人胰腺癌bcl-2基因的过度表达--免疫组织化学和原位杂交研究

目的 :应用地高辛标记bcl 2cDNA进行mRNA原位杂交和免疫组织化学分析胰腺癌中bcl 2基因的mRNA转录和蛋白翻译 ,研究bcl 2基因结构和功能表达的关系。方法 :以胰腺癌病人癌组织( 50例 )、胰腺癌转移灶组织 ( 15例 )为研究对象 ,采用免疫组织化学 (SP法 )和分子原位杂交法检测bcl 2在胰腺癌中的表达。对照组 10例为慢性胰腺炎组织。结果 :bcl 2蛋白在胰腺癌组明显高于胰腺癌转移灶组 ,Ⅰ、Ⅱ期、无转移、高分化的胰腺癌明显高于Ⅲ、Ⅳ期、有转移和低分化的胰腺癌。免疫组织化学与原位杂交技术检测胰腺癌组织中bcl 2基因表达结果一致。结论 :bcl 2蛋白可能作用于胰腺细胞的癌变阶段 ,并与胰腺导管上皮癌变有关 ;bcl 2基因在mRNA和蛋白水平是相对稳定的     

肿瘤转移抑制基因KiSS-1在胰腺癌组织中的表达及意义

目的:检测KiSS-1mRNA及KiSS-1蛋白metastin在胰腺癌组织中的表达,探讨其在胰腺癌浸润及转移中的作用.方法:采用逆转录多聚酶链反应(RT-PCR)法及免疫组织化学S-P法检测42例胰腺癌,10例慢性胰腺炎及12例正常胰腺组织中KiSS-1mRNA及KiSS-1蛋白metastin的表达情况及与各种临床病理参数的关系.结果:KiSS-1mRNA及Kiss-1蛋白metastin在胰腺癌组中的阳性表达显著低于慢性胰腺炎组及正常胰腺组(P＜0.01);KISS-1mRNA和KiSS-1蛋白metastin的表达均与胰腺癌临床分期及淋巴结转移有关(P＜0.05);二者在胰腺癌组织中的表达存在明显相关性.结论:肿瘤转移抑制基因KiSS-1在抑制胰腺癌的浸润和转移过程中可能起着重要的作用.     

神经菌毛素在胰腺癌组织及MIA PaCa-Ⅱ细胞系中的表达

目的探讨神经菌毛素（Neuropilin-1，NRP-1）在胰腺导管癌组织及MIA PaCa-Ⅱ胰腺癌细胞系中的表达及意义。方法运用免疫组化和RT-PCR法分别检测在正常胰腺组织、癌旁组织、胰腺癌组织及MIA PaCa-Ⅱ细胞系中Neuropilin-1蛋白及mRNA表达水平。结果蛋白水平：可见正常胰腺组织无表达，癌旁组织轻度表达，而胰腺癌组织及MIA PaCa-Ⅱ胰腺癌细胞中高水平表达。神经组织也可表达Neuropilin-1 mRNA水平见正常胰腺组织呈微量表达，癌旁组织中度表达，而胰腺癌组织及MIA PaCa-Ⅱ胰腺癌细胞中呈高水平表达。结论Neuropilin-1可能与参与了胰腺癌的发生发展，在胰腺癌神经转移中可能起着重要作用。     

LivinmRNA在胰腺癌中的表达及与Bcl-2相关性的研究

目的:探讨凋亡抑制基因Livin mRNA在胰腺癌组织中的表达及其与Bcl-2蛋白表达的关系.方法:采用逆转录-多聚酶链反应(RT-PCR)法检测52例胰腺癌、10例慢性胰腺炎和12例正常胰腺组织中Livin mRNA的表达;用免疫组织化学法检测胰腺癌组织中Bcl-2蛋白的表达.结果:Livin mRNA在胰腺癌中的表达率为71.2%,明显高于慢性胰腺炎和正常胰腺组织(P<0.01).Livin基因表达与胰腺癌的分化程度和淋巴结转移有关(P<0.05).Livin基因表达与Bcl-2蛋白表达显著相关.结论:Livin基因在胰腺癌的发生发展中起重要作用;Livin与Bcl-2的异常表达可能在胰腺癌的发生中起协同作用.     

胰腺癌组织转录因子GATA-6表达及其意义的探讨

目的:探讨胰腺癌组织GATA-6的表达及其对胰腺癌发生、发展的可能作用及临床意义。方法:采用免疫组化检测33例胰腺癌、15例慢性胰腺炎和6例正常胰腺组织GATA-6蛋白的表达;免疫细胞荧光法检测胰腺癌细胞株Panc-1、BxPC-3和SW1990及正常胰腺导管上皮细胞H6C7的GATA-6蛋白表达情况;Real-time RT-PCR技术检测上述细胞株的GATA-6 mRNA表达;并分析GATA-6与胰腺癌临床病理参数的关系。结果:GATA-6蛋白在胞核中表达,胰腺癌组织GATA-6的阳性表达率(88.9%)显著高于慢性胰腺炎(40.0%)和正常胰腺组织(16.7%),P<0.01,但与性别、年龄、分化程度和临床分期均无显著性相关;3种胰腺癌细胞株均有GATA-6蛋白表达,而正常胰腺导管上皮细胞无表达;3种胰腺癌细胞株GATA-6 mRNA表达也明显高于正常胰腺导管上皮细胞,P<0.05。结论:GATA-6在胰腺癌组织及细胞中呈高表达,提示GATA-6在胰腺癌的发生、发展中可能起一定作用。     

胰腺癌与mdrl基因的相关性研究(英文)

目的通过研究未化疗的原发性胰腺癌 mdrl 基因 mRNA 及其蛋白产物 P-gp 的表达与肿瘤生物学特性的关系,以便指导胰腺癌的临床治疗。方法应用免疫组织化学、原位 PCR 技术对150例胰腺石蜡包埋标本(包括97例原发胰腺癌,32例胰腺炎及21例正常胰腺组织)组织中的 mdrl mRNA 和 P-gp 进行检测。结果 mdrl mRNA 和 P-gp 染色主要分布于正常胰腺小导管及肿瘤的导管上皮细胞的细胞膜和/或细胞浆内。胰腺癌和胰腺炎及正常胰腺组织中 mdrl mRNA 和 P-gp 表达阳性率之间存在着显著性差异(P<0.05),mdrl mRNA 和 P-gp 的阳性表达率与肿瘤的分化程度、浸润性和 TNM 分期等生物学特性有关(P<0.05),而与病人的年龄、性别、肿瘤的大小及部位等临床病理资料无关(P>0.05)。结论研究结果表明胰腺癌 mdrl 基因的表达与其“天然”耐药有关,检测胰腺癌 mdrl 基因表达对预测胰腺癌的化疗敏感性具有重要指导意义。同时它可能作为判断肿瘤的恶性程度及预后的一个有效生物学指标。     

胰腺癌与mdr1基因的相关性研究

目的　　通过研究未化疗的原发性胰腺癌mdrl基因mRNA及其蛋白产物p-gP的表达与肿瘤生物学特性的关系,以便指导胰腺癌的临床治疗。方法　　应用免疫组织化学、原位PCR技术对150例胰腺石蜡包埋标本(包括97例原发胰腺癌,32例胰腺炎及21例正常胰腺组织)组织中的mdrl　mRNA和P —gp进行检测。结果　　mdrl　mRNA和p-gP染色主要分布于正常胰腺小导管及肿瘤的导管上皮细胞的细胞膜和/或细胞浆内。胰腺癌和胰腺炎及正常胰腺组织中mdrl　mRNA和P-gp表达阳性率之间存在着显著性差异(P<0.05),mdrl　mRNA和P—gP的阳性表达率与肿瘤的分化程度、浸润性和TNM分期等生物学特性有关(P<0.05),而与病人的年龄、性别、肿瘤的大小及部位等临床病理资料无关(P>0.05)。结论　　研究结果表明胰腺癌mdrl基因的表达与其“天然”耐药有关,检测胰腺癌mdrl基因表达对预测胰腺癌的化疗敏感性具有重要指导意义。同时它可能作为判断肿瘤的恶性程度及预后的一个有效生物学指标。     

紧密连接蛋白claudin-1 及ZO-1 在胰腺癌中的表达*

目的:探讨紧密连接蛋白claudin- 1 及ZO- 1 在胰腺癌中的表达及在胰腺癌侵袭转移中的意义。方法:应用免疫组织化学方法检测claudin- 1 和ZO- 1 在胰腺癌不同侵袭位点（与非肿瘤胰腺组织相交界的胰腺癌侵袭前沿区、肿瘤中央区、与周围间质相交界的胰腺癌侵袭前沿区）及淋巴结转移灶和正常胰腺组织中的表达。结果:claudin- 1 与ZO- 1 蛋白正常定位于胰腺腺泡细胞和导管上皮细胞胞膜上,而在胰腺癌组织中,claudin- 1 与ZO- 1 蛋白定位从细胞膜异位至细胞浆或表达缺失,重度异位表达率分别为66.7% 和69.2% 。在与非肿瘤胰腺组织相交界处的胰腺癌组织中,低- 未分化胰腺癌的claudin- 1 重度异位表达率（91.7%）明显高于高- 中分化胰腺癌（55.6% ,P<0.05）;与周围间质相交界处的胰腺癌侵袭前沿区claudin- 1 重度异位表达率（89.7%）高于与非肿瘤胰腺组织相交界处的胰腺癌侵袭前沿区（66.7% ,P<0.05）;淋巴结转移灶重度异位表达率最高（92.3% ,P<0.05）。 同样与周围间质相交界处的胰腺癌侵袭前沿区ZO- 1 重度异位表达率（94.9%）也高于与非肿瘤胰腺组织相交界处的胰腺癌侵袭前沿区（64.2% ,P<0.05）,淋巴结转移灶最高（100% ,P<0.05）;原发胰腺癌组织各个位点中claudin- 1 与ZO- 1 表达呈正相关关系。结论:claudin- 1 和ZO- 1 异位表达对胰腺癌的发生起促进作用;claudin- 1 异位表达与胰腺癌的分化有关;claudin- 1 和ZO- 1 异位表达率增加促进胰腺癌的侵袭与转移。      

MTDH在胰腺癌中的表达及其与E-cadherin、MVD的关系

[目的]研究MTDH在胰腺癌中的表达及其与钙黏蛋白(E-cadherin)、肿瘤微血管密度的关系及其临床意义.[方法]应用免疫组织化学SP法检测MTDH、E-cadherin、CD31在60例胰腺癌及其癌旁正常胰腺组织中的表达;并用抗CD31抗体标记微血管,计数微血管密度.应用即时荧光定量PCR检测10例胰腺癌及其癌旁正常胰腺组织中MTDH mRNA的表达水平.[结果]MTDH在胰腺癌组织中的表达率显著高于癌旁胰腺组织中的表达(P＜0.05);胰腺癌组织MTDH的表达与患者临床分期(P=0.014)、淋巴结转移(P=0.002)和远处转移(P=0.006)相关;E-cadherin的表达与患者临床分期(P=0.002)、淋巴结转移(P=-0.023)和组织学分级(P=-0.027)相关.MTDH在胰腺癌组织中的表达与E-cadherin表达负相关(P＜0.05),与MVD表达呈正相关(P＜0.05).[结论]MTDH在胰腺癌中高表达,其表达水平与肿瘤的进展和转移相关.MTDH蛋白的高表达可能通过影响肿瘤E-cadherin的表达和肿瘤微血管生成来促进胰腺癌的转移.     

乳腺癌中神经激肽受体的表达及受体拮抗剂的作用研究*

目的:研究乳腺癌中全长型、截短型神经激肽1 受体（neurokinin 1 receptor,NK1R）和神经激肽2 受体（neurokinin 2 receptor,NK2R）的表达,及受体拮抗剂对乳腺癌细胞生长的影响。方法:采用免疫组织化学法检测天津医科大学肿瘤医院51例乳腺癌及其癌旁正常组织、30例乳腺良性病变组织总NK1R（包括NK1R-FL和NK1R-Tr）和NK2R 表达,采用实时定量PCR 和免疫印迹技术检测乳腺细胞系中NK1R-FL、NK1R-Tr和NK2R 表达,建立NK1R-FL和NK1R-Tr过表达的乳腺细胞系, 在NK1R 和NK2R 拮抗剂作用下测定细胞增殖和软琼脂集落形成能力。结果:乳腺癌及癌旁正常组织、乳腺良性病变组织中均总NK1R 过表达,乳腺癌组织中 NK1R-FL、NK2R 表达相比癌旁正常组织显著降低,并与乳腺癌分型、组织学分级、淋巴结转移及 Ki-67、HER-2、ER和PR表达相关。HBL-100 细胞中NK1R-FL和NK2R 过表达、NK1R-Tr低表达,MDA-MB-231、T-47D 和MCF-7 细胞中只表达NK1R-Tr。乳腺癌细胞中NK1R-Tr低表达、NK1R-FL表达增加其对NK1R 和NK2R 受体拮抗剂的敏感度。结论:乳腺组织中NK1R-FL、NK2R 共表达,乳腺癌细胞中NK1R-Tr过表达并负反馈调节NK1R-FL和NK2R 的表达,NK1R 和NK2R 受体可能成为乳腺癌治疗的新靶标。      

Characterization of cytokeratin 20 expression in pancreatic and colorectal cancer.

Cytokeratin 20 belongs to the epithelial subgroup of the intermediate filament family. Because of its restricted range of expression in humans, it has become an important tool for detecting and identifying metastatic cancer cells by immunohistochemistry and by PCR analysis. Despite its widespread diagnostic use in colorectal cancer and occasional use in pancreatic cancer, little is known about the expression of CK 20 in these tumors in vivo. Therefore, in the present study we characterized CK 20 expression in pancreatic and colorectal cancer by comparison with its expression in the normal pancreas and colon. Tissue samples from 24 patients with pancreatic cancer and from 41 patients with colorectal cancer were examined for CK 20 expression by Northern blot analysis, immunohistochemistry, and in situ hybridization. CK 20 expression was observed in the cancer cells of both cancer types. A subgroup of the pancreatic cancers exhibited a 3.2-fold increase in CK 20 mRNA by comparison with respective normal controls. In contrast, colon cancers underexpressed CK 20 mRNA by comparison with the respective controls. In the normal tissues, CK 20 immunoreactivity was relatively faint and sparse in the pancreatic ductal cells but intense and abundant in the apical portions of the colonic mucosa. CK 20 immunoreactivity was also evident in the ductal cells from the chronic pancreatitis-like lesions adjacent to the cancer cells. Furthermore, distant metastases from pancreas carcinomas exhibited strong CK 20 immunoreactivity. It is concluded that CK 20 is overexpressed in pancreatic cancer and that it can serve as an excellent marker for metastatic pancreatic cancer.     

Cripto,a member of the epidermal growth factor family,is over-expressed in human pancreatic cancer and chronic pancreatitis

Cripto is a 188 amino-acid protein containing a central segment that shares amino-acid sequence homology with epidermal growth factor (EGF) and transforming growth factor alpha (TGF-α). The EGF receptor, EGF and TGF-α are expressed in the normal human pancreas, and are over-expressed in human pancreatic cancer. Therefore, in the present study we sought to determine whether cripto is found in the normal human pancreas and whether its expression is altered in pancreatic cancer. Because chronic pancreatitis (CP) is associated with interstitial fibrosis similar to that observed in pancreatic cancer, we also examined cripto expression in pancreatic tissues from patients with CP. In the normal pancreas, cripto immunoreactivity was found at moderate levels in most ductal cells and was present very faintly in a rare acinar cell. In 26 of 58 pancreatic cancers, cripto immunoreactivity was present in many cancer cells. Its presence was associated with advanced tumor stage, but not with shorter post-operative survival. Cripto was also present in acinar and ductal cells adjacent to the cancer cells, and in many ductal and atrophic acinar cells in the CP samples. Northern blot analysis revealed a marked increase in cripto mRNA levels in the cancer and CP samples. By densitometry, there was a 11 - and 4-fold increase in cripto mRNA levels in pancreatic cancer and CP respectively. Southern blot analysis did not reveal an increase in gene copies encoding cripto either in cancer or in CP. These findings indicate that cripto expression may contribute to disease progression in pancreatic cancer, and implicate cripto in the histopathological alterations that occur in the pancreas both in cancer and in CP.     

Differential expression of TRAIL-R3 and TRAIL-R4 in human pancreatic cancer

BACKGROUND: Pancreatic cancer is one of the most aggressive cancers, in part due to its insensitivity to most treatment modalities. This resistance towards cytotoxic therapy is thought to be caused--at least in part--by a general resistance of pancreatic cancer cells towards apoptosis. TRAIL-R3 and TRAIL-R4, which belong to the TRAIL receptor family, can inhibit TRAIL-induced apoptosis. PATIENTS AND METHODS: Seven normal pancreatic tissues and 7 pancreatic cancer tissues were analyzed using Northern blotting, Western blotting and immunohistochemistry. RESULTS: TRAIL-R3 mRNA and protein expression were generally weak in pancreatic cancers and normal pancreatic tissues. In contrast, TRAIL-R4 mRNA and protein were expressed at moderate to high levels in human pancreatic cancer tissues, but demonstrated weak to negative expression in the normal pancreas. CONCLUSION: TRAIL-R4 but not TRAIL-R3 levels were significantly different in pancreatic cancer in comparison to the normal pancreas. These findings give new insight into the resistance mechanisms of pancreatic cancer towards TRAIL-mediated apoptosis.     

Inhibition of heme oxygenase-1 increases responsiveness of pancreatic cancer cells to anticancer treatment.

Heme oxygenase-1 (HO-1) is believed to represent a key enzyme for the protection of cells against "stress." Its overexpression in different types of human cancers supports the notion that HO-1 provides a growth advantage and contributes to cellular resistance against chemotherapy and radiotherapy. Given the poor survival rates of patients with pancreatic cancer due to its aggressive growth behavior and its exceptional resistance to all known forms of anticancer treatment, we have investigated the expression of HO-1 in human pancreatic cancer cells growth behavior and prognosis. Expression of HO-1 was analyzed in human pancreatic cancer samples in comparison with normal pancreas by quantitative PCR, Western blot, and confocal microscopy. The influence of radiotherapy and chemotherapy on HO-1 expression in pancreatic cancer cell lines was evaluated. Furthermore, HO-1 expression was specifically suppressed by small interfering RNA transfection and subsequently the alterations of growth behavior and resistance to anticancer treatment were tested. Human pancreatic cancer showed a 6-fold and 3.5-fold HO-1 up-regulation in comparison to normal pancreas based on mRNA and protein level, respectively (P < 0.05). Cancer tissues revealed marked HO-1 immunoreactivity in tumor cells and in tumor associated immunocytes. Treatment of the pancreatic cancer cell lines with gemcitabine or radiation strongly induced HO-1 expression. Targeted knockdown of HO-1 expression led to pronounced growth inhibition of the pancreatic cancer cells and made tumor cells significantly more sensitive to radiotherapy and chemotherapy. Therefore, specific inhibition of HO-1 expression may be a new option in pancreatic cancer therapy and may be used as sensitizer to chemotherapy and radiotherapy.     

Adhesion molecules in human pancreatic cancer

BACKGROUND AND OBJECTIVES: Adhesion molecules are cell surface glycoproteins that are important in cell-to-cell and cell-to-extracellular matrix interactions. In the present study, we analyzed the adhesion molecules ICAM-1 (intercellular adhesion molecule-1), VCAM-1 (vascular cell adhesion molecule-1), and ELAM-1 (endothelial leukocyte adhesion molecule-1) in human pancreatic cancer. METHODS: ICAM-1, VCAM-1, and ELAM-1 were analyzed in 20 pancreatic cancer specimens and 20 normal pancreatic tissues. mRNA expression encoding ICAM-, VCAM-1, and ELAM-1 was assessed with Northern blot analysis. The distribution and localization of ICAM-1, VCAM-1, and ELAM-1 was determined in the pancreatic specimens by immunohistochemistry. RESULTS: Northern blot analysis revealed a 5.4-fold increase of ICAM-1 (P<0.01) and a 3.7-fold increase in VCAM-1 (P<0.01) mRNA expression in cancer samples in comparison with normal controls. In contrast, ELAM-1 mRNA levels did not show significant differences between the cancer and the normal tissues. Immunohistochemical analysis of cancer tissues showed strong immunostaining for ICAM-1 and VCAM-1, and faint immunostaining for ELAM-1 in the pancreatic cancer cells. Fibrotic or noncancerous pancreatic tissue adjacent to the cancer mass was devoid of any immunoreactivity for ICAM-1, ELAM-1, and VCAM-1. In contrast, the normal pancreas exhibited no immunoreactivity of ICAM-1, ELAM-1, and VCAM-1. CONCLUSIONS: Enhanced expression of ICAM-1 and VCAM-1 in human pancreatic cancers suggests a role in tumor pathogenesis. The increase of these adhesion molecules might influence the detachment of cancer cells in the primary tumor, might contribute to cancer cell migration and the spread of cancer cells to distant organs, or both.     

Aberrant expression of neuropilin-1 and -2 in human pancreatic cancer cells.

PURPOSE: Neuropilin (Np)-1 and -2 are coreceptors for vascular endothelial growth factor (VEGF). This study was designed to assess their role in pancreatic ductal adenocarcinoma (PDAC). EXPERIMENTAL DESIGN: We assessed Np-1 and Np-2 expression by real-time quantitative PCR in relation to the expression of VEGF ligands and receptors in pancreatic cancer cell lines and tissues. RESULTS: ASPC-1, CAPAN-1, and PANC-1 pancreatic cancer cells and tumor-derived, laser-captured pancreatic cancer cells exhibited higher Np-1 and Np-2 mRNA levels than VEGF receptor-1, -2, or -3 mRNA levels. Transfection of Np-1 and Np-2 cDNAs in COS-7 cells, and treatment with tunicamycin revealed that both proteins were glycosylated. Both proteins were expressed in pancreatic cancer cell lines, in the PDAC samples, and in acinar cells adjacent to the cancer cells. The normal pancreas was devoid of Np-1 immunoreactivity, whereas Np-2 immunoreactivity was present in the endocrine islets and in some acinar cells, but not in ductal cells. CONCLUSIONS: The aberrant localization of Np-1 and Np-2 in the cancer cells in PDAC suggests that in addition to exerting proangiogenic effects, these coreceptors may contribute to novel autocrine-paracrine interactions in this malignancy.     

Enhanced expression of annexin II in human pancreatic carcinoma cells and primary pancreatic cancers

Annexin II is a calcium and phospholipid binding protein anda substrate for protein-tyrosine kinases. Recent investigationshave revealed involvement of annexin II in DNA synthesis andcell proliferation. Increased levels of annexin II are observedin cancer cells and tissues. To investigate the expression ofannexin II in pancreatic adenocarcinoma cells and primary tumors,we measured the levels of annexin II mRNA and protein in normalhuman pancreas, five established human pancreatic adenocarcinomacell lines, three primary pancreatic cancers and one metastatictumor. All five cell lines examined had 5- to 15-fold higherlevels of annexin II as compared to normal pancreas. Significantelevations (2-to 8-fold) of annexin II expression were observedin the three primary pancreatic tumors and one metastatic tumorexamined. Immunocytochemical analysis indicates that the increasedexpression of annexin II is limited to proliferating ductularadenocarcinoma, and annexin II expression co-localizes withcells that express PCNA. In normal pancreas, annexin II expressionis seen in ductal and ductular cells and no expression is seenin acinar or islet cells. We conclude from these findings thatannexin II has a role in cell proliferation and its regulationis altered in pancreatic cancer.