扁塑藤素调控Notch信号通路诱导人肺癌条件重编程细胞死亡的作用机制研究

目的：应用条件性重编程细胞（conditionally reprogrammed cells，CRCs）技术建立人肺癌细胞体外长期培养体系，研究扁塑藤素对人肺癌CRCs增殖和迁移能力的影响，并探讨相关作用机制。方法：免疫组化法检测Notch 1、HES1和Cyclin D3在肿瘤组织和癌旁组织中的表达水平；使用CRCs技术分离培养非小细胞肺癌原代细胞；使用2，4，8，16 μmol·L^-1的扁塑藤素处理人肺癌CRCs，MTS法检测细胞活力，Annexin V/PI流式细胞术检测细胞凋亡，Transwell法检测细胞迁移能力，Western Blot检测细胞中Notch信号通路相关蛋白Notch 1、HES1和Cyclin D3的表达水平。结果：在非小细胞肺癌患者肿瘤组织中Notch 1、HES1和Cyclin D3的表达水平高于癌旁组织；扁塑藤素能够诱导人肺癌CRCs死亡，并在一定范围内呈现时间和浓度依赖性（P<0.05）；随着扁塑藤素作用浓度的增高，人肺癌CRCs凋亡率明显增高（P<0.05），细胞迁移能力下降（P<0.05）；扁塑藤素能够下调人肺癌CRCs中Notch 1、HES1和Cyclin D3的蛋白表达水平。结论：扁塑藤素可能通过调控Notch信号通路相关蛋白的表达水平，抑制人肺癌CRCs的增殖和迁移，并诱导其凋亡，为肺癌的治疗提供新的实验数据和理论依据。

The mechanisms of pristimerin inducing human lung cancer conditionally reprogrammed cell death via Notch signaling

OBJECTIVE To establish the long-term culture system of human lung cancer cells in vitro by conditional reprogrammed cells (CRCs) technology, and investigate the effect of pristimerin on the proliferation and migration of human lung cancer CRCs as well as explore the related mechanism.METHODS The expression levels of Notch 1, HES1 and Cyclin D3 in tumor tissues and adjacent tissues were detected by immunohistochemistry. The primary cells of non-small cell lung cancer (NSCLC) were isolated and cultured by using CRCs technique. The human lung cancer CRCs were treated with 2, 4, 8, 16 μmol·L^-1 pristimerin. The cell viability was detected by MTS method. The cell apoptosis was detected by Annexin V/PI flow cytometry. The cell migration ability was detected by Transwell method. The expression levels of Notch 1, HES1 and Cyclin D3 in the cells were detected by Western Blot.RESULTS The expression levels of Notch 1, HES1 and Cyclin D3 in NSCLC tissues were higher than those in adjacent tissues. Pristimerin inhibited the proliferation of human lung cancer CRCs in a time-and does-dependent manner (P<0.05).With the increase of pristimerin concentration, the apoptosis rate of human lung cancer CRCs was significantly higher (P<0.05), and the cell migration ability decreased (P<0.05). Furthermore, the expression of Notch 1, HES1 and Cyclin D3 was remarkably down-regulated by pristimerin (P<0.05).CONCLUSION Pristimerin may inhibit the proliferation and migration of human lung cancer CRCs and induce their apoptosis by regulating the expression level of proteins related to Notch signaling pathway, which could provide new experimental data and theoretical basis for the treatment of lung cancer.