关键词： ATF3   cholangiocarcinoma   EMT   p53 signalling pathway  

摘要： The aim of this research was to determine the underlying mechanism of activating transcription factor 3 (ATF3) on cell proliferation, invasion, migration and epithelial-mesenchymal transition (EMT). The differentially expressed mRNAs in cholangiocarcinoma (CC) and its adjacent tissues were screened by microarray analysis, and the expression of ATF3 was detected through Quantitative real time polymerase chain reaction (qRT-PCR) and Western blot. The expression of EMT markers and p53-related proteins was analysed by Western blot. Analyses using the Cell Counting Kit-8 and TUNEL were performed to assess the rate of apoptosis and cell proliferation. Scratch wound and transwell assays were performed to study cell migration and invasion. Activating transcription factor 3 was restrained in CC cell lines and tissues and inhibited EMT while activating the p53 signalling pathway. Knockdown of ATF3 promoted cell proliferation but reduced the rate of apoptosis by inhibiting p53 signalling. Cell migration and invasion can be strengthened by ATF3 through activating the p53 signalling pathway.

关键词： β-adrenergic receptors   ATF3   heart failure   PDE3A   polymorphism   transcription regulation  

摘要： Background: The phosphodiesterase 3A (PDE3A) gene encodes a PDE that regulates cardiac myocyte cyclic adenosine monophosphate (cAMP) levels and myocardial contractile function. PDE3 inhibitors (PDE3i) are used for short-term treatment of refractory heart failure (HF), but do not produce uniform long-term benefit.Objectives: The authors tested the hypothesis that drug target genetic variation could explain clinical response heterogeneity to PDE3i in HF.Methods: PDE3A promoter studies were performed in a cloned luciferase construct. In human left ventricular (LV) preparations, mRNA expression was measured by reverse transcription polymerase chain reaction, and PDE3 enzyme activity by cAMP-hydrolysis.Results: The authors identified a 29-nucleotide (nt) insertion (INS)/deletion (DEL) polymorphism in the human PDE3A gene promoter beginning 2,214 nt upstream from the PDE3A1 translation start site. Transcription factor ATF3 binds to the INS and represses cAMP-dependent promoter activity. In explanted failing LVs that were homozygous for PDE3A DEL and had been treated with PDE3i pre-cardiac transplantation, PDE3A1 mRNA abundance and microsomal PDE3 enzyme activity were increased by 1.7-fold to 1.8-fold (p < 0.05) compared with DEL homozygotes not receiving PDE3i. The basis for the selective up-regulation in PDE3A gene expression in DEL homozygotes treated with PDE3i was a cAMP response element enhancer 61 nt downstream from the INS, which was repressed by INS. The DEL homozygous genotype frequency was also enriched in patients with HF.Conclusions: A 29-nt INS/DEL polymorphism in the PDE3A promoter regulates cAMP-induced PDE3A gene expression in patients treated with PDE3i. This molecular mechanism may explain response heterogeneity to this drug class, and may inform a pharmacogenetic strategy for a more effective use of PDE3i in HF. [ABSTRACT FROM AUTHOR]

关键词： β-adrenergic receptors   ATF3   heart failure   PDE3A   polymorphism   transcription regulation  

摘要： Corresponding Author [ABSTRACT FROM AUTHOR]

关键词： Anatomy   Biochemistry   Biology and life sciences   Cardiac hypertrophy   Cardiac ventricles   Cardiology   Cardiovascular anatomy   Cardiovascular physiology   Developmental biology   Diagnostic medicine   Diagnostic radiology   DNA-binding proteins   Fibrosis   Gene expression   Gene regulation   Genetics   Heart   Imaging techniques   Immune response   Immunology   Inflammation   Magnetic resonance imaging   Medicine and health sciences   Pathology and laboratory medicine   Physiology   Proteins   Radiology and imaging   Regulatory proteins   Research and analysis methods   Research Article   Signs and symptoms   Transcription factors  

摘要： c-Jun dimerization protein (JDP2) and Activating Transcription Factor 3 (ATF3) are closely related basic leucine zipper proteins. Transgenic mice with cardiac expression of either JDP2 or ATF3 showed maladaptive remodeling and cardiac dysfunction. Surprisingly, JDP2 knockout (KO) did not protect the heart following transverse aortic constriction (TAC). Instead, the JDP2 KO mice performed worse than their wild type (WT) counterparts. To test whether the maladaptive cardiac remodeling observed in the JDP2 KO mice is due to ATF3, ATF3 was removed in the context of JDP2 deficiency, referred as double KO mice (dKO). Mice were challenged by TAC, and followed by detailed physiological, pathological and molecular analyses. dKO mice displayed no apparent differences from WT mice under unstressed condition, except a moderate better performance in dKO male mice. Importantly, following TAC the dKO hearts showed low fibrosis levels, reduced inflammatory and hypertrophic gene expression and a significantly preserved cardiac function as compared with their WT counterparts in both genders. Consistent with these data, removing ATF3 resumed p38 activation in the JDP2 KO mice which correlates with the beneficial cardiac function. Collectively, mice with JDP2 and ATF3 double deficiency had reduced maladaptive cardiac remodeling and lower hypertrophy following TAC. As such, the worsening of the cardiac outcome found in the JDP2 KO mice is due to the elevated ATF3 expression. Simultaneous suppression of both ATF3 and JDP2 activity is highly beneficial for cardiac function in health and disease.

关键词： Cancer   Cell biology  

摘要： Pancreatic ductal adenocarcinoma (PDAC) exhibits one of the worst survival rates of all cancers. While death rates show declining trends in the majority of cancers, PDAC registers rising rates. Based on the recently described crosstalk between TGF-beta 1 and Nrf2 in the PDAC development, the involvement of ATF3 and its splice variant Delta Zip2 in TGF-beta 1- and Nrf2-driven pancreatic tumorigenesis was investigated. As demonstrated here, PDAC (Panc1, T3M4) cells or premalignant H6c7 pancreatic ductal epithelial cells differentially express Delta Zip2- and ATF3, relating to stronger Nrf2 activity seen in Panc1 cells and TGF-beta 1 activity in T3M4 or H6c7 cells, respectively. Treatment with the electrophile/oxidative stress inducer tBHQ or the cytostatic drug gemcitabine strongly elevated Delta Zip2 expression in a Nrf2-dependent fashion. The differential expression of ATF3 and Delta Zip2 in response to Nrf2 and TGF-beta 1 relates to differential ATF3-gene promoter usage, giving rise of distinct splice variants. Nrf2-dependent Delta Zip2 expression confers resistance against gemcitabine-induced apoptosis, only partially relating to interference with ATF3 and its proapoptotic activity, e.g., through CHOP-expression. In fact, Delta Zip2 autonomously activates expression of cIAP anti-apoptotic proteins. Moreover, Delta Zip2 favors and ATF3 suppresses growth and clonal expansion of PDAC cells, again partially independent of each other. Using a Panc1 tumor xenograft model in SCID-beige mice, the opposite activities of ATF3 and Delta Zip2 on tumor-growth and chemoresistance were verified in vivo. Immunohistochemical analyses confirmed Delta Zip2 and Nrf2 coexpression in cancerous and PanIN structures of human PDAC and chronic pancreatitis tissues, respectively, which to some extent was reciprocal to ATF3 expression. It is concluded that depending on selective ATF3-gene promoter usage by Nrf2, the Delta Zip2 expression is induced in response to electrophile/oxidative (here

关键词： ATF3   Mitochondrial dynamics   Nerve injury   Neurodegenerative disease   Axonal transport   DRP1  

摘要： Over the last decade, mitochondrial dynamics beyond function during axon regeneration/degeneration have received attention. Axons have an effective delivery system of mitochondria shuttling between soma and axonal terminals, due to their polarized structure. The proper axonal transport of mitochondria, coordinated with mitochondrial fission/fusion and clearance, is vital for supplying high power energy in injured axons. Many researchers have studied mitochondrial dynamics using in vitro cultured cells with significant progress reported. However, the in vitro culture system is missing a physiological environment including glial cells, immune cells, and endothelial cells, whose communications are indispensable to nerve regeneration/degeneration. In line with this, the understanding of mitochondrial behavior in injured axon in vivo is necessary for promoting the physiological understanding of damaged axons and the development of a therapeutic strategy. In this review, we focus on recent insights into in vivo mitochondrial dynamics during axonal regeneration/degeneration, and introduce the advances of mouse strains to visualize mitochondria in a neuron-specific or an injury-specific manner, which are extremely useful for nerve regeneration/degeneration studies. (C) 2018 Elsevier B.V. and Japan Neuroscience Society. All rights reserved.

关键词： 黄韧带骨化   ncRNA   成骨细胞分化   miR-342-3p   ATF3  

摘要： 已有研究表明非编码RNA可参与多种生理病理过程,其在疾病的发生发展中发挥重要作用。黄韧带骨化症及骨质疏松症是常见的慢性骨科疾病,如不能得到及时的诊断和治疗,将严重影响患者的生存质量。目前,鲜有报道非编码RNA在黄韧带骨化症或骨质疏松症中的功能及机制。本研究旨在通过基因芯片或测序筛选出骨化黄韧带组织（Ossification of ligamentum flavum,OLF）中显著差异表达的m RNA、lnc RNA、mi RNA及circ RNA,并对差异RNA进行关联分析及功能探究;同时明确mi R-342-3p在成骨分化中的功能及机制,为黄韧带骨化和骨质疏松症的诊断和治疗提供新思路。方法:1.标本采集:收集术中取出的新鲜骨化的黄韧带组织和正常黄韧带组织。用液氮研磨法快速提取组织样本RNA,并通过RNA电泳和2100质量检测法进行检测。从中分别选4个合格的正常黄韧带组织和4个合格的骨化黄韧带组织的RNA进一步实验。2.筛选OLF中差异表达的m RNA:通过基因芯片分析,OLF与正常黄韧带组织（ligamentum flavum,LF）相比,筛选出差异倍数>2,p<0.05的差异m RNAs。随机挑选10个m RNAs在扩大样本中进行q PCR检测表达差异,确证芯片结果可信。进一步做差异m RNAs的GO和KEGG分析,确证这些差异m RNAs是否参与成骨分化。3.筛选OLF中差异表达的lnc RNA:通过基因芯片分析,OLF与LF相比,筛选出差异倍数>2,p<0.05的差异lnc RNAs。随机挑选10个lnc RNAs在扩大样本中进行q PCR检测表达差异,确证芯片结果可信。选择两个上调的lnc RNAs,在h MSCs向成骨分化过程中,分别敲低lnc RNA后,通过q PCR检测促成骨分化相关基因表达量的改变,并进行ALP染色,确证差异lnc RNAs在细胞成骨分化中的作用。在m RNAs芯片结果中挑选10个已知的与骨化或成骨分化相关的差异m RNAs,联合m RNAs和lnc RNAs芯片结果,筛选出与上述m RNAs共表达的差异lnc RNAs,构建共表达（conding-non-coding,CNC）网。4.筛选OLF中差异表达的mi RNA:通过基因测序技术,OLF与LF相比,筛选出差异倍数>2,p<0.05的差异mi RNAs。随机挑选10个mi RNAs在扩大样本中进行q PCR检测表达差异,确证芯片结果可信。选定一个下调的mi RNA,在h MSCs向成骨分化过程中过表达,通过q PCR检测促成骨分化相关基因的表达变化,同时进行ALP染色,确证差异mi RNA在细胞成骨分化中的作用。通过Target Scan,mi RBase和mi Randa数据库,预测差异mi RNAs的靶基因;同时与m RNAs芯片结果中差异m RNAs取交集,进一步再筛选出与mi RNAs负性差异表达关联的m RNAs。进行上述筛选出的m RNAs的GO和KEGG分析,确证其是否与骨化相关。5.筛选OLF中差异表达的circ RNA:通过基因芯片技术,OLF与LF相比,筛选出差异倍数>2,p<0.05的差异circ RNAs。随机挑选10个circ RNAs在扩大样本中进行q PCR检测表达差异,确证芯片结果可信。选择一个上调的circ RNA进行功能验证。在h MSCs向成骨分化过程中,敲低选定的circ RNA后,通过q PCR检测ALP、Col1α1、Bglap及RUNX2的表达量变化,并进行ALP染色,确证差异circ RNA在细胞成骨分化中的作用。软件预测验证的lnc RNAs、circ RNA及mi RNA是否存在关联,以此进一步构建ce RNA调节网。6.确定成骨分化过程中mi R-342-3p的表达水平:在OLF和正常黄韧带样本中,健康体检者、OLF及骨质疏松患者的血标本中,骨质疏松模型小鼠股骨组织中及细胞成骨分化诱导过程中,通过q PCR检测ALP、OCN及mi R-342-3p的水平。7.确定mi R-342-3p在细胞成骨分化中的功能:分别用mi R-342-3p抑制剂（antagomir-342-3p）或mi R-342-3p激动剂（agomir-342-3p）转染原代人骨髓间充质干细胞h MSCs,q PCR确保mi R-342-3p的干扰和过表达效果,进一步成骨诱导培养3天或6天后,提取RNA和蛋白质,通过q PCR和Western Blot检测促成骨分化相关基因的表达;诱导6天时,通过ALP染色和活性检测测定ALP的沉积和活性,茜素红染色检测钙结节的形成。同时检测mi R-342-3p在小鼠前成骨细胞MC3T3-E1向成骨细胞分化过程中的作用。8.确定mi R-342-3p的作用机制:通过mi RWal

关键词： ATF3   ferroptosis   SLC7A11   system Xc~-   metabolic stress   erastin   p53  

摘要： OBJECTIVE The amino acid antiporter system Xc;is important for the synthesis of glutathione（GSH） that functions to prevent lipid peroxidation and protect cells from nonapoptotic, iron-dependent death（ie, ferroptosis）. Inhibition of system Xc;activity by small molecules（eg, erastin） can cause a decrease in the intracellular GSH level, leading to ferroptotic cell death. While the activity of system Xc;is mainly determined by the expression level of its subunit encoded by the gene SLC7A11, the mechanism by which system Xc;is regulated during ferroptosis remains largely unknown. The goal of this study is to determine whether ATF3, a common stress sensor, can regulate system Xc;activity and promote ferroptosis. METHODS Ferroptosis was evaluated by cell viability assays and confirmed by treating cells with ferrostatin-1. Lipid peroxidation was measured by flow cytometry after cell staining with BODIPY 581/591C11. The activity of system Xc;was measured by glutamate release and GSH assay. The binding of ATF3 to the SLC7A11 promoter was determined by chromatin immunoprecipitation and DNA affinity precipitation assays. The activity of SLC7A11 promoter or mutated promoters was measured by dual luciferase reporter assays. RESULTS Overexpression of ATF3 in various cancer cells（eg, HT1080） promoted lipid peroxidation and ferroptosis induced by erastin and other ferroptosis inducers（eg, RSL3）. Conversely, knockout of ATF3 expression blocked the accumulation of lipid peroxides and prevented cells from undergoing ferroptosis. ATF3 appeared to promote ferroptosis by inhibiting system Xc;,as ATF3 expression decreased, while ATF3 knockout increased, the amount of released glutamate and the intracellular GSH level. Moreover, bypassing system Xc;by treating cells with 2-mercaptoethanol（2-ME） or N-acetylcysteine（NAC） could alleviate the effects of ATF3 on ferroptosis. Mechanistically, ATF3 strongly bound to the SLC7 A11 promoter and repressedits activity. Mutating the putative ATF3-bindin

关键词： Leiomyomas   Leiomyosarcomas   Sarcomas   Uterus   Mesenchymal tumours   MS-HRM  

摘要： Mesenchymal tumours of the corpus uteri comprise common benign lesions - leiomyomas and very rare malignant variants - sarcomas. It can be difficult to distinguish between the particular types of mesenchymal tumours pre-surgically. Primarily, leiomyomas and the very aggressive leiomyosarcomas can be easily mis-diagnosed when using only imaging devices. Therefore, a reliable non-invasive marker for these tumour types would provide greater certitude for patients that the lesion remains benign. Our collection comprises 76 native leiomyomas, an equal number of healthy myometrium samples and 49 FFPE samples of various types of sarcomas. The methylation level was assessed by MS-HRM method and we observed differences in the methylation level between healthy, benign and (semi)malignant tissues in the KLF4 and DLEC1 genes. The mean methylation levels of leiomyomas compared to myometrium and leiomyosarcomas were 70.7% vs. 6.5% vs. 39.6 % (KLF4) and 66.1% vs. 14.08% vs. 37.5% (DLEC1). The ATF3 gene was differentially methylated in leiomyomatous and myometrial tissues with 98.1% compared to 76.6%. The AUC values of the predictive logistic regression model for discrimination between leiomyomas and leiomyosarcomas based on methylation levels were 0.7829 (KLF4) and 0.7719 (DLEC1). Finally, our results suggest that there should be distinct models for the methylation events in benign leiomyomas and sarcomas, and that the KLF4 and DLEC1 genes can be considered potential methylation biomarkers for uterine leiomyomas.