Expression and localization of cyclo-oxygenase isoforms in non-small cell lung cancer.

The beneficial effects of cyclo-oxygenase (COX) inhibitors in both colon cancer and adenomatous polyps suggest a role for the prostanoid pathway in epithelial malignancy. Although variable prostanoid synthesis in non-small cell lung cancer (NSCLC) has been demonstrated in freshly obtained tissue, COX messenger ribonucleic acid (mRNA) and protein localization in such tumours had not been investigated ex vivo. Thirty-four cases of primary NSCLC were examined for both constitutive (COX-1) and inducible COX (COX-2) by means of in situ hybridization and immunohistochemistry. COX-1 mRNA expression was absent or below the level of detection via in situ hybridization. COX-1 immunohistochemistry demonstrated uniform faint cytoplasmic staining in tumour cells and stromal inflammatory cells. Semiquantitative analysis of COX-2 expression in NSCLC demonstrated the highest levels of both mRNA and protein in adenocarcinoma cells (n=10, p<0.005 compared with large cell and squamous cell carcinoma), intermediate and variable expression in large cell carcinoma (n=11) and low or absent expression in squamous cell tumours (n=13). Levels of COX-2 expression in infiltrating inflammatory cells was the same in all tumour types. In conclusion, tumour cell cyclo-oxygenase-2 rather than cyclo-oxygenase-1 expression may account for the variable prostanoid production seen in non-small cell lung cancer, and primary lung adenocarcinoma expresses the highest levels of cyclooxygenase-2. Assessment of cyclo-oxygenase-2 expression ex vivo should be performed in studies examining the potential therapeutic effects of cyclo-oxygenase inhibitors in non-small cell lung cancer.     

Prognostic value of survivin,X‐linked inhibitor of apoptosis protein and second mitochondria‐derived activator of caspases expression in advanced non‐small‐cell lung cancer patients

Background and objective: Survivin and X‐linked inhibitor of apoptosis protein (XIAP) in vitro mediate cancer cell survival and chemoresistance. Second mitochondria‐derived activator of caspases (Smac), an antagonist of XIAP, has been shown in vitro to increase chemosensitivity. This study examined the prognostic value of survivin, XIAP and Smac in advanced non‐small‐cell lung cancer (NSCLC) patients treated with cisplatin‐containing chemotherapy. Methods: Semi‐quantitative RT‐PCR was used to measure survivin, XIAP and Smac mRNA expression in transbronchial biopsy tumour specimens from 72 patients with advanced NSCLC before commencing chemotherapy. Outcome measures were response to chemotherapy, progression‐free survival (PFS) and overall survival (OS). Results: Low expression of survivin was associated with good response to chemotherapy (P = 0.028). No association was found between XIAP and Smac expression levels and response to chemotherapy (P = 0.224 and P = 0.088, respectively). Patients with low survivin expression or high Smac expression had significantly longer PFS (P = 0.012 and P = 0.029, respectively) and OS (P = 0.007 and P = 0.031, respectively) compared with patients with high expression of survivin or low expression of Smac. XIAP expression was not correlated with PFS or OS. Additionally, PFS and OS in patients with performance status of 0 or 1 and stage IIIB were significantly longer than PFS and OS in patients with performance status (PS) of 2 and stage IV disease. Multivariate Cox regression analyses demonstrated that survivin and clinical stage were independent predictors for PFS and OS. Smac was an independent prognostic factor for OS, but not for PFS. Conclusions: Our findings suggest that the expression levels of survivin and Smac, but not XIAP, predict the survival of patients with advanced NSCLC treated with chemotherapy.     

Fragile histidine triad protein expression in nonsmall cell lung cancer and correlation with Ki-67 and with p53.

Fragile histidine triad (FHIT) is a tumour suppressor gene, which is altered in a variety of epithelial tumours, including lung cancer. Biochemical and functional pathways of its tumourigenicity are not yet understood. Its role in tumour proliferation is particularly controversial. The purpose of this study was to correlate the expression of FHIT protein in nonsmall cell lung cancer (NSCLC) with tumour proliferation as estimated by Ki-67 antigen and with p53, a suppressor gene. FHIT, Ki-67 and p53 expression were evaluated by immunohistochemistry in 119 resected NSCLC. Altogether, 58 tumours were negative (expression <10%) for FHIT. The median expression in tumours was 15% positive cells, in comparison with 100% in normal matched lung tissue. The expression was as strong as in normal tissue in only 19 cases. FHIT expression was significantly lower in squamous cell carcinoma (SCC) (5%) than in adenocarcinoma (ADC) (64%). The median expression of Ki-67 was 20% and 69% of tumours were positives (expression >10%). Ki-67 expression was significantly higher in SCC (33.3%) than in ADC (10%). The loss of FHIT protein was not correlated with the expression of p53 (median: 7.5%, 58% of positive tumours for a cut-off of 10% of positive cells) or Ki-67. But percentage of labelled cells for p53 and Ki-67 were significantly correlated. The results suggest that for fragile histidine triad, the pathway of tumourigenesis is independent of p53 and of tumoural proliferation, as reported previously in vitro.     

耐多药相关蛋白在人非小细胞肺癌组织中的表达

目的　检测人非小细胞肺癌组织中耐多药相关蛋白（ＭＲＰ） 的表达,探讨其与瘤组织学类型、分化程度、临床分期及预后的关系。方法　采用免疫组化方法及逆转录聚合酶链反应（ＲＴＰＣＲ）技术,分别检测了９２ 份人非小细胞肺癌石蜡组织中ＭＲＰ的表达及１６ 份人非小细胞肺癌新鲜组织中ＭＲＰ基因的表达。结果　９２ 份人非小细胞肺癌组织中（ 鳞癌４３ 例,腺癌４９ 例）ＭＲＰ表达阳性检出率为５４％（５０／９２） ,１６ 份人非小细胞肺癌组织中ＭＲＰ基因表达阳性检出率为３１％ （５／１６）。ＭＲＰ在腺癌中的表达阳性率明显高于鳞癌（ Ｐ＜ ０－０５） ,ＭＲＰ表达与瘤分化程度、肿瘤大小及淋巴结转移无显著相关。ＭＲＰ阳性患者术后５ 年生存率为１６％ （８／５０）,ＭＲＰ 阴性患者术后５ 年生存率为５２％ （２２／４２） ,二者经统计学处理,差异有显著性（ Ｐ＜ ０－０５）。ＭＲＰ阳性患者术后５ 年生存率有随其瘤组织中该蛋白表达阳性程度的增加而降低之趋势。结论　非小细胞肺癌患者瘤组织中ＭＲＰ的表达与瘤组织学类型及预后明显相关。     

Role of myofibroblasts in tumour encapsulation of hepatocellular carcinoma in haemochromatosis

BACKGROUND/AIMS: Hepatocellular carcinoma is a common malignancy and a major complication of untreated haemochromatosis. Encapsulation of liver tumours has been associated with a better prognosis and longer disease-free periods following resection. This study investigated the source of the tumour capsule in patients with haemochromatosis and coexisting hepatocellular carcinoma and examined potential factors influencing development. METHODS: Five haemochromatosis patients with encapsulated hepatocellular carcinoma were studied. Myofibroblasts were identified using combined immunohistochemistry and in situ hybridisation for alpha-smooth muscle actin and procollagen alpha1(I) mRNA, respectively. Immunohistochemistry was also performed for transforming growth factor (TGF)-beta1, platelet-derived growth factor (PDGF)-beta receptor and malondialdehyde. RESULTS: Procollagen alpha1(I) mRNA co-localised to alpha-smooth muscle actin positive myofibroblasts. The number of myofibroblasts was maximal within the capsule and decreased away from the tumour. TGF-beta1 protein was expressed in iron-loaded cells in non-tumour liver at the interface of tumour capsule. PDGF-beta receptor expression was observed in mesenchymal cells in the tumour capsule and in portal tracts. Malondialdehyde adducts were observed in the tumour, non-tumour tissue and in the capsule. CONCLUSIONS: This study provides evidence that myofibroblasts are the cell type responsible for collagen production within the tumour capsule surrounding hepatocellular carcinoma in haemochromatosis. The production of TGF-beta1 by iron-loaded hepatic cells at the tumour capsule interface may perpetuate the myofibroblastic phenotype, resulting in the formation of the tumour capsule.     

Pathologie und molekulare Pathologie von Lungentumoren

Carcinoma of the lung is one of the most common cancers and the most frequent cause of cancer death. Owing to the introduction of innovative substances it is possible to employ a differentiated tumour therapy that promises an improvement of prognosis and quality of life. The effectiveness of these therapeutic agents is based on specific molecular characteristics of lung carcinoma and poses new challenges regarding its pathological diagnostics. In addition to histotype, molecular genetic alterations, in particular EGFR mutations, have proved to be predictive markers. Therefore, histological typing comprises not only the distinction between small cell (SCLC) and non-small cell carcinoma (NSCLC) but also subtyping of NSCLC with involvement of immunohistological and molecular investigations. This article summarizes the criteria and innovations regarding the pathohistological evaluation of lung carcinoma under consideration of the TNM classification, grading, tumour regression, immunohistological expression patterns, mutation analyses and division into neuroendocrine tumours. In addition, we present the central points of a new interdisciplinary classification of adenocarcinoma.     

肺部占位性病变经纤支镜取材Survivin检测的意义

目的探讨新的凋亡抑制基因survivin在肺部占位性病变经纤支镜取材标本中的表达，及其临床应用价值。方法用原位杂交和免疫细胞化学的方法检测60例在经纤支镜抽吸物脱落细胞中survivin mRNA及其相应蛋白的表达。结果Survivin基因在NSCLC病例纤支镜抽吸物脱落细胞中呈高表达，survivin mRNA阳性表达与其蛋白表达呈正相关。Survivin和脱落细胞学联合检测较单用脱落细胞学检测敏感性和阴性预测值均提高。结论Survivin在非小细胞肺癌的发生发展中起重要作用，联合脱落细胞学和survivin基因分子生物学检测有助于肺癌的早期诊断。     

TGF-α、TGF-β1、TIMP-2 mRNA在非小细胞肺癌中的表达及临床意义

目的从TGF-α、TGF-β配体成员(TGF-β1、TGF-β2、TGF-β3)、TIMP-1、TIMP-2、MMP-2筛选在非小细胞肺癌(NSCLC)中mRNA水平异常表达的细胞因子,并探讨其临床意义。方法实时定量PCR法检测NSCLC组织和毗邻正常组织中TGF-α、TGF-β1、TGF-β2、TGF-β3、TIMP-1、TIMP-2、MMP-2 mRNA表达量,并研究其与患者临床病理特征之间的关系。结果 NSCLC组织中TGF-α、TGF-β1表达量均明显高于正常组织(0.046 vs 0.005,P0.01;16.91 vs 11.76,P=0.04),TIMP-2水平降低(25.23 vs 207.85,P=0.02),而NSCLC中MMP-2/TIMP-2显著高于正常肺组织(0.77 vs 0.02,P=0.04)。III-IV期NSCLC组织的TGF-α水平高于III期(0.06 vs 0.02,P0.01)。有淋巴结转移组TGF-β1水平高于无淋巴结转移组(17.70 vs 10.00,P0.01)。鳞癌组织中TIMP-2水平低于腺癌组织(20.06 vs 33.51,P=0.02),无淋巴结转移组TIMP-2水平高于有淋巴结转移组(30.02 vs 20.56,P=0.03)。结论 TGF-α与NSCLC进展密切相关;TGF-β1在NSCLC的发生、发展、侵袭转移中发挥重要作用;MMP-2/TIMP-2可能是较敏感的肿瘤生物学指标,且TIMP-2与NSCLC侵袭转移相关,相对高表达的TIMP-2可能提示腺癌。     

survivin基因在非小细胞肺癌中的表达及与P53、c-myc、k-ras蛋白表达的关系

目的：探讨survivin基因在非小细胞肺癌（NSCLC）中的表达,及与P53、c-myc、k-ras蛋白表达的相互关系。方法：逆转录PCR法检测了76例NSCLC肿瘤组织,20例良性瘤样病变和21例病灶旁正常肺组织survivin mRNA表达,免疫组织化学法检测P53,c-myc,k-ras蛋白表达,并将结果进行了相关分析。结果：61％NSCLC癌组织表达survivin mRNA基因,而良性瘤样病变和正常组织则分别为30％和19％（P＜0．001）。survivin基因表达与肺癌组织细胞类型,分化程度,TNM分期及淋巴结转移无明显相关关系。P53蛋白,c-myc与survivin基因表达显著相关,k-ras蛋白表达与survivin基因表达未见明显相关。结论：（1）survivin基因在肺癌组织中表达上调,提示该基因对NSCLC发生发展起重要作用。Survivin基因有望成为肺癌基因治疗的新靶点。（2）抑癌基因P53的失活和癌基因c-myc的上调与survivin基因的表达可能在NSCLC癌变中起协同作用。survivin和k-ras基因则可能通过各自不同的机制参与NSCLC的发病机制。     

The Role of cIAP1 and XIAP in Apoptosis Induced by Tumor Necrosis Factor Alpha in Esophageal Squamous Cell Carcinoma Cells

Background

The inhibitor of apoptosis protein (IAP) family are reported to play important roles in cancer cells evading apoptosis. However, the significance of their expression in human esophageal squamous cell carcinoma (ESCC) cells remains uncertain.

Aims

The present study aimed to investigate the role of the IAP family members in tumor necrosis factor-α (TNF-α)-induced apoptosis of human ESCC cells.

Methods

Five human ESCC cell lines were pretreated with TNF-α, cycloheximide (CHX, protein synthesis inhibitor), epoxomicin (proteasome inhibitor). Apoptosis assay and protein study with Western blot testing were conducted. Knockdown experiments with IAP siRNA were conducted, and the effect on cell apoptosis was analyzed.

Results

Significant apoptosis was induced in five ESCC cell lines by TNF-α plus CHX stimulation, but not when treated with TNF-α or CHX alone. The protein expression levels of cIAP1 and XIAP were decreased by treatment with TNF-α in the presence of CHX, and the degree of cIAP1 and XIAP expression decrease was correlated with sensitivity to TNF-α plus CHX-induced apoptosis. Epoxomicin suppressed TNF-α plus CHX-induced degradation of survivin, cIAP1, and XIAP, in addition to apoptosis. A caspase inhibitor (z-VAD-fmk) suppressed TNF-α plus CHX-induced apoptosis, but did not suppress degradation of survivin, cIAP1, and XIAP. Furthermore, cIAP1 or XIAP siRNA transfected cells underwent apoptosis in response to treatment with TNF-α alone. Double knockdown of both genes resulted in further increased apoptosis.

Conclusion

cIAP1 and XIAP play an essential role in the resistance of ESCC cells against apoptosis.

     

非小细胞肺癌组织中核因子κB的活性及其与肿瘤细胞增殖的关系

目的研究非小细胞肺癌组织中核因子κB（nuclear Factor-κB，NF—κB）的活性及其与细胞增殖、自发性细胞凋亡的关系。方法2006年5至10月收集30例非小细胞肺癌组织标本及15例肺癌患者癌旁5cm肺组织标本。NF—κB活性通过凝胶电泳迁移率改变试验（EMSA）检测，用RT—PCR和Western blot方法检测CyclinD1含量，免疫组织化学染色法检测增殖细胞核抗原（PCNA）蛋白含量，TUNEL法检测细胞凋亡。结果癌旁肺组织、鳞癌组织、腺癌组织中NF—κB活性（吸光度，A值）分别为24826±3724、28028±4204、35425±5317，三组比较差异有统计学意义（F=78．96，P〈0．01）。鳞癌组织、腺癌组织中NF—κB活性高于癌旁肺组织中NF—κB活性，腺癌组织中NF—κB活性高于鳞癌组织中NF—κB活性。健康肺组织、鳞癌组织、腺癌组织中CyclinD1 mRNA表达量分别为2．04±0．24、2．91±0．37、4．13±0．36，三组比较差异有统计学意义（F=62．43，P〈0．01）。癌旁肺组织、鳞癌组织、腺癌组织中Cyclin D1蛋白表达量分别为0．31±0．06、0．43±0．07、0．58±0．08，三组比较差异有统计学意义（F=89．24，P〈0．01）。癌旁肺组织、鳞癌组织、腺癌组织中PCNA蛋白表达量分别为0．32±0．09、0．42±0．10、0．54±0．16，三组比较差异明显。癌旁肺组织、鳞癌组织、腺癌组织中凋亡指数分别为（2．58±0．39）％、（2．27±0．34）％、（2．92±0．59）％，三组比较无明显差异。鳞癌组织中NF—κB活性与CyclinD1 mRNA、CyclinD1蛋白、PCNA蛋白均呈正相关（r值分别为0．51、0．54和0．60，P均〈0．05）；腺癌组织中NF—κB活性与CyclinD1mRNA、CyclinD1蛋白、PCNA蛋白均呈正相关（r值分别为0．60、0．64和0．68，P均〈0．05）；鳞癌、腺癌组织中NF—κB活性与细胞凋亡指数无相关关系。结论在非小细胞肺癌组织中NF—κB活性增高。非小细胞肺癌组织中NF—κB的异常活化可能与细胞增殖有关，但不影响自发性细胞凋亡。     

Survivin的表达与非小细胞肺癌生物学特征及预后的关系

目的探讨凋亡抑制基因Survivin的表达与非小细胞肺癌(NSCLC)生物学特征及临床预后的关系。方法应用免疫组化检测了70例NSCLC中Survivin的表达,并选取30例癌旁组织作为对照,比较Survivin的表达与NSCLC患者生物学特征及预后的关系。结果Survivin在NSCLC中的阳性表达率为74.3%,而癌旁组织无1例表达,Survivin的表达与肺癌患者年龄、性别、病理类型、分化程度无关(P0.05),与临床分期有关(P=0.008);Survivin表达阳性者生存时间明显低于阴性表达者(P=0.000),COX回归显示Survivin与NSCLC患者预后有关(P=0.001)。结论Survivin与NSCLC的发生发展有关,过度表达提示预后不良,Survivin有望成为肺癌诊断和基因治疗的新靶点。     

肝癌细胞放射敏感性与survivin蛋白表达的关系

目的 探讨肝癌细胞放射敏感性与survivin表达的关系.方法 肝癌细胞HepG2和SMMC-7721在接受不同剂量γ射线照射后,分别采用克隆形成法、免疫细胞化学法、流式细胞术、比色法等检测细胞存活率、survivin蛋白表达、细胞周期变化和Caspase-3活性.结果 在2Gy照射下HepG2和SMMC-7721细胞的存活分数分别为0.43±0.01与0.70±0.02,SMMC-7721较HepG2放射抗拒.γ射线对SMMC-7721细胞的G2/M期阻滞时间较HepG2细胞长(48 h对24 h),在阻滞峰各剂量点SMMC-7721细胞的G2/M期比例也更高.γ射线可上调两株肝癌细胞survivin蛋白的表达,照射后48～72 h,SMMC-7721细胞的survivin蛋白表达水平显著高于HepG2细胞(t值为2.81～5.20,P值均＜0.05).而Caspase-3的活化水平在放射敏感的HepG2细胞中更高(t值为6.05～6.72,P值均＜0.01).结论 射线诱导的survivin表达上调及survivin对Caspase-3的负调控可能是SMMC-7721细胞较HepG2细胞放射抗拒的原因之一.