成纤维细胞生长因子受体抑制剂NVP-BGJ398对胶质瘤生长的影响

发布时间：2018-02-11 14:16

本文关键词： 胶质瘤成纤维细胞生长因子受体 NVP-BGJ398 血管生成拟态　出处：《南方医科大学》2017年硕士论文　论文类型：学位论文

【摘要】：研究背景:胶质细胞瘤是常见的中枢神经系统恶性肿瘤,患者的半数生存期不足两年。成纤维细胞生长因子受体(FGFR)是一类酪氨酸激酶膜受体。FGFR信号通路涉及肿瘤细胞的各种活动,如肿瘤的增殖,血管生成,侵袭及肿瘤干细胞多功能性的维持等。文献显示FGFR在胶质瘤中表达上升,提示FGFR是治疗胶质瘤的潜在靶点。NVP-BGJ398是目前可获得的强效选择性抑制FGFR的药剂。血管生成拟态是一种与血管相似的官腔结构,但它的管壁不是由血管内皮细胞构成的而是由肿瘤细胞组成的,这个官腔可以给胶质瘤提供额外的血供,促进恶性肿瘤的发展。目的:检测FGFR抑制剂NVP-BGJ398对抑制胶质瘤细胞增殖、迁移和侵袭的影响;检验NVP-BGJ398对胶质瘤细胞形成血管生成拟态的影响;探索NVP-BGJ398影响胶质瘤细胞的机制。方法:1、进行WB实验和免疫组化染色分析蛋白表达;2、运用增殖实验、平板克隆实验和皮下移植瘤模型检测胶质瘤细胞生长;3、进行侵袭实验和迁移实验分析肿瘤细胞的侵袭和迁移能力;4、运用小管生成实验和CD34/PAS双染检测肿瘤细胞血管拟态的形成;5、统计分析。结果:1、Western blotting显示NVP-BGJ398处理可以有效的抑制胶质瘤细胞中的FGFR磷酸化激活成为pFGFR和降低胶质瘤细胞中Vimentin、MMP2和MMP14 的表达(P0.05);2、CCK-8实验结果表明经过NVP-BGJ398的处理两株细胞的增殖速度明显的下降(P0.05);平板克隆实验对照组中克隆形成数量远大于经过NVP-BGJ398 实验组(P0.05);3、划痕实验中对照组U251MG细胞迁移速度明显快于NVP-BGJ398实验组(P0.05);Transwell迁移和侵袭实验结果表明穿膜细胞数在NVP-BGJ398处理后都明显的减少(P0.05);4、小管形成实验中NVP-BGJ398实验组中胶质瘤细胞小管周长和交点数较对照组明显的减少;5、U87MG皮下成瘤实验中NVP-BGJ398实验组(0.19±0.06g)肿瘤重量明显轻于对照组(0.45±0.11g)(P0.05);6、CD34/PAS双染VM计数结果显示皮下瘤模型NVP-BGJ398实验组中VM的数量(13.85±3.96)较对照组(26.40±5.06)明显的减少(P0.05);7、免疫组织染色显示NVP-BGJ398实验组中Vimentin、MMP2和MMP14平均光密度(2.50±0.36,1.89±0.95,1.26±0.15)明显浅于对照组(4.13±1.15,4.90±0.52,3.77±0.39)(P0.05)。结论:在体内外实验中发现FGFR抑制剂NVP-BGJ398具有良好的抗胶质瘤作用。NVP-BGJ398除了抑制肿瘤细胞的增殖、迁移以及侵袭,还具有抑制血管生成拟态生成的作用。其机制与能够抑制FGFR的激活下调vimentin、MMP14和MMP2的表达有关。提示成纤维细胞生长因子受体是治疗胶质瘤的一个有效靶点。
[Abstract]:Background: glioma is a common malignant tumor of the central nervous system. Fibroblast growth factor receptor (FGFR) is a type of tyrosine kinase receptor. FGFR signaling pathway involves various activities of tumor cells, such as tumor proliferation, angiogenesis, Invasion and multifunctional maintenance of tumor stem cells. The literature shows that the expression of FGFR is increased in gliomas. It is suggested that FGFR is a potential target for the treatment of glioma. NVP-BGJ398 is a potent and selective inhibitor of FGFR. However, its wall is composed not of vascular endothelial cells but of tumor cells, which can provide additional blood supply to glioma and promote the development of malignant tumor. Objective: to detect the inhibitory effect of FGFR inhibitor NVP-BGJ398 on the proliferation of glioma cells. The effects of NVP-BGJ398 on the angiogenesis of glioma cells were examined. The mechanism of NVP-BGJ398 affecting glioma cells was explored. Plate clone assay and subcutaneous tumor transplantation model were used to detect the growth of glioma cells. Invasion and migration experiments were carried out to analyze the invasion and migration ability of tumor cells. Microtubule formation test and CD34/PAS double staining were used to detect tumor blood vessels. Results NVP-BGJ398 treatment could effectively inhibit the activation of FGFR phosphorylation into pFGFR in glioma cells and decrease the expression of Vimentinine MMP2 and MMP14 in glioma cells. The results showed that NVP-BGJ398 treatment could effectively inhibit the activation of FGFR phosphorylation into pFGFR and decrease the expression of Vimentinine MMP2 and MMP14 in glioma cells. The proliferation rate of the two cells decreased significantly (P 0.05), the number of clone formation in the control group was much larger than that in the NVP-BGJ398 experimental group, and the migration rate of U251MG cells in the scratch test group was significantly faster than that in the NVP-BGJ398 experimental group. The results showed that the number of perforating cells decreased significantly after NVP-BGJ398 treatment. In the tubule formation test, the circumference and intersection number of glioma cells in the NVP-BGJ398 experimental group were significantly reduced compared with those in the control group. In the subcutaneous tumorigenesis experiment of U87MG, the number of the glioma cells in the NVP-BGJ398 experimental group was significantly reduced. Tumor weight (0.19 卤0.06g) was significantly lighter than that in control group (0.45 卤0.11g). The results of VM counting of CD34 / pas double staining showed that the number of VM in the subcutaneous tumor model NVP-BGJ398 group was 13.85 卤3.96, significantly lower than that in the control group (26.40 卤5.06g). The immunohistochemical staining showed that Vimentinine MMP2 and MMP14 average optical density in NVP-BGJ398 experimental group were 2.50 卤0.36 卤1.89 卤51.26 卤0.15). Conclusion: in vivo and in vitro, FGFR inhibitor NVP-BGJ398 has a good anti-glioma effect. NVP-BGJ398 can not only inhibit the proliferation of tumor cells, but also inhibit the proliferation of tumor cells. Migration and invasion also inhibit angiogenesis mimicopoiesis. The mechanism is related to the down-regulation of FGFR activation and down-regulation of the expression of MMP14 and MMP2. It suggests that fibroblast growth factor receptor is an effective target for the treatment of glioma.
【学位授予单位】：南方医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R739.41

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