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Title: | BCL10對於口腔癌進程之調控機制探討 The Regulation Mechanism of B Cell Lymphoma/Leukemia 10 in Oral Squamous Cell Carcinoma |
Authors: | Tai-Sheng Wu 吳泰昇 |
Advisor: | 張正琪(Cheng-Chi Chang) |
Keyword: | 口腔鱗狀上皮細胞癌,移行,侵入,生長,B細胞淋巴癌10號,S100P, oral cancer,invasion,migration,proliferation,BCL10,S100P, |
Publication Year : | 2012 |
Degree: | 碩士 |
Abstract: | 目的:BCL10已知表現量與口腔鱗狀癌細胞病患的惡性程度成正相關,所以我們的目標在於探討BCL10如何調控口腔鱗狀癌細胞的進程與以及機制的運作。
實驗設計:利用SAS、CA9-22、HSC3、TW2.6以及Cal27五株口腔癌細胞株來探討口腔癌因BCL10表現量被小髮夾型核醣核酸抑制後的移行能力、侵入能力以及生長能力是否有受到影響。在細胞移動能力的測定上,我們利用傷口癒合實驗及博登細胞移行器實驗來做分析;生長能力則是使用細胞存活率分析、細胞群落形成分析以及流式細胞儀分析等方法做測定。動物模式實驗是利用皮下細胞注射的方式來觀察抑制BCL10的表現是否會影響口腔癌細胞的致癌能力。 結果:細胞實驗中發現BCL10的表現量與細胞侵入能力成正相關,抑制SAS以及CA9-22細胞中BCL10的表現量會使細胞移行能力、侵入能力以及生長能力下降約40% (P < 0.05),並且造成細胞生長週期停滯在G0/G1期 (P < 0.05)。經由微小核醣核酸陣列分析找到下游分子S100P,S100家族的其中一員,可以顯著恢復細胞因BCL10表現量被抑制而造成的移動能力及生長能力的下降,移行以及侵入能力方面有約70%的恢復,生長速率也有30%的恢復 (P < 0.05)。在動物實驗方面,BCL10被抑制的組別其腫瘤與對照組相比縮小了約60%,而將其細胞中的S100P表現量回復成對照組則使得腫瘤大小恢復成趨近對照組的大小 (P < 0.05)。訊息傳遞方面發現BCL10被抑制時P65的活性以及表現量明顯下降,而在其細胞中加入帶有S100P的載體則可以回復P65的表現量以及活性。 結論:本研究確認了在口腔鱗狀上皮細胞癌中,BCL10可藉由活化S100P的蛋白質表現量進而使NF-kB的訊息傳遞路徑活化造成癌細胞進程。 Purpose: B-cell lymphoma/leukemia 10 (BCL10) is elevated in advanced oral squamous cell carcinoma (OSCC) patient and positive correlated with TNM stage. However, the regulatory mechanism of BCL10 in OSCC progression is completely unknown. Methods: SAS, CA9-22, HSC3, TW2.6, and Cal27 oral cancer cell lines were used as in vitro models to investigate cell phenotypes, including invasion, migration, and proliferation by BCL10 shRNA knockdown. Wound healing analysis and Boyden chamber assay were performed to check migration and invasion abilities. Proliferation ability was identified by MTT assay, colony formation, and flow cytometry in vitro. Mice were s.c. injected with shBCL10-silencing or vector control SAS cells to measure tumorigenicity. Results: BCL10 expression was positively correlated with invasion ability in OSCC cells. Transiently transfected SAS and CA9-22 cells with shBCL10 knockdown plasmid significantly reduced migration, invasion ability, and proliferation ability, approximately 40 % reduction of migration, 40 % reduction of invasion, and 30 % reduction of proliferation abilities in SAS and CA9-22 cells, compared to vector control cells (all P < 0.05). Furthermore, we established shBCL10-stable transfectants in SAS and CA9-22 cells, and found that silencing BCL10 could significantly down-regulated migration, invasion, and proliferation abilities, approximately 40% reduction of migration, invasion ,and proliferation (all P < 0.05). Cell cycle arrest was also detected in BCL10-silenced transfectants, cells were 20 % increase of G0/G1 phase and 10 % decrease of G2/M phase in SAS/shBCL10 and CA9-22/shBCL10 transfectants, compared to vector control cells (P < 0.05). Additionally, we performed high throughput mRNA microarray analysis and identified a downstream effecter S100P, a member of S100 protein family after transiently transfected with S100P expression plasmid could restore migration, invasion, and proliferation abilities in shBCL10-stable transfectant conformed (all P < 0.05). Moreover, we re-expressed S100P in shBCL10-stable transfectant could significantly restore the migration, invasion, and proliferation ability, approximately 70% induction of migration and invasion, and 30% induction of proliferation abilities in SAS/shBCL10 cells (P < 0.05). Tumor volume was decreased in s.c. injected shBCL10-stable transfectant, and restored S100P expression reversed BCL10- enhanced tumorigenicity in vivo. Conclusion: BCL10 promotes OSCC progression via induced S100P-dependent signaling. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/64693 |
Fulltext Rights: | 有償授權 |
Appears in Collections: | 口腔生物科學研究所 |
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