中文摘要：

研究甲胎蛋白（α-fetoprotein，AFP）对肝癌细胞内PTEN／AKT信息通路信号传递的影响．用Western blotting法分析全反式维甲酸（all trarts retinoic acid，ATRA）处理人肝癌Bel 7402和HepG2细胞24h后PTEN表达的变化．免疫共沉淀（Co—IP）技术研究AFP与PTEN相互作用．激光共聚焦显微镜观察AFP与PTEN在细胞共定位．RNA干扰（RNAi）技术抑制AFP表达，再用ATRA处理24h后检测细胞内PTEN表达的变化，并分析蛋白激酶B（AKT）的磷酸化．用pcDNA3．1质粒和人afp基因连接构建表达AFP的载体（称为pcDNA3．1-afp），然后转染到不表达AFP的人肝癌HLE细胞．结果显示，人肝癌Bel 7402和HepG2细胞均有PTEN的表达，ATRA（160μmol／L）处理24h后能促进这些细胞的PTEN表达．co—IP技术研究发现AFP能与PTEN结合．共聚焦显微镜观察显示AFP与PTEN共定位于细胞浆．干扰AFP表达后，PTEN表达明显提高．抑制AFP表达后，ATRA能显著促进Bel 7402细胞内PTEN的表达，并能抑制AKT的磷酸化．转染pcDNA3．1-afp载体后，HLE细胞内有AFP表达，并与PTEN结合，且发现pcDNA3．1-afp载体能增加AKT的磷酸化[p-AKT（Ser473）]，对抗ATRA抑制HLE细胞增殖．研究的结论是：肝癌细胞内表达的AFP能与PTEN结合并抑制PTEN对AKT的去磷酸化作用，肝癌细胞内高表达的AFP能激活AKT信息通路．胞浆内的AFP是肝癌细胞耐受ATRA的重要因子．

英文摘要：

α-Fetoprotein （AFP） has a property to maintain the growth of hepetocellular cancinoma cells（HCC） in vivo, but the critical functional step of AFP is still obscurity. In order to explore AFP influences on the transduction of PTEN/AKT signal in HCC, expression of PTEN in human hepatoma cells and the effect of all trans retinoic acid （ATRA） on PTEN expression in these cancer cells were detected by Western blotting. AFP interacted with PTEN was analyzed by co-immunoprecipitation （Co-IP）. Laser confocal microscopy was used to observe co-localization of AFP and PTEN. Short small RNA interfering （RNAi） was applied to knockdown the expression of AFP in Bel 7402 cells, and the phosphorylation of protein kinase B （AKT） was detected by Western blotting. Growth of Bel 7402 and HLE cells were assayed by MTT. Results showed that Bel 7402 cells expressed PTEN, and the expression of PTEN was induced by ATRA（160μmol/L） mildly. Co-IP indicated that AFP has a property to interact with PTEN. PTEN co-localization with AFP was observed in cytoplasm of Bel 7402 cells. Constructed RNAi vector could knockdown the expression of AFP, expression of PTEN was promoted and phosphorylation of AKT was decreased when the expression of AFP was interfered or the cells were treated with ATRA（160 p, mol/L）. AFP-expressed vector pcDNA3.1-afp was transfected into human hepatoma HLE cells （AFP-non production）. Co-IP analysis indicated that AFP interacted with PTEN, and expression of p-AKT（Ser473） was promoted in the tumor cells. It was also proved that pcDNA3.1-afp was able to reduce the role of Ly294002 in inhibiting the activity of AKT in HLE cells. These data provide the first evidence that AFP has a capacity to both interact with and inhibit activity of PTEN, which is also the pivotal events in the process that AFP activated the transduction of PI3FUAKT signal in hepatoma cells. Cytoplasmic AFP plays importance role on the resistance to ATRA induced apoptosis of HCC.