秦皮乙素调控AKT/SKP2/MTH1通路诱导急性髓系白血病HL-60细胞凋亡的作用研究OA
Study on the apoptosis-inducing effect of esculetin on acute myeloid leukemia HL-60 cells via regulating the AKT/SKP2/MTH1 pathway
目的 探讨秦皮乙素(Esc)通过调控蛋白激酶B(AKT)/S期激酶相关蛋白2(SKP2)/MutT同源酶1(MTH1)通路诱导急性髓系白血病(AML)HL-60细胞凋亡的作用.方法 将HL-60细胞分为Control组(常规培养)、Esc低浓度组(L-Esc组,25 μmol/L Esc)、Esc中浓度组(M-Esc组,50 μmol/L Esc)、Esc高浓度组(H-Esc组,100 μmol/L Esc)、Esc高浓度+SC79(AKT激动剂)组(100 μmol/L Esc+5 μmol/L SC79).采用MTT法和克隆形成实验检测细胞增殖能力;采用CM-H2DCFDA荧光探针法检测细胞中活性氧(ROS)水平;采用流式细胞术检测细胞凋亡情况;采用Western blot法检测细胞中凋亡相关蛋白[B细胞淋巴瘤2(Bcl-2)、Bcl-2相关X蛋白(Bax)、裂解的胱天蛋白酶3(cleaved caspase-3)]和AKT/SKP2/MTH1通路相关蛋白[磷酸化AKT(p-AKT)、AKT、SKP2、MTH1]及AKT上下游效应蛋白磷脂酰肌醇3激酶(PI3K)和周期蛋白依赖性激酶抑制剂1(P21)、周期蛋白依赖性激酶抑制剂1B(P27)的表达.结果 与Control组相比,L-Esc组、M-Esc组、H-Esc组细胞的存活率、集落数以及AKT、PI3K蛋白的磷酸化水平和SKP2、MTH1、Bcl-2蛋白表达水平均显著降低(P＜0.05),而ROS水平、细胞凋亡率和Bax、cleaved caspase-3、P21、P27蛋白表达水平均显著升高(P＜0.05),且Esc的作用具有浓度依赖性(P＜0.05);与H-Esc组相比,Esc高浓度+SC79组细胞上述指标均被显著逆转(P＜0.05).结论 Esc可能通过抑制AKT/SKP2/MTH1通路,促进HL-60细胞中ROS的大量产生及细胞凋亡程序的启动,最终有效抑制HL-60细胞增殖.
OBJECTIVE To investigate the apoptosis-inducing effect of esculetin(Esc)on acute myeloid leukemia(AML)HL-60 cells by regulating the protein kinase B(AKT)/S-phase kinase-associated protein 2(SKP2)/MutT homolog 1(MTH1)pathway.METHODS AML HL-60 cells were randomly divided into control group(routine culture),Esc low-concentration group(L-Esc group,25 μmol/L Esc),Esc medium-concentration group(M-Esc group,50 μmol/L Esc),Esc high-concentration group(H-Esc group,100 μmol/L Esc),and high-concentration of Esc+SC79(AKT agonist)group(100 μmol/L Esc+5 μmol/L SC79).Cell proliferation in each group was detected by MTT assay and colony formation assay.The level of reactive oxygen species(ROS)in cells was measured by using the CM-H2 DCFDA fluorescent probe.Cell apoptosis was analyzed by flow cytometry.Western blot assay was performed to detect the expression levels of apoptosis-related proteins[B-cell lymphoma 2(Bcl-2),Bcl-2-associated X protein(Bax),cleaved caspase-3],AKT/SKP2/MTH1 pathway-related proteins(p-AKT,AKT,SKP2,MTH1),along with the upstream and downstream proteins of AKT phosphatidylinositol 3-kinase(PI3K),cyclin-dependent kinase inhibitor 1(P21)and cyclin-dependent kinase inhibitor 1B(P27).RESULTS Compared with control group,the cell viability,colony number,and the phosphorylation levels of AKT and PI3K proteins as well as protein expressions of SKP2,MTH1 and Bcl-2 were significantly decreased(P＜0.05),while ROS level,apoptosis rate,and the expression levels of Bax,cleaved caspase-3,P21 and P27 proteins were significantly increased(P＜0.05).Moreover,the effects of Esc exhibited concentration-dependence(P＜0.05).Compared with H-Esc group,above indexes of high-concentration of Esc+SC79 group were reversed significantly(P＜0.05).CONCLUSIONS Esc may promote massive ROS production and induce activation of apoptosis in HL-60 cells by inhibiting the AKT/SKP2/MTH1 pathway,thus inhibiting the proliferation of HL-60 cells.
宋卫华;褚福营;谢玮;陈金亮;赵枰;仇宏;陶健;陈相
东南大学附属南通市第一人民医院检验科,江苏 南通 226001东南大学附属南通市第一人民医院检验科,江苏 南通 226001东南大学附属南通市第一人民医院检验科,江苏 南通 226001东南大学附属南通市第一人民医院呼吸科,江苏 南通 226001东南大学附属南通市第一人民医院检验科,江苏 南通 226001东南大学附属南通市第一人民医院药剂科,江苏 南通 226001东南大学附属南通市第一人民医院血液科,江苏 南通 226001东南大学附属南通市第一人民医院检验科,江苏 南通 226001
医药卫生
秦皮乙素急性髓系白血病HL-60细胞蛋白激酶B细胞凋亡AKT/SKP2/MTH1通路
esculetinacute myeloid leukemiaHL-60 cellsprotein kinase Bcell apoptosisAKT/SKP2/MTH1 pathway
《中国药房》 2026 (1)
36-41,6
江苏省基础研究计划(自然科学基金)面上项目(No.BK20191207)南通市科技项目(No.MS22021001)
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