端粒酶逆转录酶、P53、PCNA在非小细胞肺癌的表达及意义

目的　探讨端粒酶逆转录酶、P5 3、PCNA在非小细胞肺癌组织及癌旁正常组织中的表达及其关系。方法　取自手术切除及肺穿刺经病理证实为肺癌组织标本 6 0例 ,采用免疫组化法检测人端粒酶逆录酶、P5 3、PCNA的表达 ,并取癌旁肺组织 10例作对照。结果　肺癌组织中端粒酶逆转录酶、P5 3、阳性率分别为78 3% ( 4 7/6 0 )和 5 3 3% ( 32 /6 0 ) ,PCNA的指数为 5 8 6 3± 10 6 1。癌旁组织依次为 0 % ( 0 /10 ) ,10 % ( 1/10 )和30 3± 7 3,(P <0 0 1) ;在鳞癌中的表达依次为 72 7% ,5 1 5 % ,5 7 78± 10 6 8;在腺癌中的表达依次为88 2 % ,5 2 9% ,5 9 18± 11 4 7;在腺鳞癌中依次为 80 % ,6 0 % ,6 1 2± 10 6 7,(P >0 0 5 )。临床分期的关系为 :Ⅰ期依次为 71% ,35 5 % ,5 5 74± 10 71;Ⅱ期为 87 5 % ,70 8% ,5 9 79± 9 14 ;Ⅲ期为 80 4 % ,80 1% ,71 6±7 31 (P <0 0 1)。在淋巴结转移阳性病例中表达为 89 7% ,6 0 9% ,6 1 72± 9 74 ;在淋巴结转移阴性病例中为 6 7 7% ,38 7% ,5 5 74± 10 71,(P =0 0 2 8) ;病理分化较高组织中的表达为 70 4 % ,2 9 6 % ,5 5 6 3± 9 72 ,分化中等的为 84 2 % ,6 8 4 % ,5 8 2 6± 10 2 6 ,分化较低的组织中为 85 7% ,78 6 % ,6 4 93± 10 7     

凋亡和p53突变对非小细胞肺癌预后的影响

目的:研究细胞凋亡及凋亡相关蛋白p53在肺癌组织中的表达水平,及其对预后的影响。方法:应用DNA缺口末端标记技术和免疫组化方法检测111例肺癌患者组织中细胞凋亡和突变型p53蛋白表达水平。结果:111例肺癌组织中,细胞凋亡数量≥1%的病例有47·7%(53/111),突变型p53蛋白阳性表达率为40·5%(45/111);单因素分析肺癌患者预后的影响因素有凋亡(χ2=6·77,P<0·01)、突变型p53(χ2=6·56,P<0·05)、淋巴结转移(χ2=16·31,P<0·01)和TNM分期(χ2=15·86,P<0·01);COX模型多因素分析显示,肿瘤大小(χ2=4·12,P<0·05)、淋巴结转移(χ2=12·94,P<0·01)和细胞凋亡(χ2=4·61,P<0·05)是肺癌患者的预后不良因素。结论:凋亡及凋亡相关蛋白p53影响肺癌患者的生物学行为及预后。     

p53和PCNA与非小细胞肺癌

p53基因和增殖细胞核抗原(PCNA)均与非小细胞肺癌(NSCIE)的放疗有密切关系.两者同时高表达的NSCLC患者的生存期低于同时阴性者.研究p53基因和PCNA在NSCLC中的表达,及其与NSCLC放射敏感性和预后的关系,可以对临床制定个体化治疗方案和提高疗效起到积极作用.     

p53、Cyclin D1及PCNA在非小细胞肺癌的表达

目的 研究非小细胞肺癌（NSCLC）中p53、Cyclin D1蛋白表达及与细胞增殖的关系。方法 应用免疫组织化学技术（SP法）检测74例NSCLC中p53、Cyclin D1蛋白和增殖细胞抗原（PCNA）的表达情况。结果 p53蛋白、CyclinD1蛋白在NSCLC中阳性表达率分别为55．41％和37．84％,均明显高于正常肺组织（P1＝0．0031,P2＝0．0429）。 p53蛋白表达与淋巴结转移有密切关系（P＝0．0222）。p53蛋白、CyclinD1蛋白表达分别与PCNA指数有密切关系（P1＝0．001,P2＝0．0009）。p53蛋白与CyclinD1蛋白表达之间未见明显关系（P＞0．05）。结论 p53蛋白和CyclinD1蛋白近表达参与了NSCLC的发生发展,有促进细胞增殖的作用。p53的诊断和评价NSCLC预后的重要参数。     

非小细胞肺癌组织Survivin和p53蛋白表达临床意义的探讨

目的:探讨Survivin和p53蛋白在非小细胞肺癌(NSCLC)组织中的表达及其临床意义.方法:采用免疫组化SP法检测60例NSCLC和10例肺良性病变组织中Survivin 和p53蛋白的表达.结果:NSCLC组织中Survivin蛋白阳性表达率为61.6%,明显高于正常对照组的0,x2=13.08,P=0.000...     

p53蛋白在非小细胞肺癌中的表达及意义

目的 :探讨非小细胞肺癌中p5 3蛋白的表达及意义。方法 :应用SP免疫组化法检测 5 2例肺癌组织中p5 3蛋白的表达。结果 :非小细胞肺癌中p5 3蛋白阳性表达率为 78 8% ,其中鳞癌与腺癌分别为 75 0 %和 82 1% ,p5 3蛋白阳性病人的预后较阴性者差。结论 :p5 3蛋白在非小细胞肺癌中有较高的表达 ,有可能作为评估预后、指导治疗的指标。     

晚期非小细胞肺癌对铂类药物的化疗敏感性与p53和p73基因多态的关系

目的探讨细胞凋亡相关基因p53和p73的遗传多态,与晚期非小细胞肺癌（NSCLC）对铂类药物化疗敏感性的关系。方法以聚合酶链反应（PCR）-限制性片段长度多态性（RFLP）和突变扩增阻抑系统（ARMS）分析方法,对165例以顺铂（DDP）或卡铂（CBP）为主进行化疗的晚期NSCLC患者,进行p53第72密码子Arg→Pro多态,和p73第2外显子G4C14-A4T14多态的检测,2～3个疗程后进行效果评价。以非条件Logistic回归模型比较不同基因型与化疗疗效的关系。结果携带p53 72Pro等位基因患者的化疗敏感性,是携带p53 72Arg／Arg基因型患者的2、46倍（95％CI,1．11～5．45,P=0．026）;而携带至少1个p73突变等位基因（A4T14）的患者,其化疗敏感性是携带p73 G4C14／G4C14基因型患者的2．22倍（95％讲,1．14～4、30,P=0．019）。2个多态位点合并分析结果显示,同时携带p53和p73野生基因型的患者,化疗有效率为7．7％;而携带1个、2个和≥3个p53和p73变异等位基因的患者,化疗有效率分别为34．8％、42．2％和40、7％。结论p53和p73基因遗传多态与晚期NSCLC患者对以铂类药物为主的化疗敏感性有关。     

非小细胞肺癌循环肿瘤细胞检测进展

肺癌是世界范围内发病率最高的恶性肿瘤, 约有18%的癌症相关死亡与肺癌有关。循环肿瘤细胞(CTCs)的出现可看做是肿瘤细胞播散入血的直接体现, 而这正是导致非小细胞肺癌患者预后较差的最重要因素之一。检测外周血中循环肿瘤细胞有望成为诊断肺癌的辅助方法, 使肺癌患者得到最佳的个体化治疗。研究者们已尝试多种不同的方法从外周血中分离CTCs。虽然有关CTCs检测的研究众多, 但不同检测方法的敏感度、特异性及重复性限制了其临床意义, 本文就现有的各项技术进行综述。      

p53 regulates FAK expression in human tumor cells

Attenuation of the p53 protein is one of the most common abnormalities in human tumors. Another important marker of tumorigenesis is focal adhesion kinase (FAK), a 125-kDa tyrosine kinase that is overexpressed at the mRNA and protein levels in a variety of human tumors. FAK is a critical regulator of adhesion, motility, metastasis, and survival signaling. We have characterized the FAK promoter and demonstrated that p53 can inhibit the FAK promoter activity in vitro. In the present study, we showed that p53 can bind the FAK promoter-chromatin region in vivo by chromatin immunoprecipitation (ChIP) assay. Furthermore, we demonstrated down-regulation of FAK mRNA and protein levels by adenoviral overexpression of p53. We introduced plasmids with different mutations in the DNA-binding domain of p53 (R175H, p53 R248W and R273H) into HCTp53(-/-) cells and showed that these mutations of p53 did not bind FAK promoter and did not inhibit FAK promoter activity, unlike wild type p53. We analyzed primary breast and colon cancers for p53 mutations and FAK expression, and showed that FAK expression was increased in tumors containing mutations of p53 compared to tumors with wild type p53. In addition, tumor-derived missense mutations in the DNA-binding domain (R282, R249, and V173) also led to increased FAK promoter activity. Thus, the present data show that p53 can regulate FAK expression during tumorigenesis.     

HPV感染与p53蛋白及P-糖蛋白表达在非小细胞肺癌中的关系及其临床意义

目的:探讨人乳头瘤病毒(humanpapillomavirus,HPV)感染在非小细胞肺癌(nonsmallcelllungcancer,NSCLC)发生中的病因学意义及其与p53蛋白和P糖蛋白(Pgp)表达之间的相关性。方法:应用PCR、免疫组化方法分别检测76例NSCLC组织中HPVDNA及其E6、E7原癌蛋白、p53蛋白和Pgp的表达情况。结果:NSCLC中HPVDNA及其E6、E7原癌蛋白的检出率分别为40.8%(31/76)、43.4%(33/76),2种检测方法的符合率为78.9%(60/76);p53蛋白和Pgp的阳性率分别为63.2%(48/76)、59.2%(45/76),且HPV感染阳性组中p53蛋白的表达率为80.6%(25/31),显著高于阴性组51.1%(23/45)(P<0.05);p53蛋白表达阳性组中Pgp的表达率为68.8%(33/48),显著高于阴性组42.9%(12/28)(P<0.05);而HPV感染阳性组与阴性组间Pgp的表达无显著性差异(P>0.05)。HPV感染与高、中分化程度的NSCLC及吸烟有关。结论:HPV感染可能是NSCLC发生的另一重要病因学因素,且HPV感染可能导致p53基因突变,后者可能促进肺癌耐药性的增加。     

紫杉醇对不同p53基因型肺癌细胞作用的研究

目的:观察紫杉醇对3种不同p53基因型肺癌细胞的作用。方法:以不同浓度紫杉醇(0.1、1.0、10、100、1 000nmol/L)分别作用肺癌细胞A549、H322、H1299不同时间(4、12、24、48、96 h),实验同时设阴性对照组(不加紫杉醇)和空白对照组(只加细胞培养液)。采用MTT、流式细胞术、Western blotting检测细胞的生长及细胞周期、凋亡、乙酰化微管蛋白、p53蛋白等指标变化。结果:紫杉醇对3株肺癌细胞的生长抑制作用均随着作用时间的延长而增强(P0.05);不同浓度紫杉醇对3株细胞生长的影响也存在差异(P0.05)。3株细胞中A549细胞半数抑制浓度相对较低,但3株细胞间对紫杉醇敏感性的差异无统计学意义(P0.05)。紫杉醇10 nmol/L作用时,3株细胞G2/M期变化趋势基本一致,1 000 nmol/L作用时,A549细胞G2/M期比例与作用时间呈正相关(P0.05),H322和H1299的G2/M细胞于24 h达高峰,作用后期H1299出现异常多倍体;同浓度紫杉醇作用的各株肺癌细胞的凋亡呈时间依赖性,但1 000 nmol/L组诱导的细胞凋亡比10 nmol/L组更晚出现。浓度依赖性实验中,10 nmol/L诱导凋亡比例最高;紫杉醇浓度≤100 nmol/L时,G2/M期比例随浓度增加而增高(P0.05);紫杉醇浓度≥100 nmol/L时,各肺癌细胞株G2/M期比例间的差异无统计学意义。凋亡和G2/M期阻滞无相关性(P0.05)。紫杉醇作用后,A549细胞p53蛋白呈浓度和时间依赖性增加,H322细胞p53蛋白无明显变化。3株细胞乙酰化微管蛋白均较对照组显著上升(P0.05)。结论:紫杉醇对不同p53基因型肺癌细胞生长的抑制作用呈时间依赖性。紫杉醇可增加野生型p53蛋白表达,诱导p53依赖和非p53依赖的两种凋亡途径,p53基因型影响了高浓度紫杉醇对肺癌细胞的周期阻滞作用,但对紫杉醇化疗敏感性无显著影响。     

Dominant negative knockout of p53 abolishes ErbB2-dependent apoptosis and permits growth acceleration in human breast cancer cells

We previously reported that the ErbB2 oncoprotein prolongs and amplifies growth factor signalling by impairing ligand-dependent downregulation of hetero-oligomerised epidermal growth factor receptors. Here we show that treatment of A431 cells with different epidermal growth factor receptor ligands can cause growth inhibition to an extent paralleling ErbB2 tyrosine phosphorylation. To determine whether such growth inhibition signifies an interaction between the cell cycle machinery and ErbB2-dependent alterations of cell signalling kinetics, we used MCF7 breast cancer cells (which express wild-type p53) to create transient and stable ErbB2 transfectants (MCF7-B2). Compared with parental cells, MCF7-B2 cells are characterised by upregulation of p53, p21(WAF) and Myc, downregulation of Bcl2, and apoptosis. In contrast, MCF7-B2 cells co-transfected with dominant negative p53 (MCF7-B2/Delta p53) exhibit reduced apoptosis and enhanced growth relative to both parental MCF7-B2 and control cells. These data imply that wild-type p53 limits survival of ErbB2-overexpressing breast cancer cells, and suggest that signals of varying length and/or intensity may evoke different cell outcomes depending upon the integrity of cell cycle control genes. We submit that acquisition of cell cycle control defects may play a permissive role in ErbB2 upregulation, and that the ErbB2 overexpression phenotype may in turn select for the survival of cells with p53 mutations or other tumour suppressor gene defects.     

p53抑制干细胞的自我更新能力而增敏奥沙利铂诱导肺癌细胞凋亡的作用及机制

目的:研究p53通过增加肿瘤干细胞不等向分裂的比例而增敏奥沙利铂(Oxaliplatin,L-OHP)诱导肺癌细胞凋亡的作用及机制,探索影响肺癌化疗效果的因素.方法:用Nutlin-3(N-3)调控肺癌细胞野生型p53的表达,研究p53对L-OHP诱导细胞凋亡及对肺癌干细胞不同分裂模式的影响;使用Akt激活剂及shRNA-GSK3β调控表达的肺癌细胞系,研究p53通过Akt调控奥沙利铂诱导的肺癌细胞凋亡,并抑制肺癌干细胞自我更新能力.在肺癌标本中检测GSK3β与磷酸化Akt1的表达及与预后的关系.结果:Nutlin-3上调了A549及H460细胞中野生型p53的表达,有效抑制了细胞增殖,增强L-OHP诱导细胞凋亡效果.Nutlin-3抑制磷酸化Akt1,促进肺癌干细胞不等向分裂,进而抑制肿瘤干细胞的比例,增敏L-OHP诱导肺癌细胞凋亡.Akt1磷酸化活性在p53介导的下游GSK3β表达增加的调控中起关键作用.50nmol/L的L-OHP通过抑制Akt1活性促进凋亡,10μmol/L的Nutlin-3对L-OHP诱导的肺癌细胞凋亡有协同作用,是通过共同调控Akt1/GSK3β通路的活性实现,Akt1的激活与GSK3β的抑制均显著地抵消了p53对L-OHP诱导的肺癌干细胞比例降低.结论:p53通过抑制磷酸化Akt1的水平调控GSK3β表达,促肺癌干细胞不均等分裂,增敏L-OHP诱导凋亡.肺癌中GSK3β的高表达与磷酸化Akt1的低表达预示较好的临床治疗效果.     

p53基因突变与人非小细胞肺癌临床病理生理分析

目的 探讨ｐ５３基因点突变与人非小细胞肺癌临床生理特征的联系。方法 应用聚合酶反应－单链构象多态性分析方法（ＰＣＲ－ＳＳＣＰ０检测原发性非小细胞肺癌癌组织的ｐ５３基因第５～８外显子点突变。结果 ４０例肺癌组织中１９例（４７．５％）有点突变发生,８例良性肿瘤组织均无ｐ５３基因点突变发生,点突变发生与病理分期和淋巴结转移有明显关系（Ｐ〈０．０２５）,结论ｐ基因估变在非小细胞肺癌的发生和进展中可能起重要作用。     

Involvement of p53 in oroxylin A‐induced apoptosis in cancer cells

Oroxylin A, a naturally occurring monoflavonoid extracted from Scutellariae radix, exhibits anticancer activity and induces apoptosis in human hepatocellular carcinoma HepG2 cells according to our previous data. In this study, we investigate whether p53 is involved in oroxylin A‐triggered viability inhibition and apoptosis induction in cancer cells. In a panel of different cancer cell lines, more potent inhibitory effects of oroxylin A were observed in wtp53 cells than those in mtp53 or p53‐null cells. Moreover, p53‐siRNA‐transfected HepG2 cells showed lower levels of apoptosis induced by oroxylin A than control‐siRNA‐transfected cells. Likewise, after oroxylin A treatment, p53‐null K‐562 cells displayed promoted apoptosis rate when transfected with wtp53 plasmid. Western blot and real‐time RT‐PCR assay revealed that oroxylin A markedly upregulated p53 protein expression in HepG2 and p53‐overexpressing K‐562 cells, but had no influence on p53 mRNA synthesis. Furthermore, after co‐treatment with cycloheximide, oroxylin A still exerted a little effect on p53 expression. The negative regulator of p53, MDM2 protein was detected, and downregulated expression was observed. In the presence of MG132, an inhibitor of proteasome‐mediated proteolysis, no change in p53 expression was obtained. Additionally, the antioxidant N‐acetyl‐L‐cysteine could obviously abrogate p53 stabilization triggered by oroxylin A. Therefore, it is summarized that oroxylin A stabilized p53 expression and induced apoptosis at the posttranslational level via downregulating MDM2 expression and interfering MDM2‐modulated proteasome‐related p53 degradation. This indicated that oroxylin A could be served as a potential, novel agent candidate for cancer therapy. © 2009 Wiley‐Liss, Inc.     

环氧化酶-2及p53在非小细胞肺癌中的表达与临床意义

研究表明肺癌发生、发展是多种癌基因和抑癌基因相互作用的结果。目前认为环氧合酶- 2(COX-2)参与肺癌的发生、发展、转移等多方面。p53基因是最重要的抑癌基因之一,p53缺失突变与肺癌的发生、发展有关。p53突变与COX-2在非小细胞肺癌(NSCLC)的发生、发展中可能具有协同作用。     

Akt promotes cisplatin resistance in human ovarian cancer cells through inhibition of p53 phosphorylation and nuclear function

Resistance to cisplatin-based chemotherapy is a major cause of treatment failure in human ovarian cancer. Wild-type TP53 status is often, but not always, associated with cisplatin sensitivity, suggesting that additional factors may be involved. Overexpression/activation of the phosphatidylinositol-3-kinase/Akt pathway is commonly observed in ovarian cancer, and Akt activation is a determinant of chemoresistance in ovarian cancer cells, an effect that may be due, in part, to its inhibitory actions on p53-dependent apoptosis. To that end, we examined the role and regulation of p53 in chemosensitive ovarian cancer cells, as well as in their chemoresistant counterparts, and investigated if and how Akt influences this pathway. Cisplatin induced apoptosis in chemosensitive, but not chemoresistant cells, and this was inhibited by downregulation of p53. Cisplatin upregulated PUMA in a p53-dependent manner, and the presence of PUMA was necessary, but not sufficient for cisplatin-induced apoptosis. p53 was phosphorylated on numerous N-terminal residues, including Ser15, Ser20, in response to cisplatin in chemosensitive, but not chemoresistant cells. Furthermore, activation of Akt inhibited the cisplatin-induced upregulation of PUMA, and suppressed cisplatin-induced p53 phosphorylation, while inhibition of Akt increased total and phospho-p53 contents and sensitized p53 wild-type, chemoresistant cells to cisplatin-induced apoptosis. Finally, mutation of Ser15 and/or Ser20, but not of Ser37, to alanine significantly attenuated the ability of p53 to facilitate CDDP-induced apoptosis, and this was independent of PUMA expression. These results support the hypothesis that p53 is a determinant of CDDP sensitivity, and suggest that Akt contributes to chemoresistance, in part, by attenuating p53-mediated PUMA upregulation and phosphorylation of p53, which are essential, but independent determinants of sensitivity to CDDP-induced apoptosis.     

miR-223-3p通过调控ECT2诱导非小细胞肺癌细胞凋亡的实验研究

目的：探讨miR-223-3p对非小细胞肺癌细胞增殖与凋亡的影响及其可能的作用机制。方法：选取24例非小细胞肺癌患者的癌及癌旁组织,实时定量PCR检测miR-223-3p的相对表达水平,免疫组化检测分析上皮细胞转化序列2（ECT2）蛋白的表达水平,并对两者进行相关性分析。将人非小细胞肺癌A549细胞分为正常对照组、转染对照组、miR-223-3p过表达组,其中正常对照组细胞未作任何处理,转染对照组细胞转染空质粒载体,miR-223-3p过表达组转染miR-223-3p mimics。MTT实验和平板集落形成实验检测各组细胞的增殖率,流式细胞术检测各组细胞的凋亡率,Western blot实验检测ECT2蛋白的表达水平;采用siRNA转染法沉默A549细胞中的ECT2后,检测细胞增殖及凋亡的变化。结果：miR-223-3p在癌组织中表达显著低于癌旁组织,而癌组织中ECT2蛋白表达高于癌旁组织,二者存在负相关关系（r=-0.666,P < 0.05）;与正常对照组和转染对照组细胞相比,miR-223-3p过表达组细胞的增殖率降低（P < 0.05）,ECT2蛋白表达显著降低,而细胞凋亡率显著升高（P < 0.05）;沉默ECT2对细胞增殖和凋亡的影响与过表达miR-223-3p基本一致。结论：miR-223-3p可能通过负向调控ECT2的表达,进而抑制非小细胞肺癌细胞增殖并诱导其凋亡,其作用机制有待进一步研究。