目的 探究丹参酮ⅡA(Tan ⅡA)对乳腺癌细胞迁移、侵袭的机制.方法 使用不同浓度的Tan ⅡA对乳腺癌细胞系MCF-7进行治疗,MTT法检测细胞活力,流式细胞偶数检测细胞凋亡,Transwell法检测细胞迁移、侵袭能力.使用Hedgehog激活剂SAG激活Hedgehog/Gli1通路,使用qRT-PCR及免疫印迹检测SMO、Gli1 mRNA及蛋白表达水平.通过免疫印迹检测自噬相关蛋白LC3-Ⅰ、LC3-Ⅱ及Beclin-1表达水平.结果 50 μmol·L-1 Tan ⅡA可以抑制MCF-7细胞活力,MDA-MB-231细胞活力由(98.12±3.85)%下降至(69.32±3.68)%,MCF-7细胞活力由(99.32±3.65)%下降至(49.65±3.74)%.添加SAG后MCF-7细胞活力上升至(71.20±5.00)%(P<0.05).SAG抑制MCF-7细胞凋亡,细胞凋亡率从(20.50±2.00)%下降至(12.00±1.00)%.SAG使得MCF-7细胞迁移数量从(52.00±5.00)上升至(76.00±5.00)(P<0.05),侵袭数量从(41.00±3.00)上升至(75.00±5.00)(P<0.05).与Control组相比,Tan ⅡA组Beclin-1蛋白水平由(0.54±0.05)上升至(1.20±0.10),LC3-Ⅱ/LC3-Ⅰ水平由(0.60±0.06)上升至(1.80±0.12)(P<0.05),而添加SAG后Beclin-1蛋白水平下降至(0.70±0.07)(P<0.05),LC3-Ⅱ/LC3-Ⅰ水平下降至(0.75±0.04)(P<0.05).结论 Tan ⅡA降低乳腺癌细胞活力、迁移及侵袭能力,抑制Hedgehog/Gli1信号通路活性,且促进乳腺癌细胞自噬.
Mechanisms of tanshinone Ⅱ A promoting migration and invasion of breast cancer cells by regulating Hedgehog/Gli1 signaling pathway
Objective To observe the mechanisms of tanshinone ⅡA(Tan ⅡA)for promoting migration and invasion of breast cancer cells.Methods After the breast cancer cell line MCF-7 was treated with different concentrations of Tan ⅡA,the cell viability was detected by MTT assay,apoptosis determined by flow cytometry and the migration and invasion of the cells determined by Transwell assay.The Hedgehog activator SAG was used to activate the Hedgehog/Gli1 pathway.The expression levels of mRNA and protein of SMO and Gli1 were detected by qRT-PCR and Western blot.The expression levels of autophagy-related proteins LC3-Ⅰ,LC3-Ⅱ and Beclin-1 were determined by Western blot.Results 50 μmol·L-1 Tan ⅡA significantly inhibited the activity of MCF-7 cells,the activity of MDA-MB-231 cells decreased from(98.12±3.85)%to(69.32±3.68)%,and the activity of MCF-7 cells decreased from(99.32±3.65)%to(49.65±3.74)%.The cell viability of MCF-7 was significantly increased to(71.20±5.00)%after SAG addition(P<0.05).SAG significantly inhibited apoptosis of MCF-7 cells,and the apoptosis rate decreased from(20.50±2.00)%to(12.00±1.00)%.SAG increased the number of MCF-7 cell migration from(52.00±5.00)to(76.00±5.00)(P<0.05),and the number of invasion from(41.00±3.00)to(75.00±5.00)(P<0.05).Compared with control group,Beclin-1 protein level in Tan ⅡA group was increased from(0.54±0.05)to(1.20±0.10),LC3-Ⅱ/LC3-Ⅰlevel was increased from(0.60±0.06)to(1.80±0.12)(P<0.05).After addition of SAG,Beclin-1 protein level was significantly decreased to(0.70±0.07)(P<0.05),LC3-Ⅱ/LC3-Ⅰlevel was significantly decreased to(0.75±0.04)(P<0.05).Conclusion Tan ⅡA can reduce the viability,migration and invasion of the breast cancer cells,inhibit the activity of Hedgehog/Gli1 signaling pathway,and promote the autophagy of the cells.
Breast cancerTanshinone ⅡAAutophagyHedgehog/Gli1 signaling pathway
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