SMYD2-Methylated PPARγ Facilitates Hypoxia-Induced Pulmonary Hypertension by Activating Mitophagy.

BACKGROUND: Hyperproliferation of pulmonary arterial smooth muscle cells (PASMCs) and consequent pulmonary vascular remodeling are the crucial pathological features of pulmonary hypertension (PH). Protein methylation has been shown to be critically involved in PASMC proliferation and PH, but the underlying mechanism remains largely unknown. METHODS: PH animal models were generated by treating mice/rats with chronic hypoxia for 4 weeks. SMYD2-vTg mice (vascular smooth muscle cell-specific suppressor of variegation, enhancer of zeste, trithorax and myeloid Nervy DEAF-1 domain-containing protein 2 transgenic) or wild-type rats and mice treated with LLY-507 were used to investigate the function of SMYD2 (suppressor of variegation, enhancer of zeste, trithorax and myeloid Nervy DEAF-1 domain-containing protein 2) on PH development in vivo. Primary cultured rat PASMCs with SMYD2 knockdown or overexpression were used to explore the effects of SMYD2 on proliferation and to decipher the underlying mechanism. RESULTS: We demonstrated that the expression of the lysine methyltransferase SMYD2 was upregulated in the smooth muscle cells of pulmonary arteries from patients with PH and hypoxia-exposed rats/mice and in the cytoplasm of hypoxia-induced rat PASMCs. More importantly, targeted inhibition of SMYD2 by LLY-507 significantly attenuated hypoxia-induced pulmonary vascular remodeling and PH development in both male and female rats in vivo and reduced rat PASMC hyperproliferation in vitro. In contrast, SMYD2-vTg mice exhibited more severe PH phenotypes and related pathological changes than nontransgenic mice after 4 weeks of chronic hypoxia treatment. Furthermore, SMYD2 overexpression promoted, while SMYD2 knockdown suppressed, the proliferation of rat PASMCs by affecting the cell cycle checkpoint between S and G2 phases. Mechanistically, we revealed that SMYD2 directly interacted with and monomethylated PPARγ (peroxisome proliferator-activated receptor gamma) to inhibit the nuclear translocation and transcriptional activity of PPARγ, which further promoted mitophagy to facilitate PASMC proliferation and PH development. Furthermore, rosiglitazone, a PPARγ agonist, largely abolished the detrimental effects of SMYD2 overexpression on PASMC proliferation and PH. CONCLUSIONS: Our results demonstrated that SMYD2 monomethylates nonhistone PPARγ and inhibits its nuclear translocation and activation to accelerate PASMC proliferation and PH by triggering mitophagy, indicating that targeting SMYD2 or activating PPARγ are potential strategies for the prevention of PH.

Regulated cell death in myocardial ischemia-reperfusion injury.

Myocardial ischemia-reperfusion (I/R) injury most commonly occurs in coronary artery disease when prompt reperfusion is used to salvage the ischemic myocardium. Cardiomyocyte death is a significant component of myocardial I/R injury and its mechanism was previously thought to be limited to apoptosis and necrosis. With the discovery of novel types of cell death, ferroptosis, necroptosis, and pyroptosis have been shown to be involved in myocardial I/R. These new forms of regulated cell death cause cardiomyocyte loss and exacerbate I/R injury by affecting reactive oxygen species (ROS) generation, calcium stress, and inflammatory cascades, subsequently mediating adverse remodeling, cardiac dysfunction, and heart failure. Herein, we review the roles of ferroptosis, necroptosis, and pyroptosis in myocardial I/R and discuss their contribution to pathology.

The functions of SET domain bifurcated histone lysine methyltransferase 1 (SETDB1) in biological process and disease.

Histone methyltransferase SETDB1 (SET domain bifurcated histone lysine methyltransferase 1, also known as ESET or KMT1E) is known to be involved in the deposition of the di- and tri-methyl marks on H3K9 (H3K9me2 and H3K9me3), which are associated with transcription repression. SETDB1 exerts an essential role in the silencing of endogenous retroviruses (ERVs) in embryonic stem cells (mESCs) by tri-methylating H3K9 (H3K9me3) and interacting with DNA methyltransferases (DNMTs). Additionally, SETDB1 is engaged in regulating multiple biological processes and diseases, such as ageing, tumors, and inflammatory bowel disease (IBD), by methylating both histones and non-histone proteins. In this review, we provide an overview of the complex biology of SETDB1, review the upstream regulatory mechanisms of SETDB1 and its partners, discuss the functions and molecular mechanisms of SETDB1 in cell fate determination and stem cell, as well as in tumors and other diseases. Finally, we discuss the current challenges and prospects of targeting SETDB1 for the treatment of different diseases, and we also suggest some future research directions in the field of SETDB1 research.

BRD4770 inhibits vascular smooth muscle cell proliferation via SUV39H2, but not EHMT2 to protect against neointima formation.

The behavior of vascular smooth muscle cells (VSMCs) contributes to the formation of neointima. We previously found that EHMT2 suppressed autophagy activation in VSMCs. BRD4770, an inhibitor of EHMT2/G9a, plays a critical role in several kinds of cancers. However, whether and how BRD4770 regulates the behavior of VSMCs remain unknown. In this study, we evaluate the cellular effect of BRD4770 on VSMCs by series of experiments in vivo and ex vivo. We demonstrated that BRD4770 inhibited VSMCs' growth by blockage in G2/M phase in VSMCs. Moreover, our results demonstrated that the inhibition of proliferation was independent on autophagy or EHMT2 suppression which we previous reported. Mechanistically, BRD4770 exhibited an off-target effect from EHMT2 and our further study reveal that the proliferation inhibitory effect by BRD4770 was associated with suppressing on SUV39H2/KTM1B. In vivo, BRD4770 was also verified to rescue VIH. Thus, BRD4770 function as a crucial negative regulator of VSMC proliferation via SUV39H2 and G2/M cell cycle arrest and BRD4770 could be a molecule for the therapy of vascular restenosis.

A Novel Serum Biomarker Model to Discriminate Aortic Dissection from Coronary Artery Disease.

Background: The misdiagnosis of aortic dissection (AD) can lead to a catastrophic prognosis. There is currently a lack of stable serological indicators with excellent efficacy for the differential diagnosis of AD and coronary artery disease (CAD). A recent study has shown an association between AD and iron metabolism. Thus, we investigated whether iron metabolism could discriminate AD from CAD. Methods: This retrospective and multicenter cross-sectional study investigated the efficacy of biomarkers of iron metabolism for the differential diagnosis of AD. We collected biomarkers of iron metabolism, liver function, kidney function, and other biochemistry test, and further, logistic regression analysis was applied. Results: Between Oct. 8, 2020, and Mar. 1, 2021, we recruited 521 patients diagnosed with AD, CAD, and other cardiovascular diseases (OCDs) with the main symptoms of chest and back pain and assigned them to discovery set (n = 330) or validation set (n = 191). We found that six serum biomarkers, including serum iron, low-density lipoprotein, uric acid, transferrin, high-density lipoprotein, and estimated glomerular filtration rate, can serve as a novel comprehensive indicator (named FLUTHE) for the differential diagnosis of AD and CAD with a sensitivity of 0.954 and specificity of 0.905 to differentially diagnose AD and CAD more than 72 h past symptom onset. Conclusion: Our findings provide insight into the role of iron metabolism in diagnosing and distinguishing AD, which might in the future be a key component in AD diagnosis. Furthermore, we establish a novel model named "FLUTHE" with higher efficiency, safety, and economy, especially for patients with chest pain for more than 72 h.

JIB-04, a histone demethylase Jumonji C domain inhibitor, regulates phenotypic switching of vascular smooth muscle cells.

BACKGROUND: Vascular smooth muscle cell (VSMC) phenotype switching is critical for neointima formation, which is the major cause of restenosis after stenting or coronary artery bypass grafting. However, the epigenetic mechanisms regulating phenotype switching of VSMCs, especially histone methylation, are not well understood. As a main component of histone lysine demethylases, Jumonji demethylases might be involved in VSMC phenotype switching and neointima formation. METHODS AND RESULTS: A mouse carotid injury model and VSMC proliferation model were constructed to investigate the relationship between histone methylation of H3K36 (downstream target molecule of Jumonji demethylase) and neointima formation. We found that the methylation levels of H3K36 negatively correlated with VSMC proliferation and neointima formation. Next, we revealed that JIB-04 (a pan-inhibitor of the Jumonji demethylase superfamily) could increase the methylation levels of H3K36. Furthermore, we found that JIB-04 obviously inhibited HASMC proliferation, and a cell cycle assay showed that JIB-04 caused G2/M phase arrest in HASMCs by inhibiting the phosphorylation of RB and CDC2 and promoting the phosphorylation of CHK1. Moreover, JIB-04 inhibited the expression of MMP2 to suppress the migration of HASMCs and repressed the expression of contraction-related genes. RNA sequencing analysis showed that the biological processes associated with the cell cycle and autophagy were enriched by using Gene Ontology analysis after HASMCs were treated with JIB-04. Furthermore, we demonstrated that JIB-04 impairs autophagic flux by downregulating STX17 and RAB7 expression to inhibit the fusion of autophagosomes and lysosomes. CONCLUSION: JIB-04 suppresses the proliferation, migration, and contractile phenotype of HASMCs by inhibiting autophagic flux, which indicates that JIB-04 is a promising reagent for the treatment of neointima formation.

Targeting Ferroptosis as a Novel Approach to Alleviate Aortic Dissection.

A variety of programmed cell death types have been shown to participate in the loss of smooth muscle cells (SMCs) during the development of aortic dissection (AD), but it is still largely unclear whether ferroptosis is involved in the development of AD. In the present study, we found that the expression of key ferroptosis regulatory proteins, solute carrier family 7 member 11 (SLC7A11), ferroptosis suppressor protein 1 (FSP1) and glutathione peroxidase 4 (GPX4) were downregulated in aortas of Stanford type A AD (TAAD) patients, and liproxstatin-1, a specific inhibitor of ferroptosis, obviously abolished the ß-aminopropionitrile (BAPN)-induced development and rupture of AD in mice. Furthermore, the expression of methyltransferase-like 3 (METTL3), a major methyltransferase of RNA m6A, was remarkably upregulated in the aortas of TAAD patients, and the protein levels of METTL3 were negatively correlated with SLC7A11 and FSP1 levels in human aortas. Overexpression of METTL3 in human aortic SMCs (HASMCs) inhibited, while METTL3 knockdown promoted SLC7A11 and FSP1 expression. More importantly, overexpression of METTL3 facilitated imidazole ketone erastin- and cystine deprivation-induced ferroptosis, while knockdown of METTL3 repressed ferroptosis of HASMCs. Overexpression of either SLC7A11 or FSP1 largely abrogated the effect of METTL3 on HASMC ferroptosis. Therefore, we have revealed that ferroptosis is a critical cause of AD in both humans and mice and that METTL3 promotes ferroptosis of HASMCs by inhibiting the expression of SLC7A11 and FSP1. Thus, targeting ferroptosis or m6A RNA methylation is a potential novel strategy for the treatment of AD.

Comprehensive analysis identified a reduction in ATP1A2 mediated by ARID3A in abdominal aortic aneurysm.

Abdominal aortic aneurysm (AAA) is characterized by abdominal aorta dilatation and progressive structural impairment and is usually an asymptomatic and potentially lethal disease with a risk of rupture. To investigate the underlying mechanisms of AAA initiation and progression, seven AAA datasets related to human and mice were downloaded from the GEO database and reanalysed in the present study. After comprehensive bioinformatics analysis, we identified the enriched pathways associated with inflammation responses, vascular smooth muscle cell (VSMC) phenotype switching and cytokine secretion in AAA. Most importantly, we identified ATPase Na+ /K+ transporting subunit alpha 2 (ATP1A2) as a key gene that was significantly decreased in AAA samples of both human and mice; meanwhile, its reduction mainly occurred in VSMCs of the aorta; this finding was validated by immunostaining and Western blot in human and mouse AAA samples. Furthermore, we explored the potential upstream transcription factors (TFs) that regulate ATP1A2 expression. We found that the TF AT-rich interaction domain 3A (ARID3A) bound the promoter of ATP1A2 to suppress its expression. Our present study identified the ARID3A-ATP1A2 axis as a novel pathway in the pathological processes of AAA, further elucidating the molecular mechanism of AAA and providing potential therapeutic targets for AAA.

BRD4770 functions as a novel ferroptosis inhibitor to protect against aortic dissection.

Smooth muscle cell (SMC) loss is the characteristic feature in the pathogenesis of aortic dissection (AD), and ferroptosis is a novel iron-dependent regulated cell death driven by the excessive lipid peroxidation accumulation. However, whether targeting ferroptosis is an effective approach for SMC loss and AD treatment remains unclear. Here, we found that the iron level, ferroptosis-related molecules TFR, HOMX1, ferritin and the lipid peroxidation product 4-hydroxynonenal were increased in the aorta of AD. Then, we screened several inhibitors of histone methyltransferases and found that BRD4770 had a protective effect on cystine deprivation-, imidazole ketone erastin- or RSL3-induced ferroptosis of SMCs. The classic ferroptosis pathways, System Xc--GPX4, FSP1-CoQ10 and GCH1-BH4 pathways which were inhibited by ferroptosis inducers, were re-activated by BRD4770 via inhibiting mono-, di- and tri- methylated histone H3 at lysine 9 (H3K9me1/2/3). RNA-sequencing analysis revealed that there was a positive feedback regulation between ferroptosis and inflammatory response, and BRD4770 can reverse the effects of inflammation activation on ferroptosis. More importantly, treatment with BRD4770 attenuated aortic dilation and decreased morbidity and mortality in a ß-Aminopropionitrile monofumarate-induced mouse AD model via inhibiting the inflammatory response, lipid peroxidation and ferroptosis. Taken together, our findings demonstrate that ferroptosis is a novel and critical pathological mechanism that is involved in SMC loss and AD development. BRD4770 is a novel ferroptosis inhibitor and has equivalent protective effect to Ferrostatin-1 at the optimal concentration. Translating insights into the anti-ferroptosis effects of BRD4770 may reveal a potential therapeutic approach for targeting SMC ferroptosis in AD.

Upregulation of IRF9 Contributes to Pulmonary Artery Smooth Muscle Cell Proliferation During Pulmonary Arterial Hypertension.

Abnormal proliferation of pulmonary artery smooth muscle cells (PASMCs) is a critical pathological feature in the pathogenesis of pulmonary arterial hypertension (PAH), but the regulatory mechanisms remain largely unknown. Herein, we demonstrated that interferon regulatory factor 9 (IRF9) accelerated PASMCs proliferation by regulating Prohibitin 1 (PHB1) expression and the AKT-GSK3ß signaling pathway. Compared with control groups, the rats treated with chronic hypoxia (CH), monocrotaline (MCT) or sugen5416 combined with chronic hypoxia (SuHx), and mice challenged with CH had significantly thickened pulmonary arterioles and hyperproliferative PASMCs. More importantly, the protein level of IRF9 was found to be elevated in the thickened medial wall of the pulmonary arterioles in all of these PAH models. Notably, overexpression of IRF9 significantly promoted the proliferation of rat and human PASMCs, as evidenced by increased cell counts, EdU-positive cells and upregulated biomarkers of cell proliferation. In contrast, knockdown of IRF9 suppressed the proliferation of rat and human PASMCs. Mechanistically, IRF9 directly restrained PHB1 expression and interacted with AKT to inhibit the phosphorylation of AKT at thr308 site, which finally led to mitochondrial dysfunction and PASMC proliferation. Unsurprisingly, MK2206, a specific inhibitor of AKT, partially reversed the PASMC proliferation inhibited by IRF9 knockdown. Thus, our results suggested that elevation of IRF9 facilitates PASMC proliferation by regulating PHB1 expression and AKT signaling pathway to affect mitochondrial function during the development of PAH, which indicated that targeting IRF9 may serve as a novel strategy to delay the pathological progression of PAH.

Downregulation of Filamin a Expression in the Aorta Is Correlated With Aortic Dissection.

Filamins (FLNs) are actin cross-linking proteins, and as scaffolding proteins, FLNs are closely associated with the stabilization of the cytoskeleton. Nevertheless, the biological importance of FLNs in aortic dissection (AD) has not been well-elucidated. In this study, we first reanalyzed datasets downloaded from the Gene Expression Omnibus (GEO) database, and we found that in addition to the extracellular matrix, the actin cytoskeleton is a key structure associated with AD. Given that FLNs are involved in remodeling the cytoskeleton to affect cellular functions, we measured their expression levels in the aortas of patients with Stanford type A AD (TAAD). Our results showed that the mRNA and protein levels of FLNA were consistently decreased in dissected aortas of both humans and mice, while the FLNB protein level was upregulated despite decreased FLNB mRNA levels, and comparable expression levels of FLNC were observed between groups. Furthermore, the immunohistochemistry results demonstrated that FLNA was highly expressed in smooth muscle cells (SMCs) of aorta in non-AD samples, and downregulated in the medial layer of the dissected aortas of humans and mice. Moreover, we revealed that FOS and JUN, forming a dimeric transcription factor called AP-1 (activating protein-1), were positively correlated with the expression of FLNA in aorta. Either overexpression of FOS or JUN alone, or overexpression of FOS and JUN together, facilitated the expression of FLNA in primary cultured human aortic SMCs. In the present study, we not only detected the expression pattern of FLNs in aortas of humans and mice with or without AD, but we also found that the expression of FLNA in the AD samples was significantly reduced and that AP-1 might regulate the expression of FLNA. Our findings will contribute to the elucidation of the pathological mechanisms of AD and provide potential therapeutic targets for AD.

Posttranslational Modifications in Ferroptosis.

Ferroptosis was first coined in 2012 to describe the form of regulated cell death (RCD) characterized by iron-dependent lipid peroxidation. To date, ferroptosis has been implicated in many diseases, such as carcinogenesis, degenerative diseases (e.g., Huntington's, Alzheimer's, and Parkinson's diseases), ischemia-reperfusion injury, and cardiovascular diseases. Previous studies have identified numerous targets involved in ferroptosis; for example, acyl-CoA synthetase long-chain family member 4 (ACSL4) and p53 induce while glutathione peroxidase 4 (GPX4) and apoptosis-inducing factor mitochondria-associated 2 (AIFM2, also known as FSP1) inhibit ferroptosis. At least three major pathways (the glutathione-GPX4, FSP1-coenzyme Q10 (CoQ10), and GTP cyclohydrolase-1- (GCH1-) tetrahydrobiopterin (BH4) pathways) have been identified to participate in ferroptosis regulation. Recent advances have also highlighted the crucial roles of posttranslational modifications (PTMs) of proteins in ferroptosis. Here, we summarize the recently discovered knowledge regarding the mechanisms underlying ferroptosis, particularly the roles of PTMs in ferroptosis regulation.

EHMT2/G9a Inhibits Aortic Smooth Muscle Cell Death by Suppressing Autophagy Activation.

Although EHMT2 (also known as G9a) plays a critical role in several kinds of cancers and cardiac remodeling, its function in vascular smooth muscle cells (VSMCs) remains unknown. In the present study, we revealed a novel function of EHMT2 in regulating autophagic cell death (ACD) of VSMC. Inhibition of EHMT2 by BIX01294 or knockdown of EHMT2 resulted in reduced VSMC numbers which were independent of proliferation and apoptosis. Interestingly, EHMT2 protein levels were significantly decreased in VSMCs treated with autophagic inducers. Moreover, more autophagic vacuoles and accumulated LC3II were detected in VSMCs treated with BIX01294 or lenti-shEHMT2 than their counterparts. Furthermore, we found that EHMT2 inhibited the ACD of VSMCs by suppressing autophagosome formation. Mechanistically, the pro-autophagic effect elicited by EHMT2 inhibition was associated with SQSTM1 and BECN1 overexpression. Moreover, these detrimental effects were largely nullified by SQSTM1 or BECN1 knockdown. More importantly, similar results were observed in primary human aortic VSMCs. Overall, these findings suggest that EHMT2 functions as a crucial negative regulator of ACD via decreasing SQSTM1 or BECN1 expression and that EHMT2 could be a potent therapeutic target for cardiovascular diseases (e.g., aortic dissection).

Histone methylation and vascular biology.

The vasculature not only transports oxygenated blood, metabolites, and waste products but also serves as a conduit for hormonal communication between distant tissues. Therefore, it is important to maintain homeostasis within the vasculature. Recent studies have greatly expanded our understanding of the regulation of vasculature development and vascular-related diseases at the epigenetic level, including by protein posttranslational modifications, DNA methylation, and noncoding RNAs. Integrating epigenetic mechanisms into the pathophysiologic conceptualization of complex and multifactorial vascular-related diseases may provide promising therapeutic approaches. Several reviews have presented detailed discussions of epigenetic mechanisms not including histone methylation in vascular biology. In this review, we primarily discuss histone methylation in vascular development and maturity, and in vascular diseases.

Disturbed energy and amino acid metabolism with their diagnostic potential in mitral valve disease revealed by untargeted plasma metabolic profiling.

INTRODUCTION: Mitral valve disease (MVD), including mitral valve regurgitation (MR) and mitral valve stenosis (MS), is a chronic and progressive cardiac malady. However, the metabolic alterations in MVD is not well-understood till now. The current gold standard diagnostic test, transthoracic echocardiography, has limitations on high-throughput measurement and lacks molecular information for early diagnosis of the disease. OBJECTIVE: The present study aimed to investigate the biochemical alterations and to explore their diagnostic potential for MVD. METHODS: Plasma metabolic profile derangements and their diagnostic potential were non-invasively explored in 34 MR and 20 MS patients against their corresponding controls, using high-throughput NMR-based untargeted metabolomics. RESULTS: Eighteen differential metabolites were identified for MR and MS patients respectively, on the basis of multivariate and univariate data analysis, which were mainly involved in energy metabolism, amino acid metabolism, calcium metabolism and inflammation. These differential metabolites, notably the significantly down-regulated formate and lactate, showed high diagnostic potential for MVD by using Spearman's rank-order correlation analysis and ROC analysis. CONCLUSIONS: To the best of our knowledge, the present study is the first one that explores the metabolic derangements and their diagnostic values in MVD patients using metabolomics. The findings indicated that metabolic disturbance occurred in MVD patients, with plasma formate and lactate emerged as important candidate biomarkers for MVD.

HDAC6 is associated with the formation of aortic dissection in human.

BACKGROUND: The pathological features of aortic dissection (AD) include vascular smooth muscle cell (VSMC) loss, elastic fiber fraction, and inflammatory responses in the aorta. However, little is known about the post-translational modification mechanisms responsible for these biological processes. METHODS: A total of 72 aorta samples, used for protein detection, were collected from 36 coronary artery disease (CAD, served as the control) patients and 36 type A AD (TAAD) patients. Chromatin immunoprecipitation (ChIP)-PCR was used to identify the genes regulated by H3K23ac, and tubastatin A, an inhibitor of HDAC6, was utilized to clarify the downstream mechanisms regulated by HDAC6. RESULTS: We found that the protein level of histone deacetylase HDAC6 was reduced in the aortas of patients suffering from TAAD and that the protein levels of H4K12ac, and H3K23ac significantly increased, while H3K18ac, H4K8ac, and H4K5ac dramatically decreased when compared with CAD patients. Although H3K23ac, H3K18ac, and H4K8ac increased in the human VSMCs after treatment with the HDAC6 inhibitor tubastatin A, only H3K23ac showed the same results in human tissues. Notably, the results of ChIP-PCR demonstrated that H3K23ac was enriched in extracellular matrix (ECM)-related genes, including Col1A2, Col3A1, CTGF, POSTN, MMP2, TIMP2, and ACTA2, in the aortic samples of TAAD patients. In addition, our results showed that HDAC6 regulates H4K20me2 and p-MEK1/2 in the pathological process of TAAD. CONCLUSIONS: These results indicate that HDAC6 is involved in human TAAD formation by regulating H3K23ac, H4K20me2 and p-MEK1/2, thus, providing a strategy for the treatment of TAAD by targeting protein post-translational modifications (PTMs), chiefly histone PTMs.

EZH2 inhibits autophagic cell death of aortic vascular smooth muscle cells to affect aortic dissection.

Enhancer of zeste homolog 2 (EZH2), a methyltransferase that di- and tri-methylates lysine-27 of histone H3, largely functions as a transcriptional repressor, and plays a critical role in various kinds of cancers. Here we report a novel function of EZH2 in regulating autophagic cell death (ACD) of vascular smooth muscle cells (VSMCs) that affect aortic dissection (AD). Inhibition of EZH2 activity by UNC1999 or knockdown EZH2 resulted in VSMC loss, while overexpression of EZH2 facilitated VSMC growth, and these effects of EZH2 on VSMCs were independent of proliferation and apoptosis. Interestingly, more autophagic vacuoles and increased LC3II protein levels were identified in VSMCs with EZH2 inhibition or deficiency. Moreover, when compared with counterparts, chloroquine alone, or chloroquine with rapamycin treatment led to more LC3II accumulation in EZH2 inhibited or knockdown VSMCs, which indicated that EZH2 negatively regulated autophagosome formation. In conjunction to this, ATG5 and ATG7 protein levels were remarkably increased in EZH2 inhibited or deficient VSMCs, and ATG5 or ATG7 knockdown virtually rescued VSMC loss induced by EZH2 inhibition or knockdown. In addition, we found that the MEK-ERK1/2 signaling pathway, but not AMPKα, mTOR, or AKT pathway, is responsible for the impact of EZH2 on ACD of VSMCs. Additionally, the adverse effects of EZH2 inhibition or knockdown on VSMCs were largely reversed by PD98059, an inhibitor of MEK1. More importantly, decreased EZH2 expression levels in the aortic wall of patients with AD indicated its contribution to VSMC loss and AD occurrence. Overall, these findings revealed that EZH2 affects ACD of VSMCs and the pathologic process of AD via regulating ATG5 and ATG7 expression and MEK-ERK1/2 signaling. Our hitherto unrecognized findings indicate that EZH2 activation has therapeutic or preventive potential for AD.

The Histone Methyltransferase Mixed Lineage Leukemia (MLL) 3 May Play a Potential Role on Clinical Dilated Cardiomyopathy.

Histone modifications play a critical role in the pathological processes of dilated cardiomyopathy (DCM). While the role and expression pattern of histone methyltransferases (HMTs), especially mixed lineage leukemia (MLL) families on DCM are unclear. To this end, twelve normal and fifteen DCM heart samples were included in the present study. A murine cardiac remodelling model was induced by transverse aortic constriction (TAC). Real-time PCR was performed to detect the expression levels of MLL families in the mouse and human left ventricles. The mRNA level of MLL3 was significantly increased in the mouse hearts treated by TAC surgery. Compared with normal hearts, higher mRNA and protein level of MLL3 was detected in the DCM hearts, and its expression level was closely associated with left ventricular end systolic diameter (LVEDD) and left ventricular ejection fraction (LVEF). However, the expression level of other MLL families (MLL, MLL2, MLL4, MLL5, SETD1A, and SETD1B) had no obvious change between control and DCM hearts or remodeled mouse hearts. Furthermore, the di-methylated histone H3 lysine 4 (H3K4me2) but not H3K4me3 was significantly increased in the DCM hearts. The protein levels of Smad3, GATA4, EGR1, which might regulate by MLL3, were remarkably elevated in the DCM hearts. Our hitherto unrecognized findings indicate that MLL3 has a potential role on pathological processes of DCM via regulating H3K4me2 and the expression of Smad3, GATA4, and EGR1.