本文選題：膠質(zhì)瘤　切入點(diǎn)：PARP-1　出處：《中南大學(xué)》2014年博士論文　論文類型：學(xué)位論文

【摘要】：目的：本實(shí)驗(yàn)擬研究人腦膠質(zhì)瘤組織中PARP-1mRNA的表達(dá)；觀察給予替莫唑胺、PARP-1抑制劑BMN-673后膠質(zhì)瘤細(xì)胞中PARP-1的表達(dá)；然后對(duì)膠質(zhì)瘤U251、U87細(xì)胞系予以PARP-1抑制劑BMN-673與替莫唑胺單獨(dú)用藥及聯(lián)合用藥,通過彗星實(shí)驗(yàn)觀察細(xì)胞的DNA損傷；通過CCK-8試驗(yàn)、流式細(xì)胞術(shù)觀察細(xì)胞增殖抑制、細(xì)胞凋亡及細(xì)胞周期阻滯水平,以初步探討PARP-1抑制劑BMN-673與替莫唑胺聯(lián)用治療膠質(zhì)瘤的可能機(jī)制,并為其臨床治療提供基礎(chǔ)理論依據(jù)。 方法：收集中南大學(xué)湘雅醫(yī)院神經(jīng)外科2013年10月至2014年01月35例新鮮膠質(zhì)瘤、5例新鮮正常腦組織標(biāo)本,用定量RT-PCR測(cè)定其腫瘤正常組組織間的PARP-1mRNA表達(dá)差異及與膠質(zhì)瘤病理分級(jí)的關(guān)系。對(duì)惡性星形膠質(zhì)瘤細(xì)胞系U251、惡性膠質(zhì)母細(xì)胞瘤細(xì)胞系U87予以替莫唑胺、PARP-1抑制劑BMN-673處理,并通過RT-PCR測(cè)定其PARP-1的表達(dá)情況。對(duì)膠質(zhì)瘤細(xì)胞予以替莫唑胺、BMN-673單獨(dú)及聯(lián)合用藥,通過彗星實(shí)驗(yàn)檢測(cè)DNA損傷,CCK-8試驗(yàn)檢測(cè)細(xì)胞生長抑制、流式細(xì)胞術(shù)檢測(cè)細(xì)胞凋亡及細(xì)胞周期阻滯情況。 結(jié)果：膠質(zhì)瘤組織中PARP-1mRNA的表達(dá)水平比正常腦組織顯著升高,具有統(tǒng)計(jì)學(xué)意義(P0.01)；腫瘤不同級(jí)別間PARP-1mRNA表達(dá)量存在差異,具有統(tǒng)計(jì)學(xué)意義(P=0.01)。對(duì)膠質(zhì)瘤細(xì)胞予以替莫唑胺、BMN-673及聯(lián)合用藥后,PARP-1mRNA表達(dá)水平較正常培養(yǎng)組存在差異,具有統(tǒng)計(jì)學(xué)意義(P0.05)。替莫唑胺與BMN-673聯(lián)合用藥后,U251、U87細(xì)胞的DNA損傷加重,增殖受到明顯抑制,細(xì)胞凋亡增加,細(xì)胞周期G2/M期受到阻滯。 結(jié)論：1. PARP-1mRNA的表達(dá)在膠質(zhì)瘤組織中明顯增高,且與病理級(jí)別呈正相關(guān)；2.予以PARP-1抑制劑BMN-673并與常用化療藥物TMZ聯(lián)用,膠質(zhì)瘤細(xì)胞中PARP-1mRNA表達(dá)升高;3.PARP-1抑制劑BMN-673對(duì)替莫唑胺在膠質(zhì)瘤細(xì)胞治療中有增敏作用。PARP-1有望成為膠質(zhì)瘤分子靶向治療的一個(gè)治療靶點(diǎn),PARP-1抑制劑BMN-673有望成為DNA損傷類化療藥物的增敏劑。
[Abstract]:Objective: to study the expression of PARP-1mRNA in human glioma tissues and to observe the expression of PARP-1 in glioma cells after treatment with temozolomide PARP-1 inhibitor BMN-673. Then the U251 U87 cell line was treated with PARP-1 inhibitor BMN-673 and temozolidomide alone or in combination. The DNA damage was observed by comet assay, and the inhibition of cell proliferation was observed by CCK-8 assay and flow cytometry. In order to explore the possible mechanism of the combination of PARP-1 inhibitor BMN-673 and temozolamide in the treatment of glioma and provide the basic theoretical basis for its clinical treatment. Methods: 35 cases of fresh normal brain tissue were collected from October 2013 to January 2014 in Xiangya Hospital of Central South University. Quantitative RT-PCR was used to determine the difference of PARP-1mRNA expression in normal tissues and its relationship with pathological grade of glioma. The malignant astrocytoma cell line U251 and malignant glioblastoma cell line U87 were treated with temozolomide PARP-1 inhibitor BMN-673. The expression of PARP-1 in glioma cells was determined by RT-PCR. The glioma cells were treated with temozolamine BMN-673 alone or in combination. DNA damage was detected by comet assay and CCK-8 assay was used to detect the growth inhibition of glioma cells. Apoptosis and cell cycle arrest were detected by flow cytometry. Results: the expression of PARP-1mRNA in gliomas was significantly higher than that in normal brain tissues (P 0.01). There were significant differences in the expression of PARP-1mRNA between the glioma cells treated with temozolamide BMN-673 and those of the normal culture group. The DNA damage of U251 U87 cells was aggravated after the combination of temozolidomide and BMN-673. Proliferation was significantly inhibited, cell apoptosis was increased and G 2 / M phase of cell cycle was blocked. Conclusion the expression of PARP-1mRNA was significantly increased in glioma tissues, and was positively correlated with pathological grade. BMN-673, a PARP-1 inhibitor, was used in combination with TMZ, a common chemotherapeutic drug. Increased expression of PARP-1mRNA in glioma cells. 3. PARP-1 inhibitor BMN-673 can enhance the sensitivity of temozolamide in glioma cells. PARP-1 may become a therapeutic target for glioma molecular targeted therapy. PARP-1 inhibitor BMN-673 may become a kind of DNA damage. An sensitizer for a therapeutic drug.
【學(xué)位授予單位】：中南大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2014
【分類號(hào)】：R739.4

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本文編號(hào)：1590354