The expression of octamer binding factor 4 （Oct4） gene in bladder cancer cell line T24 and its effects on the biological characteristics of the cells were investigated. RT-PCR and Western blot were employed to detec...The expression of octamer binding factor 4 （Oct4） gene in bladder cancer cell line T24 and its effects on the biological characteristics of the cells were investigated. RT-PCR and Western blot were employed to detect the expression of Oct4 in T24 cells. The changes of biological characteristics in T24 cells were analyzed before and after gene-silencing by Boyden chamber and MTT. The results showed that the expression of Oct4 gene was detectable in T24 cells by RT-PCR and Western blot. The expression of Oct4 gene and protein was down-regulated by siRNA, and average number of transwell cells in interference group, negative control group and blank control group was 101.40±54.56, 104.20± 10.03 and 111.00±11.90, respectively. There was significant difference in the proliferation abilit,） of the cells from 48 h, 72 h to 96 h after the interference by siRNA between interference group and negative group or blank control group （P〈0.05）. It was suggested that Oct4 gene was related with proliferation ability of T24 cells, but not with invasive capability.展开更多
Objectives：To investigate the effects of adenovirus-mediated inducible nitric oxide synthase gene transfection on bladder transitional cell carcinoma T24 cells,and to provide novel insights and approaches to clinical...Objectives：To investigate the effects of adenovirus-mediated inducible nitric oxide synthase gene transfection on bladder transitional cell carcinoma T24 cells,and to provide novel insights and approaches to clinical therapies against bladder transitional cell carcinoma.Methods：Firstly,construct recombinant adenovirus vector pAd-iNOS of iNOS,followed by transfection of pAd-iNOS into HECK293 packaging cells.Thirdly,harvest recombinant adenovirus rAd-iNOS after amplification and purification procedures.Finally,transfect the recombinant adenovirus rAd-iNOS into human bladder carcinoma T24 cells and examine the effect of rAd-iNOS transfection on apoptosis of T24 and possible mechanism.Results：As shown by this study,the recombinant adenovirus rAd-iNOS was constructed successfully.The virus titer was 5.8×108 PFU/mL and recombinant was verified by PCR analysis.Transfection of adenovirus rAd-iNOS into T24 cells could induce secretion of high NO concentration,P53 protein expression upregulation,as well as promotion ofT24 cell apoptosis.Conclusions：The transfection of human bladder carcinoma T24 cells from recombinant adenovirus rAdiNOS was confirmed to induce intracellular iNOS over-expression,high production of NO,up-regulation of intracellular P53 expression and promotion of cell apoptosis.展开更多
目的:探讨氧化苦参碱能否通过第10号染色体缺失的磷酸酶及张力蛋白同源的基因/3-磷酸肌醇激酶/蛋白激酶B/哺乳动物雷帕霉素靶向基因(Phosphatase and tension homolog deleted on chromosome ten/phosphoinositide 3-kinase/protein kin...目的:探讨氧化苦参碱能否通过第10号染色体缺失的磷酸酶及张力蛋白同源的基因/3-磷酸肌醇激酶/蛋白激酶B/哺乳动物雷帕霉素靶向基因(Phosphatase and tension homolog deleted on chromosome ten/phosphoinositide 3-kinase/protein kinase B/mammalian target of rapamycin,PTEN/PI3K/Akt/mTOR)通路对人膀胱癌T24细胞的增殖、凋亡产生影响。方法:T24细胞复苏传代后与不同剂量氧化苦参碱共培养,MTT法检测不同剂量氧化苦参碱对T24细胞增殖的影响,流式细胞仪检测氧化苦参碱对T24细胞凋亡的影响,Hoechst光镜观察氧化苦参碱对T24细胞核形态的影响,Western blot检测氧化苦参碱对T24细胞PI3K、Akt、mTOR、PTEN蛋白表达的影响。结果:与0μmol/L组比较,氧化苦参碱各剂量组T24细胞的增殖率下降,凋亡率升高,PI3K、Akt、mTOR蛋白表达下降,PTEN蛋白表达上升,且在一定范围内存在剂量依赖性(P<0.05)。结论:氧化苦参碱可能通过PTEN/PI3K/Akt/mTOR通路抑制T24细胞的增殖,促进其凋亡。展开更多
目的:探讨AMP依赖的蛋白激酶α(AMP-activated protein kinaseα,AMPKα)过表达对膀胱癌T24细胞增殖、迁移、侵袭和EMT的作用及其机制。方法:建立AMPKα过表达的膀胱癌T24细胞株,依据转染质粒的不同分为T24空白组、pc-DNA空载组和pc-AMP...目的:探讨AMP依赖的蛋白激酶α(AMP-activated protein kinaseα,AMPKα)过表达对膀胱癌T24细胞增殖、迁移、侵袭和EMT的作用及其机制。方法:建立AMPKα过表达的膀胱癌T24细胞株,依据转染质粒的不同分为T24空白组、pc-DNA空载组和pc-AMPKα组。用Wb检测T24细胞AMPKα、EMT相关蛋白及EMT通路相关分子的表达水平,用Hoechst染色法检测转染后T24细胞的凋亡,CCK-8法检测T24细胞的增殖,细胞划痕愈合实验检测T24细胞的迁移,Transwell实验检测细胞的侵袭。结果:成功构建AMPKα过表达的膀胱癌T24细胞株。与T24空白组和pc-DNA空载组比较,pc-AMPKα组T24细胞上皮钙黏蛋白水平显著升高(P<0.01)、波形蛋白和神经钙黏蛋白表达水平显著降低(均P<0.01),EMT通路相关信号分子P38、STAT3活性受到显著抑制(均P<0.01),细胞发生明显凋亡、增殖能力显著减弱(均P<0.01);T24细胞迁移和侵袭能力显著降低(均P<0.01)。结论:AMPKα过表达可使EMT通路相关分子活性受到抑制,使得膀胱癌T24细胞发生明显凋亡、增殖受限并使其侵袭和迁移能力降低及伴随EMT的逆转。展开更多
文摘The expression of octamer binding factor 4 （Oct4） gene in bladder cancer cell line T24 and its effects on the biological characteristics of the cells were investigated. RT-PCR and Western blot were employed to detect the expression of Oct4 in T24 cells. The changes of biological characteristics in T24 cells were analyzed before and after gene-silencing by Boyden chamber and MTT. The results showed that the expression of Oct4 gene was detectable in T24 cells by RT-PCR and Western blot. The expression of Oct4 gene and protein was down-regulated by siRNA, and average number of transwell cells in interference group, negative control group and blank control group was 101.40±54.56, 104.20± 10.03 and 111.00±11.90, respectively. There was significant difference in the proliferation abilit,） of the cells from 48 h, 72 h to 96 h after the interference by siRNA between interference group and negative group or blank control group （P〈0.05）. It was suggested that Oct4 gene was related with proliferation ability of T24 cells, but not with invasive capability.
基金No.B2010033 Scientific Research Foundation,Health Department,Hunan Province
文摘Objectives：To investigate the effects of adenovirus-mediated inducible nitric oxide synthase gene transfection on bladder transitional cell carcinoma T24 cells,and to provide novel insights and approaches to clinical therapies against bladder transitional cell carcinoma.Methods：Firstly,construct recombinant adenovirus vector pAd-iNOS of iNOS,followed by transfection of pAd-iNOS into HECK293 packaging cells.Thirdly,harvest recombinant adenovirus rAd-iNOS after amplification and purification procedures.Finally,transfect the recombinant adenovirus rAd-iNOS into human bladder carcinoma T24 cells and examine the effect of rAd-iNOS transfection on apoptosis of T24 and possible mechanism.Results：As shown by this study,the recombinant adenovirus rAd-iNOS was constructed successfully.The virus titer was 5.8×108 PFU/mL and recombinant was verified by PCR analysis.Transfection of adenovirus rAd-iNOS into T24 cells could induce secretion of high NO concentration,P53 protein expression upregulation,as well as promotion ofT24 cell apoptosis.Conclusions：The transfection of human bladder carcinoma T24 cells from recombinant adenovirus rAdiNOS was confirmed to induce intracellular iNOS over-expression,high production of NO,up-regulation of intracellular P53 expression and promotion of cell apoptosis.
文摘目的:探讨氧化苦参碱能否通过第10号染色体缺失的磷酸酶及张力蛋白同源的基因/3-磷酸肌醇激酶/蛋白激酶B/哺乳动物雷帕霉素靶向基因(Phosphatase and tension homolog deleted on chromosome ten/phosphoinositide 3-kinase/protein kinase B/mammalian target of rapamycin,PTEN/PI3K/Akt/mTOR)通路对人膀胱癌T24细胞的增殖、凋亡产生影响。方法:T24细胞复苏传代后与不同剂量氧化苦参碱共培养,MTT法检测不同剂量氧化苦参碱对T24细胞增殖的影响,流式细胞仪检测氧化苦参碱对T24细胞凋亡的影响,Hoechst光镜观察氧化苦参碱对T24细胞核形态的影响,Western blot检测氧化苦参碱对T24细胞PI3K、Akt、mTOR、PTEN蛋白表达的影响。结果:与0μmol/L组比较,氧化苦参碱各剂量组T24细胞的增殖率下降,凋亡率升高,PI3K、Akt、mTOR蛋白表达下降,PTEN蛋白表达上升,且在一定范围内存在剂量依赖性(P<0.05)。结论:氧化苦参碱可能通过PTEN/PI3K/Akt/mTOR通路抑制T24细胞的增殖,促进其凋亡。
文摘目的:探讨AMP依赖的蛋白激酶α(AMP-activated protein kinaseα,AMPKα)过表达对膀胱癌T24细胞增殖、迁移、侵袭和EMT的作用及其机制。方法:建立AMPKα过表达的膀胱癌T24细胞株,依据转染质粒的不同分为T24空白组、pc-DNA空载组和pc-AMPKα组。用Wb检测T24细胞AMPKα、EMT相关蛋白及EMT通路相关分子的表达水平,用Hoechst染色法检测转染后T24细胞的凋亡,CCK-8法检测T24细胞的增殖,细胞划痕愈合实验检测T24细胞的迁移,Transwell实验检测细胞的侵袭。结果:成功构建AMPKα过表达的膀胱癌T24细胞株。与T24空白组和pc-DNA空载组比较,pc-AMPKα组T24细胞上皮钙黏蛋白水平显著升高(P<0.01)、波形蛋白和神经钙黏蛋白表达水平显著降低(均P<0.01),EMT通路相关信号分子P38、STAT3活性受到显著抑制(均P<0.01),细胞发生明显凋亡、增殖能力显著减弱(均P<0.01);T24细胞迁移和侵袭能力显著降低(均P<0.01)。结论:AMPKα过表达可使EMT通路相关分子活性受到抑制,使得膀胱癌T24细胞发生明显凋亡、增殖受限并使其侵袭和迁移能力降低及伴随EMT的逆转。
