METHODS: The effects of Zea on the proliferation, adhesion, migration and invasion of human GBM cell lines were detected by cell proliferation assay, cell adhesion assay and Transwell assay. The effect of Zea on angiogenesis was detected by rat aortic ring assay and human umbilical vein endothelial cells (HUVEC) in vitro tube formation assay. The effects of Zea on PARP, Caspase 3 and VEGFR2 phosphorylation as well as VEGFR2\'s downstream signaling pathway were detected by Western blot. The in vivo human GBM xenograft mouse model was employed to study the therapeutic efficacy of Zea.
RESULTS: Zea impaired the proliferation, adhesion, migration and invasion of U87 and U251 cells as well as HUVECs. Rat aortic ring experiments displayed Zea significantly inhibited angiogenesis during VEGF-induced microvascular germination. In vitro and in vivo vascular experiments verified that Zea inhibited VEGF-induced HUVEC proliferation and capillary-like tube formation. Additionally, Zea induced GBM cells apoptosis via increasing the expression of cleaved PARP and Caspase 3. In HUVECs and U251 GBM cells, Zea down-regulated VEGF-induced activation of the VEGFR2 kinase pathway. Meanwhile the expression of p-AKT, p-ERK, p-STAT3 and FAK were all attenuated in U251 cells. Moreover, the effects of Zea on GBM cells proliferation could be blocked by VEGFR2 kinase inhibitor SU5408. These results suggest that Zea may hinder GBM angiogenesis and tumor growth through down-regulating a cascade of oncogenic signaling pathways, both through the inhibition of angiogenesis and the anti-tumor mechanism of a direct cytotoxic effect. Besides, Zea inhibits GBM angiogenesis and tumor growth exemplified through a xenograft mouse model in vivo.
CONCLUSIONS: Zea impairs angiogenesis and tumor growth of GBM both in vitro and in vivo. It can be declared that Zea is a potential valuable anticancer candidate for the future treatment strategy of GBM.
方法:Zea对细胞增殖的影响,附着力,通过细胞增殖实验检测人GBM细胞系的迁移和侵袭,细胞粘附测定和Transwell测定。通过大鼠主动脉环测定法和人脐静脉内皮细胞(HUVEC)体外试管形成测定法检测Zea对血管生成的影响。Zea对PARP的影响,Westernblot检测Caspase3和VEGFR2的磷酸化以及VEGFR2的下游信号通路。采用体内人GBM异种移植小鼠模型来研究Zea的治疗功效。
结果:Zea损害了增殖,附着力,U87和U251细胞以及HUVECs的迁移和侵袭。大鼠主动脉环实验显示,在VEGF诱导的微血管萌发过程中,Zea可显着抑制血管生成。体外和体内血管实验证实Zea抑制VEGF诱导的HUVEC增殖和毛细血管样管形成。此外,Zea通过增加切割的PARP和Caspase3的表达来诱导GBM细胞凋亡。在HUVEC和U251GBM细胞中,Zea下调VEGF诱导的VEGFR2激酶途径的活化。同时p-AKT的表达,p-ERK,p-STAT3和FAK在U251细胞中均减弱。此外,VEGFR2激酶抑制剂SU5408可以阻断Zea对GBM细胞增殖的影响。这些结果表明,Zea可能通过下调致癌信号通路的级联来阻碍GBM血管生成和肿瘤生长。两者通过抑制血管生成和抗肿瘤机制的直接细胞毒作用。此外,Zea抑制GBM血管生成和肿瘤生长通过体内异种移植小鼠模型举例说明。
结论:Zea在体外和体内都会损害GBM的血管生成和肿瘤生长。可以声明Zea是GBM未来治疗策略的潜在有价值的抗癌候选物。