硫氧还蛋白与肿瘤转移

 硫氧还蛋白（Thioredoxin,Trx）是一类广泛存在于生物体内的抗氧化二硫还原蛋白,参与调节细胞内氧化还原平衡。近年研究发现,Trx在多种肿瘤组织中过表达,与肿瘤细胞增殖和凋亡以及细胞周期调控有关。Trx促进缺氧诱导因子（HIF）-1α的合成和稳定,与氧自由基的调控密切相关,参与肿瘤细胞的化疗耐药,在肿瘤转移中发挥重要作用,可能成为抑制肿瘤转移的新靶点。     

肺癌A549细胞抗脱落凋亡相关基因表达谱及其TC-1的表达和意义

目的:应用基因芯片筛选出脱落培养A549细胞抗脱落凋亡相关基因,并探讨TC-1基因的表达和意义.方法:应用基因芯片检测脱落培养A549细胞与贴壁培养A549细胞的差异基因,利用NCBI Pubmed数据库筛选出与肺癌细胞抗脱落凋亡相关的基因,从中找出差异表达倍数最大的基因,运用逆转录聚合酶链反应(RT-PCR)技术及蛋白质印迹技术测定其表达.结果:基因芯片共检测到表达差异的基因745个,筛选出63个与肺癌细胞抗脱落凋亡相关的基因,从中找出差异表达倍数最大的基因--TC-1基因,TC-1基因及其编码蛋白在脱落培养A549细胞中表达明显高于贴壁培养A549细胞,P＜0.02.结论:基因芯片技术为筛选肺癌抗脱落凋亡相关基因提供了有效方法,TC-1基因可能参与调控肺癌细胞抗脱落凋亡过程,与肺癌细胞侵袭、转移等行为相关.     

TC-1与VEGF在非小细胞肺癌肿瘤组织和淋巴结组织中表达及相关性研究

目的：检测TC-1蛋白和VEGF蛋白在NSCLC中的表达,分析二者表达的相关性及其与Wnt/β-Catenin通路的调控的相关性,为进一步研究TC-1基因在NSCLC中的作用及其机制提供依据。方法：采用免疫组化EnVinsion方法检测TC-1蛋白和VEGF蛋白在95例NSCLC中的表达,统计学分析表达差异性和蛋白间的相关性。结果：免疫组化结果显示TC-1阳性表达主要位于胞浆中和部分胞核,VEGF阳性表达主要位于胞浆中。TC-1、VEGF蛋白在鳞癌、腺癌中高表达,与相应正常组织表达相比差异显著（P﹤0.01）。TC-1、VEGF在NSCLC（鳞癌、腺癌）TNM不同分期的表达强度经检验差异性显著（P=0.014;P=0.011）。有淋巴结转移原发灶TC-1、VEGF蛋白的阳性率为90.57%（48/53）、92.45%（49/53）,无淋巴结转移原发灶阳性率为38.09%（16∕42）、28.57%（12/42）,经spearman秩相关检验差异性非常显著（P=0.000;P=0.016）。TC-1、VEGF在NSCLC中的表达率和阳性强度存在相关性（P=0.000）,相关系数rs=0.691。结论：TC-1在非小细胞肺癌（NSCLC）组织中高表达,TC-1表达高低与NSCLC总的分期、淋巴结转移与否相关;TC-1与VEGF蛋白在NSCLC及NSCLC与淋巴结转移原发灶中的表达存在很好的相关性,而VEGF基因是Wnt/β-Catenin通路的靶基因,提示TC-1蛋白可能与Wnt/β-Catenin通路的调控存在相关性。     

小鼠对IFN-α1转染的Friend白血病细胞的局部和全身反应

3C18系Frienol白血病细胞(FLC)对小鼠IFN-α/β具有完全的抵抗性,经IFN-αl基因转染后此抵抗性仍存在,且能分泌高剂量的IFN,接种小鼠后能出现持续数周的干扰素血症,故作者选用此细胞建立模型,观察接种IFN-αl基因转染FLC后小鼠的局部和全身反应,以进一步了解宿主抑制此肿瘤生长的机制。 应用逆转录病毒将IFN-αl基因转入FLC,选出克隆IFN-αlCl-11FLC,可向培养液中分泌IFN-αl256～512U/ml,对照转染的FLC(TC-2)不分泌IFN。具有免疫活性的DBA2 /bg小鼠皮下注射TC-2 FLC后可生成快速生长的肿瘤,接种后20～35天后肿瘤呈现大面积坏死,小鼠存活时间和存活数量较对照组显著增多。IFN-αl C1-11 FLC接种缺失NK细胞活性的bg/bg小鼠后呈现相似的致瘤性下降现象,用单抗去除DBA/2小鼠体内的CD4~ CD8~ 细胞或粒细胞并不影响实验组小鼠的肿瘤生长曲线、肿瘤发生率及存活时间;经一系列处理后获得完全性免疫抑制的SIA裸鼠用于实验,实验组小鼠肿瘤与对照组TC-2肿瘤生长曲线仅稍有差异,并不显著,二者在存活时间、存活数量上并无差异。光镜检查发现在第7、14天,接种TC-2细胞小鼠肿瘤在皮肤表面破溃,在肿瘤边缘仅见少量巨噬细胞和中性粒细胞;在接利IFN-αl C1-11细胞小鼠,肿瘤仍在网状胶原层,有广泛的细胞坏死和凋亡,肿     

肿瘤抑制基因ARHI的生物学功能及研究进展

肿瘤是一种基因病,抑癌基因在抵御肿瘤发生、发展中起着重要作用。ARHI(aplysia ras hom olog I)/NOEY2 是一个新发现的抑癌基因,是 ras/rap超家族成员之一,与 ras家族有50%~60%的同源性,位于人染色体1p31,属小GTP结合蛋白,是该家族第1个被报道的肿瘤抑制基因。ARHI基因编码的蛋白在人类多种组织表达,其中正常卵巢的ARHI表达最高。ARHI是一个印迹基因,印迹机制可能与其CpG岛的差异甲基化有关。ARHI 参与细胞周期调控可能作用于cyclin D1 ,使其不能与 CDK结合形成活性激酶,从而使细胞停止于 G1 期。AR HI可能通过依赖caspase和 calpain两条途径参与信号通路传导诱发细胞凋亡。该基因的异常表达跟多种肿瘤的发生、发展有关。ARHI基因参与了乳腺癌的发生和发展,该基因的表达缺失可能与乳腺癌的转移机制有关。卵巢癌、乳腺癌存在广泛的1p31缺失,其中 ARHI 基因是最常见的一个缺失区域。ARHI基因和蛋白在胰腺癌组织中有较高比例的缺失,提示该基因和蛋白在胰腺癌的发生中起一定作用。ARHI基因在膀胱癌、肝癌、前列腺癌等其他肿瘤中也有不同程度的表达异常。     

肿瘤抑制基因ARHI的生物学功能及研究进展

肿瘤是一种基因病，抑癌基因在抵御肿瘤发生、发展中起着重要作用。ARHI(aplysia ras homolog I)／NOEY2是一个新发现的抑癌基因．是ras/rap超家族成员之一．与ras家族有50％～60％的同源性．位于人染色体1p31，属小GTP结合蛋白，是该家族第1个被报道的肿瘤抑制基因。ARHI基因编码的蛋白在人类多种组织表达，其中正常卵巢的ARHI表达最高。ARHI是一个印迹基因．印迹机制可能与其CpG岛的差异甲基化有关。ARHI参与细胞周期调控可能作用于cyclin D1，使其不能与CDK结合形成活性激酶，从而使细胞停止于G1期。ARHI可能通过依赖caspase和calpain两条途径参与信号通路传导诱发细胞凋亡。该基因的异常表达跟多种肿瘤的发生、发展有关。ARHI基因参与了乳腺癌的发生和发展，该基因的表达缺失可能与乳腺癌的转移机制有关。卵巢癌、乳腺癌存在广泛的1p31缺失，其中ARHI基因是最常见的一个缺失区域。ARHI基因和蛋白在胰腺癌组织中有较高比例的缺失，提示该基因和蛋白在胰腺癌的发生中起一定作用。ARHI基因在膀胱癌、肝癌、前列腺癌等其他肿瘤中也有不同程度的表达异常。     

KiSS-1基因在肿瘤中的研究进展

侵袭和（或）远处转移是影响恶性肿瘤患者预后的关键因素。肿瘤转移涉及肿瘤细胞的侵袭力、黏附力、与间质的相互作用等异常，是多步骤、多因素参与的过程。对肿瘤转移抑制基因的研究已成为肿瘤转移机制研究的热点。Kiss-1是近年克隆的1个新的肿瘤转移抑制基因，研究表明，其表达缺失与人类多种恶性肿瘤转移有关，本文就其在肿瘤中的研究进展作一综述。     

基质金属蛋白酶-24在卵巢浆液性囊腺癌细胞株中的表达

目的探讨基质金属蛋白酶－24(MMP-24)基因在卵巢癌细胞株中的表达及临床意义。方法应用半定量RT-PCR和Western blot对卵巢浆液性囊腺癌细胞株OVCa-R₃、SKOV₃ 、HO-8910及人卵巢细胞株TC-1进行检测。结果3种卵巢癌细胞株中MMP-24基因和蛋白的表达均增强,与正常卵巢细胞相比,差异有统计学意义。结论MMP-24基因可能与卵巢癌的发生、发展有关。     

趋化因子CXCL14与肿瘤

CXCL14是趋化因子CXC家族的一个新成员,最早是从人类乳腺和肾脏组织中分离克隆出来的。CXCL14蛋白能参与调控体内许多生物学过程,如炎症免疫反应,肿瘤相关的血管生成,宿主对肿瘤特异免疫的激活以及肿瘤生长的自分泌调节等。CXCL14基因在正常组织中有表达,而在多种肿瘤细胞系及肿瘤组织中失表达。目前研究认为该基因属于肿瘤抑制基因,肿瘤中CXCL14蛋白的表达缺失与肿瘤的发生发展有关。本文综述有关CXCL14基因与肿瘤的研究进展。     

结直肠肿瘤的分子生物学及其临床意义

越来越多的证据表明结直肠癌的发生与环境和遗传因素有关。癌症发生的最关键的遗传因素是参与细胞调控、增生和分化的癌基因、肿瘤抑制基因和看家基因,了解这些基因的改变对于结直肠肿瘤的诊断和处理有重要的意义。     

Transforming properties of TC-1 in human breast cancer: interaction with FGFR2 and beta-catenin signaling pathways

Breast cancer development is associated with gene amplification and over expression that is believed to have a causative role in oncogenesis. Previous studies have demonstrated that over expression of TC-1(C8orf4) mRNA occurs in approximately 50% of breast cancer cell lines and primary tumor specimens. Here, we show that TC-1 has transforming properties in human mammary epithelial (HME) cells and its expression is mechanistically linked to FGFR signaling cascades. In vitro experiments demonstrate that TC-1 over expression mediates both anchorage-independent and growth factor-independent proliferation of HME cells. TC-1 was down regulated by the FGFR inhibitor PD173074 in the breast cancer cell line SUM-52 that also has an FGFR2 gene amplification and over expression. Furthermore, forced expression of FGFR2 in HME cells increased the level of expression of endogenous TC-1 mRNA. TC-1 has been implicated as a modulator of Wnt/beta-catenin signaling in 293 cells and in gastric cancer cells. However, while we did find increased expression of a subset of beta-catenin target genes in TC-1 over expressing cells, we did not find an association of TC-1 with global expression of beta-catenin target genes in our cells. Taken together, our data suggest that TC-1 over expression is transforming and may link with the FGFR pathway in a subset of breast cancer.     

TC-1 is a novel tumorigenic and natively disordered protein associated with thyroid cancer

A novel gene, thyroid cancer 1 (TC-1), was found recently to be overexpressed in thyroid cancer. TC-1 shows no homology to any of the known thyroid cancer-associated genes. We have produced stable transformants of normal thyroid cells that express the TC-1 gene, and these cells show increased proliferation rates and anchorage-independent growth in soft agar. Apoptosis rates also are decreased in the transformed cells. We also have expressed recombinant TC-1 protein and have undertaken a structural and functional characterization of the protein. The protein is monomeric and predominantly unstructured under conditions of physiologic salt and pH. This places it in the category of natively disordered proteins, a rapidly expanding group of proteins, many members of which play critical roles in cell regulation processes. We show that the protein can be phosphorylated by cyclic AMP-dependent protein kinase and protein kinase C, and the activity of both of these kinases is up-regulated when cells are stably transfected with TC-1. These results suggest that overexpression of TC-1 may be important in thyroid carcinogenesis.     

TC-1在非小细胞肺癌的表达及与淋巴结转移的关系

目的:探讨TC-1蛋白在非小细胞肺癌(NSCLC)组织中的表达及其在病理分类、分级、TNM分期中的差异性,为进一步研究TC-1在NSCLC中的作用及其机制提供依据。方法:采用免疫组化EnVinsion法检测TC-1蛋白在95例非小细胞肺癌中的表达,统计学分析其在病理分类、分级及临床TNM分期及淋巴结转移与否的差异性。结果:免疫组化结果显示TC-1阳性表达主要位于胞浆中和部分胞核。肺鳞癌和腺癌总阳性率分别为63.46%、72.09%,差异性不显著(P=0.154)。鳞癌高、中、低分级阳性率分别为20.00%、65.51%、72.22%,经Kruskal-Wallis H检验无明显差异性(P=0.075)。腺癌的高、中、低分级的阳性率分别是33.33%、65.21%、88.23%,差异性非常显著(P=0.005)。NSCLC(鳞癌、腺癌)的TNM分期Ⅰa、Ⅰb、Ⅱa、Ⅱb、Ⅲa、Ⅲb的表达强度经检验差异性显著(P=0.029)。有淋巴结转移者原发灶阳性率为90.57%(48/53)、无淋巴结转移者原发灶阳性率为38.09%(16/42),经Mann-Whitney U秩和检验差异性非常显著(P=0.000)。53例伴淋巴结转移的NSCLC原发灶与淋巴结转移灶阳性表达相关性用Spearman检验,有相关性(r=0.39,P=0.000)。结论:TC-1在非小细胞肺癌(NSCLC)组织中高表达,TC-1表达与病理分型、鳞癌分级无关,与腺癌分级、NSCLC TNM分期、淋巴结转移与否相关。提示TC-1在肺癌组织中表达高低与淋巴结转移密切相关。     

TC-1在非小细胞肺癌的表达及与淋巴结转移的关系

目的：探讨TC-1蛋白在非小细胞肺癌（NSCLC）组织中的表达及其在病理分类、分级、TNM分期中的差异性,为进一步研究TC-1在NSCLC中的作用及其机制提供依据。方法：采用免疫组化EnVinsion法检测TC-1蛋白在95例非小细胞肺癌中的表达,统计学分析其在病理分类、分级及临床TNM分期及淋巴结转移与否的差异性。结果：免疫组化结果显示TC-1阳性表达主要位于胞浆中和部分胞核。肺鳞癌和腺癌总阳性率分别为63.46%、72.09%,差异性不显著（P=0.154）。鳞癌高、中、低分级阳性率分别为20.00%、65.51%、72.22%,经Kruskal-Wallis H检验无明显差异性（P=0.075）。腺癌的高、中、低分级的阳性率分别是33.33%、65.21%、88.23%,差异性非常显著（P=0.005）。NSCLC（鳞癌、腺癌）的TNM分期Ⅰa、Ⅰb、Ⅱa、Ⅱb、Ⅲa、Ⅲb的表达强度经检验差异性显著（P=0.029）。有淋巴结转移者原发灶阳性率为90.57%（48/53）、无淋巴结转移者原发灶阳性率为38.09%（16/42）,经Mann-Whitney U秩和检验差异性非常显著（P=0.000）。53例伴淋巴结转移的NSCLC原发灶与淋巴结转移灶阳性表达相关性用Spearman检验,有相关性（r=0.39,P=0.000）。结论：TC-1在非小细胞肺癌（NSCLC）组织中高表达,TC-1表达与病理分型、鳞癌分级无关,与腺癌分级、NSCLC TNM分期、淋巴结转移与否相关。提示TC-1在肺癌组织中表达高低与淋巴结转移密切相关。     

Vaccination of full-length HPV16 E6 or E7 protein inhibits the growth of HPV16 associated tumors

Cervical cancer is the second most common cancer in women worldwide. Human papillomavirus (HPV) is the primary etiologic agent of cervical cancer. Two HPV16 proteins, E6 and E7, are consistently expressed in tumor cells. Most therapeutic vaccines target one or both of these proteins. Taking the advantages of safety and no human leukocyte antigen restriction, protein vaccine has become the most popular form of HPV therapeutic vaccines. Here we demonstrate that immunization with full-length HPV16 E6 or E7 protein elicited specific immunological effect and inhibition of TC-1 cell growth using TC-1 mouse model. HPV16 E6 and E7 genes were cloned into pET-28a(+) and introduced into E. coli Rosetta. Expression of the genes was induced by IPTG. Proteins were purified by Ni-NTA agarose and they were detected by SDS-PAGE and Western blotting. C57BL/6 mice were vaccinated with 1.5 nmol HPV16 E6 or E7 protein. Then they were implanted with 1x10(5) TC-1 cells. No tumor was detected in any mouse vaccinated with E7 protein. Forty days later, the tumor-free mice and control mice were challenged with 2x10(5) TC-1 cells. All control mice developed tumors 6 days later, but E7 immunized mice were tumor free until 90 days. Tumor growth was slow in the E6 immunized mice, but 83% of the mice developed tumors and the survival percentage was not significantly different from the control. An adoptive immune model was used to demonstrate the therapeutic effect. Results showed that the development of TC-1 cells was obviously reduced by transfusion of T-cells but not serum from mice immunized with E7 protein. T-cells from E7 immunized mice also induced the lysis of TC-1 cells in the cytotoxic T lymphocyte assay. These findings show that immunization with HPV16 E6 or E7 protein was able to elicit specific protective immunity against TC-1 tumor growth.     

Selenium enhances the efficacy of Radachlorin mediated-photodynamic therapy in TC-1 tumor development

Selenium, an essential trace element possessing anti-carcinogenic properties, can induce apoptosis in cancer cells. Our goal was to investigate the enhanced antitumor effects of photodynamic therapy (PDT) plus selenium in TC-1 tumor cells and implanted mice. Cell viability was evaluated at various time intervals after PDT treatment and/or selenium by methyl thiazolyl tetrazolium (MTT) assay. When only PDT treatment was administered to TC-1 tumor cells, TC-1 cell growth recovered over time. On the other hand, co-treatment of PDT and selenium extended the inhibition time of tumor cell growth. Co-treatment of PDT and selenium showed serious morphological changes in TC-1 cells and induced a more apoptotic population by FACS analysis. By signal transduction pathway SuperArray analysis, genes closely involved in the NFκB, p53 and phopholipase C pathways, such as VCAM1, MDM2 and FOS, were significantly downregulated at least 10-fold in TC-1 cells following PDT and selenium cotreatment. In an in vivo study, tumor-bearing mice were intravenously injected with Radachlorin 3 h before irradiation with 300 J/cm2 of light. Selenium was administered daily for 20 days. Combination therapy against the mouse tumors generated by TC-1 cells was more effective than PDT or selenium alone. These data suggest that selenium plus PDT can induce a significant tumor suppression response compared with PDT alone. Additionally, it can be an effective anticancer therapy strategy.     

Combination of tumor site-located CTL-associated antigen-4 blockade and systemic regulatory T-cell depletion induces tumor-destructive immune responses

Accumulating data indicate that tumor-infiltrating regulatory T cells (Treg) are present in human tumors and locally suppress antitumor immune cells. In this study, we found an increased Treg/CD8 ratio in human breast and cervical cancers. A similar intratumoral lymphocyte pattern was observed in a mouse model for cervical cancer (TC-1 cells). In this model, systemic Treg depletion was inefficient in controlling tumor growth. Furthermore, systemic CTL-associated antigen-4 (CTLA-4) blockade, an approach that can induce tumor immunity in other tumor models, did not result in TC-1 tumor regression but led to spontaneous development of autoimmune hepatitis. We hypothesized that continuous expression of an anti-CTLA-4 antibody localized to the tumor site could overcome Treg-mediated immunosuppression and locally activate tumor-reactive CD8+ cells, without induction of autoimmunity. To test this hypothesis, we created TC-1 cells that secrete a functional anti-CTLA-4 antibody (TC-1/alphaCTLA-4-gamma1 cells). When injected into immunocompetent mice, the growth of TC-1/alphaCTLA-4-gamma1 tumors was delayed compared with control TC-1 cells and accompanied by a reversion of the intratumoral Treg/CD8 ratio due to an increase in tumor-infiltrating IFNgamma-producing CD8+ cells. When local anti-CTLA-4 antibody production was combined with Treg inhibition, permanent TC-1 tumor regression and immunity was induced. Importantly, no signs of autoimmunity were detected in mice that received local CTLA-4 blockade alone or in combination with Treg depletion.     

FAP-1在卵巢癌细胞株中的表达与意义

目的 :探讨Fas相关磷酸酯酶 -1(FAP 1)在卵巢癌细胞株SKOV3、HO 8910、TC 1和 3AO中的表达与意义。方法 :采用免疫印迹法和逆转录聚合酶链式反应方法 ,检测了 4种卵巢癌细胞株中FAP 1蛋白与核酸的表达水平 ;应用四唑盐比色法和流式细胞术检测了经抗Fas激活性抗体DX2诱导凋亡后 ,FAP 1分子与 4种细胞对凋亡敏感性的关系。结果 :FAP 1蛋白与mRNA的表达结果一致 ,均在SKOV3和HO 8910细胞中有表达 ,以SKOV3表达最强 ,而在TC 1和 3AO中未见表达。当用DX2以不同时间诱导凋亡后 ,FAP 1阴性株的细胞抑制率与凋亡率均高于FAP 1阳性株 ,P<0 0 5 ,P <0 0 1;而FAP 1阳性株中 ,HO 8910的生长抑制率又高于SKOV3 ,P <0 0 5。同时 ,FAP 1阴性株对DX2的反应具有时间依赖性 ,而FAP 1阳性株却表现出对DX2的逐渐耐受性。结论 :FAP 1对细胞凋亡的敏感性具有负性调控作用 ,可能对卵巢癌的发病和耐药具有重要意义     

HPV16Z-Hsp65-E6／E7无佐剂重组蛋白疫苗的研究

目的：研究治疗性HPV16Z-Hsp65-E6／E7无佐剂重组蛋白疫苗的抗肿瘤活性。方法：通过淋巴细胞增殖实验和细胞毒性杀伤实验研究该疫苗激发的细胞免疫反应及反应强度；观察该疫苗对小鼠TC-1肿瘤细胞移植瘤的体内治疗作用和对小鼠生存期的影响。结果：重组蛋白疫苗免疫小鼠后，小鼠脾淋巴细胞与该疫苗体外混合培养增殖明显，并可特异性地在体外杀伤TC-1细胞；体内抑瘤试验显示该疫苗对HPV16病毒转化的TC-1细胞小鼠移植瘤的生长有显著的抑制作用。结论：该疫苗能激发特异性细胞免疫反应；显著抑制HPV16转化的TC-1肿瘤细胞生长。     

Immunotherapeutic Effects of Dendritic Cells Pulsed with a Coden-optimized HPV 16 E6 and E7 Fusion Gene in Vivo and in Vitro

Background: : Cervical cancer is the second most common cause of cancer related death of women. PersistentHPV infection, especially with high-risk types such as HPV16 and HPV18, has been identified to be the primarycause of cervical cancer. E6 and E7 are the major oncoproteins of high-risk HPVs, which are expressed exclusivelyin HPV infected tissues, and thereby represent ideal therapeutic targets for immunotherapy of cervical cancer.Materials and Methods: In this work, we used recombinant adenovirus expressing coden-optimized HPV16 E6and E7 fusion protein (Ad-ofE6E7) to prime dendritic cells (DC-ofE6E7), to investigate the ability of primed DCvaccine in eliciting antitumor immunity in vitro and vivo. Results: Our results indicated that DC-ofE6E7 vaccineco-culturing with splenocytes could strongly induce a tumor-specific cytotoxic T lymphocyte (CTL) responseand kill the TC-1 cells effectively in vitro. Moreover, DC-ofE6E7 vaccine induced protective immunity againstthe challenge of TC-1 cancer cells in vivo. Conclusions: The results suggested that the HPV16 ofE6E7 primedDC vaccine has potential application for cervical cancer immunotherapy.