DEN诱发大鼠肝癌形成过程中差异表达蛋白质组分析及AKR1B10的研究
|
摘要
肝细胞癌(hepatocellular carcinoma,HCC,以下简称肝癌)是一种恶性程度高、预后凶险的肿瘤,总体5年生存率只有7%左右。在全世界范围内肝癌约占全部人类恶性肿瘤的5.6%,其中一半以上发生在我国,并且发病率近年来还呈上升的趋势。虽然全世界对肝癌的研究已有多年的历史并已取得长足的进步,但它的发病机制至今仍未得到充分阐释。现已知肝癌的形成是一个多因素、多步骤、多基因参与的过程,仅用传统的单因素、单基因相关性分析无法准确反映肝癌复杂的病理机制。近年来迅速发展的蛋白质组学技术为动态、整体、定量地考察疾病发生发展过程中全部蛋白质种类和数量的变化提供了一个高效的研究平台,推动了肝癌发生机制、新的肝癌诊断标志物和治疗靶标等方面的研究进展。
本研究用二乙基亚硝胺(diethylnitrosamine,DEN)诱发大鼠肝癌形成,以γ-谷氨酰转肽酶(γ-glutamyl transpeptidase,γ-GT)染色阳性的肝细胞增生灶(γ-GT阳性灶)为标志获取肝癌前病变的组织标本,再用比较蛋白质组学技术对大鼠肝癌组织、肝癌前病变组织和正常肝组织的全蛋白质组表达谱进行差异分析,筛选出一批差异表达蛋白。随后,本研究对部分筛选出来的差异表达蛋白质如醛糖还原酶1B10(aldo-keto reductase family 1 member B10,AKR1B10)、波形蛋白(VIMENTIN)进行表达水平改变的验证,再应用RNA干扰(RNA interfering,RNAi)技术对AKR1B10进行肿瘤相关生物学功能分析和机制探讨,以及应用组织芯片和免疫组化技术观察和分析AKR1B10在较大样本量的人肝癌组织中的表达及其临床意义。研究结果表明,AKR1B10在肝癌的发生发展过程中可能通过调节肿瘤相关基因的表达来影响细胞的增殖和凋亡能力,它在肝癌组织中的表达与肝癌组织的分化程度相关,在血清甲胎蛋白(α-fetal protein, AFP)阴性患者的肝癌组织中高表达。这些结果提示AKR1B10对预测肝癌的发生和预后可能有一定的价值,是一种具有潜在应用前景的肝癌早期诊断的分子标志物。
研究分以下四部分进行。
第一部分DEN诱发大鼠肝癌发生过程中差异表达蛋白质组学研究
建立DEN诱发大鼠肝癌实验模型,应用比较蛋白质组学技术观察大鼠肝癌发生发展不同阶段的肝组织中蛋白质表达谱的改变,从中筛选出在肝癌发生过程中起重要作用的候选蛋白。
6周龄的雄性近交系Wistar大鼠随机分为实验组和对照组,实验组动物经腹腔注射DEN诱发肝癌,对照组不予DEN处理。实验过程定期处死两组部分动物,通过γ-GT组织化学染色并结合HE染色识别和获取肝癌前病变组织、肝癌组织和正常对照肝组织。三种组织标本各取6例,分别提取肝组织总蛋白,然后按组织类别将6份总蛋白等量混合,获得癌前、肝癌和正常三种组织总蛋白。经双向凝胶电泳(two-dimensional electrophoresis,2-DE)进行总蛋白分离,通过ImageMaster软件分析找出差异表达蛋白点,应用基质辅助激光解吸飞行时间串联质谱(MALDI-TOF-MS/MS)进行肽指纹谱鉴定。质谱鉴定数据送入NCBI非冗余数据库,通过MASCOT搜索引擎检索,得到差异表达蛋白质。最后通过Gene Ontology、SWISSPROT和NCBI等数据库分析差异表达蛋白质的细胞内定位、分子功能和生物过程,并应用IntACT数据库对差异表达蛋白质进行相互作用网络分析。
动物实验于30周结束,结果显示实验组共有9只动物发生肝癌,肝癌发生的最早时间为实验第21周;对照组无1例发生肝癌。γ-GT组织化学染色显示随着DEN诱癌时间延长,γ-GT阳性灶数量增多、面积增大,癌前组织中γ-GT阳性灶总面积与肝组织切片面积比达29%～43%;肝癌组织γ-GT染色全为阳性;正常肝组织内无γ-GT阳性灶。应用2-DE技术筛选出组间表达水平差异≥2倍或特异存在的蛋白点127个,质谱鉴定出AKR1B10、VIMENTIN和粘连蛋白受体1(laminin receptor 1,LAMR1)等82种蛋白质,其中肝癌组织与正常组织相比有75个差异表达蛋白质,肝癌组织与癌前组织相比有39个差异表达蛋白质,癌前组织与正常组织相比有14个差异表达蛋白质,在以上三种比较中同时差异表达的蛋白质有5个。生物信息学分析显示,这些差异表达蛋白质主要位于细胞质和线粒体;发挥酶的活性作用、氧化还原和结合等功能;参与的主要生物学过程为代谢、运输和转移、生物合成和氧化应激等。应用IntACT数据库分析差异表达蛋白质的相互作用,发现4型葡萄糖转运蛋白(Glucose transporter type 4,GLUT4)是相互作用网络中的一个共同节点。
以上结果表明通过γ-GT组织化学染色和HE染色相结合的方法有助于识别和获取大鼠肝癌前病变组织、肝癌组织及正常肝组织;大鼠肝癌发生发展不同阶段的蛋白质表达谱存在明显差别;肝癌的发生发展可能与多种蛋白质表达水平的改变有关。
第二部分差异表达蛋白AKR1B10和VIMENITN在大鼠和人肝癌组织中的表达验证
由于从2-DE到质谱鉴定技术流程长、影响因素多,为了保证以上筛选实验结果的可靠性以便对相关分子作进一步研究,本部分研究对筛选出来的部分蛋白质进行差异表达的验证。
研究方法主要为应用Western blot和RT-PCR方法分别检测AKR1B10和VIMENTIN在大鼠和人肝癌组织中的蛋白质水平和mRNA水平的表达情况,以及应用免疫组化染色观察AKR1B10在肝细胞中的定位。结果显示:①AKR1B10蛋白定位于肝细胞胞浆内,其蛋白表达水平和mRNA表达水平在大鼠正常肝、肝癌前病变、肝癌组织中依次上调;AKR1B10蛋白表达水平在人正常肝、癌旁肝及肝癌组织中也依次上调。②VIMENTIN蛋白在大鼠正常肝、肝癌前病变、肝癌组织中表达依次明显上调;VIMENTIN mRNA在大鼠肝癌组织中明显升高,在肝癌前病变组织中轻度升高但与正常肝组织间的差别无统计学意义;VIMENTIN蛋白在人正常肝、癌旁肝及肝癌组织中的表达依次上调。
以上结果显示的AKR1B10和VIMENTIN的表达变化趋势与2-DE结果基本一致,提示本研究采用的2-DE和MALDI-TOF-MS/MS蛋白质组学技术平台具有相当的稳定性和可靠性。在mRNA和蛋白质两个水平上,AKR1B10和VIMENTIN不仅在大鼠正常肝、肝癌前病变、肝癌组织中表达依次上调,并且在人正常肝、肝癌旁组织、肝癌组织中的表达水平也显著依次升高,提示这两个蛋白可能参与人类肝癌的发生发展过程。
第三部分AKR1B10在肝癌发生过程中作用及机制的初步探讨
为了进一步探讨AKR1B10在肝癌发生发展中的作用,本部分应用RNAi技术和肿瘤相关生物学功能鉴定方法,观察AKR1B10表达被抑制后对人肝癌细胞系MHCC97H相关功能的影响,并探讨其机制。
首先应用RT-PCR、Western blot检测AKR1B10 mRNA和蛋白在肝癌细胞系MHCC97L、MHCC97H、SMMC-7721和BEL-7402及正常肝细胞系Changliver中的表达以筛选AKR1B10表达量最高的细胞系,然后采用RNAi技术,将化学合成的两对针对AKR1B10的小干扰RNA(small interfering RNA,siRNA)瞬时转染至肝癌细胞MHCC97H细胞,再通过实时荧光定量PCR(real-time PCR)、细胞免疫荧光、Western blot及AKR1B10酶活性检测,筛选干扰效果最明显的一对siRNA作RNAi组,另设置阴性对照组(Mock,转染无效用的dsRNA)和空白对照组(Con,不作任何转染),最后观察AKR1B10表达下调后对MHCC97H细胞的增殖、凋亡、粘附、运动等肿瘤相关生物学功能的影响,并用Real-time PCR检测干扰AKR1B10表达后c-myc等肿瘤相关基因的表达变化。
结果显示,AKR1B10在所检测的四种肝癌细胞系中均有不同程度的表达,以在MHCC97H细胞中表达最强;在正常肝细胞Changliver中无表达。瞬时转染AKR1B10-siRNA 48h和72h后,MHCC97H细胞中的AKR1B10 mRNA和蛋白表达受到较为明显的抑制。CCK-8实验结果显示AKR1B10表达被干扰后MHCC97H细胞生长受到抑制,抑制率在24h、48h、72h和96h分别为6.5%、18.0%、26.6%和22.7%。流式细胞仪检测结果显示RNAi组的凋亡细胞比例显著高于与阴性和空白对照组,三组分别为37.30%、5.18%和5.57%(p<0.05)。粘附实验结果显示RNAi组在30min、50min和70min时的粘附细胞数与阴性对照组和空白组均无明显差异。体外运动实验结果显示RNAi组、阴性对照组和空白对照组中MHCC97H细胞穿过微孔滤膜到达下室面的细胞数分别为18±2、20±3和21±4个,三组间的差别无统计学显著性。Real-time PCR检测发现干扰AKR1B10表达后,RNAi组中癌基因c-myc、c-fos、N-ras和增殖相关基因ki-67表达下降,促凋亡基因casps-3和bax表达上调。
以上提示AKR1B10可能通过调节肿瘤相关基因的表达水平来促进细胞增殖、抑制细胞凋亡,从而引起肝细胞的恶性转化。AKR1B10与肝癌细胞的粘附和运动能力无明显关系。
第四部分AKR1B10在较大样本人肝癌组织中的表达及临床意义的研究
本部分利用组织芯片和免疫组化技术检测AKR1B10蛋白在65例人肝癌及其相应癌旁组织、14例正常人肝组织中的表达情况,分析肝癌组织中AKR1B10蛋白表达与患者的临床病理特征的关系,评估AKR1B10作为新的肝癌早期诊断分子标志物的可能性。
结果显示AKR1B10蛋白在人正常肝组织、癌旁肝组织及肝癌组织中的表达率和表达量均依次升高,并且差别具有统计学显著性(p<0.05)。AKR1B10在肝癌组织的表达水平与该组织的Edmondson病理分级相关,即其表达量在分化程度为I～II级的肝癌组织中显著高于III～IV级(p<0.05)。65例肝癌组织中AKR1B10阳性表达率为76.9%,明显高于本组患者术前血清AFP阳性率(55.4%,p<0.05)。在28例术前血清AFP阴性的肝癌组织中,21例(75.0%)显示为AKR1B10表达阳性。卡方检验显示AKR1B10在肝癌患者组织中的表达与术前血清AFP无相关性(p>0.05)。
AKR1B10蛋白在人正常肝、癌旁肝及肝癌组织中的表达依次升高,在肝癌尤其是血清AFP阴性的肝癌组织中表达率较高,这些特点提示它在肝癌的早期诊断和预后评估等方面具有潜在的临床应用价值。
结论
1.应用γ-GT组织化学染色和HE染色相结合的办法有助于识别和准确获取大鼠肝癌前病变组织用于比较蛋白质组学研究,从而实现动态观察肝癌形成过程中差异表达蛋白。
2.本研究通过2-DE和质谱技术筛查出的一系列与肝癌相关的差异表达蛋白质,尤其是那些在肝癌前病变阶段即出现表达水平改变的蛋白,有可能成为新的诊断早期肝癌的分子标志物或防治靶点。
3. AKR1B10可能通过调节相关基因的表达来影响细胞的增殖和凋亡能力,从而在肝癌的发生发展过程中发挥作用。AKR1B10在人和大鼠肝癌形成过程中表达逐步上调、在术前血清AFP阴性的人肝癌组织中高表达等特点,提示它对预测肝癌的发生有一定的临床应用价值,有可能成为肝癌早期诊断的新分子标志物或防治靶点。
潜在应用价值
1. AKR1B10有可能成为新的肝癌早期诊断的分子标志物,丰富了肝癌诊断指标的选择与组合,有助于提高肝癌早期诊断率。
2.初步探讨了AKR1B10在肝癌发生发展中的作用和机制,有助于肝癌病理机制的阐明,为肝癌的预防和治疗提供了值得进一步研究的分子靶点。
创新点
1.应用γ-GT组织化学染色和HE染色相结合的方法识别和获取大鼠肝癌前病变组织,为动态观察肝癌发生过程中蛋白质表达谱的变化提供了准确有价值的标本。
2.获得了DEN诱发的大鼠肝癌前病变组织和肝癌组织的蛋白质表达谱,共筛选出82种差异表达蛋白质,其中部分蛋白质在肝癌相关研究中为新报道。
3.探讨了AKR1B10的肿瘤相关生物学功能和机制,发现AKR1B10可能通过调节某些基因的表达水平来参与肝癌的发生发展。AKR1B10在大鼠及人的肝癌形成过程中的动态差异表达模式使其有可能成为肝癌的早期诊断标志物或防治靶点。
Hepatocellular carcinoma (HCC) is a type of highly malignant tumor with extremely poor prognosis, and its overall 5-year survival rate is only about 7%. HCC is widespread all over the world, accounting for 5.6% of human malignant tumor, and with an increasing incidence in recent years. The pathogenesis of HCC has not yet been fully explained, although significant progress has been made in the decades-long research. As the formation of HCC is a multi-factor, multi-step and multi-gene process, the study on HCC with the methods that only focus on single factor or single gene can not accurately reflect the complexity of its pathological mechanism. In recent years, the traditional research strategy has been challenged by the rapid progress of proteomics, a technology that is helpful for studying the quantitative and dynamic changes of total proteins during the initiation and progression of a disease.
This study applied two-dimensional electrophoresis (2-DE) and mass spectrometry (MALDI-TOF-MS/MS) to compare the differentially expressed proteins among the normal tissue, preneoplastic tissue and HCC tissue during rat hepatocarcinogenesis induced by diethylnitrosamine (DEN). The preneoplastic tissue was distinguished byγ-glutamyl transpeptidase (γ-GT) staining which marks the foci of liver cell proliferation positive. Among the differentially expressed proteins, aldo-keto reductase family 1 member B10 (AKR1B10) and VIMENTIN were confirmed by Western blot and RT-PCR. The biological functions and mechanism of AKR1B10 were further studied with RNA interference (RNAi) technique. Its expression in a larger scale of human HCC samples and the clinical significance were studied by tissue microarray (TMA) and immunohistochemical techniques. The results indicated that AKR1B10 probably affects cell proliferation and apoptosis by regulating the expression of tumor-associated genes, and thereby plays a role in the occurrence and development of HCC. The expression level of AKR1B10 in HCC tissue was related to the degree of HCC’s differentiation. AKR1B10 up-expressed in 75% (21/28) HCC tissues of which the preoperative serum AFP were negative. AKR1B10 therefore may be helpful to predict the occurrence and prognosis of HCC, and it potentially could be a molecular marker for early diagnosis of HCC.
The entire study includes four parts.
Part One Study on differentially expressed proteome in the rat hepatocarcinogenesis induced by DEN
The purpose of this study was to screen the protein molecules that were crucial for HCC, by comparing the protein expression profiles during the rat hepatocarcinogenesis induced by DEN.
Male Wistar rats, 6 weeks old, were divided into DEN group and control group. Rats in DEN group were injected DEN in order to induce HCC, and the rats in control group were raised normally. The animal experiment lasted 30 weeks, during which a few animals from each group were sacrificed periodically for collecting liver tissues samples. The samples were stained withγ-GT and HE to distinguish the preneoplastic lesion from HCC and the normal. Total proteins were extracted respectively from six tissue samples out of the three categories such as HCC, preneoplastic and normal. The 6 protein samples in each category were mixed equally. The mixed proteins were then separated by 2-DE, and the 2-DE gels were scanned by Image Scanner and analyzed with ImageMaster 2D Platinum 6.0 software. The differentially expressed protein sports were screened out and excised for identifying by MALDI-TOF-MS/MS. The data were then analyzed with the search engine MASCOT (Matrix Science) against a NCBI nonredundant protein sequence database. The cellular location, the molecular function and the biological process of these differentially expressed proteins were analyzed through databases such as Gene Ontology, SWISSPROT and NCBI. The analysis of protein interaction networks was carried on by IntACT database.
A total of nine rats in DEN group developed HCC until the end of animal experiment. The earliest HCC occurred at the 21st week in the animal in DEN group, while none in control group. With the extension of time by DEN-induced HCC, the number and size ofγ-GT foci in the liver of animals in DEN group increased, and the totalγ-GT positive area reached 29%～43% of the whole liver tissue area. HCC tissues wereγ-GT positive entirely. There was noγ-GT focus in control group. 127 differentially expressed proteins, which were with the expression level equal to or more than 2 fold comparing to the corresponding tissues, were screened by 2-DE. 82 proteins including AKR1B10, VIMENTIN and laminin receptor 1 (LAMR1) were identified by MALDI-TOF-MS/MS. Among them, 75 differentially expressed proteins were detected from HCC comparing the normal, 38 were detected from HCC comparing preneoplastic tissue, 14 were detected from preneoplastic tissue comparing the normal, and 5 were detected from the three categories. The proteins were mainly located in cytoplasm and mitochondria, and played roles of enzyme activity, oxidoreductase activity and binding activity. The main biological processes that they involved themselves in were metabolism, transport, biosynthesis, oxidative stress and so on. According to the analysis of proteins interaction network, glucose transporter type 4 (GLUT4) was a common point in all of interaction networks.
The results showed that the methods thatγ-GT staining combined with HE were able to distinguish the preneoplastic lesion from HCC and the normal. The protein expression profiles at different stages during rat hepatocarcinogenesis were significant differences. The occurrence and development of HCC was related to the change in the expression level of multiple proteins.
Part 2 Validation on differential expression of AKR1B10 and VIMENTIN in rat and human HCC
To ensure the reliability of the results from 2-DE and MALDI-TOF-MS/MS, some of the differentially expressed proteins, such as AKR1B10 and VIMENTIN, were validated by immunohistochemical staining, Western blot and RT-PCR.
Immunohistochemical staining showed that AKR1B10 was located in cytoplasm of hepatocyte. AKR1B10 was up-regulated at both mRNA and protein level in rat HCC compared to preneoplastic tissue, and in preneoplastic tissue compared to normal liver tissue. Also, the protein level of AKR1B10 was up-regulated in human HCC tissues compared to the corresponding adjacent liver tissues, and in adjacent liver tissues compared to normal liver tissue. The protein level of VIMENTIN was up-regulated, from rat normal liver tissue to preneoplastic tissue and then to HCC tissue, while its mRNA level was only obviously up-regulated in HCC tissue. The protein level of VIMENTIN was up-regulated from human normal liver tissue to adjacent liver tissue and then to HCC tissue.
It was confirmed that the expression trends of AKR1B10 and VIMENTIN were consistent with the results from 2-DE, indicating the proteomics technology platform used in this study has considerable stability and reliability. Further results showed that AKR1B10 and VIMENTIN were up-regulated, from human normal liver tissue to adjacent liver and then to HCC tissue, suggesting these two proteins may be involved in the occurrence and development of human HCC.
Part 3 Biological function and mechanism study of AKR1B10 in hepatocarcinogenesis
To explore the role of AKR1B10 in hepatocarcinogenesis, further study on its biological function and mechanism was carried on by RNAi technique on the HCC cell line of MHCC97H.
At first, AKR1B10 mRNA and protein levels in HCC cell lines MHCC97L, MHCC97H, SMMC-7721 and BEL-7402 and normal liver cell line Changliver were examinated by RT- PCR and Western blot. Then two pairs of chemically synthesized small interfering RNA (siRNA) targeted on AKR1B10 were transfected into MHCC97H cells. The one pair which had the relatively obvious interfering effect was selected for the following RNA interfering (RNAi) experiments which three groups were set: RNAi group, in which the MHCC97H cells were transfected with AKR1B10 specific siRNA. Mock group, in which the MHCC97H cells were transfected with the dsRNA that had no target at all. Control group, in which the MHCC97H cells were without any transfection. The abilities about cell apoptosis, proliferation, adhesion, migration and the expressions of tumor-associated genes like c-myc were observed after AKR1B10 expression was inhibited.
The results showed that AKR1B10 existed in all of the four HCC cell lines, and the highest expression was in MHCC97H, while there was no expression in the normal liver cell Changliver. The expression levels of AKR1B10 mRNA and protein were down-regulated obviously after transfecting the AKR1B10-targeted siRNA for 48h and 72h respectively. The CCK-8 assay showed that, compared to the two control groups, the cell growth inhibition rates in the RNAi group were 6.5%, 18.0%, 26.6% and 22.7% respectively after transfecting for 24h, 48h, 72h and 96h. The results from flow cytometry analysis showed the proportion of apoptotic cells in RNAi group (37.3%) was significantly higher than that in mock (5.2%) and control group (5.6%). The results from cell adhesion test showed that, after 30min, 50min and 70min, the average cell numbers in the RNAi group were not significantly different from that in the two control groups. Cell motility test showed the average cell numbers per field in the group RNAi, mock and control were 18±2,20±3 and 21±4 respectively, without significant difference from each other. The results of Real-time PCR showed that the oncogenes c-myc, c-fos, N-ras and the proliferation-associated gene ki-67 were down-regulated in MHCC97H, while the apoptosis-promoting genes caspas-3 and bax were up-regulated.
The results in this part showed AKR1B10 was overexpressed in HCC cell lines. When the expression of AKR1B10 had been interfered, the cell growth was inhibited and the cell apoptosis increased, while the abilities about cell adhesion and migration had no significant change. Further assay found some tumor-associated genes changed after the expression of AKR1B10 had been interfered, which might indicate AKR1B10 caused the malignant transformation of liver cells by regulating the expression level of some related genes to promote proliferation and inhibit apoptosis.
Part 4 The expression and clinical significance of AKR1B10 in a larger scale of human HCC samples
To explore the clinical significance of AKR1B10 expression in HCC and evaluate the potential value of AKR1B10 as a new molecular marker for diagnosis early HCC, the AKR1B10 protein level was examinated in 65 cases of human HCC and the adjacent liver tissues, as well as in 14 cases of normal human liver tissue, by tissue microarray (TMA) and immunohistochemical techniques. The correlation analysis between AKR1B10 expression and some of the clinical-pathological characters of the HCC cases was then performed.
The results showed the expression rate and level of AKR1B10 protein was up-regulated in human HCC samples compared to the corresponding adjacent liver tissues, and in adjacent liver tissues compared to normal liver tissue (p<0.05). The AKR1B10 level had significant correlation with Edmondson stage of the HCC cases, namely the AKR1B10 protein level in stage I～II was significantly higher than that in stage III～IV. The up-regulating AKR1B10 expression was found in 50 of 65 (76.9%) HCC cases, which was higher than that of preoperative serum AFP of the patients. Moreover, AKR1B10 protein was detected in 21 HCC tissues of 28 (75.0%) negative serum AFP HCCs. The statistical analysis showed the expression of AKR1B10 protein in HCC tissue was not correlated with the preoperative serum AFP level.
AKR1B10 was up-regulated, from human normal liver tissue to adjacent liver and then to HCC tissue. Its expression rate is high in HCCs, especially in negative serum AFP HCCs. These results might imply AKR1B10 had certain value of clinical applications for early diagnosis of HCC.
Conclusion
1. Byγ-GT staining combined with HE, the rat normal liver tissue, preneoplastic lesion tissue and HCC tissue can be distinguished from each other and therefore obtained for further comparative proteome analysis.
2. The differentially expressed proteins obtained through 2-DE and mass spectrometry in this study might serve as new prognostic biomarkers and/or therapeutic targets for HCC.
3. AKR1B10 might play a role in hepatocarcinogenesis by regulating the expression of some related genes. Its gradually up-regulated pattern in both rat and human hepatocarcinogenesis and the fairly high expression rate in negative serum AFP HCCs, might imply it could serve as a new diagnostic marker for early HCC.
Potential value for application
1. AKR1B10 may be a potential early diagnostic molecular marker of HCC, which can enrich the combination and selection of diagnostic markers and be helpful to improve the rate of early diagnosis for HCC.
2. It is helpful to elucidate the pathogenesis of HCC. Its evaluation may be helpful to explore new therapeutic targets.
Novelty
1. The preneoplastic tissue can be distinguished from HCC and the normal byγ-GT staining combined with HE. By this method, the accurate and valuable tissue samples can be obtained for observing the changes of protein expression profiles during hepatocarcinogenesis.
2. Protein expression profiles were obtained from the preneoplastic lesion and HCC tissue in the rat hepatocarcinogenesis induced by DEN. Eighty-two proteins were screened and identified, some of which were firstly reported.
3. Analysis on biological function and mechanism of AKR1B10 indicates it may be involved in the occurrence and development of HCC by regulating the expression level of some genes. The dynamic differential expression pattern of AKR1B10 in rat and human hepatocarcinogenesis will make AKR1B10 possible to serve as a new diagnosis marker for early HCC.
本研究用二乙基亚硝胺(diethylnitrosamine,DEN)诱发大鼠肝癌形成,以γ-谷氨酰转肽酶(γ-glutamyl transpeptidase,γ-GT)染色阳性的肝细胞增生灶(γ-GT阳性灶)为标志获取肝癌前病变的组织标本,再用比较蛋白质组学技术对大鼠肝癌组织、肝癌前病变组织和正常肝组织的全蛋白质组表达谱进行差异分析,筛选出一批差异表达蛋白。随后,本研究对部分筛选出来的差异表达蛋白质如醛糖还原酶1B10(aldo-keto reductase family 1 member B10,AKR1B10)、波形蛋白(VIMENTIN)进行表达水平改变的验证,再应用RNA干扰(RNA interfering,RNAi)技术对AKR1B10进行肿瘤相关生物学功能分析和机制探讨,以及应用组织芯片和免疫组化技术观察和分析AKR1B10在较大样本量的人肝癌组织中的表达及其临床意义。研究结果表明,AKR1B10在肝癌的发生发展过程中可能通过调节肿瘤相关基因的表达来影响细胞的增殖和凋亡能力,它在肝癌组织中的表达与肝癌组织的分化程度相关,在血清甲胎蛋白(α-fetal protein, AFP)阴性患者的肝癌组织中高表达。这些结果提示AKR1B10对预测肝癌的发生和预后可能有一定的价值,是一种具有潜在应用前景的肝癌早期诊断的分子标志物。
研究分以下四部分进行。
第一部分DEN诱发大鼠肝癌发生过程中差异表达蛋白质组学研究
建立DEN诱发大鼠肝癌实验模型,应用比较蛋白质组学技术观察大鼠肝癌发生发展不同阶段的肝组织中蛋白质表达谱的改变,从中筛选出在肝癌发生过程中起重要作用的候选蛋白。
6周龄的雄性近交系Wistar大鼠随机分为实验组和对照组,实验组动物经腹腔注射DEN诱发肝癌,对照组不予DEN处理。实验过程定期处死两组部分动物,通过γ-GT组织化学染色并结合HE染色识别和获取肝癌前病变组织、肝癌组织和正常对照肝组织。三种组织标本各取6例,分别提取肝组织总蛋白,然后按组织类别将6份总蛋白等量混合,获得癌前、肝癌和正常三种组织总蛋白。经双向凝胶电泳(two-dimensional electrophoresis,2-DE)进行总蛋白分离,通过ImageMaster软件分析找出差异表达蛋白点,应用基质辅助激光解吸飞行时间串联质谱(MALDI-TOF-MS/MS)进行肽指纹谱鉴定。质谱鉴定数据送入NCBI非冗余数据库,通过MASCOT搜索引擎检索,得到差异表达蛋白质。最后通过Gene Ontology、SWISSPROT和NCBI等数据库分析差异表达蛋白质的细胞内定位、分子功能和生物过程,并应用IntACT数据库对差异表达蛋白质进行相互作用网络分析。
动物实验于30周结束,结果显示实验组共有9只动物发生肝癌,肝癌发生的最早时间为实验第21周;对照组无1例发生肝癌。γ-GT组织化学染色显示随着DEN诱癌时间延长,γ-GT阳性灶数量增多、面积增大,癌前组织中γ-GT阳性灶总面积与肝组织切片面积比达29%～43%;肝癌组织γ-GT染色全为阳性;正常肝组织内无γ-GT阳性灶。应用2-DE技术筛选出组间表达水平差异≥2倍或特异存在的蛋白点127个,质谱鉴定出AKR1B10、VIMENTIN和粘连蛋白受体1(laminin receptor 1,LAMR1)等82种蛋白质,其中肝癌组织与正常组织相比有75个差异表达蛋白质,肝癌组织与癌前组织相比有39个差异表达蛋白质,癌前组织与正常组织相比有14个差异表达蛋白质,在以上三种比较中同时差异表达的蛋白质有5个。生物信息学分析显示,这些差异表达蛋白质主要位于细胞质和线粒体;发挥酶的活性作用、氧化还原和结合等功能;参与的主要生物学过程为代谢、运输和转移、生物合成和氧化应激等。应用IntACT数据库分析差异表达蛋白质的相互作用,发现4型葡萄糖转运蛋白(Glucose transporter type 4,GLUT4)是相互作用网络中的一个共同节点。
以上结果表明通过γ-GT组织化学染色和HE染色相结合的方法有助于识别和获取大鼠肝癌前病变组织、肝癌组织及正常肝组织;大鼠肝癌发生发展不同阶段的蛋白质表达谱存在明显差别;肝癌的发生发展可能与多种蛋白质表达水平的改变有关。
第二部分差异表达蛋白AKR1B10和VIMENITN在大鼠和人肝癌组织中的表达验证
由于从2-DE到质谱鉴定技术流程长、影响因素多,为了保证以上筛选实验结果的可靠性以便对相关分子作进一步研究,本部分研究对筛选出来的部分蛋白质进行差异表达的验证。
研究方法主要为应用Western blot和RT-PCR方法分别检测AKR1B10和VIMENTIN在大鼠和人肝癌组织中的蛋白质水平和mRNA水平的表达情况,以及应用免疫组化染色观察AKR1B10在肝细胞中的定位。结果显示:①AKR1B10蛋白定位于肝细胞胞浆内,其蛋白表达水平和mRNA表达水平在大鼠正常肝、肝癌前病变、肝癌组织中依次上调;AKR1B10蛋白表达水平在人正常肝、癌旁肝及肝癌组织中也依次上调。②VIMENTIN蛋白在大鼠正常肝、肝癌前病变、肝癌组织中表达依次明显上调;VIMENTIN mRNA在大鼠肝癌组织中明显升高,在肝癌前病变组织中轻度升高但与正常肝组织间的差别无统计学意义;VIMENTIN蛋白在人正常肝、癌旁肝及肝癌组织中的表达依次上调。
以上结果显示的AKR1B10和VIMENTIN的表达变化趋势与2-DE结果基本一致,提示本研究采用的2-DE和MALDI-TOF-MS/MS蛋白质组学技术平台具有相当的稳定性和可靠性。在mRNA和蛋白质两个水平上,AKR1B10和VIMENTIN不仅在大鼠正常肝、肝癌前病变、肝癌组织中表达依次上调,并且在人正常肝、肝癌旁组织、肝癌组织中的表达水平也显著依次升高,提示这两个蛋白可能参与人类肝癌的发生发展过程。
第三部分AKR1B10在肝癌发生过程中作用及机制的初步探讨
为了进一步探讨AKR1B10在肝癌发生发展中的作用,本部分应用RNAi技术和肿瘤相关生物学功能鉴定方法,观察AKR1B10表达被抑制后对人肝癌细胞系MHCC97H相关功能的影响,并探讨其机制。
首先应用RT-PCR、Western blot检测AKR1B10 mRNA和蛋白在肝癌细胞系MHCC97L、MHCC97H、SMMC-7721和BEL-7402及正常肝细胞系Changliver中的表达以筛选AKR1B10表达量最高的细胞系,然后采用RNAi技术,将化学合成的两对针对AKR1B10的小干扰RNA(small interfering RNA,siRNA)瞬时转染至肝癌细胞MHCC97H细胞,再通过实时荧光定量PCR(real-time PCR)、细胞免疫荧光、Western blot及AKR1B10酶活性检测,筛选干扰效果最明显的一对siRNA作RNAi组,另设置阴性对照组(Mock,转染无效用的dsRNA)和空白对照组(Con,不作任何转染),最后观察AKR1B10表达下调后对MHCC97H细胞的增殖、凋亡、粘附、运动等肿瘤相关生物学功能的影响,并用Real-time PCR检测干扰AKR1B10表达后c-myc等肿瘤相关基因的表达变化。
结果显示,AKR1B10在所检测的四种肝癌细胞系中均有不同程度的表达,以在MHCC97H细胞中表达最强;在正常肝细胞Changliver中无表达。瞬时转染AKR1B10-siRNA 48h和72h后,MHCC97H细胞中的AKR1B10 mRNA和蛋白表达受到较为明显的抑制。CCK-8实验结果显示AKR1B10表达被干扰后MHCC97H细胞生长受到抑制,抑制率在24h、48h、72h和96h分别为6.5%、18.0%、26.6%和22.7%。流式细胞仪检测结果显示RNAi组的凋亡细胞比例显著高于与阴性和空白对照组,三组分别为37.30%、5.18%和5.57%(p<0.05)。粘附实验结果显示RNAi组在30min、50min和70min时的粘附细胞数与阴性对照组和空白组均无明显差异。体外运动实验结果显示RNAi组、阴性对照组和空白对照组中MHCC97H细胞穿过微孔滤膜到达下室面的细胞数分别为18±2、20±3和21±4个,三组间的差别无统计学显著性。Real-time PCR检测发现干扰AKR1B10表达后,RNAi组中癌基因c-myc、c-fos、N-ras和增殖相关基因ki-67表达下降,促凋亡基因casps-3和bax表达上调。
以上提示AKR1B10可能通过调节肿瘤相关基因的表达水平来促进细胞增殖、抑制细胞凋亡,从而引起肝细胞的恶性转化。AKR1B10与肝癌细胞的粘附和运动能力无明显关系。
第四部分AKR1B10在较大样本人肝癌组织中的表达及临床意义的研究
本部分利用组织芯片和免疫组化技术检测AKR1B10蛋白在65例人肝癌及其相应癌旁组织、14例正常人肝组织中的表达情况,分析肝癌组织中AKR1B10蛋白表达与患者的临床病理特征的关系,评估AKR1B10作为新的肝癌早期诊断分子标志物的可能性。
结果显示AKR1B10蛋白在人正常肝组织、癌旁肝组织及肝癌组织中的表达率和表达量均依次升高,并且差别具有统计学显著性(p<0.05)。AKR1B10在肝癌组织的表达水平与该组织的Edmondson病理分级相关,即其表达量在分化程度为I～II级的肝癌组织中显著高于III～IV级(p<0.05)。65例肝癌组织中AKR1B10阳性表达率为76.9%,明显高于本组患者术前血清AFP阳性率(55.4%,p<0.05)。在28例术前血清AFP阴性的肝癌组织中,21例(75.0%)显示为AKR1B10表达阳性。卡方检验显示AKR1B10在肝癌患者组织中的表达与术前血清AFP无相关性(p>0.05)。
AKR1B10蛋白在人正常肝、癌旁肝及肝癌组织中的表达依次升高,在肝癌尤其是血清AFP阴性的肝癌组织中表达率较高,这些特点提示它在肝癌的早期诊断和预后评估等方面具有潜在的临床应用价值。
结论
1.应用γ-GT组织化学染色和HE染色相结合的办法有助于识别和准确获取大鼠肝癌前病变组织用于比较蛋白质组学研究,从而实现动态观察肝癌形成过程中差异表达蛋白。
2.本研究通过2-DE和质谱技术筛查出的一系列与肝癌相关的差异表达蛋白质,尤其是那些在肝癌前病变阶段即出现表达水平改变的蛋白,有可能成为新的诊断早期肝癌的分子标志物或防治靶点。
3. AKR1B10可能通过调节相关基因的表达来影响细胞的增殖和凋亡能力,从而在肝癌的发生发展过程中发挥作用。AKR1B10在人和大鼠肝癌形成过程中表达逐步上调、在术前血清AFP阴性的人肝癌组织中高表达等特点,提示它对预测肝癌的发生有一定的临床应用价值,有可能成为肝癌早期诊断的新分子标志物或防治靶点。
潜在应用价值
1. AKR1B10有可能成为新的肝癌早期诊断的分子标志物,丰富了肝癌诊断指标的选择与组合,有助于提高肝癌早期诊断率。
2.初步探讨了AKR1B10在肝癌发生发展中的作用和机制,有助于肝癌病理机制的阐明,为肝癌的预防和治疗提供了值得进一步研究的分子靶点。
创新点
1.应用γ-GT组织化学染色和HE染色相结合的方法识别和获取大鼠肝癌前病变组织,为动态观察肝癌发生过程中蛋白质表达谱的变化提供了准确有价值的标本。
2.获得了DEN诱发的大鼠肝癌前病变组织和肝癌组织的蛋白质表达谱,共筛选出82种差异表达蛋白质,其中部分蛋白质在肝癌相关研究中为新报道。
3.探讨了AKR1B10的肿瘤相关生物学功能和机制,发现AKR1B10可能通过调节某些基因的表达水平来参与肝癌的发生发展。AKR1B10在大鼠及人的肝癌形成过程中的动态差异表达模式使其有可能成为肝癌的早期诊断标志物或防治靶点。
Hepatocellular carcinoma (HCC) is a type of highly malignant tumor with extremely poor prognosis, and its overall 5-year survival rate is only about 7%. HCC is widespread all over the world, accounting for 5.6% of human malignant tumor, and with an increasing incidence in recent years. The pathogenesis of HCC has not yet been fully explained, although significant progress has been made in the decades-long research. As the formation of HCC is a multi-factor, multi-step and multi-gene process, the study on HCC with the methods that only focus on single factor or single gene can not accurately reflect the complexity of its pathological mechanism. In recent years, the traditional research strategy has been challenged by the rapid progress of proteomics, a technology that is helpful for studying the quantitative and dynamic changes of total proteins during the initiation and progression of a disease.
This study applied two-dimensional electrophoresis (2-DE) and mass spectrometry (MALDI-TOF-MS/MS) to compare the differentially expressed proteins among the normal tissue, preneoplastic tissue and HCC tissue during rat hepatocarcinogenesis induced by diethylnitrosamine (DEN). The preneoplastic tissue was distinguished byγ-glutamyl transpeptidase (γ-GT) staining which marks the foci of liver cell proliferation positive. Among the differentially expressed proteins, aldo-keto reductase family 1 member B10 (AKR1B10) and VIMENTIN were confirmed by Western blot and RT-PCR. The biological functions and mechanism of AKR1B10 were further studied with RNA interference (RNAi) technique. Its expression in a larger scale of human HCC samples and the clinical significance were studied by tissue microarray (TMA) and immunohistochemical techniques. The results indicated that AKR1B10 probably affects cell proliferation and apoptosis by regulating the expression of tumor-associated genes, and thereby plays a role in the occurrence and development of HCC. The expression level of AKR1B10 in HCC tissue was related to the degree of HCC’s differentiation. AKR1B10 up-expressed in 75% (21/28) HCC tissues of which the preoperative serum AFP were negative. AKR1B10 therefore may be helpful to predict the occurrence and prognosis of HCC, and it potentially could be a molecular marker for early diagnosis of HCC.
The entire study includes four parts.
Part One Study on differentially expressed proteome in the rat hepatocarcinogenesis induced by DEN
The purpose of this study was to screen the protein molecules that were crucial for HCC, by comparing the protein expression profiles during the rat hepatocarcinogenesis induced by DEN.
Male Wistar rats, 6 weeks old, were divided into DEN group and control group. Rats in DEN group were injected DEN in order to induce HCC, and the rats in control group were raised normally. The animal experiment lasted 30 weeks, during which a few animals from each group were sacrificed periodically for collecting liver tissues samples. The samples were stained withγ-GT and HE to distinguish the preneoplastic lesion from HCC and the normal. Total proteins were extracted respectively from six tissue samples out of the three categories such as HCC, preneoplastic and normal. The 6 protein samples in each category were mixed equally. The mixed proteins were then separated by 2-DE, and the 2-DE gels were scanned by Image Scanner and analyzed with ImageMaster 2D Platinum 6.0 software. The differentially expressed protein sports were screened out and excised for identifying by MALDI-TOF-MS/MS. The data were then analyzed with the search engine MASCOT (Matrix Science) against a NCBI nonredundant protein sequence database. The cellular location, the molecular function and the biological process of these differentially expressed proteins were analyzed through databases such as Gene Ontology, SWISSPROT and NCBI. The analysis of protein interaction networks was carried on by IntACT database.
A total of nine rats in DEN group developed HCC until the end of animal experiment. The earliest HCC occurred at the 21st week in the animal in DEN group, while none in control group. With the extension of time by DEN-induced HCC, the number and size ofγ-GT foci in the liver of animals in DEN group increased, and the totalγ-GT positive area reached 29%～43% of the whole liver tissue area. HCC tissues wereγ-GT positive entirely. There was noγ-GT focus in control group. 127 differentially expressed proteins, which were with the expression level equal to or more than 2 fold comparing to the corresponding tissues, were screened by 2-DE. 82 proteins including AKR1B10, VIMENTIN and laminin receptor 1 (LAMR1) were identified by MALDI-TOF-MS/MS. Among them, 75 differentially expressed proteins were detected from HCC comparing the normal, 38 were detected from HCC comparing preneoplastic tissue, 14 were detected from preneoplastic tissue comparing the normal, and 5 were detected from the three categories. The proteins were mainly located in cytoplasm and mitochondria, and played roles of enzyme activity, oxidoreductase activity and binding activity. The main biological processes that they involved themselves in were metabolism, transport, biosynthesis, oxidative stress and so on. According to the analysis of proteins interaction network, glucose transporter type 4 (GLUT4) was a common point in all of interaction networks.
The results showed that the methods thatγ-GT staining combined with HE were able to distinguish the preneoplastic lesion from HCC and the normal. The protein expression profiles at different stages during rat hepatocarcinogenesis were significant differences. The occurrence and development of HCC was related to the change in the expression level of multiple proteins.
Part 2 Validation on differential expression of AKR1B10 and VIMENTIN in rat and human HCC
To ensure the reliability of the results from 2-DE and MALDI-TOF-MS/MS, some of the differentially expressed proteins, such as AKR1B10 and VIMENTIN, were validated by immunohistochemical staining, Western blot and RT-PCR.
Immunohistochemical staining showed that AKR1B10 was located in cytoplasm of hepatocyte. AKR1B10 was up-regulated at both mRNA and protein level in rat HCC compared to preneoplastic tissue, and in preneoplastic tissue compared to normal liver tissue. Also, the protein level of AKR1B10 was up-regulated in human HCC tissues compared to the corresponding adjacent liver tissues, and in adjacent liver tissues compared to normal liver tissue. The protein level of VIMENTIN was up-regulated, from rat normal liver tissue to preneoplastic tissue and then to HCC tissue, while its mRNA level was only obviously up-regulated in HCC tissue. The protein level of VIMENTIN was up-regulated from human normal liver tissue to adjacent liver tissue and then to HCC tissue.
It was confirmed that the expression trends of AKR1B10 and VIMENTIN were consistent with the results from 2-DE, indicating the proteomics technology platform used in this study has considerable stability and reliability. Further results showed that AKR1B10 and VIMENTIN were up-regulated, from human normal liver tissue to adjacent liver and then to HCC tissue, suggesting these two proteins may be involved in the occurrence and development of human HCC.
Part 3 Biological function and mechanism study of AKR1B10 in hepatocarcinogenesis
To explore the role of AKR1B10 in hepatocarcinogenesis, further study on its biological function and mechanism was carried on by RNAi technique on the HCC cell line of MHCC97H.
At first, AKR1B10 mRNA and protein levels in HCC cell lines MHCC97L, MHCC97H, SMMC-7721 and BEL-7402 and normal liver cell line Changliver were examinated by RT- PCR and Western blot. Then two pairs of chemically synthesized small interfering RNA (siRNA) targeted on AKR1B10 were transfected into MHCC97H cells. The one pair which had the relatively obvious interfering effect was selected for the following RNA interfering (RNAi) experiments which three groups were set: RNAi group, in which the MHCC97H cells were transfected with AKR1B10 specific siRNA. Mock group, in which the MHCC97H cells were transfected with the dsRNA that had no target at all. Control group, in which the MHCC97H cells were without any transfection. The abilities about cell apoptosis, proliferation, adhesion, migration and the expressions of tumor-associated genes like c-myc were observed after AKR1B10 expression was inhibited.
The results showed that AKR1B10 existed in all of the four HCC cell lines, and the highest expression was in MHCC97H, while there was no expression in the normal liver cell Changliver. The expression levels of AKR1B10 mRNA and protein were down-regulated obviously after transfecting the AKR1B10-targeted siRNA for 48h and 72h respectively. The CCK-8 assay showed that, compared to the two control groups, the cell growth inhibition rates in the RNAi group were 6.5%, 18.0%, 26.6% and 22.7% respectively after transfecting for 24h, 48h, 72h and 96h. The results from flow cytometry analysis showed the proportion of apoptotic cells in RNAi group (37.3%) was significantly higher than that in mock (5.2%) and control group (5.6%). The results from cell adhesion test showed that, after 30min, 50min and 70min, the average cell numbers in the RNAi group were not significantly different from that in the two control groups. Cell motility test showed the average cell numbers per field in the group RNAi, mock and control were 18±2,20±3 and 21±4 respectively, without significant difference from each other. The results of Real-time PCR showed that the oncogenes c-myc, c-fos, N-ras and the proliferation-associated gene ki-67 were down-regulated in MHCC97H, while the apoptosis-promoting genes caspas-3 and bax were up-regulated.
The results in this part showed AKR1B10 was overexpressed in HCC cell lines. When the expression of AKR1B10 had been interfered, the cell growth was inhibited and the cell apoptosis increased, while the abilities about cell adhesion and migration had no significant change. Further assay found some tumor-associated genes changed after the expression of AKR1B10 had been interfered, which might indicate AKR1B10 caused the malignant transformation of liver cells by regulating the expression level of some related genes to promote proliferation and inhibit apoptosis.
Part 4 The expression and clinical significance of AKR1B10 in a larger scale of human HCC samples
To explore the clinical significance of AKR1B10 expression in HCC and evaluate the potential value of AKR1B10 as a new molecular marker for diagnosis early HCC, the AKR1B10 protein level was examinated in 65 cases of human HCC and the adjacent liver tissues, as well as in 14 cases of normal human liver tissue, by tissue microarray (TMA) and immunohistochemical techniques. The correlation analysis between AKR1B10 expression and some of the clinical-pathological characters of the HCC cases was then performed.
The results showed the expression rate and level of AKR1B10 protein was up-regulated in human HCC samples compared to the corresponding adjacent liver tissues, and in adjacent liver tissues compared to normal liver tissue (p<0.05). The AKR1B10 level had significant correlation with Edmondson stage of the HCC cases, namely the AKR1B10 protein level in stage I～II was significantly higher than that in stage III～IV. The up-regulating AKR1B10 expression was found in 50 of 65 (76.9%) HCC cases, which was higher than that of preoperative serum AFP of the patients. Moreover, AKR1B10 protein was detected in 21 HCC tissues of 28 (75.0%) negative serum AFP HCCs. The statistical analysis showed the expression of AKR1B10 protein in HCC tissue was not correlated with the preoperative serum AFP level.
AKR1B10 was up-regulated, from human normal liver tissue to adjacent liver and then to HCC tissue. Its expression rate is high in HCCs, especially in negative serum AFP HCCs. These results might imply AKR1B10 had certain value of clinical applications for early diagnosis of HCC.
Conclusion
1. Byγ-GT staining combined with HE, the rat normal liver tissue, preneoplastic lesion tissue and HCC tissue can be distinguished from each other and therefore obtained for further comparative proteome analysis.
2. The differentially expressed proteins obtained through 2-DE and mass spectrometry in this study might serve as new prognostic biomarkers and/or therapeutic targets for HCC.
3. AKR1B10 might play a role in hepatocarcinogenesis by regulating the expression of some related genes. Its gradually up-regulated pattern in both rat and human hepatocarcinogenesis and the fairly high expression rate in negative serum AFP HCCs, might imply it could serve as a new diagnostic marker for early HCC.
Potential value for application
1. AKR1B10 may be a potential early diagnostic molecular marker of HCC, which can enrich the combination and selection of diagnostic markers and be helpful to improve the rate of early diagnosis for HCC.
2. It is helpful to elucidate the pathogenesis of HCC. Its evaluation may be helpful to explore new therapeutic targets.
Novelty
1. The preneoplastic tissue can be distinguished from HCC and the normal byγ-GT staining combined with HE. By this method, the accurate and valuable tissue samples can be obtained for observing the changes of protein expression profiles during hepatocarcinogenesis.
2. Protein expression profiles were obtained from the preneoplastic lesion and HCC tissue in the rat hepatocarcinogenesis induced by DEN. Eighty-two proteins were screened and identified, some of which were firstly reported.
3. Analysis on biological function and mechanism of AKR1B10 indicates it may be involved in the occurrence and development of HCC by regulating the expression level of some genes. The dynamic differential expression pattern of AKR1B10 in rat and human hepatocarcinogenesis will make AKR1B10 possible to serve as a new diagnosis marker for early HCC.
引文
1. Tang ZY, Ye SL, Liu YK, et al. A decade's studies on metastasis of hepatocellular carcinoma. J Cancer Res Clin Oncol, 2004, 130(4): 187-196.
2. Parkin DM, Bray F, Ferlay J, et al. Global cancer statistics, 2002. CA Cancer J Clin, 2005, 55(2): 74-108.
3. Parkin DM, Bray F, Ferlay J, et al. Estimating the world cancer burden: Globocan 2000. Int J Cancer, 2001, 94(2): 153-156.
4. Wright LM, Kreikemeier JT, Fimmel CJ. A concise review of serum markers for hepatocellular cancer. Cancer Detect Prev, 2007, 31(1): 35-44.
5.张思维,李连弟,鲁凤珠,等.中国1990～1992年原发性肝癌死亡调查分析.中华肿瘤杂志, 1999(04): 5-9.
6. Ward DG, Cheng Y, N'Kontchou G, et al. Changes in the serum proteome associated with the development of hepatocellular carcinoma in hepatitis C-related cirrhosis. Br J Cancer, 2006, 94(2): 287-292.
7. Bosch FX, Ribes J, Diaz M, et al. Primary liver cancer: worldwide incidence and trends. Gastroenterology, 2004, 127(5 Suppl 1): S5-S16.
8. Wands JR. Prevention of hepatocellular carcinoma. N Engl J Med, 2004, 351(15): 1567-1570.
9.郑树森,吴应盛,梁廷波.原发性肝癌420例综合治疗报告.中华普通外科杂志, 2003(12): 3-5.
10. Daniele B, Bencivenga A, Megna AS, et al. Alpha-fetoprotein and ultrasonography screening for hepatocellular carcinoma. Gastroenterology, 2004, 127(5 Suppl 1): S108-S112.
11. Trevisani F, D'Intino PE, Morselli-Labate AM, et al. Serum alpha-fetoprotein for diagnosis of hepatocellular carcinoma in patients with chronic liver disease: influence of HBsAg and anti-HCV status. J Hepatol, 2001, 34(4): 570-575.
12. Tang ZY, Yu YQ, Zhou XD, et al. Subclinical hepatocellular carcinoma: an analysis of 391 patients. J Surg Oncol Suppl, 1993, 3:55-58.
13. Okubo H, Moriyama M, Tanaka N, et al. Detection of serum and intrahepatic hepatocyte growth factor during DEN-induced carcinogenesis in the rat. Hepatol Res, 2002, 24(4): 385-394.
14. Han KQ, Gu W, Hu X, et al. Establishment and biological characteristics of orthotopic transplantation tumor model of hepatocellular carcinoma in miceJ. Zhong Xi Yi Jie He Xue Bao, 2004, 2(5): 372-374.
15. Travis CC, Mcclain TW, Birkner PD. Diethylnitrosamine-induced hepatocarcinogenesis in rats: a theoretical study. Toxicol Appl Pharmacol, 1991, 109(2): 289-304.
16. Yoshiji H, Yoshii J, Ikenaka Y, et al. Inhibition of renin-angiotensin system attenuates liver enzyme-altered preneoplastic lesions and fibrosis development in rats. J Hepatol, 2002, 37(1): 22-30.
17. Yao DF, Dong ZZ, Yao DB, et al. Abnormal expression of hepatoma-derived gamma-glutamyltransferase subtyping and its early alteration for carcinogenesis of hepatocytes. Hepatobiliary Pancreat Dis Int, 2004, 3(4): 564-570.
18. Li Y, Yan RQ, Qin GZ, et al. Reliability of a short-term test for hepatocarcinogenesis induced by aflatoxin B1. IARC Sci Publ, 1991 (105): 431-433.
19.唐小岚,段小娴,苏建家,等.金花茶和银杏叶对2-乙基亚硝胺致大鼠肝癌前病变抑制作用的初步研究.实用癌症杂志, 2007(3): 224-227.
20.刘启福,罗丹,苏建家,等.实验性大鼠肝癌变过程中p16和Rb基因表达的动态观察.临床与实验病理学杂志, 1999(3): 41-43.
21. Fields S. Proteomics. Proteomics in genomeland. Science, 2001, 291 (5507): 1221-1224.
22. Pandey A, Mann M. Proteomics to study genes and genomes. Nature, 2000, 405(6788): 837-846.
23. Gou LT, Tong AP, Chen LJ, et al. Comparative plasma membrane-associated proteomics of immortalized human hepatocytes. Biochemistry (Mosc), 2008, 73(11):1200-1206.
24. Xu J, Khor K A, Sui J, et al. Comparative proteomics profile of osteoblasts cultured on dissimilar hydroxyapatite biomaterials: an iTRAQ-coupled 2-D LC-MS/MS analysis. Proteomics, 2008, 8(20): 4249-4258.
25. Fujii K, Kondo T, Yokoo H, et al. Proteomic study of human hepatocellular carcinoma using two-dimensional difference gel electrophoresis with saturation cysteine dye. Proteomics, 2005, 5(5): 1411-1422.
26. Cui J F, Liu Y K, Pan B S, et al. Differential proteomic analysis of human hepatocellular carcinoma cell line metastasis-associated proteins. J Cancer Res Clin Oncol, 2004, 130(10): 615-622.
27. Cui J F, Liu YK, Zhang LJ, et al. Identification of metastasis candidate proteins among HCC cell lines by comparative proteome and biological function analysis of S100A4 in metastasis in vitro. Proteomics, 2006, 6(22): 5953-5961.
28. Yi X, Luk JM, Lee NP, et al. Association of mortalin (HSPA9) with liver cancer metastasis and prediction for early tumor recurrence. Mol Cell Proteomics, 2008, 7(2): 315-325.
29. Fire A, Xu S, Montgomery MK, et al. Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature, 1998, 391(6669):806-811.
30. Sen G L, Blau H M. A brief history of RNAi: the silence of the genes. FASEB J, 2006, 20(9):1293-1299.
31. Stamps AC, Terrett JA, Adam PJ. Application of in situ reverse trancriptase-polymerase chain reaction (RT-PCR) to tissue microarrays. J Nanobiotechnology, 2003, 1(1): 3.
32. Kononen J, Bubendorf L, Kallioniemi A, et al. Tissue microarrays for high-throughput molecular profiling of tumor specimens. Nat Med, 1998, 4(7): 844-847.
33. Yan R, Zu X, Ma J, et al. Aldo-keto reductase family 1 B10 gene silencing results in growth inhibition of colorectal cancer cells: Implication for cancer intervention. Int J Cancer, 2007, 121(10): 2301-2306.
34. Fukumoto S, Yamauchi N, Moriguchi H, et al. Overexpression of the aldo-keto reductase family protein AKR1B10 is highly correlated with smokers' non-small cell lung carcinomas. Clin Cancer Res, 2005, 11 (5): 1776-1785.
1. Yoshiji H, Yoshii J, Ikenaka Y, et al. Inhibition of renin-angiotensin system attenuates liver enzyme-altered preneoplastic lesions and fibrosis development in rats. J Hepatol, 2002, 37(1): 22-30.
2. Chakraborty T, Chatterjee A, Rana A, et al. Cell proliferation and hepatocarcinogenesis in rat initiated by diethylnitrosamine and promoted by phenobarbital: potential roles of early DNA damage and liver metallothionein expression. Life Sci, 2007, 81(6): 489-499.
3. Swinbanks D. Government backs proteome proposal. Nature, 1995, 378(6558):653.
4. Gou LT, Tong AP, Chen LJ, et al. Comparative plasma membrane-associated proteomics of immortalized human hepatocytes. Biochemistry (Mosc), 2008, 73(11):1200-1206.
5. Xu J, Khor K A, Sui J, et al. Comparative proteomics profile of osteoblasts cultured on dissimilar hydroxyapatite biomaterials: an iTRAQ-coupled 2-D LC-MS/MS analysis. Proteomics, 2008, 8(20): 4249-4258.
6. Petricoin EF, Zoon KC, Kohn EC, et al. Clinical proteomics: translating benchside promise into bedside reality. Nat Rev Drug Discov, 2002, 1(9): 683-695.
7. Ward DG, Cheng Y, N'Kontchou G, et al. Changes in the serum proteome associated with the development of hepatocellular carcinoma in hepatitis C-related cirrhosis. Br J Cancer, 2006, 94(2): 287-292.
8. Ding SJ, Li Y, Tan YX, et al. From proteomic analysis to clinical significance: overexpression of cytokeratin 19 correlates with hepatocellular carcinoma metastasis. Mol Cell Proteomics, 2004, 3(1): 73-81.
9. Farber E. The sequential analysis of liver cancer induction biochem. Et Biophys Acta, 1980, 605:149- 154.
10. Li Y, Yan RQ, Qin GZ, et al. Reliability of a short-term test for hepatocarcinogenesis induced by aflatoxin B1. IARC Sci Publ, 1991(105): 431-433.
11.唐小岚,段小娴,苏建家,等.金花茶和银杏叶对2-乙基亚硝胺致大鼠肝癌前病变抑制作用的初步研究.实用癌症杂志, 2007(3): 224-227.
12. Solt D, Farber E. New principle for the analysis of chemical carcinogenesis. Nature, 1976, 263(5580):701-703.
13. Schwartzenberg-Bar-Yoseph F, Armoni M, Karnieli E. The tumor suppressor p53 down-regulates glucose transporters GLUT1 and GLUT4 gene expression. Cancer Res, 2004, 64(7): 2627-2633.
14. Miyagi T, Hori O, Koshida K, et al. Antitumor effect of reduction of 150-kDa oxygen-regulated protein expression on human prostate cancer cells. Int J Urol, 2002, 9(10): 577-585.
15. Asahi H, Koshida K, Hori O, et al. Immunohistochemical detection of the
150-kDa oxygen-regulated protein in bladder cancer. BJU Int, 2002, 90(4): 462-466.
16. Siegenthaler G, Hotz R, Chatellard-Gruaz D, et al. Characterization and expression of a novel human fatty acid-binding protein: the epidermal type(E-FABP). Biochem Biophys Res Commun, 1993, 190(2): 482-487.
17. Chmurzynska A. The multigene family of fatty acid-binding proteins (FABPs): function, structure and polymorphism. J Appl Genet, 2006, 47(1): 39-48.
18. Masouye I, Hagens G, Van Kuppevelt T H, et al. Endothelial cells of the human microvasculature express epidermal fatty acid-binding protein. Circ Res, 1997, 81(3): 297-303.
19. Loeffler-Ragg J, Skvortsov S, Sarg B, et al. Gefitinib-responsive EGFR-positive colorectal cancers have different proteome profiles from non-responsive cell lines. Eur J Cancer, 2005, 41(15): 2338-2346.
20. Uma RS, Naresh KN, D'Cruz AK, et al. Metastasis of squamous cell carcinoma of the oral tongue is associated with down-regulation of epidermal fatty acid binding protein (E-FABP).Oral Oncol, 2007, 43(1): 27 -32.
21. Rauch J, Ahlemann M, Schaffrik M, et al. Allogenic antibody-mediated identification of head and neck cancer antigens. Biochem Biophys Res Commun, 2004, 323(1):156-162.
22. Munz M, Zeidler R, Gires O. The tumour-associated antigen EpCAM upregulates the fatty acid binding protein E-FABP. Cancer Lett, 2005, 225(1): 151-157.
23. Fujii K, Kondo T, Yokoo H, et al. Proteomic study of human hepatocellular carcinoma using two-dimensional difference gel electrophoresis with saturation cysteine dye. Proteomics, 2005, 5(5): 1411 -1422.
24. Yamaguchi M, Tsurusaki Y, Misawa H, et al. Potential role of regucalcin as a specific biochemical marker of chronic liver injury with carbon tetrachloride administration in rats. Mol Cell Biochem, 2002, 241(1-2):61-67.
25. Tsurusaki Y, Yamaguchi M. Role of regucalcin in liver nuclear function: binding of regucalcin to nuclear protein or DNA and modulation of tumor-related gene expression. Int J Mol Med, 2004, 14(2): 277-281.
26. Fukaya Y, Yamaguchi M. Overexpression of regucalcin suppresses apoptotic cell death in the cloned rat hepatoma H4-II-E cells induced by a naturally occurring isothiocyanate sulforaphane. Int J Mol Med, 2005, 15(5): 853-857.
27. Izumi T, Tsurusaki Y, Yamaguchi M. Suppressive effect of endogenous regucalcin on nitric oxide synthase activity in cloned rat hepatoma H4-II-E cells overexpressing regucalcin. J Cell Biochem, 2003, 89(4): 800-807.
28. Fontanini G, Vignati S, Chine S, et al. 67-Kilodalton laminin receptor expression correlates with worse prognostic indicators in non-small cell lung carcinomas. Clin Cancer Res, 1997, 3(2): 227-231.
29. Zheng S, Ruan Y, Wu Z, et al. The relationship between 67KD laminin receptor expression and metastasis of hepatocellular carcinoma. J Tongji Med Univ, 1997, 17(4): 200-202, 224.
30.梁宏洁,韦薇,康晓楠,等. DEN诱发大鼠肝癌过程中癌前病变阶段差异表达蛋白的筛选. (已投稿)
31. Jr Brozinick J T, Berkemeier B A, Elmendorf J S. "Acting" on GLUT4: membrane & cytoskeletal components of insulin action. Curr Diabetes Rev, 2007, 3(2): 111-122.
32. Jr Brozinick JT, Mccoid SC, Reynolds TH, et al. GLUT4 overexpression in db/db mice dose-dependently ameliorates diabetes but is not a lifelong cure. Diabetes, 2001, 50(3): 593-600.
1. Yan R, Zu X, Ma J, et al. Aldo-keto reductase family 1 B10 gene silencing results in growth inhibition of colorectal cancer cells: Implication for cancer intervention. Int J Cancer, 2007, 121(10): 2301-2306.
2. Fukumoto S, Yamauchi N, Moriguchi H, et al. Overexpression of the aldo-keto reductase family protein AKR1B10 is highly correlated with smokers' non-small cell lung carcinomas. Clin Cancer Res, 2005, 11(5): 1776-1785.
3. Zu X, Yan R, Robbins S, et al. Reduced 293T cell susceptibility to acrolein due to aldose reductase-like-1 protein expression. Toxicol Sci, 2007, 97(2): 562-568.
4. Baumgart E, Cohen MS, Silva NB, et al. Identification and prognostic significance of an epithelial-mesenchymal transition expression profile in human bladder tumors. Clin Cancer Res, 2007, 13(6): 1685-1694.
5. Ngan CY, Yamamoto H, Seshimo I, et al. Quantitative evaluation of vimentin expression in tumour stroma of colorectal cancer. Br J Cancer,2007, 96(6): 986-992.
6. Penning TM, Drury JE. Human aldo-keto reductases: Function, gene regulation, and single nucleotide polymorphisms.Arch Biochem Biophys, 2007, 464(2): 241-250.
7. Zeindl-Eberhart E, Jungblut PR, Otto A, et al. Identification of tumor-associated protein variants during rat hepatocarcinogenesis. Aldose reductase. J Biol Chem, 1994, 269(20): 14589-14594.
8. Zeindl-Eberhart E, Klugbauer S, Dimitrijevic N, et al. Proteome analysis of rat hepatomas: carcinogen-dependent tumor-associated protein variants. Electrophoresis, 2001, 22(14): 3009-3018.
9. Woenckhaus M, Klein-Hitpass L, Grepmeier U, et al. Smoking and cancer-related gene expression in bronchial epithelium and non-small-cell lung cancers. J Pathol, 2006, 210(2): 192-204.
10. Breton J, Gage MC, Hay AW, et al. Proteomic screening of a cell line model of esophageal carcinogenesis identifies cathepsin D and aldo-keto reductase 1C2 and 1B10 dysregulation in Barrett's esophagus and esophageal adenocarcinoma. J Proteome Res, 2008, 7(5): 1953-1962.
11. Yang Y, Sharma R, Sharma A, et al. Lipid peroxidation and cell cycle signaling: 4-hydroxynonenal, a key molecule in stress mediated signaling. Acta Biochim Pol, 2003, 50(2): 319-336.
12. Awasthi YC, Sharma R, Cheng JZ, et al. Role of 4-hydroxynonenal in stress-mediated apoptosis signaling. Mol Aspects Med, 2003, 24(4-5): 219 -230.
13. Crosas B, Hyndman D J, Gallego O, et al. Human aldose reductase and human small intestine aldose reductase are efficient retinal reductases: consequences for retinoid metabolism. Biochem J, 2003, 373(Pt 3): 973-979.
14. Ruiz FX, Gallego O, Ardevol A, et al. Aldo-keto reductases from the AKR1B subfamily: retinoid specificity and control of cellular retinoic acid levels. Chem Biol Interact, 2009, 178(1-3): 171-177.
15. Marill J, Idres N, Capron CC, et al. Retinoic acid metabolism and mechanism of action: a review. Curr Drug Metab, 2003, 4(1): 1-10.
16. Ma J, Yan R, Zu X, et al. Aldo-keto reductase family 1 B10 affects fatty acid synthesis by regulating the stability of acetyl-CoA carboxylase-alpha in breast cancer cells. J Biol Chem, 2008, 283(6): 3418-3423.
17. Abu-Elheiga L, Matzuk MM, Kordari P, et al. Mutant mice lacking acetyl-CoA carboxylase 1 are embryonically lethal. Proc Natl Acad Sci U S A, 2005, 102(34): 12011-12016.
18. Franke WW, Grund C, Kuhn C, et al. Formation of cytoskeletal elements during mouse embryogenesis. III. Primary mesenchymal cells and the first appearance of vimentin filaments. Differentiation, 1982, 23(1): 43-59.
19. Lane E B, Hogan B L, Kurkinen M, et al. Co-expression of vimentin and cytokeratins in parietal endoderm cells of early mouse embryo. Nature, 1983, 303(5919): 701-704.
20. Nieminen M, Henttinen T, Merinen M, et al. Vimentin function in lymphocyte adhesion and transcellular migration. Nat Cell Biol, 2006, 8(2): 156-162.
21. Wang N, Stamenovic D. Mechanics of vimentin intermediate filaments.J Muscle Res Cell Motil, 2002, 23(5-6): 535-540.
22. Colucci-Guyon E, Portier MM, Dunia I, et al. Mice lacking vimentin develop and reproduce without an obvious phenotype. Cell, 1994, 79(4): 679-694.
23. Lang SH, Hyde C, Reid IN, et al. Enhanced expression of vimentin in motileprostate cell lines and in poorly differentiated and metastatic prostate carcinoma. Prostate, 2002, 52(4): 253-263.
24. Mosnier JF, Balique JG. Pleomorphic giant cell carcinoma of the esophagus with coexpression of cytokeratin and vimentin and neuroendocrine differentiation. Arch Pathol Lab Med, 2000, 124(1): 135- 138.
25. Yamamoto J, Ohshima K, Nabeshima K, et al. Comparative study of primary mammary small cell carcinoma, carcinoma with endocrine features and invasive ductal carcinoma. Oncol Rep, 2004, 11(4): 825-831.
26. Grieco V, Riccardi E, Belotti S, et al. Immunohistochemical study of porcine nephroblastoma. J Comp Pathol, 2006, 134(2-3): 143-151.
27. Hu L, Lau SH, Tzang CH, et al. Association of Vimentin overexpression and hepatocellular carcinoma metastasis. Oncogene, 2004, 23(1): 298-302.
28. Hendrix MJ, Seftor EA, Seftor RE, et al. Experimental co-expression of vimentin and keratin intermediate filaments in human breast cancer cells results in phenotypic interconversion and increased invasive behavior. Am J Pathol, 1997, 150(2): 483-495.
1. Tan FL, Yin JQ. RNAi, a new therapeutic strategy against viral infection. Cell Res, 2004, 14(6): 460-466.
2. Yano J, Hirabayashi K, Nakagawa S, et al. Antitumor activity of small interfering RNA/cationic liposome complex in mouse models of cancer. Clin Cancer Res, 2004, 10(22): 7721-7726.
3. Elbashir SM, Harborth J, Lendeckel W, et al. Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. Nature, 2001, 411(6836): 494-498.
4. Bertrand JR, Pottier M, Vekris A, et al. Comparison of antisense oligonucleotides and siRNAs in cell culture and in vivo. Biochem Biophys Res Commun, 2002, 296(4): 1000-1004.
5. Wang L, Tang ZY, Qin LX, et al. High-dose and long-term therapy with interferon-alfa inhibits tumor growth and recurrence in nude mice bearing human hepatocellular carcinoma xenografts with high metastatic potential. Hepatology, 2000, 32(1): 43-48.
6. Yan R, Zu X, Ma J, et al. Aldo-keto reductase family 1 B10 gene silencing results in growth inhibition of colorectal cancer cells: Implication for cancer intervention. Int J Cancer, 2007, 121(10): 2301-2306.
7. Cisarik-Fredenburg P. Discoveries in research on diabetic keratopathy. Optometry, 2001, 72(11):691-704.
8. Naya Y, Soh J, Ochiai A, et al. Erythrocyte aldose reductase correlates with erectile dysfunction in diabetic patients. Int J Impot Res, 2002, 14(4): 213 -216.
9. Penning TM. AKR1B10: a new diagnostic marker of non-small cell lung carcinoma in smokers. Clin Cancer Res, 2005, 11(5): 1687-1690.
10. Crosas B, Hyndman DJ, Gallego O, et al. Human aldose reductase and human small intestine aldose reductase are efficient retinal reductases: consequences for retinoid metabolism. Biochem J, 2003, 373(Pt3): 973 -979.
11. Cao D, Fan ST, Chung SS. Identification and characterization of a novel human aldose reductase-like gene. J Biol Chem, 1998, 273(19): 11429 -11435.
12. Zeindl-Eberhart E, Haraida S, Liebmann S, et al. Detection and identification of tumor-associated protein variants in human hepatocellular carcinomas. Hepatology, 2004, 39(2): 540-549.
13. Woenckhaus M, Klein-Hitpass L, Grepmeier U, et al. Smoking and cancer-related gene expression in bronchial epithelium and non-small-cell lung cancers. J Pathol, 2006, 210(2): 192-204.
14. Yoshitake H, Takahashi M, Ishikawa H, et al. Aldo-keto reductase family 1, member B10 in uterine carcinomas: a potential risk factor of recurrence after surgical therapy in cervical cancer. Int J Gynecol Cancer, 2007, 17(6): 1300-1306.
15. Breton J, Gage MC, Hay AW, et al. Proteomic screening of a cell line model of esophageal carcinogenesis identifies cathepsin D and aldo-keto reductase 1C2 and 1B10 dysregulation in Barrett's esophagus and esophageal adenocarcinoma. J Proteome Res, 2008, 7(5): 1953-1962.
16. Fire A, Xu S, Montgomery MK, et al. Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature, 1998, 391(6669):806-811.
17. Waterhouse PM. Defense and counterdefense in the plant world. Nat Genet, 2006, 38(2): 138-139.
18. Tijsterman M, Ketting RF, Plasterk RH. The genetics of RNA silencing.Annu Rev Genet, 2002, 36: 489-519.
19. Sen GL, Blau HM. A brief history of RNAi: the silence of the genes. FASEB J, 2006, 20(9): 1293-1299.
20. Lin SL, Chuong CM, Ying SY. A Novel mRNA-cDNA interference phenomenon for silencing bcl-2 expression in human LNCaP cells. Biochem Biophys Res Commun, 2001, 281(3): 639-644.
21. Onodera Y, Hashimoto S, Hashimoto A, et al. Expression of AMAP1, an ArfGAP, provides novel targets to inhibit breast cancer invasive activities. EMBO J, 2005, 24(5): 963-973.
22. Caplen NJ, Parrish S, Imani F, et al. Specific inhibition of gene expression by small double-stranded RNAs in invertebrate and vertebrate systems. Proc Natl Acad Sci U S A, 2001, 98(17): 9742-9747.
23. Fukumoto S, Yamauchi N, Moriguchi H, et al. Overexpression of the aldo-keto reductase family protein AKR1B10 is highly correlated with smokers' non-small cell lung carcinomas. Clin Cancer Res, 2005, 11(5): 1776-1785.
24. Zu X, Yan R, Robbins S, et al. Reduced 293T cell susceptibility to acrolein due to aldose reductase-like-1 protein expression. Toxicol Sci, 2007, 97(2): 562-568.
25. Yang Y, Sharma R, Sharma A, et al. Lipid peroxidation and cell cycle signaling: 4-hydroxynonenal, a key molecule in stress mediated signaling. Acta Biochim Pol, 2003, 50(2): 319-336.
26. Awasthi YC, Sharma R, Cheng JZ, et al. Role of 4-hydroxynonenal in stress-mediated apoptosis signaling. Mol Aspects Med, 2003, 24(4-5): 219- 230.
27. Abu-Elheiga L, Matzuk MM, Kordari P, et al. Mutant mice lackingacetyl-CoA carboxylase 1 are embryonically lethal. Proc Natl Acad Sci U S A, 2005, 102(34): 12011-12016.
28. Zang Y, Wang T, Xie W, et al. Regulation of acetyl CoA carboxylase and carnitine palmitoyl transferase-1 in rat adipocytes. Obes Res, 2005, 13(9): 1530-1539.
29. Chajes V, Cambot M, Moreau K, et al. Acetyl-CoA carboxylase alpha is essential to breast cancer cell survival. Cancer Res, 2006, 66(10): 5287 -5294.
30. Brusselmans K, De Schrijver E, Verhoeven G, et al. RNA interference-mediated silencing of the acetyl-CoA-carboxylase-alpha gene induces growth inhibition and apoptosis of prostate cancer cells. Cancer Res, 2005, 65(15): 6719-6725.
31. Ma J, Yan R, Zu X, et al. Aldo-keto reductase family 1 B10 affects fatty acid synthesis by regulating the stability of acetyl-CoA carboxylase-alpha in breast cancer cells. J Biol Chem, 2008, 283(6): 3418-3423.
32. Liu Z, Zhong L, Krishack PA, et al. Structure and promoter characterization of aldo-keto reductase family 1 B10 gene. Gene, 2009, 437(1-2): 39-44.
33. Lin CP, Liu JD, Chow JM, et al. Small-molecule c-Myc inhibitor, 10058-F4, inhibits proliferation, downregulates human telomerase reverse transcriptase and enhances chemosensitivity in human hepatocellular carcinoma cells. Anticancer Drugs, 2007, 18(2): 161-170.
34. Spector M, Nguyen VA, Sheng X, et al. Activation of mitogen-activated protein kinases is required for alpha1-adrenergic agonist-induced cell scattering in transfected HepG2 cells. Exp Cell Res, 2000, 258(1): 109-120.
35.郑杰,武忠弼,阮幼冰,等.大鼠实验性肝癌最早可测的分子标志.癌症,1996,04:28-30.
36. Mazumder S, Plesca D, Almasan, A. Caspase-3 activation is a critical determinant of genotoxic stress-induced apoptosis. Methods Mol Biol, 414: 13-21.
37. Korsmeyer, SJ. BCL-2 gene family and the regulation of programmed cell death. Cancer Res, 1999, 59(7 Suppl): 1693-1700.
1. Kononen J, Bubendorf L, Kallioniemi A, et al. Tissue microarrays for high-throughput molecular profiling of tumor specimens. Nat Med, 1998, 4(7): 844-847.
2. Gambarin-Gelwan M, Wolf DC, Shapiro R, et al. Sensitivity of commonly available screening tests in detecting hepatocellular carcinoma in cirrhotic patients undergoing liver transplantation. Am J Gastroenterol, 2000, 95(6): 1535-1538.
3. Duffy MJ. Role of tumor markers in patients with solid cancers: A criticalreview. Eur J Intern Med, 2007, 18(3): 175-184.
4. Okura H, Kononen J, Bubendorf L, et al. Usefulness of tumor marker in early diagnosis; Tissue microarrays for high-throughput molecular profiling of tumor specimens. Nippon Naika Gakkai Zasshi; Nat Med, 2005, 94; 4(12; 7): 2479-2485, 844-847.
5. Kumar S, Mohan A, Guleria R. Biomarkers in cancer screening, research and detection: present and future: a review. Biomarkers, 2006, 11(5): 385 -405.
6. Zolg W. The proteomic search for diagnostic biomarkers: lost in translation? Mol Cell Proteomics, 2006, 5(10): 1720-1726.
7. Chignard N, Beretta L. Proteomics for hepatocellular carcinoma marker discovery. Gastroenterology, 2004, 127(5 Suppl 1): S120-S125.
8. Feng JT, Liu YK, Song HY, et al. Heat-shock protein 27: a potential biomarker for hepatocellular carcinoma identified by serum proteome analysis. Proteomics, 2005, 5(17): 4581-4588.
9. Li C, Tan YX, Zhou H, et al. Proteomic analysis of hepatitis B virus-associated hepatocellular carcinoma: Identification of potential tumor markers. Proteomics, 2005, 5(4): 1125-1139.
10. Wang JX, Zhang B, Yu JK, et al. Application of serum protein fingerprinting coupled with artificial neural network model in diagnosis of hepatocellular carcinoma. Chin Med J (Engl), 2005, 118(15): 1278-1284.
11. Daniele B, Bencivenga A, Megna AS, et al. Alpha-fetoprotein and ultrasonography screening for hepatocellular carcinoma. Gastroenterology, 2004, 127(5 Suppl 1): S108-S112.
12. Trevisani F, D'Intino PE, Morselli-Labate AM, et al. Serum alpha-fetoprotein for diagnosis of hepatocellular carcinoma in patients withchronic liver disease: influence of HBsAg and anti-HCV status. J Hepatol, 2001, 34(4): 570-575.
13. Giardina MG, Matarazzo M, Morante R, et al. Serum alpha-L-fucosidase activity and early detection of hepatocellular carcinoma: a prospective study of patients with cirrhosis. Cancer, 1998, 83(12): 2468-2474.
14. Marrero JA, Su GL, Wei W, et al. Des-gamma carboxyprothrombin can differentiate hepatocellular carcinoma from nonmalignant chronic liver disease in american patients. Hepatology, 2003, 37(5): 1114-1121.
15. Capurro M, Wanless I R, Sherman M, et al. Glypican-3: a novel serum and histochemical marker for hepatocellular carcinoma. Gastroenterology, 2003, 125(1): 89-97.
16. Teramoto R, Minagawa H, Honda M, et al. Protein expression profile characteristic to hepatocellular carcinoma revealed by 2D-DIGE with supervised learning. Biochim Biophys Acta, 2008, 1784(5): 764-772.
1. Penning TM, Drury JE. Human aldo-keto reductases: Function, gene regulation, and single nucleotide polymorphisms. Arch Biochem Biophys, 2007, 464(2):241-250.
2. Hyndman D, Bauman DR, Heredia VV, et al. The aldo-keto reductase superfamily homepage. Chem Biol Interact, 2003, 143 -144: 621-631.
3. Ho HT, Chung SK, Law JW, et al. Aldose reductase-deficient mice develop nephrogenic diabetes insipidus. Mol Cell Biol, 2000, 20(16): 5840-5846.
4. Cisarik-Fredenburg P. Discoveries in research on diabetic keratopathy. Optometry, 2001, 72(11): 691-704.
5. Naya Y, Soh J, Ochiai A, et al. Erythrocyte aldose reductase correlates with erectile dysfunction in diabetic patients. Int J Impot Res, 2002, 14(4): 213-216.
6. Takahashi M, Fujii J, Miyoshi E, et al. Elevation of aldose reductase gene expression in rat primary hepatoma and hepatoma cell lines: implication in detoxification of cytotoxic aldehydes. Int J Cancer, 1995, 62(6): 749-754.
7. Cao D, Fan ST, Chung SS. Identification and characterization of a novelhuman aldose reductase-like gene. J Biol Chem, 1998, 273(19): 11429-11435.
8. Martin HJ, Maser E. Role of human aldo-keto-reductase AKR1B10 in the protection against toxic aldehydes. Chem Biol Interact, 2009, 178(1-3): 145-150.
9. Crosas B, Hyndman D J, Gallego O, et al. Human aldose reductase and human small intestine aldose reductase are efficient retinal reductases: consequences for retinoid metabolism. Biochem J, 2003, 373(Pt 3): 973-979.
10. Zeindl-Eberhart E, Jungblut PR, Otto A, et al. Identification of tumor-associated protein variants during rat hepatocarcinogenesis. Aldose reductase. J Biol Chem, 1994, 269(20): 14589-14594.
11. Zeindl-Eberhart E, Klugbauer S, Dimitrijevic N, et al. Proteome analysis of rat hepatomas: carcinogen-dependent tumor-associated protein variants. Electrophoresis, 2001, 22(14): 3009-3018.
12. Zeindl-Eberhart E, Jungblut PR, Otto A, et al. Further characterization of a rat hepatoma-derived aldose-reductase-like protein--organ distribution and modulation in vitro. Eur J Biochem, 1997, 247(3): 792-800.
13. Woenckhaus M, Klein-Hitpass L, Grepmeier U, et al. Smoking and cancer-related gene expression in bronchial epithelium and non-small-cell lung cancers. J Pathol, 2006, 210(2): 192-204.
14. Fukumoto S, Yamauchi N, Moriguchi H, et al. Overexpression of the aldo-keto reductase family protein AKR1B10 is highly correlated with smokers' non-small cell lung carcinomas. Clin Cancer Res, 2005, 11(5): 1776-1785.
15. Yoshitake H, Takahashi M, Ishikawa H, et al. Aldo-keto reductase family 1, member B10 in uterine carcinomas: a potential risk factor of recurrenceafter surgical therapy in cervical cancer. Int J Gynecol Cancer, 2007, 17(6): 1300-1306.
16. Breton J, Gage MC, Hay AW, et al. Proteomic screening of a cell line model of esophageal carcinogenesis identifies cathepsin D and aldo-keto reductase 1C2 and 1B10 dysregulation in Barrett's esophagus and esophageal adenocarcinoma. J Proteome Res, 2008, 7(5): 1953-1962.
17. Li CP, Goto A, Watanabe A, et al. AKR1B10 in usual interstitial pneumonia: expression in squamous metaplasia in association with smoking and lung cancer. Pathol Res Pract, 2008, 204(5): 295-304.
18. Zeindl-Eberhart E, Haraida S, Liebmann S, et al. Detection and identification of tumor-associated protein variants in human hepatocellular carcinomas. Hepatology, 2004, 39(2): 540-549.
19. Martin HJ, Breyer-Pfaff U, Wsol V, et al. Purification and characterization of akr1b10 from human liver: role in carbonyl reduction of xenobiotics. Drug Metab Dispos, 2006, 34(3): 464-470.
20. Zu X, Yan R, Robbins S, et al. Reduced 293T cell susceptibility to acrolein due to aldose reductase-like-1 protein expression. Toxicol Sci, 2007, 97(2): 562-568.
21. Yang Y, Sharma R, Sharma A, et al. Lipid peroxidation and cell cycle signaling: 4-hydroxynonenal, a key molecule in stress mediated signaling. Acta Biochim Pol, 2003, 50(2):319-336.
22. Awasthi YC, Sharma R, Cheng JZ, et al. Role of 4-hydroxynonenal in stress-mediated apoptosis signaling. Mol Aspects Med, 2003, 24 (4-5): 219-230.
23. Ou H, Haendeler J, Aebly MR, et al. Retinoic acid-induced tissue transglutaminase and apoptosis in vascular smooth muscle cells. Circ Res,2000, 87(10): 881-887.
24. Dawson MI, Zhang XK. Discovery and design of retinoic acid receptor and retinoid X receptor class- and subtype-selective synthetic analogs of all-trans-retinoic acid and 9-cis-retinoic acid. Curr Med Chem, 2002, 9(6): 623-637.
25. Marill J, Idres N, Capron CC, et al. Retinoic acid metabolism and mechanism of action: a review. Curr Drug Metab, 2003, 4(1): 1-10.
26. Gumireddy K, Sutton L N, Phillips PC, et al. All-trans-retinoic acid-induced apoptosis in human medulloblastoma: activation of caspase-3/poly (ADP-ribose) polymerase 1 pathway. Clin Cancer Res, 2003, 9(11): 4052-4059.
27. Brabender J, Metzger R, Salonga D, et al. Comprehensive expression analysis of retinoic acid receptors and retinoid X receptors in non-small cell lung cancer: implications for tumor development and prognosis. Carcinogenesis, 2005, 26(3): 525-530.
28. Ruiz FX, Gallego O, Ardevol A, et al. Aldo-keto reductases from the AKR1B subfamily: retinoid specificity and control of cellular retinoic acid levels. Chem Biol Interact, 2009, 178(1-3): 171-177.
29. Bannasch P. Pathogenesis of hepatocellular carcinoma: sequential cellular, molecular, and metabolic changes. Prog Liver Dis, 1996, 14: 161-197.
30. Stevens MJ, Henry DN, Thomas TP, et al. Aldose reductase gene expression and osmotic dysregulation in cultured human retinal pigment epithelial cells. Am J Physiol, 1993, 265(3P1): E428-E438.
31. Yan R, Zu X, Ma J, et al. Aldo-keto reductase family 1 B10 gene silencing results in growth inhibition of colorectal cancer cells: Implication for cancer intervention. Int J Cancer, 2007, 121 (10): 2301-2306.
32. Ruef J, Liu S Q, Bode C, et al. Involvement of aldose reductase in vascular smooth muscle cell growth and lesion formation after arterial injury. Arterioscler Thromb Vasc Biol, 2000, 20(7): 1745 -1752.
33. Ramana KV, Bhatnagar A, Srivastava S, et al. Mitogenic responses of vascular smooth muscle cells to lipid peroxidation-derived aldehyde 4-hydroxy-trans-2-nonenal (HNE): role of aldose reductase-catalyzed reduction of the HNE-glutathione conjugates in regulating cell growth. J Biol Chem, 2006, 281(26): 17652-17660.
34. Ramana KV, Chandra D, Srivastava S, et al. Aldose reductase mediates the mitogenic signals of cytokines. Chem Biol Interact, 2003, 143-144: 587-596.
35. Swinnen JV, Roskams T, Joniau S, et al. Overexpression of fatty acid synthase is an early and common event in the development of prostate cancer. Int J Cancer, 2002, 98(1): 19-22.
36. Rossi S, Graner E, Febbo P, et al. Fatty acid synthase expression defines distinct molecular signatures in prostate cancer. Mol Cancer Res, 2003, 1(10): 707-715.
37. Yahagi N, Shimano H, Hasegawa K, et al. Co-ordinate activation of lipogenic enzymes in hepatocellular carcinoma. Eur J Cancer, 2005, 41(9): 1316-1322.
38. Rouquette-Jazdanian AK, Pelassy C, Breittmayer JP, et al. Metabolic labelling of membrane microdomains/rafts in Jurkat cells indicates the presence of glycerophospholipids implicated in signal transduction by the CD3 T-cell receptor. Biochem J, 2002, 363(Pt3): 645-655.
39. Swinnen JV, Heemers H, Van De Sande T, et al. Androgens, lipogenesis and prostate cancer. J Steroid Biochem Mol Biol, 2004, 92(4): 273-279.
40. Ma J, Yan R, Zu X, et al. Aldo-keto reductase family 1 B10 affects fatty acidsynthesis by regulating the stability of acetyl-CoA carboxylase-alpha in breast cancer cells. J Biol Chem, 2008, 283(6): 3418-3423.
41. Zang Y, Wang T, Xie W, et al. Regulation of acetyl CoA carboxylase and carnitine palmitoyl transferase-1 in rat adipocytes. Obes Res, 2005, 13(9): 1530-1539.
42. Swinnen JV, Brusselmans K, Verhoeven G. Increased lipogenesis in cancer cells: new players, novel targets. Curr Opin Clin Nutr Metab Care, 2006, 9(4):358-365.
43. Abu-Elheiga L, Matzuk MM, Kordari P, et al. Mutant mice lacking acetyl-CoA carboxylase 1 are embryonically lethal. Proc Natl Acad Sci U S A, 2005, 102(34): 12011-12016.
44. Chajes V, Cambot M, Moreau K, et al. Acetyl-CoA carboxylase alpha is essential to breast cancer cell survival. Cancer Res, 2006, 66(10): 5287-5294.
45. Brusselmans K, De Schrijver E, Verhoeven G, et al. RNA interference-mediated silencing of the acetyl-CoA-carboxylase-alpha gene induces growth inhibition and apoptosis of prostate cancer cells. Cancer Res, 2005, 65(15): 6719-6725.
46. Tong L, Jr Harwood H J. Acetyl-coenzyme A carboxylases: versatile targets for drug discovery. J Cell Biochem, 2006, 99(6): 1476-1488
47. Smith J A, Gaikwad A, Ramondetta LM, et al. Determination of the mechanism of gemcitabine modulation of cisplatin drug resistance in panel of human endometrial cancer cell lines. Gynecol Oncol, 2006, 103(2): 518-522.
48. Kijima H, Ueyama Y. Molecular mechanism of drug resistance in colorectal cancer. Nippon Rinsho, 2003, 61 Suppl 7: 303-309.
49. Sun CA, Wang LY, Chen CJ, et al. Genetic polymorphisms of glutathione S-transferases M1 and T1 associated with susceptibility to aflatoxin-related hepatocarcinogenesis among chronic hepatitis B carriers: a nested case-control study in Taiwan. Carcinogenesis, 2001, 22(8): 1289-1294.
50. Ishimura H. Chemosensitivity test on hepatocellular carcinoma (HCC) and drug resistance. Hokkaido Igaku Zasshi, 1996, 71(6): 689-698.
51. Lee KW, Ko BC, Jiang Z, et al. Overexpression of aldose reductase in liver cancers may contribute to drug resistance. Anticancer Drugs, 2001, 12(2):129-132.
52.王建,万腊香,吴孟津,等.醛糖还原酶相似基因转染细胞对含碳酰基抗癌药物耐药的研究.中华肝脏病杂志, 2002(06): 46-49.
53.杨向东,唐大年,王建,等.转ARL-1基因HepG2细胞耐药相关基因的筛选.癌症, 2003(12): 1289-1295.
54. Penning TM. AKR1B10: a new diagnostic marker of non-small cell lung carcinoma in smokers. Clin Cancer Res, 2005, 11(5): 1687 -1690.
55. Hamada Y, Nakamura J. Clinical potential of aldose reductase inhibitors in diabetic neuropathy. Treat Endocrinol, 2004, 3(4): 245- 255.
56. Gallego O, Ruiz FX, Ardevol A, et al. Structural basis for the high all-trans-retinaldehyde reductase activity of the tumor marker AKR1B10. Proc Natl Acad Sci U S A, 2007, 104(52): 20764-20769.
57. Ko Ec, Wang X, Ferron S. Immunotherapy of malignant diseases. Challenges and strategies. Int Arch Allergy Immunol, 2003, 132(4): 294-309.
58. Weber WA, Grosu AL, Czernin J. Technology Insight: advances in molecular imaging and an appraisal of PET/CT scanning. Nat Clin Pract Oncol, 2008, 5(3): 160-170.
2. Parkin DM, Bray F, Ferlay J, et al. Global cancer statistics, 2002. CA Cancer J Clin, 2005, 55(2): 74-108.
3. Parkin DM, Bray F, Ferlay J, et al. Estimating the world cancer burden: Globocan 2000. Int J Cancer, 2001, 94(2): 153-156.
4. Wright LM, Kreikemeier JT, Fimmel CJ. A concise review of serum markers for hepatocellular cancer. Cancer Detect Prev, 2007, 31(1): 35-44.
5.张思维,李连弟,鲁凤珠,等.中国1990～1992年原发性肝癌死亡调查分析.中华肿瘤杂志, 1999(04): 5-9.
6. Ward DG, Cheng Y, N'Kontchou G, et al. Changes in the serum proteome associated with the development of hepatocellular carcinoma in hepatitis C-related cirrhosis. Br J Cancer, 2006, 94(2): 287-292.
7. Bosch FX, Ribes J, Diaz M, et al. Primary liver cancer: worldwide incidence and trends. Gastroenterology, 2004, 127(5 Suppl 1): S5-S16.
8. Wands JR. Prevention of hepatocellular carcinoma. N Engl J Med, 2004, 351(15): 1567-1570.
9.郑树森,吴应盛,梁廷波.原发性肝癌420例综合治疗报告.中华普通外科杂志, 2003(12): 3-5.
10. Daniele B, Bencivenga A, Megna AS, et al. Alpha-fetoprotein and ultrasonography screening for hepatocellular carcinoma. Gastroenterology, 2004, 127(5 Suppl 1): S108-S112.
11. Trevisani F, D'Intino PE, Morselli-Labate AM, et al. Serum alpha-fetoprotein for diagnosis of hepatocellular carcinoma in patients with chronic liver disease: influence of HBsAg and anti-HCV status. J Hepatol, 2001, 34(4): 570-575.
12. Tang ZY, Yu YQ, Zhou XD, et al. Subclinical hepatocellular carcinoma: an analysis of 391 patients. J Surg Oncol Suppl, 1993, 3:55-58.
13. Okubo H, Moriyama M, Tanaka N, et al. Detection of serum and intrahepatic hepatocyte growth factor during DEN-induced carcinogenesis in the rat. Hepatol Res, 2002, 24(4): 385-394.
14. Han KQ, Gu W, Hu X, et al. Establishment and biological characteristics of orthotopic transplantation tumor model of hepatocellular carcinoma in miceJ. Zhong Xi Yi Jie He Xue Bao, 2004, 2(5): 372-374.
15. Travis CC, Mcclain TW, Birkner PD. Diethylnitrosamine-induced hepatocarcinogenesis in rats: a theoretical study. Toxicol Appl Pharmacol, 1991, 109(2): 289-304.
16. Yoshiji H, Yoshii J, Ikenaka Y, et al. Inhibition of renin-angiotensin system attenuates liver enzyme-altered preneoplastic lesions and fibrosis development in rats. J Hepatol, 2002, 37(1): 22-30.
17. Yao DF, Dong ZZ, Yao DB, et al. Abnormal expression of hepatoma-derived gamma-glutamyltransferase subtyping and its early alteration for carcinogenesis of hepatocytes. Hepatobiliary Pancreat Dis Int, 2004, 3(4): 564-570.
18. Li Y, Yan RQ, Qin GZ, et al. Reliability of a short-term test for hepatocarcinogenesis induced by aflatoxin B1. IARC Sci Publ, 1991 (105): 431-433.
19.唐小岚,段小娴,苏建家,等.金花茶和银杏叶对2-乙基亚硝胺致大鼠肝癌前病变抑制作用的初步研究.实用癌症杂志, 2007(3): 224-227.
20.刘启福,罗丹,苏建家,等.实验性大鼠肝癌变过程中p16和Rb基因表达的动态观察.临床与实验病理学杂志, 1999(3): 41-43.
21. Fields S. Proteomics. Proteomics in genomeland. Science, 2001, 291 (5507): 1221-1224.
22. Pandey A, Mann M. Proteomics to study genes and genomes. Nature, 2000, 405(6788): 837-846.
23. Gou LT, Tong AP, Chen LJ, et al. Comparative plasma membrane-associated proteomics of immortalized human hepatocytes. Biochemistry (Mosc), 2008, 73(11):1200-1206.
24. Xu J, Khor K A, Sui J, et al. Comparative proteomics profile of osteoblasts cultured on dissimilar hydroxyapatite biomaterials: an iTRAQ-coupled 2-D LC-MS/MS analysis. Proteomics, 2008, 8(20): 4249-4258.
25. Fujii K, Kondo T, Yokoo H, et al. Proteomic study of human hepatocellular carcinoma using two-dimensional difference gel electrophoresis with saturation cysteine dye. Proteomics, 2005, 5(5): 1411-1422.
26. Cui J F, Liu Y K, Pan B S, et al. Differential proteomic analysis of human hepatocellular carcinoma cell line metastasis-associated proteins. J Cancer Res Clin Oncol, 2004, 130(10): 615-622.
27. Cui J F, Liu YK, Zhang LJ, et al. Identification of metastasis candidate proteins among HCC cell lines by comparative proteome and biological function analysis of S100A4 in metastasis in vitro. Proteomics, 2006, 6(22): 5953-5961.
28. Yi X, Luk JM, Lee NP, et al. Association of mortalin (HSPA9) with liver cancer metastasis and prediction for early tumor recurrence. Mol Cell Proteomics, 2008, 7(2): 315-325.
29. Fire A, Xu S, Montgomery MK, et al. Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature, 1998, 391(6669):806-811.
30. Sen G L, Blau H M. A brief history of RNAi: the silence of the genes. FASEB J, 2006, 20(9):1293-1299.
31. Stamps AC, Terrett JA, Adam PJ. Application of in situ reverse trancriptase-polymerase chain reaction (RT-PCR) to tissue microarrays. J Nanobiotechnology, 2003, 1(1): 3.
32. Kononen J, Bubendorf L, Kallioniemi A, et al. Tissue microarrays for high-throughput molecular profiling of tumor specimens. Nat Med, 1998, 4(7): 844-847.
33. Yan R, Zu X, Ma J, et al. Aldo-keto reductase family 1 B10 gene silencing results in growth inhibition of colorectal cancer cells: Implication for cancer intervention. Int J Cancer, 2007, 121(10): 2301-2306.
34. Fukumoto S, Yamauchi N, Moriguchi H, et al. Overexpression of the aldo-keto reductase family protein AKR1B10 is highly correlated with smokers' non-small cell lung carcinomas. Clin Cancer Res, 2005, 11 (5): 1776-1785.
1. Yoshiji H, Yoshii J, Ikenaka Y, et al. Inhibition of renin-angiotensin system attenuates liver enzyme-altered preneoplastic lesions and fibrosis development in rats. J Hepatol, 2002, 37(1): 22-30.
2. Chakraborty T, Chatterjee A, Rana A, et al. Cell proliferation and hepatocarcinogenesis in rat initiated by diethylnitrosamine and promoted by phenobarbital: potential roles of early DNA damage and liver metallothionein expression. Life Sci, 2007, 81(6): 489-499.
3. Swinbanks D. Government backs proteome proposal. Nature, 1995, 378(6558):653.
4. Gou LT, Tong AP, Chen LJ, et al. Comparative plasma membrane-associated proteomics of immortalized human hepatocytes. Biochemistry (Mosc), 2008, 73(11):1200-1206.
5. Xu J, Khor K A, Sui J, et al. Comparative proteomics profile of osteoblasts cultured on dissimilar hydroxyapatite biomaterials: an iTRAQ-coupled 2-D LC-MS/MS analysis. Proteomics, 2008, 8(20): 4249-4258.
6. Petricoin EF, Zoon KC, Kohn EC, et al. Clinical proteomics: translating benchside promise into bedside reality. Nat Rev Drug Discov, 2002, 1(9): 683-695.
7. Ward DG, Cheng Y, N'Kontchou G, et al. Changes in the serum proteome associated with the development of hepatocellular carcinoma in hepatitis C-related cirrhosis. Br J Cancer, 2006, 94(2): 287-292.
8. Ding SJ, Li Y, Tan YX, et al. From proteomic analysis to clinical significance: overexpression of cytokeratin 19 correlates with hepatocellular carcinoma metastasis. Mol Cell Proteomics, 2004, 3(1): 73-81.
9. Farber E. The sequential analysis of liver cancer induction biochem. Et Biophys Acta, 1980, 605:149- 154.
10. Li Y, Yan RQ, Qin GZ, et al. Reliability of a short-term test for hepatocarcinogenesis induced by aflatoxin B1. IARC Sci Publ, 1991(105): 431-433.
11.唐小岚,段小娴,苏建家,等.金花茶和银杏叶对2-乙基亚硝胺致大鼠肝癌前病变抑制作用的初步研究.实用癌症杂志, 2007(3): 224-227.
12. Solt D, Farber E. New principle for the analysis of chemical carcinogenesis. Nature, 1976, 263(5580):701-703.
13. Schwartzenberg-Bar-Yoseph F, Armoni M, Karnieli E. The tumor suppressor p53 down-regulates glucose transporters GLUT1 and GLUT4 gene expression. Cancer Res, 2004, 64(7): 2627-2633.
14. Miyagi T, Hori O, Koshida K, et al. Antitumor effect of reduction of 150-kDa oxygen-regulated protein expression on human prostate cancer cells. Int J Urol, 2002, 9(10): 577-585.
15. Asahi H, Koshida K, Hori O, et al. Immunohistochemical detection of the
150-kDa oxygen-regulated protein in bladder cancer. BJU Int, 2002, 90(4): 462-466.
16. Siegenthaler G, Hotz R, Chatellard-Gruaz D, et al. Characterization and expression of a novel human fatty acid-binding protein: the epidermal type(E-FABP). Biochem Biophys Res Commun, 1993, 190(2): 482-487.
17. Chmurzynska A. The multigene family of fatty acid-binding proteins (FABPs): function, structure and polymorphism. J Appl Genet, 2006, 47(1): 39-48.
18. Masouye I, Hagens G, Van Kuppevelt T H, et al. Endothelial cells of the human microvasculature express epidermal fatty acid-binding protein. Circ Res, 1997, 81(3): 297-303.
19. Loeffler-Ragg J, Skvortsov S, Sarg B, et al. Gefitinib-responsive EGFR-positive colorectal cancers have different proteome profiles from non-responsive cell lines. Eur J Cancer, 2005, 41(15): 2338-2346.
20. Uma RS, Naresh KN, D'Cruz AK, et al. Metastasis of squamous cell carcinoma of the oral tongue is associated with down-regulation of epidermal fatty acid binding protein (E-FABP).Oral Oncol, 2007, 43(1): 27 -32.
21. Rauch J, Ahlemann M, Schaffrik M, et al. Allogenic antibody-mediated identification of head and neck cancer antigens. Biochem Biophys Res Commun, 2004, 323(1):156-162.
22. Munz M, Zeidler R, Gires O. The tumour-associated antigen EpCAM upregulates the fatty acid binding protein E-FABP. Cancer Lett, 2005, 225(1): 151-157.
23. Fujii K, Kondo T, Yokoo H, et al. Proteomic study of human hepatocellular carcinoma using two-dimensional difference gel electrophoresis with saturation cysteine dye. Proteomics, 2005, 5(5): 1411 -1422.
24. Yamaguchi M, Tsurusaki Y, Misawa H, et al. Potential role of regucalcin as a specific biochemical marker of chronic liver injury with carbon tetrachloride administration in rats. Mol Cell Biochem, 2002, 241(1-2):61-67.
25. Tsurusaki Y, Yamaguchi M. Role of regucalcin in liver nuclear function: binding of regucalcin to nuclear protein or DNA and modulation of tumor-related gene expression. Int J Mol Med, 2004, 14(2): 277-281.
26. Fukaya Y, Yamaguchi M. Overexpression of regucalcin suppresses apoptotic cell death in the cloned rat hepatoma H4-II-E cells induced by a naturally occurring isothiocyanate sulforaphane. Int J Mol Med, 2005, 15(5): 853-857.
27. Izumi T, Tsurusaki Y, Yamaguchi M. Suppressive effect of endogenous regucalcin on nitric oxide synthase activity in cloned rat hepatoma H4-II-E cells overexpressing regucalcin. J Cell Biochem, 2003, 89(4): 800-807.
28. Fontanini G, Vignati S, Chine S, et al. 67-Kilodalton laminin receptor expression correlates with worse prognostic indicators in non-small cell lung carcinomas. Clin Cancer Res, 1997, 3(2): 227-231.
29. Zheng S, Ruan Y, Wu Z, et al. The relationship between 67KD laminin receptor expression and metastasis of hepatocellular carcinoma. J Tongji Med Univ, 1997, 17(4): 200-202, 224.
30.梁宏洁,韦薇,康晓楠,等. DEN诱发大鼠肝癌过程中癌前病变阶段差异表达蛋白的筛选. (已投稿)
31. Jr Brozinick J T, Berkemeier B A, Elmendorf J S. "Acting" on GLUT4: membrane & cytoskeletal components of insulin action. Curr Diabetes Rev, 2007, 3(2): 111-122.
32. Jr Brozinick JT, Mccoid SC, Reynolds TH, et al. GLUT4 overexpression in db/db mice dose-dependently ameliorates diabetes but is not a lifelong cure. Diabetes, 2001, 50(3): 593-600.
1. Yan R, Zu X, Ma J, et al. Aldo-keto reductase family 1 B10 gene silencing results in growth inhibition of colorectal cancer cells: Implication for cancer intervention. Int J Cancer, 2007, 121(10): 2301-2306.
2. Fukumoto S, Yamauchi N, Moriguchi H, et al. Overexpression of the aldo-keto reductase family protein AKR1B10 is highly correlated with smokers' non-small cell lung carcinomas. Clin Cancer Res, 2005, 11(5): 1776-1785.
3. Zu X, Yan R, Robbins S, et al. Reduced 293T cell susceptibility to acrolein due to aldose reductase-like-1 protein expression. Toxicol Sci, 2007, 97(2): 562-568.
4. Baumgart E, Cohen MS, Silva NB, et al. Identification and prognostic significance of an epithelial-mesenchymal transition expression profile in human bladder tumors. Clin Cancer Res, 2007, 13(6): 1685-1694.
5. Ngan CY, Yamamoto H, Seshimo I, et al. Quantitative evaluation of vimentin expression in tumour stroma of colorectal cancer. Br J Cancer,2007, 96(6): 986-992.
6. Penning TM, Drury JE. Human aldo-keto reductases: Function, gene regulation, and single nucleotide polymorphisms.Arch Biochem Biophys, 2007, 464(2): 241-250.
7. Zeindl-Eberhart E, Jungblut PR, Otto A, et al. Identification of tumor-associated protein variants during rat hepatocarcinogenesis. Aldose reductase. J Biol Chem, 1994, 269(20): 14589-14594.
8. Zeindl-Eberhart E, Klugbauer S, Dimitrijevic N, et al. Proteome analysis of rat hepatomas: carcinogen-dependent tumor-associated protein variants. Electrophoresis, 2001, 22(14): 3009-3018.
9. Woenckhaus M, Klein-Hitpass L, Grepmeier U, et al. Smoking and cancer-related gene expression in bronchial epithelium and non-small-cell lung cancers. J Pathol, 2006, 210(2): 192-204.
10. Breton J, Gage MC, Hay AW, et al. Proteomic screening of a cell line model of esophageal carcinogenesis identifies cathepsin D and aldo-keto reductase 1C2 and 1B10 dysregulation in Barrett's esophagus and esophageal adenocarcinoma. J Proteome Res, 2008, 7(5): 1953-1962.
11. Yang Y, Sharma R, Sharma A, et al. Lipid peroxidation and cell cycle signaling: 4-hydroxynonenal, a key molecule in stress mediated signaling. Acta Biochim Pol, 2003, 50(2): 319-336.
12. Awasthi YC, Sharma R, Cheng JZ, et al. Role of 4-hydroxynonenal in stress-mediated apoptosis signaling. Mol Aspects Med, 2003, 24(4-5): 219 -230.
13. Crosas B, Hyndman D J, Gallego O, et al. Human aldose reductase and human small intestine aldose reductase are efficient retinal reductases: consequences for retinoid metabolism. Biochem J, 2003, 373(Pt 3): 973-979.
14. Ruiz FX, Gallego O, Ardevol A, et al. Aldo-keto reductases from the AKR1B subfamily: retinoid specificity and control of cellular retinoic acid levels. Chem Biol Interact, 2009, 178(1-3): 171-177.
15. Marill J, Idres N, Capron CC, et al. Retinoic acid metabolism and mechanism of action: a review. Curr Drug Metab, 2003, 4(1): 1-10.
16. Ma J, Yan R, Zu X, et al. Aldo-keto reductase family 1 B10 affects fatty acid synthesis by regulating the stability of acetyl-CoA carboxylase-alpha in breast cancer cells. J Biol Chem, 2008, 283(6): 3418-3423.
17. Abu-Elheiga L, Matzuk MM, Kordari P, et al. Mutant mice lacking acetyl-CoA carboxylase 1 are embryonically lethal. Proc Natl Acad Sci U S A, 2005, 102(34): 12011-12016.
18. Franke WW, Grund C, Kuhn C, et al. Formation of cytoskeletal elements during mouse embryogenesis. III. Primary mesenchymal cells and the first appearance of vimentin filaments. Differentiation, 1982, 23(1): 43-59.
19. Lane E B, Hogan B L, Kurkinen M, et al. Co-expression of vimentin and cytokeratins in parietal endoderm cells of early mouse embryo. Nature, 1983, 303(5919): 701-704.
20. Nieminen M, Henttinen T, Merinen M, et al. Vimentin function in lymphocyte adhesion and transcellular migration. Nat Cell Biol, 2006, 8(2): 156-162.
21. Wang N, Stamenovic D. Mechanics of vimentin intermediate filaments.J Muscle Res Cell Motil, 2002, 23(5-6): 535-540.
22. Colucci-Guyon E, Portier MM, Dunia I, et al. Mice lacking vimentin develop and reproduce without an obvious phenotype. Cell, 1994, 79(4): 679-694.
23. Lang SH, Hyde C, Reid IN, et al. Enhanced expression of vimentin in motileprostate cell lines and in poorly differentiated and metastatic prostate carcinoma. Prostate, 2002, 52(4): 253-263.
24. Mosnier JF, Balique JG. Pleomorphic giant cell carcinoma of the esophagus with coexpression of cytokeratin and vimentin and neuroendocrine differentiation. Arch Pathol Lab Med, 2000, 124(1): 135- 138.
25. Yamamoto J, Ohshima K, Nabeshima K, et al. Comparative study of primary mammary small cell carcinoma, carcinoma with endocrine features and invasive ductal carcinoma. Oncol Rep, 2004, 11(4): 825-831.
26. Grieco V, Riccardi E, Belotti S, et al. Immunohistochemical study of porcine nephroblastoma. J Comp Pathol, 2006, 134(2-3): 143-151.
27. Hu L, Lau SH, Tzang CH, et al. Association of Vimentin overexpression and hepatocellular carcinoma metastasis. Oncogene, 2004, 23(1): 298-302.
28. Hendrix MJ, Seftor EA, Seftor RE, et al. Experimental co-expression of vimentin and keratin intermediate filaments in human breast cancer cells results in phenotypic interconversion and increased invasive behavior. Am J Pathol, 1997, 150(2): 483-495.
1. Tan FL, Yin JQ. RNAi, a new therapeutic strategy against viral infection. Cell Res, 2004, 14(6): 460-466.
2. Yano J, Hirabayashi K, Nakagawa S, et al. Antitumor activity of small interfering RNA/cationic liposome complex in mouse models of cancer. Clin Cancer Res, 2004, 10(22): 7721-7726.
3. Elbashir SM, Harborth J, Lendeckel W, et al. Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. Nature, 2001, 411(6836): 494-498.
4. Bertrand JR, Pottier M, Vekris A, et al. Comparison of antisense oligonucleotides and siRNAs in cell culture and in vivo. Biochem Biophys Res Commun, 2002, 296(4): 1000-1004.
5. Wang L, Tang ZY, Qin LX, et al. High-dose and long-term therapy with interferon-alfa inhibits tumor growth and recurrence in nude mice bearing human hepatocellular carcinoma xenografts with high metastatic potential. Hepatology, 2000, 32(1): 43-48.
6. Yan R, Zu X, Ma J, et al. Aldo-keto reductase family 1 B10 gene silencing results in growth inhibition of colorectal cancer cells: Implication for cancer intervention. Int J Cancer, 2007, 121(10): 2301-2306.
7. Cisarik-Fredenburg P. Discoveries in research on diabetic keratopathy. Optometry, 2001, 72(11):691-704.
8. Naya Y, Soh J, Ochiai A, et al. Erythrocyte aldose reductase correlates with erectile dysfunction in diabetic patients. Int J Impot Res, 2002, 14(4): 213 -216.
9. Penning TM. AKR1B10: a new diagnostic marker of non-small cell lung carcinoma in smokers. Clin Cancer Res, 2005, 11(5): 1687-1690.
10. Crosas B, Hyndman DJ, Gallego O, et al. Human aldose reductase and human small intestine aldose reductase are efficient retinal reductases: consequences for retinoid metabolism. Biochem J, 2003, 373(Pt3): 973 -979.
11. Cao D, Fan ST, Chung SS. Identification and characterization of a novel human aldose reductase-like gene. J Biol Chem, 1998, 273(19): 11429 -11435.
12. Zeindl-Eberhart E, Haraida S, Liebmann S, et al. Detection and identification of tumor-associated protein variants in human hepatocellular carcinomas. Hepatology, 2004, 39(2): 540-549.
13. Woenckhaus M, Klein-Hitpass L, Grepmeier U, et al. Smoking and cancer-related gene expression in bronchial epithelium and non-small-cell lung cancers. J Pathol, 2006, 210(2): 192-204.
14. Yoshitake H, Takahashi M, Ishikawa H, et al. Aldo-keto reductase family 1, member B10 in uterine carcinomas: a potential risk factor of recurrence after surgical therapy in cervical cancer. Int J Gynecol Cancer, 2007, 17(6): 1300-1306.
15. Breton J, Gage MC, Hay AW, et al. Proteomic screening of a cell line model of esophageal carcinogenesis identifies cathepsin D and aldo-keto reductase 1C2 and 1B10 dysregulation in Barrett's esophagus and esophageal adenocarcinoma. J Proteome Res, 2008, 7(5): 1953-1962.
16. Fire A, Xu S, Montgomery MK, et al. Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature, 1998, 391(6669):806-811.
17. Waterhouse PM. Defense and counterdefense in the plant world. Nat Genet, 2006, 38(2): 138-139.
18. Tijsterman M, Ketting RF, Plasterk RH. The genetics of RNA silencing.Annu Rev Genet, 2002, 36: 489-519.
19. Sen GL, Blau HM. A brief history of RNAi: the silence of the genes. FASEB J, 2006, 20(9): 1293-1299.
20. Lin SL, Chuong CM, Ying SY. A Novel mRNA-cDNA interference phenomenon for silencing bcl-2 expression in human LNCaP cells. Biochem Biophys Res Commun, 2001, 281(3): 639-644.
21. Onodera Y, Hashimoto S, Hashimoto A, et al. Expression of AMAP1, an ArfGAP, provides novel targets to inhibit breast cancer invasive activities. EMBO J, 2005, 24(5): 963-973.
22. Caplen NJ, Parrish S, Imani F, et al. Specific inhibition of gene expression by small double-stranded RNAs in invertebrate and vertebrate systems. Proc Natl Acad Sci U S A, 2001, 98(17): 9742-9747.
23. Fukumoto S, Yamauchi N, Moriguchi H, et al. Overexpression of the aldo-keto reductase family protein AKR1B10 is highly correlated with smokers' non-small cell lung carcinomas. Clin Cancer Res, 2005, 11(5): 1776-1785.
24. Zu X, Yan R, Robbins S, et al. Reduced 293T cell susceptibility to acrolein due to aldose reductase-like-1 protein expression. Toxicol Sci, 2007, 97(2): 562-568.
25. Yang Y, Sharma R, Sharma A, et al. Lipid peroxidation and cell cycle signaling: 4-hydroxynonenal, a key molecule in stress mediated signaling. Acta Biochim Pol, 2003, 50(2): 319-336.
26. Awasthi YC, Sharma R, Cheng JZ, et al. Role of 4-hydroxynonenal in stress-mediated apoptosis signaling. Mol Aspects Med, 2003, 24(4-5): 219- 230.
27. Abu-Elheiga L, Matzuk MM, Kordari P, et al. Mutant mice lackingacetyl-CoA carboxylase 1 are embryonically lethal. Proc Natl Acad Sci U S A, 2005, 102(34): 12011-12016.
28. Zang Y, Wang T, Xie W, et al. Regulation of acetyl CoA carboxylase and carnitine palmitoyl transferase-1 in rat adipocytes. Obes Res, 2005, 13(9): 1530-1539.
29. Chajes V, Cambot M, Moreau K, et al. Acetyl-CoA carboxylase alpha is essential to breast cancer cell survival. Cancer Res, 2006, 66(10): 5287 -5294.
30. Brusselmans K, De Schrijver E, Verhoeven G, et al. RNA interference-mediated silencing of the acetyl-CoA-carboxylase-alpha gene induces growth inhibition and apoptosis of prostate cancer cells. Cancer Res, 2005, 65(15): 6719-6725.
31. Ma J, Yan R, Zu X, et al. Aldo-keto reductase family 1 B10 affects fatty acid synthesis by regulating the stability of acetyl-CoA carboxylase-alpha in breast cancer cells. J Biol Chem, 2008, 283(6): 3418-3423.
32. Liu Z, Zhong L, Krishack PA, et al. Structure and promoter characterization of aldo-keto reductase family 1 B10 gene. Gene, 2009, 437(1-2): 39-44.
33. Lin CP, Liu JD, Chow JM, et al. Small-molecule c-Myc inhibitor, 10058-F4, inhibits proliferation, downregulates human telomerase reverse transcriptase and enhances chemosensitivity in human hepatocellular carcinoma cells. Anticancer Drugs, 2007, 18(2): 161-170.
34. Spector M, Nguyen VA, Sheng X, et al. Activation of mitogen-activated protein kinases is required for alpha1-adrenergic agonist-induced cell scattering in transfected HepG2 cells. Exp Cell Res, 2000, 258(1): 109-120.
35.郑杰,武忠弼,阮幼冰,等.大鼠实验性肝癌最早可测的分子标志.癌症,1996,04:28-30.
36. Mazumder S, Plesca D, Almasan, A. Caspase-3 activation is a critical determinant of genotoxic stress-induced apoptosis. Methods Mol Biol, 414: 13-21.
37. Korsmeyer, SJ. BCL-2 gene family and the regulation of programmed cell death. Cancer Res, 1999, 59(7 Suppl): 1693-1700.
1. Kononen J, Bubendorf L, Kallioniemi A, et al. Tissue microarrays for high-throughput molecular profiling of tumor specimens. Nat Med, 1998, 4(7): 844-847.
2. Gambarin-Gelwan M, Wolf DC, Shapiro R, et al. Sensitivity of commonly available screening tests in detecting hepatocellular carcinoma in cirrhotic patients undergoing liver transplantation. Am J Gastroenterol, 2000, 95(6): 1535-1538.
3. Duffy MJ. Role of tumor markers in patients with solid cancers: A criticalreview. Eur J Intern Med, 2007, 18(3): 175-184.
4. Okura H, Kononen J, Bubendorf L, et al. Usefulness of tumor marker in early diagnosis; Tissue microarrays for high-throughput molecular profiling of tumor specimens. Nippon Naika Gakkai Zasshi; Nat Med, 2005, 94; 4(12; 7): 2479-2485, 844-847.
5. Kumar S, Mohan A, Guleria R. Biomarkers in cancer screening, research and detection: present and future: a review. Biomarkers, 2006, 11(5): 385 -405.
6. Zolg W. The proteomic search for diagnostic biomarkers: lost in translation? Mol Cell Proteomics, 2006, 5(10): 1720-1726.
7. Chignard N, Beretta L. Proteomics for hepatocellular carcinoma marker discovery. Gastroenterology, 2004, 127(5 Suppl 1): S120-S125.
8. Feng JT, Liu YK, Song HY, et al. Heat-shock protein 27: a potential biomarker for hepatocellular carcinoma identified by serum proteome analysis. Proteomics, 2005, 5(17): 4581-4588.
9. Li C, Tan YX, Zhou H, et al. Proteomic analysis of hepatitis B virus-associated hepatocellular carcinoma: Identification of potential tumor markers. Proteomics, 2005, 5(4): 1125-1139.
10. Wang JX, Zhang B, Yu JK, et al. Application of serum protein fingerprinting coupled with artificial neural network model in diagnosis of hepatocellular carcinoma. Chin Med J (Engl), 2005, 118(15): 1278-1284.
11. Daniele B, Bencivenga A, Megna AS, et al. Alpha-fetoprotein and ultrasonography screening for hepatocellular carcinoma. Gastroenterology, 2004, 127(5 Suppl 1): S108-S112.
12. Trevisani F, D'Intino PE, Morselli-Labate AM, et al. Serum alpha-fetoprotein for diagnosis of hepatocellular carcinoma in patients withchronic liver disease: influence of HBsAg and anti-HCV status. J Hepatol, 2001, 34(4): 570-575.
13. Giardina MG, Matarazzo M, Morante R, et al. Serum alpha-L-fucosidase activity and early detection of hepatocellular carcinoma: a prospective study of patients with cirrhosis. Cancer, 1998, 83(12): 2468-2474.
14. Marrero JA, Su GL, Wei W, et al. Des-gamma carboxyprothrombin can differentiate hepatocellular carcinoma from nonmalignant chronic liver disease in american patients. Hepatology, 2003, 37(5): 1114-1121.
15. Capurro M, Wanless I R, Sherman M, et al. Glypican-3: a novel serum and histochemical marker for hepatocellular carcinoma. Gastroenterology, 2003, 125(1): 89-97.
16. Teramoto R, Minagawa H, Honda M, et al. Protein expression profile characteristic to hepatocellular carcinoma revealed by 2D-DIGE with supervised learning. Biochim Biophys Acta, 2008, 1784(5): 764-772.
1. Penning TM, Drury JE. Human aldo-keto reductases: Function, gene regulation, and single nucleotide polymorphisms. Arch Biochem Biophys, 2007, 464(2):241-250.
2. Hyndman D, Bauman DR, Heredia VV, et al. The aldo-keto reductase superfamily homepage. Chem Biol Interact, 2003, 143 -144: 621-631.
3. Ho HT, Chung SK, Law JW, et al. Aldose reductase-deficient mice develop nephrogenic diabetes insipidus. Mol Cell Biol, 2000, 20(16): 5840-5846.
4. Cisarik-Fredenburg P. Discoveries in research on diabetic keratopathy. Optometry, 2001, 72(11): 691-704.
5. Naya Y, Soh J, Ochiai A, et al. Erythrocyte aldose reductase correlates with erectile dysfunction in diabetic patients. Int J Impot Res, 2002, 14(4): 213-216.
6. Takahashi M, Fujii J, Miyoshi E, et al. Elevation of aldose reductase gene expression in rat primary hepatoma and hepatoma cell lines: implication in detoxification of cytotoxic aldehydes. Int J Cancer, 1995, 62(6): 749-754.
7. Cao D, Fan ST, Chung SS. Identification and characterization of a novelhuman aldose reductase-like gene. J Biol Chem, 1998, 273(19): 11429-11435.
8. Martin HJ, Maser E. Role of human aldo-keto-reductase AKR1B10 in the protection against toxic aldehydes. Chem Biol Interact, 2009, 178(1-3): 145-150.
9. Crosas B, Hyndman D J, Gallego O, et al. Human aldose reductase and human small intestine aldose reductase are efficient retinal reductases: consequences for retinoid metabolism. Biochem J, 2003, 373(Pt 3): 973-979.
10. Zeindl-Eberhart E, Jungblut PR, Otto A, et al. Identification of tumor-associated protein variants during rat hepatocarcinogenesis. Aldose reductase. J Biol Chem, 1994, 269(20): 14589-14594.
11. Zeindl-Eberhart E, Klugbauer S, Dimitrijevic N, et al. Proteome analysis of rat hepatomas: carcinogen-dependent tumor-associated protein variants. Electrophoresis, 2001, 22(14): 3009-3018.
12. Zeindl-Eberhart E, Jungblut PR, Otto A, et al. Further characterization of a rat hepatoma-derived aldose-reductase-like protein--organ distribution and modulation in vitro. Eur J Biochem, 1997, 247(3): 792-800.
13. Woenckhaus M, Klein-Hitpass L, Grepmeier U, et al. Smoking and cancer-related gene expression in bronchial epithelium and non-small-cell lung cancers. J Pathol, 2006, 210(2): 192-204.
14. Fukumoto S, Yamauchi N, Moriguchi H, et al. Overexpression of the aldo-keto reductase family protein AKR1B10 is highly correlated with smokers' non-small cell lung carcinomas. Clin Cancer Res, 2005, 11(5): 1776-1785.
15. Yoshitake H, Takahashi M, Ishikawa H, et al. Aldo-keto reductase family 1, member B10 in uterine carcinomas: a potential risk factor of recurrenceafter surgical therapy in cervical cancer. Int J Gynecol Cancer, 2007, 17(6): 1300-1306.
16. Breton J, Gage MC, Hay AW, et al. Proteomic screening of a cell line model of esophageal carcinogenesis identifies cathepsin D and aldo-keto reductase 1C2 and 1B10 dysregulation in Barrett's esophagus and esophageal adenocarcinoma. J Proteome Res, 2008, 7(5): 1953-1962.
17. Li CP, Goto A, Watanabe A, et al. AKR1B10 in usual interstitial pneumonia: expression in squamous metaplasia in association with smoking and lung cancer. Pathol Res Pract, 2008, 204(5): 295-304.
18. Zeindl-Eberhart E, Haraida S, Liebmann S, et al. Detection and identification of tumor-associated protein variants in human hepatocellular carcinomas. Hepatology, 2004, 39(2): 540-549.
19. Martin HJ, Breyer-Pfaff U, Wsol V, et al. Purification and characterization of akr1b10 from human liver: role in carbonyl reduction of xenobiotics. Drug Metab Dispos, 2006, 34(3): 464-470.
20. Zu X, Yan R, Robbins S, et al. Reduced 293T cell susceptibility to acrolein due to aldose reductase-like-1 protein expression. Toxicol Sci, 2007, 97(2): 562-568.
21. Yang Y, Sharma R, Sharma A, et al. Lipid peroxidation and cell cycle signaling: 4-hydroxynonenal, a key molecule in stress mediated signaling. Acta Biochim Pol, 2003, 50(2):319-336.
22. Awasthi YC, Sharma R, Cheng JZ, et al. Role of 4-hydroxynonenal in stress-mediated apoptosis signaling. Mol Aspects Med, 2003, 24 (4-5): 219-230.
23. Ou H, Haendeler J, Aebly MR, et al. Retinoic acid-induced tissue transglutaminase and apoptosis in vascular smooth muscle cells. Circ Res,2000, 87(10): 881-887.
24. Dawson MI, Zhang XK. Discovery and design of retinoic acid receptor and retinoid X receptor class- and subtype-selective synthetic analogs of all-trans-retinoic acid and 9-cis-retinoic acid. Curr Med Chem, 2002, 9(6): 623-637.
25. Marill J, Idres N, Capron CC, et al. Retinoic acid metabolism and mechanism of action: a review. Curr Drug Metab, 2003, 4(1): 1-10.
26. Gumireddy K, Sutton L N, Phillips PC, et al. All-trans-retinoic acid-induced apoptosis in human medulloblastoma: activation of caspase-3/poly (ADP-ribose) polymerase 1 pathway. Clin Cancer Res, 2003, 9(11): 4052-4059.
27. Brabender J, Metzger R, Salonga D, et al. Comprehensive expression analysis of retinoic acid receptors and retinoid X receptors in non-small cell lung cancer: implications for tumor development and prognosis. Carcinogenesis, 2005, 26(3): 525-530.
28. Ruiz FX, Gallego O, Ardevol A, et al. Aldo-keto reductases from the AKR1B subfamily: retinoid specificity and control of cellular retinoic acid levels. Chem Biol Interact, 2009, 178(1-3): 171-177.
29. Bannasch P. Pathogenesis of hepatocellular carcinoma: sequential cellular, molecular, and metabolic changes. Prog Liver Dis, 1996, 14: 161-197.
30. Stevens MJ, Henry DN, Thomas TP, et al. Aldose reductase gene expression and osmotic dysregulation in cultured human retinal pigment epithelial cells. Am J Physiol, 1993, 265(3P1): E428-E438.
31. Yan R, Zu X, Ma J, et al. Aldo-keto reductase family 1 B10 gene silencing results in growth inhibition of colorectal cancer cells: Implication for cancer intervention. Int J Cancer, 2007, 121 (10): 2301-2306.
32. Ruef J, Liu S Q, Bode C, et al. Involvement of aldose reductase in vascular smooth muscle cell growth and lesion formation after arterial injury. Arterioscler Thromb Vasc Biol, 2000, 20(7): 1745 -1752.
33. Ramana KV, Bhatnagar A, Srivastava S, et al. Mitogenic responses of vascular smooth muscle cells to lipid peroxidation-derived aldehyde 4-hydroxy-trans-2-nonenal (HNE): role of aldose reductase-catalyzed reduction of the HNE-glutathione conjugates in regulating cell growth. J Biol Chem, 2006, 281(26): 17652-17660.
34. Ramana KV, Chandra D, Srivastava S, et al. Aldose reductase mediates the mitogenic signals of cytokines. Chem Biol Interact, 2003, 143-144: 587-596.
35. Swinnen JV, Roskams T, Joniau S, et al. Overexpression of fatty acid synthase is an early and common event in the development of prostate cancer. Int J Cancer, 2002, 98(1): 19-22.
36. Rossi S, Graner E, Febbo P, et al. Fatty acid synthase expression defines distinct molecular signatures in prostate cancer. Mol Cancer Res, 2003, 1(10): 707-715.
37. Yahagi N, Shimano H, Hasegawa K, et al. Co-ordinate activation of lipogenic enzymes in hepatocellular carcinoma. Eur J Cancer, 2005, 41(9): 1316-1322.
38. Rouquette-Jazdanian AK, Pelassy C, Breittmayer JP, et al. Metabolic labelling of membrane microdomains/rafts in Jurkat cells indicates the presence of glycerophospholipids implicated in signal transduction by the CD3 T-cell receptor. Biochem J, 2002, 363(Pt3): 645-655.
39. Swinnen JV, Heemers H, Van De Sande T, et al. Androgens, lipogenesis and prostate cancer. J Steroid Biochem Mol Biol, 2004, 92(4): 273-279.
40. Ma J, Yan R, Zu X, et al. Aldo-keto reductase family 1 B10 affects fatty acidsynthesis by regulating the stability of acetyl-CoA carboxylase-alpha in breast cancer cells. J Biol Chem, 2008, 283(6): 3418-3423.
41. Zang Y, Wang T, Xie W, et al. Regulation of acetyl CoA carboxylase and carnitine palmitoyl transferase-1 in rat adipocytes. Obes Res, 2005, 13(9): 1530-1539.
42. Swinnen JV, Brusselmans K, Verhoeven G. Increased lipogenesis in cancer cells: new players, novel targets. Curr Opin Clin Nutr Metab Care, 2006, 9(4):358-365.
43. Abu-Elheiga L, Matzuk MM, Kordari P, et al. Mutant mice lacking acetyl-CoA carboxylase 1 are embryonically lethal. Proc Natl Acad Sci U S A, 2005, 102(34): 12011-12016.
44. Chajes V, Cambot M, Moreau K, et al. Acetyl-CoA carboxylase alpha is essential to breast cancer cell survival. Cancer Res, 2006, 66(10): 5287-5294.
45. Brusselmans K, De Schrijver E, Verhoeven G, et al. RNA interference-mediated silencing of the acetyl-CoA-carboxylase-alpha gene induces growth inhibition and apoptosis of prostate cancer cells. Cancer Res, 2005, 65(15): 6719-6725.
46. Tong L, Jr Harwood H J. Acetyl-coenzyme A carboxylases: versatile targets for drug discovery. J Cell Biochem, 2006, 99(6): 1476-1488
47. Smith J A, Gaikwad A, Ramondetta LM, et al. Determination of the mechanism of gemcitabine modulation of cisplatin drug resistance in panel of human endometrial cancer cell lines. Gynecol Oncol, 2006, 103(2): 518-522.
48. Kijima H, Ueyama Y. Molecular mechanism of drug resistance in colorectal cancer. Nippon Rinsho, 2003, 61 Suppl 7: 303-309.
49. Sun CA, Wang LY, Chen CJ, et al. Genetic polymorphisms of glutathione S-transferases M1 and T1 associated with susceptibility to aflatoxin-related hepatocarcinogenesis among chronic hepatitis B carriers: a nested case-control study in Taiwan. Carcinogenesis, 2001, 22(8): 1289-1294.
50. Ishimura H. Chemosensitivity test on hepatocellular carcinoma (HCC) and drug resistance. Hokkaido Igaku Zasshi, 1996, 71(6): 689-698.
51. Lee KW, Ko BC, Jiang Z, et al. Overexpression of aldose reductase in liver cancers may contribute to drug resistance. Anticancer Drugs, 2001, 12(2):129-132.
52.王建,万腊香,吴孟津,等.醛糖还原酶相似基因转染细胞对含碳酰基抗癌药物耐药的研究.中华肝脏病杂志, 2002(06): 46-49.
53.杨向东,唐大年,王建,等.转ARL-1基因HepG2细胞耐药相关基因的筛选.癌症, 2003(12): 1289-1295.
54. Penning TM. AKR1B10: a new diagnostic marker of non-small cell lung carcinoma in smokers. Clin Cancer Res, 2005, 11(5): 1687 -1690.
55. Hamada Y, Nakamura J. Clinical potential of aldose reductase inhibitors in diabetic neuropathy. Treat Endocrinol, 2004, 3(4): 245- 255.
56. Gallego O, Ruiz FX, Ardevol A, et al. Structural basis for the high all-trans-retinaldehyde reductase activity of the tumor marker AKR1B10. Proc Natl Acad Sci U S A, 2007, 104(52): 20764-20769.
57. Ko Ec, Wang X, Ferron S. Immunotherapy of malignant diseases. Challenges and strategies. Int Arch Allergy Immunol, 2003, 132(4): 294-309.
58. Weber WA, Grosu AL, Czernin J. Technology Insight: advances in molecular imaging and an appraisal of PET/CT scanning. Nat Clin Pract Oncol, 2008, 5(3): 160-170.
© 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 |