口腔癌是台灣第六大常見癌症，而在男性中更位居第四。如同其它許多癌症，口腔鱗狀細胞癌也被推測含有一亞群的腫癌起始細胞，其擁有幹細胞的特性，也就是所謂的癌症幹細胞。為了鑑定這群細胞的存在，我們使用五個可能性的標定物，分別為β1 integrin, CD44, active β-catenin, SOX2以及Oct 3/4，並利用免疫化學組織染色(immunohistochemistry, IHC)來觀察其在十位口腔癌病人的組織切片上之表現。我們認為β1 integrin是其中最適合作為研究口腔癌細胞的標記，並希望進一步確認是否也能成為口腔癌幹細胞的標誌。我們使用幾種鑑定癌症幹細胞的方法，來找出β1 integrin和口腔癌幹細胞的相關性，像是之前已有研究利用細胞株中不同的細胞形態和標誌表現來區分幹細胞。因此，比較β1 integrin在不同形態上的表現，我們發現β1 integrin在被認為含有幹細胞的holoclone上的表現量的確較其它形態高。而在分析細胞形成懸浮球體的實驗中，我們看到β1 integrin在形成球體的細胞表現量確實也較多。此外，我們使用BrdU的長期標定來鑑定幹細胞的存在，並利用免疫組織化學染色發現β1 integrin和BrdU在口腔癌病人的組織切片中有共同的表現位置，證實幹細胞可能就位於表現有β1 integrin的口腔癌細胞中。接著，我們想更進一步觀察β1 integrin對於癌細胞致癌力的影響。我們利用流式細胞儀來將口腔癌細胞分成β1 integrin的高表現群及低表現群，藉由在免疫缺乏的老鼠皮下注入1×104個細胞量，比較腫瘤的生成變化。經由二個月的實驗觀察，我們發現注射高表現β1 integrin的地方都有腫瘤的形成，雖然大小並不一致，但平均上都大於注射β1 integrin低表現群的細胞。我們將腫瘤取下做包埋切片，在免疫組織染色中看到高表現β1 integrin細胞長成的腫瘤，其β1 integrin表現量也比較多。我們進一步利用二株口腔細胞株，OECM-1和SCC 15來探討β1 integrin在in vitro所扮演的角色。我們利用siRNA來降低β1 integrin在細胞株的表現，發現此二株細胞在β1 integrin被抑制的情況下，細胞功能有著不同的改變。我們認為β1 integrin在不同細胞株上的影響可能和其主要的integrin二聚體組成有關，畢竟β1 integrin也只是二聚體其中的一個單位。同時，我們也研究laminin和β1 integrin在口腔癌上的相關性，尤其它們互為ligand和receptor，而且也有文獻指出laminin和癌症是有相關的。最後，我們想比較β1 integrin的表現量在口腔癌臨床上分期的相關性。從初步結果發現，隨著分期越嚴重，β1 integrin在口腔癌細胞中表現的比例也越高。總而言之，β1 integrin在口腔癌細胞上不論是in vitro或是in vivo都有著一定的角色，然而是否在口腔癌幹細胞也扮演著重要角色則有待更深入的探索。

Oral squamous cell carcinoma (OSCC) has become the sixth most common prevalent cancer in Taiwan, and the fourth prevalent cancer among men. Like many other tumors, it has been speculated that OSCC contains a subpopulation of tumor-initiating cells that possess the characteristics of stem cells, therefore so called cancer stem cells (CSCs). To identify the existence of this group of cells, we used five candidate markers, β1 integrin, CD44, active β-catenin, SOX2, and Oct 3/4 to investigate their expression on ten oral cancer patients by immunohistochemistry (IHC). We found that β1 integrin is a putative marker for oral cancer cells among the five molecules, and we tried to further confirm whether β1 integrin is a marker for oral cancer stem cells.
Previous studies had demonstrated the stem cells patterns in cell lines by the heterogeneous patterns of cell morphology, and marker expression. Therefore, we compared the expression of β1 integrin on different morphology, and found that β1 integrin indeed has higher expression on holoclones which are thought to contain stem cells. In the sphere assay, we also observed the higher expression of β1 integrin on cancer cells which can form spheres. Moreover, we identified stem cells by long-term labelling with bromodeoxyuridine (BrdU), and found the co-localization of BrdU and β1 integrin on oral cancer cells derived from OSCC patients by immunohistochemistry. This finding proved the possibility of oral cancer stem cells in oral cancer cells expressed β1 integrin.
Next, we wanted to further observe the carcinogenicity of cancer cells with different expression of β1 integrin. We separated oral cancer cells into two different groups by flow cytometry: one has higher expression of β1 integrin and the other has lower expression. We tried to compare the tumor size grown in defferent groups by subcutaneous injection of 1×104 cells into SCID mice. After two month observation, we found all the sites of SCID mice injected cancer cells with high expression of β1 integrin had tumer formation. Although the tumor size was varied, the average tumor size was bigger in mice injected high expression of β1 integrin. In addtion, the expression of β1 integrin in tumor grown from mice injected cancer cells with high expression of β1 integrin was higher than the other group.
We further investigated its role in vitro using two oral cancer cells, OECM-1 and SCC15. We used siRNA to knockdown the expression of β1 integrin, and found the different results on cell function assay. We considered the effect of β1 integrin in two different cell lines are associatd with the function of main heterodimer subunits, since β1 integrin is only one subunit of integrins heterodimers. Meanwhile, we also investigated the correlation of laminin with β1 integrin in oral cancer, for they also exist the relationship of the ligands and the receptors, and previous studies revealed the correlation between laminin and cancer.
Finally, we tried to find the correlation of expression of β1 integrin with the clinical stage of oral cancer patients. In the fundamental data, we could see the percentage of cancer cells expressing β1 integrin became higher when the clinical stage became worse. Overall, β1 integrin has a role in oral cancer cells both in vitro and in vivo. Whether it also plays an important role in oral cancer stem cells needs to be further explored.

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