目的:探討TGF-β1抑制的microRNAs及其靶基因蛋白通過MAPK信號通路調(diào)節(jié)人類橫紋肌肉瘤（RMS）肌分化及其分子機制,為RMS分化治療提供新的治療靶點。方法:（1）采用Real-time PCR技術檢測TGF-β1抑制的microRNAs在不同RMS細胞株和組織樣本中的表達,篩選出差異最明顯、最廣泛的microRNA作為研究對象。（2）利用MTT、3H胸腺嘧啶核苷摻入法檢測microRNA對RMS細胞株增殖的影響。（3）構建裸鼠成瘤性模型,觀察microRNA對活體腫瘤生長的影響;采用免疫組織化學染色檢測瘤體ki-67蛋白表達情況。（4）運用生物信息學技術及網(wǎng)絡資源預測miR-411-5p的靶基因（5）運用Real-time PCR技術、雙熒光素酶報告系統(tǒng)及蛋白印跡法驗證靶基因。（6）構建Elk-1-Luc、c-Jun-Luc熒光報告質(zhì)粒,SPRY4-siRNA或siRNA-ctrl轉染PKC?表達質(zhì)粒處理的RMS細胞株,檢測Elk-1-Luc、c-Jun-Luc熒光信號;PKC?表達質(zhì)粒和SPRY4-siRNA/siRNA-ctrl共轉染RMS細胞株,檢測P-p38蛋白的變... 

【文章來源】：蘇州大學江蘇省 211工程院校

【文章頁數(shù)】：83 頁

【學位級別】：碩士

【部分圖文】：

驗證TGF-β1基因敲低效果

TGF-β1/miR-411-5p/SPRY4和 MAPK信號通路參與橫紋肌肉瘤分化調(diào)表達明顯增加 (圖 2 )。miR-411-5p 和 miR-411-3p 在 RMS、更普遍，因此選擇 miR-411-5p 在 RMS 組織中進一步驗

RT-PCR檢測miR-411-5p六對低/高TGF-β1表達的ARMS和八對ERMS組織標本

【參考文獻】：
期刊論文
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[2]Inhibition of Differentiation by Transforming Growth Factor β1 in Rhabdomyosarcoma Cells[J]. Shouli Wang1 Huihua Yao2 Zhenghong Qin3 Shigang Li1 Yizhong Feng1 Xiuzhen Wang1 Min Deng1 Lingling Guo1 Lifeng Zhang1 1 Molecular Pathology Laboratory,Medical School of Soochow University,Suzhou 215123,China. 2 Department of Surgery,the Second Affiliated Hospital of Soochow University,Suzhou 215123,China. 3 Pharmacology Laboratory,Medical School of Soochow University,Suzhou 215123,China..  Chinese Journal of Clinical Oncology. 2007(05)
[3]Smad4 and ERK2 stimulated by transforming growth factor betal in rhabdomyosarcoma[J]. GUO Hua ZHANG Hong-ying WANG Shou-li YE Lü YANG Guang-hua BU Hong Department of Pathology,West China Hospital,Sichuan University,Chengdu 610041,China (Zhang HY,Wang SL,Ye L,Yang GH and Bu H)Department of Pathology,Peking University First Hospital,Beijing 100034,China (Guo H)Department of Pathology,Medical School,Soochow University,Suzhou 215007,China (Wang SL).  Chinese Medical Journal. 2007(06)



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