醣蛋白B7H3在口腔癌細胞的表現與作用

Expression and Effects of Glycoprotein B7H3 in Oral Cancer Cells

顧可立(Ko-Li Ku)

指導教授 ： 陳敏慧

國立臺灣大學/牙醫專業學院/口腔生物科學研究所/碩士(2010年)

https://doi.org/10.6342/NTU.2010.02684

摘要

口腔癌居全球癌症發病率及死亡率的第六名，且每年的死亡率不斷的攀升，而當口腔癌發生轉移時，能治癒的機會又是微乎其微，因此，研究口腔癌轉移的機制是非常重要的。近年來的研究發現在許多癌症中，醣蛋白B7H3會有過量的表現，且這些表現可能與癌症的癌化 (cancer progression)、轉移以及低存活率有關。B7H3是一種位於細胞膜上的醣蛋白，除了會表現於免疫細胞，許多癌細胞也有B7H3蛋白的表現。本實驗主要探討口腔鱗狀細胞癌細胞株中B7H3的表現量，並以siRNA將B7H3抑制後，觀察癌細胞的移動、貼附以及增生能力是否改變。
首先，由西方墨點法發現口腔癌細胞株Ca-9-22、SAS、CAL27以及HSC3的醣蛋白表現普遍比正常細胞S-G多，之後藉由質譜儀分析以及西方墨點法找出並確定B7H3在口腔癌細胞株有大量的蛋白質表現。為了更進一步探討B7H3於癌細胞株的作用為何，本實驗利用siRNA轉染的方式抑制癌細胞Ca9-22和SAS表現B7H3，並觀察此處理對癌細胞株移動、貼附以及增生能力的影響。本實驗首先以細胞傷口癒合分析 (wound healing assay)觀察細胞的平移能力，發現轉染後細胞的平移能力有下降。接著利用細胞遷移能力分析 (migration assay) 以及侵襲能力 (invasion assay) 進行分析，結果顯示B7H3抑制後對癌細胞株Ca9-22以及SAS的遷移和侵襲能力是有抑制效果的。在細胞貼附能力方面，Ca9-22和SAS癌細胞株對fibronectin與laminin的貼附能力有下降的趨勢，而在collagen Ⅳ方面的貼附能力雖然有降低，但於統計上並無顯著差異 (p > 0.05)。另外，細胞增生實驗 (MTT assay) 的結果顯示，B7H3對細胞的增生能力並沒有影響。總結上述實驗結果可知，醣蛋白B7H3對口腔癌轉移可能扮演著重要的角色，極有潛力成為口腔癌轉移的標記而對其診斷治療有很大的幫助。

關鍵字

口腔癌 ； 醣蛋白 ； B7H3 ； siRNA ； 細胞轉移 ； 細胞侵襲 ； 細胞貼附

並列摘要

The incidence and mortality of oral cancer is the sixth in the worldwide, and the mortality is continuing rising each year. Once metastasis is occurred in oral cancer, the opportunity for patient to be cured becomes indistinct. Therefore, investigating of the mechanism of oral cancer metastasis is very important. There are some studies report the over-expression of glycoprotein B7H3 in many cancers. It was also indicated that the over-expression of B7H3 might be related with the progression, metastasis, and low survival rate of cancer. B7H3 is a membrane glycoprotein, it can be expressed in immune cells and many cancer cells. In this study, the expression of B7H3 in oral squamous cell carcinoma cell lines was investigated. Small interfering RNA (siRNA) was applied for inhibiting the expression of B7H3 and the changes of mobility, adhesion, and proliferation of cancer cells were also investigated.
In this study, we found that the expression of glycoprotein in Ca-9-22, SAS, CAL27, HSC3 oral cancer cells is more than that in S-G normal cells with significant difference. Identification and confirmation of glycoprotein B7H3 was performed by mass analysis and western blot analysis. For further investigating the function of B7H3 in cancer cells, siRNA was transfected into cancer cells in Ca9-22 and SAS to knockdown B7H3, and the changes of the ability of mobility, adhesion, and proliferation of cancer cells were observed. The results of wound healing assay showed that the migration ability of transfected cells were lower than that of control cancer cells without siRNA transfection. Furthermore, migration and invasion assay also showed that B7H3 knockdown could reduce the migration and invasion potentials of Ca9-22 and SAS cancer cells. It was also found that cell adhesion in fibronectin and laminin is decreased. Whether B7H3 was knockdown in cancer cells, there is no significant effect on adhesion to collagen Ⅳ (p > 0.05). Besides, the MTT data showed that B7H3 has no effect on cell proliferation. In conclusion, glycoprotein B7H3 plays an important role in oral cancer cells especially in cell migration and invasion. We suggest that B7H3 is a potential marker and might be useful for oral cancer therapy.

並列關鍵字

oral cancer ； glycoprotein ； B7H3 ； siRNA ； migration ； invasion ； adhesion

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