杨梅素对人肺腺癌细胞增殖的影响及其机制

目的研究杨梅素(myricetin,Myr)对人肺腺癌(A549)细胞增殖的影响及其作用机制。方法以人肺腺癌系A549细胞为离体研究对象,通过噻唑蓝(MTT)法研究不同浓度的Myr对A549细胞生长的抑制作用并测定其半数抑制浓度(IC50);采用Westernblot法检测Myr对蛋白激酶B(Akt)磷酸化水平的影响。结果Myr对A549细胞具有明显的增殖抑制作用,随着Myr浓度的增加,细胞的存活率明显降低。Myr作用72h的IC50值为41.7μg/ml。32μg/ml杨梅素无血清作用A54912h即可显著抑制AktSer473的磷酸化水平。结论Myr能够剂量依赖性抑制A549细胞增殖,Akt的活性下调可能是Myr体外诱导人A549肺腺癌细胞增殖抑制作用的分子靶点。     

百草枯诱导A549细胞间质转化及四氢吡咯二硫代氨基甲酸酯的干预作用

目的探讨体外培养的人肺腺癌细胞(A549)在百草枯(PQ)诱导下能否发生上皮-间质转化(epithelial-mesenchymal transition,EMT)及四氢吡咯二硫代氨基甲酸酯(PDTC)的干预作用和可能机制。方法体外培养A549细胞,实验分为对照A549细胞组(Control)、PQ诱导A549细胞组(PQ)、PDTC 3个浓度干预组(PQ+PDTC1,2,3;终浓度分别为10,20,40μmol/L)。用噻唑蓝(MTT)测定细胞毒性;活性氧荧光定量法(DCFH-DA)测定细胞内活性氧(ROS)水平;酶联免疫吸附试验(ELISA)测定转化生长因子β1(TGF-β1)表达水平;实时定量RCR(Real Time,RT-PCR)测定EMT相关标志物上皮细胞钙粘蛋白(Ecad)、紧密连接蛋白(ZO-1)、N-钙粘蛋白(N-cad)和α-平滑肌肌动蛋白(α-SMA)mRNA表达。结果与对照组比较,PQ(600μmol/L)作用A549细胞24h后,细胞存活率降低,细胞内ROS水平明显升高,TGF-β1表达水平明显升高,上皮细胞标志物E-cad、ZO-1基因表达下调,间质细胞标志物N-cad、α-SMA基因表达明显上调(P<0.05);PDTC干预组能够降低PQ诱导的A549细胞毒性,明显抑制上述作用,其中对TGF-β1表达、E-cad、N-cad基因表达的干预作用呈剂量依赖方式(P<0.05)。结论PQ诱导A549细胞过度表达TGF-β1,继而导致上皮细胞向间质细胞转化,这可能是PQ致肺纤维化的重要机制之一;PDTC的有效干预可能基于其抗氧化和核因子-kappaB(nuclear factor-kappaB,NF-κB)抑制作用。     

K2Cr2O7染毒对脱氢抗坏血酸预处理的A549细胞hMLH1和p53 mRNA基因表达的影响

目的 研究K2Cr2O7对A549细胞hMLH1和p53mRNA基因表达的影响.方法 体外培养A549细胞,实验组用脱氢抗坏血酸(DHA)孵育90min以模拟体内环境,对照组常规培养,然后分别用0.00、1.25、2.50和5.00 μol/L的K2Cr2O7溶液染毒细胞,用实时聚合酶链反应(Real-time PCR)检测细胞内hMLH1及p53mRNA的表达.结果 K2Cr2O7可剂量依赖性地降低A549细胞hMLH1和p53mRNA的表达;DHA预处理可进一步降低染毒A549细胞hMLH1的表达.结论 K2Cr2O7染毒可降低体内条件下A549细胞hMLH1和P53mRNA的表达,从而参与肺癌的细胞癌变过程.     

“卡宁”抗骨肿瘤的机理初探

目的体外观察抗癌中药“卡宁”对人成骨肉瘤细胞株MG-63的凋亡诱导基因fas影响,探讨其抗肿瘤治疗的作用机制。方法采用MTT比色法观察含药血清对体外培养的人骨肿瘤细胞株的抑制能力,PCR检测对凋亡诱导基因fas的影响。结果体外实验显示中药组fas的活性明显高于对照组(p=0.038),MTT结果显示细胞生长抑制率与含药血清作用时间及浓度呈正相关,p<0.05,在15%含药血清72h时作用最强。体外实验表明,“卡宁”对MG-63细胞的fas活性有促进作用。结论对肿瘤细胞fas的促进可能是抗癌中药“卡宁”诱导肿瘤细胞凋亡作用机制的一部分。     

大豆异黄酮联合顺铂对A549细胞增殖和凋亡的影响

目的探讨大豆异黄酮(ISF)联合顺铂(DDP)化疗对A549肺癌细胞增殖和凋亡的影响。方法采用噻唑蓝(MTT)法观察ISF和DDP联用对A549肺癌细胞增殖的影响,并计算联合指数(Q值)判定联合用药的相互作用,同时采用TUNEL染色法检测二者对A549细胞凋亡的影响。结果 ISF和DDP单独使用均可时间和浓度依赖性地抑制肺癌A549细胞的增殖。DDP浓度在3 mg/L时,ISF和DDP联用后抗细胞增殖作用增强,二者有相加或协同作用,且在一定浓度范围内,联合用药对细胞生长的抑制作用呈量效关系。ISF单用或和DDP联用均可浓度依赖性地诱导肺癌细胞凋亡。结论 ISF可通过诱导A549细胞凋亡而发挥抗肺癌作用;ISF和DDP联用可增强对肺癌A549细胞增殖的抑制和凋亡的诱导,二者具有协同抗肿瘤作用。     

肉桂醛对肺癌细胞A549具有体外抑制作用

目的探讨肉桂醛对肺癌细胞株A549的生长抑制作用。方法采用噻唑蓝比色法检测肉桂醛对体外培养的人肺癌细胞系A-549 细胞株的生长抑制作用,计算半数抑制浓度。结果肉桂醛能抑制人肺癌细胞系A-549 细胞增殖,且呈剂量依赖性,IC50值为0.36 mg/ml。结论在体外,肉桂醛对肺癌细胞株A549具有明显抗肿瘤活性。     

羟基喜树碱对肺癌细胞A549抑制作用的研究

目的:研究羟基喜树碱对人肺癌A549细胞的抑制效果,探讨抗癌药物对细胞凋亡与周期的调控作用.方法:CCK-8法测定不同浓度的HCPT对A549细胞的生长抑制作用,AO/EB染色观察细胞生长情况;琼脂糖凝腔电泳验证细胞调亡的特征性DNA条带;用流式细胞仪测定抗癌药物羟基喜树碱对细胞凋亡和细胞周期的影响.结果:CCK-8结果表明不同浓度的HCPT对A549细胞的生长抑制作用不同;琼脂糖凝胶电泳可见调亡细胞DNA的梯状条带;荧光显微镜下观察到经AO/EB染色的典型凋亡细胞;在相同浓度下,随时间的延长细胞的凋亡率也逐渐增加,1μmol/l的HCFT作用于A549细胞对其抑制效果最为明显,细胞凋亡率为32.03%.结论:抗癌药物羟基喜树碱对A549有抑制效果,促进其细胞凋亡,参与细胞周期调控.     

MMP-9ASODN对肺腺癌A549细胞增殖与凋亡的影响及其机制初步探讨

目的 探讨基质金属蛋白酶-9反义寡核苷酸(MMP-9ASODN)对肺腺癌A549细胞增殖与凋亡的影响,并初步探讨其机制.方法 MTT法检测MMP-9ASODN转染对A549细胞生长增殖的影响;流式细胞术检测MMP-9ASODN转染对A549细胞周期和凋亡比率的影响;RT-PCR法与Western blot法分别检测MMP-9ASODN转染对A549细胞内MMP-9mRNA及蛋白表达的影响.结果 在一定范围内,MMP-9ASODN对A549细胞生长抑制呈浓度和时间依赖性,反叉寡核苷酸浓度为600 nmo/L作用48 h时其抑制作用最明显;MMP-9ASODN转染A549细胞48 h后G1期细胞数明显增多,S期细胞数明显减少,G2期细胞数则无明显变化,其凋亡百分比明显升高;与对照组比较差异有统计学意义(P＜0.01);细胞内MMP-9mRNA及其蛋白的相对表达量明显低于对照组(P＜0.01).结论 MMP-9ASODN转染能够有效能够抑制肺腺癌A549细胞的增殖同时诱导其凋亡,其机制可能是通过下调MMP-9mRNA及蛋白的表达.     

体外及活体环境下蜈蚣提取物联合顺铂对人肺癌A549细胞的影响

目的 观察蜈蚣提取物(ECP)与顺铂(DDP)联合运用对人肺癌A549在体外及活体环境下的影响.方法 体外环境下人肺癌A549细胞培养;活体采用裸鼠皮下移植瘤进行试验.分别观察单用ECP或DDP以及两者联用对A549细胞的作用.结果 DDP联用ECP与ECP联用DDP,对A549细胞的抑制作用比单用ECP或DDP时分别增效1.80倍和2.60倍,两药的合并指数为0.93.DDP联用ECP对抑制移植瘤的体积变化与单用DDP比较,差异有统计学意义(P<0.05).结论 蜈蚣提取物与顺铂联用在体外及活体环境下均有协同作用;两者联用的作用优于单用.     

纳米与微米二氧化钛对A549细胞毒性的比较

目的 比较纳米二氧化钛和微米二氧化钛对体外培养的人肺上皮细胞A549的毒性作用.方法 用不同浓度的纳米(粒径4 nm)和微米(粒径1μm)二氧化钛处理体外培养的A549细胞,通过倒置相差显微镜观察其形态,噻唑蓝(MTT)、CCK-8和台盼蓝排斥法测定细胞生长活性,并测定培养液上清中乳酸脱氢酶活力.结果 MTT法检测发现随着剂量的增加,微米二氧化钛组细胞存活率逐渐下降,与对照组比较,在剂量为200 μg/ml时,差异有统计学意义(P＜0.05),纳米二氧化钛组未见明显影响.CCK-8法、台盼蓝排斥法及乳酸脱氢酶活力测定结果表明纳米与微米二氧化钛组均未见明显影响.结论 在本实验条件下,纳米二氧化钛和微米二氧化钛对A549细胞没有显示明显的细胞毒性作用.     

Inhibitory Effect of Liquiritigenin on Migration Via Downregulation ProMMP-2 and PI3K/Akt Signaling Pathway in Human Lung Adenocarcinoma A549 cells

Liquiritigenin (LQ) is a flavanone extracted from Glycyrrhizae, which has multiple biological effects, such as antiinflammation and anticancer. This study is the first to investigate the effect of LQ on the migration of human lung adenocarcinoma A549 cells in vitro. First, LQ exhibited inhibitory effects on the adhesion and migration of A549 cells in the absence of cytotoxicity. Gelatin zymography and Western blot analysis showed that LQ significantly reduced the expression of promatrix metalloproteinase-2 (proMMP-2) in A549 cells in terms of both activity and protein level. Second, LQ inhibited the phosphorylation of Akt and activated the phosphorylation of extracellular signal-regulated kinase 1 and 2 (ERK1/2). Furthermore, the treatment of inhibitors specific for Akt (LY294002) and ERK1/2 (U0126) to A549 cells resulted in reduced activity of proMMP-2. These results suggested that the inhibition on proMMP-2 expression by LQ may be through suppression on PI3K/Akt signaling pathway, which in turn led to the inhibition of lung adenocarcinoma A549 cells migration. However, activation of ERK might not be involved in the regulation of proMMP-2. Taken together, LQ may be considered as a potential interfering agent of cancer progression.     

Inhibitory effect of liquiritigenin on migration via downregulation proMMP-2 and PI3K/Akt signaling pathway in human lung adenocarcinoma A549 cells

Liquiritigenin (LQ) is a flavanone extracted from Glycyrrhizae, which has multiple biological effects, such as antiinflammation and anticancer. This study is the first to investigate the effect of LQ on the migration of human lung adenocarcinoma A549 cells in vitro. First, LQ exhibited inhibitory effects on the adhesion and migration of A549 cells in the absence of cytotoxicity. Gelatin zymography and Western blot analysis showed that LQ significantly reduced the expression of promatrix metalloproteinase-2 (proMMP-2) in A549 cells in terms of both activity and protein level. Second, LQ inhibited the phosphorylation of Akt and activated the phosphorylation of extracellular signal-regulated kinase 1 and 2 (ERK1/2). Furthermore, the treatment of inhibitors specific for Akt (LY294002) and ERK1/2 (U0126) to A549 cells resulted in reduced activity of proMMP-2. These results suggested that the inhibition on proMMP-2 expression by LQ may be through suppression on PI3K/Akt signaling pathway, which in turn led to the inhibition of lung adenocarcinoma A549 cells migration. However, activation of ERK might not be involved in the regulation of proMMP-2. Taken together, LQ may be considered as a potential interfering agent of cancer progression.     

The inhibitory effect of dibutyryl cyclic AMP on docosahexaenoic acid-induced apoptosis in HL-60 cells through activation of the phosphatidylinositol-3 kinase pathway

Objective Docosahexaenoic acid (DHA) is known as a chemopreventive substance for cancers. Previously we reported that DHA induces apoptosis in HL-60 cells. The aim of this study was to clarify the role of phosphatidylinositol 3-kinase (PI3-kinase)/Akt signaling during DHA-induced apoptosis in HL-60 cells. Methods The inhibitory effects of dibutyryl cAMP (db-cAMP) or LY294002 (a specific inhibitor of the PI3-kinase/Akt pathway) on DHA-induced apoptosis in HL-60 cells were evaluated by the appearance of apoptosis, and from the activities of caspases (3 and 8), the phospholylation of Akt, and cleavage of Bid using DNA indexes, emzymatic measurement of fragmented substrates, and Western blotting, respectively. Results The pre-incubation of db-cAMP reduced the activation of caspasses (3 and 8) during the occurrence of DHA-induced apoptosis in HL-60. However, the inhibition of PI3-kinase/Akt signaling by LY294002 resulted in recovery of the caspases’ activities, appearance of apoptotic cells, and cleavage of the Bid molecule when LY294002 was co-treated with db-cAMP before the occurrence of DHA-induced apoptosis in HL-60. It was also confirmed that LY294002 strongly inhibited phospholylation of Akt during db-cAMP induced-reduction of DHA-induced apoptosis in HL-60. Conclusion We demonstrated that DHA-induced apoptosis was sensitive to the modulation of PI3-kinase activity by treatment with db-cAMP or LY294002. These results may provide new insights into the mechanisms of the anti-cancer activity of DHA.     

玉米紫色植株色素体外诱导人肺腺癌A549细胞凋亡的初步研究

[目的]研究玉米紫色植株色素(PMPP)对人肺腺癌A549细胞增殖、凋亡、细胞周期的影响。[方法]MTT法比较不同浓度的PMPP对体外培养的肺癌细胞增殖的影响,流式细胞术检测细胞凋亡率和细胞周期,同时应用倒置显微镜观察凋亡细胞的形态学特征。[结果]不同浓度PMPP溶液对A549细胞的增殖均有抑制作用,浓度为50mg/ml时的抑制率最大,约为85.35%。形态学表现为单位视野内细胞数量明显减少,细胞间隙增大,细胞体积明显缩小,细胞失去原有的形态,胞体皱缩变圆,出现膜发泡现象。流式细胞术结果表明,当PMPP浓度为40mg/ml时,处于S期的肿瘤细胞增多,凋亡率为41.52%。[结论]玉米紫色植株色素可通过诱导部分细胞凋亡抑制人肺腺癌A549细胞增殖。     

白藜芦醇抑制IGF-I促人肺腺癌A549细胞增殖作用及其机制的研究

目的观察白藜芦醇对IGF-I介导人肺腺癌细胞增殖的影响及其IGF-I受体、AKT、p-AKT、ERK1/2、p-ERK1/2的表达变化。方法应用MTT方法测定细胞增殖,采用RT-PCR检测IGF-I受体mRNA表达,并应用免疫印迹方法检测AKT、p-AKT及ERK1/2、p-ERK1/2蛋白表达水平。结果不同浓度IGF-I(2.5～15nM)可显著促进A549细胞增殖(F=22.321,P=0.011),白藜芦醇(6.25～50μM)与10nMIGF-I共作用于A549细胞,均可抑制IGF-I的促A549细胞增殖作用(F=11.211,P=0.032),其中尤以25μM、50μM白藜芦醇的抑制增殖作用显著(P=0.034;P=0.012),并且,白藜芦醇可有效降低IGF-I受体mRNA表达以及降低IGF-I增强的AKT及ERK1/2磷酸化。结论白藜芦醇能够抑制IGF-I介导的A549肺癌细胞增殖,其作用机制可能与白藜芦醇降低A549肺癌细胞IGF-I受体mRNA表达及AKT、ERK1/2磷酸化相关。     

Carbon ion irradiation suppresses metastatic potential of human non-small cell lung cancer A549 cells through the phosphatidylinositol-3-kinase/Akt signaling pathway

We previously showed that carbon ion irradiation can inhibit the expression of the anillin (ANLN) gene, which is regulated by the activation of the phosphatidylinositol-3-kinase (PI3K)/Akt signaling pathway associated with metastasis. The purpose of this study is to compare the effects of carbon ion irradiation on the PI3K/Akt signaling pathway to those of photon irradiation. Our study showed that carbon ion irradiation of human lung adenocarcinoma cells A549 decreased their invasion more effectively than photon irradiation did. We found that carbon ion irradiation reduced the nuclear localization of ANLN at lower dose, but did not affect its expression. Low-dose carbon ion irradiation also reduced the level of phosphorylated Akt compared to untreated controls, whereas photon irradiation did not. These results suggest that carbon ion irradiation effectively suppresses the metastatic potential of A549 cells by suppressing the PI3K/Akt signaling pathway.     

多烯脂肪酸对海马神经元细胞脂肪酸构成和生长的作用

目的 :　观察长链多不饱和脂肪酸花生四烯酸 (AA)和 /或二十二碳六烯酸 (DHA)对体外培养新生大鼠海马神经元脂肪酸构成和生长发育的作用。方法 :　采用无血清培养液培养新生大鼠海马神经元 ,实验组分别在对照组的无血清培养液中添加 4μmol/L AA、4μ mol/L DHA或总浓度为 4μmol/L、比例为 1∶ 2～ 1 6∶ 1的 AA和 DHA。采用毛细管气相色谱法分析神经元中脂肪酸的构成 ,NSE染色鉴定神经元并利用图像分析技术 ,测量神经元胞体大小和突起长度。结果 :　当培养液中 AA和 DHA总浓度为 4μmol/L、比例为 1∶ 2～ 1 6∶ 1 ,海马神经元中 AA百分含量、AA/DHA比值与培养液中 AA和 DHA比例均呈正相关 ;当培养液中 AA和 DHA总浓度为4μmol/L,比例为 2∶ 1或 4∶ 1时 ,海马神经元的胞体面积、最大长径、最大短径和平均突起长度均显著高于单一添加 4μ mol/L AA、4μmol/L DHA、对照组以及其余各比例组。结论 :　 AA和DHA具有显著促进体外培养海马神经元生长发育的作用     

二十二碳六烯酸对神经细胞生长发育的作用

目的 :　观察二十二碳六烯酸 ( DHA)对神经细胞生长发育的作用。方法 :　将无血清培养的新生大鼠大脑神经细胞分为实验组和对照组 ,实验组加入 0 .33mol/ L( 5mol/ 1 5L)乳化DHA40 μl。对两组细胞进行神经细胞活力、蛋白质含量、神经特异性烯醇化酶 ( NSE)活性测定及神经细胞形态学观察。结果 :　实验组大脑神经细胞突起生长速度、胞体面积和直径、神经细胞存活率、NSE和神经细胞活力及蛋白含量均显著高于对照组。结论 :　DHA对神经细胞的生长有促进作用 ,且这一作用可能与促进神经细胞蛋白质合成有关。     

Saponins from the Roots of Platycodon grandiflorum Suppresses TGFβ1-Induced Epithelial-Mesenchymal Transition Via Repression of PI3K/Akt,ERK1/2 and Smad2/3 Pathway in Human Lung Carcinoma A549 Cells

Transforming growth factor β (TGFβ) is a multifunctional cytokine that induces growth arrest, tissue fibrosis, and epithelial-mesenchymal transition (EMT) through activation of Smad and non-Smad signaling pathways. EMT is the differentiation switch by which polarized epithelial cells differentiate into contractile and motile mesenchymal cells. Our previous studies have shown that saponins from the roots of Platycodon grandiflorum (CKS) have antiinflammatory, antioxidant, antimetastatic, and hepatoprotective effects. In this study, we investigated the inhibitory effect of CKS on TGFβ1-induced alterations characteristic of EMT in human lung carcinoma A549 cells. We found that CKS-treated cells displayed inhibited TGFβ1-mediated E-cadherin downregulation and Vimentin upregulation and also retained epithelial morphology. Furthermore, TGFβ1-increased Snail expression, a repressor of E-cadherin and an inducer of the EMT, was reduced by CKS. CKS inhibited TGFβ1-induced phosphorylation of Akt, ERK1/2, and glycogen synthase kinase-3β (GSK-3β). Inhibition of PI3K/Akt and ERK1/2 also blocked TGFβ1-induced GSK-3β phosphorylation and Snail activation. Furthermore, TGFβ1-increased Snail expression was reduced by selective inhibitors of Akt and ERK1/2. Moreover, CKS treatment attenuated TGFβ1-induced Smad2/3 phosphorylation and upregulated Smad7 expression. These results indicate that pretreatment with the CKS inhibits the TGFβ1-induced EMT through PI3K/Akt, ERK1/2, GSK-3β and Smad2/3 in human lung carcinoma cells.