RNAi沉默整合素连接激酶基因表达对结肠癌细胞HT-29增殖的影响

目的:利用RNA干扰(RNA interference,RNAi)技术,阻断结肠癌细胞系HT-29中整合素连接激酶(integrin-linked kinase,ILK)基因的表达,研究ILK基因沉默后对HT-29细胞系增殖产生的影响。方法:构建针对ILK基因的真核表达质粒pSUPER-neo-EGFP-ILK(pSNE-ILK),在脂质体介导下稳定转染人结肠癌细胞系HT-29细胞(HT-29/pSNE-ILK组),同时以无关质粒pSUPER-neo-EGFP-C(pSNE-C)转染HT-29细胞(HT-29/pSNE-C组)和未转染细胞(HT-29组)作为对照,分别应用RT-PCR技术及蛋白印迹法检测ILK mRNA及蛋白表达水平,四甲基偶氮唑盐(MTT)比色法检测细胞增殖状况。结果:稳定转染后,HT-29/pSNE-ILK组ILK mRNA及蛋白表达水平分别为0.16±0.11和0.24±0.07,HT-29/pSNE-C组分别为0.53±0.10和0.56±0.08,HT-29组分别为0.64±0.11和0.77±0.02,前组ILK mRNA及蛋白表达水平分别与后两组比较,差异均有统计学意义(P<0.05);稳定转染pSNE-ILK质粒后HT-29细胞中ILK mRNA及蛋白表达抑制率分别为69.8%和57.1%。MTT比色法检测显示,HT-29/pSNE-ILK组细胞增殖率明显下降(P<0.05)。结论:RNA干扰技术能有效抑制靶基因ILK的表达,进而可抑制结肠癌细胞系HT-29的增殖。     

Id1对结肠癌HT-29细胞VEGF表达及增殖的影响

目的探讨分化抑制因子１(inhibitor of differentiation-1,Id1)对结肠癌HT-29细胞VEGF表达的影响及Id1在结肠癌细胞增殖中的作用。方法HT-29转染Id1表达质粒及Id-1干扰序列后,采用RT-PCR方法和Western blot法检测VEGF mRNA和蛋白的表达,MTT法检测各组细胞增殖情况。结果 Id1过表达能显著上调VEGF mRNA和蛋白表达(P<0.01),促进细胞的生长(P<0.01);Id1抑制后能显著下调VEGFmRNA和蛋白表达(P<0.01),减缓细胞的生长(P<0.01)。结论在结肠癌HT-29细胞中Id1为VEGF的上游基因,Id1可通过正性调控VEGF影响结肠癌细胞增殖。     

CDX2基因通过NF-κB信号通路抑制结肠癌细胞增殖

目的 观察过表达CDX2基因对人结肠癌细胞株增殖影响,并初步探讨其作用机制。方法 通过基因转染技术将CDX2基因转染入结肠癌HT-29细胞株,经G418筛选后获得稳定转染细胞株,RT-PCR和Western blot检测CDX2基因表达。采用四甲基偶氮唑盐(MTT)法、流式细胞仪分析转染细胞的生物学行为。Western blot检测CDX2过表达对HT-29细胞内Cyclin D1和NF-κB磷酸化水平的影响。结果 经脂质体转染和筛选,建立了稳定过表达CDX2人结肠癌HT-29细胞株。转染CDX2组与未转染组和空白质粒组相比,细胞的生长速度减慢(P＜0.05),细胞周期中G₁/G₀期比例增加(P＜0.05),而S期比例则减少(P＜0.05);与未转染组和空白质粒组比较,转染CDX2基因组HT-29细胞的Cyclin D1和p-NF-κB活性明显降低。结论 CDX2基因可能通过抑制NF-κB通路抑制Cyclin D1活性,从而抑制人结肠癌HT-29细胞增殖。     

RNAi沉默CDX2基因对急性髓细胞白血病细胞增殖影响的实验研究

目的:探讨RNA干扰技术沉默CDX2基因对人急性髓细胞白血病细胞系THP1增殖影响的意义.方法:针对CDX2mRNA序列设计合成2对编码小干扰RNA(siRNA)的DNA模板,构建pGenSi1-CDX2siRNA重组质粒,转染THP1细胞.采用蛋白质印迹法检测重组质粒对CDX2蛋白表达的影响,MTT法观察THP1细胞增殖情况,Annexin-V-PI双染法流式细胞术检测转染重组质粒后细胞凋亡状况.结果:成功构建pGenSil-CDX2 siRNA重组质粒,成功转染THP1细胞,并能特异地抑制CDX2基因的表达;转染重组质粒后,THP1的活力分别降低为(39.8±5.0)%,与空质粒组组间差异有统计学意义,P=0.016.重组质粒组的THP1细胞凋亡率为37.3%～42.1%.结论:pGenSil-CDX2 siRNA重组质粒明显下调XDX2在急性髓细胞白血病细胞中的表达,并抑制肿瘤细胞生长,促进其凋亡.     

阻断肝再生增强因子的表达对人肝癌细胞株HepG2增殖的抑制作用

背景与目的:前期研究中已发现人肝再生增强因子(humanaugmenterofliverregeneration,hALR)在肝癌细胞中呈高表达,在正常肝细胞中几乎不表达,并在体外能刺激肝癌细胞株增殖,而对正常肝细胞无影响。本研究通过运用hALR的siRNA和hALR单克隆抗体阻断人肝癌细胞株HepG2表达,探讨hALR对细胞增殖的影响。方法:设计、构建siRNA表达质粒pSIALR-A及其阴性对照质粒pSIALR-B。将构建的pSIALR-A和pSIALR-B分别转染HepG2细胞。荧光显微镜观察绿色荧光蛋白的表达,以计算转染效率。转染48h后,免疫细胞化学及RT-PCR法检测HepG2细胞中hALR的表达。用3H-TdR掺入法检测hALR的siRNA和特异性单克隆抗体对HepG2细胞增殖的影响。结果:HepG2细胞中有hALR的表达。成功构建了针对hALR基因编码区的siRNA表达质粒pSIALR-A。转染入细胞后发现,pSIALR-A能明显抑制hALR的表达(mRNA的表达量减少83%),而随机序列的siRNA却无此作用。hALR的siRNA在体外能显著抑制人肝癌细胞株HepG2增殖,分别转染质粒pSIALR-A和pSIALR-B后HepG2细胞的cpm值为67687±6548和104807±5713(P<0.05)。抗hALR单克隆抗体特异性地阻断hALR的作用后,可部分抑制肿瘤细胞自主性生长(P<0.05)。结论:HepG2细胞高表达hALR;针对hALR的siRNA能显著、特异性地抑制hALR表达,进而抑制HepG2细胞的生长。抗hALR单克隆抗体也能部分抑制HepG2细胞自主性生长。     

NDRG 1基因转染对HT-29人结肠癌细胞株增殖、侵袭和迁移能力的影响

目的研究NDRG1基因转染对HT-29人结肠癌细胞株体外增殖、侵袭和迁移能力的影响。方法将重组真核表达质粒pEGFP-NDRG1-N3和空载体pEGFP-N3采用阳离子脂质体Lipofectamine转染HT-29人结肠癌细胞株,通过荧光倒置显微镜检测绿色荧光蛋白的表达。免疫组织化学染色检测转染前后HT-29细胞NDRG1的表达。并通过多种体外实验（流式细胞术法、24-transwell法、扫描电镜和透射电镜等）研究NDRG1对HT-29人结肠癌细胞株体外增殖、侵袭和迁移能力及表面和超微结构的影响。结果流式细胞术结果显示,与未转染组和转染pEGFP-N3空载体组比较,pEGFP-NDRG1-N3组细胞生长周期G0/G1期比例增高,S期明显减低,差异有统计学意义（P〈0.05）。细胞侵袭和迁移实验的结果显示,转染pEGFP-NDRG1-N3组细胞与空载体和空白对照组比较,穿过微孔膜的细胞数均明显减少,差异有统计学意义（P〈0.05）。扫描和透射电镜结果显示,转染pEGFP-NDRG1-N3组HT-29细胞生长增殖受抑制,细胞分化程度有增高趋势。结论 NDRG1基因可明显抑制HT-29人结肠癌细胞的体外增殖活性,降低其侵袭和迁移能力,促进其分化,提示其可作为一种候选的肿瘤转移抑制基因。     

siRNA干扰HIF-1α抑制人外周血内皮祖细胞分化

背景与目的:低氧诱导因子鄄1α(hypoxiainduciblefactor鄄1alpha,HIF鄄1α)在促血管新生中起关键作用,通过基因沉默方法研究促肿瘤发展因子对细胞分化的影响可为体内抗癌治疗提供指导。本研究构建了HIF鄄1α的RNA干扰(RNAi)质粒,在体外研究其对人外周血内皮祖细胞(endothelialprogenitorcells,EPCs)向血管内皮细胞(endothelialcell,EC)分化的影响。方法:质粒重组技术构建siRNA鄄HIF鄄1α质粒,电穿孔转染质粒,计算转染效率;RT鄄PCR法检测HIF鄄1α、HIF鄄1β及血管内皮生长因子(vascularendothelialgrowthfactor,VEGF)mRNA在质粒转染前后含量变化;免疫组化方法检测在常氧/低氧下和siRNA鄄HIF鄄1α质粒转染后12hHIF鄄1α蛋白表达情况;流式细胞术测定FITC鄄CD31 的EPCs/EC在各组细胞转染3～10天后的组间差异;一氧化氮酶法鉴定各组细胞VEGF依赖性一氧化氮合酶(endothelialnitricoxidesynthase,eNOS)含量;镜下观察转染前后细胞形态及分化程度。结果:重组子质粒测序鉴定证实siRNA鄄HIF鄄1α构建与设计相符,电转效率约20%;转染6h各组HIF鄄1αmRNA在常氧中及低氧中均表达,并可被siRNA鄄HIF鄄1α不同程度抑制(P<0.05),相应的下游靶基因VEGF表达下调(P<0.05),HIF鄄1β表达在各组中无显著性差异(P>0.05)。免疫组化示常氧及低氧1hHIF鄄1α蛋白无表达,低氧3h成倍出现,6h达高峰,24h降至基线水平;siRNA鄄HIF1α抑制后低氧12h,HIF鄄1α蛋白表达被显著抑制。流式细胞仪检测发现,经siRNA鄄HIF鄄1α抑制后CD31 细胞较对照组在常氧及低氧中表达下调(P<0.05)。siRNA减少eNOS含量,并与培养时间及VEGF刺激浓度呈反比(P<0.01)。细胞形态学观察示低氧可诱导并加速EPCs向EC定向分化、扩增,但可被siRNA鄄HIF鄄1α抑制。结论:固有的或低氧诱导的HIF鄄1α表达足以影响下游靶基因表达,siRNA能抑制促血管新生的靶基因,抑制EPCs向EC分化。     

阿司匹林对人结肠癌细胞株SW480、HT29中VEGF、β-catenin表达的影响

目的 探讨阿司匹林对人结肠癌细胞株HT-29、SW480生长增殖的影响及其β-catenin、VEGF蛋白和mRNA的表达,并为结直肠癌的预防和治疗提供新的实验和理论依据。方法 采用MTT法检测阿司匹林对SW480、HT-29细胞生长增殖的影响;Real-time PCR和Western blot法检测HT-29、SW480细胞中VEGF、β-catenin mRNA及蛋白的表达情况。结果 在SW480、HT-29细胞的生长增殖过程中,阿司匹林对其有着显著的抑制作用,且具有量效、时效关系;除6 h干预组外,余各组阿司匹林均可抑制VEGF、β-catenin蛋白和mRNA的表达（P＜0.05）。结论 阿司匹林对人结直肠癌细胞株SW480、HT-29的生长增殖具有显著的抑制作用,并对SW480、HT-29中VEGF、β-catenin mRNA和蛋白质的表达具有显著的下调效应;而阿司匹林通过抑制β-catenin的表达,同时调节VEGF抑制肿瘤血管的生成可能是其预防和治疗结直肠癌的作用机制之一。     

短发夹RNA抑制肝癌细胞HepG2 COX-2基因表达的探讨

目的：研究shRNA对肝癌细胞株HepG2中COX-2基因表达的抑制作用,同时检测对细胞周期和细胞生长的影响。方法：设计、合成一对环氧化酶-2（COX-2）基因编码的反向重复序列,中间间隔9个核苷酸序列,定向克隆至质粒载体pGenesil-1,构建抑制COX-2基因的短发夹RNA（short hairpin RNA shRNA）的重组表达质粒pshRNA-COX-2,用脂质体Lipofectamine^TM2000转染肝癌细胞HepG2,经RT-PCR和Westernblot分别检测pshRNA-COX-2重组表达质粒对COX-2mRNA和蛋白抑制效应,流式细胞仪检测细胞周期、细胞计数检测细胞生长变化。结果：pshRNA-COX-2重组表达质粒能够抑制COX-2基因表达。与未转染肝癌细胞HepG2相比,pshRNA-COX-2重组表达质粒转染肝癌细胞后,COX-2 mRNA和蛋白表达明显降低,抑制率分别为69.9%和50.3%;G0-G1期细胞由55.4%上升为77.9%,S期细胞由31.1%下降为16.2%;细胞生长明显减慢。结论：构建的pshRNA-COX-2重组表达载体能够显著抑制COX-2基因表达;引起G0-G1期细胞增多,S期细胞减少,从而阻止肝癌细胞生长。     

RNAi沉默CD133基因表达对结肠癌细胞株HT-29影响的研究

目的:探讨通过RNA干扰沉默CD133基因对结肠癌细胞株HT-29的增殖和侵袭力的影响。方法:将CD133siRNA用脂质体转染入HT-29细胞,以逆转录-聚合酶链反应(RT-PCR)检测CD133mRNA的变化,蛋白质印迹法检测转染前后蛋白的表达水平,用MTT比色法和Transwell小孔实验分别检测癌细胞的增殖及侵袭力变化。结果:CD133siRNA转染成功,与正常对照组相比,转染组CD133mRNA及蛋白的表达水平均有明显的抑制,并呈现浓度时间依赖性。MTT实验证明转染组细胞随时间延长,增殖逐渐受到抑制,P<0.05。Tran-swell小孔实验中经转染处理的穿膜细胞数明显减少(P<0.05),RNA干扰降低了细胞的侵袭性。结论:CD133在HT-29癌细胞增殖及侵袭过程中起重要作用,RNA干扰可以有效抑制其CD133的表达,并能有效降低癌细胞的增殖及侵袭力。     

Specific inhibition of cyclooxygenase-2 (COX-2) expression by dietary curcumin in HT-29 human colon cancer cells

Curcumin, a major yellow pigment and active component of turmeric, has been shown to possess anti-inflammatory and anti-cancer activities. Cyclooxygenase (COX)-2 plays an important role in colon carcinogenesis. To investigate the effect of curcumin on COX-2 expression, we treated HT-29 human colon cancer cells with various concentrations of curcumin. Curcumin inhibited the cell growth of HT-29 cells in a concentration- and time-dependent manner. Curcumin markedly inhibited the mRNA and protein expression of COX-2, but not COX-1. These data suggest that a non-toxic concentration of curcumin has a significant effect on the in vitro growth of HT-29 cells, specifically inhibits COX-2 expression, and may have value as a safe chemopreventive agent for colon cancer.     

Cyclooxygenase-2 is up-regulated by interleukin-1 beta in human colorectal cancer cells via multiple signaling pathways

Overexpression of cyclooxygenase-2 (COX-2) has been observed in human colorectal cancer. COX-2 expression in human tumors can be induced by growth factors, cytokines, oncogenes, and other factors. The mechanisms regulating COX-2 expression in human colon cancer have not been completely elucidated. We hypothesized that the proinflammatory cytokine interleukin-1 beta (IL-1 beta) mediates COX-2 expression in HT-29 human colon cancer cells. Treatment of HT-29 cells with IL-1 beta induced expression of COX-2 mRNA and protein in a time- and dose-dependent manner. Inhibitors of the extracellular signal-regulated kinase 1/2, c-Jun NH(2)-terminal kinase, P38 mitogen-activated protein kinase, and nuclear factor-kappa B (NF-kappa B) signaling pathways blocked the ability of IL-1 beta to induce COX-2 mRNA. In contrast, Wortmannin, a phosphoinositide 3-kinase inhibitor upstream of protein kinase B/Akt, led to a slight increase in COX-2 mRNA expression after IL-1 beta treatment. Electrophoretic mobility shift assay on nuclear extracts demonstrated that IL-1 beta induced NF-kappa B DNA binding activity in HT-29 cells, and the activated NF-kappa B complex was eliminated after treatment with an inhibitor of NF-kappa B. Supershift assay indicated that the two NF-kappa B subunits, p65 and p50, were involved in activation of NF-kappa B complex by IL-1 beta stimulation. The stability of COX-2 mRNA was not altered by IL-1 beta treatment. These data demonstrate that IL-1 beta induces COX-2 expression in HT-29 cells through multiple signaling pathways and NF-kappa B.     

藤黄酸联合肿瘤坏死因子相关凋亡诱导配体诱导人结肠癌HT-29细胞凋亡的效果和机制

背景与目的：肿瘤坏死因子相关凋亡诱导配体（tumor necrosis factor-related apoptosis-inducing ligand,TRAIL）能选择性地杀伤肿瘤细胞,但多种肿瘤对其耐药。该研究旨在探讨藤黄酸（gambognic acid,GA）与TRAIL联合对人结肠癌HT-29细胞裸鼠移植瘤生长的影响及GA联合TRAIL抗结肠癌的作用机制。方法: 建立人结肠癌HT-29细胞裸鼠皮下移植瘤模型,测定TRAIL和（或）GA对裸鼠移植瘤的影响,采用H-E染色观察肿瘤组织形态结构改变,TUNEL法检测细胞凋亡状况;HT-29细胞经过siRNA干扰Nrf2表达后,通过AnnexinⅤ-FITC/PI双染检测细胞凋亡情况,流式细胞术检测细胞内活性氧（reactive oxygen species,ROS）含量,RT-PCR法检测Nrf2、Bcl-2、Bax和DR5 mRNA的表达水平。结果: 联合应用GA显著促进了TRAIL对裸鼠皮下移植瘤的生长抑制（抑瘤率达67.0%）和诱导凋亡作用,下调了组织中Nrf2、Bcl-2的表达,增强了Bax和DR5的表达;与阴性对照siRNA相比,Nrf2干扰能明显上调TRAIL诱导HT-29细胞的凋亡率,提高ROS含量,下调Nrf2和Bcl-2表达,上调Bax和DR5表达。结论: GA可能是通过下调Nrf2表达,使ROS水平升高而激活线粒体凋亡途径与死亡受体途径,从而逆转人结肠癌HT-29细胞体内外对TRAIL的耐药性。     

靶向SMC1A基因慢病毒介导的RNA干扰对奥沙利铂治疗敏感性的影响

目的 探讨沉默结直肠癌细胞株染色体结构维持-1A（SMC1A）基因对奥沙利铂（L-OHP）敏感性的影响。方法 构建SMC1A基因的小分子RNA干扰表达质粒。与慢病毒包装质粒一起转染239T细胞制备pGCSIL-GFP／SMC1A-siRNA慢病毒载体,并将重组质粒转染结直肠癌SW480、HT-29细胞株。RT-PCR、Western blotting法检测SMC1A mRNA和蛋白的表达情况。CCK-8法检测不同浓度L-OHP对细胞的抑制作用。 结果 pGCSIL-GFP／SMC1A-siRNA慢病毒载体构建成功,RT-PCR产物为343bp。RT-PCR显示重组质粒转染的SW480、HT-29细胞SMC1A mRNA表达量明显低于其对应的未转染细胞（0.13±0.02 vs. 1.02±0.06,0.182±0.019 vs. 1.114±0.032;P＜0.05）;Western blotting 显示干扰后的细胞SMC1A蛋白表达受到明显抑制;不同浓度的L-OHP处理细胞48h后,CCK-8显示转染SMC1A-siRNA细胞生长抑制率较未转染细胞高,差异具有统计学意义（P＜0.05）。结论 沉默结直肠癌细胞株SMC1A基因对奥沙利铂的敏感性显著增加,为临床进一步提高结直肠癌的疗效提供了研究基础。     

Colon cancer cells with high invasive potential are susceptible to induction of apoptosis by a selective COX-2 inhibitor

Cyclooxygenase-2 (COX-2) expression has been shown to correlate with the invasiveness of colon cancer cells. To further investigate this positive correlation and its possible therapeutic implications, a selective COX-2 inhibitor, etodolac, was tested on three variants of HT-29 colon cancer cell lines, HT-29/lnv1, HT-29/ Inv2 and HT-29/lnv3, with graded increases of in vitro Matrigel invasive potential and COX-2 expression levels. HT-29 variants with higher invasive potential were found to be more sensitive to etodolac by in vitro growth inhibition assays, the estimated LD₅₀ being 0.5 m M for highly invasive HT-29/lnv2 and HT-29/lnv3 cells, 0.6 m M for slightly less invasive HT-29/lnv1, and 1.8 m M for the parental HT-29. Treatment of the highly invasive HT-29/lnv2 and Inv3 variants with as little as 0.1 m M etodolac in the growth medium produced signs of apoptosis, as detected by DNA fragmentation and TUNEL (terminal deoxynucleotidyl transferase dUTP-biotin nick end labeling) assay. In vivo experiments in SCID mice showed that etolodac inhibited the growth of subcutaneous tumors induced by HT-29/lnv3 cells significantly more than those by the parental HT-29 cells. These results suggest that COX-2 inhibitors have a potential role in prevention of tumor invasion in colon cancer patients. (Cancer Sci 2003; 94: 253–258)     

改变细胞膜脂肪酸组成防治大肠癌的基因治疗研究*

目的:利用脂肪酸去饱和酶基因fat- 1 改变细胞膜脂肪酸组成,进行大肠癌的基因治疗研究。方法:将fat- 1 基因插入腺病毒载体中,与骨架载体同源重组,构建腺病毒重组载体（Ad-GFP-fat1）,通过包装细胞系（293）产生的腺病毒,感染人大肠癌株HT- 29细胞。提取细胞的总RNA,以fat- 1 基因的反义mRNA 作探针,用Northern Blot检测fat- 1 基因在HT- 29细胞内的表达。以流式细胞仪对HT- 29细胞G0/G1 期、S 期、G2/M期所占比例进行检测,分析fat- 1 基因对HT- 29细胞增殖和凋亡的影响。以气相色谱分析仪分析fat- 1 基因对HT- 29细胞细胞膜n-6 PUFAs 和n-3 PUFAs 含量及n-6/n- 3PUFAs 比例的影响。将HT- 29细胞皮下接种于裸鼠右前肢腋下,建立裸鼠HT- 29大肠癌细胞皮下移植瘤模型。成瘤后进行治疗实验,经连续5 次治疗,于最后一次治疗后第3 天处死小鼠,取肿瘤称重。分析fat- 1 基因裸鼠体内抗肿瘤效果。结果:通过基因重组技术,得到高滴度的含fat- 1 基因的重组病毒;腺病毒介导的fat- 1 基因能够在HT- 29细胞中有效表达;fat- 1 基因的表达可降低HT- 29细胞膜n-6/n- 3PUFAs 的比例,有效抑制HT- 29细胞增殖,促进细胞凋亡并能抑制裸鼠移植瘤的发展。结论:fat- 1 基因的表达,可抑制HT- 29细胞的体内外增殖并诱导细胞凋亡,在大肠癌基因治疗中可能具有良好利用价值。      

n-3 PUFAs reduce VEGF expression in human colon cancer cells modulating the COX-2/PGE2 induced ERK-1 and -2 and HIF-1alpha induction pathway

n-3 Polyunsaturated fatty acids (PUFAs) inhibit the development of microvessels in mammary tumors growing in mice. Human colorectal tumors produce vascular endothelial growth factor (VEGF) whose expression is up-regulated in tumor cells by both cyclooxygenase-2 (COX-2) and PGE(2) and directly correlated to neoangiogenesis and clinical outcome. The goal of this study was to examine the capability of n-3 PUFAs to regulate VEGF expression in HT-29 human colorectal cells in vitro and in vivo. Constitutive VEGF expression was augmented in cultured HT-29 cells by serum starvation and the effects of eicosapentaenoic (EPA) or docosahexaenoic acid (DHA) on VEGF, COX-2, phosphorylated extracellular signal-regulated kinase (ERK)-1 and -2 and hypoxia-inducible-factor 1-alpha (HIF-1alpha) expression and PGE(2) levels were assessed. Tumor growth, VEGF, COX and PGE(2) analysis were carried out in tumors derived from HT-29 cells transplanted in nude mice fed with either EPA or DHA. Both EPA and DHA reduced VEGF and COX-2 expression and PGE(2) levels in HT-29 cells cultured in vitro. Moreover, they inhibited ERK-1 and -2 phosphorylation and HIF-1alpha protein over-expression, critical steps in the PGE(2)-induced signaling pathway leading to the augmented expression of VEGF in colon cancer cells. EPA always showed higher efficacy than DHA in vitro. Both fatty acids decreased the growth of the tumors obtained by inoculating HT-29 cells in nude mice, microvessel formation and the levels of VEGF, COX-2 and PGE(2) in tumors. The data provide evidence that these n-3 PUFAs are able to inhibit VEGF expression in colon cancer cells and suggest that one possible mechanism involved may be the negative regulation of the COX-2/PGE(2) pathway. Their potential clinical application as anti-angiogenic compounds in colon cancer therapy is proposed.