Rab27a对人脑胶质瘤细胞生物活性的影响及其在老年脑胶质瘤中的临床应用

发布时间：2018-04-08 09:38

  本文选题：Rab27a　切入点：脑胶质瘤　出处：《安徽医科大学》2014年博士论文

【摘要】：目的:探讨Rab27a的表达对人脑胶质瘤U251细胞生长、增殖、凋亡及侵袭迁移等生物学行为的影响,为将来人脑胶质瘤的生物治疗提供实验基础。方法:构建pc DNA3.1-Rab27a真核重组表达载体,利用脂质体将pc DNA3.1-Rab27a和Rab27a-sh RNA转染U251,Western blot检测Rab27a蛋白的表达;MTT法检测细胞活力;流式细胞术检测Rab27a异位表达并分析其对U251细胞增殖、凋亡的影响;Transwell侵袭小室等方法检测U251细胞侵袭性;通过检测Rab27a的异位表达,分析对组织蛋白酶D分泌的影响;应用荧光素酶报告基因系统验证Rab27a是mi R-124的靶基因。结果:Western blot结果显示:与对照组相比,pc DNA3.1-Rab27a转染组Rab27a的表达显著增加(p0.01),而Rab27a-sh RNA转染组Rab27a的表达显著降低(p0.01)。MTT、流式细胞术和Transwell侵袭小室检测结果显示:与对照组相比,pc DNA3.1-Rab27a转染组细胞活力(p0.01)、增殖指数(p0.05)、迁移能力显著增加(p0.01),Rab27a-sh RNA转染组细胞活力(p0.01)、增殖指数(p0.05)、迁移能力显著降低(p0.01)。流式细胞术分析凋亡过程,结果显示:与对照组相比,pc DNA3.1-Rab27a转染组细胞凋亡降低(p0.05),而Rab27a-sh RNA转染组细胞凋亡增加(p0.05)。组织蛋白酶D检测结果显示:与正常对照组相比,pc DNA3.1-Rab27a转染组中组织蛋白酶D的分泌量高于正常对照组(p0.05)。结论:Rab27a基因具有促进脑胶质瘤细胞U251细胞的活力、增殖、迁移,并抑制其凋亡作用,其作用机制可能通过促进组织蛋白酶D的分泌,和受mi R-124的调控来发挥作用。抑制Rab27a表达的治疗策略有望使胶质瘤患者受益。目的:根据Rab27a在脑胶质瘤组织中的表达情况在老年脑胶质瘤患者中进行个体化化疗,观察其治疗效果、不良反应和生存期。方法:入组时间:2011年7月至2013年5月。年龄≥65岁;经手术切除或活检后的组织病理学确诊的脑胶质瘤患者,测定组织Rab27a表达水平。Rab27a表达者设为治疗组,应用单药替莫唑胺化疗,治疗剂量:150~200mg/m2,口服,每日一次,d1~5;每28天为一周期,至少治疗2周期。Rab27a不表达者设为对照组,仅对症处理,未进行化疗。按照WHO标准评价疗效、CTCAE 4.0标准评价药物不良反应,其它观察指标为OS和PFS。结果:按照入组标准,治疗组共有32例患者进入临床研究;中位年龄70岁,中位KPS:70分。可评价病例29例,其中9例PR(31%)、12例SD(41%)、8例PD(28%)。自诊断开始的中位OS为7.4个月,中位PFS为6.1个月。所有入组患者共进行了160周期单药替莫唑胺化疗,中位为4周期。不良反应以Ⅰ~Ⅱ度为主,包括:恶心、呕吐、疲劳、便秘、骨髓毒性等。Ⅲ~Ⅳ度中性粒细胞减少的出现率为9.38%(3/32)患者,Ⅳ度血小板下降的出现率为6.25%(2/32)。对照组共入组21例,中位年龄68岁,中位KPS:70分,自诊断开始的中位OS为5.7个月,中位PFS为4.0个月。结论:Rab27a指导下的替莫唑胺化疗在脑胶质瘤有广阔的应用前景,替莫唑胺优点在于:疗效显著;大多数副作用可耐受;口服给药方便;改善其生活质量。Rab27a对患者生存期的预测价值有待进一步研究。
[Abstract]:Objective: To investigate the growth and proliferation of U251 human glioma cells Rab27a expression, apoptosis and invasion influence the biological behavior of migration, to provide the experimental basis for the biological treatment of glioma in the future. Methods: to construct the PC DNA3.1-Rab27a eukaryotic expression vector by liposome PC DNA3.1-Rab27a and Rab27a-sh RNA U251 transfection, the expression of Rab27a protein detection of Western blot; the cell viability was measured by MTT; flow cytometry analysis of ectopic expression of Rab27a and the proliferation of U251 cells, apoptosis; Transwell assay detected U251 cell invasion; by ectopic expression of Rab27a analysis of effect on secretion of cathepsin D using luciferase reporter system verification; Rab27a is the target gene mi R-124. Results: Western blot showed that: compared with the control group, the expression of PC DNA3.1-Rab27a transfection group Rab27a increased significantly Plus (P0.01), and the expression of Rab27a-sh RNA transfection group significantly decreased Rab27a (P0.01).MTT, flow cytometry and Transwell assay. The results showed that: compared with the control group, PC transfection group DNA3.1-Rab27a cell viability (P0.01), proliferation index (P0.05), the migration ability was significantly increased (P0.01), Rab27a-sh RNA transfection activity (P0.01), proliferation index (P0.05), the migration ability decreased significantly (P0.01). Analysis of the process of apoptosis, flow cytometry results showed that: compared with the control group, the apoptosis of PC cells in DNA3.1-Rab27a transfection group decreased (P0.05), and the apoptosis of Rab27a-sh cells in RNA transfection group increased (P0.05). The detection of cathepsin D the results showed that: compared with normal control group, PC secretion in DNA3.1-Rab27a transfection group of cathepsin D is higher than the normal control group (P0.05). Conclusion: Rab27a gene can promote glioma cell viability of U251 cells, proliferation, migration, and suppression The role of apoptosis, its mechanism may promote the secretion of cathepsin D, MI and R-124 of the control to play the role of therapeutic strategies to inhibit the expression of Rab27a is expected to make the glioma patients. Objective: according to the expression of Rab27a in glioma tissues of individual chemotherapy in elderly patients with cerebral glioma, observation the therapeutic effect, adverse reaction and survival. Methods: in the group of time: from July 2011 to May 2013. 65 years of age or older; the patients with glioma confirmed histological pathology after surgical excision or biopsy, determination of tissue Rab27a expression levels of.Rab27a expression were divided into treatment group, application of single drug temozolomide that dose: 150~200mg/m2, oral, once daily, d1~5; every 28 days for a period of at least 2 cycles of treatment, the expression of.Rab27a were set as control group, only symptomatic treatment, no chemotherapy. According to WHO evaluation criteria, C TCAE 4.0鏍囧噯璇勪环鑽�鐗╀笉鑹�鍙嶅簲,鍏跺畠瑙傚療鎸囨爣涓篛S鍜孭FS.缁撴灉:鎸夌収鍏ョ粍鏍囧噯,娌荤枟缁勫叡鏈，

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