筋骨草总环烯醚萜对乳腺癌干细胞特性的干预作用及其机制分析

Intervention Effect of Ajuga decumbens Iridoids on Characteristics of Breast Cancer Stem Cells via ERK MAPK and PI3K/Akt Signaling Pathways

目的:研究筋骨草总环烯醚萜(iridoids in Ajuga decumbens,ADI)对乳腺癌干细胞特性的干预作用及分子机制。方法:采用无血清悬浮微球体形成实验,transwell迁移和侵袭实验检测乳腺癌干细胞自我更新能力、运动和侵袭能力。流式细胞法分析CD44+CD24-/low细胞亚群比例。蛋白免疫印迹法(Western blot)检测肿瘤干细胞干性相关蛋白,细胞外信号调节激酶(ERK)和磷脂酰肌醇3激酶(PI3K)/蛋白激酶B(PKB或Akt)信号通路相关蛋白的表达。结果:5~80 mg·L-1ADI能浓度依赖性降低乳腺癌MCF-7细胞微球体形成的数量和体积,减少CD44+CD24-/low细胞亚群比例(P<0.05,P<0.01)。10~40 mg·L-1ADI对乳腺癌MCF-7干细胞的迁移和侵袭能力有明显抑制作用(P<0.05,P<0.01)。进一步研究发现,ADI能明显降低p-ERK,p-Akt,干性标志物八聚体结合转录因子-3/4(Oct-3/4),SRY相关的HMG盒转录因子-2(Sox-2)和胚胎干细胞转录因子(Ecat4或Nanog)蛋白表达(P<0.05,P<0.01)。结论:筋骨草总环烯醚萜能有效地抑制乳腺癌干细胞的自我更新和转移潜能等"干性"特性,作用机制可能与其调控ERK1/2丝裂原活化蛋白激酶(MAPK)和PI3K/Akt信号通路,下调相关干性标志物表达有关。

Objective： To investigate the effect of iridoids in Ajuga decumbens（ ADI） on the characteristics of breast cancer stem cells（ BCSCs） and its relevant mechanism. Method： We conducted several assays,including mammosphere formation assay,transwell migration,transwell invasion assay and flow cytometry to evaluated the cancer stem cell load and metastasis. Western blot was employed to detect the expression of associated proteins of BCSC markers,extracellular signal-regulated kinase 1/2（ ERK1/2）,mitogen-activated protein kinase（ MAPK） and phosphatidylinositol kinase 3-kinase（ PI3K）/protein kinase B（ Akt） signaling pathways. Result： The 5-80 mg·L-1ADI could significantly inhibit the mammosphere formation,percentage of CD44＋CD24-/lowsubpopulation,migration and invasion in a dose-dependent manner（ P〈0. 05,P〈0. 01）.Further studies found that ADI could markedly suppress p-ERK,p-Akt,octamer transcription factor-3/4（ Oct-3/4）,SRY-related HMG box-2（ Sox-2） and ES cell-associated transcripts 4（ Ecat4 or Nanog） of BCSCs（ P〈0. 05,P〈0. 01）. Conclusion： These results indicated that ADI could inhibit the characteristics of BCSCs by suppressing the ERK1/2 MAPK and PI3K/Akt pathways.