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標題: 利用Nuclear Factor-κB活體分子影像及轉錄體學技術探討放化療對宿主之影響
Assessment of Host Responses to Radio-Chemotherapy by Nuclear Factor-κB Bioluminescence Imaging-Guided Transcriptomic Analysis
作者: 張仲達
Chang, Chung-Ta
關鍵字: 5-FU;ionizing radiation;5-ASA;NF-κB;分子影像;DNA微陣列晶片;游離輻射;小腸黏膜炎;nuclear factor-κB;bioluminescence imaging;DNA microarray;5-fluorouracil;intestinal mucositis;5-aminosalicylic acid
出版社: 生命科學系所
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放射治療與化學治療在癌症治療中扮演重要的角色,但是他們與生物體之間的交互作用仍有許多待解的難題。本論文主要是利用nuclear factor-κB (NF-κB)基因轉殖小鼠作為平台,藉助活體冷光分子影像系統監測放射治療與化學治療與生物體之間的交互作用,進一步利用基因微陣列技術,解釋它們的作用機轉。
我們首先構築可以受到NF-κB調控的冷光酵素基因轉殖鼠,再以放射線或化學治療藥物處理並觀測基因轉殖鼠體內NF-κB的活動度,結果發現,經8.5格雷(Gy)放射線照射的小鼠在肝、小腸、及腦組織有較強的NF-κB活動度。接著,我們利用基因微陣列技術,分析這些器官的基因表現圖譜,發現放射線照射主要影響代謝及免疫調控途徑。因為fatty acid binding protein 4, serum amyloid A2及 serum amyloid A3等基因和發炎反應及脂質代謝有關,我們認為放射線照射會造成這些基因的上調(upregulation)並誘發代謝及發炎反應途徑。另外,我們認為chemokine (CC-motif) ligand 5, chemokine (CC-motif) ligand 20及Jagged 1等基因活性上升和放射線腸病變應有關連。
5-Fluorouracil (5-FU)是一種很常用的化學治療藥物,但是約有80%病患使用後會發生消化道黏膜炎。我們將100 mg/kg 5-FU經由腹腔注射誘發NF-κB基因轉殖小鼠產生消化道黏膜炎,接著利用基因網絡分析及活體冷光系統試圖發現誘發5-FU黏膜炎的重要分子(molecule)。結果發現,5-FU腸炎(5-FU-induced intestinal mucositis)除了造成小腸壁變厚、腺窩(crypt)長度增長、絨毛(villus)變短等組織變化外,也會造成組織中骨髓過氧化酶(myeloperoxidase)活性上升及血清中發炎前驅細胞激素產生。綜合基因網絡分析、活體冷光系統、及免疫組織化學染色分析的結果,我們認為NF-κB是5-FU腸炎上游重要的調控因子,而且,當我們用5-aminosalicylic acid (5-ASA)抑制NF-κB的活性之後,與5-FU腸炎相關的發炎前驅細胞激素活性下降,光學顯微鏡下的組織學變化也獲得顯著改善。

Radiotherapy and chemotherapy both play important roles in current cancer therapy, but there are still many unsolved problems about host-therapeutics interaction. The aim of this study was to investigate the host responses to these therapeutics by nuclear factor-κB (NF-κB) bioluminescence imaging-guided transcriptomic tool.
Transgenic mice carrying the NF-κB-driven luciferase gene were exposed to a single dose of 8.5 Gy total-body irradiation. In vivo imaging showed that a maximal NF-κB-dependent bioluminescent intensity was observed at 3 h after irradiation and ex vivo imaging showed that liver, intestine, and brain displayed strong NF-κB activations. Microarray analysis of these organs showed that irradiation altered gene expression signatures in an organ-specific manner and several pathways associated with metabolism and immune systems were significantly altered. Additionally, the upregulation of fatty acid binding protein 4, serum amyloid A2, and serum amyloid A3 genes, which are participated in both inflammation and lipid metabolism, suggested that irradiation might affect the cross pathways of metabolism and inflammation. Moreover, the alteration of chemokine (CC-motif) ligand 5, chemokine (CC-motif) ligand 20, and Jagged 1 genes, which are involved in the inflammation and enterocyte proliferation, suggested that these genes might be involved in the radiation enteropathy.
5-Fluorouracil (5-FU) is a commonly used drug for the treatment of malignant cancers. However, approximately 80% of patients undergoing 5-FU treatment suffer from gastrointestinal mucositis. We wanted to identify the drug target for the 5-FU-induced intestinal mucositis. 5-FU-induced intestinal mucositis was established by intraperitoneally administering mice with 100 mg/kg 5-FU. Network analysis of gene expression profile and bioluminescent imaging were applied to identify the critical molecule associated with 5-FU-induced mucositis. Our data showed that 5-FU induced inflammation in the small intestine, characterized by the increased intestinal wall thickness and crypt length, the decreased villus height, and the increased myeloperoxidase activity in tissues and proinflammatory cytokine production in sera. Network analysis of 5-FU-affected genes by transcriptomic tool showed that the expression of genes was regulated by NF-κB, and NF-κB was the central molecule in the 5-FU-regulated biological network. NF-κB activity was activated by 5-FU in the intestine, which was judged by in vivo bioluminescence imaging and immunohistochemical staining. However, 5-aminosalicylic acid (5-ASA) inhibited 5-FU-induced NF-κB activation and proinflammatory cytokine production. Moreover, 5-FU-induced histological changes were improved by 5-ASA.
In conclusion, this study described the comprehensive evaluation of host responses to ionizing radiation and chemotherapy. Our findings provided the fundamental information about the in vivo NF-κB activity and transcriptomic pattern after irradiation and chemotherapy. We also suggested that NF-κB was the critical molecule associated with the pathogenesis of 5-FU-induced mucositis, and inhibition of NF-κB activity ameliorated the mucosal damage caused by 5-FU. Moreover, novel targets involved in radiation injury were also suggested.
其他識別: U0005-1007201209490000
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