关键词： NITRIC-OXIDE DONOR   ACTIVATING TRANSCRIPTION FACTOR-3   CANCER-CELL-MIGRATION   JS-K   ACID SPHINGOMYELINASE   TISSUE INHIBITORS   CYSTEINE SWITCH   APOPTOSIS   PROLIFERATION   METASTASIS  

摘要： Glioblastoma is associated with poor survival and a high recurrence rate in patients due to inevitable uncontrolled infiltrative tumor growth. The elucidation of the molecular mechanisms may offer opportunities to prevent relapses. In this study we investigated the role of the activating transcription factor 3 (ATF3) in migration of GBM cells in vitro. RNA microarray revealed that gene expression of ATF3 is induced by a variety of chemotherapeutics and experimental agents such as the nitric oxide donor JS-K (O2-(2,4-dinitrophenyl) 1-[(4-ethoxycarbonyl)piperazin-1-yl]diazen-1-ium-1,2-diolate). We found NF kappa B and STAT3 to be downstream targets inhibited by overexpression of ATF3. We demonstrate that ATF3 is directly involved in the regulation of matrix metalloproteinase expression and activation. Overexpression of ATF3 therefore leads to a significantly reduced migration capacity and induction of tissue inhibitors of matrix metalloproteinases. Our study for the first time identifies ATF3 as a potential novel therapeutic target in glioblastoma.

关键词： 结缔组织病   肺间质病变   吡非尼酮   TGF-β1   ATF3  

摘要： 研究背景弥漫性结缔组织病(connective tissue diseases,CTDs)是一类由自身免疫异常所致机体损伤的疾病总称,这类疾病几乎可累及到机体的每个器官,肺脏是最常受累的器官之一。其中,肺间质病变(Interstitial lung disease,ILD)是危害最大的并发症,可导致肺通气功能受损甚至呼吸功能衰竭,严重影响患者寿命,迄今为止,CTD-ILD的发病机制尚未完全阐明,缺乏根治性治疗方法。因此,本文拟通过研究新型药物吡非尼酮在体内及体外实验及临床治疗中的抗纤维作用,阐明ILD的部分发生机制及治疗对策,为肺间质病变治疗提供新的证据和思路。研究目的1.观察吡非尼酮治疗CTD-ILD的临床疗效和不良反应。2.了解吡非尼酮对博来霉素诱导肺纤维化小鼠的抗纤维化作用。3.阐明吡非尼酮对HLFs增殖、迁移侵袭、分泌功能和表型转化的影响及其机制。研究方法纳入确诊为CTD-ILD的患者,根据患者本人意愿用或不用吡非尼酮进行12个月治疗观察。在0月、3月、6月和12月时随访进行实验室检查、肺功能测试、6分钟步行距离和HRCT评分,吡非尼酮用药组(PFD)和对照组(CTR)间比较采用协方差分析,其中各组间不同时间点的比较采用重复测量数据的方差分析。8周龄雄性C57BL/6小鼠建立博来霉素诱导鼠肺纤维化模型,采用直视下气管内注射博来霉素或生理盐水,分别给予吡非尼酮或相应体积的溶剂灌胃治疗,均在第28天处死后取肺脏组织,随后进行H&E染色、Masson染色和组织免疫荧光染色观察肺组织病理改变和α-SMA、ATF3的表达。手术获取正常肺组织(经病理证实)进行分离原代人肺成纤维细胞(HLFs),CT引导下经皮肺穿刺活检获取CTD-ILD肺脏病变组织。MTT法检测细胞存活率,EdU法检测细胞的增殖能力,Transwell实验检测细胞的迁移和侵袭能力,细胞免疫荧光检测α-SMA和ATF3,免疫印迹法检测α-SMA、ATF3和p-Smad3的蛋白水平,实时定量PCR检测α-SMA、ATF3和Ⅰ、Ⅲ型胶原蛋白的mRNA表达水平。应用慢病毒载体携带ATF3shRNA或对照shRNA感染体外培养的HLFs并进行验证。研究结果PFD组与CTR组相比,PFD组患者在第12月时FVC、TLC和DLco均得到显著改善(p值分别为0.044、0.017和0.001),而在第6月DLco明显增加(p值为0.033)。在随访期间与初始时间点相比,PFD组TLC在第3月即开始有明显改善,随后维持相同水平(p值分别为0.046、0.030和0.056),FVC和DLco在第6月开始有明显改善(p<0.05)。用吡非尼酮治疗3月后PFD组影像炎症评分均明显低于对照组(p值分别为0.021、0.010和0.036)。在随访期间PFD组炎症评分逐渐降低,而对照组则未观察到这种变化。吡非尼酮用药期间,患者最常出现的不良反应为乏力、呼吸困难、咳嗽、皮疹和消化不良,在临床对症处理下可减轻或消失。在博来霉素诱导小鼠肺间质纤维化模型中,预灌胃吡非尼酮可减少小鼠肺部结构破坏程度,降低胶原沉积,改善肺部纤维化。造模组小鼠肺组织中α-SMA的表达明显高于正常组和吡非尼酮干预组小鼠。CTD-ILD肺组织和造模组小鼠肺组织ATF3的表达明显高于正常组,然而吡非尼酮能降低ATF3在肺组织的表达。在TGF-β1诱导下HLFs的增殖、迁移侵袭能力和分泌功能能力显著增强,p-Smad3和ATF3的蛋白和mRNA表达水平明显高于对照组。吡非尼酮预处理HLFs能有效地抑制TGF-β1诱导的增殖、迁移侵袭能力、分泌功能和运动功能,下调p-Smad3和ATF3的蛋白和mRNA表达水平。ATF3敲低的HLFs其对TGF-β1刺激后的增殖、迁移侵袭能力和α-SMA和p-Smad3的蛋白水平均下降,其反应能力均明显弱于正常的HLFs。研究结论吡非尼酮能改善CTD-ILD患者肺功能结果和影像学评分,减少博来霉素肺纤维化鼠模型肺结构破坏、胶原沉积和纤维化程度,其治疗作用可能是通过抑制ATF3/Smad3通路从而减少HLFs表型转换来实现的。

关键词： 尿道下裂   WT1蛋白   ATF3基因   细胞病理学  

摘要： 目的：　　研究WT1(Wilms tumor1)基因，ATF3（Activating Transcription Factor3）基因在儿童尿道下裂包皮中的表达及它们与尿道下裂严重程度的联系。　　方法：　　尿道下裂组22例（I°，II°，III°分别为11例，7例，4例）的包皮组织来自在温州医科大学附属第二医院行“尿道成形术”的患儿，以及对照组22例非尿道下裂（包括10例包茎，5例包皮过长，7例隐匿阴茎）分别在我院儿童外科行“包皮环切术”，“阴茎延长术”的患儿。　　结果：　　WT1蛋白在尿道下裂患儿中的表达显著高于对照组（包茎，包皮过长，隐匿阴茎）的表达，I°、II°、III°的平均光密度值分别为（1.23E-03±5.11E-05）、（1.97E-03±4.46E-05）、（3.32E-03±2.63E-05），且两组差异具有统计学意义（P＜0.05），而且阳性表达位于尿道下裂腹侧包皮基底层的细胞核中。ATF3蛋白在重度尿道下裂患儿的表达高于在对照组（包茎，包皮过长，隐匿阴茎）的表达，I°、II°、III°的平均光密度值分别为（6.51E-06±3.21E-05）、（1.30E-03±4.51E-05）、（2.72E-03±2.23E-05），两组之间差别具有统计学意义（P＜0.05），尿道下裂腹侧包皮基底层的细胞核中能看到棕褐色颗粒。而对照组未见明显阳性表达结果。　　结论：　　WT1蛋白在尿道下裂患儿腹侧包皮组织中的表达明显上调，提示WT1基因的过表达可能与人类尿道下裂严重程度存在相关性。ATF3蛋白在II°、III°尿道下裂患儿中的表达量显著高于在对照组中的表达量，I°尿道下裂与对照组之间无明显差异（P＞0.05），病例组之间ATF3蛋白的表达量随着病情的加重，表达量明显上升。

关键词： Apoptosis   Activating Transcription Factor 3   Bcl-2   Colorectal Cancer   Vitex rotundifolia L   ACTIVATING TRANSCRIPTION FACTOR-3   ANTICANCER ACTIVITY   ATF3 EXPRESSION   LINOLEIC-ACID   UP-REGULATION   PATHOGENESIS   INHIBITION   DITERPENES   CASTICIN   NAG-1  

摘要： Fruit from Vitex rotundifolia L. (VF) has been reported to initiate apoptosis in human colorectal cancer cells through the accumulation of reactive oxygen species. Since various regulatory factors are involved in the apoptotic pathway, further study of the potential mechanisms of VF associated with the induction of apoptosis may be important despite the fact that the molecular target of VF for apoptosis has already been elucidated. In this study, we showed a new potential mechanism for the relationship between VF-mediated ATF3 expression and apoptosis to better understand the apoptotic mechanism of VF in human colorectal cancer cells. VF reduced the cell viability and induced apoptosis in human colorectal cancer cells. VF treatment increased both the protein and mRNA level of ATF3 and upregulated ATF3 promoter activity. The cis-element responsible for ATF3 transcriptional activation by VF was CREB which is located between -147 to -85 of ATF3 promoter. Inhibitions of ERK1/2, p38, JNK and GSK3 beta blocked VF-mediated ATF3 expression. ATF3 knockdown by ATF3 siRNA attenuated the cleavage of PARP by VF, while ATF3 overexpression increased VF-mediated cleaved PARP. ATF3 knockdown also attenuated VF-mediated cell viability and cell death. In addition, VF downregulated Bcl-2 expression at both protein and mRNA level. ATF3 knockdown by ATF3 siRNA blocked VF-mediated downregulation of Bcl-2. In conclusion, VF may activate ATF3 expression through transcriptional regulation and subsequently suppress Bcl-2 expression as an anti-apoptotic protein, which may result in the induction of apoptosis in human colorectal cancer cells.

关键词： Activating transcription factor 3   Early growth response protein 1   HNSCC   Human mini-organ culture model   Laryngopharyngeal reflux   SQUAMOUS-CELL CARCINOMA   ACTIVATING TRANSCRIPTION FACTOR-3   INDUCED APOPTOSIS   CANCER   ATF3   EXPRESSION   REFLUX   GENE   HEAD   PATHWAY  

摘要： Background/Aims: Activating transcription factor 3 (ATF3) and early growth response protein 1 (EGR1) are reported to interact, but their use as prognostic factors in cancer is discussed controversially. Methods: We measured ATF3 and EGR1 gene expression changes in human mini-organ cultures (MOCs) of healthy nasal epithelia, UM-SCC-22B, and FADU DD cells after acid reflux exposure. Next, ATF3 and EGR1 gene expression was analysed in tumour tissues and related to the median expression of autologous reference tissue samples. Results: ATF3 and EGR1 mRNA expression was significantly reduced after consecutive exposure of MOCs at pH <7.0 to artificial gastric juice (refluxate). In contrast, ATF3 mRNA was upregulated significantly within the first hour of incubation. EGR1 mRNA exhibited no significant changes. The analysed cell lines exhibited a cell line-specific alteration. In FADU DD cells, the upregulation of EGR1 was significant after refluxate exposure, but in HN-SCC 22B, no significant changes were detected. The analysis of the HNSCC samples confirmed the heterogeneous data of the literature. Conclusion: The data maintain the hypothesis that ATF3 and EGR1 are involved in the beginning of inflammatory processes. Whether these two transcription factors act as tumour suppressors or promoters is context dependent and warrants analysis in further studies. (C) 2017 S. Karger AG, Basel

关键词： NF-KAPPA-B   GENE-EXPRESSION   GLUCOSE-UPTAKE   ATF3   INFLAMMATION   MACROPHAGES   ACTIVATION   RECEPTOR   AMPK   SEPSIS  

摘要： Excessive activation of the TLR4 signalling pathway is critical for inflammation-associated disorders, while negative regulators play key roles in restraining TLR4 from over-activation. Naringenin is a citrus flavonoid with remarkable anti-inflammatory activity, but the mechanisms underlying its inhibition of LPS/TLR4 signalling are less clear. This study investigated the molecular targets and therapeutic effects of naringenin in vitro and in vivo. In LPS-stimulated murine macrophages, naringenin suppressed the expression of TNF-alpha, IL-6, TLR4, inducible NO synthase (iNOS), cyclo-oxygenase-2 (COX2) and NADPH oxidase-2 (NOX2). Naringenin also inhibited NF-kappa B and mitogen-activated protein kinase (MAPK) activation. However, it did not affect the IRF3 signalling pathway or interferon production, which upregulate activating transcription factor 3 (ATF3), an inducible negative regulator of TLR4 signalling. Naringenin was demonstrated to directly increase ATF3 expression. Inhibition of AMPK and its upstream calcium-dependent signalling reduced ATF3 expression and dampened the anti-inflammatory activity of naringenin. In murine endotoxaemia models, naringenin ameliorated pro-inflammatory reactions and improved survival. Furthermore, it induced AMPK activation in lung tissues, which was required for ATF3 upregulation and the enhanced anti-inflammatory activity. Overall, this study reveals a novel mechanism of naringenin through AMPK-ATF3-dependent negative regulation of the LPS/TLR4 signalling pathway, which thereby confers protection against murine endotoxaemia.

关键词： 急性肺损伤   活化转录因子3   基因芯片   差异基因表达  

摘要： 目的利用基因芯片技术筛选LPS诱导的急性肺损伤小鼠肺组织中与活化转录因子3(acting transcription factor 3,ATF3)相关的差异基因表达,以发现与ATF3可能相互作用的基因。方法采用基因芯片技术筛选野生型C57小鼠和ATF3敲基因小鼠经LPS处理后肺组织中的差异表达基因。进一步采用RT-PCR技术对重点基因的表达进行验证。结果与野生型C57小鼠比较,ATF3敲基因小鼠共筛选出1 692个差异基因,有21个与炎症调控相关的基因表达差异明显,其中,Myo18a、Mest、Gpr124、Rcan2、Tnfsf15、Ltbp3、Cdh2、Ppm1f、Spns2、Rarres2、Cgn、Mmrn2、Crb3、Adamts2、Mrc2、Sh3tc2、Cdk5rap1、Cenpf共18个基因在ATF3敲基因小鼠肺组织中表达明显上调;Padi4、Wfdc82个基因表达明显下调。对重点基因Tnfsf15(又称为TL1A)进一步验证,RT-PCR检测结果显示,表达趋势与基因芯片一致。结论 TL1A可能参与了ATF3对LPS诱导的急性肺损伤的炎症保护作用。

关键词： beta-catenin   noncoding RNA   ASBEL   colorectal tumorigenesis   ATF3   LONG NONCODING RNAS   TUMOR-SUPPRESSOR   COLON-CANCER   STEM-CELLS   E-CADHERIN   TRANSCRIPTION   CHROMATIN   PROTEINS   IDENTIFICATION   METASTASIS  

摘要： Wnt/beta-catenin signaling plays a key role in the tumorigenicity of colon cancer. Furthermore, it has been reported that lncRNAs are dysregulated in several steps of cancer development. Here we show that beta-catenin directly activates the transcription of the long noncoding RNA (lncRNA) ASBEL [antisense ncRNA in the ANA (Abundant in neuroepithelium area)/BTG3 (B-cell translocation gene 3) locus] and transcription factor 3 (TCF3), both of which are required for the survival and tumorigenicity of colorectal cancer cells. ASBEL interacts with and recruits TCF3 to the activating transcription factor 3 (ATF3) locus, where it represses the expression of ATF3. Furthermore, we demonstrate that ASBEL-TCF3-mediated down-regulation of ATF3 expression is required for the proliferation and tumorigenicity of colon tumor cells. ATF3, in turn, represses the expression of ASBEL. Our results reveal a pathway involving an lncRNA and two transcription factors that plays a key role in Wnt/beta-catenin-mediated tumorigenesis. These results may provide insights into the variety of biological and pathological processes regulated by Wnt/beta-catenin signaling.

关键词： ATF3   Human labour   Inflammation   Infection   Fetal membranes   ACTIVATING TRANSCRIPTION FACTOR-3   PRETERM LABOR   PROSTATE-CANCER   INTRAUTERINE INFECTION   SIGNALING PATHWAYS   PREMATURE RUPTURE   HUMAN MYOMETRIUM   EXPRESSION   PARTURITION   INTERLEUKIN-1  

摘要： Introduction: Infection and inflammation stimulate pro-inflammatory cytokines, prostaglandins and matrix metalloproteinase (MMP)-9, which play a central role in myometrial contractions and rupture of fetal membranes. In human and mouse immune cells, activating transcription factor 3 (ATF3) is a negative regulator of inflammation. No studies have examined the role of ATF3 in human labour. Methods: Primary amnion cells were used to determine the effect of interleukin (IL)-1 beta and the bacterial product fibroblast-stimulating lipopeptide (fsl-1) on ATF3 expression, and the effect of ATF3 siRNA on pro-labour mediators. ATF3 expression was assessed in fetal membranes from non-labouring and labouring women at term and preterm, and after preterm pre-labour rupture of membranes (PPROM). Results: IL-1 beta and fsl-1 significantly increased ATF3 expression. Silencing ATF3 significantly increased IL-1 beta- or fsl-1-induced expression of pro-inflammatory cytokines (TNF-alpha, IL-1 alpha, IL-1 beta, IL-6) and chemokines (IL-8 and monocyte chemoattractant protein-1 (MCP-1)); cyclooxygenase-2 (COX-2) mRNA expression and prostaglandin PGF(2 alpha) release; and MMP-9 expression. ATF3 expression was decreased in fetal membranes with term labour. There was no effect of preterm labour or PPROM on ATF3 expression. Discussion: ATF3 is a negative regulator of inflammation in human fetal membranes; in primary amnion cells, ATF3 expression is induced by IL-1 beta and fsl-1, and ATF3 silencing further exacerbates the inflammatory response when stimulated with these factors. Subsequently, ATF3 expression is decreased in fetal membranes after term labour and with preterm chorioamnionitis, conditions closely associated with inflammation and infection. Our data suggest that ATF3 may play a role in the terminal processes of human labour and delivery. (C) 2016 Elsevier Ltd. All rights reserved.

关键词： cAMP response element-binding protein   glucocorticoid   immunosuppression   liver metabolism   S100 proteins   IMMUNE SUPPRESSION   LIVER-DISEASE   FATTY LIVER   ATF3   INFLAMMATION   DIFFERENTIATION   ACCUMULATION   REGULATOR   EXPANSION   MICE  

摘要： Glucocorticoids (GCs) used as inflammation suppressors have harmful side effects, including induction of hepatic steatosis. The underlying mechanisms of GC-promoted dysregulation of lipid metabolism, however, are not fully understood. GCs could facilitate the accumulation of myeloid-derived suppressor cells (MDSC) in the liver of animals, and the potential role of MDSCs in GC-induced hepatic steatosis was therefore investigated in this study. We demonstrated that granulocytic (G)-MDSC accumulation mediated the effects of GCs on the fatty liver, in which activating transcription factor 3 (ATF3)/S100A9 signaling plays an important role. ATF3-deficient mice developed hepatic steatosis and displayed expansion of G-MDSCs in the liver and multiple immune organs, which shared high similarity with the phenotype observed in GC-treated wild-type littermates. Adoptive transfer of GC-induced or ATF3-deficient G-MDSCs promoted lipid accumulation in the liver, whereas depletion of G-MDSCs alleviated these effects. Mechanistic studies showed that in MDSCs, ATF3 was transrepressed by the GC receptor GR through direct binding to the negative GR-response element. S100A9 is the major transcriptional target of ATF3 in G-MDSCs. Silencing S100A9 clearly alleviated G-MDSCs expansion and hepatic steatosis caused by ATF3 deficiency or GC treatment. Our study uncovers an important role of G-MDSCs in GC-induced hepatic steatosis, in which ATF3 may have potential therapeutic implications.