检测粪便p53在胰腺癌诊断中应用的初步探讨

目的：本研究旨在探讨粪便p53基因突变检测方法在胰腺癌诊断中的潜在价值。方法：对连续就诊的胰腺癌62例患者的粪便，采用改进的两阶段酚－氯仿抽提和高浓度蛋白酶K消化法提取DNA。随后，采用PCR－SSCP方法检测粪便p53外显子5－8的突变情况。结果：以测定胰液p53突变率作为对照，粪便p53检测的敏感性为0．8，特异性为0．68，准确性为0．71，阳性预测值为0．36，阴性预测值为0．94。胰腺癌患者手术切除标本和粪便p53变检测结果一致率为63．6％（7／11），不一致率为36．4％（4／11），阳性预测值为0．33，阴性预测值为0．75。胰腺癌患者粪便p53突变率为37．1％（23．62），良性消化道疾病为12．8％（5／39），正常人为5．5％（3／55）。结论：粪便p53突变检测是一项具有潜在应用价值的对胰腺癌高危人群进行筛查的方法。     

p53在食管癌中的研究进展

p53作为抑癌基因,通过调节DNA修复、调控细胞周期、抑制血管生成及诱导细胞凋亡等机制,在机体组织细胞的生长、发育和分化过程中起着重要作用。食管癌是中国消化系统常见恶性肿瘤之一,大量研究证实,p53对食管鳞癌的作用机制、早期诊断、治疗和评估预后具有重要意义,此文就这些方面的研究进展作一综述。     

粪便p53基因突变检测在结直肠癌诊断中的应用

目的 探讨粪便p53基因突变检测对结直肠癌的诊断价值,旨在建立一种非侵入性的结直肠癌筛检的方法。方法 应用PCR-单链构象多态性分析-溴乙锭染色方法,检测40例结直肠癌、20例结直肠腺瘤及15例胃癌患者肿瘤组织和粪便标本p53基因突变。结果 40例结直肠癌患者粪便p53相应外显子DNA片段扩增率为90％,20例结直肠腺瘤为85％,15例胃癌为93％。所有标本总的扩增率为89％。40例结直肠癌患者组织标本中29例存在p53突变(72．5％),其中23例粪便测及p53突变,检测敏感性为57．5％,明显高于粪便隐血及血癌胚抗原检测的阳性率(P＜0．05);20例结直肠腺瘤患者组织标本中3例存在p53突变(15．0％),且均测及粪便p53基因突变。15例胃癌患者组织标本中10例存在p53突变(67．7％),粪便均未测及p53突变。结论 粪便p53基因突变检测诊断结直肠癌的敏感性较高,有望成为早期诊断大肠癌的一个相对敏感、特异、有效的指标,尤其在大范围人群筛检时。     

粪便中p53与APC突变检测在大肠癌诊断中的意义

目的探讨在大肠癌患者粪便中检测p53、APC基因突变的可行性及其应用前景和意义。方法从36例大肠癌患者、10例大肠腺瘤患者以及30例正常对照者的粪便中分别提取DNA,应用PCR-SSCP法检测粪便中p53、APC基因突变情况。结果36例大肠癌患者粪便中p53及/或APC基因突变检出率为77.78%(19/36),二者突变率分别为52.78%(19/36)和36.11%(13/36);10例大肠腺瘤中p53基因突变检出率为0%,APC为20%;30例正常对照粪便中p53、APC基因突变检出率均为0%。p53的突变随大肠癌分化程度的降低而增高(P<0.05);APC基因突变与大肠癌组织学类型无关(P>0.05)。结论联合检测粪便中p53与APC突变在大肠癌诊断和筛查中有潜在的应用价值。     

肿瘤抑癌基因p53与原发性肝癌

１９８８年,Ｂｅｎ－Ｄａｖｉｄ［１］在小鼠Ｆｒｉｅｎｄ病毒感染的红细胞中发现了Ｐ５３基因的频发失活,提出了Ｐ５３是一个肿瘤抑制基因、研究发现,将人野生型Ｐ５３基因转染到体外培养的肿瘤细胞中,可抑制肿瘤细胞生长。野生型Ｐ５３基因在正常细胞分裂过程中可能并无作用,但在细胞损伤后修复及不可修复细胞的清除过程中却必不可少。１　ｐ５３因异常与肿瘤的发生 ｐ５３基因是许多恶性肿瘤十分常见的共同基因损伤靶位,它的结构改变与表达异常可能是这些肿瘤发生的中心环节。点突变是ｐ５３基因异常表现的主要形式之一。大多数结…     

错配修复基因hMSH2与突变p53在散发性消化道肿瘤中的表达

人类ＤＮＡ错配修复系统 (ｍｉｓｍａｔｃｈｒｅｐａｉｒｓｙｓｔｅｍ ,ＭＭＲ )是由一系列特异性修复ＤＮＡ碱基错配的酶分子组成 ,此系统的存在 ,能避免遗传物质产生突变 ,保证ＤＮＡ复制的高保真度[1] 。ｈＭＳＨ2是目前研究较广泛的ＤＮＡ错配修复基因 ,它的失活可导致ＤＮＡ错配修复能力的降低 ,引起微卫星不稳定性 (ｍｉｃｒｏｓａｔｅｌｌｉｔｅｉｎｓｔａｂｉｌｉｔｙ ,ＭＳＩ) ,可使癌基因激活或抑癌基因失活 ,诱发细胞癌变[2 ] 。我们对 30例散发性消化道肿瘤ＤＮＡ上ｈＭＳＨ2基因与肿瘤组织中突变ｐ5 3的表达进行研究 ,以探讨ｈ…     

粪便中p53与APC突变检测在大肠癌诊断中的意义

目的：探讨在大肠癌患者粪便中检测p53、APC基因突变的可行性及其应用前景和意义。 方法：选取2005年6月-11月在郑州大学第一附属医院行全结肠镜检查并经病理证实的大肠癌患者36例、大肠腺瘤患者10例，经全结肠镜检查并证实的正常对照30例，共76例。其中3例为手术前大肠癌患者，余73例均为门诊全结肠镜检查病人。收集患者肠镜检查前或手术前新鲜粪便标本。所有粪便标本收集后立即保存于-20℃，并在3h内转移至-70℃保存备用。对标本进行预处理后，采用上海生工UNIQ-10柱式临床样品基因组抽提试剂盒提取DNA，并于-20℃保存。应用PCR-SSCP银染法检测粪便中p53、APC基因突变情况。实验结果采用SPSS10．0统计软件处理，基因频率采用直接计数法，各组间率的比较采用X^2检验和Fisher’s精确概率法。检验水准取α=0．05，P〈0．05有统计学意义。     

MicroRNAs与p53基因在肿瘤中相互关系的研究进展

肿瘤的发生、发展是多基因共同参与的多步骤的复杂过程,包括癌基因的异常激活和抑癌基因的失活.MicroRNAs(miRNAs)是一组真核细胞内源性产生的单链小RNA分子.研究发现miRNAs的表达水平与人类肿瘤发生有着密切的关系,可能参与调控癌基因和抑癌基因的异常表达.抑癌基因p53的突变是人类肿瘤中最常见的基因异常现象,目前已证实人类大约有50%的肿瘤与之相关.近来发现miRNAs的表达水平与p53的抑癌活性相关,本文就这一研究进展作一综述.     

p53与人类肺癌研究进展

ｐ５３基因是一种多功能的抑癌基因,近年来研究表明ｐ５３基因在人类支气管肺癌的发生、发展、转移、转归演化过程中发挥了重要角色。ｐ５３与肺癌关系的分子生物学和肿瘤发生学的深入研究为肺癌的基因诊断、基因治疗提供了科学依据,因而具有重大的临床应用价值。     

细胞老化和p53基因的研究进展

研究发现抑癌基因在抑制肿瘤发生的同时，还调控着细胞老化。细胞老化最初是在40年前防止培养的正常人纤维原细胞分化生长过程中发现的。细胞老化和凋亡被认为在防止类似发生于肿瘤中的不良细胞增殖有相同作用，但细胞凋亡是杀死并消除癌变倾向的细胞，而细胞老化仅仅是不可逆地使其生长停滞。在癌基因刺激应答过程中，细胞老化经常发生，最终导致细胞生长停滞，而不是死亡。细胞老化的机制还不完全清楚，从某种程度上可理解为一系列基因活动变化的结果。     

Clinical implication of screening p53 gene mutations in head and neck squamous cell carcinomas

The role of the tumour-suppressor gene p53 in the tumorigenesis of head and neck cancer has been well established, but the clinical significance of p53 alteration is still unclear. A group of 50 patients with head and neck squamous cell carcinoma (HNSCC) were investigated for p53 alterations. DNA was extracted from fresh tumour samples and polymerase chain reaction/single-strand conformation polymorphism analysis was used to detect p53 gene mutations in the region from exon 5 to exon 9. In addition, p53 protein overexpression was assessed by immunohistochemistry using the monoclonal antibody DO-7 on paraffin-embedded tissue sections. p53 gene mutations were found in 45% and p53 protein expression was detected in 61.2% of tumour samples. While p53 protein expression was not correlated with any clinical factors, p53 gene mutations indicated local regional recurrences of HNSCC. The risk of locoregional recurrence was significantly greater in patients with a p53 gene mutation than in patients with the wild-type p53 gene (P = 0.001). Multivariate analysis confirmed p53 gene mutation to be an independently predictive factor for the tumour recurrence (P = 0.0064). When we analysed p53 gene mutation in 12 patients with primary and recurrent tumours, we found that 4 patients (33.3%) had a different p53 gene mutation in the recurrent tumour from that in the original primary tumour. The results indicate that p53 gene mutations and not protein overexpression are valuable predictors for tumour recurrences and for differential diagnosis of a second primary HNSCC. Received: 20 January 1998 / Accepted: 20 March 1998     

原发性胃癌p53基因突变

目的 p53基因是当前抑癌基因研究中的热点之一。迄今,有关 p53基因异常与胃癌临床病理学参数如大体类型、临床分期、组织分化程度,浸润深度及淋巴结转移之间的关系尚无定论。Tumura 报告p53基因改变主要发生于异倍体瘤,国内尚无报道。本实验目的主要是分析中国人原发性胃癌 p53基因突变与这些病理参数,包括 DNA 倍体之间的关系。方法用聚合酶链式反应—单构象多态分析(PCR—SSCP)技术对20例原发性胃癌 p53基因外显子5—8突变进行检测。结果 8例(40%)发生了突变,其中2例发生在外显子7,4例发生在外显子8。0至Ⅲ期均有突变存在。66.7%(6/9)的异倍体瘤检测到了p53突变,而二倍体瘤中只有18.2%(2/11)发生了 p53突变。结论 p53基因突变与胃癌临床病理参数如大体类型、分期、组织分化程度、浸润深度及淋巴结转移之间无明显关系,而与胃癌 DNA 倍体改变有关。     

p53 overexpression and K-ras gene mutations in primary sclerosing cholangitis-associated biliary tract cancer

Cholangiocarcinoma occurs frequently in patients with primary sclerosing cholangitis (PSC). We evaluated the incidence and prognostic significance of p53 protein overexpression and K-ras gene mutations in patients with biliary tract cancer and PSC. p53 protein expression was determined in specimens from 12 patients with biliary tract cancer, using the antibody, D07. K-ras mutations were detected using DNA sequencing and a mutation ligation assay. Accumulation of p53 protein was detected in 6 of 12 tumors (50%). K-ras mutations were detected in 4 of 12 tumors (33%). Overall survival in patients with p53-negative tumors was significantly longer (P < 0.05) than that in patients with p53-positive (mutant) tumors. Similarly, overall survival was significantly longer (P < 0.05) in the absence of a K-ras mutation than in patients with a tumor containing a K-ras mutation. Mean interval from the time of diagnosis of PSC until the diagnosis of biliary tract cancer was significantly shorter (P < 0.05) in patients with p53 overexpression than in those patients without p53 overexpression (2 versus 47 months). p53 overexpression and K-ras mutations occur commonly in patients with PSC and biliary tract cancer and are associated with a shortened survival. Patients with longstanding PSC are less likely to have these genetic alterations and may have a better prognosis.     

结肠癌合并血吸虫感染患者癌细胞p53基因的表达

目的 研究原发性结肠癌细胞p53基因mRNA的表达水平及合并血吸虫感染后的差异,探讨其与患者临床病理特征的关系。 方法 38例原发性结肠癌患者分为两组,A组（合并血吸虫感染）20例和B组（未合并血吸虫感染）18例。应用实时荧光定量 PCR和相对定量分析法检测患者肿瘤组织中的p53 mRNA。 结果 p53 mRNA在两组中均可检出,A组中的基因表达水平显著高于B组（P＜0.05）。p53基因mRNA表达水平与年龄、性别相关无显著性,与肿瘤大小、有无淋巴结转移相关具有显著性。结论 p53基因mRNA的高水平表达与结肠癌的侵袭和发展具有显著性差异,血吸虫感染可能对结肠癌患者p53基因的突变有一定影响。     

联合检测粪便中k-ras与p53基因诊断大肠癌

目的探讨粪便中k-ras与p53基因突变的检测用于大肠癌诊断及筛查的可行性。方法应用聚合酶链反应-单链构象多态性分析-银染法检测大肠癌组织及粪便中k-ras与p53基因突变,并与粪便潜血实验比较。结果31例大肠癌患者组织中k-ras基因突变9例,粪便中检测出7例;p53突变12例,粪便中检测到p53突变10例,二者符合率分别为77%（7/9）和83%（10/12）,而在正常组织及正常人粪便中均未检测到突变,特异性100%。粪便中k-ras与p53基因突变与肿瘤分化程度,Dukes分期,肿瘤部位,粪便潜血,癌胚抗原无关。结论粪便中突变基因检测有望成为一种新的大肠癌无创性筛查方法。     

胰管刷检标本P53蛋白检测在胰腺癌诊断中的价值

目的：探讨胰管刷检标本p53蛋白检测在胰腺癌诊断中的价值。方法：应用免疫组化法检测26例胰腺及壶腹疾病患者胰管刷检标本p53蛋白的表达,并与常规细胞学检查作比较。结果：苏木精－伊红染色常规细胞学检查诊断胰腺癌的敏感性为53％,特异性为100％,准确性为70％。胰管刷检标本p53蛋白检测诊断胰腺癌的敏感性为59％,特异性为100％, 准确性为74％。二者联合诊断胰腺癌的敏感性为71％,特异性为100％,准确性为81％,与单项细胞学检查相比差异有非常显著性（P＜0．01）。结论：胰管刷检标本细胞学检查的同时,进行p53检测可提高胰腺癌的诊断率,有助于胰腺良、恶性疾病的鉴别。     

K-ras and p53 mutations in DNA extracted from colonic epithelial cells exfoliated in faeces of patients with colorectal cancer

BACKGROUND: Exfoliated colonic epithelial cells in faeces provide a source of human DNA which may be analysed for the presence of tumour-induced modification. AIM: In the present study we investigated K-ras and p53 mutations in faeces of patients with colorectal carcinoma, to verify whether analysis of these mutations might identify a high percentage of patients with colorectal cancer. PATIENTS AND METHODS: Faeces, tumour and normal mucosa samples were taken from 26 patients. Polymerase chain reaction amplification and restriction enzyme analysis were performed to detect K-ras mutations; p53 gene mutations were identified by using polymerase chain reaction amplification and single strand conformation polymorphism. RESULTS: We were able to amplify the K-ras gene and exons 5-9 of the p53 gene in 100% of the faecal samples studied. K-ras and p53 gene mutations were detected in faeces in 26.9% and 50% of the cases, respectively. The two mutations were present together in 5 out of 26 patients. There was full agreement between the K-ras and p53 pattern observed in faecal DNA and that in tumour tissue DNA. CONCLUSIONS: Application of K-ras and p53 mutation gene analysis in the faeces may have clinical applications in the future. Since this genetic analysis is able to detect only 57.7% of patients with colorectal cancer, the study of other genes involved in colorectal carcinogenesis is necessary.     

Usefulness of p53 gene mutations in the supernatant of bile for diagnosis of biliary tract carcinoma: comparison with K- ras mutation

Background: The sensitivity of bile cytology for the diagnosis of biliary tract carcinoma (BTCa) is still low. In addition, the incidence of detection of genetic mutations in the bile of BTCa is not satisfactory yet. To improve the molecular diagnosis of BTCa, we analyzed p53 and K-ras mutations in DNA extracted from not only the sediment but the supernatant of bile samples. Methods: Polymerase chain reaction-single-strand conformation polymorphism and direct sequencing were used for analyses of p53 mutations in exons 5 through 8. K-ras mutations at codon 12 were examined by mutant allele-specific amplification. Results: In bile supernatant from patients with BTCa, p53 and K-ras mutations were detected in 50.0% (15/30) and 56.7% (17/30) of cases, respectively. The incidence of p53 and K-ras mutations in the sediment was 33.3% and 43.3%, respectively. When a combination assay with both genes was used, molecular abnormalities were detected in 80.0% of cases, including 3 in which p53 alone was positive. In addition, either p53 or K-ras mutations were detected in 12 of 15 (80.0%) cases of BTCa in which the cytologic diagnoses were negative. p53 mutations were detected in neither supernatant nor sediment in 20 patients with cholelithiasis, although the incidence of K-ras mutations in the sediment was 20%. Conclusions: The incidence of p53 and K-ras mutations is higher in the supernatant than in the sediment, and simultaneous analyses of p53 and K-ras in the two bile fractions could enhance the genetic diagnosis of BTCa. Notably, the specificity of p53 mutations for cancer was very high in bile samples, and the sensitivity was also relatively high. Received: November 19, 2001 / Accepted: February 22, 2002 Acknowledgments. This work was supported in part by Grants-in-Aid for Scientific Research from the Japanese Ministry of Education, Science, Sports, and Culture. Reprint requests to: N. Sawabu