自噬对替莫唑胺治疗胶质瘤的影响

发布时间：2018-05-23 16:23

本文选题：自噬 + 胶质瘤　；参考：《吉林大学》2014年硕士论文

【摘要】：胶质瘤是最常见的脑原发性肿瘤[1]，占颅内原发肿瘤的35.26%～60.96%[2]。颅内胶质瘤会逐渐向四周扩散生长，每个位置都存在独特的病理特征以及不同的恶性程度。临床可采用手术、放射、化学药物、生物及其他方式开展治疗，但该病的肿瘤细胞具有很强的增殖能力，即便肿瘤有效的全切后，患者的预后效果仍然很难让人满意[3]。国内外对该疾病的研究成果较多，文献多以手术、放疗、药物、生物等联合的治疗方式为主，针对疾病的病理特性提出针对性的综合治疗方案，但多数研究仅能够保证患者的短期生存时间，，无法起到有效的根治作用，该病已逐渐成为世界医学范围内公认的治疗难题。因此，我们在以往研究工作的基础上，采用体外培养胶质瘤细胞等方法，探讨了自噬在胶质瘤细胞耐受替莫唑胺治疗过程中的作用. 实验目的:探讨自噬对胶质瘤细胞耐受替莫唑胺治疗中的作用拮抗自噬对替莫唑胺治疗胶质瘤细胞的影响及机制实验方法: 1、通过MTT法检测TMZ诱导胶质瘤细胞SHG44死亡情况，并探索3-MA对该种细胞死亡的作用。 2、通过流式细胞术进一步检测TMZ诱导胶质瘤细胞SHG44的凋亡。 3、通过吖啶橙染色及MDC染色证明TMZ诱导胶质瘤细胞SHG44死亡的过程中伴随强烈的自噬产生。 4、通过透射电镜直接观察TMZ诱导胶质瘤细胞产生的自噬情况。实验结果: MTT实验结果证实：与control组比较，TMZ90μmol/L组、TMZ90μmol/L+3-MA组细胞存活率明显降低（p0.05），TMZ90μmol/L+3-MA组与TMZ90μmol/L组比较，细胞存活率降低有统计学差异（p0.05）。流式细胞术实验结果进一步证实了上述结果。吖啶橙染色实验结果显示：TMZ90μmol/L组， TMZ90μmol/L+3-MA组与Control组及Control+3-MA组相比，胞浆内红色颗粒明显增多，比较证实TMZ90μmol/L+3-MA组比TMZ90μmol/L组，胞浆内红色颗粒明显减少。MDC染色实验结果显示：TMZ90μmol/L组， TMZ90μmol/L+3-MA组与Control组及Control+3-MA组相比，胞浆内绿色颗粒明显增多，并且TMZ90μmol/L+3-MA组与TMZ90μmol/L组比较，胞浆内绿色颗粒明显减少，荧光强度明显降低。实验结论: 1、TMZ可降低肿瘤细胞生存率，同时产生保护性自噬，导致胶质瘤细胞耐药。 2、抑制自噬能够提高TMZ对胶质瘤细胞的杀伤作用。
[Abstract]:Glioma is the most common primary brain tumor, accounting for 35.2696% of primary intracranial tumors. Intracranial gliomas gradually proliferate and grow around each location with unique pathological features and different degrees of malignancy. Surgery, radiation, chemotherapeutic, biological and other methods can be used in clinical treatment, but the tumor cells of the disease have strong proliferative ability, even after the effective total resection of the tumor, the prognosis of the patients is still very difficult to be satisfactory [3]. There are many research results on the disease at home and abroad. The literature is mainly combined with surgery, radiotherapy, medicine, biology and so on. According to the pathological characteristics of the disease, a targeted comprehensive treatment scheme is put forward. However, most studies can only guarantee the short life time of the patients, and can not play an effective role in radical cure. The disease has gradually become a medical problem in the world. Therefore, on the basis of previous studies, we investigated the role of autophagy in the treatment of glioma cell tolerance to temozolamide by means of in vitro culture of glioma cells. Objective: to investigate the role of autophagy in the treatment of glioma cell tolerance to temozolamide. Effect of antagonistic autophagy on the treatment of glioma cells with temozolamide and its mechanism Experimental methods: 1. TMZ induced SHG44 death in glioma cells was detected by MTT assay, and the effect of 3-MA on SHG44 cell death was explored. 2. The apoptosis of SHG44 cells induced by TMZ was further detected by flow cytometry. 3. Acridine orange staining and MDC staining showed that TMZ induced SHG44 death in glioma cells with strong autophagy. 4. The autophagy of glioma cells induced by TMZ was observed by transmission electron microscope. Experimental results: The results of MTT assay showed that compared with control group, the cell survival rate of TMZ90 渭 mol/L 3-MA group was significantly lower than that of TMZ90 渭 mol/L group. The cell survival rate of TMZ90 渭 mol/L 3-MA group was significantly lower than that of TMZ90 渭 mol/L group. The results of flow cytometry further confirmed the above results. The results of acridine orange staining showed that the number of red granules in the cytoplasm of the TMZ90 渭 mol/L 3-MA group was significantly higher than that of the Control group and Control 3-MA group. The results of acridine orange staining showed that the red granules in the cytoplasm of the TMZ90 渭 mol/L 3-MA group were significantly higher than that of the TMZ90 渭 mol/L group. The results of cytoplasmic red granules reduction. MDC staining showed that the green granules in the cytoplasm were significantly increased in the TMZ90 渭 mol/L 3-MA group compared with the Control group and the Control 3-MA group, and the green granules in the cytoplasm in the TMZ90 渭 mol/L 3-MA group were significantly lower than those in the TMZ90 渭 mol/L group, and in the TMZ90 渭 mol/L 3-MA group compared with the Control and Control 3-MA groups, the number of green granules in the cytoplasm was significantly lower than that in the TMZ90 渭 mol/L group. The fluorescence intensity decreased obviously. The experimental conclusions are as follows: TMZ can reduce the survival rate of tumor cells and produce protective autophagy, leading to drug resistance of glioma cells. 2. Inhibition of autophagy can enhance the killing effect of TMZ on glioma cells.
【学位授予单位】：吉林大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R739.41

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