-lethally irradiated healthy congenic CD45.1+ mice. After confirmation of engraftment at 2 weeks post-irradiation, mice were treated with vehicle or gilteritinib for 4 weeks. Animals treated with gilteritinib demonstrated increased neutrophil and decreased stem/progenitor cell populations, recapitulating the clinically observed increase in granulocytic differentiation of the FLT3-ITD clone.We next sought to understand the molecular mechanism of FLT3i-induced differentiation. We used a proteomic-based screen in a human AML cell line treated with FLT3i to identify novel targets of FLT3-ITD that could be potential mediators of the differentiation response. We identified downregulation of Enhancer of Zeste Homolog 2 (EZH2), the catalytic component of the Polycomb Repressive Complex 2 (PRC2). EZH2 and PRC2 were previously shown to be required for leukemic maintenance in mouse models of MLL-AF9 AML. We treated murine Flt3 ITD/ITDDnmt3a -/- cKIT+ leukemic splenocytes with FLT3i or the EZH1/2 inhibitor (EZH1/2i). Both promoted myeloid differentiation to similar degrees as assessed by colony morphology in this model. We hypothesized that FLT3-ITD regulates EZH2 to maintain leukemia cells in a stem/progenitor cell state. We, therefore, characterized the effect of FLT3i on PRC2 in more detail. We confirmed that FLT3i decreases EZH2 protein levels in FLT3-ITD cell lines and primary human AML within 24 hours of treatment as suggested by our proteomic data (Figure 1A-B). We found that the mechanism of EZH2 downregulation is complex with both transcriptional effects and a decrease in EZH2 protein half-life. ChIP-Seq for H3K27me3 demonstrated decreased peaks at the transcription start sites of PRC2 target genes (Figure 1C). RNA-Seq gene expression profiles of FLT3i- and EZH1/2i-treated human AML cells overlapped at 253 differentially expressed genes (Figure 1D). Critically, both FLT3i and EZH1/2i expression profiles enriched in differentiated myeloid cell gene signatures.Overall, we found that EZH2 is a novel, unexpected, and clinically relevant target of FLT3-ITD. Our data suggest that reduced EZH2 activity following FLT3 inhibition promotes myeloid differentiation of FLT3-ITD leukemic cells, providing a mechanistic explanation for the FLT3i-induced differentiation response seen in patients. These data demonstrate that FLT3-ITD has at least two functions in leukemogenesis, the well described activation of signaling pathways, and second, a previously undefined, regulation of PRC2 to maintain a myeloid stem cell state. Our results may lead to improved approaches to therapy for FLT3 mutated AML. [Display omitted] DisclosuresBernt: Syndax: Research Funding; Merck: Other: Spouse is an employee of Merck.. Carroll: Incyte Pharmaceuticals: Research Funding; Janssen Pharmaceutical: Consultancy.

Combination of curcumin with N-n-butyl haloperidol iodide inhibits hepatocellular carcinoma malignant proliferation by downregulating enhancer of zeste homolog 2 (EZH2) - lncRNA H19 to silence Wnt/β-catenin signaling

BackgroundHepatocellular carcinoma (HCC) is one of the most common cause of cancer-related death worldwide. Curcumin (C) has been extensively investigated in different types of malignancies, including hepatocellular carcinoma, but its physicochemical properties have significantly influenced its clinical use. Several approaches are being explored to enhance curcumin's therapeutic response, including its combination with various drugs. PurposeThis study aimed to evaluate the anti-tumor effect of curcumin (C) in combination with F2 (N-n-butyl haloperidol iodide) on hepatocellular carcinoma and its potential underlying mechanism in vitro and in vivo. MethodsCell proliferation was evaluated by CCK-8 and colony formation assays, and apoptosis was measured by flow cytometry. The migratory and invasive abilities of Hep3B and SMMC-7721 cells were measured by wound-healing and matrigel transwell assays. In order to investigate the molecular pathways, various experiments such as western blotting, qPCR, RNA-seq, immunostaining and transfection were performed. To evaluate the anti-HCC effects in vivo, a xenograft tumor model was used. ResultsOur findings showed that the combination of curcumin (C) & F2 (F2C) strongly inhibited malignant proliferation and migration in SMMC-7721 and Hep3B cells. The F2C treatment downregulates enhancer of zeste homolog 2 (EZH2) transcription and protein expression, which is key epigenetic regulator responsible for HCC development. Moreover, the inhibition of EZH2 by F2C led to Wnt/β-catenin signaling inhibition by decreasing tri-methylation of histone H3 at lysine 27 (H3K27me3) and long non-coding RNA H19 expression. The inhibition of F2C was associated with the suppression of tumorigenicity in xenograft HCC models. ConclusionThese findings suggested that, F2C inhibited HCC formation, migration and its modulatory mechanism seemed to be associated with downregulation of EZH2, silencing Wnt/β-catenin signaling by interacting with H19, suggesting that F2C may be a promising drug in the clinical treatment of HCC.

RETRACTED: LncRNA HOTAIR regulates the expression of E-cadherin to affect nasopharyngeal carcinoma progression by recruiting histone methylase EZH2 to mediate H3K27 trimethylation

Background/AimThere has been increasing evidence for the function of long non-coding RNA (lncRNA) in nasopharyngeal carcinoma (NPC). We aim to delve into the position of lncRNA HOX antisense intergenic RNA (HOTAIR), together with enhancer of zeste homolog 2 (EZH2), E-cadherin and trimethylation of lysine 27 on histone H3 (H3K27me3) in NPC. MethodsHOTAIR, EZH2, and E-cadherin expression in NPC tissues and cells were tested. NPC cell biological functions were examined through gain-of and loss-of function assays. The mechanism of lncRNA HOTAIR/E-cadherin/EZH2/H3K27 axis in NPC was decoded. ResultsLncRNA HOTAIR and EZH2 were highly expressed in NPC, and E-cadherin was lowly expressed. Down-regulation of HOTAIR or EZH2 inhibited NPC cell progression and tumor growth. HOTAIR recruited histone methylase EZH2 to mediate trimethylation of H3K27 and regulated E-cadherin expression. ConclusionHOTAIR inhibits E-cadherin by stimulating the trimethylation of H3K27 to promote NPC cell progression through recruiting histone methylase EZH2.

Enhancer of zeste homolog 2 participates in the process of atherosclerosis by modulating microRNA-139-5p methylation and signal transducer and activator of transcription 1 expression.

Atherosclerosis (AS) is the main cause of coronary heart disease, in which enhancer of zeste homolog 2 (EZH2) has been implied to participate in this process. Thus, this work proposed to explore the effect of EZH2 on AS from microRNA-139-5p (miR-139-5p)/signal transducer and activator of transcription 1 (STAT1) axis. EZH2, miR-139-5p, and STAT1 expression in arterial tissues of AS patients were detected. Human arterial smooth muscle cells (HASMCs) induced with oxidized low-density lipoprotein (ox-LDL) and the mice fed with high fat diet were treated with silenced EZH2 or upregulated miR-139-5p to explore their roles in proliferation and apoptosis of HASMCs, together with inflammation response and oxidative stress of mice. Chromatin immunoprecipitation experiment was applied to verify the regulatory effect of EZH2 on miR-139-5p through methylation of H3K27me3. The targeting relationship between miR-139-5p and STAT1 was verified by online website and luciferase activity assay. Reduced miR-139-5p and overexpressed EHZ2 and STAT1 were found in AS. Silenced EZH2 or elevated miR-139-5p decreased the production of cholesterol and inhibited inflammation reaction in serum of mice with AS. Silenced EZH2 or elevated miR-139-5p facilitated proliferation and restrained apoptosis of ox-LDL-treated HASMCs, and restrained oxidative stress and cell apoptosis in arterial tissues of AS mice. EZH2 regulated miR-139-5p through H3K27me3, and miR-139-5p targeted STAT1. miR-139-5p silencing antagonized the effects of EZH2 down-regulation on AS. This study manifests that down-regulated EZH2 or elevated miR-139-5p inhibits ox-LDL-induced HASMCs apoptosis, plaque formation, and inflammatory response in AS mice, which may be related to down-regulated STAT1.

EZH2 regulates sFRP4 expression without affecting the methylation of sFRP4 promoter DNA in colorectal cancer cell lines.

Abnormal activation of the Wnt signaling pathway is found in 90% of colorectal cancers (CRCs). Secreted frizzled-related protein 4 (sFRP4) serves as an antagonist of the canonical Wnt signaling pathway. Epigenetic alterations, including changes in DNA methylation and histone methylation, may influence the expression of sFRP4. Polycomb group (PcG) proteins are epigenetic transcriptional repressors that selectively repress gene expression by forming polycomb repressive complexes (PRCs). Enhancer of zeste homolog 2 (EZH2), the core component of PRC2, is a histone-lysine N-methyltransferase that interacts with DNA methyltransferases. In the present study, the promoter DNA methylation status of sFRP4 in CRC cell lines was analyzed and the underlying mechanisms of action governing this modification was investigated. Firstly, the DNA methylation status of the sFRP4 promoter in CRC cell lines was assessed using methylation-specific PCR. Subsequently, the mRNA and protein levels of sFRP4 were measured using real-time qPCR and western blot analysis, respectively, to determine whether the DNA methylation status of the sFRP4 promoter is correlated with its transcriptional levels. To screen for important epigenetic modifiers that may regulate the promoter DNA methylation status of sFRP4, the expression levels of PcG proteins were examined by gene array analysis. ChIP-qPCR was performed to test whether the selected PcG proteins directly bind the promoter region of sFRP4. Finally, the downregulated PcG proteins EZH2, chromobox 7 (CBX7) and jumonji and AT-rich interaction domain containing 2 (JARID2) were identified and their association with sFRP4 expression levels and Wnt/β-catenin signaling pathway activity were investigated. The present study revealed that sFRP4 was hypermethylated in the promoter region and downregulated during the progression of the CRC cell lines from Dukes A to Dukes C. Expression levels of PcG proteins EZH2, CBX7 and JARID2 were upregulated and positively associated with the aberrantly activated Wnt signaling pathway in the CRC cell lines. EZH2, CBX7 and JARID2 were all enriched in the sFRP4 promoter region in CRC cells. EZH2 downregulation did not affect the promoter DNA methylation status of sFRP4 but increased its expression levels and decreased CRC cell proliferation. DNA methylation controls the expression of sFRP4. EZH2 regulates sFRP4 expression without affecting the DNA hypermethylation of the sFRP4 promoter and influences CRC cell proliferation and Wnt/β-catenin signaling pathway activities.

Knockdown of Enhancer of Zeste Homolog 2 Affects mRNA Expression of Genes Involved in the Induction of Resistance to Apoptosis in MOLT-4 Cells.

The Enhancer of Zeste Homolog 2 (EZH2) is a subunit of the polycomb repressive complex 2 that silences the gene transcription via H3K27me3. Previous studies have shown that EZH2 has an important role in the induction of the resistance against the Tumor necrosis factor-Related Apoptosis-Inducing Ligand (TRAIL)-Induced Apoptosis (TIA) in some leukemia cells. The aim of this study was to determine the effect of silencing EZH2 gene expression using RNA interference on the expression of death receptors 4 and 5 (DR4/5), Preferentially expressed Antigen in Melanoma (PRAME), and TRAIL human lymphoid leukemia MOLT-4 cells. Quantitative RT-PCR was used to detect the EZH2 expression and other candidate genes following the siRNA knockdown in MOLT-4 cells. The toxicity of the EZH2 siRNA was evaluated using Annexin V/PI assay following the transfection of the cells by 80 pM EZH2 siRNA at 48 hours. Based on the flow-cytometry results, the EZH2 siRNA had no toxic effects on MOLT-4 cells. Also, the EZH2 inhibition increased the expression of DR4/5 but reduced the PRAME gene expression at the mRNA levels. Moreover, the EZH2 silencing could not change the TRAIL mRNA in the transfected cells. Our results revealed that the down-regulation of EZH2 in MOLT-4 cells was able to affect the expression of important genes involved in the induction of resistance against TIA. Hence, we suggest that the silencing of EZH2 using RNA interference can be an effective and safe approach to help defeat the MOLT-4 cell resistance against TIA.

Increased EZH2 Levels in Anterior Cingulate Cortex Microglia Aggravate Neuropathic Pain by Inhibiting Autophagy Following Brachial Plexus Avulsion in Rats.

After brachial plexus avulsion (BPA), microglia induce inflammation, initiating and maintaining neuropathic pain. EZH2 (enhancer of zeste homolog 2) has been implicated in inflammation and neuropathic pain, but the mechanisms by which it regulates neuropathic pain remain unclear. Here, we found that EZH2 levels were markedly upregulated during BPA-induced neuropathic pain in vivo and in vitro, stimulating pro-inflammatory cytokines (IL-1β, TNF-α, and IL-6) secretion in vivo. In rats with BPA-induced neuropathic pain, mechanical and cold hypersensitivities were induced by EZH2 upregulation and inhibited by EZH2 downregulation in the anterior cingulate cortex. Microglial autophagy was also significantly inhibited, with EZH2 inhibition activating autophagy and reducing neuroinflammation in vivo. However, this effect was impaired by inhibiting autophagy with 3-methyladenine, suggesting that the MTOR signaling pathway is a functional target of EZH2. These data suggest that EZH2 regulates neuroinflammation and neuropathic pain via a novel MTOR-mediated autophagy signaling pathway, providing a promising approach for managing neuropathic pain.

A positive feedback loop between EZH2 and NOX4 regulates nucleus pulposus cell senescence in age-related intervertebral disc degeneration

BackgroundThe senescence of nucleus pulposus (NP) cells plays a vital role in the pathogenesis of intervertebral disc (IVD) degeneration (IDD). NADPH oxidase 4 (NOX4)-associated oxidative stress has been shown to induce premature NP cell senescence. Enhancer of zeste homolog 2 (EZH2) is a crucial gene regulating cell senescence. The aim of this study was to investigate the roles of EZH2 in NOX4-induced NP cell senescence and a feedback loop between EZH2 and NOX4.ResultsThe down-regulation of EZH2 and the up-regulation of NOX4 and p16 were observed in the degenerative discs of aging rats. EZH2 regulated NP cell senescence via the H3K27me3-p16 pathway. Also, EZH2 regulated the expression of NOX4 in NP cells through the histone H3 lysine 27 trimethylation (H3K27me3) in the promoter of NOX4 gene. Furthermore, NOX4 down-regulated EZH2 expression in NP cells via the canonical Wnt/β-catenin pathway.ConclusionsA positive feedback loop between EZH2 and NOX4 is involved in regulating NP cell senescence, which provides a novel insight into the mechanism of IDD and a potential therapeutic target for IDD.

EZH2 deficiency attenuates Treg differentiation in rheumatoid arthritis

The chromatin modifier enhancer of zeste homolog 2 (EZH2) methylates lysine 27 of histone H3 (H3K27) and regulates T cell differentiation. However, the potential role of EZH2 in the pathogenesis of rheumatoid arthritis (RA) remains elusive. We analyzed EZH2 expression in PBMC, CD4+ T cells, CD19+ B cell, and CD14+ monocytes from active treatment-naïve RA patients and healthy controls (HC). We also suppressed EZH2 expression using EZH2 inhibitor GSK126 and measured CD4+ T cell differentiation, proliferation and apoptosis. We further examined TGFβ-SMAD and RUNX1 signaling pathways in EZH2-suppressed CD4+ T cells. Finally, we explored the regulation mechanism of EZH2 by RA synovial fluid and fibroblast-like synoviocyte (FLS) by neutralizing key proinflammatory cytokines. EZH2 expression is lower in PBMC and CD4+ T cells from RA patients than those from HC. EZH2 inhibition suppressed regulatory T cells (Tregs) differentiation and FOXP3 transcription, and downregulated RUNX1 and upregulated SMAD7 expression in CD4+ T cells. RA synovial fluid and fibroblast-like synoviocytes suppressed EZH2 expression in CD4+ T cells, which was partially neutralized by anti-IL17 antibody. Taken together, EZH2 in CD4+ T cells from RA patients was attenuated, which suppressed FOXP3 transcription through downregulating RUNX1 and upregulating SMAD7 in CD4+ T cells, and ultimately suppressed Tregs differentiation. IL17 in RA synovial fluid might promote downregulation of EZH2 in CD4+ T cells. Defective EZH2 in CD4+ T cells might contribute to Treg deficiency in RA.

LncRNA SNHG7 affects malignant tumor behaviors through downregulation of EZH2 in uveal melanoma cell lines.

Previous studies have demonstrated that the long non-coding RNA, small nucleolar RNA host gene 7 (SNHG7) plays an important role in several types of cancer; however, its role in the development of uveal melanoma (UM) remains unclear. The present study investigated the effect of SNHG7 on the prognosis of UM, as well as on cell proliferation, cell cycle and apoptosis of UM cell lines. Furthermore, the present study aimed to determine the molecular mechanisms underlying these effects. The association between SNHG7 and prognosis of UM was analyzed using detailed SNHG7 mRNA expression data and clinical information from The Cancer Genome Atlas database. Reverse transcription-quantitative PCR was used in order to detect the differential expression of SNHG7 in UM tissues and cell lines. Cell proliferation was detected using Cell Counting Kit-8 assays, following overexpression of SNHG7. A cell cycle assay was performed using propidium iodide/RNase staining. An apoptosis assay was performed using the Annexin-V-Fluorescein isothiocyanate apoptosis detection kit. The expression of enhancer of zeste homolog 2 (EZH2) was measured via western blotting. The results of the present study indicated that low expression of SNHG7 was associated with poor prognosis. Furthermore, increasing the expression of SNHG7 inhibited the proliferation of UM cells, suppressed cell cycle progression and promoted apoptosis. Western blot analysis results revealed that overexpression of SNHG7 downregulated EZH2 protein expression levels in UM cell lines. The results of the present study demonstrated that SNHG7 inhibited malignant transformation of UM cells by regulating EZH2 expression.

DZNep-mediated apoptosis in B-cell lymphoma is independent of the lymphoma type, EZH2 mutation status and MYC, BCL2 or BCL6 translocations.

Enhancer of zeste homolog 2 (EZH2) tri-methylates histone 3 at position lysine 27 (H3K27me3). Overexpression and gain-of-function mutations in EZH2 are regarded as oncogenic drivers in lymphoma and other malignancies due to the silencing of tumor suppressors and differentiation genes. EZH2 inhibition is sought to represent a good strategy for tumor therapy. In this study, we treated Burkitt lymphoma (BL) and diffuse large B-cell lymphoma (DLBCL) cell lines with 3-deazaneplanocin—A (DZNep), an indirect EZH2 inhibitor which possesses anticancer properties both in-vitro and in-vivo. We aimed to address the impact of the lymphoma type, EZH2 mutation status, as well as MYC, BCL2 and BCL6 translocations on the sensitivity of the lymphoma cell lines to DZNep-mediated apoptosis. We show that DZNep inhibits proliferation and induces apoptosis of these cell lines independent of the type of lymphoma, the EZH2 mutation status and the MYC, BCL2 and BCL6 rearrangement status. Furthermore, DZNep induced a much stronger apoptosis in majority of these cell lines at a lower concentration, and within a shorter period when compared with EPZ-6438, a direct EZH2 inhibitor currently in phase II clinical trials. Apoptosis induction by DZNep was both concentration-dependent and time-dependent, and was associated with the inhibition of EZH2 and subsequent downregulation of H3K27me3 in DZNep-sensitive cell lines. Although EZH2, MYC, BCL2 and BCL6 are important prognostic biomarkers for lymphomas, our study shows that they poorly influence the sensitivity of lymphoma cell lines to DZNep-mediated apoptosis.

EZH2 Regulates Intestinal Inflammation and Necroptosis Through the JNK Signaling Pathway in Intestinal Epithelial Cells.

Inflammatory bowel disease (IBD) is a common disorder of chronic intestinal inflammation that can be caused by the disruption of intestinal immune homeostasis. We aimed to evaluate the role of enhancer of zeste homolog 2 (EZH2) in the inflammatory response and explore the association between EZH2 and necroptosis in human epithelial colorectal adenocarcinoma cell lines. In both in vitro and in vivo models, expression of EZH2 in intestinal tissues was verified by histology. The expression of inflammatory cytokines in cell lines treated with EZH2 siRNA with or without stimulus was analyzed by quantitative real-time polymerase chain reaction. An intestinal necroptosis cell model was established to elucidate whether EZH2 is involved in necroptosis. Our present data indicated that EZH2 expression was decreased in in vitro and in vivo models and in patients with inflammatory bowel disease. EZH2 downregulation increased the expression of inflammatory factors, including TNF-α, IL-8, IL-17, CCL5, and CCL20 in a Caco-2 cell model. The JNK pathway was activated with the reduction of EZH2. In the necroptosis model, downregulation of EZH2 was detected with the upregulation of necroptotic markers RIP1 and RIP3. In addition, EZH2 knockdown with siRNA increased p-JNK and p-c-Jun. Our data suggest that EZH2 plays an important role in the development of intestinal inflammation and necroptosis. Hence, EZH2 could be a potential therapeutic target for IBD.

Diosgenin exhibits tumor suppressive function via down-regulation of EZH2 in pancreatic cancer cells

ABSTRACT Pancreatic cancer (PC) is one of the most aggressive and lethal malignancies worldwide. Although significant progress has been made in oncology treatment, this refractory disease is still become intractable. Natural herb product diosgenin is described to exhibit vast range of pharmacological activities in preclinical studies, including anti-cancer activities. Accumulating data demonstrated that Enhancer of zeste homolog 2 (EZH2) as an oncogenic protein is over-expressed in various human cancers, including PC. However, the underlying mechanism has not been fully understood. In this study, we aim to investigate the anti-cancer properties and molecular basis of diosgenin in PC cells. Significant inhibition of cell proliferation was observed in diosgenin treated Patu8988 and Panc-1 cells in a dose- and time-dependent manner. Apoptotic cell death and G2/M phase arrest were also induced by diosgenin treatment in PC cells. Moreover, obvious inhibition of cell migration and invasive capacities was detected in diosgenin treated PC cells. Mechanistically, the expression levels of EZH2 and its target Vimentin were reduced, and PTEN was promoted after diosgenin exposure. Our results further supported that EZH2 signaling was closely associated with the anti-tumor characteristics of diosgenin in PC cells. Therefore, inhibition of EZH2 by diosgenin could be a promising therapeutic method for PC treatment.

The Attenuation of Trophoblast Invasion Caused by the Downregulation of EZH2 Is Involved in the Pathogenesis of Human Recurrent Miscarriage

Recurrent miscarriage (RM) is currently defined as two or more losses of a clinically established intrauterine pregnancy. Despite years of research, RM continues to be a clinically frustrating challenge for patients and physicians, and its etiology remains poorly understood. Accumulating evidence has suggested that epigenetic modifications are involved in early embryogenesis, and defects in epigenetic patterning contribute to the development of RM. Here, we studied the role of enhancer of zeste homolog 2 (EZH2) in the pathogenesis of RM and found that the EZH2 expression was significantly decreased in the villi from women with RM compared with that in control villi. EZH2 promoted the invasion of trophoblast cells. Moreover, EZH2 could promote epithelial-mesenchymal transition by epigenetically silencing CDX1. Both chromatin immunoprecipitation (ChIP)-PCR and dual-luciferase report assays demonstrated that EZH2 repressed CDX1 transcription via direct binding to its promoter region and then trimethylating Histone3-Lysine27. Furthermore, we discovered that progesterone, which is used extensively in the treatment of miscarriage and RM, increased the expression of EZH2 via the extracellular signaling-regulated kinase (ERK1/2) pathway. These findings revealed that EZH2 may regulate trophoblast invasion as an epigenetic factor, suggesting that EZH2 might be a potential therapeutic target for RM.