首页|藏红花素通过PI3K/Akt/GSK-3β通路诱导人乳腺癌细胞凋亡的初步研究

藏红花素通过PI3K/Akt/GSK-3β通路诱导人乳腺癌细胞凋亡的初步研究

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目的:探讨藏红花素对人乳腺癌细胞凋亡的影响及相关调控机制.方法:以人正常乳腺MCF-10A细胞和人乳腺癌MCF-7细胞为受试细胞,分别以不同浓度藏红花素0(空白对照组)、200、400、800 mg/L和LY294002(PI3K特异性抑制剂)15 mg/L干预对数生长期MCF-10A细胞和MCF-7细胞48 h,采用四甲基偶氮唑蓝(MTT)法、克隆形成实验分别检测各组细胞增殖抑制率、细胞克隆能力,Annexin V-FITC染色法检测细胞凋亡水平,运用蛋白免疫印迹法(Western blot)检测磷脂酰肌醇3-激酶(phosphatidylinositol 3-kinase,PI3K)、磷酸化PI3K(p-PI3K)、蛋白激酶B(protein kinase B,Akt)、磷酸化Akt(p-Akt)、糖原合成酶激酶-3β(glycogen synthase kinase-3β,GSK-3β)、磷酸化GSK-3β(p-GSK-3β)、半胱氨酸天门冬氨酸蛋白酶9(cys-teine aspartic proteases-9,Caspase-9)、激活型半胱氨酸天门冬氨酸蛋白酶3(cleaved cysteine as-partate protease-3,Cleaved caspase-3)、B淋巴细胞瘤2(B-cell lymphoma-2,Bcl-2)、Bcl-2相关X蛋白(bcl-2 related X protein,Bax)蛋白表达.结果:与空白对照组比较,200、400、800 mg/L藏红花素组和LY294002组MCF-10A细胞增殖抑制率、克隆数目、凋亡率,差异均无统计学意义(P＞0.05);400、800 mg/L藏红花素组和LY294002组MCF-7细胞增殖抑制率升高、克隆数目降低、凋亡率升高(P＜0.01);p-PI3K、p-Akt、p-GSK-3β、Bcl-2表达下调且Caspase-9、Cleaved Caspase-3、Bax表达上调(P＜0.01),磷酸化率p-PI3K/PI3K、p-Akt/Akt、p-GSK-3β/GSK-3β降低(P＜0.01),Bax/Bcl-2表达比值升高(P＜0.01).与LY294002组比较,800 mg/L藏红花素组MCF-7细胞增殖抑制率升高、克隆数目降低、凋亡率升高(P＜0.05或P＜0.01);Caspase-9、Cleaved Caspase-3、Bax表达上调(P＜0.05或P＜0.01),Bax/Bcl-2 比值升高(P＜0.01),其他指标两组间比较差异无统计学意义(P＞0.05).结论:藏红花素具有诱导人乳腺癌细胞凋亡的作用,其机制可能与抑制PI3K/Akt/GSK-3β信号通路有关.
Preliminary Study of Saffronin-induced Apoptosis of Human Breast Cancer Cells via PI3K/Akt/GSK-3β Pathway
Objective:To explore the effect of saffronin on the apoptosis of human breast cancer cells and related regulatory mechanisms.Methods:Human normal breast MCF-10A cells and human breast cancer MCF-7 cells were used as test cells,and different concentrations of saffronin 0(blank control group),200,400,800 mg/L and LY294002(PI3K-specific inhibitor)15 mg/L were used to intervene in the logarithmic growth phase MCF-10A cells and MCF-7 cells for 48 h.Tetramethyl azole blue(MTT)assay and clonal formation assay were adopted to detect the inhibition rate of cell proliferation and clonogenic ability of cells in each group respectively,Annexin V-FITC staining to detect the level of apoptosis,Western blot was applied to detect the expressions of PI3K,p-PI3K,Akt,p-Akt,GSK-3β,p-GSK-3β,Caspase-9,Cleaved caspase-3,Bcl-2 and Bax.Results:Compared with the blank control group,the difference had no statistical meaning in the proliferation inhibition rate,clone number and apoptosis rate of MCF-10A cells in the 200,400 and 800 mg/L saffronin group and the LY294002 group(P＞0.05);the proliferation inhibition rate of MCF-7 cells in 400,800 mg/L saffronin group and LY294002 group increased,the number of clones decreased,and the apoptosis rate increased(P＜0.01);the expression of p-PI3K,p-Akt,p-GSK-3β,Bcl-2 was down-regulated,and the expression of Caspase-9,Cleaved Caspase-3,and Bax was up-regulated(P＜0.01).The phosphorylation rates of p-PI3K/PI3K,p-Akt/Akt,p-GSK-3β/GSK-3β were decreased(P＜0.01),and the Bax/Bcl-2 expression ratio was increased(P＜0.01).Compared with the LY294002 group,MCF-7 cells in the 800 mg/L saffronin group showed higher inhibition of cell proliferation,lower number of clones,and higher apoptosis rate(P＜0.05 or P＜0.01);Caspase-9,Cleaved Caspase-3 and Bax expressions were up-regulated(P＜0.05 or P＜0.01),and the Bax/Bcl-2 ratio was elevated(P＜0.01),while the differences of the other indexes were not statistically significant between the two groups(P＞0.05).Conclusion:saffronin could induce the apoptosis of human breast cancer cells,and its mechanism might be related to inhibiting p-PI3K/Akt/GSK-3β signaling pathway.

breast cancersaffroninproliferationcellular apoptosisp-PI3K/Akt/GSK-3β signaling pathway

赵宁、夏利敏、宋涛

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邯郸市中心医院,河北 邯郸 056001

乳腺癌 藏红花素 增殖 细胞凋亡 PI3K/Akt/GSK-3β信号通路

河北省医学科学研究课题计划

20191643

2024

10.12174/j.issn.2096-9600.2024.04.04

西部中医药
甘肃中医药研究院

西部中医药

CSTPCD
影响因子：0.98
ISSN：1004-6852
年,卷(期)：2024.37(4)
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