胚胎幹細胞是屬於具有多能性的幹細胞，主要是由囊胚層裡的內細胞層而 來，而且可以分化成三個胚層和大部份的細胞類型，為了要維持幹細胞的多能性 與自我更新需要外在的調控因子如LIF、BMP，和內在的因子如Oct4 和Nanog。 Oct4 又稱Oct3 是屬於POU 的轉錄因子，之前的研究指出Oct4 只表現在幹細胞中。 Nanog 為一 homoebox 的轉錄因子，在胚胎發育的時候Nanog 在內細胞層的多能 性與細胞命運的決定中扮演著重要的腳色，我們在 C2C12 中大量表現 Oct4 和 Nanog 後發現在細胞進行分化過程中，其肌管的數目明顯下降，其中在 C2C12-Oct4 的穩定細胞株中其分化能力比 control 低，而在同時表現 Oct4 和 Nanog 的穩定細胞株中，並沒有觀察到肌管的產生。在 fusion index 的數據中， 在大量表現 Oct4 時和 control 相比下降了 60% 而且在同時表現 Oct4 和 Nanog 時幾乎趨近於零。 在RT-PCR 的資料中我們也觀察到 MRF 基因家族在 大量表現 Oct4 的穩定細胞株中表現量下降，而在同時表現oct4 和Nanog 時表 現量的下降有更為增強的趨勢，在胚胎幹細胞的特異性基因的表現情況中發現在 大量表現 Oct4 和 Nanog 的穩定細胞株中其表現量上升。Pax7 在大量表現Oct4 的穩定細胞株中表現量下降，中胚層的marker CD34 和血液幹細胞的marker Sca1 在C2C12-Oct4-py-Nanog 細胞株中表現量顯著的上昇。在未來的研究中我們希望 找出Oct4 和Nanog 影響C2C12 分化的分子機制。 Embryonic stem (ES) cells are pluripotent cell derived from the inner cell mass (ICM) of blastocysts and can differentiate into most cell types. To maintaine pluripotency and self-renewal they need extrinsic regulators, such as LIF and BMP, and intrinsic factors, like Oct4 and Nanog. Oct4, also know as Oct3, belong to the POU (pit-oct-uno) transcription factor family. Recent studies indicate that Oct4 is only expressed in stem cells. Nanog is a unique homoebox transcription factor and plays a critical role in regulating cell fate of the pluripotent ICM during embryonic development. We over-expressed Oct4 and Nanog in myoblasts and found that their myogenic differentiation was morphologically repressed. C2C12-Oct4 stable clone differentiated poorer than control and no myotubes was detected in cells overexpressing Oct4 and Nanog . The fusion index in C2C12-Oct4 was repressed by 60% and almost not detectable in Oct4/Nanog stable clone. Using RT-PCR, we found MRF families were down-regulated in Oct4 stable clone as compared with control and their expression were repressed farther in double expression stable clone. ES cell marker expression was increased in double expression stable clone. Pax7 is repressed in C2C12-Oct4 stable clone. Mesoderm marker CD34 and hematopoietic stem cell marker gene Sca1 were significantly enhanced in C2C12-Oct4-py-Nanog cells. In the future, we like to find out the underlying molecular mechanisms by which Oct4 and Nanog influence C2C12 differentiation.
