吲哚美辛通过E-Cad逆转高糖诱导的胰腺癌细胞增殖作用

目的:研究吲哚美辛对高糖环境下胰腺癌细胞增殖能力的影响，并探讨相关机制。方法:体外培养胰腺癌细胞株BXPC-3和 Panc-1，应用MTT法检测肿瘤细胞在不同干预条件下的增殖能力；应用Real time-PCR和Western blot检测细胞中E-Cad和COX-2的表达。应用小干扰RNA敲除肿瘤细胞中E-Cad的表达。结果:高糖可促进胰腺癌细胞增殖能力；而吲哚美辛可逆转高糖条件促进胰腺癌细胞的增殖，同时吲哚美辛可调控肿瘤细胞中E-Cad的表达，且小干扰胰腺癌细胞E-Cad后可逆转吲哚美辛对高糖引起的增殖抑制作用。结论:吲哚美辛可以抑制高糖诱导的胰腺癌细胞增殖，且该过程是通过上调E-Cad实现的，而非通过COX-2途径。     

选择性环氧合酶2抑制剂对人膀胱癌细胞T24增殖和凋亡的作用

背景与目的:环氧合酶2(cyclooxygenase-2,COX-2)与膀胱癌的发生发展密切相关,因此COX-2抑制剂对于膀胱癌可能具有潜在的抗肿瘤效应.本实验旨在探讨选择性COX-2抑制剂对人膀胱癌细胞T24增殖和凋亡的抑制作用.方法:用四甲基偶氮唑蓝还原法(MTT)研究选择性COX-2抑制剂SC-58125、塞来昔布(celecoxib)和非选择性COX抑制剂吲哚美辛(indomethacin)对T24细胞增殖的影响;用流式细胞术、DNA电泳和Hoechst33258荧光染色3种方法检测SC-58125作用下T24细胞的凋亡,同时逆转录聚合酶链反应(RT-PCR)检测细胞凋亡相关基因Bcl-2和Bax基因的变化.结果:在12.5 μmol/L至200 μmol/L的剂量范围内,SC-58125、塞来昔布及吲哚美辛均能不同程度地抑制T24细胞的增殖,以SC-58125的抑制作用最强,IC50在25 μmol/L～50 μmol/L之间;3种凋亡检测方法均观察到SC-58125作用后凋亡细胞的比例增加,其中100 μmol/L的SC-58125作用于T24细胞6 h和12 h后,流式细胞仪测得凋亡细胞的比例分别为(7.95±1.88)%和(12.5±2.42)%,较对照组增加(P＜0.05),但细胞内与凋亡相关的Bcl-2和Bax基因表达没有变化.结论:选择性COX-2抑制剂体外可抑制T24细胞的增殖,并诱导细胞凋亡.     

吲哚美辛对小鼠肝癌细胞免疫的影响

吲哚美辛是一重要的生物反应调节剂，广泛用于各种肿瘤的实验性研究和临床辅助治疗.本采用微量肝癌细胞悬液移植法复制了肝内移植型小鼠肝癌模型．并采用此模型观察吲哚美辛对肝癌小鼠细胞免疫的影响。结果：肝癌模型的建立成功率为100％，无自发性消退。吲哚美辛对肝癌小鼠细胞免疫的影响:显示可明显提高宿主外周血中T辅助淋巴细胞，降低T抑制淋巴细胞的水平，使T辅/T抑比值增高，并且增加外周血中T淋巴细胞数．由此表明，吲哚美辛明显增强肝癌宿主细胞免疫，其作用的实现可能与抑制肝癌组织中前列腺素E2的合成有关。     

乳腺癌患者髓系来源抑制性细胞的鉴定与免疫作用研究

目的:探讨乳腺癌患者体内一群髓系来源抑制性细胞(MDSCs)的表型特点及它对淋巴细胞的抑制作用.方法:收集20例乳腺癌患者外周血及肿瘤组织,10例体检正常健康者外周血,通过流式和免疫组织化学染色技术检测MDSCs的表型特点及比例.采用免疫磁珠技术分选乳腺癌患者肿瘤中的MDSCs,通过瑞氏染色观察细胞形态,分别利用MTT法、流式细胞术和ELISA法检测其对自体淋巴细胞增殖、凋亡和细胞因子分泌的影响.结果:流式细胞仪检测结果显示乳腺癌患者肿瘤组织及外周血中有一群具有髓系前体细胞形态的MDSCs,表面标志为CD45+ CD13+CD33+CD14-CD15-.实验结果显示肿瘤组织来源的MDSCs显著抑制自体淋巴细胞增殖,促进其调亡,并显著抑制其分泌IFN-γ,促进TGF-β、IL-10释放.结论:乳腺癌患者肿瘤组织中存在一群具有独特表型特点的MDSCs,不仅抑制自体淋巴细胞增殖和因子分泌,同时促进细胞凋亡.     

吲哚美辛诱导肝癌HepG2细胞凋亡及其对cyclin E蛋白表达的影响

目的　观察吲哚美辛诱导肝癌 Hep G2细胞凋亡 ,及其对 cyclin E蛋白的影响。方法　采用 MTT比色法观察吲哚美辛对肝癌 Hep G2细胞增殖的影响 ;采用流式细胞仪检测吲哚美辛对 Hep G2细胞周期分布的影响 ,同时结合透射电子显微镜观察吲哚美辛诱导肝癌 Hep G2细胞凋亡的作用 ;免疫细胞化学方法观察吲哚美辛对Hep G2细胞周期调控蛋白 cyclin E的影响。结果　吲哚美辛可抑制肝癌 Hep G2细胞的增殖 ,诱导其凋亡 ,改变细胞周期分布 ,使 G0 /G1 期细胞比例增高 ,S期比例降低 ,同时还可使 cyclin E蛋白表达下降。上述作用具有剂量和时间依赖性 (P<0 .0 5 )。结论　吲哚美辛可诱导肝癌 Hep G2细胞凋亡 ,改变细胞周期分布 ,影响细胞周期调控蛋白表达 ,从而抑制细胞增殖。     

肝细胞肝癌组织COX-2和VEGF及NF-κB的表达及相关性研究

目的:分析肝细胞肝癌(HCC)组织中环氧化酶-2(COX-2)与血管内皮生长因子(VEGF)、核转录因子κB(NF-κB)的表达及相关性,并探讨癌组织微环境中炎症反应与HCC发生、进展的关系。方法:采用免疫组织化学方法检测53例HCC组织中COX-2、VEGF、NF-κB与Ki-67的表达,并建立HCC组织微环境炎症级别标准。结果:HCC组织微环境炎症级别与COX-2表达水平相关(P<0.01),COX-2表达水平与VEGF、NF-κB相关(P<0.05),NF-κB表达水平与VEGF、Ki-67相关,P<0.05。结论:HCC组织中COX-2的表达与肿瘤微环境中炎症反应程度密切相关,COX-2在HCC的发生、发展和转移过程中起重要作用,可能与血管生成和NF-κB信号通路有关。     

塞来昔布对胃癌抑制作用的实验研究

目的 :探讨塞来昔布和吲哚美辛对胃癌细胞增殖的抑制作用及可能的机制。方法 :MTT比色实验检测药物对 SGC790 1细胞和 AGS细胞增殖的影响 ,裸鼠移植瘤实验检测药物对胃癌细胞成瘤性的影响 ;流式细胞术检测药物对胃癌细胞凋亡的影响 ;电镜观察药物作用前后胃癌细胞的超微结构的变化。结果 :塞来昔布和吲哚美辛抑制 SGC790 1细胞和 AGS细胞增殖 ,效应呈剂量依赖性 ,两者抑制率比较差异有显著性 (P<0 .0 5 ) ;塞来昔布和吲哚美辛抑制荷瘤裸鼠肿瘤增殖 ,瘤结节重量分别为 (48.3± 2 .88) mg和 (83.3± 2 0 .81) m g,对照组为 (82 6 .7± 77.72 ) mg(P<0 .0 5 ) ;塞来昔布和吲哚美辛可诱导细胞凋亡 ,凋亡细胞分别增加了 30 .4 % (34.5 %比 4 .1% )和 2 3.9% (2 8.0 %比 4 .1% )。塞来昔布可使胃癌细胞超微结构发生良性转化 ,可见有凋亡特征的细胞。结论 :塞来昔布和吲哚美辛可抑制胃癌体内外增殖 ,前者作用更强 ,诱导细胞凋亡是其可能的作用机制之一。     

吲哚美辛对结肠癌细胞中细胞周期蛋白的影响

目的体外观察吲哚美辛对含突变型p53的SW480细胞及经野生型p53转染的结肠癌细胞株SW480(wtp53/SW480)的影响,探讨吲哚美辛抗结肠癌作用的分子机制.方法不同浓度的吲哚美辛作用于细胞株,采用Western斑点免疫印迹方法,分别检测SW480细胞及wtp53/SW480细胞中CDK2、CDK4和p21WAF1/PIC1蛋白的表达水平.结果吲哚美辛作用于wtp53/SW480细胞24 h后,其CDK2、CDK4蛋白表达水平随吲哚美辛浓度的增加而逐渐下调,呈浓度依赖性,以600μmol/L时抑制作用最强;而p21WAF1/PIC1蛋白表达水平上调,其表达水平随着一定范围内吲哚美辛浓度的增加而逐渐上调,以400μmol/L吲哚美辛时作用最强,600μmol/L时回到基础水平.在未经转染的SW480细胞中CDK2、CDK4和p21WAF1/PIC1蛋白表达没有发生明显改变.结论吲哚美辛抗结肠癌作用部分分子机制可能是通过下调CDK2、CDK4蛋白的表达和上调p21WAF1/PIC1蛋白来实现且存在p53-p21WAF1/CIP1依赖途径.     

乳腺癌髓系来源抑制细胞中IDO对T淋巴细胞免疫抑制作用初探

  目的   检测乳腺癌患者肿瘤原位组织的一群髓系来源抑制细胞（MDSCs）中吲哚胺2,3双加氧酶（indoleamine-2,3-dioxygenase,IDO）的表达情况,探讨IDO对MDSCs介导T淋巴细胞免疫抑制作用的影响。   方法   收集30例乳腺癌患者的肿瘤组织和外周血及30例健康供者外周血,将肿瘤组织制成单细胞悬液,采用免疫磁珠技术分选肿瘤单细胞悬液中CD33+ MDSCs和健康供者外周血中的CD33+细胞,应用Western blot和PCR方法检测MDSCs中IDO的表达情况。将肿瘤组织来源MDSCs和异体T淋巴细胞按照1:1比例混合培养3天,在加用和不加IDO特异性抑制剂1-MT条件下,利用Annexin-V凋亡试剂盒检测各组T淋巴细胞凋亡率,利用ELISA法检测各组T淋巴细胞分泌的细胞因子量。   结果   Western blot和PCR检测发现MDSCs中IDO过表达。T细胞单独培养时凋亡率为（2.40±0.66）%,MDSCs和T细胞共孵育组中T细胞凋亡率为（12.30±0.80）%,比T细胞单独培养时显著升高（P < 0.05）,在共孵育过程中加用1-MT组的T细胞凋亡率为（3.30±0.58）%,与不加1-MT组比较差异具有统计学意义（P < 0.05）。细胞因子检测的结果发现MDSCs促进T淋巴细胞TGF-β、IL-10的释放,抑制IFN-γ的分泌,而对IL-4和IL-12的分泌影响并不明显,而加用1-MT后MDSCs和T淋巴细胞共孵育组中TGF-β、IL-10的分泌水平与未加1-MT组相比显著降低,IFN-γ的分泌显著增加（P < 0.05）。   结论   在乳腺癌患者中,原位肿瘤组织来源的MDSCs对T细胞具有明显的免疫抑制作用;IDO在此群细胞中有过表达,MDSCs发挥免疫抑制作用与IDO密切相关。      

人体肿瘤髓系来源抑制性细胞的研究进展

髓系来源抑制性细胞(MDSC)是一群来源于髓系的异质性未分化成熟的细胞;在感染、创伤、败血症,尤其是肿瘤等病理状态下,MDSC在血液、淋巴器官、脾脏及肿瘤等组织中大量聚集;MDSC可以通过多种机制抑制肿瘤免疫,MDSC在患者外周血的数量与肿瘤的分期、负荷、远处转移及患者的预后密切相关.然而关于MDSC的研究结果大部分来自小鼠实验,由于肿瘤患者MDSC的表型比较复杂,其研究进展相对缓慢.     

PGE(2)-induced CXCL12 production and CXCR4 expression controls the accumulation of human MDSCs in ovarian cancer environment

Signals mediated by CXCL12 (SDF1) and its receptor CXCR4 are centrally involved in cancer progression, both directly by activating cancer cells and indirectly by inducing angiogenesis plus recruiting T regulatory and plasmacytoid dendritic immune cells. Here, we show that in ascites isolated from ovarian cancer patients, both CXCL12 and CXCR4 are controlled by the tumor-associated inflammatory mediator prostaglandin E(2) (PGE(2)), which attracts myeloid-derived suppressor cells (MDSC) into the ascites microenvironment. In this setting, PGE(2) was essential both for expression of functional CXCR4 in cancer-associated MDSCs and for production of its ligand CXCL12. Frequencies of CD11b(+)CD14(+)CD33(+)CXCR4(+) MDSCs closely correlated with CXCL12 and PGE(2) levels in patient ascites. MDSCs migrated toward ovarian cancer ascites in a CXCR4-dependent manner that required COX2 activity and autocrine PGE(2) production. Inhibition of COX2 or the PGE(2) receptors EP2/EP4 in MDSCs suppressed expression of CXCR4 and MDSC responsiveness to CXCL12 or ovarian cancer ascites. Similarly, COX2 inhibition also blocked CXCL12 production in the ovarian cancer environment and its ability to attract MDSCs. Together, our findings elucidate a central role for PGE(2) in MDSC accumulation triggered by the CXCL12-CXCR4 pathway, providing a powerful rationale to target PGE(2) signaling in ovarian cancer therapy.     

Induction of myeloid‐derived suppressor cells by tumor exosomes

Myeloid‐derived suppressor cells (MDSCs) promote tumor progression. The mechanisms of MDSC development during tumor growth remain unknown. Tumor exosomes (T‐exosomes) have been implicated to play a role in immune regulation, however the role of exosomes in the induction of MDSCs is unclear. Our previous work demonstrated that exosomes isolated from tumor cells are taken up by bone marrow myeloid cells. Here, we extend those findings showing that exosomes isolated from T‐exosomes switch the differentiation pathway of these myeloid cells to the MDSC pathway (CD11b⁺Gr‐1⁺). The resulting cells exhibit MDSC phenotypic and functional characteristics including promotion of tumor growth. Furthermore, we demonstrated that in vivo MDSC mediated promotion of tumor progression is dependent on T‐exosome prostaglandin E2 (PGE2) and TGF‐β molecules. T‐exosomes can induce the accumulation of MDSCs expressing Cox2, IL‐6, VEGF, and arginase‐1. Antibodies against exosomal PGE2 and TGF‐β block the activity of these exosomes on MDSC induction and therefore attenuate MDSC‐mediated tumor‐promoting ability. Exosomal PGE2 and TGF‐β are enriched in T‐exosomes when compared with exosomes isolated from the supernatants of cultured tumor cells (C‐exosomes). The tumor microenvironment has an effect on the potency of T‐exosome mediated induction of MDSCs by regulating the sorting and the amount of exosomal PGE2 and TGF‐β available. Together, these findings lend themselves to developing specific targetable therapeutic strategies to reduce or eliminate MDSC‐induced immunosuppression and hence enhance host antitumor immunotherapy efficacy. © 2008 Wiley‐Liss, Inc.     

COX-2 blockade suppresses gliomagenesis by inhibiting myeloid-derived suppressor cells

Epidemiologic studies have highlighted associations between the regular use of nonsteroidal anti-inflammatory drugs (NSAID) and reduced glioma risks in humans. Most NSAIDs function as COX-2 inhibitors that prevent production of prostaglandin E? (PGE?). Because PGE? induces expansion of myeloid-derived suppressor cells (MDSC), we hypothesized that COX-2 blockade would suppress gliomagenesis by inhibiting MDSC development and accumulation in the tumor microenvironment (TME). In mouse models of glioma, treatment with the COX-2 inhibitors acetylsalicylic acid (ASA) or celecoxib inhibited systemic PGE? production and delayed glioma development. ASA treatment also reduced the MDSC-attracting chemokine CCL2 (C-C motif ligand 2) in the TME along with numbers of CD11b(+)Ly6G(hi)Ly6C(lo) granulocytic MDSCs in both the bone marrow and the TME. In support of this evidence that COX-2 blockade blocked systemic development of MDSCs and their CCL2-mediated accumulation in the TME, there were defects in these processes in glioma-bearing Cox2-deficient and Ccl2-deficient mice. Conversely, these mice or ASA-treated wild-type mice displayed enhanced expression of CXCL10 (C-X-C motif chemokine 10) and infiltration of cytotoxic T lymphocytes (CTL) in the TME, consistent with a relief of MDSC-mediated immunosuppression. Antibody-mediated depletion of MDSCs delayed glioma growth in association with an increase in CXCL10 and CTLs in the TME, underscoring a critical role for MDSCs in glioma development. Finally, Cxcl10-deficient mice exhibited reduced CTL infiltration of tumors, establishing that CXCL10 limited this pathway of immunosuppression. Taken together, our findings show that the COX-2 pathway promotes gliomagenesis by directly supporting systemic development of MDSCs and their accumulation in the TME, where they limit CTL infiltration.     

Cyclooxygenase inhibitors decrease the growth and induce regression of human esophageal adenocarcinoma xenografts in nude mice

Cyclooxygenase (COX) inhibition has been shown to prevent the development of esophageal adenocarcinoma (EAC). However, the potential of this approach for treatment of established cancer has been poorly investigated. Our objective was to determine whether non-selective or selective inhibition of the COX pathway affects the growth of esophageal adenocarcinoma xenografts in nude mice. A human esophageal adenocarcinoma xenograft model was established by subcutaneous inoculation of OE33 cells in nude mice. Small tumor slices harvested from four OE33 xenografts were implanted in the flanks of new mice that were randomized to different treatments (6 animals per group): indomethacin (3 mg/kg/day), parecoxib (0.11 and 0.22 mg/kg/day) or a selective prostaglandin E? receptor antagonist (AH-23848B, 1 mg/kg/day). For each treatment, a control group of 6 animals (vehicle) carrying xenografts from the same OE33 tumor was included. Tumor growth was measured twice a week. After 8 weeks mice were euthanized. Tumors were assessed by histological analysis, mRNA expression of COX isoenzymes, PGE? receptors and PGE? content. All OE33 tumors were poorly differentiated esophageal adenocarcinomas. Tumors expressed COX-2, EP?, EP? and EP? receptor mRNA. Treatment with parecoxib, higher dose or indomethacin significantly inhibited tumor growth. Furthermore, indomethacin induced tumor regression (74 vs 582% in control animals; p<0.01). However, AH-23848B or parecoxib low dose failed to affect tumor growth significantly. PGE? content in tumors was significantly decreased by high-dose parecoxib and indomethacin. Indomethacin and parecoxib inhibit the growth of human esophageal adenocarcinoma xenografts in nude mice, which suggests a potential role for NSAIDs or selective COX-2 inhibitors for EAC chemotherapy.     

Cyclooxygenase inhibitors retard murine mammary tumor progression by reducing tumor cell migration, invasiveness and angiogenesis

Tumor-derived prostaglandins (PGs) have been implicated in the progression of murine and human breast cancer. Chronic treatment with a non-selective PG inhibitor indomethacin was shown in this laboratory to retard the development and metastasis of spontaneous mammary tumors in C3H/HeJ female retired breeder mice. The present study examined the role of endogenous prostaglandins in the proliferation/survival, the migratory and invasive behavior and angiogenic ability of a highly metastatic murine mammary tumor cell line, C3L5, originally derived from a C3H/HeJ spontaneous mammary tumor. This cell line was shown to express high levels of cyclooxygenase (COX) -2 mRNA and protein as detected by Northern and Western blotting as well as immunostaining. PGE(2) production by C3L5 cells was primarily owing to COX-2, since this was blocked similarly with non-selective COX inhibitor indomethacin and selective COX-2 inhibitor NS-398, but unaffected with the selective COX-1 inhibitor valeryl salicylate. C3L5 cell proliferation/survival in vitro was not influenced by PGs, since their cellularity remained unaffected in the presence of PGE(2) or NS-398 or PG-receptor (EP1/EP2) antagonist AH6809; a marginal decline was noted only at high doses of indomethacin, which was not abrogated by addition of exogenous PGE(2). Migratory and invasive abilities of C3L5 cells, as quantitated with in vitro transwell migration/invasion assays, were inhibited with indomethacin or NS-398 or AH6809 in a dose-dependent manner; the indomethacin and NS-398-mediated inhibition was partially reversed upon addition of exogenous PGE(2). An in vivo angiogenesis assay that used subcutaneous implants of growth factor-reduced matrigel inclusive of tumor cells showed a significant inhibition of blood vessel formation in these implants in animals treated with indomethacin compared with animals receiving vehicle alone. These studies show that selective and nonselective COX-2 inhibitors retarded tumor progression in this COX-2-expressing murine mammary tumor model by inhibiting tumor cell migration, invasiveness and tumor-induced angiogenesis. The inhibitory effects were not entirely PG dependent; some PG-independent effects were also noted.     

Cyclooxygenase-2 inhibitors suppress the growth of human hepatocellular carcinoma implants in nude mice

Cyclooxygenase (COX)-2 is expressed in hepatocellular carcinomas (HCCs) and HCC cell lines. COX-2 inhibition strongly suppresses growth of HCC cells in vitro by inducing apoptosis and reducing proliferation. Here, we evaluate the in vivo effects and mechanism of COX-2 inhibition of human HCC cell line derived xenotransplanted tumors in nude mice. Firstly, nude mice were treated with a COX-2 specific inhibitor (meloxicam) or a non-specific inhibitor (sulindac) starting 5 days prior to tumor cell injection. After 35 days mice were killed and tumors were analyzed morphologically and assayed for proliferation (Ki67), apoptosis (M30) and COX-2 expression. Secondly, mice were treated with meloxicam or sulindac after tumors had reached a diameter of at least 0.2 cm. COX-2 expression was maintained in implant tumors at levels comparable with parental cells. Selective COX-2 inhibition led to a significant reduction of tumor growth and weight. COX-2 inhibition had a significant anti-proliferative and pro-apoptotic effect on tumor cells. These results demonstrate that under experimental conditions selective COX-2 inhibition suppresses solid HCC growth in vivo and, therefore may have preventive and therapeutic potential for human HCCs.     

Cyclooxygenase-2 pathway correlates with vascular endothelial growth factor expression and tumor angiogenesis in hepatitis B virus-associated hepatocellular carcinoma

Evidence indicates that cyclooxygenase (COX)-2-derived prostaglandins (PGs) contribute to tumor growth by inducing angiogenesis. We investigated the role of COX-2 in hepatitis B virus (HBV)-associated hepatocellular carcinoma (HCC). COX-2 and vascular endothelial growth factor (VEGF) expressions were examined by immunohistochemistry in 24 HBV-associated HCC. Tumor micro-vessel density (MVD) was assessed using CD34 immunohistochemistry. Hep3B HCC cell line, which carries integrated HBV genome, was stably transfected with human COX-2 cDNA. COX-2 and VEGF expressions were determined by Western blot while PG level was determined by ELISA. The effects of PGs on VEGF expression were also investigated. Expression of COX-2 and VEGF in HCC cells were observed in 19 (79%) and 16 (67%) cases, respectively. Well-differentiated HCC expressed COX-2 more strongly than less-differentiated HCC (p<0.001). COX-2 expression was found to correlate with VEGF expression and MVD (p=0.003 and 0.004, respectively). COX-2 overexpressing Hep3B clone had higher VEGF expression as compared to non-COX-2 expressing clone and parental cells. Treatment of the COX-2 overexpressing cells with a COX-2-selective inhibitor, NS-398 (10 microM), decreased PGE2 level and attenuated VEGF expression. Addition of PGE2 (10 microM) and the stable analog of PGI2, carbaprostacyclin (5 microM), to Hep3B cells also increased VEGF expression. Up-regulation of COX-2 correlates with VEGF expression and tumor angiogenesis in HBV-associated HCC. Moreover, COX-2 up-regulates VEGF expression in HCC cells, possibly via PGs production. Selective inhibition of COX-2 may block HCC associated angiogenesis and thus provides a rational approach for treatment of this malignancy.     

The role of cyclooxygenase in n-6 and n-3 polyunsaturated fatty acid mediated effects on cell proliferation,PGE(2) synthesis and cytotoxicity in human colorectal carcinoma cell lines

This study was conducted to investigate the role of the enzyme cyclooxygenase (COX) and its prostaglandin product PGE(2) in n-6 and n-3 polyunsaturated fatty acid (PUFA)-mediated effects on cellular proliferation of two human colorectal carcinoma cell lines. The long chain PUFAs eicosapentaenoic acid (EPA; 20:5n-3) and arachidonic acid (AA; 20:4n-6) both inhibited cell proliferation of Caco-2 cells compared with the long chain fatty acids alpha-linolenic acid (ALA; 18:3n-3) and linoleic acid (LA; 18:2n-6). Neither incubation with PGE(2) nor reduction in PGE(2) synthesis by EPA compared with AA led to differential effects on cell proliferation in Caco-2 cells. This suggests that n-6 and n-3 PUFA-mediated cell proliferation in Caco-2 cells is not regulated via PGE(2) levels. AA and EPA had no effect on growth of HT-29 colon cancer cells with a low COX activity. However, stimulation of COX-2 activity by IL-1 beta resulted in a decrease in cell proliferation and an induction of cytotoxicity by AA as well as by EPA. Both inhibition of the COX pathway by indomethacin as well as inhibition of direct lipid peroxidation by antioxidants such as vitamin E and C diminished the anti-proliferative effects of AA as well as EPA. Also, malondialdehyde, a product of lipid peroxidation and COX-activity was decreased by addition of vitamin E and partially decreased by indomethacin. These data support the hypothesis that growth inhibitory and cytotoxic effects of PUFAs with methylene-interrupted double bonds such as AA and EPA are due to peroxidation products that are generated during lipid peroxidation and COX activity.