关键词： 食管癌   激活转录因子3   细胞生长  

摘要： 背景与目的:激活转录因子3(activating transcription factor 3,ATF3)在食管癌中表达异常下调,但其功能仍不清楚。本研究拟探讨ATF3在食管癌细胞过表达对癌细胞生长及裸鼠成瘤的影响。材料与方法:利用分子克隆技术将ATF3基因完整编码区克隆至真核细胞表达载体pcDNA3中,获得重组表达质粒;将该表达质粒转染食管癌EC109细胞并用G418筛选稳定表达的细胞克隆;用Western blot检测ATF3的过表达效果;对ATF3过表达的细胞与相应对照细胞进行细胞克隆形成实验和裸鼠体内成瘤实验,以分析ATF3过表达对食管癌细胞生长的影响。结果:ATF3的过表达在体外可以降低食管癌细胞的克隆形成能力;在体内可以抑制食管癌细胞在雌鼠中的成瘤能力,但对雄鼠中的成瘤能力没有明显影响。结论:ATF3的过表达可以抑制食管癌细胞的生长,ATF3可能在食管癌的发生发展中发挥重要作用。

关键词： Fibroid   myometrium   jun   fos   ATF3   racial disparity   stress   UTERINE LEIOMYOMA   INDUCIBLE GENE   RISK-FACTORS   APOPTOSIS   FIBROIDS   PROGESTERONE   PROTEIN   GROWTH   WOMEN   ATF3  

摘要： Objective: To determine whether expression of the stress response gene ATF3 and related members of activator protein complex-1, cJun and cFos, were altered in leiomyoma compared with myometrium, and whether this difference might correlate with leiomyoma size or race. Design: Laboratory study. Setting: University hospital. Patient(s): Fifteen women undergoing hysterectomy for symptomatic leiomyoma. Intervention(s): Tissue procurement, RNA isolation, reverse-transcriptase polymerase chain reaction, real-time reverse-transcriptase polymerase (RT-PCR) chain reaction, immunohistochemistry, Western blot. Main Outcome Measure(s): Expression of mRNA and protein in leiomyoma and patient-matched myometrium. Result(s): mRNA transcripts of ATF3 were decreased in leiomyoma compared with matched myometrium by both RT-PCR and real-time RT-PCR. The decrease was greater than fivefold in a majority of samples. The reduction seen in ATF3 mRNA expression did not show a correlation with race and leiomyoma size. Surprisingly, immunohistochemistry and Western blot analysis demonstrated an elevation of ATF3 protein expression by a mean of 2.9-fold. Transcripts of related AP-1 genes, cJun and cFos, were significantly decreased by a mean of -29.57 for cJun and -23.78 for cFos, but there was no significant change in protein expression of the two transcription factors. Conclusions: Alterations in ATF3 gene expression resemble the response to mechanical and ischemic stress reported in other tissues. Results suggested that ATF3 protein expression was increased in leiomyoma, and may reflect increased tissue stress. (Fertil Steril (R) 2009;92:748-55. (C)2009 by American Society for Reproductive Medicine.)

关键词： 肝细胞癌ATF3   MMP-2   maspin   组织芯片  

摘要： 目的:检测ATF3、MMP-2及maspin在肝细胞癌(hepatocellular carcinoma,HCC)组织芯片中的表达情况,探讨ATF3、MMP-2及maspin表达与HCC发生、发展的关系及其意义。方法:对手工制作组织芯片的方法进行改良,构建含HCC、癌旁肝组织、远癌肝组织、正常肝组织及肝内转移灶的组织芯片,应用免疫组织化学和图像分析技术检测ATF3、MMP-2及maspin的相对表达量,并分析其与各临床病理参数的关系及其意义。结果:ATF3及MMP-2在HCC中表达明显高于癌旁肝组织、远癌肝组织及正常肝组织(P<0.01),而maspin在HCC中的表达明显低于癌旁肝组织、远癌肝组织及正常肝组织(P<0.01)。ATF3蛋白在女性HCC患者中的表达量高于男性患者(P<0.01),在转移组中表达高于无转移组(P<0.01);MMP-2蛋白随HCC病理学分级的升高,相对表达量降低(P<0.05),MMP-2蛋白在直径大于3cm的HCC中的相对表迭量高于直径小于3cm的表达量(P<0.05),包膜侵犯组高于无包膜侵犯组(P<0.05),肝内转移灶表达量高于原发灶(P<0.05);ATF3和MMP-2均随着HCC的进展表达量逐渐增高,maspin却随着HCC病理学分级的升高,相对表达量降低(P<0.05)。ATF3蛋白表达与MMP-2表达呈正相关(r_s=0.13,P<0.05),与maspin表达呈负相关(r_s=-0.23,P<0.01);MMP-2蛋白表达与maspm表达呈负相关(r_s=-0.17,P<0.01)。结论:在HCC组织中,ATF3、MMP-2及maspin的表达存在相关关系,ATF3、MMP-2高表达及maspin缺失表达与HCC的发生密切相关,他们可能在HCC发生发展、侵袭转移过程中发挥重要作用。选择性抑制ATF3有可能成为治疗HCC新的靶点。

关键词： Adult T-cell leukemia   Human T-lymphotropic virus type I   Activating transcriptional factor 3   Testican 3   VIRUS TYPE-I   ACTIVATING TRANSCRIPTION FACTOR-3   MATRIX METALLOPROTEINASES   ANTIBODIES   GENE   ATF3   SUPPRESSION   INDUCER   EMMPRIN   DISEASE  

摘要： Adult T-cell leukemia (ATL) is an aggressive disease characterized by visceral invasion, and ATL regulates matrix metalloproteinase (MMP) activities of the endothelial cells. The controlling system of MMP activities in ATL is regulated by various factors such as Emmprin and tissue inhibitor of MMP. In this study, we demonstrated that Testican 3 expression in ATL decreased the activity of MMP. Furthermore, we showed that the expression of Testican 3 was regulated by activating transcriptional factor 3 in human T-lymphotropic virus type I-related cell lines. Thus, Testican 3 is a novel regulator to reduce the activity of MMP in ATL (C) 2008 Elsevier Ltd. All rights reserved.

关键词： 膀胱尿路上皮癌   ATF3   免疫组织化学   RT—PCR  

摘要： 目的探讨膀胱尿路上皮癌中转录因子ATF3的表达及其临床意义。方法采用逆转录聚合酶链反应(RT-PCR)和免疫组织化学方法检测32例膀胱癌组织和14例正常膀胱粘膜中ATF3 mRNA和ATF3蛋白的表达,分析其表达与肿瘤病理分级、临床分期的关系。结果膀胱癌组织中ATF3 mRNA相对表达量稍高于正常膀胱粘膜组织,浸润膀胱癌组织中ATF3 mRNA相对表达量稍高于浅表膀胱癌组织,但差别均无显著性。ATF3蛋白在膀胱癌组织和正常膀胱粘膜中的阳性表达率分别为93.7%(30/32)、21.4%(3/14),差异有显著性。结论ATF3表达参与了膀胱尿路上皮癌的发病过程,进一步研究其功能有可能揭示膀胱癌的发病机制。

关键词： ATF3   shRNA   肾小球系膜细胞  

摘要： 目的:构建针对大鼠ATF3(activating transcription factor3,ATF3)基因的特异性短发卡RNA(small hairpinRNA,sh RNA)真核表达质粒。方法:用DNA重组技术将针对大鼠ATF3基因不同位点所设计的5个shRNA序列克隆到真核表达质粒pGenesil-1中。在酶切分析及序列测定后,用脂质体将5个shRNA分别转染至大鼠肾小球系膜细胞(glomerular mesangial cells,GMC),随后用Western blot检测蛋白的相对表达量来筛选最佳沉默效率的shRNA。结果:限制性酶切及核酸序列分析证明重组质粒构建正确。Western blot证实在重组质粒中,shATF3-1具有最佳的沉默效率。结论:成功构建了针对ATF3基因的shRNA的真核表达质粒,并且该shRNA可以特异性地降低大鼠肾小球系膜细胞ATF3的蛋白表达。

关键词： DNA repair   ATF3   p15(paf)   PCNA   UV   ACTIVATING TRANSCRIPTION FACTOR-3   NUCLEOTIDE EXCISION-REPAIR   PCNA-ASSOCIATED FACTOR   DNA-REPAIR   INDUCED APOPTOSIS   DOWN-REGULATION   GENE   IRRADIATION   EXPRESSION   INDUCTION  

摘要： After genotoxic stress, normal cells trigger DNA repair or, if unable to repair, undergo apoptosis to eradicate the cells that bear the risk of becoming tumorigenic. Here we show that repression of the transcription factor, activating transcription factor 3 (ATF3), after ultraviolet (UV)-mediated genotoxic stress impairs the DNA repair process. We provide evidence that ATF3 directly regulates the proliferating cell nuclear antigen (PCNA)-associated factor KIAA0101/p15(PAF). We further show that the expressions of ATF3 and p15(PAF) is sufficient to trigger the DNA repair machinery, and that attenuation of their expression alters DNA repair mechanisms. We show that the expression of p15(PAF) compensates for a lack of ATF3 expression, thereby constituting a major effector of ATF3 in the DNA repair process. In addition, we provide evidence that p15(PAF) expression is required for the correct function of PCNA during DNA repair, as prevention of their interaction significantly alters DNA repair mechanisms. Finally, defective DNA repair, because of the downregulation of p15(PAF) expression, rendered the cells more sensitive to UV-induced cell death. Therefore, our results suggest ATF3 and p15(PAF) as novel gatekeepers of genomic integrity after UV exposure.

关键词： Diarylheptanoid   Cell cycle arrest   Apoptosis   p53   ATF3   ACTIVATING TRANSCRIPTION FACTOR-3   ALPINIA-OFFICINARUM HANCE   DNA-DAMAGE   NEUROBLASTOMA-CELLS   SIGNALING PATHWAYS   MOLECULAR-BIOLOGY   UP-REGULATION   IN-VIVO   KAPPA-B   KINASE  

摘要： Purpose The aim of the study is to dissect the cytotoxic mechanisms of 1-(4-hydroxy-3-methoxyphenyl)-7-(3,4dihydroxyphenyl)-4E-en-3-heptanone (compound 1) in SH-SY5Y cells and therefore to provide new insight into neuroblastoma chemotherapy. Methods Nine diarylheptanoids were isolated from Alpinia officinarum by chromatography and their cytotoxicity was evaluated by an MTS assay. Flow cytometry, BrdU incorporation assay and fluorescence staining were employed to investigate cytostatic and apoptotic effects induced by the compound 1. In addition, Western blot, qPCR and siRNA techniques were used to elucidate the molecular mechanisms of the cytotoxicity. Results The study to elucidate the cytotoxic mechanisms of compound 1, the most potent diarylheptanoid showed that cell cycle-related proteins, cyclins, CDKs and CDKIs, as well as two main apoptotic related families, caspase and Bcl 2 were involved in S phase arrest and apoptosis in neuroblastoma cell line SH-SY5Y. Furthermore, following the drug treatment, the protein expression of p53, phospho-p53 (Ser20) as well as the p53 transcriptional activated genes ATF3, puma and Apaf-1 were increased dramatically; MDM2 and Aurora A, the two p53 negative regulators were decreased; the p53 protein stability was enhanced, whereas the p53 mRNA expression level slightly decreased and ATF3 mRNA expression apparently increased. In addition, the knockdown of ATF3 gene by siRNA partially suppressed p53, caspase 3, S phase arrest and apoptosis triggered by compound 1. Conclusion These results suggest that compound 1 induces S phase arrest and apoptosis via up regulation of ATF3 and stabilization of p53 in SH-SY5Y cell line. Therefore, compound 1 might be a promising lead structure for neuroblastoma therapy.

关键词： Ribotoxic stress   Activating transcription factor 3 (ATF3)   Epithelial cell survival   p38 MAP kinase   Double-stranded RNA (dSRNA)-activated protein kinase (PKR)   DOUBLE-STRANDED-RNA   PROTEIN-KINASE-R   BETA-CATENIN DEGRADATION   JUN NH2-TERMINAL KINASE   INDUCED APOPTOSIS   GENE-EXPRESSION   INHIBITOR SB203580   ENDOTHELIAL-CELLS   HUMAN NEUTROPHILS   SHIGA TOXIN-1  

摘要： Ribotoxic stress responses lead to the expression of genes important for cellular homeostasis by modulating cell survival, proliferation and differentiation. ATF3 was investigated for its modulation of the epithelial cellular integrity in response to mucosal ribotoxic stress. ATF3 expression was up-regulated by chemical agents causing ribotoxic stress such as deoxynivalenol. and anisomycin in different types of intestinal epithelial cells. Moreover, reduction of ATF3 gene expression promoted ribotoxic stress-triggered programmed cell death, implicating a protective role of ATF3 in epithelial cell survival. Mechanistically, stabilization of ATF3 messenger RNA and protein played a critical role in maintaining enhanced levels of ATF3 production in response to the ribotoxic chemical agent. For ATF3 mRNA stability, p38 specific inhibitor SB203580 was the most efficient agent for suppression, suggesting the involvement of the p38 MAP kinase in ATF3 mRNA stabilization. in addition, the p38 MAP kinase as well as its downstream mediator glycogen synthase kinase 30 (GSK3 beta) was involved in ATF3 protein stabilization caused by chemical ribotoxic stress in human epithelial cells. As another separate signaling cascade, double-stranded RNA (dsRNA)-activated protein kinase (PKR) was demonstrated to translationally modulate ATF3 expression and contribute to the epithelial cell survival. PKR interference caused cells to be more susceptible to cell death caused by the chemical ribotoxic stress. The results of this study showed that enhanced ATF3 production was associated with cellular defenses by maintaining the epithelial survival after ribotoxic mucosal insults. (C) 2009 Published by Elsevier Inc.