iPS cells是指誘導性多能性幹細胞，具有與胚胎幹細胞(ES cells)相似的功能，目前已經可以從人類體組織製造出來，解決了胚胎幹細胞一直以來牽涉的倫理議題。研究指出，將Oct3/4、Sox2、c-Myc和Klf4四種基因一起送入人體的纖維母細胞後，能使人類體細胞進行重新組成，產生類似胚胎幹細胞的多能性幹細胞。先前在本實驗室研究癌症幹細胞時，在免疫組織化學染色法(immunohistochemistry；IHC)檢測下，發現人體的正常口腔黏膜組織有表現Oct3/4，所以在本實驗中，想更進一步確認人體口腔黏膜組織是否的確會產生Oct3/4，甚至更同時表現了其他iPS細胞特定的因子，進而研究口腔黏膜纖維母細胞(oral mucosa fibroblast；OMF)是否能成為製造iPS之好的來源。
我們用免疫組織化學染色法檢測口腔黏膜組織切片，發現口腔黏膜組織有Oct3/4和Sox2的陽性反應。接著，我們培養口腔黏膜纖維母細胞，進一部探討是否口腔黏膜纖維母細胞也會表現Oct3/4與Sox2。我們從細胞收取蛋白質後，利用西方點墨法(Western blotting)做檢測，發現細胞並不表現Oct3/4及Sox2。儘管如此，由於口腔的細胞取得容易，所以我們假設，如果口腔細胞可以成功產生iPS細胞，對於製造iPS細胞而言，口腔細胞仍舊是一個很好的來源。因此我們想要利用慢病毒(lentivirus)系統，藉由病毒感染的方式將Oct3/4、Sox2、c-Myc和Klf4四種基因送入口腔纖維母細胞，期望能利用口腔的細胞成功產生iPS細胞，以致將來有助於研究iPS細胞對於疾病的治療。
此外，我們也同時利用現有的老鼠iPS細胞，探討iPS細胞在疾病治療上的應用。我們使用帶有綠色螢光基因的老鼠iPS細胞，先將iPS細胞誘導分化成為造骨細胞(osteoblast)，再將此分化後的細胞利用人工骨移植材料(骨粉)填補到老鼠頭蓋骨的缺損上，使之運用在老鼠頭蓋骨缺損的修復，接著使用免疫組織化學染色法觀察頭蓋骨的修復情形，並使用綠色螢光蛋白(GFP)染色，驗證這些細胞是從帶有綠色螢光基因的老鼠iPS細胞分化而來的。另外，我們也利用此老鼠iPS細胞帶有綠色螢光基因的特性，使用非侵入性活體影像偵測系統(in vivo imaging system；IVIS)，觀察iPS細胞分化後的造骨細胞在老鼠體內分部的位置，進一步了解iPS細胞在牙科及顏面外科治療上的應用。

Induced pluripotent stem (iPS) cells, which show similar functions to embryonic stem cells, can now be derived from human adult somatic tissues. Human embryonic stem cells research involves many ethical issues all the time, and the production of iPS cells now can avoid these issues. Recent researches have shown that introduction of Oct3/4, Sox2, c-Myc, and Klf4 into human fibroblast can reprogram human somatic cells, and generate ES-like pluripotent stem cells. In the previous studies of cancer stem cells in our laboratory, we found human normal oral mucosa tissues expressed Oct3/4 based on immunohistochemistry (IHC). In this research, we want to further confirm that whether human oral mucosa tissues can indeed produce Oct3/4, even more other defined factors, to study if oral mucosa fibroblast (OMF) is a good source to produce iPS cells.
At first, we analyzed oral mucosa tissues by IHC and found that oral mucosa tissues expressed Oct3/4 and Sox2 by IHC. Then we used cell culture to further study whether oral mucosa fibroblast also expressed Oct3/4 and Sox2. We collected cell lysate from oral mucosa fibroblast to perform Western blotting and found OMF didn’t express Oct3/4 and Sox2 distinctly. Even so, we considered that it’s easy to get the oral cells. We hypothesized that if we can produce iPS cells by oral mucosa fibroblast successfully, oral mucosa fibroblast are still a good source to produce iPS cells. We tried to produce lentivirus to introduce Oct3/4, Sox2, c-Myc, and Klf4 into oral fibroblast. We expected to produce iPS successfully by oral cells and it will be useful to study disease therapy.
On the other hand, we used mouse iPS cells, which we bought from Japan, to study the application of iPS cells on disease therapy. We tried to induce the mouse iPS cells, which expressed green fluorescence protein (GFP), to differentiate into osteoblast. Then we supplied the osteoblast to cranial osseous defect on mice by HA-TCP to enhance the regeneration of bony defect. We analyzed the repair of bony tissue by IHC, and we stained the bony tissue with green fluorescent protein to confirm these cells were differentiated by iPS cells which expressed green fluorescent protein. In addition, we also studied the distribution of these cells in the animal model by observing the position of the green fluorescence by using IVIS (in vivo imaging system). By this research, we’ll further understand the application of iPS cells on oral and maxillofacial surgery therapy.

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