Fas信号通路促进胃癌细胞的侵袭及转移

目的探讨Fas通路的非凋亡信号在胃癌细胞株AGS中促进增殖转移的作用。方法流式细胞仪检测AGS细胞表面Fas及FasL表达情况;根据FasL不同作用浓度梯度,绘制细胞生长曲线;shRNA沉默AGS的Fas表达后,或NOK-1拮抗FasL后,利用transwell实验检测AGS的侵袭能力;免疫组化检测FasL表达与E-cadherin关系。结果低浓度的FasL（12.5ng/ml）刺激对细胞增殖无明显抑制作用,反而能提高肿瘤细胞的侵袭能力。在沉默Fas受体或经NOK-1拮抗FasL后,该效应减弱。FasL和E-cadherin在胃癌组织中的表达呈负性相关。结论低剂量的FasL不能诱导AGS产生凋亡效应,反而通过Fas信号通路促进其侵袭转移能力。     

GSK-3β通过调节 Snail 参与 Fas 诱导的EMT过程

目的：探讨ERK、GSK3β和Snail在Fas诱导的EMT中的作用。方法选用结肠癌SW480细胞及胃癌AGS细胞作为研究对象,使用U0126以阻断其ERK/MAPK通路的激活,稳定转染GSK3βS9A以抑制其GSK3βSer9位点的磷酸化及信号转导,稳定转染Snail shRNA以抑制其Snail活性,并对接受上述处理的细胞分别给予低剂量FasL刺激后,再利用侵袭试验、免疫荧光、免疫印迹、RT-PCR、qRT-PCR、免疫共沉淀及荧光素酶报告基因等方式检测细胞的形态、功能变化。结果 Fas信号通路的激活可抑制E-cadherin的转录表达,且这一过程依赖于ERK/MAPK通路。通过转染稳定敲除Snail表达后,Fas对E-cadherin的转录抑制作用显著减弱。在细胞内过表达突变型GSK3βS9A可显著降低Fas通路激活后Snail的表达上调水平、E-cadherin的转录抑制水平及细胞的侵袭能力增加程度。免疫共沉淀提示, GSK3β与ERK、Snail在细胞核存在相互作用。结论在消化道肿瘤细胞中,Fas诱导EMT的调控机制包括ERK/MAPK通路的激活、ERK对GSK3β的磷酸化（Ser9位点）失活、GSK3β失活导致的Snail表达上调及核易位,以及Snail作用下的E-cadherin转录水平下调。     

桂皮醛通过TGF-beta信号通路调控结直肠癌细胞侵袭和转移研究

目的研究桂皮醛通过转化生长因子-β(TGF-β)信号通路调控结直肠癌细胞侵袭和转移的机制。方法用不同浓度桂皮醛处理人结肠癌SW480细胞,观察SW480细胞增殖活性、细胞凋亡、侵袭、转移能力及凋亡、上皮间质转化(EMT)、TGF-β/Smad3信号途径关键蛋白表达情况;用10 ng/m L的TGF-β1处理诱导SW480细胞EMT,并用80 mg/L的桂皮醇进行干预,观察SW480细胞中EMT、TGF-β/Smad3信号途径关键蛋白表达。结果桂皮醛处理后SW480细胞Bcl-2/Bax比率、细胞侵袭率、划痕愈合率、波形蛋白(Vimentin)、N-钙黏蛋白(N-cadherin)、基质金属蛋白酶-9(MMP-9)、TGF-β、Smad3、Snail蛋白表达降低,细胞增殖抑制率、细胞凋亡率、E-cadherin蛋白表达升高,差异有统计学意义(P 0. 05);桂皮醛干预后能够缓解TGF-β1处理后诱导的Vimentin、N-cadherin、MMP-9、Smad3、Snail蛋白表达升高,E-cadherin蛋白表达降低,差异有统计学意义(P 0. 05)。结论桂皮醛可能通过抑制人结肠癌细胞系SW480中TGF-beta/Smad3信号途径来抑制细胞增殖,促进细胞凋亡,并抑制EMT,降低其侵袭转移能力。     

COX-2抑制剂NS398调控PPARs信号转导通路抑制结肠癌细胞增殖的分子机制

目的:观察选择性COX-2抑制剂NS-398对结肠癌细胞系SW480中PPARs信号转导通路的影响,以期初步阐明选择性COX-2抑制剂抗结直肠癌非COX-2依赖性途径的作用机制．方法:应用RT-PCR检测结肠癌细胞系SW480中COX-2 mRNA表达水平,用选择性COX-2抑制剂NS-398处理结肠癌细胞系SW480．Western blot检测PPARs信号转导通路成员表达,四甲基偶氮唑盐(MTT)法检测细胞增殖状态,流式细胞技术检测细胞周期与凋亡情况．结果:结肠癌细胞系SW480中未检测到COX-2 mRNA表达,NS-398(75μmol／L)作用于SW480细胞72 h后,G1期细胞比率由31．2％上升至40．6％,S期细胞比率由52．8％下降至41．2％,细胞增殖受抑制．PPARα,PPARδ,PPARγ,cyclin D1与Bcl-xl表达水平随NS-398作用时间延长而下降．结论:选择性COX-2抑制剂NS-398可能通过非COX-2依赖途径影响结肠癌细胞的增殖．     

姜黄素对结肠癌细胞SW480 FasL基因表达和侵袭能力的影响

目的研究姜黄素对人结肠癌SW480细胞FasL mRNA表达及对其侵袭能力的影响,为中药抗肿瘤提供实验依据。方法根据MTT法得到姜黄素对SW480细胞的半数有效抑制浓度（IC50）,确定药物作用浓度。SW480细胞分别经姜黄素不同浓度（0．5 IC50、IC50）作用后,应用逆转录-聚合酶链反应（RT-PCR）法检测姜黄素作用前后人结肠癌SW480细胞FasL mRNA的变化;应用Transwell细胞侵袭试验检测姜黄素对SW480细胞侵袭能力的影响。结果姜黄素处理后SW480细胞FasL mRNA表达水平均明显高于对照组（P〈0．01）;而且FasL mRNA表达水平随姜黄素作用浓度增加显著上调,姜黄素不同浓度组比较均有显著性差异（P〈0．01）;随姜黄素作用浓度升高,SW480细胞侵袭能力明显增强,不同浓度组比较均有显著性差异（P〈0．01）。结论姜黄素在一定时间内均可上调人结肠癌SW480细胞FasL mRNA的表达,而且这种上调作用在一定范围内呈剂量依赖性,可使结肠癌细胞的侵袭能力增强。     

氧化苦参碱诱导人结肠癌SW480细胞凋亡的机制

目的探讨氧化苦参碱诱导人结肠癌SW480细胞凋亡的作用及机制。方法以不同剂量的氧化苦参碱作用于人结肠癌细胞株SW480,四甲基偶氮唑蓝法(MTT)检测细胞的增殖能力,Hoechst 33258染色法观测细胞凋亡。免疫印迹法测定B淋巴细胞瘤-2(Bcl-2)、Bcl-2相关X蛋白(Bax)的表达,荧光法测定半胱氨酸蛋白酶(caspase)-3的活性。结果氧化苦参碱可剂量依赖性地抑制细胞的增殖,促进细胞凋亡。Western印迹方法显示氧化苦参碱可抑制抗凋亡蛋白Bcl-2的表达,提高促凋亡蛋白Bax的表达,同时激活凋亡效应分子caspase-3。结论氧化苦参碱能够抑制结肠癌SW480细胞的增殖,通过提高Bax的表达、抑制Bcl-2的表达、激活caspase-3发挥其诱导细胞凋亡的作用。     

TPARM对人结直肠癌细胞株SW480增殖、凋亡、侵袭及迁移的影响

目的 探讨四肽重复结构域(TTC)12,也被称为TPARM,其表达对人结肠癌细胞SW480增殖、凋亡、侵袭及迁移的影响。方法 将人结肠癌SW480细胞株分为3组：NC组、TPARM-siRNA组和Scr-siRNA组。采用实时荧光定量聚合酶链反应(qPCR)法和Western印迹法检测各组TPARM mRNA和蛋白表达。细胞计数试剂盒(CCK)8法、平板克隆法测定各组TPARM表达对SW480细胞增殖的影响。Western印迹法检测TPARM表达对各组周期相关蛋白及凋亡相关蛋白的表达。流式细胞术检测各组TPARM表达对SW480细胞凋亡的影响。Transwell小室侵袭实验和划痕实验观察TPARM表达对SW480细胞侵袭、迁移能力的影响。结果 与NC组、Scr-siRNA组比较,转染TPARM-siRNA后,SW480细胞中TPARM mRNA及蛋白表达水平均明显降低(P<0.05)。分别与NC组和Scr-siRNA组比较,TPARM-siRNA转染组SW480细胞增殖明显减少(P<0.05),周期蛋白依赖性激酶(CDK)2、CDK4、CDK6的蛋白表达明显下降(P<...     

结肠癌细胞增殖和侵袭过程中miR-92a和miR-29c的表达及其意义

目的探讨miR-92a和miR-29c表达水平对结肠癌细胞增殖和侵袭能力的影响。方法应用miR-92ainhibitor、miR-29c mimic在体外转染结肠癌细胞系SW480,分别构建miR-92a的干扰表达和miR-29c的过表达,设置转染无义序列组作为对照组。(1)采用反转录合成cDNA和荧光定量PCR,以U6表达作为内对照,检测细胞中miR-92a和miR-29c的表达水平;(2)采用CCK-8法,通过检测转染结肠癌细胞后不同时期的450nm处吸光度值,测定细胞增殖能力;(3)采用Transwell小室侵袭实验检测细胞侵袭能力,分析miR-92a和miR-29c的相对表达量与结肠癌细胞增殖和侵袭的关系。结果 (1)与对照组相比,转染miR-92ainhibitor后,结肠癌细胞系SW480的miR-92a表达水平明显降低(P0.01),增殖能力和侵袭能力均受到明显抑制(P0.01);(2)转染miR-29cmimic后,结肠癌细胞系SW480中miR-29c表达水平明显升高(P0.01),但SW480细胞系的增殖能力和侵袭能力均受到明显抑制(P0.01)。结论 miR-92a在结肠癌细胞中过表达能提高肿瘤细胞的增殖能力和侵袭能力,miR-29c则发挥着相反的作用。     

野生型p53基因对肠癌细胞增殖及凋亡的影响

目的研究人野生型p53基因对肠癌细胞系SW480增殖及凋亡的影响。方法阳离子脂质体介导转染人野生型p53基因至SW480细胞系,利用MTT检测该基因对SW480细胞增殖的影响;利用Western blot检测CyclinD1及Bcl-2、Bax蛋白表达水平。结果与对照组比较,转染人野生型p53基因后的SW480细胞的增殖能力受到抑制;Cy-clinD1蛋白水平下调,Bcl-2、Bax表达呈负相关趋势。结论野生型p53基因在一定程度上可诱导肿瘤细胞凋亡。     

敲减SATB1抑制结肠癌细胞增殖、迁移、侵袭

目的 研究慢病毒介导敲减核基质结合区结合蛋白质(SATB)1对结肠癌细胞增殖、凋亡、迁移和侵袭的影响.方法 结肠癌SW480细胞感染阴性对照慢病毒和SATB1 shRNA重组慢病毒设置为SW480-sh-NC、SW480-sh-SATB1组,将不进行慢病毒感染的SW480细胞设置为SW480-WT组.Realtime ...     

Inhibition of CXCR4 activity with AMD3100 decreases invasion of human colorectal cancer cells in vitro

AIM： To investigate the effect and mechanism of blockade of the CXC chemokine receptor-4 （CXCR4） signaling pathway by AMD3100, a small non-peptide CXCR4 inhibitor, on invasion and metastasis of colorectal cancer cells in vitro. METHODS： Human colorectal cancer cell line SW480 was treated with AMD3100 at different final concentrations. 3-（4,5-dimethylthiazole-2-yl）-2.5-dipheny-ltetrazolium bromide （MTT） assay was used to detect the effect of AMD3100 on cell proliferation. The invasion ability of SW480 cells was determined by cell invasion assay kit. In the presence of AMD3100, the CXCL12-mediated migratory response of SW480 cells was tested by classical chemotaxis assays. RT-PCR analysis and Western blotting were used to detect the expression of vascular endothelial growth factor （VEGF）, matrix metalloproteinase-2 （MMP-2） and -9 （MMP-9） in SW480 cells. RESULTS： Cell viability was significantly suppressed by AMD3100 in a dose-dependent manner. AMD3100 （100 and 1000 ng/mL） significantly inhibited the invasion ability of SW480 cells. Treatment with AMD3100 markedly reduced the expression of VEGF and MMP-9 but not MMP-2 in SW480 cells. CONCLUSION： The CXCL12/CXCR4 system is an important mediator of proliferation and invasion of CXCR4-expressing colorectal cancer cells. AMD3100 inhibited invasion and metastasis activity of the colorectalcancer cell line SW480 through down-regulation of VEGF and MMP-9 expression.     

Mechanism of counterattack of colorectal cancer cell by Fas/Fas ligand system

AIM: To determine the role of Fas/Fas ligand (FasL) in the immune escape of colon cancer cells. METHODS: Immunohistochemistry was used to observe the expression of Fas and FasL in the tissues of colon cancer patients. In situ hybridization was used to detect the localization of FasL mRNA expression in cancer tissues. Terminal deoxynucleotide transferase-mediated dUTP nick end labeling (TUNEL) assay and CD45 staining were performed to detect the apoptosis of tumor-infiltrating lymphocytes (TILs). Co-culture assays of colon cancer cells (SW480) and Jurkat cells (Fas-sensitive cells) were performed to observe the counterattack of colon cancer cells to lymphocytes. RESULTS: Of 53 cases of colon carcinomas, 23 cases (43.4%) expressed Fas which was significantly lower as compared to the normal colonic mucosa (73.3%, P<0.01), and 45 cases (84.9%) of colon carcinomas expressed FasL, whereas only two cases (3.75%) in normal mucosa expressed FasL. FasL expression in the colon cancer cells was found to be associated with increased cell death of TILs. The apoptotic rate of TIL in the FasL-positive staining regions of tumor cells was significantly higher than that in the FasL-negative staining region (54.84±2.79% vs 25.73±1.98%, P<0.01). The co-culture of SW480 cells and Jurkat cells confirmed the function of FasL on the SW480 cells. The apoptotic rates of Jurkat cells were found to be related with the amount of SW480 cells. CONCLUSION: Colon cancer cells can escape the immune surveillance and killing via decreasing Fas expression, and can counterattack the immune system via increasing FasL expression. Fas/FasL can serve as potential targets for effective antitumor therapy.     

Special AT-rich sequence-binding protein 1 promotes cell growth and metastasis in colorectal cancer

AIM: To evaluate the expression of special AT-rich sequence-binding protein 1 (SATB1 ) gene in colorectal cancer and its role in colorectal cancer cell proliferation and invasion.METHODS: Immunohistochemistry was used to detect the protein expression of SATB1 in 30 colorectal cancer (CRC) tissue samples and pair-matched adjacent nontumor samples. Cell growth was investigated after enhancing expression of SATB1. Wound-healing assay and Transwell assay were used to investigate the impact of SATB1 on migratory and invasive abilities of SW480 cells in vitro . Nude mice that received subcutaneous implantation or lateral tail vein were used to study the effects of SATB1 on tumor growth or metastasis in vivo . RESULTS: SATB1 was over-expressed in CRC tissues and CRC cell lines. SATB1 promotes cell proliferation and cell cycle progression in CRC SW480 cells. SATB1 over-expression could promote cell growth in vivo . In addition, SATB1 could significantly raise the ability of cell migration and invasion in vitro and promote the ability of tumor metastasis in vivo . SATB1 could up-regulate matrix metalloproteases 2, 9, cyclin D1 and vimentin, meanwhile SATB1 could down-regulate E-cadherin in CRC. CONCLUSION: SATB1 acts as a potential growth and metastasis promoter in CRC. SATB1 may be useful as a therapeutic target for CRC.     

Syndecan-1对结肠癌细胞增殖、迁移和侵袭的影响

背景:Syndecan-1是表达于上皮细胞表面的一种硫酸乙酰肝素蛋白聚糖,在多种肿瘤细胞中的表达下降,与肿瘤的多种生物学行为密切相关,但其在细胞水平上对结直肠癌发展的作用目前尚无深入的研究。目的:评估Syndecan-1对人结肠癌细胞株增殖、迁移和侵袭能力的影响,并初步探讨可能的分子机制。方法:分别将Syndecan-1高表达质粒和Syndecan-1 siRNA转染人结肠癌细胞株SW620和SW480,分别以转染空载质粒或无关序列作为阴性对照。采用CCK-8法检测细胞增殖,Transwell细胞迁移实验和Matrigel细胞侵袭实验分别检测细胞迁移、侵袭能力,蛋白质印迹法检测上皮细胞间质转化主要分子标记物E-cadherin的蛋白表达。结果:与阴性对照组相比,转染Syndecan-1高表达质粒的SW620细胞生长速度、迁移细胞数和侵袭细胞数均明显降低,同时E-cadherin表达升高;转染Syndecan-1 siRNA的SW480细胞生长速度、迁移细胞数和侵袭细胞数均明显升高,同时E-cadherin表达降低。结论:Syndecan-1能抑制结肠癌细胞的增殖、迁移和侵袭能力,其机制可能与上皮细胞间质转化有关。     

Antitumor bioactivity of adenovirus-mediated p27mt in colorectal cancer cell line SW480

AIM： To explore the antitumor bioactivity of adenovirusmediated mutant type p27^kip1 gene in a colorectal cancer cell line SW480. METHODS： We constructed recombinant adenovirus vector expressing a mutant type p27^kip1 gene （ad- p27mt）, with mutation of Thr-187/Pro-188 （ACGCCC） to Met-187/Ile-188 （ATGATC）, and transduced into SW480 cells. Then we detected expression of p27, Bcl-2 and Bax protein in the transductants by Western blotting, cell cycle of transductants by a digital flow cytometric system, migrating potential with Boyden Chamber and SW480 tumor cell growth inhibition in vitro and in vivo. RESULTS： We found that a recombinant adenovirus vector of expressing ad-p27mt, with mutation of Thr-187/Pro-188 （ACGCCC） to Met-187/Ile-188 （ATGATC） has potent inhibition of SW480 tumor cell growth in vitro and in vivo. Furthermore, ad-p27mt induced cell apoptosis via regulating bax and bcl-2 expressions, and G1/S arrest in SW480 cells and inhibited cell migration. CONCLUSION： ad-p27mt has a strong anti-tumor bioactivity and has the potential to develop into new therapeutic agents for colorectal cancer.     

藤梨根提取物通过调控miR-192-5p/ARPP19轴影响结直肠癌细胞的增殖和凋亡

背景 藤梨根提取物(rattan root extract,RRE)可抑制胃癌、肺癌等肿瘤细胞增殖,具有一定抗肿瘤作用,但其是否影响结直肠癌细胞的恶性表型还未知.miR-192-5p在结直肠癌组织中表达降低,且其低表达与肿瘤大小等临床病理特征相关,可作为结直肠癌诊治的潜在生物学标志物.StarBase生物信息学软件预测...     

溶血磷脂酸协同TGFβ1促进结肠癌细胞上皮间质转化的研究

目的探讨溶血磷脂酸(LPA)协同转化生长因子1(TGFβ1)的促上皮间质转化(EMT)作用。 方法观察TGFβ1及LPA诱导SW480细胞后细胞形态变化,Western blotting方法检测E-Cadherin、Vimentin的表达变化以及细胞增殖活性变化。 结果TGFβ1及LPA均能使SW480细胞形态发生EMT改变;两者联合诱导细胞EMT改变更为明显。空白对照组、LPA组、TGFβ1组、LPA组＋TGFβ1组,E-Cadherin表达依次下降;Vimentin表达未见明显差异。 结论LPA可能协同TGF-β1产生EMT作用,下调LPA的表达或可抑制肿瘤侵袭转移。     

Mechanisms underlying aspirin-mediated growth inhibition and apoptosis induction of cyclooxygenase-2 negative colon cancer cell line SW480

AIM： To investigate the effects of aspirin （acetylsalicylic acid） on proliferation and apoptosis of colorectal cancer cell line SW480 and its mechanism. METHODS： Cyclooxygenase （COX）-2 negative colorectal cancer cell line SW480 was treated with aspirin at concentrations of 2.5 mmol/L, 5.0 mmol/L, 10.0 mmol/L for different periods in v/tro. Anti-proliferation effect of aspirin on SW480 was detected by 3-（4,5-dimeth- ylthiazol-2-yl）-2,5-diphenyltetrazolium bromide （MTT） assay. Cell cycle and apoptosis were observed by flow cytometry （FCM）. Transmission electron microscope （TEM） was used for morphological study. Apoptosis-associated genes were detected by immunohistochemical staining and Western blotting. RESULTS： Aspirin inhibited SW480 proliferation and induced apoptosis in a dose- and time-dependent manner. Treatment with different concentrations of aspirin significantly increased the proportions of cells at the G0/G1 phase and decreased the proportions of cells at the S- and G2/M phases in a concentration-dependent manner. Aspirin not only induced apoptosis but also caused cell necrosis at a high concentration as well. After treatment with aspirin, SW480 cells displayed typically morphological features of apoptosis and necrosis under TEM, and increased the Bcl-2 expression in cells, but the expression of Bax was down regulated. CONCLUSION： Aspirin inhibits proliferation and induces apoptosis of SW480 cells. Its anti-tumor mechanism may arrest cell cycle and shift Bax/Bcl-2 balance in cells.