罗非昔布对胰腺癌 BXPC-3细胞裸鼠移植瘤血管形成的影响

背景与目的:肿瘤血管形成在肿瘤细胞浸润性生长中起了重要作用,增加表达的环氧合酶-2与快速生长肿瘤中的血管形成有关.本研究目的是观察选择性环氧合酶-2抑制剂-罗非昔布抑制胰腺癌生长的体内效应,以及对胰腺癌移植瘤相关血管形成的影响.方法:将表达有环氧合酶-2的人胰腺癌细胞 BXPC-3种植入裸鼠皮下,形成移植瘤.经口灌入罗非昔布 30 mg· (kg· d)- 1,共 8周,记录肿瘤大小,采用Ⅷ因子免疫组化显示血管密度, BXPC-3细胞上的血管内皮生长因子 (VEGF)检测亦用免疫细胞化学染色.采用 RT-PCR和明胶酶法检测胰腺癌基质金属蛋白酶-2(MMP-2)mRNA的表达及酶活性的变化.结果:罗非昔布对裸鼠胰腺癌移植瘤的肿瘤重量抑瘤率为 73.64%,肿瘤体积抑瘤率为 87.74%.实验组胰腺癌组织微血管密度 [(1.5± 0.2)个 /200倍放大视野 ]明显低于对照组 [(4.7± 1.5)个 /200倍放大视野 ].与对照组比较,罗非昔布显著降低了胰腺癌 BXPC-3细胞中 VEGF、 MMP-2 mRNA的表达以及酶活性.结论:减少胰腺癌相关的血管形成是罗非昔布阻止胰腺癌生长的机制之一.     

奥曲肽对胃癌细胞转录活化蛋白-1的抑制作用

背景与目的:生长抑素是一种多功能神经肽,其本身及其类似物能抑制多种内分泌肿瘤及某些胃肠肿瘤的生长,但是否能抑制胃癌的生长并不清楚。因此,本研究拟探讨生长抑素类似物奥曲肽对胃癌细胞生长的影响及其机制。方法:不同浓度的奥曲肽作用于人胃癌SGC-7901细胞24h后,用免疫印迹法检测奥曲肽对SGC-7901细胞中c-Fos、ERK蛋白表达的影响。建立人胃癌细胞裸鼠原位移植瘤模型,奥曲肽治疗8周,观察奥曲肽对胃癌生长的影响;免疫组化法检测裸鼠人胃癌组织c-Fos和ERK表达的变化。在此基础上,采用EMSA技术检测奥曲肽对胃癌细胞AP-1活化的影响。结果:1×10-5mol/L奥曲肽可明显降低胃癌细胞的3H-TdR的掺入;奥曲肽能抑制人胃癌细胞裸鼠原位移植瘤的生长,抑瘤率为62.3%。SGC-7901胃癌细胞经不同浓度奥曲肽作用后,其c-Fos和ERK的表达水平均有不同程度下降;组织标本检测亦有类似变化。EMSA分析显示,胃癌细胞有基础水平的AP-1活化,20%血清能刺激AP-1的结合力,奥曲肽能抑制这种刺激作用。结论:奥曲肽通过抑制胃癌c-Fos、ERK蛋白的表达而抑制AP-1的结合活性,从而抑制胃癌的生长。     

塞来昔布联合顺铂对人舌鳞癌Tca8113细胞移植瘤的生长抑制作用

目的:通过动物实验观察COX-2抑制剂塞菜昔布与顺铂联合应用后对人舌鳞癌Tca8113细胞移植瘤的生长抑制作用.方法:将Tca8113细胞接种于裸鼠皮下,分别给予塞来昔布、顺铂及两者联合应用,35 d后处死裸鼠,测移植瘤质量,计算抑瘤率,光镜及电镜观察移植瘤组织学变化,免疫组化染色观察COX-2蛋白的表达,RT-PCR检测COX-2 mRNA表达.结果:COX-2抑制剂塞来昔布除抑制Tca8113细胞移植瘤生长及COX-2蛋白的表达外,还显著增强顺铂对移植瘤的生长抑制作用.塞来昔布组、顺铂组及塞来昔布+顺铂组的抑瘤率分别为15.63%、37.50%和82.81%,与对照组相比差异有统计学意义,P＜0.01;塞来昔布+顺铂组与塞来昔布及顺铂单独用药组相比,其抑制移植瘤生长差异有统计学意义,P＜0.01;塞来昔布对Tca8113细胞COX-2mRNA表达的抑制作用较弱,与对照组相比差异无统计学意义,P=0.073.结论:塞来昔布除了可以抑制裸鼠移植瘤生长外,还可以增强顺铂对Tca8113细胞裸鼠移植瘤生长抑制作用,其作用机制可能与抑制COX-2蛋白的表达有关,为进一步探索抑制COX-2酶的活性与防治头颈肿瘤的作用机制方面,提供了有益的参考.     

肝、胆

AZT对肝癌细胞端粒酶活性及相关蛋白表达的影响；RNA干扰SMYD3基因衰达对诱导肝癌细胞凋亡的影响；血管内皮生长因子和基质金属蛋白酶-2在肝细胞肝癌中的表达殛临床意义；肝细胞生长因子对卵巢癌细胞侵袭的促进作用及信号转导途径；环氧合酶-2抑制剂塞米昔布对人肝癌HepG2裸小鼠移植瘤生长和肿瘤血管生成的抑制作用。     

米非司酮对子宫内膜癌HHUA细胞裸小鼠移植瘤生长作用的研究

目的探讨米非司酮(RU486)对人子宫内膜癌HHUA细胞裸小鼠移植瘤生长的抑制作用。方法体外培养人子宫内膜癌HHUA细胞,裸小鼠皮下接种HHUA细胞建立裸小鼠移植瘤动物模型,将荷瘤裸小鼠随机分为两组,分别应用米非司酮、精制花生油治疗4周。治疗期间观察各组裸小鼠生长情况、移植瘤体积、增殖率和形态,治疗结束后应用流式细胞术(FCM)检测各组裸小鼠移植瘤组织细胞周期时相的变化和细胞凋亡率;应用免疫组化技术检测基因Bcl-2和Bax蛋白的表达。结果米非司酮组裸小鼠移植瘤体积第3周明显减小,肿瘤增殖率明显降低(P<0.01)。4周治疗结束后米非司酮组裸小鼠移植瘤细胞G0/G1期比例升高(38.17±3.02)%,S期细胞比例(SPF)降低(43.87±4.29)%,与对照组比较差异均有显著性(P<0.05),移植瘤细胞凋亡率为(15.03±2.19)%,明显高于对照组(3.06±1.13)%(P<0.01);治疗组移植瘤细胞Bcl-2蛋白表达明显降低(1.9±0.61),而Bax呈现过度表达(4.0±1.28)(P<0.05)。结论抗孕激素米非司酮(RU486)能显著抑制人子宫内膜癌HHUA细胞裸小鼠移植瘤的生长,其抗肿瘤作用与调控细胞周期时相和诱导细胞凋亡相关。     

塞来昔布联合氟尿嘧啶对MFC细胞615小鼠原位移植瘤COX-2、Fas表达的影响及意义

[目的]通过测定在塞来昔布及氟尿嘧啶干预下的小鼠前胃癌细胞(MFC)615小鼠原位移植瘤中环氧合酶(COX-2)、Fas的表达,对COX-2抑制剂联合氟尿嘧啶对胃癌的影响及意义进行探讨.[方法]制备MFC细胞615小鼠原位移植胃癌模型.造模成功后分为塞来昔布组、联合用药组及对照组,其中塞来昔布25mg/kg灌胃,氟尿嘧啶50mg/kg腹腔注射.观察各组小鼠一般情况.干预3周后处死小鼠,计算抑瘤率.SP免疫组织化学法检测各组肿瘤COX-2、Fas表达.[结果]造模成功率100.00％(30/30).联合用药组、塞来昔布组的小鼠一般情况优于对照组.实验结束后3组比较,塞来昔布组、联合用药组比对照组肿瘤体积和瘤重的抑制率均明显升高(P＜0.05).与塞来昔布组相比,联合用药组肿瘤体积更小、瘤重减轻(P＜0.05).COX-2蛋白在塞来昔布组、联合用药组表达阳性率明显低于对照组,而Fas蛋白的表达则明显高于对照组(P＜0.05).[结论]塞来昔布及氟尿嘧啶对胃癌有抑制作用,其机制与调节COX-2、Fas蛋白表达有关.     

TNF-α通过JNK和AP-1途径调节乳腺癌MCF-7细胞VEGF的表达

目的: 探讨肿瘤坏死因子(tumor necrosis factor-α, TNF-α)诱导血管内皮生长因子(vascular endothelial growth factor,VEGF)表达的机制.方法:以20 ng/ml TNF-α处理MCF-7细胞,用Western blotting检测MAPK(JNK,p38,ERK)信号通路中蛋白磷酸化水平的变化以及AP-1家族(c-Jun,Jun-B,c-Fos,Fra-1,Fra-2,JunD)的蛋白表达及磷酸化水平的变化;以免疫共沉淀法检测激活后的AP-1存在形式;以RT-PCR以及Western blotting检测VEGF mRNA和蛋白表达水平;以MAPK抑制剂预处理后,检测VEGF蛋白表达水平;运用ChIP的方法验证p-c-Jun结合在VEGF启动子区.结果:TNF-α通过激活JNK信号转导通路活化AP-1;被TNF-α激活后AP-1以p-c-Jun-c-Jun和p-c-Jun-JunB同源二聚体形式存在;TNF-α通过激活转录因子AP-1促进VEGF的转录,并增强VEGF的蛋白表达水平;p-c-jun通过与VEGF启动子AP-1结合参与对VEGF转录的调控.结论:在TNF-α作用下,AP-1通过p-c-jun同源二聚体结合在VEGF启动子的AP-1结合位点上,直接对VEGF转录进行调控.     

ERRγ对子宫内膜癌细胞移植瘤组织中Akt、ERK表达的影响

目的:以ERα（+）子宫内膜癌细胞株Ishikawa为研究对象,建立子宫内膜癌细胞的裸鼠皮下移植瘤模型,进一步研究在子宫内膜癌细胞移植瘤中ERK、p-ERK、Akt、p-Akt蛋白的表达及其意义。方法:利用转染前后的Ishikawa细胞建立子宫内膜癌细胞裸鼠皮下移植瘤模型,采用免疫组织化学链霉亲和素-生物素过氧化物酶复合物(SABC法)检测裸鼠移植瘤中ERRγ蛋白的表达情况。Western blot方法检测移植瘤组织中Akt、p-Akt、ERK、p-ERK蛋白的表达情况。结果:免疫组化检测Ishikawa 细胞移植瘤组与no-silence Ishikawa细胞移植瘤组（空载体组）ERRγ蛋白表达阳性率无显著性差异（P＞0.05）,Ishikawa 细胞移植瘤组与ERRγ-shRNA Ishikawa细胞移植瘤组阳性率具有显著性差异（P＜0.05）。Western blot方法检测3组组织的活化情况:ERRγ-shRNA Ishikawa细胞移殖瘤组中Akt、p-Akt、ERK、p-ERK表达量均低于Ishikawa 细胞移殖瘤及no-silence Ishikawa细胞移殖瘤组（P＜0.05）。结论:沉默ERRγ基因的瘤组织的ERK及Akt的表达呈现低表达,从而可以推测:通过ERRγ的表达来调节ERK及Akt信号通路的表达,进一步抑制子宫内膜癌Ishikawa细胞增殖,促进其凋亡。     

影响人大肠癌裸小鼠移植瘤生长的因素分析

目的:探索影响人大肠癌移植瘤裸小鼠肿瘤生长的相关因素.方法:12只BALB/C裸小鼠建立人大肠癌原位移植瘤模型,于接种后8周处死.观察记录裸小鼠瘤质重、瘤体积,并采用RT-PCR或免疫组化法检测肿瘤组织AP-1、NF-kb、COX-2、PGE2、VEGF、EGFR,以及STAT3、Bcl-2、Survivin、Caspase9、Caspase8和Caspase3等与细胞凋亡相关的因子表达.采用多元逐步回归模型分析各指标与肿瘤生长(瘤质重和瘤体积)的关系.结果:经多元逐步回归模型分析,瘤体积以及肿瘤组织Caspase3、VEGF、COX-2的表达与瘤质重有关.瘤质重以及肿瘤组织Caspase 8、STAT3、EGFR、VEGF的表达与瘤体积相关.结论:人移植瘤裸小鼠的生长与包括凋亡因子、COX-2、VEGF、EGFR在内的多种因子有密切关系,大肠癌的发生发展是一个多因子、多信号通路参与的过程.     

罗非昔布对裸鼠人胃癌组织中新生血管及血管内皮生长因子表达的影响

目的探讨罗非昔布对裸鼠人胃癌组织中新生血管及血管内皮生长因子（vascular endothelial growthfactor,VEGF）表达的影响。方法建立BALB/C nu/nu裸鼠人胃腺癌SGC-7901细胞株原位种植转移模型,在种植术后1周以罗非昔布0.2 mL,5 mg.kg-1.d-1给裸鼠灌胃,连续8周后,对荷瘤鼠行彩色多普勒探查,观察胃肿瘤新生血管的形态和数量,通过免疫组织化学Envision法及RT-PCR方法检测胃癌组织中的VEGF蛋白及VEGFmRNA的表达。结果各治疗组肿瘤血管体积[罗非昔布组（9.96±3.58）,5-FU组（9.74±2.83）及联合组（8.25±3.14）]与生理盐水对照组（13.10±4.00）比较,差异有统计学意义（P〈0.05）。VEGF蛋白表达及VEGFmRNA检测结果显示,各治疗组与生理盐水对照组比较,差异有统计学意义（P〈0.05）。结论环氧合酶-2抑制剂罗非昔布可以通过降低胃癌肿瘤组织中VEGF蛋白及VEGFmRNA的表达,减少肿瘤血管数量,抑制胃癌细胞的生长和转移。     

Knockdown of c-Fos suppresses the growth of human colon carcinoma cells in athymic mice

Here we have investigated whether inhibition of c-Fos expression in RKO human colon carcinoma cells (HCCCs) would result in reduced TGFβ1 expression and suppression of tumor growth in athymic mice. We stably transfected RKO cells with c-Fos small interfering RNA (siRNA) or with the corresponding control siRNA. Using these stable cell lines, we demonstrated that siRNA-c-Fos significantly suppressed both AP-1 binding, promoter reporter activity at the proximal AP-1 site in the TGFβ1 promoter, and TGFβ1 production. Further, we established colon cancer xenografts with each of RKO-siRNA-EV, RKO-siRNA-Ctrl and RKO-siRNA-c-Fos cells. By 24 days, the tumor size of RKO-siRNA-c-Fos xenografts was 40% that of either RKO-EV or RKO-siRNA-Ctrl. Immunohistochemistry (IHC) of tumor xenografts demonstrated that siRNA-c-Fos significantly blocked c-Fos expression, and consequently expression of TGFβ1. However, expression of TGFβ2 and TGFβ3 were unaffected. Overall, our results demonstrate that blockade of TGFβ1 production by siRNA-c-Fos effectively suppressed tumor growth in vivo.     

Enhanced inhibitive effects of combination of rofecoxib and octreotide on the growth of human gastric cancer

Our previous studies indicated that cyclooxygenase-2 inhibitor or octreotide could suppress the proliferation of gastric adenocarcinoma in vitro or in vivo. The present study was aimed to find whether rofecoxib combined with octreotide could enhance the inhibitive effects on the growth of gastric cancer. The effect of rofecoxib or octreotide on proliferation of gastric cancer cell line was determined by 3H-thymidine ribotide incorporation. The TdT-mediated dUTP nick end-labeling assay was used to detect the apopotosis. To determine their synergic antineoplastic effects, the interaction between rofecoxib and octreotide on SGC-7901 cell was evaluated by the median effect plot. After orthotopical implantion of xenografts of human gastric cancer in stomach, nude mice were given rofecoxib plus octreotide for 8 weeks. Cyclooxygenase-2 in gastric cancer tissues was measured by immunohistochemistry. Combination of rofecoxib and octreotide presented synergistic effect (combination index < 1) in the majority of responses. The inhibitory rate for xenografts in nude mice was 89.7% in rofecoxib group. Combination of rofecoxib and octreotide enhanced inhibitory rate to 98.8%. The combination greatly increased the apoptotic index (78.20% +/- 6.45%) of the xenografts as compared with that of using rofecoxib alone (46.60% +/- 3.42%); the difference was very significant (p < 0.001). Rofecoxib could inhibit the activity of cyclooxygenase-2 in the tissue of gastric adenocarcinomas of nude mice. Our results indicate that combination of rofecoxib and octreotide significantly enhances the antiproliferative effect in gastric adenocarcinoma, which might have potential therapeutic value.     

Psoriasin interacts with Jab1 and influences breast cancer progression

Psoriasin (S100A7) is expressed at low levels in normal breast epithelial cells but is highly expressed in preinvasive ductal carcinoma in situ. Persistent psoriasin expression occurs in some invasive carcinomas and is associated with poor prognostic factors. Whereas there is evidence that secreted psoriasin can act as a chemotactic factor for CD-4-positive lymphocytes in psoriatic skin lesions, an intracellular biological function is unknown. We have found that psoriasin physically interacts with Jab1 (c-jun activation-domain binding protein 1) in the yeast two-hybrid assay and confirmed this by coimmunoprecipitation assay in breast cancer cells. Psoriasin-transfected breast cancer cells showed increased nuclear Jab1 and demonstrated several features consistent with an alteration in Jab1 activity including an increase in activator protein-1 (AP-1) activity, increased expression of AP-1 and HIF-1-dependent genes, and reduced expression of the cell-cycle inhibitor p27(Kip1). Psoriasin overexpression was also associated with alteration of cellular functions that are associated with increased malignancy, including increased growth, decreased adhesion, and increased invasiveness in vitro, as well as increased tumorigenicity in vivo in nude mice. We conclude that intracellular psoriasin influences breast cancer progression and that this may occur through stimulation of Jab1 activity.     

Tumor-necrosis-factor and cell-death in tumors (review)

The prognosis of pancreatic cystadenocarcinoma is known to be relatively favorable, unlike poor prognosis of the majority of pancreatic cancers. However, little is known about the pathogenesis and mode of progression of this cancer. We report on a newly established pancreatic cystadenocarcinoma cell line, OCUP-1. Its characteristics were compared to those of cells derived from pancreatic ductal cell carcinoma. OCUP-1 was established by successive culture of cancer cells obtained during an operation for pancreatic cystadenocarcinoma. This cell line demonstrated mono-layered proliferation and a doubling time of 80.3 h, which was 2.1 to 5.3 times longer than that of six ductal cell carcinoma-derived cell lines. Cell cycle time agreed with the doubling time. Tumors could be produced in all nude mice by inoculating cells from the 6 different ductal cell carcinoma derived cell lines without pre-treatment. However, with OCUP-1 cell line inoculation, the nude mice had to be pre-treated with asialo-GM1 - an inhibitor of natural killer cell activity - for tumors to be produced. Although the newly established OCUP-1 cell line demonstrated low proliferative activity, its genetic characteristics (point mutation at codon 12) were similar to those of other cells derived from ductal cell carcinomas. The OCUP-1 cell line may be used to investigate the pathogenesis and progression of pancreatic cystadenocarcinomas.