解放军医学院学报
解放軍醫學院學報
해방군의학원학보
Academic Journal of Chinese Pla Medical School
2013年
1期
78-81
,共4页
郭恩凯%李冰%申晶%程愈%郝好杰%韩庆旺%伍志强%韩为东%母义明
郭恩凱%李冰%申晶%程愈%郝好傑%韓慶旺%伍誌彊%韓為東%母義明
곽은개%리빙%신정%정유%학호걸%한경왕%오지강%한위동%모의명
Oct4%肝细胞%去分化
Oct4%肝細胞%去分化
Oct4%간세포%거분화
Oct4%hepatocyte%dedifferentiation
目的通过Oct4介导小鼠肝细胞去分化为Sox17阳性细胞,探索肝胰重编程新途径.方法构建慢病毒载体pWPT-Oct4,包装和浓缩病毒;改良两步胶原酶灌流法分离小鼠原代肝细胞;Oct4-慢病毒浓缩液感染小鼠原代肝细胞,RT-PCR检测肝细胞及内胚层转录因子的表达.结果通过酶切、测序鉴定,证实成功构建包含Oct4慢病毒表达载体.感染肝细胞后,外源性Oct4出现表达,成熟的肝细胞转录因子(Alb、Ttr、Afp)表达下降,并表达内胚层早期标记物Sox17,而没有Oct4、Nanog及Sox2等多能性基因的内源性表达.结论 Oct4介导肝细胞去分化,并出现Sox17阳性细胞,为进一步将Sox17阳性细胞分化为胰岛素产生细胞奠定基础.
目的通過Oct4介導小鼠肝細胞去分化為Sox17暘性細胞,探索肝胰重編程新途徑.方法構建慢病毒載體pWPT-Oct4,包裝和濃縮病毒;改良兩步膠原酶灌流法分離小鼠原代肝細胞;Oct4-慢病毒濃縮液感染小鼠原代肝細胞,RT-PCR檢測肝細胞及內胚層轉錄因子的錶達.結果通過酶切、測序鑒定,證實成功構建包含Oct4慢病毒錶達載體.感染肝細胞後,外源性Oct4齣現錶達,成熟的肝細胞轉錄因子(Alb、Ttr、Afp)錶達下降,併錶達內胚層早期標記物Sox17,而沒有Oct4、Nanog及Sox2等多能性基因的內源性錶達.結論 Oct4介導肝細胞去分化,併齣現Sox17暘性細胞,為進一步將Sox17暘性細胞分化為胰島素產生細胞奠定基礎.
목적통과Oct4개도소서간세포거분화위Sox17양성세포,탐색간이중편정신도경.방법구건만병독재체pWPT-Oct4,포장화농축병독;개량량보효원매관류법분리소서원대간세포;Oct4-만병독농축액감염소서원대간세포,RT-PCR검측간세포급내배층전록인자적표체.결과통과매절、측서감정,증실성공구건포함Oct4만병독표체재체.감염간세포후,외원성Oct4출현표체,성숙적간세포전록인자(Alb、Ttr、Afp)표체하강,병표체내배층조기표기물Sox17,이몰유Oct4、Nanog급Sox2등다능성기인적내원성표체.결론 Oct4개도간세포거분화,병출현Sox17양성세포,위진일보장Sox17양성세포분화위이도소산생세포전정기출.
Objective To study the new approaches to liver and pancreas reprogramming by observing the Oct4-mediated dedifferentiation of liver cells into Sox17 positive cells. Methods A lentiviral vector, pWPT-Oct4, was constructed, and lentivirus was packed and concentrated. Primary liver cells in mice were isolated by the modified two-step collagenase perfusion and infected with the concentrated Oct4 lentivirus fluid. Expression of endoderm and hepatic cell transcription factors was detected by RT-PCR. Results Whether the expression vector containing Oct4 lentivirus was successfully constructed was verified by enzyme digestion and sequencing. Exogenous Oct4 was expressed, the expression level of mature transcription factors in liver cells (Alb, Ttr and Afp) decreased, early endoderm marker Sox17 was expressed while endogenous pluripotency genes, Oct4, Nanog, and Sox2 were not expressed in infected primary liver cells. Conclusion Oct4 mediates dedifferentiation of liver cells in which Sox17 positive cells can be observed, thus laying a foundation for the further differentiation of Sox17 positive cells into insulin-producing cells.