白藜芦醇对血虚证小鼠模型的改善作用及诱导红系分化机制研究  

Effect of resveratrol on mouse model with blood deficiency syndrom and mechanism in inducing erythroid differentiation of K562 cells

作　　者：李佳奇 何睿 张一童 刘海静[1,2,4] 王嫦鹤 LI Jia-qi;HE Rui;ZHANG Yi-tong;LIU Hai-jing;WANG Chang-he(Shaanxi University of Traditional Chinese Medicine,Xianyang Shaanxi 712046;Shaanxi Institute for Food and Drug Control,Xi’an 710065;Department of Pharmacy,Honghui Hospital Affiliated to Xi’an Jiaotong University,Xi’an 710054;Shaanxi Key Laboratory of Food and Drug Safety Monitoring,Xi’an 710065)

机构地区：[1]陕西中医药大学,陕西咸阳712046 [2]陕西省食品药品检验研究院,西安710065 [3]西安交通大学附属红会医院药剂科,西安710054 [4]陕西省食品药品安全监测重点实验室,西安710065

出　　处：《中南药学》2024年第6期1505-1511,共7页Central South Pharmacy

基　　金：陕西省秦创原“科学家+工程师”队伍建设项目(No.2023KXJ-128);陕西省创新能力支撑计划(No.2020PT-041)。

摘　　要：目的 以血虚证小鼠模型和K562细胞为对象,研究白藜芦醇促进骨髓造血恢复及诱导红系分化的作用机制。方法 采用环磷酰胺诱导血虚证小鼠模型,BALB/c小鼠随机分为对照组,模型组,白藜芦醇低、中、高剂量组(50、100、200 mg·kg^(-1)),每组8只,连续给药10 d后,采用全自动血细胞分析仪检测小鼠外周血血常规;采用酶联免疫法(ELISA)分析小鼠血浆中白细胞介素-6(IL-6)、促红细胞生成素(EPO)浓度;流式细胞术检测不同浓度白藜芦醇对K562细胞凋亡、周期分布及细胞表面红系标志蛋白(CD71、CD235a)的影响;RT-qPCR检测红系分化相关基因(CD235a,CD71,GATA1,γ-globin,HBA1,HBB 和 ZFPM1)的表达,Western blot法检测JNK1/2/3和ERK1/2表达。结果 与模型组比较,白藜芦醇可不同程度提高模型组小鼠外周血中白细胞(WBC)、红细胞(RBC)、血小板(PLT)、网织红细胞(RET)数量与血红蛋白(HGB)浓度(P<0.05),升高血浆中的IL-6、EPO浓度(P<0.05)。流式细胞术结果表明,白藜芦醇能够通过S期阻滞抑制K562细胞增殖,诱导其凋亡;与对照组相比,白藜芦醇能显著增加红系标志抗原CD71、CD235a阳性表达率,且存在浓度依赖性(P<0.05)。RT-qPCR结果表明,白藜芦醇可呈剂量依赖性上调CD235a、CD71、GATA1、γ-globin、HBA1、HBB等红系分化相关基因的表达(P<0.05)。Western blot结果表明,白藜芦醇可上调ERK1/2和JNK1/2/3蛋白的磷酸化水平。结论 白藜芦醇能够通过调节JNK通路和ERK通路,增加细胞表面抗原CD71、CD235a阳性表达,增加红系分化关键基因的表达,诱导K562细胞红系分化,并能改善小鼠由环磷酰胺所致的血虚证症状,为白藜芦醇促进造血的作用机制研究奠定基础。Objective To determine the effect of resveratrol(RES)on the hematopoietic function in mice with blood deficiency syndrome induced by cyclophosphamide,and the mechanism of RES in inducing erythroid differentiation of K562 cells.Methods Cyclophosphamide was used to induce the mouse model of blood deficiency syndrome.After the modeling,BALB/c mice were randomly divided into a control group,a model group,and low,medium,and high dose RES groups(50,100 and 200 mg·kg^(-1)).The mice were given the drug continuously for 10 days.On the 11th day,the peripheral blood cells of mice were analyzed with a blood cell analyzer.The concentration of erythropoietin(EPO)and interleukin-6(IL-6)in the plasma of mice were determined by enzyme linked immunosorbent assay.The apoptosis,cell cycle distribution and the expression of erythroid differentiation marker proteins CD71 and CD235a of K562 cells induced by different concentrations of RES were measured by flow cytometry.The mRNA expressions of CD235a,CD71,GATA1,γ-globin,HBA1,HBB and ZFPM1 were detected by RT-qPCR,and the expressions of JNK1/2 and ERK1/2 were detected by Western blot.Results Compared with the model group,the white blood cells,red blood cells,platelets,reticulocyte and hemoglobin in the blood of mice in RES groups were all increased(P<0.05).The expressions of EPO and IL-6 in the RES groups were significantly increased(P<0.05).Flow flow cytometry showed that compared with the control group,RES inhibited the proliferation via S phase block and induced the apoptosis of K562 cells,and greatly increased the positive expression rate of erythroid surface marker proteins CD235a and CD71 in a dose-dependent manner(P<0.05).RT-qPCR showed that the expressions of erythroid differentiation related genes such as CD235a,CD71,GATA1,γ-globin,HBA1,and HBB in K562 cells were significantly increased after the treatment with different concentrations of RES,also in a dose-dependent manner(P<0.05).Western blot showed that RES up-regulated the phosphorylation levels of ERK1/2 and JNK1/2/3 pr

关 键 词：白藜芦醇 血虚证 K562细胞 红系分化 ERK通路 JNK通路 

分 类 号：R286[医药卫生—中药学]