臺灣博碩士論文加值系統

本研究的目的主要是建立一個人工轉移的模型，來檢視胸部放射線（6 MV X-ray）照射是否會影響腫瘤在肺部的轉移。我們採用C3H/HeN小鼠做為實驗動物，每隻老鼠以靜脈注射（i.v.）的方式注入1x10E5 顆NFsa腫瘤細胞，在注入細胞後的第8天犧牲老鼠，並計數肺表面的肺群落數。對於照射組，我們給予小鼠胸部20 Gy X-ray的劑量，並在照射後立刻以靜脈注射的方式注入腫瘤細胞。在8次重覆的實驗中，照射組肺表面的肺群落數是未照射組的3.18±1.26倍（p = 0.02）；而從肺的切片來看，照射組的每單位面積的正常肺組織中轉移腫瘤的個數以及全部腫瘤面積佔全部肺切面面積的比例都比未照射組高，這和我們所觀察到的肺表面群落數的結果相似。為了測試是否預先胸部放射線照射可以促進腫瘤的成長速率，我們將肺臟切片以H&E染色後，計算腫瘤群落的大小分布，結果發現照射組比未照射組有出現較大腫瘤的傾向。為了測試劑量對轉移的影響，我們從0 Gy、6 Gy、12 Gy到20 Gy逐漸增加胸部劑量，結果發現即使在6 Gy的較低劑量，預先給予胸部照射仍然可以促進肺群落的生成，但效果並沒有20 Gy來的強。為了測試這腫瘤被促進的現象是否與時間有關，我們在照射後的不同時間，注射NFsa腫瘤細胞，結果發現在照射後立刻或者是在照射後第1天注射腫瘤細胞都會促進腫瘤在肺部的轉移，而隨著照射放射線與注射腫瘤細胞之間的時間間隔加長，轉移的能力有下降的趨勢，當時間間隔長到1個月時，照射組的肺部轉移並沒有顯著的增加。最後，我們發現aspirin並不能有效地抑制由胸部放射線照射所引起的肺群落數增加的現象。

The aim of this research was to establish an artificial metastatic model to study the influence of pre-irradiation (6 MV X-ray) of thorax on the process of metastasis in lung. C3H/HeN mice were injected i.v. with 1x10E5 NFsa cells/mouse, sacrificed at the 8th day after injection. The number of lung colony at lung surface was counted. For irradiated group, the mice’ thorax was irradiated 20 Gy X-ray and injected i.v. NFsa tumors cells immediately after irradiation. In 8 repeated experiments, the ratio of the number of lung colonies at surface between irradiated group and non-irradiated group was 3.18±1.26 (p = 0.02). In lung sections, the number of metastatic tumors per normal lung area and the ratio between total tumor area and total normal lung area in irradiated group were both higher than those of the non-irradiated. The ratios were similar to that we observed in the number of lung colonies at lung surface. To evaluate if pre-irradiation promote the growth rate of tumor, the lung section was stained with H&E and the distribution of the size of lung tumor was calculated. We observed that irradiated group had more potential to get bigger tumors than those of non-irradiated group. To evaluate the dose-response, we gave the grading radiation doses including 0 Gy, 6 Gy, 12 Gy and 20 Gy. The results showed that even at lower dose of 6 Gy, the formation of lung colony was still promoted by the pre-irradiation of thorax but the effect was less obvious than that at 20 Gy. To evaluate if the promotion of tumor is time-dependent, we injected NFsa tumor cells at various time after irradiation. Immediately or at the first day after irradiation of thorax, we found both can promote metastasis in lung. When the time interval between irradiation and injection of tumor cells became longer; the ability of metastasis was lower and no significant increase of lung metastasis was found if time interval was up to 1 month. Finally, we found that aspirin cannot inhibit the promotion of metastasis induced by pre-irradiation of thorax.

目 錄
英文摘要……………………………………………………… I
中文摘要……………………………………………………… II
誌謝…………………………………………………………… III
目錄…………………………………………………………… IV
圖目錄………………………………………………………… VI
表目錄……………………………………………………… VII
第一章 緒論……………………………………………… 1
1.1 腫瘤的轉移過程……………………………… 1
1.1.1 癌症與治療………………………… 1
1.1.2 癌症的轉移…………………………… 1
1.1.3 在肺部發生的轉移事件……………… 3
1.1.4 放射線照射與轉移的關係…………… 4
1.2 胸部放射線照射………………………… 5
1.2.1 放射線照射………………………. 5
1.2.2 肺部放射線照射引起的發炎和纖維化…… 5
1.3 研究目的………………………… 7
第二章 實驗方法……………………………………… 8
2.1 即時肺部放射線照射對NFsa腫瘤在肺部轉移的影響 8
2.1.1 分組…………………………………….. 8
2.1.2 細胞培養………………………………… 8
2.1.3 細胞的準備………………………………… 8
2.1.4 小鼠的肺部照射……………………………… 9
2.1.5 從靜脈注射腫瘤……………………… 9
2.1.6 樣品收集和處理…………………… 9
2.1.7 組織包埋及切片……………………………… 10
2.1.8 數位照相系統及影像分析……………………… 10
2.2 不同的肺部照射劑量對NFsa腫瘤在肺部轉移的影響 10
2.3 照射後不同時間對NFsa腫瘤在肺部轉移的影響 11
2.4 Aspirin抗發炎藥對NFsa腫瘤在肺部轉移的影響 11
2.5 NSAID對細胞株raw 264.7中介白素一號（IL-1β）分泌量的影響 11
2.5.1 細胞培養…………………………………………. 12
2.5.2 實驗方法………………………………. 12
2.5.3 介白素一號（IL-1β）的酵素連結免疫吸附法（enzyme-linked immunosorbent assay，ELISA）………………… 12
2.5.4 BCA protein assay……………………………… 13
2.6 Total RNA的萃取…………………………………… 14
2.6.1 使用Solution D來萃取肺的total RNA………………… 14
2.6.2 使用TRIZOL來萃取raw 264.7 cell的total RNA……… 14
2.7 統計方法……………………………………………. 15
第三章 結果……………………………………………… 16
3.1 肺部照射後NFsa腫瘤轉移至肺表面的數目…………… 16
3.2 肺部照射的劑量與NFsa腫瘤轉移到肺表面數目的關係… 16
3.3 照射後與腫瘤轉移的時間曲線（time course） 17
3.4 胸部照射後肺內部腫瘤轉移的情形………………… 18
3.5 Aspirin及放射線對轉移的影響………………………… 19
3.6 Dexamethasone對raw 264.7細胞株分泌mIL-1β的影響… 20
第四章 討論……………………………………………. 21
第五章 未來展望…………………………………. 25
第六章 參考資料………………………………………………. 27
第七章 附錄………………………………………………… 29
圖 目 錄
圖一C3H/HeN小鼠被施打NFsa腫瘤細胞株後，肺表面及肺內部的肺群落照片
(照射組及未照射組)……………………………….. 30
圖二施打腫瘤前給予胸部20 Gy照射引起肺表面的肺群落數目 31
圖三不同照射劑量對NFsa腫瘤在C3H/HeN小鼠肺部的轉移能力. 32
圖四不同時間胸部照射對NFsa腫瘤在C3H/HeN小鼠肺部的轉移能力 33
圖五 NFsa腫瘤群落在肺切片中的大小分佈圖……………. 34
圖六 胸部放射線照射及Aspirin對肺表面群落數的影響 35
表 目 錄
表一 胸部照射放射線後的肺表面及肺內部肺群落的大小及數目在照射組和未照射組之間的比例關係表…………………………… 36

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