小檗碱下调蛋白激酶C-α逆转K562/A02细胞耐药作用机制研究  

Mechanism of berberine reverses drug resistance of K562/A02 cells by down-regulating protein kinase C-alpha

作　　者：刘牧 薄海美 杨夏茵 陈建立 Liu Mu;Bo Haimei;Yang Xiayin;Chen Jianli(Pediatrics Department,the Affiliated Hospital of North China University of Science and Technology,Tangshan 063000,China;Department of Cardiovascular,Clinical Medical College,North China University of Science and Technology,Tangshan 063000,China)

机构地区：[1]华北理工大学附属医院儿科,唐山063000 [2]华北理工大学临床医学院心血管科,唐山063000

出　　处：《国际中医中药杂志》2022年第5期535-540,共6页International Journal of Traditional Chinese Medicine

基　　金：河北省政府资助专科能力建设和专科带头人培养项目(361036)。

摘　　要：目的研究小檗碱对白血病耐药细胞株K562/A02阿霉素耐药性及蛋白激酶C-α(PRKCA)的影响,探讨其作用机制。方法体外培养人慢性粒细胞白血病K562细胞、阿霉素耐药株K562/A02,使用2.5~50.0μmol/L的阿霉素处理,检测K562、K562/A02对阿霉素的耐药性,计算药物半抑制浓度(IC_(50));采用终浓度5μmol/L的阿霉素溶液处理K562/A02细胞,并将K562/A02细胞分为对照组、抑制剂组(50μmol/L PRKCA抑制剂)及小檗碱低、中、高剂量组,采用细胞计数(CCK-8)法检测各组细胞增殖抑制率,通过流式细胞仪检测各组细胞凋亡情况,实时荧光定量PCR法检测PRKCA、多药耐药相关基因(MDR1)水平,Western blot法检测PRKCA、多药耐药相关蛋白(MRP)、P-糖蛋白(P-gp)表达情况。结果与K562比较,K562/A02对阿霉素的IC_(50)升高(P<0.05);与对照组比较,抑制剂组及小檗碱低、中、高剂量组细胞增殖抑制率、凋亡率升高(P<0.05),PRKCA mRNA[(0.45±0.08)、(0.92±0.10)、(0.57±0.05)、(0.35±0.04)比(1.00±0.12)]、MDR1 mRNA[(0.73±0.08)、(0.87±0.09)、(0.65±0.07)、(0.41±0.05)比(1.00±0.11)]及PRKCA蛋白[(0.59±0.09)、(0.78±0.12)、(0.61±0.11)、(0.42±0.07)比(0.96±0.14)]、MRP蛋白[(0.62±0.08)、(0.79±0.13)、(0.62±0.10)、(0.41±0.06)比(0.98±0.14)]、P-gp[(0.55±0.08)、(0.75±0.12)、(0.59±0.09)、(0.35±0.06)比(0.92±0.15)]表达降低(P<0.05),且小檗碱呈药物剂量依赖性(P<0.05);过表达PRKCA可显著抑制小檗碱逆转K562/A02细胞耐药的作用。结论小檗碱可能通过下调PRKCA逆转人白血病耐药株K562/A02对阿霉素的耐药性。Objective To observe the effect of berberine on leukemia drug-resistant cell strain K562/A02 to Adriamycin resistance and protein kinase C-alpha(PRKCA)and explore its possible mechanism.Methods The leukemia K562 cells of human chronic myeloid and Adriamycin resistant strain K562/A02 were cultured in vitro with 2.5-50.0μmol/L doxorubicin to treat thoese cells and drug resistance of K562 and K562/A02 to Adriamycin was detected,the 50%inhibitory concentration(IC_(50))of the drug was calculatedthe resistance of K562 and K562/A02 to doxorubicin was detectd,and,K562/A02 cells were treated with doxorubicin solution at a final concentration of 5μmol/L,and K562/A02 cells were divided into control group,inhibitor group(50μmol/L PRKCA inhibitor),low dose berberine group,medium dose berberine group and high dose berberine group.Cell counting(CCK-8)method was used to detect the inhibition rate of cell proliferation,the apoptosis was detected by flow cytometry,real-time fluorescent quantitative PCR assay detects PRKCA,MRP,multidrug resistance related genes(MDR1)levels,and the protein expressions of protein kinase C-α(PRKCA),multidrug resistance related protein(MRP),P-glycoprotein(P-gp)were detected by Western blotting.Results The IC_(50) concentration of K562/A02 to Adriamycin was significantly higher than K562.Compared with the control group,the inhibition rate of cell proliferation and the apoptosis rate in the inhibitor group,low-dose berberine group,medium-dose berberine group,and high-dose berberine group were significantly increased(P<0.05),the expression of PRKCA mRNA(0.45±0.08,0.92±0.10,0.57±0.05,0.35±0.04 vs.1.00±0.12),MDR1 gene(0.73±0.08,0.87±0.09,0.65±0.07,0.41±0.05 vs.1.00±0.11)and PRKCA(0.59±0.09,0.78±0.12,0.61±0.11,0.42±0.07 vs.0.96±0.14),MRP(0.62±0.08,0.79±0.13,0.62±0.10,0.41±0.06 vs.0.98±0.14),P-gp(0.55±0.08,0.75±0.12,0.59±0.09,0.35±0.06 vs.0.92±0.15)were significantly reduced(P<0.05),and berberine was dose-dependent(P<0.05);Overexpression of PRKCA can inhibit the effect of berberin

关 键 词：小檗碱 蛋白激酶C-Α K562细胞 耐药 

分 类 号：R285[医药卫生—中药学]