AIM: To assess the impact of eukaryotic elongation factor 1 alpha 2 (eEF1A2) on hepatocellular carcinoma (HCC) cell proliferation, apoptosis, migration and invasion, and determine the underlying mechanisms.METHODS: eE...AIM: To assess the impact of eukaryotic elongation factor 1 alpha 2 (eEF1A2) on hepatocellular carcinoma (HCC) cell proliferation, apoptosis, migration and invasion, and determine the underlying mechanisms.METHODS: eEF1A2 levels were detected in 62 HCC tissue samples and paired pericarcinomatous specimens, and the human HCC cell lines SK-HEP-1, HepG2 and BEF-7402, by real-time PCR and immunohistochemistry. Experimental groups included eEF1A2 silencing in BEL-7402 cells with lentivirus eEF1A2-shRNA (KD group) and eEF1A2 overexpression in SK-HEP-1 cells with eEF1A2 plasmid (OE group). Non-transfected cells (control group) and lentivirus-based empty vector transfected cells (NC group) were considered control groups. Cell proliferation (MTT and colony formation assays), apoptosis (Annexin V-APC assay), cell cycle (DNA ploidy assay), and migration and invasion (Transwell assays) were assessed. Protein levels of PI3K/Akt/NF-κB signaling effectors were evaluated by Western blot.RESULTS: eEF1A2 mRNA and protein levels were significantly higher in HCC cancer tissue samples than in paired pericarcinomatous and normal specimens. SK-HEP-1 cells showed lower eEF1A2 mRNA levels; HepG2 and BEL-7402 cells showed higher eEF1A2 mRNA levels, with BEL-7402 cells displaying the highest amount. Efficient eEF1A2 silencing resulted in reduced cell proliferation, migration and invasion, increased apoptosis, and induced cell cycle arrest. The PI3K/Akt/NF-κB signaling pathway was notably inhibited. Inversely, eEF1A2 overexpression resulted in promoted cell proliferation, migration and invasion.CONCLUSION: eEF1A2, highly expressed in HCC, is a potential oncogene. Its silencing significantly decreases HCC tumorigenesis, likely by inhibiting PI3K/Akt/NF-κB signaling.展开更多
Enterovirus A71(EV-A71) is the major pathogen responsible for the severe hand, foot and mouth disease worldwide, for which few effective antiviral drugs are presently available. Interferon-a(IFN-a) has been used in an...Enterovirus A71(EV-A71) is the major pathogen responsible for the severe hand, foot and mouth disease worldwide, for which few effective antiviral drugs are presently available. Interferon-a(IFN-a) has been used in antiviral therapy for decades;it has been reported that EV-A71 antagonizes the antiviral activity of IFN-a based on viral 2 Apro-mediated reduction of the interferon-alpha receptor 1(IFNAR1);however, the mechanism remains unknown. Here, we showed a significant increase in IFNAR1 protein induced by IFN-a in RD cells, whereas EV-A71 infection caused obvious downregulation of the IFNAR1 protein and blockage of IFN-a signaling. Subsequently, we observed that EV-A71 2 Apro inhibited IFNAR1 translation by cleavage of the eukaryotic initiation factor 4 GI(eIF4GI), without affecting IFNAR1 m RNA levels induced by IFN-a. The inhibition of IFNAR1 translation also occurred in puromycin-induced apoptotic cells when caspase-3 cleaved e IF4 GI. Importantly, we verified that 2 Aprocould activate cellular caspase-3, which was subsequently involved in e IF4 GI cleavage mediated by 2 Apro. Furthermore, inhibition of caspase-3 activation resulted in the partial restoration of IFNAR1 in cells transfected with 2 A or infected with EV-A71, suggesting the pivotal role of both viral 2 Aproand caspase-3 activation in the disturbance of IFN-a signaling. Collectively, we elucidate a novel mechanism by which cellular caspase-3 contributes to viral 2 Apro-mediated down-regulation of IFNAR1 at the translation level during EV-A71 infection, indicating that caspase-3 inhibition could be a potential complementary strategy to improve clinical anti-EV-A71 therapy with IFN-a.展开更多
目的探讨真核翻译延长因子1α1(eTEF1α1)基因在反式二羟环氧苯并芘致癌机制中所起的作用。方法采用逆转录聚合酶链反应(RT-PCR)方法扩增真核翻译延长因子1α1基因全长,插入pcDNATM3.1 D irectional TOPO表达载体,以脂质体转染介导的技...目的探讨真核翻译延长因子1α1(eTEF1α1)基因在反式二羟环氧苯并芘致癌机制中所起的作用。方法采用逆转录聚合酶链反应(RT-PCR)方法扩增真核翻译延长因子1α1基因全长,插入pcDNATM3.1 D irectional TOPO表达载体,以脂质体转染介导的技术和G418细胞筛选法转染人支气管上皮细胞,构建转基因稳定表达的细胞株。用半定量RT-PCR方法分析转基因表达产物,对转基因细胞株,进行双层软琼脂试验以确定基因的恶性特性。结果成功扩增出eTEF1α1基因全长,构建出稳定表达真核翻译延长因子1α1基因的细胞株。稳定转染eTEF1α1基因的细胞株,能在双层软琼脂中形成克隆。结论真核翻译延长因子1α1基因与反式二羟环氧苯并芘的恶性转化有关。展开更多
基金Supported by the Middle-Young Age Backbone Talent Cultivation Program of Fujian Health System,No.2013-ZQNJC-2Key Projects of Science and Technology Plan of Fujian Province,No.2014Y0009
文摘AIM: To assess the impact of eukaryotic elongation factor 1 alpha 2 (eEF1A2) on hepatocellular carcinoma (HCC) cell proliferation, apoptosis, migration and invasion, and determine the underlying mechanisms.METHODS: eEF1A2 levels were detected in 62 HCC tissue samples and paired pericarcinomatous specimens, and the human HCC cell lines SK-HEP-1, HepG2 and BEF-7402, by real-time PCR and immunohistochemistry. Experimental groups included eEF1A2 silencing in BEL-7402 cells with lentivirus eEF1A2-shRNA (KD group) and eEF1A2 overexpression in SK-HEP-1 cells with eEF1A2 plasmid (OE group). Non-transfected cells (control group) and lentivirus-based empty vector transfected cells (NC group) were considered control groups. Cell proliferation (MTT and colony formation assays), apoptosis (Annexin V-APC assay), cell cycle (DNA ploidy assay), and migration and invasion (Transwell assays) were assessed. Protein levels of PI3K/Akt/NF-κB signaling effectors were evaluated by Western blot.RESULTS: eEF1A2 mRNA and protein levels were significantly higher in HCC cancer tissue samples than in paired pericarcinomatous and normal specimens. SK-HEP-1 cells showed lower eEF1A2 mRNA levels; HepG2 and BEL-7402 cells showed higher eEF1A2 mRNA levels, with BEL-7402 cells displaying the highest amount. Efficient eEF1A2 silencing resulted in reduced cell proliferation, migration and invasion, increased apoptosis, and induced cell cycle arrest. The PI3K/Akt/NF-κB signaling pathway was notably inhibited. Inversely, eEF1A2 overexpression resulted in promoted cell proliferation, migration and invasion.CONCLUSION: eEF1A2, highly expressed in HCC, is a potential oncogene. Its silencing significantly decreases HCC tumorigenesis, likely by inhibiting PI3K/Akt/NF-κB signaling.
基金grants from Beijing Natural Science Foundation(No.19G10290)National Natural Science Foundation of China(No.81772184).
文摘Enterovirus A71(EV-A71) is the major pathogen responsible for the severe hand, foot and mouth disease worldwide, for which few effective antiviral drugs are presently available. Interferon-a(IFN-a) has been used in antiviral therapy for decades;it has been reported that EV-A71 antagonizes the antiviral activity of IFN-a based on viral 2 Apro-mediated reduction of the interferon-alpha receptor 1(IFNAR1);however, the mechanism remains unknown. Here, we showed a significant increase in IFNAR1 protein induced by IFN-a in RD cells, whereas EV-A71 infection caused obvious downregulation of the IFNAR1 protein and blockage of IFN-a signaling. Subsequently, we observed that EV-A71 2 Apro inhibited IFNAR1 translation by cleavage of the eukaryotic initiation factor 4 GI(eIF4GI), without affecting IFNAR1 m RNA levels induced by IFN-a. The inhibition of IFNAR1 translation also occurred in puromycin-induced apoptotic cells when caspase-3 cleaved e IF4 GI. Importantly, we verified that 2 Aprocould activate cellular caspase-3, which was subsequently involved in e IF4 GI cleavage mediated by 2 Apro. Furthermore, inhibition of caspase-3 activation resulted in the partial restoration of IFNAR1 in cells transfected with 2 A or infected with EV-A71, suggesting the pivotal role of both viral 2 Aproand caspase-3 activation in the disturbance of IFN-a signaling. Collectively, we elucidate a novel mechanism by which cellular caspase-3 contributes to viral 2 Apro-mediated down-regulation of IFNAR1 at the translation level during EV-A71 infection, indicating that caspase-3 inhibition could be a potential complementary strategy to improve clinical anti-EV-A71 therapy with IFN-a.
文摘目的探讨真核翻译延长因子1α1(eTEF1α1)基因在反式二羟环氧苯并芘致癌机制中所起的作用。方法采用逆转录聚合酶链反应(RT-PCR)方法扩增真核翻译延长因子1α1基因全长,插入pcDNATM3.1 D irectional TOPO表达载体,以脂质体转染介导的技术和G418细胞筛选法转染人支气管上皮细胞,构建转基因稳定表达的细胞株。用半定量RT-PCR方法分析转基因表达产物,对转基因细胞株,进行双层软琼脂试验以确定基因的恶性特性。结果成功扩增出eTEF1α1基因全长,构建出稳定表达真核翻译延长因子1α1基因的细胞株。稳定转染eTEF1α1基因的细胞株,能在双层软琼脂中形成克隆。结论真核翻译延长因子1α1基因与反式二羟环氧苯并芘的恶性转化有关。
