硫氧还蛋白过氧化物酶Peroxiredoxin Ⅱ与前列腺癌关系的研究
|
摘要
研究背景与目的
硫氧还蛋白过氧化物酶家族最初发现于酵母中,构成了一个与既往的抗氧化蛋白无同源性的抗氧化物酶家族。目前从细菌到哺乳动物细胞中报道有一百多个具有高度保守蛋白结构域的同系物。硫氧还蛋白过氧化物酶家族在多种细胞功能中发挥重要作用,如抗氧化损伤保护蛋白和脂质,细胞增殖、分化,调节凋亡过程中细胞内信号传导等。增加细胞内抗氧化酶水平可抗凋亡。硫氧还蛋白过氧化物酶家族是红细胞中主要的抗氧还剂,并且介导早期红细胞系的分化。该家族所包含的一类过氧化物酶,能够利用其活性半胱氨酸残基清除过氧化物及羟基。
根据氨基酸序列的差异,硫氧还蛋白过氧化物酶家族有六个亚型,并被分为两大亚家族。亚家族一含有两个保守性半胱氨酸残基,包括Ⅰ、Ⅱ、Ⅲ、Ⅳ四个亚型,亚家族二拥有一个保守性半胱氨酸残基,包括Ⅴ和Ⅵ两个亚型。Ⅰ和Ⅱ亚型位于胞浆中。与细胞增殖、分化、抗凋亡及肿瘤相关。研究表明,硫氧还蛋白过氧化物酶Ⅱ在红细胞及K-562红白血病细胞系中作为巯基特异性的抗氧还剂。
硫氧还蛋白过氧化物酶家族在某些肿瘤组织中高表达。硫氧还蛋白过氧化物酶Ⅰ、Ⅱ和Ⅲ在人乳腺癌组织中过表达,并与肿瘤的发展和分级相关。硫氧还蛋白过氧化物酶Ⅰ在肺癌、甲状腺瘤及口腔肿瘤中也有高表达,是潜在的肿瘤标记物。
硫氧还蛋白过氧化物酶Ⅱ是酵母的硫氧还蛋白过氧化物酶的人类同源体,又名硫氧还蛋白依赖的过氧化物还原酶Ⅰ(thioredoxin-dependent peroxide reductase 1 ,TDPX1)或自然杀伤增强因子B(natural killer-enhancing factor-B ,NEKF-B),位于染色体13q12,在人神经元细胞有表达,但在神经胶质细胞不表达。
硫氧还蛋白过氧化物酶Ⅱ可利用硫氧还蛋白作为直接的电子供体发挥降解过氧化氢及其他活性氧类物质的作用。其过氧化物酶活性能保护细胞免于活性氧的损伤。硫氧还蛋白过氧化物酶Ⅱ因其对细胞内过氧化氢浓度调节作用,参与了生长因子和肿瘤坏死因子α的细胞内信号转导。
目前研究表明,硫氧还蛋白过氧化物酶Ⅱ具有硫氧还蛋白依赖的过氧化物酶活性,能够抑制细胞凋亡,与细胞膜结合,与红细胞中钙离子激活的钾离子转运相关。在既往的研究中,我们发现在皮肤组织中至少有三种硫氧还蛋白过氧化物酶亚型存在,而硫氧还蛋白过氧化物酶Ⅱ主要存在于真皮层的上皮细胞及其附属结构中(毛囊、汗腺及皮脂腺中),我们还发现,硫氧还蛋白过氧化物酶Ⅱ存在于正常皮肤的血管内皮细胞中。硫氧还蛋白过氧化物酶Ⅱ存在于红细胞及人脐内皮细胞的胞浆中,同时表达于人体表皮肤的良恶性血管瘤。因其高表达于血管内的红细胞中,甚至可作为检测组织样本中的血管标记物。
硫氧还蛋白过氧化物酶Ⅱ不仅能发挥抗氧化应激的细胞保护作用,同时赋予肿瘤细胞抗过氧化氢、顺铂和抗辐射的能力。硫氧还蛋白过氧化物酶Ⅱ可在过氧化氢及顺铂的作用下表达上调,并能抑制顺铂、促甲状腺激素、血浆剥夺、神经酰胺及鬼臼乙叉甙引起的细胞凋亡,因此硫氧还蛋白过氧化物酶Ⅱ的过表达可导致化疗耐药。研究表明,硫氧还蛋白过氧化物酶Ⅱ还能介导辐射抵抗。
在既往的研究中,我们通过二维电泳和质谱分析发现Prx II蛋白在高转移潜能的肿瘤细胞系PC-3M-1E8的表达水平较低转移潜能的肿瘤细胞系PC-3M-2B4明显下调。为了研究前列腺癌的分子机制,阐明Prx II基因和前列腺癌的相关性,我们对Prx II基因进行了深入的研究。基于Prx II基因在不同转移潜能的前列腺癌细胞中存在表达异常和功能障碍,通过药物或者遗传修饰等手段控制Prx II基因的功能或它们的调节通路,将是一个令人振奋但又出人意料的分子治疗靶点。
方法
运用RT-PCR、免疫细胞化学及Western blot法检测配对高低转移前列腺癌细胞系中Prx II mRNA及蛋白的表达水平;采用基因工程技术构建Prx II全长真核表达载体,G418筛选稳定表达Prx II的前列腺癌细胞克隆株;应用RT-PCR、cfse-流式细胞测定和体外细胞迁移实验等方法观察1E8/Prx II细胞增殖及其运动能力的变化;应用RT-PCR、Western blot法检测1E8/Prx II细胞和1E8/pcDNA3.1细胞中AKT、P-AKT、E-cadherin、β-catenin和Bcl-X/L的表达变化;采用细胞迁移实验、Transwell体外侵袭实验和细胞黏附实验检测1E8/Prx II细胞和1E8/pcDNA3.1细胞的增殖、细胞运动、体外侵袭和黏附能力。另外,为了筛选在前列腺癌细胞中与PrxⅡ蛋白相互作用的蛋白,我们应用酵母双杂交体系,构建人PrxⅡ蛋白酵母双杂交诱饵载体,将该基因片段与pGBKT7载体定向重组,将序列正确的重组质粒pGBKT7-PrxⅡ转化入AH109酵母菌株,在缺陷性培养基上观察pGBKT7-PrxⅡ在AH109中的表达情况,检测诱饵载体有无毒性作用和单倍体及二倍体自激活功能。
结果
1. Prx II mRNA和蛋白水平在高转移潜能的前列腺癌细胞系PC-3M-1E8的表达水平较低转移潜能的前列腺癌细胞系PC-3M-2B4明显下调,表明Prx II基因的异常表达与肿瘤转移密切相关,
2.成功构建了Prx II基因的全长真核表达载体,获得稳定表达Prx II的前列腺癌细胞克隆株1E8/Prx II和转染空载体pcDNA3.1的对照细胞克隆株1E8/pcDNA3.1。
3.通过转染技术使人前列腺癌PC-3M-1E8细胞过表达Prx II,能够抑制细胞体外增殖、运动和侵袭能力,同时,Prx II的过表达促进丁基过氧化氢(TBHP)引起的前列腺癌细胞凋亡,抑制细胞增殖,可能是通过下调PI4K/AKT胞内信号对PC-3M-1E8细胞发挥作用的。
4.过表达外源性Prx II,通过使AKT失活、调控内化的E-cadherin的再循环,增加E-cadherin/catenin复合物形成,改变了细胞极性和组织结构,抑制人前列腺癌细胞系PC-3M-1E8细胞的转移表型。
5.成功构建pGBKT7-PrxⅡ重组质粒。转化有重组质粒和pGBKT7空载体的酵母菌都能在SD/-Trp/X-α-gal平板上长出粉红色菌落,在SD/-His /-Trp /X-α-gal, SD/-Ade/-Trp/X-α-gal平板上不能生长,并将pGBKT7-PrxⅡ诱饵质粒成功转化入酵母菌AH109,且对酵母菌株AH109无毒性,不具自主激活报告基因的功能。
结论
验证PrxⅡ在不同高低转移配对前列腺癌细胞株的表达显示,PrxⅡ基因的异常表达与肿瘤转移密切相关,使人前列腺癌PC-3M-1E8细胞过表达Prx II,能够抑制细胞体外增殖、运动和侵袭能力,同时,Prx II的过表达促进丁基过氧化氢(TBHP)引起的前列腺癌细胞凋亡,抑制细胞增殖。
进一步探讨PrxⅡ基因参与前列腺癌侵袭转移的分子机制发现:外源性PrxⅡ过表达,下调PI4K/AKT胞内信号,抑制人前列腺癌PC-3M-1E8细胞的体外增殖和运动能力;PrxⅡ的过表达抑制AKT活化,调控内化的E-cadherin的再循环,从而改变E-cadherin/catenin复合物维持细胞极性和组织结构的功能,抑制人前列腺癌PC-3M-1E8细胞的转移表型,PrxⅡ的过度表达逆转前列腺癌细胞的转移表型。另外,成功获得了在酵母细胞中正确表达,并对酵母细胞无毒性且未自主激活报告基因的诱饵表达载体pGBKT7-PrxⅡ,该载体可作为酵母双杂交系统中的“诱饵”,该诱饵载体可应用于酵母双杂交系统3筛选PrxⅡ相互作用蛋白。
Background and Objective
Peroxiredoxins (Prxs), initially characterized in yeast, constitute a family of antioxidant enzymes with no homology with conventional antioxidant proteins. More than a hundred homologues have been described from bacteria to mammals, sharing highly conserved protein domains. Prxs play a key role in several cellular functions including protein and lipid protection against oxidative injury, cell proliferation, differentiation and intracellular signaling pathways regulating apoptosis. Increased cellular levels of antioxidant enzymes confer protection against apoptosis. Prxs as reported to be one of major antioxidants in RBCs, and was induced at early stages of erythroid differentiation. The Prx family consists of a kind of peroxidase that is able to remove both peroxide and hydroxyl radical– using reactive cysteine.
Six Prx isoforms have been identified which, based on the amino acid sequences, are generally divided into two subfamilies; groups I, II, III, and IV with two conserved cysteines and groups V and VI with one conserved cysteine. Prx I and Prx II proteins are known to be located in cytoplasm. Prx isotypes (I-VI) are associated with the cellular proliferation, differentiation, and antiapoptotic effect, and their expression is relevant to cancer. Prx II was identified as a thiol-specific antioxidant in the K-562 erythroleukemia cell line and erythrocytes, respectively.
Prx over-expression is frequently observed in certain types of cancer tissues. Three types of Prx (I, II, and III) have been shown to be over-expressed in the case of human breast cancer, and it has been suggested that their over-expressions are related to cancer development or progression. The increased expression of Prx I is also detected in lung cancer, thyroid tumors and oral cancer, and is suggested to constitute a potential tumor marker.
Prx II is a human homologue of yeast thioredoxin peroxidase (TPX), also named thioredoxin-dependent peroxide reductase 1 (TDPX1) or natural killer-enhancing factor-B (NEKF-B). It is located on chromosome 13q12 and is expressed in human neurons but not in glial cells.
Prx II is an antioxidant enzyme that reduces H2O2 and other reactive oxygen species using thioredoxin as the immediate electron donor, and its peroxidase activity prevents cells from reactive oxygen species insult. Prx II is also involved in the cellular signaling pathways of growth factors and tumor necrosis factor-a, by virtue of its regulation of intracellular H2O2.
Several properties of Prx II, such as thioredoxin- dependent peroxidase activity, inhibition of apoptosis, characteristic binding to membrane, and correlation with Ca2+-activated potassium transport in erythrocytes, have been reported. In previous studies, we found that at least 3 isotypes of Prx have been identified in the skin, and Prx II was identified at the epidermal cells and epidermal appendage structures (hair follicle, eccrine gland, and sebaceous gland) of the dermis. We also found that Prx II was expressed in vascular endothelial cells in normal skin. Prx II has been reported to be present in the cytoplasm of RBCs and human umbilical endothelial cells. In a previous study, Prx II is expressed in endothelial cells of benign and malignant vascular tumors in human skin in vivo. In addition to biologic availability of Prx II as an endothelial marker, it has an advantage in detecting vascular spaces in tissue specimens because it is strongly expressed in intravascular erythrocytes.
Prx II is known not only to protect cells from oxidative damage caused by hydrogen peroxide (H2O2), but also to endow cancer cells with resistance to both H2O2 and cisplatin and to grant them radioresistance. Prx II is up-regulated by H2O2 and cisplatin treatment, and its increased expression inhibits the apoptosis induced by cisplatin, thyrotropin, serum deprivation, ceramide, or etoposide, thereby rendering tumor cells resistant to some chemotherapeutic agents. We demonstrated earlier that Prx II was involved in radioresistance.
In previous studies, we found that Prx II expression was notably down-regulated in tumor cell lines PC-3M-1E8 with high metastasis potentials compared with lowly metastatic cancer cell lines PC-3M-2B4 through two-dimensional gel electrophoresis (2-DE) and mass spectrometry analyses。To better understand the molecular mechanisms of prostate cancer and demonstrate the relativity of Prx II and prostate cancer, the function of Prx II gene was further investigated. Based on the prevalence of altered Prx II expression and function in different deseases, it was of interest to consider the therapeutic potential of controlling Prx II function or their regulated pathways through pharmacologic or genetic modulation.
Methods
Prx II mRNA and protein expression of matched-pair of prostate cancer cell lines with high and low metastasis potentials were detected by reverse transcriptase-polymerase chain reaction (RT-PCR), immunocytochemistry and western blotting . Prx II full-length eukaryotic expression vectors were constructed using the technique of gene engineering; the prostate cancer cell clones stable expressing Prx II were screened by G418. After transfection, the changes of cell proliferation and motility were detected by CFSE-flow cytometry and monolayer wound healing assay, respectively. The changes of AKT、P-AKT、E-cadherin、β-catenin and Bcl-X/L expression in 1E8/Prx II and 1E8/pcDNA3.1 cell clones were detected by RT-PCR and western blotting. The capabilities of cell growth, cell motility, in vitro invasion and cell adhesion in 1E8/Prx II and 1E8/pcDNA3.1 cell clones were performed by monolayer wound healing assay, Transwell migration assay and cell adhesion assay, respectively. In addition, To construct and identify a yeast two hybrid system bait vector for screening of homo sapiens PrxⅡbinding proteins in prostate cancer, full fragment of ORF of PrxⅡcDNA was amplified using PCR and directly ligated to the pGBKT7 vector. Insert-contained plasmid was confirmed by restriction endonuclease analysis and DNA sequencing. The plasmid was transformed into the yeast cell AH109, and its toxicity and transcriptional activation was tested by both the phenotype assay and the color assay.
Results
1. In contrast, Prx II mRNA and protein expression was notably down-regulated in prostate tumor cell line PC-3M-1E8 with high metastasis potentials compared with lowly metastatic cancer cell line PC-3M-2B4; These results indicated that abnormal-expression of Prx II gene closely correlated with neoplasm metastasis. Morever, its over-expression inhibited cancer metastasis in prostate cancer;
2. Constructed Prx II full-length eukaryotic expression vector successfully, and obtained prostate cancer cell clone(1E8/Prx II) which stably expressed Prx II. At the same time, we obtained control cell clone 1E8/pcDNA3.1.
3. After transferring Prx II gene into human prostate cancer cell line PC-3M-1E8, Prx II decreased the abilities of cell proliferation and motility of PC-3M-1E8 cells, Prx II is up-regulated by H2O2(TBHP) treatment, and its over-expression increases the apoptosis induced by TBHP. These functions might be regulated by activating PI4K/AKT intracellular signaling pathway
4. Prx II gene over-expression inhibited AKT phosphorylated at Ser473 site and regulated the recycling of internalized E-cadherin. Because of changing the function of E-cadherin /catenin complex retaining cell polarity and tissue structure, Prx II gene over-expression reversed the malignant phenotypes of PC-3M-1E8 cell lines.
5. The fragments of PrxⅡprotein were successfully obtained. The recombinant pGBKT7-PrxⅡplasmids and empty pGBKT7 vector could grow white colonies on SD/-Trp/X-α-gal plates and none could survive on SD/-His /-Trp /X-α-gal, SD/-Ade/-Trp/X-α-gal plates. The bait plasmid pGBKT7-PrxⅡconstructed expresses correctly, and can not activate the transcription of reporter gene alone.
Conclusion
Identification of PrxⅡmRNA and protein expression in matched-pairs of human prostate cancer cell lines with high and low metastasis potentials showed that abnormal-expression of PrxⅡgene closely correlated with neoplasm metastasis, moreover, in prostate cancer PrxⅡover-expression inhibited cancer metastasis and decreased the abilities of cell proliferation and motility of PC-3M-1E8 cell with high metastasis potentials. Prx II is upregulated by H2O2(TBHP) treatment, and its over-expression increases the apoptosis induced by TBHP.
To further investigate the molecular mechanism of PrxⅡin prostate cancer invasion and metastasis, the present studies showed that forced PrxⅡexpression inhibited the capabilities of cell growth and motility of Skov-3 cells through down-regulating the PI4K/AKT signaling; Over-expression of that gene inhibited AKT phosphorylated at Ser473 site and regulated the recycling of internalized E-cadherin. Because of changing the function of E-cadherin/catenin complex retaining cell polarity and tissue structure, Prx II gene over-expression reversed the malignant phenotypes of PC-3M-1E8 cell lines. Moreover, the bait plasmid pGBKT7-PrxⅡconstructed expresses correctly, and can not activate the transcription of reporter gene alone. The yeast two hybrid GAL4 system3 can be utilized to fish PrxⅡr egion interacting protein.
硫氧还蛋白过氧化物酶家族最初发现于酵母中,构成了一个与既往的抗氧化蛋白无同源性的抗氧化物酶家族。目前从细菌到哺乳动物细胞中报道有一百多个具有高度保守蛋白结构域的同系物。硫氧还蛋白过氧化物酶家族在多种细胞功能中发挥重要作用,如抗氧化损伤保护蛋白和脂质,细胞增殖、分化,调节凋亡过程中细胞内信号传导等。增加细胞内抗氧化酶水平可抗凋亡。硫氧还蛋白过氧化物酶家族是红细胞中主要的抗氧还剂,并且介导早期红细胞系的分化。该家族所包含的一类过氧化物酶,能够利用其活性半胱氨酸残基清除过氧化物及羟基。
根据氨基酸序列的差异,硫氧还蛋白过氧化物酶家族有六个亚型,并被分为两大亚家族。亚家族一含有两个保守性半胱氨酸残基,包括Ⅰ、Ⅱ、Ⅲ、Ⅳ四个亚型,亚家族二拥有一个保守性半胱氨酸残基,包括Ⅴ和Ⅵ两个亚型。Ⅰ和Ⅱ亚型位于胞浆中。与细胞增殖、分化、抗凋亡及肿瘤相关。研究表明,硫氧还蛋白过氧化物酶Ⅱ在红细胞及K-562红白血病细胞系中作为巯基特异性的抗氧还剂。
硫氧还蛋白过氧化物酶家族在某些肿瘤组织中高表达。硫氧还蛋白过氧化物酶Ⅰ、Ⅱ和Ⅲ在人乳腺癌组织中过表达,并与肿瘤的发展和分级相关。硫氧还蛋白过氧化物酶Ⅰ在肺癌、甲状腺瘤及口腔肿瘤中也有高表达,是潜在的肿瘤标记物。
硫氧还蛋白过氧化物酶Ⅱ是酵母的硫氧还蛋白过氧化物酶的人类同源体,又名硫氧还蛋白依赖的过氧化物还原酶Ⅰ(thioredoxin-dependent peroxide reductase 1 ,TDPX1)或自然杀伤增强因子B(natural killer-enhancing factor-B ,NEKF-B),位于染色体13q12,在人神经元细胞有表达,但在神经胶质细胞不表达。
硫氧还蛋白过氧化物酶Ⅱ可利用硫氧还蛋白作为直接的电子供体发挥降解过氧化氢及其他活性氧类物质的作用。其过氧化物酶活性能保护细胞免于活性氧的损伤。硫氧还蛋白过氧化物酶Ⅱ因其对细胞内过氧化氢浓度调节作用,参与了生长因子和肿瘤坏死因子α的细胞内信号转导。
目前研究表明,硫氧还蛋白过氧化物酶Ⅱ具有硫氧还蛋白依赖的过氧化物酶活性,能够抑制细胞凋亡,与细胞膜结合,与红细胞中钙离子激活的钾离子转运相关。在既往的研究中,我们发现在皮肤组织中至少有三种硫氧还蛋白过氧化物酶亚型存在,而硫氧还蛋白过氧化物酶Ⅱ主要存在于真皮层的上皮细胞及其附属结构中(毛囊、汗腺及皮脂腺中),我们还发现,硫氧还蛋白过氧化物酶Ⅱ存在于正常皮肤的血管内皮细胞中。硫氧还蛋白过氧化物酶Ⅱ存在于红细胞及人脐内皮细胞的胞浆中,同时表达于人体表皮肤的良恶性血管瘤。因其高表达于血管内的红细胞中,甚至可作为检测组织样本中的血管标记物。
硫氧还蛋白过氧化物酶Ⅱ不仅能发挥抗氧化应激的细胞保护作用,同时赋予肿瘤细胞抗过氧化氢、顺铂和抗辐射的能力。硫氧还蛋白过氧化物酶Ⅱ可在过氧化氢及顺铂的作用下表达上调,并能抑制顺铂、促甲状腺激素、血浆剥夺、神经酰胺及鬼臼乙叉甙引起的细胞凋亡,因此硫氧还蛋白过氧化物酶Ⅱ的过表达可导致化疗耐药。研究表明,硫氧还蛋白过氧化物酶Ⅱ还能介导辐射抵抗。
在既往的研究中,我们通过二维电泳和质谱分析发现Prx II蛋白在高转移潜能的肿瘤细胞系PC-3M-1E8的表达水平较低转移潜能的肿瘤细胞系PC-3M-2B4明显下调。为了研究前列腺癌的分子机制,阐明Prx II基因和前列腺癌的相关性,我们对Prx II基因进行了深入的研究。基于Prx II基因在不同转移潜能的前列腺癌细胞中存在表达异常和功能障碍,通过药物或者遗传修饰等手段控制Prx II基因的功能或它们的调节通路,将是一个令人振奋但又出人意料的分子治疗靶点。
方法
运用RT-PCR、免疫细胞化学及Western blot法检测配对高低转移前列腺癌细胞系中Prx II mRNA及蛋白的表达水平;采用基因工程技术构建Prx II全长真核表达载体,G418筛选稳定表达Prx II的前列腺癌细胞克隆株;应用RT-PCR、cfse-流式细胞测定和体外细胞迁移实验等方法观察1E8/Prx II细胞增殖及其运动能力的变化;应用RT-PCR、Western blot法检测1E8/Prx II细胞和1E8/pcDNA3.1细胞中AKT、P-AKT、E-cadherin、β-catenin和Bcl-X/L的表达变化;采用细胞迁移实验、Transwell体外侵袭实验和细胞黏附实验检测1E8/Prx II细胞和1E8/pcDNA3.1细胞的增殖、细胞运动、体外侵袭和黏附能力。另外,为了筛选在前列腺癌细胞中与PrxⅡ蛋白相互作用的蛋白,我们应用酵母双杂交体系,构建人PrxⅡ蛋白酵母双杂交诱饵载体,将该基因片段与pGBKT7载体定向重组,将序列正确的重组质粒pGBKT7-PrxⅡ转化入AH109酵母菌株,在缺陷性培养基上观察pGBKT7-PrxⅡ在AH109中的表达情况,检测诱饵载体有无毒性作用和单倍体及二倍体自激活功能。
结果
1. Prx II mRNA和蛋白水平在高转移潜能的前列腺癌细胞系PC-3M-1E8的表达水平较低转移潜能的前列腺癌细胞系PC-3M-2B4明显下调,表明Prx II基因的异常表达与肿瘤转移密切相关,
2.成功构建了Prx II基因的全长真核表达载体,获得稳定表达Prx II的前列腺癌细胞克隆株1E8/Prx II和转染空载体pcDNA3.1的对照细胞克隆株1E8/pcDNA3.1。
3.通过转染技术使人前列腺癌PC-3M-1E8细胞过表达Prx II,能够抑制细胞体外增殖、运动和侵袭能力,同时,Prx II的过表达促进丁基过氧化氢(TBHP)引起的前列腺癌细胞凋亡,抑制细胞增殖,可能是通过下调PI4K/AKT胞内信号对PC-3M-1E8细胞发挥作用的。
4.过表达外源性Prx II,通过使AKT失活、调控内化的E-cadherin的再循环,增加E-cadherin/catenin复合物形成,改变了细胞极性和组织结构,抑制人前列腺癌细胞系PC-3M-1E8细胞的转移表型。
5.成功构建pGBKT7-PrxⅡ重组质粒。转化有重组质粒和pGBKT7空载体的酵母菌都能在SD/-Trp/X-α-gal平板上长出粉红色菌落,在SD/-His /-Trp /X-α-gal, SD/-Ade/-Trp/X-α-gal平板上不能生长,并将pGBKT7-PrxⅡ诱饵质粒成功转化入酵母菌AH109,且对酵母菌株AH109无毒性,不具自主激活报告基因的功能。
结论
验证PrxⅡ在不同高低转移配对前列腺癌细胞株的表达显示,PrxⅡ基因的异常表达与肿瘤转移密切相关,使人前列腺癌PC-3M-1E8细胞过表达Prx II,能够抑制细胞体外增殖、运动和侵袭能力,同时,Prx II的过表达促进丁基过氧化氢(TBHP)引起的前列腺癌细胞凋亡,抑制细胞增殖。
进一步探讨PrxⅡ基因参与前列腺癌侵袭转移的分子机制发现:外源性PrxⅡ过表达,下调PI4K/AKT胞内信号,抑制人前列腺癌PC-3M-1E8细胞的体外增殖和运动能力;PrxⅡ的过表达抑制AKT活化,调控内化的E-cadherin的再循环,从而改变E-cadherin/catenin复合物维持细胞极性和组织结构的功能,抑制人前列腺癌PC-3M-1E8细胞的转移表型,PrxⅡ的过度表达逆转前列腺癌细胞的转移表型。另外,成功获得了在酵母细胞中正确表达,并对酵母细胞无毒性且未自主激活报告基因的诱饵表达载体pGBKT7-PrxⅡ,该载体可作为酵母双杂交系统中的“诱饵”,该诱饵载体可应用于酵母双杂交系统3筛选PrxⅡ相互作用蛋白。
Background and Objective
Peroxiredoxins (Prxs), initially characterized in yeast, constitute a family of antioxidant enzymes with no homology with conventional antioxidant proteins. More than a hundred homologues have been described from bacteria to mammals, sharing highly conserved protein domains. Prxs play a key role in several cellular functions including protein and lipid protection against oxidative injury, cell proliferation, differentiation and intracellular signaling pathways regulating apoptosis. Increased cellular levels of antioxidant enzymes confer protection against apoptosis. Prxs as reported to be one of major antioxidants in RBCs, and was induced at early stages of erythroid differentiation. The Prx family consists of a kind of peroxidase that is able to remove both peroxide and hydroxyl radical– using reactive cysteine.
Six Prx isoforms have been identified which, based on the amino acid sequences, are generally divided into two subfamilies; groups I, II, III, and IV with two conserved cysteines and groups V and VI with one conserved cysteine. Prx I and Prx II proteins are known to be located in cytoplasm. Prx isotypes (I-VI) are associated with the cellular proliferation, differentiation, and antiapoptotic effect, and their expression is relevant to cancer. Prx II was identified as a thiol-specific antioxidant in the K-562 erythroleukemia cell line and erythrocytes, respectively.
Prx over-expression is frequently observed in certain types of cancer tissues. Three types of Prx (I, II, and III) have been shown to be over-expressed in the case of human breast cancer, and it has been suggested that their over-expressions are related to cancer development or progression. The increased expression of Prx I is also detected in lung cancer, thyroid tumors and oral cancer, and is suggested to constitute a potential tumor marker.
Prx II is a human homologue of yeast thioredoxin peroxidase (TPX), also named thioredoxin-dependent peroxide reductase 1 (TDPX1) or natural killer-enhancing factor-B (NEKF-B). It is located on chromosome 13q12 and is expressed in human neurons but not in glial cells.
Prx II is an antioxidant enzyme that reduces H2O2 and other reactive oxygen species using thioredoxin as the immediate electron donor, and its peroxidase activity prevents cells from reactive oxygen species insult. Prx II is also involved in the cellular signaling pathways of growth factors and tumor necrosis factor-a, by virtue of its regulation of intracellular H2O2.
Several properties of Prx II, such as thioredoxin- dependent peroxidase activity, inhibition of apoptosis, characteristic binding to membrane, and correlation with Ca2+-activated potassium transport in erythrocytes, have been reported. In previous studies, we found that at least 3 isotypes of Prx have been identified in the skin, and Prx II was identified at the epidermal cells and epidermal appendage structures (hair follicle, eccrine gland, and sebaceous gland) of the dermis. We also found that Prx II was expressed in vascular endothelial cells in normal skin. Prx II has been reported to be present in the cytoplasm of RBCs and human umbilical endothelial cells. In a previous study, Prx II is expressed in endothelial cells of benign and malignant vascular tumors in human skin in vivo. In addition to biologic availability of Prx II as an endothelial marker, it has an advantage in detecting vascular spaces in tissue specimens because it is strongly expressed in intravascular erythrocytes.
Prx II is known not only to protect cells from oxidative damage caused by hydrogen peroxide (H2O2), but also to endow cancer cells with resistance to both H2O2 and cisplatin and to grant them radioresistance. Prx II is up-regulated by H2O2 and cisplatin treatment, and its increased expression inhibits the apoptosis induced by cisplatin, thyrotropin, serum deprivation, ceramide, or etoposide, thereby rendering tumor cells resistant to some chemotherapeutic agents. We demonstrated earlier that Prx II was involved in radioresistance.
In previous studies, we found that Prx II expression was notably down-regulated in tumor cell lines PC-3M-1E8 with high metastasis potentials compared with lowly metastatic cancer cell lines PC-3M-2B4 through two-dimensional gel electrophoresis (2-DE) and mass spectrometry analyses。To better understand the molecular mechanisms of prostate cancer and demonstrate the relativity of Prx II and prostate cancer, the function of Prx II gene was further investigated. Based on the prevalence of altered Prx II expression and function in different deseases, it was of interest to consider the therapeutic potential of controlling Prx II function or their regulated pathways through pharmacologic or genetic modulation.
Methods
Prx II mRNA and protein expression of matched-pair of prostate cancer cell lines with high and low metastasis potentials were detected by reverse transcriptase-polymerase chain reaction (RT-PCR), immunocytochemistry and western blotting . Prx II full-length eukaryotic expression vectors were constructed using the technique of gene engineering; the prostate cancer cell clones stable expressing Prx II were screened by G418. After transfection, the changes of cell proliferation and motility were detected by CFSE-flow cytometry and monolayer wound healing assay, respectively. The changes of AKT、P-AKT、E-cadherin、β-catenin and Bcl-X/L expression in 1E8/Prx II and 1E8/pcDNA3.1 cell clones were detected by RT-PCR and western blotting. The capabilities of cell growth, cell motility, in vitro invasion and cell adhesion in 1E8/Prx II and 1E8/pcDNA3.1 cell clones were performed by monolayer wound healing assay, Transwell migration assay and cell adhesion assay, respectively. In addition, To construct and identify a yeast two hybrid system bait vector for screening of homo sapiens PrxⅡbinding proteins in prostate cancer, full fragment of ORF of PrxⅡcDNA was amplified using PCR and directly ligated to the pGBKT7 vector. Insert-contained plasmid was confirmed by restriction endonuclease analysis and DNA sequencing. The plasmid was transformed into the yeast cell AH109, and its toxicity and transcriptional activation was tested by both the phenotype assay and the color assay.
Results
1. In contrast, Prx II mRNA and protein expression was notably down-regulated in prostate tumor cell line PC-3M-1E8 with high metastasis potentials compared with lowly metastatic cancer cell line PC-3M-2B4; These results indicated that abnormal-expression of Prx II gene closely correlated with neoplasm metastasis. Morever, its over-expression inhibited cancer metastasis in prostate cancer;
2. Constructed Prx II full-length eukaryotic expression vector successfully, and obtained prostate cancer cell clone(1E8/Prx II) which stably expressed Prx II. At the same time, we obtained control cell clone 1E8/pcDNA3.1.
3. After transferring Prx II gene into human prostate cancer cell line PC-3M-1E8, Prx II decreased the abilities of cell proliferation and motility of PC-3M-1E8 cells, Prx II is up-regulated by H2O2(TBHP) treatment, and its over-expression increases the apoptosis induced by TBHP. These functions might be regulated by activating PI4K/AKT intracellular signaling pathway
4. Prx II gene over-expression inhibited AKT phosphorylated at Ser473 site and regulated the recycling of internalized E-cadherin. Because of changing the function of E-cadherin /catenin complex retaining cell polarity and tissue structure, Prx II gene over-expression reversed the malignant phenotypes of PC-3M-1E8 cell lines.
5. The fragments of PrxⅡprotein were successfully obtained. The recombinant pGBKT7-PrxⅡplasmids and empty pGBKT7 vector could grow white colonies on SD/-Trp/X-α-gal plates and none could survive on SD/-His /-Trp /X-α-gal, SD/-Ade/-Trp/X-α-gal plates. The bait plasmid pGBKT7-PrxⅡconstructed expresses correctly, and can not activate the transcription of reporter gene alone.
Conclusion
Identification of PrxⅡmRNA and protein expression in matched-pairs of human prostate cancer cell lines with high and low metastasis potentials showed that abnormal-expression of PrxⅡgene closely correlated with neoplasm metastasis, moreover, in prostate cancer PrxⅡover-expression inhibited cancer metastasis and decreased the abilities of cell proliferation and motility of PC-3M-1E8 cell with high metastasis potentials. Prx II is upregulated by H2O2(TBHP) treatment, and its over-expression increases the apoptosis induced by TBHP.
To further investigate the molecular mechanism of PrxⅡin prostate cancer invasion and metastasis, the present studies showed that forced PrxⅡexpression inhibited the capabilities of cell growth and motility of Skov-3 cells through down-regulating the PI4K/AKT signaling; Over-expression of that gene inhibited AKT phosphorylated at Ser473 site and regulated the recycling of internalized E-cadherin. Because of changing the function of E-cadherin/catenin complex retaining cell polarity and tissue structure, Prx II gene over-expression reversed the malignant phenotypes of PC-3M-1E8 cell lines. Moreover, the bait plasmid pGBKT7-PrxⅡconstructed expresses correctly, and can not activate the transcription of reporter gene alone. The yeast two hybrid GAL4 system3 can be utilized to fish PrxⅡr egion interacting protein.
引文
1. Chae, HZ, Robison K, Poole LB, et al. Cloning and sequencing of thiol-specific antioxidant from mammalian brain: alkyl hydroperoxide reductase and thiol-specific antioxidant define a large family of antioxidant enzymes. Proc. Natl Acad. Sci. USA,1994,91, 7017–7021
2. Chae, HZ, Chung SJ, Rhee SG. Thioredoxin-dependent peroxide reductase from yeast. J. Biol. Chem,1994,269, 27670–27678
3. Boucher IW, McMillan PJ, Gabrielsen M,et al. Structural and biochemical characterization of a mitochondrial peroxiredoxin from Plasmodium falciparum[J]. Mol Microbiol,2006,61(4):948-959
4. Molina Lopez J, Jimenez L, Ochoa Sanchez A, et al. Molecular cloning and characterization of a 2-Cys peroxiredoxin from Taenia solium [J]. J Parasitol.,2006,92(4): 796-802
5. Chae H. Z., Kim L.-H., Kim K., and Rhe S. G. (1993) Cloning, sequencing, and mutation of thiol-specific antioxidant gene of Saccharomyces cerevisiae. J. Biol. Chem. 268: 16815–16821
6. Yim M. B., Chae H. Z., Rhee S. G., Chock P. B. and Stadtman E. R. (1994) On the protective mechanism of the thiol-specific antioxidant enzyme against the oxidative damage of biomacromolecules. J. Biol. Chem. 269: 1621–1626
7. Chae H. Z., Kim H. J., Kang S. W. and Rhee S. G. (1999) Characterization of three isoforms of mammalian peroxiredoxin that reduce peroxides in the presence of thioredoxin. Diabetes Res. Clin. Pract. 45: 101–112
8. Chen JW., Dodia C., Feinstein S I., Jai M. K. and Fisher A. B. (2000) 1-Cysperoxiredoxin, a bifunctional enzyme with glutathione peroxidase and phospholipase A2 activities. J. Biol. Chem 275: 28421–28427
9. Bryk, R.Griffin P, Nathan C. Peroxynitrite reductase activity of bacterial peroxiredoxins. Nature,2000, 407, 211–215
10 Peshenko, I.V. and Shichi, H. (2001) Oxidation of active center cysteine of bovine 1-Cys peroxiredoxin sulfenic acid form by peroxide and peroxynitrite. Free Radic. Biol. Med. 31, 292–303
11. Shau H, Merino A, Chen L, et al. Induction of peroxiredoxins in transplanted livers and demonstration of their in vitro cytoprotection activety[J]. Antioxid Redox Signal,2000,2:347-354.
12. Yim M. B., Chae H. Z., Rhee S. G., Chock P. B. and Stadtman E. R. (1994) On the protective mechanism of the thiol-specific antioxidant enzyme against the oxidative damage of biomacromolecules. J. Biol. Chem. 269: 1621–1626
13. Netto L. E. S., Chae H. Z., Kang S.-W., Rhee S. G. and Stadtman E. R. (1996) Removal of hydrogen peroxide by thiol-specific antioxidant enzyme (TSA) is involved with its antioxidant properties. J. Biol. Chem. 271: 15315–15321
14. Sarafian T. A., Rajper N., Grigorian B., Kim A. and Shau H. (1997) Cellular antioxidant properties of human natural killer enhancing factor B. Free Radic. Res. 26: 281–289
15. Lim M. J., Chae H. Z., Rhee S. G., Yu D.-Y., Lee K.-K. and Yeom Y. I. (1998) The type II peroxiredoxin gene family of the mouse: molecular structure, expression and evolution. Gene 216: 197–205
16. Kang S. W., Chae H. Z., Seo M. S., Kim K., Baines I. C. and Rhee S. G. (1998) Mammalian peroxiredoxin isoforms can reduce hydrogen peroxide generated in response to growth factors and tumor necrosis factor-a. J. Biol. Chem. 273: 6297–6302
17. Kim H., Lee T.-H., Park E. S., Suh J. M., Park S. J., Chung H. K. et al. (2000) Role of peroxideroxins in regulating intracellular hydrogen peroxide and hydrogen peroxide-induced apoptosis in thyroid cells. J. Biol. Chem. 275: 18266–18270
18. Rabilloud T, Berthier R, Vincon M, Ferbus D, Goubin G, Lawrenve J-J. Earlyevents in erythroid differentiation: accumulation of the acidic peroxiredoxin (PRP/thiol-specific antioxidant /NKEF-B). Biochem J 1995;312:699-705.
19. Lim YS, Cha MK, Yun CH, Kim HK, Kim K, Kim IH. Purification and characterization of thiol-specific antioxidant protein from human red blood cell: a new type of antioxidant protein. Biochem Biophys Res Commun 1994;199:199-206.
20. Shau H, Kim A. Identification of natural killer enhancing factor as a major antioxidant in human red blood cells. Biochem Biophys Res Commun 1994;199:83-8.
21. Lim YS, Cha MK, Kim HK, Uhm TB, Park JW, Kim K, et al. Removals of hydrogen peroxide and hydroxyl radical by thiol-specific antioxidant protein as a possible role in vivo. Biochem Biophys Res Commun 1993;192:273-80.
22. Butterfield LH, Merino A, Golb SH, Shau H. From cytoprotection to tumor suppression: the multifactorial role of peroxiredoxins. Antiox Redox Signal 1999;1:385-402
23. Chae HZ, Kang SW, Rhee SG. Isoforms of Mammalian Peroxiredoxin That Reduce Peroxides in Presence of Thioredoxin. Methods Enzymol 1999;300:219-6
24. Chae H. Z., Kim L.-H., Kim K., and Rhe S. G. (1993) Cloning, sequencing, and mutation of thiol-specific antioxidant gene of Saccharomyces cerevisiae. J. Biol. Chem. 268: 16815–16821
25. Lim Y. S., Cha M. K., Kim H. K., Uhm T. B., Park J. W., Kim K. et al. (1993) Removals of hydrogen peroxide and hydroxyl radical by thiol-specific antioxidant protein as a possible role in vivo. Biochem. Biophys. Res. Commun. 192: 273–280
26. Chae H. Z., Chung S. J. and Rhee S. G. (1994) Thioredoxin-dependent peroxide reductase from yeast. J. Biol. Chem. 269: 27670–27678
27. Chae H. Z., Robison K., Poole L. B., Church G., Storz G. and Rhee S. G. (1994) Cloning and sequencing of thiol-specific antioxidant from mammalian brain: alkyl hydroperoxide reductase and thiol-specific antioxidant define a large family of antioxidant enzymes. Proc. Natl. Acad. Sci. USA 91: 7017–7021
28. Chae H. Z., Uhm T. B. and Rhee S. G. (1994) Dimerization of thiol-specific antioxidant and the essential role of cysteine 47. Proc. Natl. Acad. Sci. USA91:7022–7026
29. Yim M. B., Chae H. Z., Rhee S. G., Chock P. B. and Stadtman E. R. (1994) On the protective mechanism of the thiol-specific antioxidant enzyme against the oxidative damage of biomacromolecules. J. Biol. Chem. 269: 1621–1626
30. Netto L. E. S., Chae H. Z., Kang S.-W., Rhee S. G. and Stadtman E. R. (1996) Removal of hydrogen peroxide by thiol-specific antioxidant enzyme (TSA) is involved with its antioxidant properties. J. Biol. Chem. 271: 15315–15321
31. Sarafian T. A., Rajper N., Grigorian B., Kim A. and Shau H. (1997) Cellular antioxidant properties of human natural killer enhancing factor B. Free Radic. Res. 26: 281–289
32. Pahl P., Berger R., Hart I., Chae H. Z., Rhee S. G. and Patterson D. (1995) Localization of TDPX1, a human homologue of the yeast thioredoxin-dependent peroxide reductase gene (TPX), to chromosome 13q12. Genomics 26: 602–606
33. Sarafian T. A., Verity M. A., Vinters H. V., Shih C. C.-Y., Shi L., Ji X. D. et al. (1999) Differential expression of peroxiredoxin subtypes in human brain cell types. J. Neurosci. Res. 56: 206–212
34. Kang SW, Bains IC, Rhee SG. Characterization of a mammalian peroxiredoxin that contains one conserved cysteine. J Biol Chem 1998;273:6303-11.
35. Chae HZ, Kang SW, Rhee SG. Isoforms of mammalian peroxiredoxin that reduce peroxides in presence of thioredoxin. Methods Enzymol 1999;300:219-26.
36. Zhang P, Liu B, Kanf SW, Seo MS, Rhee SG, Obeid LM. Thioredoxin peroxidase is a novel inhibitor of apoptosis with a mechanism distinct from that of Bcl-2. J Biol Chem 1997;272:30615-8.
37. Plishker GA, Chevalier D, Seinsoth L, Moore RB. Calcium-activated potassium transport and high molecular weight forms of calpromotin. J Biol Chem 1992;267:21839-43.
38. Schroder E, Willis AC, Ponting CP. Porcine natural-killer-enhancing factor-B: oligomerisation and identification as a calpain substrate in vitro. Biochim Biophys Acta 1998;1383:279-91.
39. Lee SC, Chae HZ, Lee JE, Kwon BD, Lee JB, Won YH, et al. Peroxiredoxin isubiquitously expressed in rat skin: isotype-specific expression in the epidermis and hair follicle. J Invest Dermatol 2000;115:1108-14.
40. Lee JB, Yun SJ, Chae HZ, Won YH, Kim YP, Lee SC. Expression of peroxiredoxin and thioredoxin in dermatological disorders. Br J Dermatol 2002;146:710-2.
41. Mitsumoto A, Takanezawa Y, Okawa K, Iwamatsu A, Nakagawa Y. Variants of peroxiredoxins expression in response to hydroperoxide stress. Free Radic Biol Med 2001;30:625-35.
42.Karihtala P, Mantyniemi A, Kang SW, Kinnula VL, Soini Y. Peroxiredoxins in Breast Carcinoma. Clin Cancer Res. 2003,15;9(9):3418-24.
43.Young DY, Young MC, Jong KP, Chul MA, Sung KK, Hyung JK. Synergistic effect of peroxiredoxin II antisense on cisplatininduced cell death[J]. Experimental and Molecular Medicine,2002;34:273-277
44. Wulfkuhle JD, Sgroi DC, Krutzsch H, et al. Proteomics of human breast ductal carcinoma in situ[J]. Cancer Research,2002;62:6740-6749
45. Keegan L, Gill G, Ptashne M. Separation of DNA binding from the transcription-activating function of a eukaryotic regulatory protein. Science,1986,231:699-704
46.Chung YM, Yoo YD, Park JK, Kim YT, Kim HJ. Increased expression of peroxiredoxin II confers resistance to cisplatin. Anticancer Res 2001;21:1129-34
47.Kim H, Lee TH, Park ES, Suh JM, Park SJ, Chung HK, Kwon OY, Kim YK, Ro HK, Shong M. Role of peroxiredoxins in regulating intracellular hydrogen peroxide and hydrogen peroxide-induced apoptosis in thyroid cells. J Biol Chem 2000;275:18266-70
48.Zhang P, Liu B, Kang SW, Seo MS, Rhee SG, Obeid LM. Thioredoxin peroxidase is a novel inhibitor of apoptosis with a mechanism distinct from that of Bcl-2. J Biol Chem 1997;272: 30615-8
49.Park SH, Chung YM, Lee YS, Kim HJ, Kim JS, Chae HZ, Yoo YD. Antisense of human peroxiredoxin II enhances radiationinduced cell death. Clin Cancer Res 2000;6:4915-20
50. Wu MF, Bai XY, Xu G, et al. Proteome analysis of human androgen-independent prostate cancer cell lines: Variable metastatic potentials correlated with vimentin expression.Proteomics,2007,in print
51. Chen CD , Welsbie DS , Tran C , et al. Molecular determinants of resistance to anti androgen therapy. Nat Med , 2004 , 10∶33
52. Martel CL,Gumerlock PH,Meyers FJ,et al.current strategies in the treatment ofhormone refractory prostate cancer[J]. Cancer Treat Rev,2003;29:171-187
53. Liu Yuxin, Zheng Jie, Fang Weigang, You Jiangfeng, Wang Jieliang, Cui Xianglin, Wu Bingquan, et al. Isolation and characterization of human prostate cancer cell subclones with different metastatic potential.[J]. Chin J Pathol, 1999,28:361-364
54. Fields S, Song O. A novel genetic system to detect protein-protein interactions[J]. Nature,1989,340(6230):245-246
55. Jemal A ,Thomas A ,Murray T , et al . Cancer statistics ,2002 [J ] . CA Cancer J Clin ,2002 ,52(1) :23-47.
56. Bubley GJ , Carducci M, and Dahut W. Eligibility and response guidelines for phase II clinical trials in androgen independent prostate cancer : Recommendations from the prostate specific antigen working group. J Clin Oncol , 1999 , 17∶3461
57. Beck MA. Selenium and host defence towards viruses. Proc Nutr Soc,1999,58(3):707-711
58. Ozaki M, Deshpande SS, Angkeow P, et al. Inhibition of the Rac1 GT-Pase protects against nonlethal ischrmia /reperfusion-induced necrosis and apoptosis in vivo. FASEB J,2000,14:418-429
1. Valster A, Tran NL, Nakda M, et al. Cell migration and invasion assays[J]. Methods, 2005, 37(2):208-215.
2. Eposti, M. Measuring mitochondrial reactive oxygen species. Methods Enzymol., 26: 335– 340, 2002.
3. Chae H. Z., Kim L.-H., Kim K., and Rhe S. G. (1993) Cloning, sequencing, and mutation of thiol-specific antioxidant gene of Saccharomyces cerevisiae. J. Biol. Chem. 268: 16815–16821
4. Lim Y. S., Cha M. K., Kim H. K., Uhm T. B., Park J. W., Kim K. et al. (1993) Removals of hydrogen peroxide and hydroxyl radical by thiol-specific antioxidantprotein as a possible role in vivo. Biochem. Biophys. Res. Commun. 192: 273–280
5. Chae H. Z., Chung S. J. and Rhee S. G. (1994) Thioredoxin-dependent peroxide reductase from yeast. J. Biol. Chem. 269: 27670–27678
6. Chae H. Z., Robison K., Poole L. B., Church G., Storz G. and Rhee S. G. (1994) Cloning and sequencing of thiol-specific antioxidant from mammalian brain: alkyl hydroperoxide reductase and thiol-specific antioxidant define a large family of antioxidant enzymes. Proc. Natl. Acad. Sci. USA 91: 7017–7021
7. Chae H. Z., Uhm T. B. and Rhee S. G. (1994) Dimerization of thiol-specific antioxidant and the essential role of cysteine 47. Proc. Natl. Acad. Sci. USA 91:7022–7026
8. Yim M. B., Chae H. Z., Rhee S. G., Chock P. B. and Stadtman E. R. (1994) On the protective mechanism of the thiol-specific antioxidant enzyme against the oxidative damage of biomacromolecules. J. Biol. Chem. 269: 1621–1626
9. Netto L. E. S., Chae H. Z., Kang S.-W., Rhee S. G. and Stadtman E. R. (1996) Removal of hydrogen peroxide by thiol-specific antioxidant enzyme (TSA) is involved with its antioxidant properties. J. Biol. Chem. 271: 15315–15321
10. Sarafian T. A., Rajper N., Grigorian B., Kim A. and Shau H. (1997) Cellular antioxidant properties of human natural killer enhancing factor B. Free Radic. Res. 26: 281–289
11. Pahl P., Berger R., Hart I., Chae H. Z., Rhee S. G. and Patterson D. (1995) Localization of TDPX1, a human homologue of the yeast thioredoxin-dependent peroxide reductase gene (TPX), to chromosome 13q12. Genomics 26: 602–606
12. Sarafian T. A., Verity M. A., Vinters H. V., Shih C. C.-Y., Shi L., Ji X. D. et al. (1999) Differential expression of peroxiredoxin subtypes in human brain cell types. J. Neurosci. Res. 56: 206–212
13. Kang SW, Bains IC, Rhee SG. Characterization of a mammalian peroxiredoxin that contains one conserved cysteine. J Biol Chem 1998;273:6303-11.
14. Chae HZ, Kang SW, Rhee SG. Isoforms of mammalian peroxiredoxin that reduce peroxides in presence of thioredoxin. Methods Enzymol 1999;300:219-26.
15. Zhang P, Liu B, Kanf SW, Seo MS, Rhee SG, Obeid LM. Thioredoxinperoxidase is a novel inhibitor of apoptosis with a mechanism distinct from that of Bcl-2. J Biol Chem 1997;272:30615-8.
16. Plishker GA, Chevalier D, Seinsoth L, Moore RB. Calcium-activated potassium transport and high molecular weight forms of calpromotin. J Biol Chem 1992;267:21839-43.
17. Schroder E, Willis AC, Ponting CP. Porcine natural-killer-enhancing factor-B: oligomerisation and identification as a calpain substrate in vitro. Biochim Biophys Acta 1998;1383:279-91.
18. Lee SC, Chae HZ, Lee JE, Kwon BD, Lee JB, Won YH, et al. Peroxiredoxin is ubiquitously expressed in rat skin: isotype-specific expression in the epidermis and hair follicle. J Invest Dermatol 2000;115:1108-14.
19. Lee JB, Yun SJ, Chae HZ, Won YH, Kim YP, Lee SC. Expression of peroxiredoxin and thioredoxin in dermatological disorders. Br J Dermatol 2002;146:710-2.
20. Mitsumoto A, Takanezawa Y, Okawa K, Iwamatsu A, Nakagawa Y. Variants of peroxiredoxins expression in response to hydroperoxide stress. Free Radic Biol Med 2001;30:625-35.
21. Rabilloud T, Berthier R, Vincon M, Ferbus D, Goubin G, Lawrenve J-J. Early events in erythroid differentiation: accumulation of the acidic peroxiredoxin (PRP/thiol-specific antioxidant /NKEF-B). Biochem J 1995;312:699-705.
22. Lim YS, Cha MK, Yun CH, Kim HK, Kim K, Kim IH. Purification and characterization of thiol-specific antioxidant protein from human red blood cell: a new type of antioxidant protein. Biochem Biophys Res Commun 1994;199:199-206.
23. Shau H, Kim A. Identification of natural killer enhancing factor as a major antioxidant in human red blood cells. Biochem Biophys Res Commun 1994;199:83-8.
24.Karihtala P, Mantyniemi A, Kang SW, Kinnula VL, Soini Y. Peroxiredoxins in Breast Carcinoma. Clin Cancer Res. 2003,15;9(9):3418-24.
25.Young DY, Young MC, Jong KP, Chul MA, Sung KK, Hyung JK. Synergistic effect of peroxiredoxin II antisense on cisplatininduced cell death[J]. Experimentaland Molecular Medicine,2002;34:273-277
26. Wulfkuhle JD, Sgroi DC, Krutzsch H, et al. Proteomics of human breast ductal carcinoma in situ[J]. Cancer Research,2002;62:6740-6749
27. Keegan L, Gill G, Ptashne M. Separation of DNA binding from the transcription-activating function of a eukaryotic regulatory protein. Science,1986,231:699-704
28.Chung YM, Yoo YD, Park JK, Kim YT, Kim HJ. Increased expression of peroxiredoxin II confers resistance to cisplatin. Anticancer Res 2001;21:1129-34
29.Kim H, Lee TH, Park ES, Suh JM, Park SJ, Chung HK, Kwon OY, Kim YK, Ro HK, Shong M. Role of peroxiredoxins in regulating intracellular hydrogen peroxide and hydrogen peroxide-induced apoptosis in thyroid cells. J Biol Chem 2000;275:18266-70
30.Zhang P, Liu B, Kang SW, Seo MS, Rhee SG, Obeid LM. Thioredoxin peroxidase is a novel inhibitor of apoptosis with a mechanism distinct from that of Bcl-2. J Biol Chem 1997;272: 30615-8
31.Park SH, Chung YM, Lee YS, Kim HJ, Kim JS, Chae HZ, Yoo YD. Antisense of human peroxiredoxin II enhances radiationinduced cell death. Clin Cancer Res 2000;6:4915-20
32. Beck MA. Selenium and host defence towards viruses. Proc Nutr Soc,1999,58(3):707)711
33. Ozaki M, Deshpande SS, Angkeow P, et al. Inhibition of the Rac1 GT-Pase protects against nonlethal ischrmia /reperfusion-induced necrosis and apoptosis in vivo. FASEB J,2000,14:418-429
34. Lennon SV, Martin SJ, Cotter TG Does-dependent induction of apoptosis in human tumor or cell lines by widely diverging stiumuli. Cell prolif,1991,24:203-214
35.高颖,许彩民,杨洋.氧化诱导K652细胞凋亡机制的初步探讨.中国生物化学与分子生物学学报,1998,14(3):295-299
36. Ishisaka R, Utsumi K, Utsumi T. Involvement of lysosomal cysteine protease in hydrogen peroxide - induced apoptosis in HL - 60 cells. Biosci Biotechnol Biochem,2002,66(9):1865-1872
37. Aruoma, O. I.; Halliwell, B.; Gajewski, E.; Dizdaroglu, M. Copperion-dependent damage to the bases in DNA in the presence of hydrogen peroxide. Biochem. J. 1991,273:601– 604;.
38. Termini, J. Hydroperoxide-induced DNA damage and mutations. Mutat. Res. 2000.450:107– 124;
39. Guidarelli, A.; Cattabeni, F.; Cantoni, O. Alternative mechanisms for hydroperoxide-induced DNA single strand breakage. Free Radic. Res. 1997.26:537– 547;
40. Cooke, M. S.; Evans, M. D.; Dizdaroglu, M.; Lunec, J. Oxidative DNA damage: mechanisms, mutation and disease. FASEB J. 2003.17:1195– 1214;
41.J in X , Han C S , Yu F Q , et al . Anti apoptic action of stem cell fact or on oocytes in primordial follicles and its signal transduction[J ] . Mol Rep rod Dev ,2005 ,70 (1) :82290.
42. Hu X M , Christian P , Sipes I G , et al . Exp ression and redist ribution of cellular Bad , Bax , and Bcl-X/L p rotein is associated wit h VCD2induced ovot oxicity in rats [ J ] . Biol Rep rod , 2001 ,65 (5) :148921495.
43. Yan W , Samson M , J egou B , et al . Bak/ Bcl2W cor resp ond to spermat ogonial and spermat ocyte ap opt os Bcl2W forms complexes wit h Bax and Bak , and elevated ratio of Bak/ Bcl2W and is in t he testis [J ] . Mol Endocrinol , 2000 ,14 (5) :6822699.
44. Kimura M , It oh N , Takagi S , et al . Balance of ap opt osis and prolif eration of germ cells related t o spermat ogenesis in aged men[J ] . J Androl ,2003 ,24 (2) :185291.
45. Lionel Larue,Alfonso Bellacosa. Epithelial–mesenchymal transition in development and cancer: role of phosphatidylinositol 3 kinase/AKT pathways[J]. Oncogene, 2005,24, 7443-7454.
46. Sylvia Julien Grille, Alfonso Bellacosa, John Upson, et al. The Protein Kinase Akt Induces Epithelial Mesenchymal Transition and Promotes Enhanced Motility and Invasiveness of Squamous Cell Carcinoma Lines[J]. Cancer Research, 2003,63, 2172–2178.
47. Felipe Palacios, Jogender S. Tushir, Yasuyuki Fujita, et al. Lysosomal Targeting of E-Cadherin: a Unique Mechanism for the Down-Regulation of Cell-Cell Adhesion during Epithelial to Mesenchymal Transitions[J]. Molecular and Cell Biology, 2005,25(1):389-402.
1 Chae, HZ, Robison K, Poole LB, et al. Cloning and sequencing of thiol-specific antioxidant from mammalian brain: alkyl hydroperoxide reductase and thiol-specific antioxidant define a large family of antioxidant enzymes[J]. Proc. Natl Acad. Sci. USA,1994,91:7017–7021
2 Chae, HZ, Chung SJ, Rhee SG. Thioredoxin-dependent peroxide reductase from yeast[J]. J. Biol. Chem,1994,269:27670–27678
3 Bryk, R.Griffin P, Nathan C. Peroxynitrite reductase activity of bacterial peroxiredoxins[J]. Nature,2000, 407: 211–215
4.Peshenko IV, Shichi H. Oxidation of active center cysteine of bovine 1-Cys peroxiredoxin sulfenic acid form by peroxide and peroxynitrite[J]. Free Radic Biol Med,2001, 31: 292–303
5.Shau H, Merino A, Chen L, et al. Induction of peroxiredoxins in transplanted livers and demonstration of their in vitro cytoprotection activety[J]. Antioxid Redox Signal,2000,2:347-354.
6.Liu Yuxin, Zheng Jie, Fang Weigang, You Jiangfeng, Wang Jieliang, Cui Xianglin, Wu Bingquan, et al. Isolation and characterization of human prostate cancer cell subclones with different metastatic potential.[J]. Chin J Pathol, 1999,28:361-364
7. Bartel PL, Fields S. The yeast two-hybrid system[M]. New York: Oxford University Press, Oxford,1997:425-428
8. Schiestl RH, Gietz RD. High efficiency transformation of intact yeast cells using single stranded nucleic acids as a carrier.[J] Curr Genet,1989,16(5-6):339-346
9. Printen JA, Sprague GF Jr. Protein-protein interactions in the yeast pheromone response pathway: Ste5p interacts with all members of the MAP kinase cascade[J].Genetics, 1994,138(3):609-19.
10. Schiestl RH, Gietz RD. High efficiency transformation of intact yeast cells using single stranded nucleic acids as a carrier.[J] Curr Genet,1989,16:339-346.
11. Boucher IW, McMillan PJ, Gabrielsen M,et al. Structural and biochemical characterization of a mitochondrial peroxiredoxin from Plasmodium falciparum[J]. Mol Microbiol,2006,61(4):948-959
12. Molina Lopez J, Jimenez L, Ochoa Sanchez A, et al. Molecular cloning and characterization of a 2-Cys peroxiredoxin from Taenia solium [J]. J Parasitol.,2006,92(4): 796-802
13. Zhang P, Liu B, Kanf SW, Seo MS, Rhee SG, Obeid LM. Thioredoxin peroxidase is a novel inhibitor of apoptosis with a mechanism distinct from that of Bcl-2.[J]. J Biol Chem,1997,272:30615-8.
14. Noh DY, Ahn SJ, Lee RA, Kim SW, Park ZA, Chae HZ. Overexpression of peroxiredoxin in human breast cancer[J]. Anticancer Res,2001,21:2085-90.
15. Okazaki S, Naganuma A, Kuge S. Peroxiredoxin-mediated redox regulation of the nuclear localization of Yap1, a transcription factor in budding yeast[J].Antioxid Redox Signal,2005,7(3-4): 327-334
16. Plishker GA, Chevalier D, Seinsoth L, Moore RB. Calcium-activated potassiumtransport and high molecular weight forms of calpromotin[J]. J Biol Chem,1992,267:21839-43.
17. Schroder E, Willis AC, Ponting CP. Porcine natural-killer-enhancing factor-B: oligomerisation and identification as a calpain substrate in vitro[J]. Biochim Biophys Acta,1998,1383:279-91.
18. Lee SC, Na YP, Lee JB. Expression of peroxiredoxin II in vascular tumors of the skin: A novel vascular marker endothelial cells[J]. J Am Acad Dermatol,2003,49(3):487-491
19.Karihtala P, Mantyniemi A, Kang SW, Kinnula VL, Soini Y. Peroxiredoxins in Breast Carcinoma[J]. Clin Cancer Res,2003,15;9(9):3418-24.
20. Young DY, Young MC, Jong KP, Chul MA, Sung KK, Hyung JK. Synergistic effect of peroxiredoxin II antisense on cisplatininduced cell death[J]. Exp Mol Med,2002,34(4):273-277
21. Wulfkuhle JD, Sgroi DC, Krutzsch H, et al. Proteomics of human breast ductal carcinoma in situ[J]. Cancer Research,2002,62:6740-6749
22. Fields S, Song O. A novel genetic system to detect protein-protein interactions[J]. Nature,1989,340(6230):245-246
23.Keegan L, Gill G, Ptashne M. Separation of DNA binding from the transcription-activating function of a eukaryotic regulatory protein[J]. Science,1986,231:699-704
1. Chen CD , Welsbie DS , Tran C , et al. Molecular determinants of resistance to anti androgen therapy. Nat Med , 2004 , 10∶33
2. Bubley GJ , Carducci M, and Dahut W. Eligibility and response guidelines for phase II clinical trials in androgen independent prostate cancer : Recommendations from the prostate specific antigen working group. J Clin Oncol , 1999 , 17∶3461
3. McDonnell , TJ . Expression of the protooncogene bcl-2 in the prostate and its association with emergence of androgen independent prostate cancer. Cancer Res , 1992 , 52 , 6940
4. Yeh S , et al. From HER22/ Neu signal cascade to androgen receptor and its coactivators : A novel pathway by induction of androgen target genes through MAP kinase in prostate cancer cells.Proc Natl Acad Sci USA , 1999 , 96∶5458
5. Putz T, et al. Epidermal growth factor ( EGF) receptor blockade inhibits the action of EGF , insulin2like growth factor I , and a protein kinase A activator on the mitogen2activated protein kinase pathway in prostate cancer cell lines. Cancer Res , 1999 ,59∶227
6. Sherwood ER , et al. Epidermal growth factor receptor activation in androgen independent but not androgen stimulated growth of human prostatic carcinoma cells. Br J Cancer , 1998 , 77∶855
7. Craft N , Shostak Y, Carey M, et al. Mechanism for hormone independent prostate cancer through modulation of androgen receptor signaling by the HER - 2/ neu tyrosine kinase. Nature Med , 1999 , 5∶280
8. Yeh S. et al. From HER2/ Neu signal cascade to androgen receptor and its coactivators : A novel pathway by induction of androgen target genes through MAP kinase in prostate cancer cells. Proc Natl Acad Sci USA , 1999 , 96∶5458
9. Wen Y. et al. HER - 2/ neu promotes androgen independent survival and growth of prostate cancer cells through the AKT path way. Cancer Res , 2000 , 60∶6841
10. Zhou BP , et al. HER - 2/ neu blocks tumor necrosis factor in duced apoptosis via the AKT/ NF-kB pathway. J Biol Chem , 2000 , 275∶8027
11. McMenamin ME , Soung P , Perera S , et al. Loss of PTEN expression inparaffin embedded primary prostate cancer correlates with high Gleason score and advanced stage. Cancer Res , 1999 , 59∶4291
12. Vivanco I , Sawyers CL. The phosphatidylinositol 3– kinase AKT pathway in human cancer. Nat Rev Cancer , 2002 , 2∶489
13. Fizazi K,Martinez LA ,Sikes CR , et al . The association of p21 (WAF21/ CIPI)with progression to androgen2independent prostate cancer [J ] .Clin Cancer Res ,2002 ,8 (3) :775-781.
14. Taplin ME ,Ho SM. Clinical review 134 :The endocrinology of prostate cancer[J ] . J Clin Endocrinol Metab ,2001 ,86(8) :3467-3477.
15. Agus DB ,Cordon2Cardo C , Fox W, et al . Prostate cancer cell cycle regulators :response to androgen withdrawal and development of androgen independence[J ] . J Natl Cancer Inst ,1999 ,91(21) :1869-1876.
16. Gao X,Chen YQ ,Wu N , et al . Somatic mutations of the WAF1/ CIP1 gene in primary prostate cancer [J ] . Oncogene ,1995 ,11 (7) : 1395 -1398.
17. Chang BD ,Watanabe K,Broude EV, et al . Effects of p21Waf1/ Cip1/Sdi1 on cellular gene expression : implications for carcinogenesis ,senescence ,and age2related diseases[J ] . Proc Natl Acad Sci U S A ,2000 ,97 (8) :4291-4296.
18. Ling MT ,Wang X,Lee DT , et al . Id21 expression induces androgen independent prostate cancer cell growth through activation of epidermal growth factor receptor ( EGF2R) [ J ] . Carcinogenesis , 2004 , 25 (4) :517-525.
19. Palayoor ST , Youmell MY, Calderwood SK, et al . Constitutive activation of Ikappa B kinase alpha and NF-kappa B in prostate cancer cells is inhibited by ibuprofen[J ] . Oncogene , 1999 , 18 (51) :7389-7394.
20. Kimura K, Markowski M, Bowen C , et al . Androgen blocks apoptosis of hormone2dependent prostate cancer cells [ J ] . Cancer Res ,2001 , 61 (14) :5611-5618.
21. Chen CD , Sawyers CL. NF-kappa B activates prostate-specific antigen expression and is upregulated in androgen-independent prostate cancer[J ] . Mol Cell Biol , 2002 , 22 (8) :2862-2870.
22. Kasof GM, Lu JJ , Liu D , et al . Tumor necrosis factor-alpha induces theexpression of DR6 , a member of the TNF receptor family , through activation of NF-kappa B[J ] . Oncogene , 2001 , 20 (55) :7965-7975.
23. Yu R , Mandlekar S , Ruben S , et al . Tumor necrosis factor-related apoptosis-inducing ligand-mediated apoptosis in androgen-independent prostate cancer cells [J ] . Cancer Res , 2000 , 60(9) :2384-2389.
24. Catz SD , Johnson JL. Transcriptional regulation of Bcl-2 by nuclear factor kappa B and its significance in prostate cancer[J ] . Oncogene , 2001 , 20 (50) :7342-7351.
25. Huang S , Pettaway CA , Uehara H , et al . Blockade of NF-kappa B activity in human prostate cancer cells is associated with suppression of angiogenesis, invasion , and metastasis [ J ] . Oncogene , 2001 , 20(31) :4188-4197.
26. Lindholm PF , Bub J , Kaul S , et al . The role of constitutive NF-kappa B activity in PC-3 human prostate cancer cell invasive behavior [J ] . Clin Exp Metastasis , 2000 , 18 (6) :471-479.
2. Chae, HZ, Chung SJ, Rhee SG. Thioredoxin-dependent peroxide reductase from yeast. J. Biol. Chem,1994,269, 27670–27678
3. Boucher IW, McMillan PJ, Gabrielsen M,et al. Structural and biochemical characterization of a mitochondrial peroxiredoxin from Plasmodium falciparum[J]. Mol Microbiol,2006,61(4):948-959
4. Molina Lopez J, Jimenez L, Ochoa Sanchez A, et al. Molecular cloning and characterization of a 2-Cys peroxiredoxin from Taenia solium [J]. J Parasitol.,2006,92(4): 796-802
5. Chae H. Z., Kim L.-H., Kim K., and Rhe S. G. (1993) Cloning, sequencing, and mutation of thiol-specific antioxidant gene of Saccharomyces cerevisiae. J. Biol. Chem. 268: 16815–16821
6. Yim M. B., Chae H. Z., Rhee S. G., Chock P. B. and Stadtman E. R. (1994) On the protective mechanism of the thiol-specific antioxidant enzyme against the oxidative damage of biomacromolecules. J. Biol. Chem. 269: 1621–1626
7. Chae H. Z., Kim H. J., Kang S. W. and Rhee S. G. (1999) Characterization of three isoforms of mammalian peroxiredoxin that reduce peroxides in the presence of thioredoxin. Diabetes Res. Clin. Pract. 45: 101–112
8. Chen JW., Dodia C., Feinstein S I., Jai M. K. and Fisher A. B. (2000) 1-Cysperoxiredoxin, a bifunctional enzyme with glutathione peroxidase and phospholipase A2 activities. J. Biol. Chem 275: 28421–28427
9. Bryk, R.Griffin P, Nathan C. Peroxynitrite reductase activity of bacterial peroxiredoxins. Nature,2000, 407, 211–215
10 Peshenko, I.V. and Shichi, H. (2001) Oxidation of active center cysteine of bovine 1-Cys peroxiredoxin sulfenic acid form by peroxide and peroxynitrite. Free Radic. Biol. Med. 31, 292–303
11. Shau H, Merino A, Chen L, et al. Induction of peroxiredoxins in transplanted livers and demonstration of their in vitro cytoprotection activety[J]. Antioxid Redox Signal,2000,2:347-354.
12. Yim M. B., Chae H. Z., Rhee S. G., Chock P. B. and Stadtman E. R. (1994) On the protective mechanism of the thiol-specific antioxidant enzyme against the oxidative damage of biomacromolecules. J. Biol. Chem. 269: 1621–1626
13. Netto L. E. S., Chae H. Z., Kang S.-W., Rhee S. G. and Stadtman E. R. (1996) Removal of hydrogen peroxide by thiol-specific antioxidant enzyme (TSA) is involved with its antioxidant properties. J. Biol. Chem. 271: 15315–15321
14. Sarafian T. A., Rajper N., Grigorian B., Kim A. and Shau H. (1997) Cellular antioxidant properties of human natural killer enhancing factor B. Free Radic. Res. 26: 281–289
15. Lim M. J., Chae H. Z., Rhee S. G., Yu D.-Y., Lee K.-K. and Yeom Y. I. (1998) The type II peroxiredoxin gene family of the mouse: molecular structure, expression and evolution. Gene 216: 197–205
16. Kang S. W., Chae H. Z., Seo M. S., Kim K., Baines I. C. and Rhee S. G. (1998) Mammalian peroxiredoxin isoforms can reduce hydrogen peroxide generated in response to growth factors and tumor necrosis factor-a. J. Biol. Chem. 273: 6297–6302
17. Kim H., Lee T.-H., Park E. S., Suh J. M., Park S. J., Chung H. K. et al. (2000) Role of peroxideroxins in regulating intracellular hydrogen peroxide and hydrogen peroxide-induced apoptosis in thyroid cells. J. Biol. Chem. 275: 18266–18270
18. Rabilloud T, Berthier R, Vincon M, Ferbus D, Goubin G, Lawrenve J-J. Earlyevents in erythroid differentiation: accumulation of the acidic peroxiredoxin (PRP/thiol-specific antioxidant /NKEF-B). Biochem J 1995;312:699-705.
19. Lim YS, Cha MK, Yun CH, Kim HK, Kim K, Kim IH. Purification and characterization of thiol-specific antioxidant protein from human red blood cell: a new type of antioxidant protein. Biochem Biophys Res Commun 1994;199:199-206.
20. Shau H, Kim A. Identification of natural killer enhancing factor as a major antioxidant in human red blood cells. Biochem Biophys Res Commun 1994;199:83-8.
21. Lim YS, Cha MK, Kim HK, Uhm TB, Park JW, Kim K, et al. Removals of hydrogen peroxide and hydroxyl radical by thiol-specific antioxidant protein as a possible role in vivo. Biochem Biophys Res Commun 1993;192:273-80.
22. Butterfield LH, Merino A, Golb SH, Shau H. From cytoprotection to tumor suppression: the multifactorial role of peroxiredoxins. Antiox Redox Signal 1999;1:385-402
23. Chae HZ, Kang SW, Rhee SG. Isoforms of Mammalian Peroxiredoxin That Reduce Peroxides in Presence of Thioredoxin. Methods Enzymol 1999;300:219-6
24. Chae H. Z., Kim L.-H., Kim K., and Rhe S. G. (1993) Cloning, sequencing, and mutation of thiol-specific antioxidant gene of Saccharomyces cerevisiae. J. Biol. Chem. 268: 16815–16821
25. Lim Y. S., Cha M. K., Kim H. K., Uhm T. B., Park J. W., Kim K. et al. (1993) Removals of hydrogen peroxide and hydroxyl radical by thiol-specific antioxidant protein as a possible role in vivo. Biochem. Biophys. Res. Commun. 192: 273–280
26. Chae H. Z., Chung S. J. and Rhee S. G. (1994) Thioredoxin-dependent peroxide reductase from yeast. J. Biol. Chem. 269: 27670–27678
27. Chae H. Z., Robison K., Poole L. B., Church G., Storz G. and Rhee S. G. (1994) Cloning and sequencing of thiol-specific antioxidant from mammalian brain: alkyl hydroperoxide reductase and thiol-specific antioxidant define a large family of antioxidant enzymes. Proc. Natl. Acad. Sci. USA 91: 7017–7021
28. Chae H. Z., Uhm T. B. and Rhee S. G. (1994) Dimerization of thiol-specific antioxidant and the essential role of cysteine 47. Proc. Natl. Acad. Sci. USA91:7022–7026
29. Yim M. B., Chae H. Z., Rhee S. G., Chock P. B. and Stadtman E. R. (1994) On the protective mechanism of the thiol-specific antioxidant enzyme against the oxidative damage of biomacromolecules. J. Biol. Chem. 269: 1621–1626
30. Netto L. E. S., Chae H. Z., Kang S.-W., Rhee S. G. and Stadtman E. R. (1996) Removal of hydrogen peroxide by thiol-specific antioxidant enzyme (TSA) is involved with its antioxidant properties. J. Biol. Chem. 271: 15315–15321
31. Sarafian T. A., Rajper N., Grigorian B., Kim A. and Shau H. (1997) Cellular antioxidant properties of human natural killer enhancing factor B. Free Radic. Res. 26: 281–289
32. Pahl P., Berger R., Hart I., Chae H. Z., Rhee S. G. and Patterson D. (1995) Localization of TDPX1, a human homologue of the yeast thioredoxin-dependent peroxide reductase gene (TPX), to chromosome 13q12. Genomics 26: 602–606
33. Sarafian T. A., Verity M. A., Vinters H. V., Shih C. C.-Y., Shi L., Ji X. D. et al. (1999) Differential expression of peroxiredoxin subtypes in human brain cell types. J. Neurosci. Res. 56: 206–212
34. Kang SW, Bains IC, Rhee SG. Characterization of a mammalian peroxiredoxin that contains one conserved cysteine. J Biol Chem 1998;273:6303-11.
35. Chae HZ, Kang SW, Rhee SG. Isoforms of mammalian peroxiredoxin that reduce peroxides in presence of thioredoxin. Methods Enzymol 1999;300:219-26.
36. Zhang P, Liu B, Kanf SW, Seo MS, Rhee SG, Obeid LM. Thioredoxin peroxidase is a novel inhibitor of apoptosis with a mechanism distinct from that of Bcl-2. J Biol Chem 1997;272:30615-8.
37. Plishker GA, Chevalier D, Seinsoth L, Moore RB. Calcium-activated potassium transport and high molecular weight forms of calpromotin. J Biol Chem 1992;267:21839-43.
38. Schroder E, Willis AC, Ponting CP. Porcine natural-killer-enhancing factor-B: oligomerisation and identification as a calpain substrate in vitro. Biochim Biophys Acta 1998;1383:279-91.
39. Lee SC, Chae HZ, Lee JE, Kwon BD, Lee JB, Won YH, et al. Peroxiredoxin isubiquitously expressed in rat skin: isotype-specific expression in the epidermis and hair follicle. J Invest Dermatol 2000;115:1108-14.
40. Lee JB, Yun SJ, Chae HZ, Won YH, Kim YP, Lee SC. Expression of peroxiredoxin and thioredoxin in dermatological disorders. Br J Dermatol 2002;146:710-2.
41. Mitsumoto A, Takanezawa Y, Okawa K, Iwamatsu A, Nakagawa Y. Variants of peroxiredoxins expression in response to hydroperoxide stress. Free Radic Biol Med 2001;30:625-35.
42.Karihtala P, Mantyniemi A, Kang SW, Kinnula VL, Soini Y. Peroxiredoxins in Breast Carcinoma. Clin Cancer Res. 2003,15;9(9):3418-24.
43.Young DY, Young MC, Jong KP, Chul MA, Sung KK, Hyung JK. Synergistic effect of peroxiredoxin II antisense on cisplatininduced cell death[J]. Experimental and Molecular Medicine,2002;34:273-277
44. Wulfkuhle JD, Sgroi DC, Krutzsch H, et al. Proteomics of human breast ductal carcinoma in situ[J]. Cancer Research,2002;62:6740-6749
45. Keegan L, Gill G, Ptashne M. Separation of DNA binding from the transcription-activating function of a eukaryotic regulatory protein. Science,1986,231:699-704
46.Chung YM, Yoo YD, Park JK, Kim YT, Kim HJ. Increased expression of peroxiredoxin II confers resistance to cisplatin. Anticancer Res 2001;21:1129-34
47.Kim H, Lee TH, Park ES, Suh JM, Park SJ, Chung HK, Kwon OY, Kim YK, Ro HK, Shong M. Role of peroxiredoxins in regulating intracellular hydrogen peroxide and hydrogen peroxide-induced apoptosis in thyroid cells. J Biol Chem 2000;275:18266-70
48.Zhang P, Liu B, Kang SW, Seo MS, Rhee SG, Obeid LM. Thioredoxin peroxidase is a novel inhibitor of apoptosis with a mechanism distinct from that of Bcl-2. J Biol Chem 1997;272: 30615-8
49.Park SH, Chung YM, Lee YS, Kim HJ, Kim JS, Chae HZ, Yoo YD. Antisense of human peroxiredoxin II enhances radiationinduced cell death. Clin Cancer Res 2000;6:4915-20
50. Wu MF, Bai XY, Xu G, et al. Proteome analysis of human androgen-independent prostate cancer cell lines: Variable metastatic potentials correlated with vimentin expression.Proteomics,2007,in print
51. Chen CD , Welsbie DS , Tran C , et al. Molecular determinants of resistance to anti androgen therapy. Nat Med , 2004 , 10∶33
52. Martel CL,Gumerlock PH,Meyers FJ,et al.current strategies in the treatment ofhormone refractory prostate cancer[J]. Cancer Treat Rev,2003;29:171-187
53. Liu Yuxin, Zheng Jie, Fang Weigang, You Jiangfeng, Wang Jieliang, Cui Xianglin, Wu Bingquan, et al. Isolation and characterization of human prostate cancer cell subclones with different metastatic potential.[J]. Chin J Pathol, 1999,28:361-364
54. Fields S, Song O. A novel genetic system to detect protein-protein interactions[J]. Nature,1989,340(6230):245-246
55. Jemal A ,Thomas A ,Murray T , et al . Cancer statistics ,2002 [J ] . CA Cancer J Clin ,2002 ,52(1) :23-47.
56. Bubley GJ , Carducci M, and Dahut W. Eligibility and response guidelines for phase II clinical trials in androgen independent prostate cancer : Recommendations from the prostate specific antigen working group. J Clin Oncol , 1999 , 17∶3461
57. Beck MA. Selenium and host defence towards viruses. Proc Nutr Soc,1999,58(3):707-711
58. Ozaki M, Deshpande SS, Angkeow P, et al. Inhibition of the Rac1 GT-Pase protects against nonlethal ischrmia /reperfusion-induced necrosis and apoptosis in vivo. FASEB J,2000,14:418-429
1. Valster A, Tran NL, Nakda M, et al. Cell migration and invasion assays[J]. Methods, 2005, 37(2):208-215.
2. Eposti, M. Measuring mitochondrial reactive oxygen species. Methods Enzymol., 26: 335– 340, 2002.
3. Chae H. Z., Kim L.-H., Kim K., and Rhe S. G. (1993) Cloning, sequencing, and mutation of thiol-specific antioxidant gene of Saccharomyces cerevisiae. J. Biol. Chem. 268: 16815–16821
4. Lim Y. S., Cha M. K., Kim H. K., Uhm T. B., Park J. W., Kim K. et al. (1993) Removals of hydrogen peroxide and hydroxyl radical by thiol-specific antioxidantprotein as a possible role in vivo. Biochem. Biophys. Res. Commun. 192: 273–280
5. Chae H. Z., Chung S. J. and Rhee S. G. (1994) Thioredoxin-dependent peroxide reductase from yeast. J. Biol. Chem. 269: 27670–27678
6. Chae H. Z., Robison K., Poole L. B., Church G., Storz G. and Rhee S. G. (1994) Cloning and sequencing of thiol-specific antioxidant from mammalian brain: alkyl hydroperoxide reductase and thiol-specific antioxidant define a large family of antioxidant enzymes. Proc. Natl. Acad. Sci. USA 91: 7017–7021
7. Chae H. Z., Uhm T. B. and Rhee S. G. (1994) Dimerization of thiol-specific antioxidant and the essential role of cysteine 47. Proc. Natl. Acad. Sci. USA 91:7022–7026
8. Yim M. B., Chae H. Z., Rhee S. G., Chock P. B. and Stadtman E. R. (1994) On the protective mechanism of the thiol-specific antioxidant enzyme against the oxidative damage of biomacromolecules. J. Biol. Chem. 269: 1621–1626
9. Netto L. E. S., Chae H. Z., Kang S.-W., Rhee S. G. and Stadtman E. R. (1996) Removal of hydrogen peroxide by thiol-specific antioxidant enzyme (TSA) is involved with its antioxidant properties. J. Biol. Chem. 271: 15315–15321
10. Sarafian T. A., Rajper N., Grigorian B., Kim A. and Shau H. (1997) Cellular antioxidant properties of human natural killer enhancing factor B. Free Radic. Res. 26: 281–289
11. Pahl P., Berger R., Hart I., Chae H. Z., Rhee S. G. and Patterson D. (1995) Localization of TDPX1, a human homologue of the yeast thioredoxin-dependent peroxide reductase gene (TPX), to chromosome 13q12. Genomics 26: 602–606
12. Sarafian T. A., Verity M. A., Vinters H. V., Shih C. C.-Y., Shi L., Ji X. D. et al. (1999) Differential expression of peroxiredoxin subtypes in human brain cell types. J. Neurosci. Res. 56: 206–212
13. Kang SW, Bains IC, Rhee SG. Characterization of a mammalian peroxiredoxin that contains one conserved cysteine. J Biol Chem 1998;273:6303-11.
14. Chae HZ, Kang SW, Rhee SG. Isoforms of mammalian peroxiredoxin that reduce peroxides in presence of thioredoxin. Methods Enzymol 1999;300:219-26.
15. Zhang P, Liu B, Kanf SW, Seo MS, Rhee SG, Obeid LM. Thioredoxinperoxidase is a novel inhibitor of apoptosis with a mechanism distinct from that of Bcl-2. J Biol Chem 1997;272:30615-8.
16. Plishker GA, Chevalier D, Seinsoth L, Moore RB. Calcium-activated potassium transport and high molecular weight forms of calpromotin. J Biol Chem 1992;267:21839-43.
17. Schroder E, Willis AC, Ponting CP. Porcine natural-killer-enhancing factor-B: oligomerisation and identification as a calpain substrate in vitro. Biochim Biophys Acta 1998;1383:279-91.
18. Lee SC, Chae HZ, Lee JE, Kwon BD, Lee JB, Won YH, et al. Peroxiredoxin is ubiquitously expressed in rat skin: isotype-specific expression in the epidermis and hair follicle. J Invest Dermatol 2000;115:1108-14.
19. Lee JB, Yun SJ, Chae HZ, Won YH, Kim YP, Lee SC. Expression of peroxiredoxin and thioredoxin in dermatological disorders. Br J Dermatol 2002;146:710-2.
20. Mitsumoto A, Takanezawa Y, Okawa K, Iwamatsu A, Nakagawa Y. Variants of peroxiredoxins expression in response to hydroperoxide stress. Free Radic Biol Med 2001;30:625-35.
21. Rabilloud T, Berthier R, Vincon M, Ferbus D, Goubin G, Lawrenve J-J. Early events in erythroid differentiation: accumulation of the acidic peroxiredoxin (PRP/thiol-specific antioxidant /NKEF-B). Biochem J 1995;312:699-705.
22. Lim YS, Cha MK, Yun CH, Kim HK, Kim K, Kim IH. Purification and characterization of thiol-specific antioxidant protein from human red blood cell: a new type of antioxidant protein. Biochem Biophys Res Commun 1994;199:199-206.
23. Shau H, Kim A. Identification of natural killer enhancing factor as a major antioxidant in human red blood cells. Biochem Biophys Res Commun 1994;199:83-8.
24.Karihtala P, Mantyniemi A, Kang SW, Kinnula VL, Soini Y. Peroxiredoxins in Breast Carcinoma. Clin Cancer Res. 2003,15;9(9):3418-24.
25.Young DY, Young MC, Jong KP, Chul MA, Sung KK, Hyung JK. Synergistic effect of peroxiredoxin II antisense on cisplatininduced cell death[J]. Experimentaland Molecular Medicine,2002;34:273-277
26. Wulfkuhle JD, Sgroi DC, Krutzsch H, et al. Proteomics of human breast ductal carcinoma in situ[J]. Cancer Research,2002;62:6740-6749
27. Keegan L, Gill G, Ptashne M. Separation of DNA binding from the transcription-activating function of a eukaryotic regulatory protein. Science,1986,231:699-704
28.Chung YM, Yoo YD, Park JK, Kim YT, Kim HJ. Increased expression of peroxiredoxin II confers resistance to cisplatin. Anticancer Res 2001;21:1129-34
29.Kim H, Lee TH, Park ES, Suh JM, Park SJ, Chung HK, Kwon OY, Kim YK, Ro HK, Shong M. Role of peroxiredoxins in regulating intracellular hydrogen peroxide and hydrogen peroxide-induced apoptosis in thyroid cells. J Biol Chem 2000;275:18266-70
30.Zhang P, Liu B, Kang SW, Seo MS, Rhee SG, Obeid LM. Thioredoxin peroxidase is a novel inhibitor of apoptosis with a mechanism distinct from that of Bcl-2. J Biol Chem 1997;272: 30615-8
31.Park SH, Chung YM, Lee YS, Kim HJ, Kim JS, Chae HZ, Yoo YD. Antisense of human peroxiredoxin II enhances radiationinduced cell death. Clin Cancer Res 2000;6:4915-20
32. Beck MA. Selenium and host defence towards viruses. Proc Nutr Soc,1999,58(3):707)711
33. Ozaki M, Deshpande SS, Angkeow P, et al. Inhibition of the Rac1 GT-Pase protects against nonlethal ischrmia /reperfusion-induced necrosis and apoptosis in vivo. FASEB J,2000,14:418-429
34. Lennon SV, Martin SJ, Cotter TG Does-dependent induction of apoptosis in human tumor or cell lines by widely diverging stiumuli. Cell prolif,1991,24:203-214
35.高颖,许彩民,杨洋.氧化诱导K652细胞凋亡机制的初步探讨.中国生物化学与分子生物学学报,1998,14(3):295-299
36. Ishisaka R, Utsumi K, Utsumi T. Involvement of lysosomal cysteine protease in hydrogen peroxide - induced apoptosis in HL - 60 cells. Biosci Biotechnol Biochem,2002,66(9):1865-1872
37. Aruoma, O. I.; Halliwell, B.; Gajewski, E.; Dizdaroglu, M. Copperion-dependent damage to the bases in DNA in the presence of hydrogen peroxide. Biochem. J. 1991,273:601– 604;.
38. Termini, J. Hydroperoxide-induced DNA damage and mutations. Mutat. Res. 2000.450:107– 124;
39. Guidarelli, A.; Cattabeni, F.; Cantoni, O. Alternative mechanisms for hydroperoxide-induced DNA single strand breakage. Free Radic. Res. 1997.26:537– 547;
40. Cooke, M. S.; Evans, M. D.; Dizdaroglu, M.; Lunec, J. Oxidative DNA damage: mechanisms, mutation and disease. FASEB J. 2003.17:1195– 1214;
41.J in X , Han C S , Yu F Q , et al . Anti apoptic action of stem cell fact or on oocytes in primordial follicles and its signal transduction[J ] . Mol Rep rod Dev ,2005 ,70 (1) :82290.
42. Hu X M , Christian P , Sipes I G , et al . Exp ression and redist ribution of cellular Bad , Bax , and Bcl-X/L p rotein is associated wit h VCD2induced ovot oxicity in rats [ J ] . Biol Rep rod , 2001 ,65 (5) :148921495.
43. Yan W , Samson M , J egou B , et al . Bak/ Bcl2W cor resp ond to spermat ogonial and spermat ocyte ap opt os Bcl2W forms complexes wit h Bax and Bak , and elevated ratio of Bak/ Bcl2W and is in t he testis [J ] . Mol Endocrinol , 2000 ,14 (5) :6822699.
44. Kimura M , It oh N , Takagi S , et al . Balance of ap opt osis and prolif eration of germ cells related t o spermat ogenesis in aged men[J ] . J Androl ,2003 ,24 (2) :185291.
45. Lionel Larue,Alfonso Bellacosa. Epithelial–mesenchymal transition in development and cancer: role of phosphatidylinositol 3 kinase/AKT pathways[J]. Oncogene, 2005,24, 7443-7454.
46. Sylvia Julien Grille, Alfonso Bellacosa, John Upson, et al. The Protein Kinase Akt Induces Epithelial Mesenchymal Transition and Promotes Enhanced Motility and Invasiveness of Squamous Cell Carcinoma Lines[J]. Cancer Research, 2003,63, 2172–2178.
47. Felipe Palacios, Jogender S. Tushir, Yasuyuki Fujita, et al. Lysosomal Targeting of E-Cadherin: a Unique Mechanism for the Down-Regulation of Cell-Cell Adhesion during Epithelial to Mesenchymal Transitions[J]. Molecular and Cell Biology, 2005,25(1):389-402.
1 Chae, HZ, Robison K, Poole LB, et al. Cloning and sequencing of thiol-specific antioxidant from mammalian brain: alkyl hydroperoxide reductase and thiol-specific antioxidant define a large family of antioxidant enzymes[J]. Proc. Natl Acad. Sci. USA,1994,91:7017–7021
2 Chae, HZ, Chung SJ, Rhee SG. Thioredoxin-dependent peroxide reductase from yeast[J]. J. Biol. Chem,1994,269:27670–27678
3 Bryk, R.Griffin P, Nathan C. Peroxynitrite reductase activity of bacterial peroxiredoxins[J]. Nature,2000, 407: 211–215
4.Peshenko IV, Shichi H. Oxidation of active center cysteine of bovine 1-Cys peroxiredoxin sulfenic acid form by peroxide and peroxynitrite[J]. Free Radic Biol Med,2001, 31: 292–303
5.Shau H, Merino A, Chen L, et al. Induction of peroxiredoxins in transplanted livers and demonstration of their in vitro cytoprotection activety[J]. Antioxid Redox Signal,2000,2:347-354.
6.Liu Yuxin, Zheng Jie, Fang Weigang, You Jiangfeng, Wang Jieliang, Cui Xianglin, Wu Bingquan, et al. Isolation and characterization of human prostate cancer cell subclones with different metastatic potential.[J]. Chin J Pathol, 1999,28:361-364
7. Bartel PL, Fields S. The yeast two-hybrid system[M]. New York: Oxford University Press, Oxford,1997:425-428
8. Schiestl RH, Gietz RD. High efficiency transformation of intact yeast cells using single stranded nucleic acids as a carrier.[J] Curr Genet,1989,16(5-6):339-346
9. Printen JA, Sprague GF Jr. Protein-protein interactions in the yeast pheromone response pathway: Ste5p interacts with all members of the MAP kinase cascade[J].Genetics, 1994,138(3):609-19.
10. Schiestl RH, Gietz RD. High efficiency transformation of intact yeast cells using single stranded nucleic acids as a carrier.[J] Curr Genet,1989,16:339-346.
11. Boucher IW, McMillan PJ, Gabrielsen M,et al. Structural and biochemical characterization of a mitochondrial peroxiredoxin from Plasmodium falciparum[J]. Mol Microbiol,2006,61(4):948-959
12. Molina Lopez J, Jimenez L, Ochoa Sanchez A, et al. Molecular cloning and characterization of a 2-Cys peroxiredoxin from Taenia solium [J]. J Parasitol.,2006,92(4): 796-802
13. Zhang P, Liu B, Kanf SW, Seo MS, Rhee SG, Obeid LM. Thioredoxin peroxidase is a novel inhibitor of apoptosis with a mechanism distinct from that of Bcl-2.[J]. J Biol Chem,1997,272:30615-8.
14. Noh DY, Ahn SJ, Lee RA, Kim SW, Park ZA, Chae HZ. Overexpression of peroxiredoxin in human breast cancer[J]. Anticancer Res,2001,21:2085-90.
15. Okazaki S, Naganuma A, Kuge S. Peroxiredoxin-mediated redox regulation of the nuclear localization of Yap1, a transcription factor in budding yeast[J].Antioxid Redox Signal,2005,7(3-4): 327-334
16. Plishker GA, Chevalier D, Seinsoth L, Moore RB. Calcium-activated potassiumtransport and high molecular weight forms of calpromotin[J]. J Biol Chem,1992,267:21839-43.
17. Schroder E, Willis AC, Ponting CP. Porcine natural-killer-enhancing factor-B: oligomerisation and identification as a calpain substrate in vitro[J]. Biochim Biophys Acta,1998,1383:279-91.
18. Lee SC, Na YP, Lee JB. Expression of peroxiredoxin II in vascular tumors of the skin: A novel vascular marker endothelial cells[J]. J Am Acad Dermatol,2003,49(3):487-491
19.Karihtala P, Mantyniemi A, Kang SW, Kinnula VL, Soini Y. Peroxiredoxins in Breast Carcinoma[J]. Clin Cancer Res,2003,15;9(9):3418-24.
20. Young DY, Young MC, Jong KP, Chul MA, Sung KK, Hyung JK. Synergistic effect of peroxiredoxin II antisense on cisplatininduced cell death[J]. Exp Mol Med,2002,34(4):273-277
21. Wulfkuhle JD, Sgroi DC, Krutzsch H, et al. Proteomics of human breast ductal carcinoma in situ[J]. Cancer Research,2002,62:6740-6749
22. Fields S, Song O. A novel genetic system to detect protein-protein interactions[J]. Nature,1989,340(6230):245-246
23.Keegan L, Gill G, Ptashne M. Separation of DNA binding from the transcription-activating function of a eukaryotic regulatory protein[J]. Science,1986,231:699-704
1. Chen CD , Welsbie DS , Tran C , et al. Molecular determinants of resistance to anti androgen therapy. Nat Med , 2004 , 10∶33
2. Bubley GJ , Carducci M, and Dahut W. Eligibility and response guidelines for phase II clinical trials in androgen independent prostate cancer : Recommendations from the prostate specific antigen working group. J Clin Oncol , 1999 , 17∶3461
3. McDonnell , TJ . Expression of the protooncogene bcl-2 in the prostate and its association with emergence of androgen independent prostate cancer. Cancer Res , 1992 , 52 , 6940
4. Yeh S , et al. From HER22/ Neu signal cascade to androgen receptor and its coactivators : A novel pathway by induction of androgen target genes through MAP kinase in prostate cancer cells.Proc Natl Acad Sci USA , 1999 , 96∶5458
5. Putz T, et al. Epidermal growth factor ( EGF) receptor blockade inhibits the action of EGF , insulin2like growth factor I , and a protein kinase A activator on the mitogen2activated protein kinase pathway in prostate cancer cell lines. Cancer Res , 1999 ,59∶227
6. Sherwood ER , et al. Epidermal growth factor receptor activation in androgen independent but not androgen stimulated growth of human prostatic carcinoma cells. Br J Cancer , 1998 , 77∶855
7. Craft N , Shostak Y, Carey M, et al. Mechanism for hormone independent prostate cancer through modulation of androgen receptor signaling by the HER - 2/ neu tyrosine kinase. Nature Med , 1999 , 5∶280
8. Yeh S. et al. From HER2/ Neu signal cascade to androgen receptor and its coactivators : A novel pathway by induction of androgen target genes through MAP kinase in prostate cancer cells. Proc Natl Acad Sci USA , 1999 , 96∶5458
9. Wen Y. et al. HER - 2/ neu promotes androgen independent survival and growth of prostate cancer cells through the AKT path way. Cancer Res , 2000 , 60∶6841
10. Zhou BP , et al. HER - 2/ neu blocks tumor necrosis factor in duced apoptosis via the AKT/ NF-kB pathway. J Biol Chem , 2000 , 275∶8027
11. McMenamin ME , Soung P , Perera S , et al. Loss of PTEN expression inparaffin embedded primary prostate cancer correlates with high Gleason score and advanced stage. Cancer Res , 1999 , 59∶4291
12. Vivanco I , Sawyers CL. The phosphatidylinositol 3– kinase AKT pathway in human cancer. Nat Rev Cancer , 2002 , 2∶489
13. Fizazi K,Martinez LA ,Sikes CR , et al . The association of p21 (WAF21/ CIPI)with progression to androgen2independent prostate cancer [J ] .Clin Cancer Res ,2002 ,8 (3) :775-781.
14. Taplin ME ,Ho SM. Clinical review 134 :The endocrinology of prostate cancer[J ] . J Clin Endocrinol Metab ,2001 ,86(8) :3467-3477.
15. Agus DB ,Cordon2Cardo C , Fox W, et al . Prostate cancer cell cycle regulators :response to androgen withdrawal and development of androgen independence[J ] . J Natl Cancer Inst ,1999 ,91(21) :1869-1876.
16. Gao X,Chen YQ ,Wu N , et al . Somatic mutations of the WAF1/ CIP1 gene in primary prostate cancer [J ] . Oncogene ,1995 ,11 (7) : 1395 -1398.
17. Chang BD ,Watanabe K,Broude EV, et al . Effects of p21Waf1/ Cip1/Sdi1 on cellular gene expression : implications for carcinogenesis ,senescence ,and age2related diseases[J ] . Proc Natl Acad Sci U S A ,2000 ,97 (8) :4291-4296.
18. Ling MT ,Wang X,Lee DT , et al . Id21 expression induces androgen independent prostate cancer cell growth through activation of epidermal growth factor receptor ( EGF2R) [ J ] . Carcinogenesis , 2004 , 25 (4) :517-525.
19. Palayoor ST , Youmell MY, Calderwood SK, et al . Constitutive activation of Ikappa B kinase alpha and NF-kappa B in prostate cancer cells is inhibited by ibuprofen[J ] . Oncogene , 1999 , 18 (51) :7389-7394.
20. Kimura K, Markowski M, Bowen C , et al . Androgen blocks apoptosis of hormone2dependent prostate cancer cells [ J ] . Cancer Res ,2001 , 61 (14) :5611-5618.
21. Chen CD , Sawyers CL. NF-kappa B activates prostate-specific antigen expression and is upregulated in androgen-independent prostate cancer[J ] . Mol Cell Biol , 2002 , 22 (8) :2862-2870.
22. Kasof GM, Lu JJ , Liu D , et al . Tumor necrosis factor-alpha induces theexpression of DR6 , a member of the TNF receptor family , through activation of NF-kappa B[J ] . Oncogene , 2001 , 20 (55) :7965-7975.
23. Yu R , Mandlekar S , Ruben S , et al . Tumor necrosis factor-related apoptosis-inducing ligand-mediated apoptosis in androgen-independent prostate cancer cells [J ] . Cancer Res , 2000 , 60(9) :2384-2389.
24. Catz SD , Johnson JL. Transcriptional regulation of Bcl-2 by nuclear factor kappa B and its significance in prostate cancer[J ] . Oncogene , 2001 , 20 (50) :7342-7351.
25. Huang S , Pettaway CA , Uehara H , et al . Blockade of NF-kappa B activity in human prostate cancer cells is associated with suppression of angiogenesis, invasion , and metastasis [ J ] . Oncogene , 2001 , 20(31) :4188-4197.
26. Lindholm PF , Bub J , Kaul S , et al . The role of constitutive NF-kappa B activity in PC-3 human prostate cancer cell invasive behavior [J ] . Clin Exp Metastasis , 2000 , 18 (6) :471-479.
© 2004-2018 中国地质图书馆版权所有 京ICP备05064691号 京公网安备11010802017129号 地址：北京市海淀区学院路29号 邮编：100083 电话：办公室：(+86 10)66554848；文献借阅、咨询服务、科技查新：66554700 |