摘要
目的研究葛根素对人肾癌786-O细胞凋亡的影响及其机制。方法分别以DMSO(空白对照组)、不同浓度葛根素(10、20、40 mg/L)和PI3K抑制剂LY29400215 mg/L干预对数生长期人肾癌786-O细胞48 h,采用CCK-8法检测细胞增殖抑制率,流式细胞术检测细胞凋亡水平,Western blot法检测磷脂酰肌醇3-激酶(PI3K)、磷酸化PI3K(Tyr317)[p-PI3K(Tyr317)]、蛋白激酶B(Akt)、磷酸化Akt(Ser473)[p-Akt(Ser473)]、糖原合成酶激酶-3β(GSK-3β)、磷酸化GSK-3β(Ser21)[p-GSK-3β(Ser21)]、半胱氨酸天冬氨酸蛋白酶-9(Caspase-9)、激活型Caspase-3(Cleaved Caspase-3)、B淋巴细胞瘤-2(Bcl-2)、Bcl-2相关X蛋白(Bax)表达。结果与空白对照组比较,葛根素10、20、40 mg/L组和LY29400215 mg/L组786-O细胞增殖抑制率升高[(17.04±2.92)%、(26.48±4.77)%、(42.79±6.81)%、(35.73±6.12)%比(3.47±0.53)%,P<0.01]、凋亡率升高[(13.72±2.38)%、(30.41±4.05)%、(55.39±6.72)%、(36.08±4.32)%比(2.64±0.51)%,P<0.01]。与空白对照组比较,葛根素20、40 mg/L组和LY29400215 mg/L组786-O细胞p-PI3K(Tyr317)、p-Akt(Ser473)、p-GSK-3β(Ser21)、Bcl-2表达下调[p-PI3K(Tyr317):(0.58±0.13)、(0.32±0.07)、(0.38±0.08)比(1.03±0.24);p-Akt(Ser473):(0.25±0.06)、(0.09±0.03)、(0.11±0.03)比(0.51±0.12);p-GSK-3β(Ser21):(0.12±0.03)、(0.07±0.02)、(0.09±0.03)比(0.58±0.13);Bcl-2:(0.31±0.08)、(0.18±0.05)、(0.21±0.07)比(0.48±0.12),P均<0.01],Caspase-9、Cleaved Caspase-3、Bax表达上调[Caspase-9:(0.37±0.08)、(0.66±0.13)、(0.58±0.11)比(0.17±0.04);Cleaved Caspase-3:(0.25±0.06)、(0.72±0.16)、(0.43±0.12)比(0.11±0.03);Bax:(0.36±0.08)、(0.62±0.12)、(0.20±0.05)比(0.08±0.02),P均<0.01],PI3K、Akt、GSK-3β磷酸化率降低[PI3K磷酸化率:(0.55±0.15)、(0.29±0.08)、(0.34±0.09)比(0.92±0.28);Akt磷酸化率:(0.24±0.07)、(0.09±0.03)、(0.11±0.04)比(0.53±0.14);GSK-3β磷酸化率:(0.14±0.04)、(0.08±0.03)、(0.10±0.04)比(0.62±0.15),P均<0.01],Bax/Bcl-2比值升高[(1.16±0.27)、(3.41±0.65)、
Objective To assess the effect and the underlying mechanism of Puerarin on the induction of human renal clear cell carcinoma cell apoptosis.Methods Human renal carcinoma 786-O cells were grown and subjected to the treatment with dimethyl sulfoxide(DMSO;a blank control),different doses of Puerarin(10,20,40 mg/L),and the PI3K specific inhibitor LY294002(15 mg/L,a positive control).Changes in tumor cell viability were measured using the cell counting kit-8(CCK-8)assay,while tumor cell apoptosis was detected using flow cytometry and expression of PI3K,p-PI3K(Tyr317),Akt,p-Akt(Ser473),GSK-3β,p-GSK-3β(Ser21),Caspase-9,Cleaved Caspase-3,Bcl-2,Bax was assessed using Western blot.Results Compared to the blank control,Puerarin at 10,20,or 40 mg/L dose and LY294002(15 mg/L)reduced tumor cell viability(17.04±2.92,26.48±4.77,42.79±6.81,and 35.73±6.12 vs.3.47±0.53%,respectively;All P<0.01),but induced tumor cell apoptosis(13.72±2.38,30.41±4.05,55.39±6.72 and 36.08±4.32 vs.2.64±0.51%,respectively;All P<0.01).Compared to the blank control,treatment with 20 or 40 mg/L Puerarin and 15 mg/L LY294002 downregulated expression of p-PI3K(Tyr317)(0.58±0.13,0.32±0.07,and 0.38±0.08 vs.1.03±0.24,respectively),p-Akt(Ser473)(0.25±0.06,0.09±0.03,and 0.11±0.03 vs.0.51±0.12,respectively),p-GSK-3β(Ser21)(0.12±0.03,0.07±0.02,and 0.09±0.03 vs.0.58±0.13,respectively),and Bcl-2(0.31±0.08,0.18±0.05,and 0.21±0.07 vs.0.48±0.12,respectively)in 786-O cells(all P<0.01).In contrast,these treatments upregulated expression of Caspase 9(0.37±0.08,0.66±0.13,and 0.58±0.11 vs.0.17±0.04,respectively),Cleaved Caspase 3(0.25±0.06,0.72±0.16,and 0.43±0.12 vs.0.11±0.03,respectively),and Bax(0.36±0.08,0.62±0.12,and 0.20±0.05 vs.0.08±0.02,respectively;all P<0.01).In addition,these treatments reduced the phosphorylation of PI3K(0.55±0.15,0.29±0.08,and 0.34±0.09 vs.0.92±0.28,respectively),Akt(0.24±0.07,0.09±0.03,and 0.11±0.04 vs.0.53±0.14,respectively),and GSK-3βproteins(0.14±0.04,0.08±0.03,and 0.10±0.04 vs.0.62±0.15,r
作者
竺东杰
邹杰
余史丹
陈潇
李红霞
ZHU Dong-jie;ZOU Jie;YU Shi-dan;CHEN Xiao;LI Hong-xia(Department of Pharmacy,The Number 906 Hospital of The PLA Joint Logistic Support Force,Ningbo 315100,Zhejiang,China;Department of Pharmacy,The Number 903 Hospital of The PLA Joint Logistic Support Force,Hangzhou,Zhejiang 310013,China;Department of Oncology,The Number 906 Hospital of The PLA Joint Logistic Support Force,Ningbo,Zhejiang 315100,China)
出处
《浙江中西医结合杂志》
2023年第10期893-898,共6页
Zhejiang Journal of Integrated Traditional Chinese and Western Medicine
基金
浙江省宁波市科技局计划项目(No.202002N4913)。
