穿心莲内酯通过Notch信号通路抑制前列腺癌骨转移细胞体外生物学行为的研究

Andrographolide inhibits biological behavior of prostate cancer bone metastasis cells in vitro via Notch signaling pathway

目的:研究穿心莲内酯(ANDRO)对前列腺癌骨转移细胞株PC-3和C4-2B体外培养生物学行为的影响及其调控机制.方法:用ANDRO处理人源性的前列腺癌骨转移细胞株PC-3、C4-2B,MTS检测ANDRO对PC-3、C4-2B细胞的半抑制浓度IC 50,将实验分为IC 50药物浓度干预的ANDRO组和对照组,通过MTS检测不同时间下ANDRO对细胞增殖的影响;流式细胞术检测ANDRO对细胞周期的影响;Transwell检测细胞迁移和侵袭能力;纤维连接蛋白检测细胞黏附能力;Western blot检测Notch信号通路Notch-1、NICD、Hes-1,基质金属蛋白酶MMP2、MMP9及ICAM-1的蛋白表达情况.结果:ANDRO作用48 h的IC 50均约15μmol/L,且随浓度和时间的增加ANDRO对PC-3与C4-2B细胞增殖的抑制作用越显著(P<0.01).ANDRO组PC-3与C4-2B细胞周期分布G1期比例较对照组均呈升高,S期比例相应降低(P<0.05).ANDRO组PC-3与C4-2B细胞相较对照组的迁移、侵袭和黏附能力均明显下降(P<0.01).Western blot检测结果显示,ANDRO能显著抑制PC-3与C4-2B细胞Notch信号通路相关因子Noth-1、Hes-1和NICD的表达,同时也降低肿瘤转移相关蛋白MMP2、MMP9及ICAM-1的表达(P<0.01).结论:ANDRO可以抑制前列腺癌骨转移细胞株体外增殖、迁移、侵袭、黏附的能力,并影响其细胞周期分布,其作用机制可能与ANDRO抑制Notch信号通路蛋白及肿瘤转移相关蛋白表达有关.

Objective:To investigate the effects of andrographolide(ANDRO)on the biological behavior of prostate cancer bone metastasis cell lines PC-3 and C4-2B in vitro and its regulatory mechanism.Methods:Human prostate cancer bone metastasis cell line PC-3 and C4-2B were treated with ANDRO,and the half inhibitory concentration IC 50 of ANDRO on PC-3 and C4-2B cells was detected by MTS.Then,the experiment was divided into ANDRO group with IC 50 drug concentration and control group.MTS was used to detect the effects of ANDRO on cell proliferation at different times;flow cytometry to detect the effects of ANDRO on cell cycle;Transwell to detect cell migration and invasion ability;fibronectin to detect cell adhesion;Western blot to detect Notch signaling pathway Notch-1,NICD,Hes-1,the protein expression of matrix metalloproteinases MMP2,MMP9 and ICAM-1.Results:The IC 50 of ANDRO for 48 h was about 15μmol/L,and the inhibitory effect of ANDRO on the proliferation of PC-3 and C4-2B cells was more significant with the increase of concentration and time(P<0.01).The proportion of PC-3 and C4-2B cell cycle distribution in G1 phase in the ANDRO group was higher than that in the control group,and the proportion of S phase was correspondingly lower(P<0.05).The migration,invasion and adhesion abilities of PC-3 and C4-2B cells in the ANDRO group compared with the control group were significantly decreased(P<0.01).Western blot results showed that ANDRO can significantly inhibit the expression of Notch signaling pathway related factors Notch-1,Hes-1 and NICD in PC-3 and C4-2B cells,and also reduce the expression of tumor metastasis-related proteins MMP2,MMP9 and ICAM-1(P<0.01).Conclusion:ANDRO can inhibit the proliferation,migration,invasion and adhesion of prostate cancer bone metastasis cell lines in vitro,and affect their cell cycle distribution.The mechanism may be related to the inhibition of Notch signaling pathway protein and tumor metastasis-related protein expression by ANDRO.