DEK基因沉默对宫颈癌CaSki细胞增殖和凋亡影响的研究

目的:通过观察DEK基因沉默后对CaSki细胞增殖、细胞周期及凋亡影响,探讨DEK原癌基因siRNA对宫颈癌细胞的影响和机制.方法:将DEK siRNA真核表达载体转入CaSki细胞,转染48 h后,采用MTT法检测细胞增殖、流式细胞仪检测细胞凋亡和细胞周期改变情况.结果:与对照组和未转染组相比,转染psiRNA-hH...     

凋亡抑制新基因Livin与宫颈癌的关系

凋亡蛋白抑制因子(IAP)是一类抑制凋亡的调节分子,其成员Livin是新发现的凋亡抑制基因,能够与内源IAP抑制剂SMAC和caspase-3、caspase-7、caspase-9结合,发挥抗细胞凋亡的作用.近年来,研究发现Livin在宫颈癌细胞及组织中高表达,而且通过检测癌细胞凋亡有助于确定宫颈癌的发生机制及其疗效评价,因此研究Livin结构与功能及作用机制对于宫颈癌的发生、发展、诊断与治疗有积极意义.     

小干扰RNA沉默survivin基因对结肠癌Caco-2细胞增殖和凋亡的影响

目的:观察survivin在人结肠癌细胞Caco-2中的表达及小干扰RNA(small interfering RNA,siRNA)沉默survivin基因对survivin蛋白表达、细胞增殖及凋亡的影响.方法:以脂质体LipofectAMINE 2000为载体,将针对特异靶点survivin基因的siRNA转染入人结肠癌细胞Caco-2.应用免疫细胞化学SP法和Western印迹法检测survivin蛋白表达的变化,MTT法检测细胞增殖,FCM法检测细胞凋亡.结果:免疫细胞化学检测结果显示,survivin蛋白在Caco-2细胞空白对照组、脂质体对照组及阴性错配对照组的细胞质中均呈强阳性表达,而在survivin-siRNA转染组细胞质中呈弱阳性表达;Western印迹法检测结果显示,survivin-siRNA转染组细胞的蛋白条带亮度明显低于空白对照组、脂质体对照组和阴性错配对照组;MTT检测结果显示,与阴性错配对照组相比, survivin-siRNA转染组细胞生长出现明显的抑制(P＜0.05),且不同时段(24、48和72 h)的肿瘤细胞增殖抑制率之间差异有统计学意义(P＜0.05);FCM检测结果显示,survivin-siRNA转染组细胞出现明显的亚二倍体峰,细胞凋亡率(9.72%)明显高于空白对照组(P＜0.01).结论:siRNA沉默survivin基因可抑制结肠癌细胞增殖,促进细胞凋亡.推测survivin基因可能成为结肠癌基因治疗的一个新靶点.     

凋亡抑制新基因Livin与宫颈癌的关系

凋亡蛋白抑制因子(IAP)是一类抑制凋亡的调节分子,其成员Livin是新发现的凋亡抑制基因,能够与内源IAP抑制剂SMAC和caspase-3、caspase-7、caspase-9结合,发挥抗细胞凋亡的作用.近年来,研究发现Livin在宫颈癌细胞及组织中高表达,而且通过检测癌细胞凋亡有助于确定宫颈癌的发生机制及其疗效评价,因此研究Livin结构与功能及作用机制对于宫颈癌的发生、发展、诊断与治疗有积极意义.     

siRNA介导的survivin基因沉默诱导人膀胱癌BIU-87细胞凋亡的研究

[目的]研究siRNA（small interference RNA）对人膀胱癌BIU-87细胞凋亡和survivin基因表达的影响。[方法]利用Ambion公司设计软件，设计并体外转录合成针对survivin基因的3个特异性siRNA，通过脂质体将siRNA转入BIU-87细胞，分别采用MTT和DNA原位末端标记（TUNEL）法检测siRNA对BIU-87细胞生长抑制率（IR）和凋亡指数（AI），半定量逆转录聚合酶链反应（RT-PCR）和Western Blotting检测siRNA对survivin mRNA及其蛋白表达的影响。[结果]转染后的siRNA1～3组的BIU-87细胞的IR（41.84%、56.21%、54.13%）和AI（21.98%、36.54%、31.34%）均分别显著高于正常对照组（1.98%和3.17%）（P＜0.05），survivin mRNA及其蛋白表达水平均显著低于对照组；其中siRNA2～3对BIU-87细胞的IR、AI和survivin表达的抑制作用均显著高于siRNA1。[结论]体外转录合成的siRNA可抑制BIU-87细胞survivin的表达，诱导BIU-87细胞凋亡，从而抑制BIU-87细胞生长，为siRNA介导的膀胱肿瘤基因沉默提供实验依据。     

肺腺癌组织中RGC32基因的表达及其对肺腺癌A549细胞增殖和凋亡的影响

 目的　检测RGC32基因在肺腺癌组织中的表达及其对肺腺癌A549细胞增殖和凋亡的影响。方法　实时荧光定量PCR检测36例肺腺癌及相应癌旁组织中RGC32基因的表达;应用RNA干扰技术抑制A549细胞中RGC32基因的表达,实时荧光定量PCR检测抑制效果,MTT法测定A549细胞生长抑制率,流式细胞术检测A549细胞凋亡。结果　RGC32基因在肺腺癌组织中表达明显上调,为癌旁组织表达量的2.2736倍（t＝-29.185,P＝0.001）。RNA干扰能够显著抑制A549细胞中RGC32基因的表达。RGC32基因表达降低后,A549细胞的凋亡率由（2.47±0.17）%提高至（4.65±0.26）%（t＝-202.868,P＝0.000）,生长抑制率由2.9 %提高至45.4 %（t＝-37.915,P＝0.001）,细胞增殖活力明显降低。结论　RGC32基因在肺腺癌组织中的表达明显上调,RGC32基因表达降低能够促进A549细胞的凋亡并抑制细胞增殖。     

重组pcDNA3-UpIb promoter-Smac质粒转染靶向膀胱癌的促凋亡效应

目的：探讨转染新构建的UpIb启动子调控携Smac基因的真核表达质粒pcDNA3-UpIb promoter—Smac对膀胱癌细胞的促凋亡效用一方法：用脂质体将质粒转染到膀胱癌BIU-87细胞系中，2dh后用RT—PCR检测Smac的表达，以低剂量丝裂霉素C诱导凋亡，利用倒置光学显微镜、Wrighs—Gimesa染色、DNA凝胶电泳技术、TUNEL-荧光标记技术、流式细胞术检测凋亡。结果：丝裂霉素C处理的各组在倒置光学显微镜和Wrighs—Gimesa染色油镜下可见到典型的凋亡形态学改变，DNA凝胶电泳呈特征性的梯形条带jTUNEL-荧光标记技术及流式细胞术检测转染pcDNA3-UpIb promoter—Smac组凋亡率显著高于单用丝裂霉素C组。结论：转染pcDNA3-Up Ibpromoter—Smac能够通过提高Smac的表达而有效促进丝裂霉素C诱导的膀胱癌细胞凋亡，提示该质粒配合凋亡诱导药物可能成为膀胱癌新的靶向基因治疗手段．     

沉默SEPT9基因对肝癌HepG2细胞增殖及凋亡的影响

目的 探讨SEPT9基因与肝癌的关系,进一步揭示SEPT9基因在肝癌发生发展过程中的作用.方法 将构建的针对SEPT9基因的shRNA表达载体通过脂质体LipofectamineTM 2000转染人肝癌HepG2细胞,荧光显微镜下观察细胞的转染效率,RT-PCR、Western blot法分别检测SEPT9 mRNA和蛋白质的表达抑制情况,CCK-8检测细胞的生长抑制情况,流式细胞术检测细胞凋亡情况.结果 荧光显微镜观察细胞的转染效率达到70％以上;SEPT9 mRNA及蛋白表达抑制率均受到明显抑制.CCK-8法检测结果显示,实验组HepG2细胞的生长受到明显抑制;流式细胞术检测结果显示实验组HepG2细胞凋亡明显(P＜0.05).结论 抑制SEPT9基因的表达可有效抑制HepG2细胞的增殖,并对细胞凋亡有明显的促进作用.     

RNA干扰REG1A基因抑制人膀胱癌T24细胞增殖

目的:观察小干扰RNA(small interfering RNA,siRNA)沉默REG1A (regenerating islet-derived 1 alpha)基因的表达对人膀胱癌T24细胞增殖及细胞周期的影响.方法:化学合成针对REG1A基因的3对siRNA,使用脂质体将其转染至T24细胞中.应用实时荧光定量PCR和蛋白质印迹法分别检测REG1A siRNA转染后T24细胞REG1A mRNA和蛋白的表达,CCK-8(cell counting kit-8)法检测转染后的细胞增殖情况,FCM检测转染后细胞周期的变化.结果:与空白对照组比较,REG1A siRNA转染T24细胞后,REG1A mRNA和蛋白表达分别下降了(83.10±0.06)％和(55.81±0.16)％,差异有统计学意义(P＜0.05);细胞增殖受到抑制(P＜0.05),G0/G1期细胞百分比明显增加(P＜0.05),细胞增殖指数下降(P＜0.05).结论:REG1A siRNA能有效抑制膀胱癌T24细胞REG1A mRNA和蛋白的表达,并抑制细胞增殖.     

KDR基因沉默对胃癌MGC-803细胞增殖和凋亡的影响

[目的]探讨KDR基因对胃癌MGC-803细胞增殖和凋亡的影响。[方法]设计及合成KDR的siRNA序列,LipofectamineTM 2000转染MGC-803细胞。通过RT-PCR、Western Blot检测KDR在干扰后的mRNA和蛋白的表达情况,利用流式细胞仪检测细胞周期,WST-1法检测细胞增殖活性,TUNEL法检测细胞凋亡情况。[结果]KDR mRNA和蛋白表达,观察组较对照组和空白组显著降低,差异有统计学意义(P<0.05)。观察组细胞的生长速度明显减慢,细胞周期被阻滞在G0/G1期,S期细胞数减少,差异有统计学意义(P<0.05)。转染KDRsiRNA的MGC-803细胞的增殖能力明显受到抑制(P<0.05),且明显促进了细胞的凋亡,差异有统计学意义(P<0.05)。[结论]特异性干扰KDR基因表达可抑制胃癌MGC-803细胞的增殖,并促进肿瘤细胞凋亡。KDR的siRNA序列可能成为治疗胃癌的有效靶点。     

转染血管内皮生长因子C反义寡核苷酸对子宫颈癌Hela细胞增生和凋亡的影响

Objective To explore effect on the proliferation and apoptosis of Hela cells in vitro by using vascular endothelial growth factor-C(VEGF-C) antisense oligonucleotides (ASODN). Methods VEGF-C ASODN was transfected into Hela cells by liposome-mediated; the cells transfected with the oligodeoxynuclecotide (SODN) and saline were used as control groups. The efficiency of transfection was detected by fluorescence microscope. The inhibitive rate of cell growth was detected with MTT methods. Apoptosis and cell cycle were evaluated using FCM. The mRNA expression of VEGF-C was determined by RT-PCR, and the expression of VEGF-C was determined by western blotting. Results VEGF-C ASODN had been transfected into Hela cells by liposome-mediated. The index of apoptosis after transfection 48 h was (19.39±1.81)%. G2/M phase cell increased after transfection, meanwhile the expression of VEGF-C degraded on the level of mRNA and protein (P<0.05). Conclusion Transfeeted with VEGF-C ASODN could down-regulate the expression of VEGF on the level of mRNA and protein, block the cell circle, inhibit cell proliferation and induce apoptosis in Hela cells in vitro.     

转染血管内皮生长因子C反义寡核苷酸对子宫颈癌Hela细胞增生和凋亡的影响

Objective To explore effect on the proliferation and apoptosis of Hela cells in vitro by using vascular endothelial growth factor-C(VEGF-C) antisense oligonucleotides (ASODN). Methods VEGF-C ASODN was transfected into Hela cells by liposome-mediated; the cells transfected with the oligodeoxynuclecotide (SODN) and saline were used as control groups. The efficiency of transfection was detected by fluorescence microscope. The inhibitive rate of cell growth was detected with MTT methods. Apoptosis and cell cycle were evaluated using FCM. The mRNA expression of VEGF-C was determined by RT-PCR, and the expression of VEGF-C was determined by western blotting. Results VEGF-C ASODN had been transfected into Hela cells by liposome-mediated. The index of apoptosis after transfection 48 h was (19.39±1.81)%. G2/M phase cell increased after transfection, meanwhile the expression of VEGF-C degraded on the level of mRNA and protein (P<0.05). Conclusion Transfeeted with VEGF-C ASODN could down-regulate the expression of VEGF on the level of mRNA and protein, block the cell circle, inhibit cell proliferation and induce apoptosis in Hela cells in vitro.     

转染血管内皮生长因子C反义寡核苷酸对子宫颈癌Hela细胞增生和凋亡的影响

目的体外研究脂质体介导转染血管内皮生长因子C（VEGF—C）反义寡核苷酸（VEGF—C,ASODN）对子宫颈癌Hela细胞增生和凋亡的影响。方法实验组将VEGF—CASODN（优化筛选）用脂质体导人Hela细胞,设正义（SODN）和空白对照组进行比较。用荧光显微镜观察转染率,MTT比色法检测细胞增生,流式细胞术检测细胞凋亡及细胞周期变化,RT—PCR、Western blotting法检测VEGF—C mRNA和蛋白的表达。结果经脂质体介导可成功将VEGF—C反义核酸转染至子宫颈癌Hela细胞;细胞的增生受到明显抑制,G2／M期细胞比例增多,转染作用48h时凋亡率最高,达（19．39±1．81）％;VEGF—C在mRNA和蛋白水平的表达均明显减少,与对照组比较差异有统计学意义（P〈0．05）。结论体外转染VEGF—CASODN可在mRNA和蛋白水平下调Hela细胞VEGF—C的表达,阻滞并同步化细胞周期,抑制细胞增生,诱导细胞凋亡。     

转染血管内皮生长因子C反义寡核苷酸对子宫颈癌Hela细胞增生和凋亡的影响

Objective To explore effect on the proliferation and apoptosis of Hela cells in vitro by using vascular endothelial growth factor-C(VEGF-C) antisense oligonucleotides (ASODN). Methods VEGF-C ASODN was transfected into Hela cells by liposome-mediated; the cells transfected with the oligodeoxynuclecotide (SODN) and saline were used as control groups. The efficiency of transfection was detected by fluorescence microscope. The inhibitive rate of cell growth was detected with MTT methods. Apoptosis and cell cycle were evaluated using FCM. The mRNA expression of VEGF-C was determined by RT-PCR, and the expression of VEGF-C was determined by western blotting. Results VEGF-C ASODN had been transfected into Hela cells by liposome-mediated. The index of apoptosis after transfection 48 h was (19.39±1.81)%. G2/M phase cell increased after transfection, meanwhile the expression of VEGF-C degraded on the level of mRNA and protein (P<0.05). Conclusion Transfeeted with VEGF-C ASODN could down-regulate the expression of VEGF on the level of mRNA and protein, block the cell circle, inhibit cell proliferation and induce apoptosis in Hela cells in vitro.     

转染血管内皮生长因子C反义寡核苷酸对子宫颈癌Hela细胞增生和凋亡的影响

Objective To explore effect on the proliferation and apoptosis of Hela cells in vitro by using vascular endothelial growth factor-C(VEGF-C) antisense oligonucleotides (ASODN). Methods VEGF-C ASODN was transfected into Hela cells by liposome-mediated; the cells transfected with the oligodeoxynuclecotide (SODN) and saline were used as control groups. The efficiency of transfection was detected by fluorescence microscope. The inhibitive rate of cell growth was detected with MTT methods. Apoptosis and cell cycle were evaluated using FCM. The mRNA expression of VEGF-C was determined by RT-PCR, and the expression of VEGF-C was determined by western blotting. Results VEGF-C ASODN had been transfected into Hela cells by liposome-mediated. The index of apoptosis after transfection 48 h was (19.39±1.81)%. G2/M phase cell increased after transfection, meanwhile the expression of VEGF-C degraded on the level of mRNA and protein (P<0.05). Conclusion Transfeeted with VEGF-C ASODN could down-regulate the expression of VEGF on the level of mRNA and protein, block the cell circle, inhibit cell proliferation and induce apoptosis in Hela cells in vitro.     

转染血管内皮生长因子C反义寡核苷酸对子宫颈癌Hela细胞增生和凋亡的影响

Objective To explore effect on the proliferation and apoptosis of Hela cells in vitro by using vascular endothelial growth factor-C(VEGF-C) antisense oligonucleotides (ASODN). Methods VEGF-C ASODN was transfected into Hela cells by liposome-mediated; the cells transfected with the oligodeoxynuclecotide (SODN) and saline were used as control groups. The efficiency of transfection was detected by fluorescence microscope. The inhibitive rate of cell growth was detected with MTT methods. Apoptosis and cell cycle were evaluated using FCM. The mRNA expression of VEGF-C was determined by RT-PCR, and the expression of VEGF-C was determined by western blotting. Results VEGF-C ASODN had been transfected into Hela cells by liposome-mediated. The index of apoptosis after transfection 48 h was (19.39±1.81)%. G2/M phase cell increased after transfection, meanwhile the expression of VEGF-C degraded on the level of mRNA and protein (P<0.05). Conclusion Transfeeted with VEGF-C ASODN could down-regulate the expression of VEGF on the level of mRNA and protein, block the cell circle, inhibit cell proliferation and induce apoptosis in Hela cells in vitro.     

转染血管内皮生长因子C反义寡核苷酸对子宫颈癌Hela细胞增生和凋亡的影响

Objective To explore effect on the proliferation and apoptosis of Hela cells in vitro by using vascular endothelial growth factor-C(VEGF-C) antisense oligonucleotides (ASODN). Methods VEGF-C ASODN was transfected into Hela cells by liposome-mediated; the cells transfected with the oligodeoxynuclecotide (SODN) and saline were used as control groups. The efficiency of transfection was detected by fluorescence microscope. The inhibitive rate of cell growth was detected with MTT methods. Apoptosis and cell cycle were evaluated using FCM. The mRNA expression of VEGF-C was determined by RT-PCR, and the expression of VEGF-C was determined by western blotting. Results VEGF-C ASODN had been transfected into Hela cells by liposome-mediated. The index of apoptosis after transfection 48 h was (19.39±1.81)%. G2/M phase cell increased after transfection, meanwhile the expression of VEGF-C degraded on the level of mRNA and protein (P<0.05). Conclusion Transfeeted with VEGF-C ASODN could down-regulate the expression of VEGF on the level of mRNA and protein, block the cell circle, inhibit cell proliferation and induce apoptosis in Hela cells in vitro.     

转染血管内皮生长因子C反义寡核苷酸对子宫颈癌Hela细胞增生和凋亡的影响

Objective To explore effect on the proliferation and apoptosis of Hela cells in vitro by using vascular endothelial growth factor-C(VEGF-C) antisense oligonucleotides (ASODN). Methods VEGF-C ASODN was transfected into Hela cells by liposome-mediated; the cells transfected with the oligodeoxynuclecotide (SODN) and saline were used as control groups. The efficiency of transfection was detected by fluorescence microscope. The inhibitive rate of cell growth was detected with MTT methods. Apoptosis and cell cycle were evaluated using FCM. The mRNA expression of VEGF-C was determined by RT-PCR, and the expression of VEGF-C was determined by western blotting. Results VEGF-C ASODN had been transfected into Hela cells by liposome-mediated. The index of apoptosis after transfection 48 h was (19.39±1.81)%. G2/M phase cell increased after transfection, meanwhile the expression of VEGF-C degraded on the level of mRNA and protein (P<0.05). Conclusion Transfeeted with VEGF-C ASODN could down-regulate the expression of VEGF on the level of mRNA and protein, block the cell circle, inhibit cell proliferation and induce apoptosis in Hela cells in vitro.     

转染血管内皮生长因子C反义寡核苷酸对子宫颈癌Hela细胞增生和凋亡的影响

Objective To explore effect on the proliferation and apoptosis of Hela cells in vitro by using vascular endothelial growth factor-C(VEGF-C) antisense oligonucleotides (ASODN). Methods VEGF-C ASODN was transfected into Hela cells by liposome-mediated; the cells transfected with the oligodeoxynuclecotide (SODN) and saline were used as control groups. The efficiency of transfection was detected by fluorescence microscope. The inhibitive rate of cell growth was detected with MTT methods. Apoptosis and cell cycle were evaluated using FCM. The mRNA expression of VEGF-C was determined by RT-PCR, and the expression of VEGF-C was determined by western blotting. Results VEGF-C ASODN had been transfected into Hela cells by liposome-mediated. The index of apoptosis after transfection 48 h was (19.39±1.81)%. G2/M phase cell increased after transfection, meanwhile the expression of VEGF-C degraded on the level of mRNA and protein (P<0.05). Conclusion Transfeeted with VEGF-C ASODN could down-regulate the expression of VEGF on the level of mRNA and protein, block the cell circle, inhibit cell proliferation and induce apoptosis in Hela cells in vitro.     

转染血管内皮生长因子C反义寡核苷酸对子宫颈癌Hela细胞增生和凋亡的影响

Objective To explore effect on the proliferation and apoptosis of Hela cells in vitro by using vascular endothelial growth factor-C(VEGF-C) antisense oligonucleotides (ASODN). Methods VEGF-C ASODN was transfected into Hela cells by liposome-mediated; the cells transfected with the oligodeoxynuclecotide (SODN) and saline were used as control groups. The efficiency of transfection was detected by fluorescence microscope. The inhibitive rate of cell growth was detected with MTT methods. Apoptosis and cell cycle were evaluated using FCM. The mRNA expression of VEGF-C was determined by RT-PCR, and the expression of VEGF-C was determined by western blotting. Results VEGF-C ASODN had been transfected into Hela cells by liposome-mediated. The index of apoptosis after transfection 48 h was (19.39±1.81)%. G2/M phase cell increased after transfection, meanwhile the expression of VEGF-C degraded on the level of mRNA and protein (P<0.05). Conclusion Transfeeted with VEGF-C ASODN could down-regulate the expression of VEGF on the level of mRNA and protein, block the cell circle, inhibit cell proliferation and induce apoptosis in Hela cells in vitro.