機械循環拉伸對肺癌細胞功能的影響之研究

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姓名

黃元蓉(Yuan-Jung Huang) 查詢紙本館藏

畢業系所

系統生物與生物資訊研究所

論文名稱

機械循環拉伸對肺癌細胞功能的影響之研究
(The effect of mechanical circulation stretching on the function of lung cancer cells)

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摘要(中)

在過去的文章我們知道機械循環拉伸會重新定位細胞骨架結構以及改變細胞的侵襲及爬行速率，所以本實驗的目的是探討機械循環拉伸對人類肺癌細胞功能的影響，而細胞功能的改變是由哪些基因在做調控。本實驗對象分別是人類肺腺癌細胞A549以及人類非小細胞肺癌細胞H322m，將這兩株細胞經由Boxer TM Cyclic Stretch Culture System (ATMS-Boxer)機械拉伸儀器進行拉伸實驗，拉伸的時間設定在0h、4h和24h，頻率設定在15%，0.5HZ。對拉伸與未拉伸的細胞分別做細胞侵襲、細胞遷移和細胞形成細胞群之能力以及DNA微陣列的分析。從實驗結果我們發現A549和H322m在經由機械循環拉伸四小時會促進細胞的侵襲能力(p-value < 0.05)，接著我們藉由DNA微陣列的分析，分析出CXCL1、 FOS、 CCL2、 CCL20、 BIRC3、TNFAIP3這6個基因在肺癌細胞拉伸四小時後與未拉伸的細胞相比有顯著性的差異(fold change > 2 or < -2 )。而這些基因大部分都是促進細胞侵襲能力，DNA微陣列的分析結果我們發現在拉伸四小時的細胞中都是表現量較高，這些拉力調控基因在路徑分析中發現可能藉由TNF信號通路來調控細胞的侵襲能力。

摘要(英)

In the past, we known that mechanical cyclic stretching can induce reorganization of cytoskeleton and changed the cell migration and invasion. Therefor we identified the cyclic stretch-regulated gene expression in human lung cancer cell lines. Human lung cancer cell line A549 and H322m were subjected to cyclic strain (the frequency set in 15%, 0.5HZ) on 0h, 4h and 24h. We compared invasion assay, migration assay, colony formation and microarray assay in stretching and non-stretching groups. The results showed that the cell invasion in A549 and H322m were increased after 4hr stretching (p-value < 0.05). The microarray data showed that CXCL1, FOS, CCL2, CCL20, BIRC3, and TNFAIP3 were significant differences after 4hr stretching (fold change >2 or <-2). These genes have been reported to enhance cell invasion and these genes were cyclic stretch-regulated genes. Pathway enrichment analysis have found that these genes might regulate cells invasion through the TNF signaling pathway.

關鍵字(中)

★ 肺癌細胞
★ 機械循環拉伸
★ 細胞侵襲
★ 長片段非編碼RNA

關鍵字(英)

★ lung cancer cells
★ mechanical circulation stretching
★ cell invasion
★ long non-coding RNA

論文目次

目錄
中文摘要 ii
Abstract iii
目錄 iv
圖目錄 vi
表目錄 vii
一、緒論 1
1-1 非小細胞肺癌 1
1-2 非小細胞肺癌肺癌成因 1
1-3 肺腺癌治療方法 2
1-4 細胞機械循環拉伸 3
1-5 DNA微陣列分析 5
1-6 KEGG資料庫 5
1-7 長片段非編碼RNA 6
二、實驗方法與材料 7
2-1實驗方法 7
2-1-1細胞循環拉伸實驗 7
2-1-2 細胞侵入 7
2-1-3 細胞爬行 7
2-1-4 細胞群落的形成 8
2-1-5 細胞生長速率 8
2-1-6 qPCR sybr green 8
2-1-7 西方墨點法 9
2-1 實驗材料 10
2-2-1 細胞培養 10
2-2-2 週期性細胞拉伸系統 10
2-2-3 細胞功能和RNA萃取套件 11
三、實驗流程 12
四、實驗結果 14
4-1 細胞拉伸實驗結果 14
4-1-1 A549和H322m 經由機械循環拉伸四小時後會促進細胞的侵襲能力 14
4-1-2 H322m 經由機械循環拉伸後會抑制細胞的遷移能力 15
4-1-3 機械循環拉伸對於細胞形成群落的速率 18
4-1-4 機械循環拉伸影響細胞的生長速率 20
4-2 共五十九個基因在A549和H322m拉伸四小時後表現量的方向性一致 21
4-3 在這五十九個基因有6個基因都位在TNF signaling pathway的下游 25
4-4 利用qPCR以及西方墨點法來應證DNA微陣列分析的結果 33
4-5 細胞在拉伸四小時後LINC00707、LINC00941和LINC01296的表現量上升 34
4-6 LINC00707促進肺腺癌細胞的增生以及促進癌細胞的遷移能力 35
4-7 LINC00941的過表達會促進細胞發生上皮變間質型的轉換 35
4-8 LINC01296會促進細胞侵襲和遷移，抑制細胞凋亡 35
4-9 機械拉伸對人類肺癌是否有持續性的影響 37
五、討論與結論 38
5-1 TNF signaling pathway與細胞侵襲能力之間的關係 38
5-2 長時間機械循環拉伸抑制細胞的生長速率以及侵襲速率 39
5-3 機械循環拉對細胞有持續性的影響 40
5-4 lncRNA透過調控目標基因來調節細胞功能 40
5-5 結論 41
六、參考文獻 42

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指導教授

許藝瓊(Yi-Chiung Hsu)

審核日期

2018-8-21

推文