氢气对慢性脑缺血大鼠脑组织铁代谢相关蛋白表达的影响  

Effects of hydrogen on expression of iron metabolism-related proteins in brain tissues of rats with chronic cerebral ischemia

作　　者：谭永星 凌秀梅[3] 董庆华[2] 粟立斌[4] 郝怡萌 谢克亮 于泳浩[1] Tan Yongxing;Ling Xiumei;Dong Qinghua;Su Libin;Hao Yimeng;Xie Keliang;Yu Yonghao(Department of Anesthesiology,Tianjin Medical University General Hospital Tianjin Research Institute of Anesthesiology,Tianjin 300052,China;Surgical Intensive Care Unit,Affiliated Hospital of Guilin Medical University,Guilin 541001,China;Department of Radiology,Affiliated Hospital of Guilin Medical University,Guilin 541001,China;Intensive Care Unit,Guanyang County People′s Hospital,Guanyang 541600,China)

机构地区：[1]天津医科大学总医院麻醉科,天津市麻醉学研究所,300052 [2]桂林医学院附属医院SICU,541001 [3]桂林医学院附属医院放射科,541001 [4]灌阳县人民医院ICU,541600

出　　处：《中华麻醉学杂志》2020年第12期1511-1515,共5页Chinese Journal of Anesthesiology

基　　金：广西壮族自治区自然科学基金(2015GXNSFAA139130);桂医科[2018]5号中青年教职工科研能力提升项目(2018glmcy069);广西壮族自治区卫生健康委员会自筹课题(Z20180612,Z20190270)。

摘　　要：目的探讨氢气对慢性脑缺血(CCI)大鼠脑组织铁代谢相关蛋白表达的影响。方法健康成年雄性Wistar大鼠36只,体重300~400 g,采用随机数字表法分为3组(n=12):假手术组(Sham组)、CCI组、富氢盐水组(H组)。采用改良的双侧颈总动脉永久性结扎法制备大鼠CCI模型。CCI组及H组在大鼠建模4周后每天固定时点,分别腹腔注射生理盐水或富氢盐水10 ml/kg,1次/d,持续2周。CCI建模第6周时,处死大鼠,取脑组织,免疫组化法检测特异性神经元核蛋白(NeuN)、谷胱甘肽过氧化物酶4(GPX-4)的表达,采用改良Perl染色法检测病灶脑组织铁沉积量,采用比色法测定MDA、谷胱甘肽(GSH)、谷胱甘肽过氧化物酶(GSH-px)水平,采用ELISA法检测4-羟基壬烯酸(4-HNE)水平,采用流式细胞术检测神经细胞ROS水平,采用RT-PCR法检测转铁蛋白(TF)、转铁蛋白受体1(TFR1)、铁蛋白轻链(FTL)、铁蛋白重链-1(FTH1)、二价金属离子转运体(DMT1)、铁反应元件结合蛋白2(IREB2)的mRNA表达。结果与Sham组比较,CCI组及H组海马CA1区NeuN、GPX-4染色阳性细胞百分比、GSH、GSH-px水平降低,铁染色阳性细胞百分比、MDA、4-HNE及神经细胞ROS水平升高,FTL mRNA、FTH1 mRNA表达下调,TF mRNA、TFR1 mRNA、DMT1 mRNA和IREB2 mRNA表达上调(P<0.05);与CCI组比较,H组海马CA1区NeuN、GPX-4染色阳性细胞百分比、GSH和GSH-px水平升高,铁染色阳性细胞百分比、MDA、4-HNE及神经细胞ROS水平降低,FTL mRNA、FTH1 mRNA表达上调,TF mRNA、TFR1 mRNA、DMT1 mRNA和IREB2 mRNA表达下调(P<0.05)。结论氢气对CCI大鼠可产生神经保护作用,机制可能与其调控CCI后铁代谢及转运相关蛋白的表达,减轻神经细胞铁超载,进而抑制氧化应激有关。Objective To investigate the effects of hydrogen on the expression of iron metabolism-related proteins in brain tissues of rats with chronic cerebral ischemia(CCI).Methods Thirty-six healthy adult male Wistar rats,weighing 300-400 g,were divided into 3 groups(n=12 each)by a random number table method:sham operation group(Sham group),CCI group and hydrogen-rich saline group(H group).The model of CCI was established by permanent occlusion of bilateral common carotid arteries in anesthetized rats.In CCI group and H group,normal saline and hydrogen-rich saline 10 ml/kg were intraperitoneally injected once a day at a fixed time point every day for 2 consecutive weeks starting from 4 weeks after establishing the model.At the 6th week after establishing the model,the rats were sacrificed and brain tissues were obtained for determination of the expression of specific neuronal nuclear protein(NeuN)and glutathione peroxidase 4(GPX-4)(by immunohistochemistry),iron deposition in brain tissues of the lesion(by improved Perl staining),levels of malondialdehyde(MDA),glutathione(GSH)and glutathione peroxidase(GSH-px)(by colorimetric method),level of 4-hydroxynonenoic acid(4-HNE)(by enzyme-linked immunosorbent assay),and level of ROS in nerve cells(by flow cytometry),and expression of transferrin(TF),transferrin receptor 1(TFR1),ferritin light chain(FTL),ferritin heavy chain-1(FTH1),divalent metal-ion transporter 1(DMT1)and iron-responsive element binding protein 2(IREB2)mRNA(by real-time polymerase chain reaction).Results Compared with Sham group,the percentage of NeuN and GPX-4 positive cells in hippocampal CA1 region and levels of GSH and GSH-px were significantly decreased,the percentage of iron staining positive cells,levels of MDA and 4-HNE,and level of ROS in nerve cells were increased,the expression of FTL and FTH1 mRNA was down-regulated,and the expression of TF,TFR1,DMT1 and IREB2 mRNA was up-regulated in CCI group and H group(P<0.05).Compared with CCI group,the percentage of NeuN and GPX-4 positive cells in hippocampal

关 键 词：氢 脑缺血 铁 

分 类 号：R743.3[医药卫生—神经病学与精神病学]