上皮細胞黏著分子EpCAM在胚胎幹細胞的功能EpCAM as a reprogramming factor for non-pluripotent cells

專利類型

發明

專利國別 (專利申請國家)

美國

專利申請案號

12/930,154

專利證號

US 8,709,806 B2

專利獲證名稱

上皮細胞黏著分子EpCAM在胚胎幹細胞的功能EpCAM as a reprogramming factor for non-pluripotent cells

專利所屬機關 (申請機關)

中央研究院

獲證日期

2014/04/29

技術說明

人類胚胎幹細胞為一群具全功能性的細胞，能自我更新並分化成三個胚層。本研究中，我們利用一株對抗上皮細胞黏著分子 (EpCAM) 單株抗體OC98-1，透過免疫螢光顯微、西方點漬法與流式細胞分析廣泛地研究EpCAM的表現。我們發現EpCAM高度專一地表現在未分化的幹細胞，而不表現在已分化的細胞中。其蛋白與轉錄體 (transcript) 在幹細胞進行分化時即減少。此一調控機制與徹底地轉換組織蛋白 (histone) 後修飾的作用緊密相關。進一步地，我們證明兩個調控組織蛋白3離氨酸27三甲基化 (lysine 27 trimethylation of histone 3 ; H3K27me3) 的分子，SUZ12與JMJD3，可作用在EpCAM起始子上。此外，我們也利用核染質免疫沈澱分析 (chromatin immunoprecipitation; ChIP) 證明EpCAM可直接調控數個改編基因 (reprogramming gene)，包括c-Myc、Oct-4、Nanog、Sox2及Klf4，以幫助維持幹細胞特性。EpCAM在幹細胞內的表現，是透過表觀遺傳機制 (epigenetic mechanism) 調控，維持幹細胞特性。綜合以上結果，我們認為EpCAM可作為幹細胞的表面標記抗原及製備誘導式多能性幹細胞(induced pluripotent stem cells, iPS cells)。 Human embryonic stem cells (hESCs) are unique pluripotent cells capable of self-renewal and differentiation into all three germ layers. We found epithelial cell adhesion molecule (EpCAM) to be highly and selectively expressed by undifferentiated rather than differentiated hESCs. The protein and transcript level of EpCAM rapidly diminished as soon as hESC had differentiated. Moreover, we demonstrated that the dynamic pattern of lysine 27 trimethylation of histone 3 (H3K27me3) was conferred by the interplay of SUZ12 and JMJD3, both of which were involved in maintaining hESC pluripotency. In addition, we used chromatin immunoprecipitation (ChIP) analysis to elucidate EpCAM’s direct regulation of several reprogramming genes, including c-Myc, Oct-4, Nanog, Sox2 and Klf4, to help maintain the stemness of hESCs. Collectively, our results indicate that EpCAM can be used as a stem cell surface marker and useful for generation of induced pluripotent stem (iPS) cells.

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