NS-398和顺铂联合作用对人肺腺癌增殖的影响及其机制初步探讨

背景与目的　近年的研究结果显示COX 2抑制剂具有抑制肿瘤生长、增强化疗药物疗效的作 用。NS 398是一种选择性COX 2抑制剂,可抑制肺癌细胞的增殖并增强化疗药物的疗效。但NS 398与顺 铂联合作用对肺腺癌细胞增殖的影响及其作用机制目前尚不清楚。本研究旨在明确NS 398对顺铂肺腺癌 细胞增殖抑制作用的影响,探讨NS 398与顺铂的联合作用对细胞凋亡以及细胞周期的影响。方法　采用 MTT比色法检测NS 398和顺铂联合作用对肺腺癌细胞增殖的影响,吖啶橙和溴化乙啶荧光染色法观察肺 腺癌细胞凋亡情况,流式细胞仪PI染色法分析两药联合处理的肺腺癌细胞凋亡率和细胞周期的改变。结果 　联合NS 398后SPC A 1细胞顺铂IC50降低百分率为43.34%～69.61%,A549细胞为48.25%～62.44%, 两药联合为轻度协同～协同作用。NS 398与顺铂联合组作用24和48小时诱导肺腺癌细胞的凋亡率显著增 高,联合组48小时和72小时的S期细胞比例亦明显增高。NS 398和顺铂联合作用诱导的肺腺癌细胞凋亡 率与S期细胞比例呈正相关(r=0.882,P=0.01)。结论　NS 398可以增强顺铂对肺腺癌细胞增殖的抑制作 用,两种药物具有协同作用。NS 398与顺铂的协同作用可能与凋亡机制有关。     

人肺腺癌细胞耐顺铂株A549/CDDP的比较蛋白质组学研究

背景与目的 化疗是肺癌综合治疗的重要部分,但是肿瘤细胞耐药常导致化疗失败.本研究采用蛋白组学方法研究人肺腺癌细胞A549对顺铂产生耐药性前后的蛋白表达差异,筛选有价值的靶位.方法 以顺铂为诱导药物,人肺腺癌细胞系A549为诱导对象,采用逐步增加剂量与大剂量冲击相结合的方法,诱导建立耐顺铂细胞株A549/CDDP.对A549与A549/CDDP进行蛋白组学研究,即利用双向凝胶电泳(2-DE)分离两组总蛋白后,通过图像分析寻找表达差异的蛋白,对其进行MALDI-TOF质谱分析.结果 比较两者2-DE图谱,得到差异蛋白点82个;对其中6个差异蛋白点进行肽质量指纹图分析,鉴定出葡萄糖调节蛋白75、核糖体蛋白S4、线粒体F1-ATP合酶β亚单位、免疫球蛋白重链可变区;以上差异蛋白均在耐药株中过表达,而在对照组细胞中表达下调或不表达.结论 所鉴定的差异蛋白将为阐明肺癌细胞对顺铂耐药性产生的机制提供线索,也为筛选具体潜在价值的肺癌化疗靶位提供理论依据.     

人Ⅰ期肺腺癌及癌旁组织蛋白质双向电泳图谱的差异分析

目的筛选并鉴定存在明显表达差异的蛋白质,为人肺腺癌发病机制及其早期诊断的研究提供理论依据。方法采用固相pH梯度双向凝胶电泳技术分离12例Ⅰ期肺腺癌及其相配的癌旁正常肺组织可溶性总蛋白,建立人Ⅰ期肺腺癌及相配癌旁正常肺组织蛋白质的双向电泳凝胶图谱;PDQuest凝胶图像分析软件比较分析,筛选出差异表达的蛋白质点;基质辅助激光解吸电离飞行时间质谱获得相应蛋白质点的肽质量指纹图谱;搜索蛋白质数据库鉴定差异表达的蛋白质。结果建立了重复性较好的人Ⅰ期肺腺癌及相配癌旁正常肺组织蛋白质的双向电泳凝胶图谱;筛选出存在明显表达差异的蛋白质点26个,挖取其中的9个蛋白质点进行质谱分析,9个蛋白质点均得到了满意的肽质量指纹图谱;搜索蛋白质数据库鉴定出4种蛋白质,分别是:60S核糖体蛋白P2、组织蛋白酶B1、载脂蛋白A-I前体和La4.1蛋白。结论成功建立了人Ⅰ期肺腺癌及相配癌旁正常肺组织蛋白质的双向电泳凝胶图谱,鉴定出4种在肺腺癌发生早期出现明显表达变化的蛋白质,为进一步探索人肺腺癌的发病机制及寻找特异性的早期分子标志物提供了理论依据。     

人大肠腺瘤和腺癌之间相关差异蛋白的蛋白质组学研究

目的　应用蛋白质组学的方法,建立人大肠腺癌组织、大肠腺瘤组织、正常大肠黏膜组织的二维电泳图谱,筛选并鉴定三者之间的差异蛋白,发现人大肠癌的潜在生物学标志物。方法　收集同时患有大肠腺瘤的大肠腺癌患者１０例,术前及术后病理证实均为Ｄｕｋｅｓ’Ｂ期。取同一患者术后大肠癌组织、距大肠癌组织１５ｃｍ以上正常大肠黏膜组织以及肠镜下切除大肠腺瘤组织,提取不同组织总蛋白进行二维电泳,用ＩｍａｇｅＭａｓｔｅｒ２ＤＰｌａｔｉｎｕｍ５.０凝胶图像分析软件对双向电泳图依次进行凝胶点的检测、背景消减和编号。识别不同组织之间的差异蛋白点,对差异蛋白点用胶内酶解后行质谱分析和网上数据库鉴定。结果　大肠腺癌组织、大肠腺瘤组织和正常大肠黏膜组织之间存在９０个差异蛋白点。对这些差异点进行质谱分析,经数据库鉴定出７１个蛋白质。经分析,大肠腺瘤组织与正常大肠黏膜组织相比,１７个蛋白表达上调,９个蛋白表达下调;大肠腺癌组织与正常大肠黏膜组织比较,４７个蛋白表达上调,１２个蛋白表达下调;大肠腺癌组织与大肠腺瘤组织相比,３５个蛋白表达上调,８个蛋白表达下调。结论　大肠腺癌组织、大肠腺瘤组织和正常大肠黏膜组织之间存在差异蛋白表达,可能与大肠腺癌的发生发展有关;应用蛋白质组学方法筛选人大肠腺癌发生发展相关蛋白是可行的。     

rAd-p53联合顺铂体外抑制肺腺癌细胞增殖的最佳作用时序研究

目的:研究rAd-p53(商品名:今又生)与顺铂以不同序贯方式联合应用对肺腺癌细胞的增殖抑制作用,明确rAd-p53与顺铂联合的协同效应关系和最佳作用时序.方法:采用MTT法测定不同感染强度rAd-p53联合顺铂以及rAd-p53不同时序联合顺铂对人肺腺癌细胞A549的生长抑制率;采用流式细胞术检测细胞周期.结果:rAd-p53具有明显的肿瘤抑制作用,随着感染强度增加抑制作用增强;在较低感染强度下联合顺铂,即有显著的相加或协同作用(Q>0.85或Q>1.15),与单药组比较差异显著(P＜0.05);两药不同序贯方式联合抑制作用均大于各时间点单药作用(P＜0.01),尤以rAd-053后48h加入顺铂Q值最高(Q=1.38),该时间细胞被更多的阻滞在G0/G1期,而S期细胞比例减少.结论:rAd-p53与顺铂联合对肺腺癌细胞具有显著的协同抑制作用,并随剂量增加协同作用增强;联合应用时顺铂在rAd-p53作用48h时加入可最大发挥两者的协同抑制作用,其机制与细胞周期阻滞有关.     

环氧化酶-2抑制剂塞来昔布联合顺铂对A549人肺腺癌细胞株增殖影响的体外实验研究

目的观察环氧化酶-2（COX-2）抑制剂塞来昔布（Celecoxib）与化疗药物顺铂（DDP）联合应用对A549人肺腙癌细胞增殖的影响。方法应用MTS法检测Celecoxib与DDP联用对人肺腺癌A549细胞增殖的影响。结果在一定浓度范围内，Celecoxib和DDP均可抑制A549人肺腺癌细胞的生长，其抑制作用呈量-效关系。Celecoxib（0．35mg／L）与DDP联用可增强对A549细胞生长的抑制作用，DDP浓度≥1mg／L时两者具有协同或相加作用；且在一定浓度范围内，Celecoxib与DDP联用对细胞的生长抑制作用呈时-效关系。结论Celecoxib与DDP联合可增强对A549人肺腺癌细胞的生长抑制效应。     

紫杉醇抗胆管癌细胞差异表达蛋白的分离、质谱鉴定及生物信息学分析

目的:筛选紫杉醇诱导胆管癌细胞凋亡的相关蛋白并鉴定差异表达的蛋白点以寻找抗胆管癌药物作用的新靶点,筛选胆管癌新的肿瘤标志物,为胆管癌的早期诊断、疗效观察提供基础理论依据.方法:应用比较蛋白质组学技术对紫杉醇诱导凋亡的胆管癌QBC939细胞进行蛋白分离和蛋白表达谱差异分析,筛选并鉴定其相关蛋白并测定其胶内酶解后的肽指纹图谱.结果:本研究发现对照组和实验组蛋白点匹配率分别为(90.1±0.14)%和(87.1±0.22)%(P＜0.05),匹配后在干预组中共发现68个差异表达蛋白点,其中47个表达下调和21个上调;共获得28张PMF,初步质谱鉴定发现11个与凋亡相关的差异表达蛋白,其中6种蛋白表达上调,5种蛋白表达下调.结论:应用蛋白质组学技术分析所获得的68个差异蛋白质点可能在QBC939细胞发生凋亡过程中发挥重要作用;被鉴定出的11个与细胞凋亡相关的差异蛋白点提示紫杉醇是通过多个重要蛋白,发挥其诱导细胞凋亡的抗癌作用机制的.     

三七总皂苷对顺铂肾损害大鼠肾组织差异表达蛋白质的影响

目的 探讨三七总皂苷（panax notoginsenosides,PNS）对顺铂肾损害大鼠肾组织差异表达蛋白质的影响。方法 实验大鼠随机分为正常对照组、顺铂模型组和PNS治疗组,对大鼠给药处理10 d后,检测大鼠血清尿素氮（BUN）、肌酐（Scr）和尿N-乙酰-β-氨基葡萄糖苷酶（NAG）的水平,并做肾脏病理检查;采用SELDI-TOF-MS技术筛选大鼠肾组织的差异表达蛋白质,并通过MALDI-TOF-MS/MS、Western blot实验予以鉴定。结果 顺铂模型组大鼠血清BUN、Scr和尿NAG的水平均显著高于正常对照组（P均〈0.05）。电镜下可见肾小管上皮细胞的线粒体明显损伤,说明顺铂肾损害大鼠模型制作成功。PNS干预可使大鼠血清BUN、Scr和尿NAG的水平显著低于顺铂模型组（P〈0.05）,肾小管上皮细胞的线粒体损害程度较顺铂模型组明显改善,提示PNS对其有保护作用。筛选出顺铂模型组与正常对照组肾组织差异表达的蛋白质20个,其中7个蛋白质在顺铂模型组中的表达下调2倍以上;顺铂模型组与PNS治疗组肾组织差异表达的蛋白质18个,其中11个蛋白质在PNS治疗组中的表达下调;有6个共同的差异表达蛋白质在顺铂模型组较正常对照组的表达上调或下调,在PNS治疗组可回调到接近正常对照组水平。差异表达蛋白质m/z 10815.42被鉴定为线粒体热休克蛋白,m/z 16021.67被鉴定为血红蛋白β1亚基、血红蛋白β2亚基。结论 顺铂肾损害可伴随多种蛋白质的表达变化,这些差异表达蛋白质可能与顺铂损害肾脏以及PNS的保护作用有关。通过对其分离和鉴定,进一步了解其性质,将有助于全面、系统地探讨顺铂肾损害以及PNS保护作用的机制。     

NS-398联合表阿霉素对肝癌HepG2细胞增殖及survivin表达的影响

[目的]探讨表阿霉素联合COX-2抑制剂NS-398对HepG2细胞增殖及survivin表达的影响.[方法]不同浓度表阿霉素单药或联合不同浓度NS-398分别作用于肝癌HepG2细胞株,MTT法测定表阿霉素联合NS-398对细胞生长的抑制率,RT-PCR和免疫荧光法分别检测HepG2细胞中survivin mRNA与survivin蛋白表达量的变化.[结果]单药表阿霉素对HepG2细胞有显著的抑制作用,且与NS-398联合应用后抑制作用更明显.正常无药对照组HepG2细胞中survivin蛋白和mRNA均呈强表达,表达水平分别为68.970±1.156和0.919±0.021;且在胞浆中survivin蛋白表达强于胞核.表阿霉素组HepG2细胞中survivin蛋白(47.630±0.863)和mRNA (0.749±0.014)水平均明显降低,联合组survivin蛋白(27.544±0.748)和mRNA(0.481±0.012)水平降低更显著(F=1065.233,P＜0.05;F=304.553,P＜0.05).[结论]NS-398联合表阿霉素可明显抑制HepG2细胞生长,NS-398可显著增强表阿霉素的抗癌作用;两者联合可明显下调HepG2细胞株中survivin蛋白和mRNA的表达.     

基于TCGA数据库筛选肺腺癌顺铂耐药的分子标志物及功能验证

目的 探究在肺腺癌顺铂耐药过程中起关键调控作用的相关基因。方法 利用生物信息学方法在TCGA数据库和GDSC数据库下载肺腺癌患者顺铂敏感组与耐药组之间的差异表达基因,对差异基因进行GO功能分析和KEGG通路富集分析,构建蛋白质互作网络,并进行层次聚类分析,筛选得到关键基因并利用实时荧光定量PCR和ELISA法在细胞水平进行验证。通过siRNA沉默A549/DDP细胞中CXCL10基因的表达并检测其对顺铂的敏感度。结果 筛选得到178个差异表达基因,经过聚类分析最终获得肺腺癌顺铂耐药的关键基因CXCL9、CXCL10、NKX2-1以及SFTPA1。暂时选择CXCL10进行后续验证及功能实验。实时荧光定量PCR结果显示CXCL10在A549/DDP细胞中的mRNA表达量显著高于A549细胞（P<0.001）,A549/DDP细胞上清液中CXCL10蛋白的表达量高于A549细胞,均与生物信息学预测一致。MTT结果显示沉默CXCL10表达后,A549/DDP细胞对DDP的敏感度增加。结论 CXCL10是调控肺腺癌顺铂耐药的关键基因,下调CXCL10表达可成为逆转肺腺癌顺铂耐药的潜在靶点。     

Inhibition of COX-2 and activation of peroxisome proliferator-activated receptor γ synergistically inhibits proliferation and induces apoptosis of human pancreatic carcinoma cells

Although inhibition of cyclooxygenase-2 (COX-2) or activation of peroxisome proliferators-activated receptor γ (PPAR-γ) leads to growth inhibition in malignancies, the synergistic anti-tumor effects of combination of COX-2 inhibitor (NS-398) and PPAR-γ agonist (rosiglitazone) on the human pancreatic cancer cells remains unknown. Here, we evaluated the effects of NS-398 and/or rosiglitazone on the cell proliferation and apoptosis in a pancreatic cancer cell line, SW1990. NS-398 and rosiglitazone decreased cell proliferation in a dose- and time-dependent manner. Proliferating cell nuclear antigen (PCNA) labeling index significantly decreased in the cells treated with either NS-398 or rosiglitazone. Both NS-398 and rosiglitazone alone induced apoptotic cell death of SW1990. The combination of NS-398 and rosiglitazone exerted synergistic effects on proliferation inhibition, and apoptosis induction in SW1990 cells, with down-regulation of Bcl-2 and up-regulation of Bax expression. Our results indicate that simultaneous targeting of COX-2 and PPAR-γ inhibits pancreatic cancer development more effectively than targeting each molecule alone.     

Effect of non-steroidal anti-inflammatory drugs on colon carcinoma Caco-2 cell responsiveness to topoisomerase inhibitor drugs

Numerous studies demonstrate that the chemopreventive effect of non-steroidal anti-inflammatory drugs on colon cancer is mediated through inhibition of cell growth and induction of apoptosis. For these effects non-steroidal anti-inflammatory drugs have been recently employed as sensitising agents in chemotherapy. We have shown previously that treatments with aspirin and NS-398, a cyclo-oxygenase-2 selective inhibitor, affect proliferation, differentiation and apoptosis of the human colon adenocarcinoma Caco-2 cells. In the present study, we have evaluated the effects of aspirin and NS-398 non-steroidal anti-inflammatory drugs on sensitivity of Caco-2 cells to irinotecan (CPT 11) and etoposide (Vp-16) topoisomerase poisons. We find that aspirin co-treatment is able to prevent anticancer drug-induced toxicity, whereas NS-398 co-treatment poorly affects anticancer drug-induced apoptosis. These effects correlate with the different ability of aspirin and NS-398 to interfere with cell cycle during anticancer drug co-treatment. Furthermore, aspirin treatment is associated with an increase in bcl-2 expression, which persists in the presence of the anticancer drugs. Our data indicate that aspirin, but not NS-398, determines a cell cycle arrest associated with death suppression. This provides a plausible mechanism for the inhibition of apoptosis and increase in survival observed in anticancer drug and aspirin co-treatment.     

托瑞米芬协同顺铂对人肺癌细胞株A549的影响

目的：研究托瑞米芬（TOR）对人肺腺癌细胞系A549的毒性作用及其与顺铂（DDP）联用的协同效应,探讨肺癌综合治疗的方向。方法：用MTT显色法检测TOR及与DDP联用后对A549细胞的毒性作用,测定其吸光度（A）值。用流式细胞仪检测细胞DNA含量,Western blot法检测p21蛋白表达。结果：TOR能直接抑制A549细胞的生长,≥5μmol/L的TOR可明显增强DDP的细胞毒性作用。TOR可加强DDP对S期、G2期及M期细胞的作用,且DDP＋TOR后p21蛋白表达增加。结论：≥5μmol/L的TOR与DDP联用对A549细胞具有显著的协同抗肿瘤效应。     

Dose-dependent modulation of apoptosis and cyclooxygenase-2 expression in human 1547 osteosarcoma cells by NS-398, a selective cyclooxygenase-2 inhibitor

A selective cyclooxygenase-2 (COX-2) inhibitor, NS-398, was shown to produce an anti-proliferative and pro-apoptotic effect on different types of cell lines. We describe the presence of COX-1 and COX-2 pathways in the human osteosarcoma 1547 cell line, as well as the conflicting effects of NS-398 (10, 50 and 100 microM) on programmed cell death, PGE2 release and COX-2 expression in 1547 cells cultured under apoptotic conditions. We demonstrate a link between the effects of 10 and 100 microM NS-398 on cell apoptosis, PGE2 release, and expression of COX-2 in 1547 cells undergoing apoptosis. At 10 microM, NS-398 acted as a selective COX-2 inhibitor moderately increasing apoptosis without any effect on COX-2 expression. In contrast, at 100 microM, NS-398 induced a cell cycle slowing or arrest, strongly enhanced COX-2 expression which was associated with a high PGE2 release and a marked decrease in apoptosis. This latter property of NS-398 at 100 microM in 1547 human osteosarcoma cells is novel compared to the described NS-398 pro-apoptotic effect on other cell lines.     

Cyclooxygenase-2 inhibitors demonstrate anti-proliferative effects in oesophageal cancer cells by prostaglandin E(2)-independent mechanisms

The incidence of oesophageal cancer (OC) has risen in recent decades, with survival rates remaining poor despite surgical treatment and adjuvant chemotherapy. Studies have reported cyclooxygenase-2 (COX-2) overexpression in OC and current evidence suggests NSAIDs have major potential for chemoprevention through COX-2 inhibition. However, several reports have questioned the specificity of these inhibitors, suggesting they may act through mechanisms other than COX-2. We evaluated the effects of specific COX-2 inhibitors, NS-398 and nimesulide, on cell lines of both histological types of OC. COX-2 protein expression varied in the cell lines and corresponded with levels of prostaglandin E(2) (PGE(2)) production. Following treatment with low concentrations of NS-398 (0.1 microM), PGE(2) production was reduced dramatically, indicating inhibition of COX-2 activity. Examination of cellular morphology, caspase-3 activity and mitochondrial membrane integrity found no major induction of apoptotic cell death at concentrations below 100 microM. Tumour cell proliferation was significantly reduced at high concentrations (50-100 microM) of both inhibitors over 6 days. Cellular responses were more evident in NS-398-treated adenocarcinoma cells. However, concentrations required to inhibit proliferation were up to 1000-fold higher than those needed to inhibit enzyme activity. Addition of exogenous PGE(2) to NS-398-treated adenocarcinoma cells failed to reverse the inhibitory effects, indicating PG and COX-2 independence. It remains possible that in vivo COX-2 is the primary target, as enzyme inhibition can be achieved at low concentrations, however, inhibition of proliferation is not the primary mechanism of their anti-tumour activity.     

ＮＳ-３９８对乳腺癌细胞ＭＣＦ-７增殖和凋亡的影响及其作用机制

目的　探讨环氧合酶-２（ＣＯＸ-２）选择性抑制剂ＮＳ-３９８对乳腺癌细胞ＭＣＦ-７增殖和凋亡的影响及其可能的作用机制。方法　用不同浓度ＮＳ-３９８作用ＭＣＦ-７细胞后,采用四甲基唑蓝法（ＭＴＴ）检测ＮＳ-３９８对ＭＣＦ-７细胞增殖的抑制率;流式细胞仪（ＦＣＭ）检测细胞周期、细胞凋亡以及ＣＯＸ-２、Ｂｃｌ-２和Ｂａｘ蛋白的表达;用放射免疫分析（ＲＩＡ）检测培养液上清中前列腺素Ｅ２（ＰＧＥ２）含量。结果　ＮＳ-３９８可显著抑制ＭＣＦ-７细胞的生长,并随药物的浓度升高、时间延长其抑制作用增强。ＮＳ-３９８使ＭＣＦ-７Ｇ_０／Ｇ_１期细胞显著增多（Ｐ＜０.０１）,Ｓ期及Ｇ_２／Ｍ期细胞显著减少（Ｐ＜０.０１）,并引起显著的细胞凋亡。ＮＳ-３９８使ＭＣＦ-７细胞ＣＯＸ-２和Ｂｃｌ-２表达显著降低（Ｐ＜０.０１）,而Ｂａｘ表达显著增高（Ｐ＜０.０１）,并使ＭＣＦ-７细胞产生的ＰＧＥ２显著降低（Ｐ＜０.０１）。结论　ＮＳ-３９８可抑制乳腺癌细胞ＭＣＦ-７的增殖并可诱导其凋亡,其作用机制可能通过抑制ＣＯＸ-２活性,使ＰＧＥ_２产量降低,从而下调Ｂｃｌ-２表达及激活Ｂａｘ表达。     

NS-398对人胃癌细胞系SGC-7901增殖、凋亡和COX-2表达的影响

目的 探讨环氧合酶-2(COX-2)抑制剂NS-398对人胃癌细胞系SGC-7901增殖、凋亡及COX-2表达的影响,并进一步探讨其作用的可能机制.方法 采用四甲基偶氮唑蓝(MTT)法检测NS-398对SGC -7901细胞的杀伤抑制作用;免疫细胞化学法检测SGC-7901细胞内COX-2的蛋白表达情况;ELISA法检测NS-398作用于SGC-7901细胞后PGE2释放水平;流式细胞仪检测SGC-7901细胞的凋亡情况.结果 NS-398对胃癌SGC-7901细胞具有较强的抑制作用,且这种抑制作用随浓度和时间的增加而增强,呈剂量-时间双效应关系(P＜0.05);不同浓度NS-398作用下的SGC-7901细胞中,COX-2的表达明显减弱,且呈剂量梯度下降(P＜ 0.05);NS-398可抑制PGE2释放,并且这种抑制作用呈剂量效应关系,与对照组相比差异有统计学意义(P＜ 0.05);NS-398作用于SGC-7901细胞48 h后,细胞凋亡率升高,且呈剂量效应(P＜0.05).结论 NS-398通过COX-2依赖途径抑制SGC-7901细胞增殖并促进其凋亡.     

Cyclooxygenase-2 inhibitor NS-398 suppresses cell growth and constitutive production of granulocyte-colony stimulating factor and granulocyte macrophage-colony stimulating factor in lung cancer cells

We previously established two lung cancer cell lines, OKa-C-1 and MI-4, which constitutively produce abundant granulocyte-colony stimulating factor (G-CSF) and granulocyte macrophage-colony stimulating factor (GM-CSF). Inflammatory cytokines, tumor necrosis factor-alpha (TNF-alpha) and interleukin (IL)-1beta stimulated the expression of G-CSF, GM-CSF, and cyclooxygenase (COX)-2 in the two cell lines. It is known that increased COX-2 activity promotes tumor growth and induces G-CSF and GM-CSF expression in non-malignant cells, and that selective COX-2 inhibitors inhibit the growth of some types of malignant cells. Therefore, we hypothesized that inhibition of COX-2 activity might suppress constitutive production of G-CSF or GM-CSF in addition to reducing the growth of malignant cells. We confirmed that the selective COX-2 inhibitor, NS-398 suppressed the constitutive production of G-CSF and GM-CSF, and the cell growth in both OKa-C-1 and MI-4 cell lines. Prostaglandin E2 (PGE2) reversed the inhibitions of G-CSF and GM-CSF expression, as well as cell growth, by NS-398. This result confirms that the effects of NS-398 are based on the inhibition of COX activity. Some studies have indicated that nuclear factor kappa B (NF-kappaB) or MAPK (mitogen-activated protein kinase) activation is related to upregulation of G-CSF, GM-CSF or COX-2 expression in some types of cells. Therefore, we examined if the actions of NS-398 might be mediated by the MAP kinase pathway or NF-kappaB activity in OKa-C-1 and MI-4 cells. We found that NS-398 inhibits G-CSF and GM-CSF production and cell growth through an extracellular signal-regulated kinase kinase (MEK) signaling pathway in these cell lines. The prognosis of non-small cell lung cancer showing G-CSF gene expression is significantly worse. G-CSF overproduction by tumor cells is observed at an advanced clinical stage. Our findings imply that a COX-2 inhibitor might improve the prognosis of patients with lung cancer through the reduction of G-CSF or GM-CSF.     

The effect of non-steroidal anti-inflammatory drugs on human colorectal cancer cells: evidence of different mechanisms of action

Non-steroidal anti-inflammatory drugs (NSAIDs) inhibit proliferation and induce apoptosis in human colorectal cancer cells in vitro. It remains unclear whether individual NSAIDs act by cyclooxygenase-2 (COX-2) inhibition and how NSAIDs exert their anti-proliferative effects. We investigated the effects of NS-398 (a selective COX-2 inhibitor), indomethacin (a non-selective COX inhibitor) and aspirin on four human colorectal cancer cell lines (HT29.Fu, HCA-7, SW480 and HCT116). NS-398 completely inhibited proliferation, induced G1 arrest and promoted apoptosis in COX-2-expressing cells (HT29.Fu and HCA-7). However, indomethacin had similar effects on all cells, regardless of COX-2 expression. NS-398 also had anti-proliferative activity on COX-2-negative cell lines (SW480 and HCT116). Aspirin inhibited proliferation of all cell lines but did not induce apoptosis. Indomethacin decreased beta-catenin protein expression in all cells (unlike NS-398 or aspirin). NSAIDs act on human colorectal cancer cells via different mechanisms. Decreased beta-catenin protein expression may mediate the anti-proliferative effects of indomethacin.