Objective： To establish the method of real time fluorescence quantitative RT-PCR for detecting the expression of Survivin mRNA in nasopharyngeat carcinoma （NPC） tissues. Methods： The total RNA was extracted from N...Objective： To establish the method of real time fluorescence quantitative RT-PCR for detecting the expression of Survivin mRNA in nasopharyngeat carcinoma （NPC） tissues. Methods： The total RNA was extracted from NPC cell line CNE-2 and tissues with Trizol and then been transcribed reversely to cDNA, a method of real time fluorescence quantitative RT-PCR for detecting the expression of Survivin mRNA in NPC tissues had been established, in which chronic nasopharyn-gitis patients＇ nasopharynx tissues treated as control group. Results： The expression of Survivin mRNA all could be detected either in CNE-2 cells, NPC tissues or in chronic nasopharyngitis patients＇ nasopharynx tissues, and there was higher the expression level of Survivin mRNA in NPC tissues than which in chronic nasopharyngitis patients＇ nasopharynx tissues, the difference was significant （P 〈 0.01）. The expression of Survivin mRNA could be detected both in stage Ⅰ ＋ Ⅱ and stage Ⅲ ＋ Ⅳ NPC, and there was no significant difference in relative quantifications of gene expression between these two groups （P 〉 0.05）. There was no relationship between Survivin mRNA expression and age and sex of NPC patients （P 〉 0.05）. Conclusion： Real time fluorescence quantitative RT-PCR is a rapid, effective and high sensitive method for detecting the expression of Survivin mRNA in NPC tissues. The overexpression of Survivin mRNA may play some roles in pathogenesis of NPC.展开更多
Objective:The aim of this study was to investigate the molecular mechanism of anti-apoptotic action of survivin to the hepatoma-cellular carcinoma cell line HepG2.Methods:Design and synthesize siRNA gene sequence spec...Objective:The aim of this study was to investigate the molecular mechanism of anti-apoptotic action of survivin to the hepatoma-cellular carcinoma cell line HepG2.Methods:Design and synthesize siRNA gene sequence specifically targeting at HepG2 cell.HepG2 cells cultures were divided into five groups:blank control group,negative control group,low dose group,medium dose group and high dose group.HepG2 cells were treated respectively by pshRNA-survivin-387 of different concentrations.The apoptosis index(AI) was determined by flow cytometry(FCM).Cells were stained with rhodomine-123(Rh123) to detect changes of mitochondrial membrane potentials.The concentration of cytoplasmic cytochrome C(Cyt.C) was continuously determined by ELISA.Relative activities of caspase-9 and caspase-3 were assessed by colorimetric assay.Results:Compared with the control group,due to the function of short interference RNAs(SiRNAs) that suppresses the survivin gene expression,the apoptotic index of transfected groups were significantly higher than those of control groups(F = 13568.68,q = 110.47-327.16,P < 0.01),the apoptosis index of high concentration of transfected cells was higher than the low concentration transfected group(q = 39.63-168.22,P < 0.01).The apoptosis index of high concentrations transfected HepG2 cells was 25.54%,higher than that of blank control group,negative control group,low dose group and medium dose group(5.24%,6.61%,12.63% and 15.64%,respectively).HepG2 cells transfected with SiRNA exhibit gradually decreasing mitochondrial membrane potentials,which then lead to the releasing of Cyt C,following it were the activation of caspase-9 and caspase-3.Conclusion:Survivin performs the function of anti-apoptosis to the HepG2 cells via modulating the apoptosis of mitochondrial.HepG2 cells transfected with SiRNA survivin can significantly induce apoptosis.展开更多
基金the National Natural Science Foundation of China (No. 30460145).
文摘Objective： To establish the method of real time fluorescence quantitative RT-PCR for detecting the expression of Survivin mRNA in nasopharyngeat carcinoma （NPC） tissues. Methods： The total RNA was extracted from NPC cell line CNE-2 and tissues with Trizol and then been transcribed reversely to cDNA, a method of real time fluorescence quantitative RT-PCR for detecting the expression of Survivin mRNA in NPC tissues had been established, in which chronic nasopharyn-gitis patients＇ nasopharynx tissues treated as control group. Results： The expression of Survivin mRNA all could be detected either in CNE-2 cells, NPC tissues or in chronic nasopharyngitis patients＇ nasopharynx tissues, and there was higher the expression level of Survivin mRNA in NPC tissues than which in chronic nasopharyngitis patients＇ nasopharynx tissues, the difference was significant （P 〈 0.01）. The expression of Survivin mRNA could be detected both in stage Ⅰ ＋ Ⅱ and stage Ⅲ ＋ Ⅳ NPC, and there was no significant difference in relative quantifications of gene expression between these two groups （P 〉 0.05）. There was no relationship between Survivin mRNA expression and age and sex of NPC patients （P 〉 0.05）. Conclusion： Real time fluorescence quantitative RT-PCR is a rapid, effective and high sensitive method for detecting the expression of Survivin mRNA in NPC tissues. The overexpression of Survivin mRNA may play some roles in pathogenesis of NPC.
文摘Objective:The aim of this study was to investigate the molecular mechanism of anti-apoptotic action of survivin to the hepatoma-cellular carcinoma cell line HepG2.Methods:Design and synthesize siRNA gene sequence specifically targeting at HepG2 cell.HepG2 cells cultures were divided into five groups:blank control group,negative control group,low dose group,medium dose group and high dose group.HepG2 cells were treated respectively by pshRNA-survivin-387 of different concentrations.The apoptosis index(AI) was determined by flow cytometry(FCM).Cells were stained with rhodomine-123(Rh123) to detect changes of mitochondrial membrane potentials.The concentration of cytoplasmic cytochrome C(Cyt.C) was continuously determined by ELISA.Relative activities of caspase-9 and caspase-3 were assessed by colorimetric assay.Results:Compared with the control group,due to the function of short interference RNAs(SiRNAs) that suppresses the survivin gene expression,the apoptotic index of transfected groups were significantly higher than those of control groups(F = 13568.68,q = 110.47-327.16,P < 0.01),the apoptosis index of high concentration of transfected cells was higher than the low concentration transfected group(q = 39.63-168.22,P < 0.01).The apoptosis index of high concentrations transfected HepG2 cells was 25.54%,higher than that of blank control group,negative control group,low dose group and medium dose group(5.24%,6.61%,12.63% and 15.64%,respectively).HepG2 cells transfected with SiRNA exhibit gradually decreasing mitochondrial membrane potentials,which then lead to the releasing of Cyt C,following it were the activation of caspase-9 and caspase-3.Conclusion:Survivin performs the function of anti-apoptosis to the HepG2 cells via modulating the apoptosis of mitochondrial.HepG2 cells transfected with SiRNA survivin can significantly induce apoptosis.
