本文選題：紫草素 + NB4細胞��； 參考：《重慶醫(yī)科大學》2017年碩士論文

【摘要】：目的:研究紫草素對人急性早幼粒細胞白血病NB4細胞增殖與凋亡的影響及其相關作用機制。方法:1.以NB4細胞株作為研究對象,用梯度濃度的紫草素分別作用于NB4細胞不同時間后,CCK-8實驗測定細胞增殖情況;2.以0.3μmol/L紫草素作用于NB4細胞,培養(yǎng)1 d后,通過流式細胞術檢測細胞周期的改變和凋亡的改變;3.經Hochest 33342實驗,通過熒光顯微鏡檢測NB4細胞的胞核的形態(tài)學改變;4.Western blot檢測細胞c-Myc,PARP,Caspase 3,Cleaved PARP,Cleaved Caspase 3,ERK,p-ERK,JNK,p-JNK,p38MAPK及p-p38MAPK蛋白表達情況;5.構建NB4細胞裸鼠移植瘤模型,通過描繪腫瘤生長曲線分析紫草素對裸鼠腫瘤發(fā)展情況的作用;6.通過免疫組化技術檢測紫草素對抑制瘤組織中增殖指標c-Myc表達的影響。結果:1.NB4細胞經紫草素處理后,細胞活力受到抑制(P0.01),并且具有時間依賴性及濃度依賴性;2.流式細胞術檢測到NB4細胞經紫草素處理后,G1期細胞比例上升(P0.01),G2期和S期細胞比例下降(P0.01);細胞凋亡率明顯增加(P0.01);3.紫草素處理NB4后,經Hochest 33342著色,在熒光顯微鏡下發(fā)現(xiàn)細胞表現(xiàn)出核固縮,染色質聚集凝集,核破碎等現(xiàn)象;4.紫草素作用細胞1 d后,Western blot檢測到凋亡相關蛋白Cleaved PARP,Cleaved Caspase 3表達明顯增多(P0.01),p-JNK和p-p38MAPK蛋白水平明顯升高(P0.01),p-ERK和c-Myc蛋白表達水平明顯降低(P0.05);5.紫草素能夠顯著抑制移植瘤生長(P0.05);6.免疫組化結果顯示,紫草素處理組增殖蛋白c-Myc表達與對照組相比顯著降低(P0.05)。結論:紫草素可能通過調節(jié)MAPK通路和c-Myc表達水平來抑制NB4細胞增殖誘導其凋亡,并可抑制NB4細胞裸鼠移植瘤生長。
[Abstract]:Aim: to study the effect of porphyrin on proliferation and apoptosis of human acute promyelocytic leukemia (NB4) cells and its related mechanism.Method 1: 1.NB4 cell line was used as the research object. The proliferation of NB4 cells was measured by CCK-8 assay after treated with the gradient concentration of porphyrin for different time.NB4 cells were treated with 0.3 渭 mol/L porphyrin for 1 day. The cell cycle and apoptosis were detected by flow cytometry.The morphological changes of the nucleus of NB4 cells were detected by Hochest 33342 assay. 4. Western blot was used to detect the expression of c-Myccine paspase 3 and Cleaved PARPASE p38 MAPK and p-p38MAPK protein in NB4 cells.The model of transplanted tumor in nude mice with NB4 cells was constructed, and the effect of porphyrin on tumor development in nude mice was analyzed by depicting tumor growth curve.The effect of Shikonin on the expression of c-Myc in tumor tissue was detected by immunohistochemical technique.Results: 1. The cell viability of Nb4 cells was inhibited after treated with porphyrin, and the cell viability was time-dependent and concentration-dependent.Flow cytometry (FCM) showed that the proportion of NB4 cells in G _ 1 phase increased after treatment with Shikonin, and the proportion of P0.01G _ 2 and S phase cells decreased (P _ (0.01)), and the apoptosis rate increased significantly (P _ (0.01)).After NB4 was treated with porphyrin, the cells were stained with Hochest 33342 and the cells showed nuclear pyknosis, chromatin aggregation and nuclear fragmentation under fluorescence microscope.One day after treatment with porphyrin, the expression of apoptosis-related protein Cleaved, p0.01, p-JNK and p-p38MAPK was significantly increased by Western blot. The expression level of P0.01PERK and c-Myc decreased significantly.Shikonin could significantly inhibit the growth of transplanted tumor.Immunohistochemical results showed that the expression of proliferating protein (c-Myc) in the treated group was significantly lower than that in the control group (P 0.05).Conclusion: Shikonin may inhibit the proliferation and apoptosis of NB4 cells by regulating the MAPK pathway and the expression of c-Myc, and inhibit the growth of NB4 xenografts in nude mice.
【學位授予單位】：重慶醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R733.7

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