本文選題：維康醇　切入點：前列腺癌　出處：《中國人民解放軍醫(yī)學(xué)院》2014年博士論文　論文類型：學(xué)位論文

【摘要】：目的： 對于去勢抵抗性前列腺癌目前尚無有效的治療手段,亟待開發(fā)一種新型的化療藥物。本研究主要通過體外實驗來評估新型天然化合物維康醇對于惡性前列腺細(xì)胞的抗腫瘤作用,并進一步探討其藥物作用機制。 材料與方法： 通過蛋白免疫印跡及流式細(xì)胞計數(shù)等實驗方法評估維康醇在不同前列腺癌細(xì)胞系及正常前列腺上皮誘導(dǎo)凋亡的作用,然后通過反應(yīng)活性氧(Reactive oxygen species,ROS)熒光染料檢測三株具有代表性的細(xì)胞系PC3,DU145,BPH1添加不同種類ROS清除劑后細(xì)胞內(nèi)ROS水平的變化,探討其作用途徑。最后為了證明維康醇誘導(dǎo)的細(xì)胞凋亡是Bax蛋白依賴性的,我們通過對PC3細(xì)胞系進行RNA干擾及外源性導(dǎo)入Bax蛋白天然缺失的細(xì)胞系DU145,評估維康醇處理細(xì)胞后的凋亡指標(biāo)。最后通過透射電鏡及應(yīng)用不同種類Caspase蛋白酶抑制劑探討了Bax蛋白可能的下游途徑。 結(jié)果： 通過對凋亡標(biāo)志物的檢測,如Caspase-3及PARP確認(rèn)了維康醇誘導(dǎo)的細(xì)胞死亡方式為凋亡。ROS熒光染色結(jié)果發(fā)現(xiàn)維康醇誘導(dǎo)了前列腺癌細(xì)胞內(nèi)廣泛的氧化應(yīng)激并活化凋亡途徑最終導(dǎo)致細(xì)胞死亡。然而,維康醇誘導(dǎo)的這種細(xì)胞凋亡可被ROS清除劑N-乙酰-L-半胱氨酸(NAC)及二氫硫辛酸(DHLA)完全阻斷。我們同時證明了Bax蛋白在維康醇誘導(dǎo)的細(xì)胞凋亡中激活并發(fā)揮著重要的作用,維康醇誘導(dǎo)的細(xì)胞凋亡均伴有Bax蛋白的激活。對于Bax缺失的前列腺癌細(xì)胞系DU145在外源性導(dǎo)入Bax質(zhì)粒后,其對維康醇的抗藥性便會消失,再次證明維康醇引起的凋亡是Bax依賴途徑。激活的Bax蛋白最終導(dǎo)致線粒體損傷,最后通過Caspase依賴及非依賴途徑誘導(dǎo)前列腺癌細(xì)胞的凋亡。這些結(jié)果表明為維康醇將來可能作為一種新型治療晚期前列腺癌化療藥物奠定了理論基礎(chǔ)。 結(jié)論： 維康醇可誘導(dǎo)前列腺癌細(xì)胞內(nèi)廣泛的氧化應(yīng)激反應(yīng)最終導(dǎo)致Bax依賴性的細(xì)胞凋亡。
[Abstract]:Objective:. There is no effective treatment for ovariectomized prostate cancer. It is urgent to develop a new chemotherapeutic drug. The aim of this study was to evaluate the antitumor effect of a new natural compound, veconol, on malignant prostate cells in vitro, and to further explore the mechanism of its action. Materials and methods:. The effects of veconol on apoptosis in different prostate cancer cell lines and normal prostate epithelium were evaluated by Western blotting and flow cytometry. Then the changes of ROS levels in three representative cell lines PC3DU145BPH1 were detected by reactive oxygen speciesrosis (Ros) after adding different kinds of ROS scavengers. Finally, in order to prove that the apoptosis induced by veconol is Bax protein-dependent, We evaluated the apoptosis index of PC3 cell line by RNA interference and exogenous introduction of Bax protein deletion cell line DU145. finally, we applied different kinds of Caspase protease inhibitor by transmission electron microscope and different kinds of Caspase protease inhibitor. The possible downstream pathway of Bax protein was discussed. Results:. By detecting the markers of apoptosis, For example, Caspase-3 and PARP confirmed that Vicol induced cell death was apoptosis. Ros fluorescence staining results showed that Vicol induced extensive oxidative stress and activated apoptosis pathway in prostate cancer cells, and eventually resulted in cell death. The apoptosis induced by veconol can be completely blocked by ROS scavenger N-acetyl-L-cysteine) and dihydrolipoxylic acid (DHla). We have also demonstrated that Bax protein is activated and plays an important role in the apoptosis induced by veconol. The apoptosis induced by veconol was accompanied by the activation of Bax protein. For prostate cancer cell line DU145 with Bax deletion, its resistance to veconol disappeared after exogenous transfection into Bax plasmid. It is proved that the apoptosis induced by veconol is Bax dependent pathway. The activated Bax protein eventually leads to mitochondrial damage. Finally, apoptosis of prostate cancer cells was induced by Caspase dependent and non-dependent pathway. These results suggest that Viconol may be a new chemotherapeutic agent for advanced prostate cancer in the future. Conclusion:. Vicol can induce extensive oxidative stress in prostate cancer cells and eventually lead to Bax-dependent apoptosis.
【學(xué)位授予單位】：中國人民解放軍醫(yī)學(xué)院
【學(xué)位級別】：博士
【學(xué)位授予年份】：2014
【分類號】：R737.25

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本文編號：1600991