转铁蛋白受体对铁过载所致大鼠心肌细胞铁死亡的作用及其机制  

Effect and mechanism of TFRC on ferroptosis induced by iron overload in cardiac myocytes of rats

作　　者：杨岚婷 徐涛[1,2] 杨福情 肖丹丹 宋林 王建勋 Yang Lan-Ting;Xu Tao;Yang Fu-Qing;Xiao Dan-Dan;Song Lin;Wang Jian-Xun(School of Basic Medicine,Qingdao University,Qingdao,Shandong 266000,China;Central Laboratory,Qingdao Agricultural University,Qingdao,Shandong 266109,China)

机构地区：[1]青岛大学基础医学院,山东青岛266000 [2]青岛农业大学中心实验室,山东青岛266109

出　　处：《解放军医学杂志》2022年第8期781-788,共8页Medical Journal of Chinese People's Liberation Army

基　　金：国家自然科学基金(81970253,82000290)。

摘　　要：目的观察铁过载对小鼠心脏功能及大鼠心肌细胞铁死亡的影响,探讨铁转运蛋白受体(TFRC)在大鼠心肌细胞铁死亡中的作用机制。方法C57BL/6J小鼠14只,随机分为正常饮食组和高铁饮食组各7只,分别给予常规饲料和高铁饲料喂食8周后,采用小动物超声检测其心脏功能,酶标仪检测心脏组织丙二醛(MDA)含量,Masson染色检测心脏纤维化情况。通过碘化丙啶(PI)染色和亲脂荧光染料(C11)染色检测细胞死亡率和脂质过氧化物含量,观察柠檬酸铁铵(AIC)和铁死亡抑制剂Ferrostatin-1(Fer-1)对H9C2大鼠心肌细胞的作用。AIC和铁死亡诱导剂Erastin处理H9C2细胞后,Western blotting检测TFRC、谷胱甘肽过氧化物酶4(GPX4)蛋白表达水平,qRT-PCR检测mRNA表达水平;相同条件下,小干扰RNA敲低TFRC的表达,检测H9C2细胞死亡率和脂质过氧化物含量。结果与正常饮食组比较,高铁饮食组小鼠心室间隔厚度(IVSd)、左心室后壁厚度(LVPWd)、心脏组织MDA相对含量和胶原沉积相对面积均明显增大[(0.96±0.12)mm vs.(0.73±0.09)mm,(1.18±0.28)mm vs.(0.84±0.07)mm,2.08±0.80 vs.1.00±0.50,4.04±0.60 vs.1.00±0.21,P<0.05],但左室射血分数和左心室缩短分数差异无统计学意义(P>0.05)。经500μmol/L AIC处理后,H9C2细胞死亡率和脂质过氧化物含量明显升高(33.73%±1.20%vs.2.30%±1.73%,5.36±0.06 vs.1.00±0.19,P<0.05),加用Fer-1处理后H9C2细胞死亡率和脂质过氧化物含量明显降低(19.63%±0.81%vs.33.73%±1.2%,2.03±0.12 vs.5.36±0.06,P<0.05);500μmol/L AIC处理后,H9C2细胞TFRC表达量明显升高(P<0.05),TFRC敲低后,脂质过氧化物含量和细胞死亡率明显下降(P<0.05)。Erastin诱导下H9C2细胞TFRC和GPX4蛋白表达量分别明显上调和下调(P<0.05);TFRC敲低后,细胞死亡率明显下降(P<0.05)。结论高铁饮食可诱导大鼠心肌细胞发生铁死亡损伤,TFRC参与了高铁诱导的心肌细胞铁死亡并可能成为心脏疾病治疗研究的新靶点。Objective To observe the effect of iron overload on the cardiac function of mice and cardiac myocytes ferroptosis of rats, and explore the mechanism of action of iron transporter receptor(TFRC) in cardiac myocytes ferroptosis of rats. Methods Fourteen C57BL/6J mice were randomly divided into normal-iron diet(NID) group and high-iron diet(HID) group(7 each), fed with normal diet or high-iron diet for 8 weeks, respectively. The mice cardiac function was then detected by echocardiography;The content of malondialdehyde(MDA) in the heart was detected with ELISA;Masson staining was performed to detect the cardiac fibrosis;Propidium iodide(PI) staining and lipophilic fluorescent dye(C11) were used to detect the cell mortality and the content of lipid peroxide. The effect was observed of ferric ammonium citrate(AIC) and ferroptosis inhibitor ferrostatin-1(Fer-1) on the cardiac myocytes of H9C2 rat. H9C2 cells were treated with AIC and ferroptosis inducer Erastin, Western blotting was performed to detect the expression level of TFRC and glutathione peroxidase 4(GPX4) protein, qRT-PCR was used to detect the mRNA level;at the same condition, small interfering RNA was applied to knock-down the expression of TFRC, detect the mortality of H9C2 cells and the content of lipid peroxide. Results Compared with mice in NID group, the mice in HID group showed obviously increased interventricular septal thickness(IVSd), left ventricular posterior wall thickness(LVPWd), MDA relative content and area of collagen deposition(0.96±0.12 vs. 0.73±0.09, 1.18±0.28 vs. 0.84±0.07, 2.08±0.8 vs. 1.00±0.50, 4.04±0.60 vs. 1.00±0.21, P<0.05);But no significant difference between the two groups on left ventricular ejection fraction and shortening fraction. Treated with 500 μmol/L AIC, the death rate and lipid peroxide content of H9C2 increased obviously(33.73%±1.20% vs. 2.30%±1.73%, 5.36±0.06 vs. 1.00±0.19, P<0.05), while after treatment with Fer-1, the death rate and lipid peroxide content of H9C2 decreased markedly(19.63%±0.81% vs. 33.

关 键 词：铁过载 心肌细胞 铁死亡 转铁蛋白受体 谷胱甘肽过氧化物酶4 

分 类 号：R542.2[医药卫生—心血管疾病]