本文選題：神經(jīng)膠質(zhì)瘤 + 化療藥物敏感性　； 參考：《福建醫(yī)科大學(xué)》2014年碩士論文

【摘要】：目的： 1.研究Notch1基因?qū)θ四z質(zhì)瘤U251細(xì)胞株化療藥物敏感性的影響。 2.人膠質(zhì)瘤中Notch1與多藥耐藥基因MDR、MRP1表達(dá)的相關(guān)性。 方法： 1.構(gòu)建NICD高表達(dá)質(zhì)粒載體和Notch1sh-RNA質(zhì)粒載體，通過(guò)慢病毒包裝、轉(zhuǎn)染、篩選，分別獲得高表達(dá)NICD的U251細(xì)胞株和Notch1基因干擾的U251細(xì)胞株。 2. MTT法檢測(cè)上述模型細(xì)胞及對(duì)照U251細(xì)胞對(duì)化療藥物BCNU、VM-26的敏感性。 3. Q-PCR、Western blot、ICC方法檢測(cè)U251細(xì)胞、NICD高表達(dá)的U251細(xì)胞及Notch1基因干擾的U251細(xì)胞中多藥耐藥基因MDR1、MRP1的表達(dá)，以及BCNU、VM-26處理后，各組細(xì)胞Notch1、MDR1、MRP1基因表達(dá)的變化，并分析其相關(guān)性。 4. IHC檢測(cè)人膠質(zhì)瘤組織標(biāo)本中Notch1、MDR1、MRP1基因表達(dá)，分析其相關(guān)性。結(jié)果： 1.成功建立了高表達(dá)NICD和Notch1基因干擾的U251細(xì)胞株。 2. MTT實(shí)驗(yàn)表明NICD高表達(dá)U251細(xì)胞（plvx-NICD組）與空載對(duì)照細(xì)胞（plvx組）相比，其藥物的半數(shù)抑制濃度IC50值升高，而Notch1基因干擾的U251細(xì)胞（sh-NA組）與對(duì)照組細(xì)胞（sh-NC組）相比，其IC50值降低，表明高表達(dá)NICD使U251細(xì)胞有更強(qiáng)的藥物抗性，低表達(dá)Notch1細(xì)胞其耐藥性減弱，，對(duì)兩種化療藥物更加敏感。 3. Q-PCR、Western Blot、免疫熒光染色檢測(cè)均表明NICD高表達(dá)U251細(xì)胞中耐藥基因MDR1、MRP1的表達(dá)水平均高于對(duì)照組細(xì)胞，而Notch1基因干擾的U251細(xì)胞，MDR1、MRP1表達(dá)均下調(diào)。 4.免疫組化實(shí)驗(yàn)結(jié)果顯示人腦膠質(zhì)瘤組織中存在多藥耐藥基因MDR1、MRP1的表達(dá)，表達(dá)水平與膠質(zhì)瘤病理級(jí)別呈正相關(guān)。在Ⅰ、Ⅱ、Ⅲ級(jí)膠質(zhì)瘤中與Notch1基因的表達(dá)具有相關(guān)性。 結(jié)論： 1. Notch1基因參與調(diào)節(jié)神經(jīng)膠質(zhì)瘤對(duì)化療藥物BCNU、VM-26的敏感性。 2.在人膠質(zhì)瘤U251細(xì)胞中Notch1基因可以影響多藥耐藥基因MDR1、MRP1的表達(dá)，并可能通過(guò)調(diào)節(jié)多藥耐藥基因MDR1、MRP1的表達(dá)導(dǎo)致腫瘤細(xì)胞的耐藥。 3.人腦膠質(zhì)瘤組織中存在多藥耐藥基因MDR1、MRP1的表達(dá)，且在Ⅰ、Ⅱ、Ⅲ級(jí)膠質(zhì)瘤中與Notch1基因的表達(dá)呈正相關(guān)。
[Abstract]:Objective: 1. To study the effect of Notch1 gene on chemosensitivity of human glioma cell line U251. 2. Correlation between Notch1 and multidrug resistance gene MDR- MRP1 expression in human glioma. Methods: 1. NICD high expression plasmid vector and Notch1sh-RNA plasmid vector were constructed. U251 cell lines with high expression of NICD and U251 cell lines interfered with Notch1 gene were obtained by lentivirus packaging, transfection and screening. 2. The sensitivity of U251 cells to BCNUVM-26 was detected by MTT assay. 3. The expression of multidrug resistance gene MDR1 and MRP1 in U251 cells with high expression of NICD and interference with Notch1 gene was detected by Q-PCR- Western blotnICC, and the expression of Notch1, MDR1, MRP1 was analyzed after treated with BCNUVM-26, and the correlation between MDR1 and MDR1MRP1 in U251 cells was analyzed. 4. IHC was used to detect the expression of Notch1, MDR1 and MRP1 gene in human glioma tissues and its correlation was analyzed. Results: 1. U251 cell lines with high expression of NICD and Notch1 genes were successfully established. 2. MTT assay showed that the half inhibitory concentration (IC50) of U251 cells with high expression of NICD (plvx-NICD) was higher than that of unloaded control cells (plvx), but the IC50 value of Notch1 gene interfering U251 cells sh-na) was lower than that of control group (sh-NC). The results showed that high expression of NICD resulted in stronger drug resistance in U251 cells and decreased drug resistance of low expression Notch1 cells, which was more sensitive to two chemotherapeutic drugs. 3. Q-PCR Western blot and immunofluorescence staining showed that the expression level of MDR1- MRP1 in U251 cells was significantly higher than that in control cells, but the expression of MDR1MRP1 was down-regulated in U251 cells with interference of Notch1 gene. 4. The results of immunohistochemistry showed that there was a positive correlation between the expression of MDR1and MRP1 in human glioma tissues and the pathological grade of gliomas. The expression of Notch1 gene in grade 鈪

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