Objective:To explore the influence of EBP50(ezrin-radixin-moesin-binding phospho-protein-50) on microfilament cytoskeleton content and distribution in cultured Hela cells, and to investigate the relationship between t...Objective:To explore the influence of EBP50(ezrin-radixin-moesin-binding phospho-protein-50) on microfilament cytoskeleton content and distribution in cultured Hela cells, and to investigate the relationship between the changes in microfilament cytoskeleton localization and EBP50 after PDGF(platelet-derived growth factor) stimulation, and to further clarify the molecular mechanism by which EBP50 suppresses tumor cell proliferation and migration.Methods:pBK-CMV-HAEBP50 wild type recombinant plasmid and pBK-CMV-HA empty vector were transfected into Hela cells.G418 at 350 mg/L was used to screen for cell clones stably expressing EBP50.Western blot was carried out to detect EBP50 expression.Similarities and differences in microfilament cytoskeleton content and distribution in Hela cells transfected with pBK-CMV-HA-EBP50 wild type recombinant plasmid and pBK-CMV-HA empty vector were also treated with PDGF(10 ng/mL and 20 ng/mL, 37 ℃, 15 min) and stained by rhodamine-labeled phalloidin to observe the distribution of microfilament cytoskeleton in the two groups.EBP50 protein distribution in PDGF-stimulated Hela cells was detected by immunofluorescence.Results:Western blot results confirmed that the EBP50 cDNA fragment could express EBP50 in cultured Hela cell lines and that cell lines stably expressing EBP50 were successfully obtained.Western blot and fluorescence results showed that in the cell line transfected with empty vector, the microfilament cytoskeleton was thick, loose, multidirectional and displayed crossing arrangements.The content of microfilament cytoskeleton in the cell line transfected with pBK-CMV-HA-EBP50 was different from that found in the cell line transfected with empty vector.EBP50 expression enhanced microfilament cytoskeleton polymerization into compact thin filaments.Under the stimulation of PDGF, EBP50 migrated to the cell membrane from the cytosol together with microfilament cytoskeleton and co-localized there.Conclusion:EBP50 can change the distribution of microfilament cytoskeleton in cult展开更多
目的探讨EBP50基因在胰腺癌中的表达及临床意义。方法应用Real-time PCR检测20例胰腺癌及其癌旁组织和10例正常胰腺组织中的EBP50 m RNA水平,并采用量子点技术对120例胰腺癌及其癌旁组织,10例正常胰腺组织中EBP50蛋白的表达进行检测,并...目的探讨EBP50基因在胰腺癌中的表达及临床意义。方法应用Real-time PCR检测20例胰腺癌及其癌旁组织和10例正常胰腺组织中的EBP50 m RNA水平,并采用量子点技术对120例胰腺癌及其癌旁组织,10例正常胰腺组织中EBP50蛋白的表达进行检测,并对120例胰腺癌患者的5年生存率进行分析。结果 EBP50 m RNA及蛋白表达率在胰腺癌组织中明显低于癌旁非癌组织及正常胰腺组织;与正常胰腺组织比较,癌旁组织EBP50 m RNA及蛋白表达率均降低(P<0.05);EBP50蛋白高表达胰腺癌患者的预后生存期明显长于低表达患者(P<0.05)。结论 EBP50在胰腺癌中低表达,表明其是一种抑癌基因,可作为胰腺癌治疗的新靶点,值得深入研究。展开更多
基金Supported by grants from the National Natural Science Foundation ofChina (No. 30572183 and 30772573)Beijing Educational Committee Foundation (No. KZ200610025013)+2 种基金the New Century Excellent Talentsin University of China (No. NCEF-06-0184)Excellent Talents Foundation in Beijing (No. 20071D0501800253)Beijing Novagramme Foundation (No. 2008B58 [MS1]).
文摘Objective:To explore the influence of EBP50(ezrin-radixin-moesin-binding phospho-protein-50) on microfilament cytoskeleton content and distribution in cultured Hela cells, and to investigate the relationship between the changes in microfilament cytoskeleton localization and EBP50 after PDGF(platelet-derived growth factor) stimulation, and to further clarify the molecular mechanism by which EBP50 suppresses tumor cell proliferation and migration.Methods:pBK-CMV-HAEBP50 wild type recombinant plasmid and pBK-CMV-HA empty vector were transfected into Hela cells.G418 at 350 mg/L was used to screen for cell clones stably expressing EBP50.Western blot was carried out to detect EBP50 expression.Similarities and differences in microfilament cytoskeleton content and distribution in Hela cells transfected with pBK-CMV-HA-EBP50 wild type recombinant plasmid and pBK-CMV-HA empty vector were also treated with PDGF(10 ng/mL and 20 ng/mL, 37 ℃, 15 min) and stained by rhodamine-labeled phalloidin to observe the distribution of microfilament cytoskeleton in the two groups.EBP50 protein distribution in PDGF-stimulated Hela cells was detected by immunofluorescence.Results:Western blot results confirmed that the EBP50 cDNA fragment could express EBP50 in cultured Hela cell lines and that cell lines stably expressing EBP50 were successfully obtained.Western blot and fluorescence results showed that in the cell line transfected with empty vector, the microfilament cytoskeleton was thick, loose, multidirectional and displayed crossing arrangements.The content of microfilament cytoskeleton in the cell line transfected with pBK-CMV-HA-EBP50 was different from that found in the cell line transfected with empty vector.EBP50 expression enhanced microfilament cytoskeleton polymerization into compact thin filaments.Under the stimulation of PDGF, EBP50 migrated to the cell membrane from the cytosol together with microfilament cytoskeleton and co-localized there.Conclusion:EBP50 can change the distribution of microfilament cytoskeleton in cult
文摘目的探讨EBP50基因在胰腺癌中的表达及临床意义。方法应用Real-time PCR检测20例胰腺癌及其癌旁组织和10例正常胰腺组织中的EBP50 m RNA水平,并采用量子点技术对120例胰腺癌及其癌旁组织,10例正常胰腺组织中EBP50蛋白的表达进行检测,并对120例胰腺癌患者的5年生存率进行分析。结果 EBP50 m RNA及蛋白表达率在胰腺癌组织中明显低于癌旁非癌组织及正常胰腺组织;与正常胰腺组织比较,癌旁组织EBP50 m RNA及蛋白表达率均降低(P<0.05);EBP50蛋白高表达胰腺癌患者的预后生存期明显长于低表达患者(P<0.05)。结论 EBP50在胰腺癌中低表达,表明其是一种抑癌基因,可作为胰腺癌治疗的新靶点,值得深入研究。
