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目的探讨载蟾毒灵Pluronic-PEI纳米微囊通过抑制miR-497介导的IGF1-R-PI3K-Akt信号通路对大肠癌Lovo细胞体外及在裸鼠体内生长的影响及其作用机制。方法利用慢病毒miR-497转染大肠癌Lovo细胞,观察载蟾毒灵Pluronic-PEI纳米微囊对Lovo细胞体外增殖和侵袭转移能力的影响及miR-497介导的IGF1-R-PI3K-Akt信号通路相关基因和蛋白的调控作用。建立裸鼠原位移植瘤模型,8周后处死动物,观测对成瘤的影响。结果 (1)与对照组比较,载蟾毒灵的纳米微囊对过表达miR-497的Lovo细胞的抑制率最显著并呈时间-剂量依赖性;并使其凋亡率从(28.93±0.24)%提高至(75.20±0.29)%,明显高于相同浓度作用的Lovo细胞和Lovo NC细胞(P<0.01);划痕实验中过表达miR-497的Lovo细胞的迁移率从(0.083±0.019)(25 nmol/L)降低到(-0.242±0.035)(400 nmol/L),呈剂量依赖性;(2)Transwell结果提示载蟾毒灵纳米微囊在25 nmol/L就能够显著抑制过表达miR-497的Lovo细胞的侵袭作用;(3)免疫印迹分析提示载蟾毒灵的纳米微囊和LY 294002作用Lovo NC细胞和过表达miR-497的Lovo细胞,两种细胞的p-Akt Ser 473和p-Akt Thr 308蛋白表达明显下降,而pan-Akt变化不明显。(4)体内实验显示载蟾毒灵纳米微囊抑制瘤体转移优于蟾毒灵组。结论载蟾毒灵Pluronic-PEI纳米微囊可以通过抑制miR-497介导的IGF1-R-PI3K-Akt信号通路,在体外、体内都可发挥抗结肠癌侵袭和转移的作用。
OBJECTIVE: To investigate the effects of bufalin-Pluronic-PEI nanocapsules on the growth of colorectal cancer Lovo cells in vitro and in nude mice by inhibiting the IGF1-R-PI3K-Akt signaling pathway mediated by miR-497. Methods Lentiviral miR-497 cells were transfected with Lovo cells to observe the effects of bufalin-Pluronic-PEI nanocapsules on the proliferation, invasion and metastasis of Lovo cells and the effects of miR-497-mediated IGF1-R-PI3K-Akt Signaling pathway related genes and proteins regulation. Nude mice xenografts model was established. After 8 weeks, the animals were sacrificed and the effect on tumorigenesis was observed. Results (1) Compared with the control group, buprenorphine-loaded nanocapsules showed the most significant and time-dose-dependent inhibition on Lovo cells overexpressing miR-497 from (28.93 ± 0.24 ) Was increased to (75.20 ± 0.29)%, which was significantly higher than that of Lovo cells and Lovo NC cells (P <0.01). Migration of Lovo cells overexpressing miR-497 in the scratch assay ranged from (0.083 ± 0.019) (25 nmol / L) decreased to (-0.242 ± 0.035) (400 nmol / L) in a dose-dependent manner. (2) Transwell results suggested that Bufalin microcapsules could significantly inhibit (3) Immunoblotting analysis showed that bupropion-loaded nanocapsules and LY294002 could affect Lovo NC cells and Lovo cells overexpressing miR-497, and p-Akt Ser 473 and p-Akt Thr 308 protein expression decreased significantly, while the pan-Akt change was not obvious. (4) In vivo experiments showed that bufalin nanocapsules inhibited tumor metastasis better than bufalin group. Conclusion Bufalin-Pluronic-PEI nanocapsules can inhibit the invasion and metastasis of colon cancer in vitro and in vivo by inhibiting the IGF1-R-PI3K-Akt signaling pathway mediated by miR-497.