胰腺癌组织相关蛋白质分子的差异表达

目的用双向凝胶电泳的方法建立胰腺癌、癌旁组织和正常胰腺组织的蛋白质组二维图谱并分析差异蛋白质点。方法提取人正常胰腺组织、胰腺癌和癌旁组织总蛋白质，双向电泳分离蛋白，比较不同组织中蛋白质表达的差异。每份组织均重复电泳3次。结果在同一份组织的3张银染的凝胶蛋白质图谱上均可辨识1000个左右的蛋白质点，蛋白质点在形状、位置和密度上一致，重复性好。在不同组织之间发现了22个明显差异的蛋白质点，初步确定了其等电点和分子量。正常胰腺组织有一个高表达的蛋白质点，癌和癌旁组织有21个高表达的蛋白质点。结论正常胰腺组织、胰腺癌、癌旁组织蛋白质组二维图谱存在明显的差异，部分差异蛋白质可能具有诊断应用价值。     

胰腺癌组织相关蛋白质分子的差异表达

目的用双向凝胶电泳的方法建立胰腺癌、癌旁组织和正常胰腺组织的蛋白质组二维图谱并分析差异蛋白质点.方法提取人正常胰腺组织、胰腺癌和癌旁组织总蛋白质,双向电泳分离蛋白,比较不同组织中蛋白质表达的差异.每份组织均重复电泳3次.结果在同一份组织的3张银染的凝胶蛋白质图谱上均可辨识1 000个左右的蛋白质点,蛋白质点在形状、位置和密度上一致,重复性好.在不同组织之间发现了22个明显差异的蛋白质点,初步确定了其等电点和分子量.正常胰腺组织有一个高表达的蛋白质点,癌和癌旁组织有21个高表达的蛋白质点.结论正常胰腺组织、胰腺癌、癌旁组织蛋白质组二维图谱存在明显的差异,部分差异蛋白质可能具有诊断应用价值.     

Ultrastructural differentiation and CEA expression of butyrate-treated human pancreatic carcinoma cells

The effects of butyrate (a biological response modifier) on cellular morphologic features and carcinoembryonic antigen (CEA) expression of human pancreatic carcinoma cells were studied and compared in a well-differentiated, CEA-producing cell line (CAPAN-1), and a poorly differentiated cell line (PANC-1). Butyrate treatment resulted in the acquisition of phenotypic traits commonly attributed to increased "differentiation," including a twofold increase in doubling time, decreased saturation densities, and approximately 50% reduction in colony forming efficiency in both cell lines. Elongation and flattening of cells with extending cellular processes were seen by light microscopy. Significant ultrastructural changes were seen only in the PANC-1 cells, including an increased number of intercellular desmosomes, tonofilaments, and lipid droplets. In contrast, to the coarsely clumped nuclear chromatin (heterochromatin) of untreated PANC-1 cells, the nuclei of the butyrate-treated cells consisted of finely dispersed chromatin (euchromatin). CAPAN-1 cells responded to butyrate with increased CEA synthesis and release. This effect was greatest in the stationary growth phase. Butyrate had no effect on the already low rate of CEA synthesis by PANC-1 cells. These studies suggest that CEA synthesis and state of differentiation are affected independently by butyrate treatment and that the original tumor phenotype plays an important role in response to such treatment.     

晚期糖基化终产物受体在肝细胞癌患者中的表达

目的 研究晚期糖基化终产物受体(RAGE)在原发性肝细胞癌(PHC)中的表达.方法 收集10例PHC患者的肝癌组织、癌旁组织、血清及正常人的血清;用RT-PCR及Western blot分别检测组织中RAGE基因及蛋白的表达;用ELISA检测血清RAGE表达,同时用化学发光法检测血清中甲胎蛋白(AFP)表达.结果 RA...     

MK615 decreases RAGE expression and inhibits TAGE-induced proliferation in hepatocellular carcinoma cells

AIM:To investigate the proliferative effect of advanced glycation end-products(AGEs) and the role of their cellular receptor(RAGE) on hepatocellular carcinoma(HCC) cells,and the inhibitory effects of MK615,an extract from Japanese apricot,against AGEs were also evaluated.METHODS:Two HCC cell lines,HuH7 and HepG2,were used.Expression of RAGE was investigated by poly-merase chain reaction,Western blotting,and flow cytemetry(FACS).The effect of MK615 on RAGE expression was also evaluated by FACS.The proliferat...     

血小板衍化内皮细胞生长因子在胰腺癌组织中的表达及其临床意义

目的:观察血小板衍化内皮细胞生长因子(PD-ECGF)在胰腺癌组织中的表达,探讨其临床意义.方法:利用Elivison免疫组织化学法检测36例手术切除的胰腺癌及21例癌旁正常胰腺组织,以及胃癌、食管癌、肝癌、结肠癌、肺癌及乳腺癌组织各10例中PD-ECGF的表达.分析PD-ECGF与胰腺癌大小、分化程度及淋巴结转移的相关性.结果:PD-ECGF在胰腺癌组织中的表达阳性率显著高于在癌旁正常胰腺组织(88.9% vs 28.6%,P<0.01).PD-ECGF在结肠癌、肝癌、乳腺癌、食管癌、胃癌及肺癌组织中的表达阳性率为60%、70%、80%、90%、90%及80%.Ⅱ-Ⅳ期胰腺癌组织中PD-ECGF的表达阳性率显著高于Ⅰ期胰腺癌组织(100% vs 75%,P<0.05).结论:PD-ECGF为一种非特异性的肿瘤相关因子,其可能与胰腺癌的病程进展有关.     

大鼠胰腺上皮内瘤变和胰腺癌模型血清蛋白质谱的差异表达

目的:利用表面增强激光解析离子化飞行时间质谱(SELDI-TOF MS)技术,分析由7,12-二甲基苯并蒽(DMBA)诱导建立的大鼠胰腺上皮内瘤变(PanIN)和胰腺癌(PC)模型血清蛋白质谱的差异表达.方法:40只♂清洁级SD大鼠为模型组,DMBA胰腺局部种植建立PanIN和PC模型,根据PanIN标准进行病理学分级.26只♂清洁级SD大鼠为正常对照组.采用SELDI-TOF MS和铜离子鳌合芯片(IMAC-Cu2 芯片)检测大鼠血清蛋白质谱, Biomarker Wizard 3.0软件分析比较对照组、PanIN组和PC组之间的差异表达蛋白.结果:DMBA胰腺局部种植后共获得PC 11例,PanIN 18例.与对照组比较, PanIN组和PC组表达强度显著上调(P<0.001)的蛋白质峰有19个,显著下调(P<0.001)的蛋白质峰有11个;其中质荷比分别为5835.2、4087.3、4786.5、4800.5、3932.2、5765.9、5924.8、5001.9、3913.7的9个蛋白质峰表达强度在对照组、PanIN组和PC组呈逐级递增趋势, 质荷比分别为1096.9、1478.9、8572.9、1007.1的4个蛋白质峰表达强度呈逐级递减趋势.结论:与正常大鼠比较,PanIN和PC模型大鼠血清蛋白质谱表达发生显著变化,这些差异表达蛋白质在胰腺癌中的作用值得进一步深入研究.     

Differential effects of nucleoside analogs on oxidative phosphorylation in human pancreatic cells

Although nucleoside analogs as a group inhibit mtDNA replication, individually they target specific organs for toxicity. For example, dideoxyinosine(ddI) is most closely associated with clinical pancreatitis and dideoxycytosine (ddC) with peripheral neuropathy. Comparison of the differential effects of these analogs on mitochondrial function in relevant human cell lines could provide general clues as to the mechanisms of their differential toxicity. We compared the effects of ddI [and its intracellular metabolite dideoxyadenosine (ddA)], with other nucleoside analogs ddC, Azidothymidine (AZT) and didehydrodeoxythymidine(d4T) on mtDNA elongation, cytotoxicity, oxidative phosphorylation, and cellular ATP concentration in a human pancreatic cell line, Capan-1 cells. AZT, like all the other analogs tested, altered mtDNA elongation, but had no other effect on these cells. Both ddC and d4T, but not ddI (20 m and 50 M), reduced total dish protein (a measure of cell numbers) in cells grown to confluence. The effect of ddA was intermediate. All (except AZT) increased lactate concentration in the cell culture medium. Dideoxycytosine (ddC) and d4T did not significantly affect cell oxygen consumption, expressed as a fraction of total dish protein. By contrast, ddI and ddA reduced basal and/or FCCP-stimulated oxygen consumption. %Dideoxycytosine (ddC) but not ddI or ddA (50 M) was cytotoxic to cells after six days of growth. Nevertheless, the ATP content (expressed as a fraction of surviving cells) for ddC-, ddI-, and ddA-treated cells was similar to control cells. Cytotoxicity was apparent for ddI, ddA, as well as ddC after seven days. Paradoxically, cell ATP content was now significantly higher than control cells. Electron microscopy of cells treated with ddI confirmed significant ultrastructural changes affecting the inner mitochondria membrane and cristae. In conclusion, these data suggest that nucleoside analogs uniformly induce damage to mtDNA. However, the mitochondrial phenotypic damage induced by ddI and ddA appear to result in less Capan-1 cytotoxicity than ddC and d4T. The link between these differential effects and ddI pancreatitis is unclear.     

Differential expression of cell cycle proteins during ageing of pancreatic islet cells

One of the major challenges for developmental biologists and investigators in the field of diabetes over the last few decades has been to dissect the origin of pancreatic endocrine cells and to accurately understand the mechanisms that regulate islet cell regeneration. While significant advances have been made recently, there continues to be a paucity of knowledge regarding the growth factor signalling pathways that directly regulate the proteins involved in islet cell cycle control. We will discuss recent work in these areas and provide insights from our studies into age-dependent alterations in the expression of growth factor signalling proteins and cell cycle proteins in islet cells.     

LY294002联合吉西他滨对胰腺癌PANC-1细胞p-Akt和MRP表达的影响

目的:研究LY294002联合吉西他滨对体外培养的人胰腺癌PANC-1细胞内p-Akt和MRP表达的作用.方法:半定量RT-PCR和Western blot检测不同浓度的LY294002联合吉西他滨用药后PANC-1细胞内MRP mRNA以及p-Akt和MRP蛋白表达水平的变化.结果:LY294002联合吉西他滨能显著抑制PANC-1细胞内MRP mRNA的表达(1.47±0.03,1.31±0.05,1.02±0.04,0.76±0.06,0.37±0.02,P<0.05),亦显著抑制p-Akt和MRP蛋白的表达,并且这种抑制作用与药物浓度显著相关(p-Akt:0.80±0.02,0.63±0.01,0.52±0.01,0.41±0.02,0.35±0.02,P<0.05;MRP:0.93±0.05,0.87±0.03,0.81±0.03,0.71±0.02,0.40±0.03,P<0.05),在其浓度最大组抑制效应达到最大.结论:LY294002可能通过抑制PI3K/Akt信号途径抑制MRP mRNA和蛋白的表达,逆转肿瘤的耐药.     

Differential gene expression in human hepatocellular carcinoma Hep3B cells induced by apoptosis-related gene BNIPL-2

AIM:Bd-2/adenovirus E1B 19 ku interacting protein 2-like(BNIPL-2) is a novel protein recently identified in ourlaboratory.BNIPL-2 is homologous to human BNIP-2,apotentially proapoptotic protein,and can interact with Bcl-2 and Cdc42GAP and promote apoptosis in BEL-7402 cells.Here we report the gene-expression profile regulated byBNIPL-2 in human hepatocarcinoma Hep3B cells and theanalysis of its potential roles in cell apoptosis.METHODS:BNIPL-2 was overexpressed in Hep3B cellsusing tetracycline inducible or Tet-on system.Screened byWestern blot,the cells with low background and highinduction fold of BNIPL-2 were obtained.We performedAtlas human cDNA expression array hybridization on thesecells and analyzed the data with Quantarray~(?) software toidentify BNIPL-2-regulated genes and their expressionprofile.RT-PCR was used to confirm the altered expressionlevel of part of genes identified by the Atlas array hybridization.RESULTS:Fifteen of 588 genes spotted on the Atlasmembrane showed altered expression levels in BNIPL-2-transfected Hep3B-Tet-on cells,in which 8 genes involvedin cell apoptosis or growth inhibition were up-regulatedand 7 genes involved in cellular proliferation were down-regulated following overexpression of BNIPL-2.CONCLUSION:cDNA array is a powerful tool to exploregene expression profiles under inducible conditions.Thedata obtained using the cDNA expression microarraytechnology indicates that BNIPL-2 may play its roles inapoptosis through regulating the expression of genesassociated with cell apoptosis,growth inhibition and cellproliferation.     

Differential gene expression in human hepatocellular carcinoma Hep3B cells induced by apoptosis-related gene BNIPL-2

AIM: Bcl-2/adenovirus E1B 19 ku interacting protein 2-like (BNIPL-2) is a novel protein recently identified in our laboratory. BNIPL-2 is homologous to human BNIP-2, a potentially proapoptotic protein, and can interact with Bcl-2 and Cdc42GAP and promote apoptosis in BEL-7402 cells. Here we report the gene-expression profile regulated by BNIPL-2 in human hepatocarcinoma Hep3B cells and the analysis of its potential roles in cell apoptosis. METHODS: BNIPL-2 was overexpressed in Hep3B cells using tetracycline inducible or Tet-on system. Screened by Western blot, the cells with low background and high induction fold of BNIPL-2 were obtained. We performed Atlas human cDNA expression array hybridization on these cells and analyzed the data with Quantarray software to identify BNIPL-2-regulated genes and their expression profile. RT-PCR was used to confirm the altered expression level of part of genes identified by the Atlas array hybridization. RESULTS: Fifteen of 588 genes spotted on the Atlas membrane showed altered expression levels in BNIPL-2-transfected Hep3B-Tet-on cells, in which 8 genes involved in cell apoptosis or growth inhibition were up-regulated and 7 genes involved in cellular proliferation were down-regulated following overexpression of BNIPL-2. CONCLUSION: cDNA array is a powerful tool to explore gene expression profiles under inducible conditions. The data obtained using the cDNA expression microarray technology indicates that BNIPL-2 may play its roles in apoptosis through regulating the expression of genes associated with cell apoptosis, growth inhibition and cell proliferation.     

Expression of receptor for advanced glycation end products (RAGE) and MMP-9 in human pancreatic cancer cells

BACKGROUND/AIMS: Amphoterin is considered as a regulator for the ability of invasion and migration in tumor cells and embryonic neurons through binding to receptor for advanced glycation end products (RAGE), a multiligand cell surface molecule of the immunoglobulin superfamily. As matrix metalloproteinase-9 (MMP-9, gelatinase B) has been reported to play a critical role in tumor progression and metastasis, we have examined the relation of RAGE and MMP in human pancreatic cancer. METHODOLOGY: Three representative human pancreatic carcinoma cells were rendered for the study which show different metastatic potential, PANC-1 and MIA PaCa-2 as the cells with high ability, BxPC-3 as with low. The expression of RAGE was examined by RT-PCR. The expression of MMP-9 protein was examined by Western blotting. RESULTS: RAGE was strongly expressed in MIA PaCa-2 and PANC-1 that have high metastatic ability. On the contrary, RAGE was expressed little in BxPC-3 that has low ability. Similarly, expression of MMP-9 showed almost the same tendency. RAGE and MMP-9 are expressed concordant with the metastatic ability of the human pancreatic cancer cells. CONCLUSIONS: Control of these molecules could be a key to regulating the metastatic ability of pancreatic cancer and this may be exploited in targeted therapy of this cancer.     

Experimental pancreatic carcinoma as a model of human pancreatic carcinoma

Experimental pancreatic carcinoma induced by BHP in hamsters and by DMBA in rats are compared with human pancreatic carcinoma. Tubular adenocarcinoma and papillary adenocarcinoma in human pancreas and hamster pancreas appeared to be similar. However, poorly differentiated adenocarcinoma and acinar cell carcinoma were not observed in the experimental hamster model. Poorly differentiated adenocarcinoma and acinar cell carcinoma in human and rat pancreas were similar but experimentally induced papillary adenocarcinoma and cystoadenocarcinoma were not observed in the rat. Mucin producing cells were usually observed in human and experimental pancreatic carcinoma in the hamster but were rarely noted in the rat. These results indicate that experimental pancreatic carcinoma either in rats or hamsters is a good model for the understanding of both the histogenesis and the carcinogenesis of human pancreatic carcinoma.     

Differential expression of the amv gene in human hematopoietic cells.

Total cellular RNAs from a variety of fresh and culture-derived human hematopoietic neoplastic cell types at various stages of differentiation and human sarcoma, carcinoma, melanoma, and glioblastoma cell lines were enriched for poly(A)- containing sequences, fractionated by gel electrophoresis, and blot hybridized to a cloned DNA probe containing the transforming sequences (v-amv) of avian myeloblastosis virus (AMV), a virus known to cause myeloid leukemias in chickens. Expression of RNA sequences homologous to AMV was detected in all immature myeloid and lymphoid T cells in addition to the single erythroid cell line examined, but not in mature T cells or in B cells, including lymphoblast cell lines derived from patients with Burkitt lymphoma. In addition, induction of the cell line HL60, a promyelocytic leukemia line, to differentiate with dimethyl sulfoxide or retinoic acid resulted in a reduction of the level of expression of the human cellular gene c-amv homologous to v-amv. There was no detectable c-amv mRNA in any of the solid tumor cell lines examined. Thus, expression of the human c-amv gene could be correlated with the stage of differentiation of different hematopoietic cell types determined by morphologic and marker studies. Expression of c-amv could not be correlated with the extent of methylation in HL60 and in HL60 induced to differentiate with dimethyl sulfoxide.     

Identification and characterization of muscarinic receptors in cultured human pancreatic carcinoma cells

Five human pancreatic carcinoma cell lines were screened for the presence of muscarinic cholinergic receptors (mAChRs), using [3H]N-methylscopolamine ([3H]NMS). T3M4 and COLO-357 cells exhibited specific, high-affinity binding to mAChRs. A small amount of [3H]NMS also bound in PANCI and ASPC-I cells, but not in MIA PaCa-2 cells. Atropine, pirenzepine (PZ), and 11-[[2-[(diethylamino) methyly]-1-piperidinyl] acetyl]-5, 11-dihydro-6H-pyrido-[2, 3-b] [1, 4] benzodiazepine-6-one (AF-DX 116) inhibited [3H]NMS binding and carbachol-mediated [3H]inositol monophosphate formation in both T3M4 and COLO-357 cells. The order of inhibition was: atropine greater than PZ greater than AF-DX 116. Carbachol did not alter [3H]inositol monophosphate formation in the other cell lines. These findings suggest that the mAChRs expressed in some human pancreatic cancer cells exhibit the pharmacologic characteristics of a muscarinic receptor subtype with an intermediate affinity for PZ and a lower affinity for AF-DX 116 and are functionally coupled to activation of phospholipid hydrolysis.     

Differential expression and action of Toll-like receptors in human adrenocortical cells

During sepsis, an intact adrenal gland glucocorticoid stress response is critical for survival. Recently, we have shown that Toll-like receptors, particularly TLR2 and TLR4, are crucial in HPA axis regulation following inflammation, establishing a direct link between bacterial and viral ligands and the endocrine stress response. However, the exact role which TLRs play in adrenal homeostasis and malfunction is not yet sufficiently known. Using quantitative real-time PCR, confocal microscopy and the NF-κB reporter gene assay, we aimed to analyse both, expression and function of all relevant TLRs in the human adrenocortical cell line—NCI-H295R and adrenal cells in primary culture. Our results demonstrate a differential expression pattern of TLR1–9 in human adrenocortical cells as compared to immune cells and adrenocortical cancer cells. Consequently, activation of these cells by bacterial ligands leads to differential induction of cytokines including IL6, IL8 and TNF-α. Therefore, Toll-like receptors expression and function is a novel feature of the adrenal stress system contributing to adrenal tissue homeostasis, regeneration and tumorigenesis.     

凋亡抑制蛋白XIAP在胰腺癌组织中的表达及意义

目的 通过检测XIAP在胰腺癌中的表达,探讨其对胰腺癌发生、发展的可能作用及临床意义.方法 应用免疫组化SP法检测10例正常胰腺组织、14例胰腺良性病变和42例胰腺癌组织中XIAP的表达,并分析其与胰腺癌临床病理参数的关系.结果 XIAP在正常胰腺组织、胰腺良性病变及胰腺癌中均有表达.正常胰腺组织中,高表达占20%,良性病变中占21.4%,而胰腺癌中,高表达占59.5%,较前二者均有显著性差异(P ＜ 0.05).42例胰腺癌组织中,XIAP表达强度与性别、年龄、肿瘤部位、肿瘤大小、分化程度、临床分期及淋巴结转移均无显著相关.结论 XIAP在正常胰腺组织、胰腺的良性病变及胰腺癌中均有表达,但在胰腺癌中呈高表达,表明XIAP在胰腺癌的发生中起到一定作用;XIAP的表达与胰腺癌各项临床病理参数无相关性.