LPIN1/PPARA通过抑制SLC47A1介导的神经元铁死亡缓解帕金森病模型大鼠病情进展的机制研究  

Mechanism Study on LPIN1/PPARA Alleviating the Progression of Parkinson’s Disease in Rats by Inhibiting SLC47A1-Mediated Ferroptosis of Neurons

作　　者：曹明月 王巍[1] 周美宁 CAO Mingyue;WANG Wei;ZHOU Meining(Department of Clinical Laboratory,Daqing Longnan Hospital/the Fifth Affiliated Hospital of Qiqihar Medical College,Heilongjiang Daqing 163453,China;Department of Neurology,the Third Affiliated Hospital of Xi’an Medical University,Xi’an 710068,China)

机构地区：[1]大庆龙南医院、齐齐哈尔医学院第五附属医院检验科,黑龙江大庆163453 [2]西安医学院第三附属医院神经内科,西安710068

出　　处：《现代检验医学杂志》2024年第4期63-71,共9页Journal of Modern Laboratory Medicine

摘　　要：目的探究脂蛋白1(lipin1,LPIN1)对帕金森病(Parkinson’s disease,PD)模型大鼠病情进展的影响及其调控的可能分子机制。方法采用6-羟基多巴胺(6-hydroxydopamine hydrobromide,6-OHDA)注射大鼠单侧的内侧前脑束构建PD大鼠模型,并稳定转染LPIN1过表达腺病毒,评估大鼠行为学变化;检测大鼠脑组织中Fe^(2+)和还原型谷胱甘肽(glutathione,GSH)含量及酪氨酸羟化酶(tyrosine hydroxylase,TH)蛋白水平;HE染色观察大鼠脑组织病理变化。构建体外PD细胞模型,检测细胞中TH,α-突触核蛋白(α-synuclein,α-syn),LPIN1蛋白水平及细胞活力;转染LPIN1小干扰siRNA序列和过表达载体及过氧化物酶增殖物激活受体A(peroxisome proliferator-activated receptor A,PPARA)小干扰RNA(small interfering RNA,siRNA)和过表达载体,或用铁死亡诱导剂Erastin处理细胞24 h,后用6-OHDA处理细胞48 h。检测各组细胞中Fe^(2+)含量、活性氧(reactive oxygen species,ROS)、丙二醛(malondialdehyde,MDA)、GSH和炎症因子水平以评估铁死亡;CCK-8检测细胞增殖活力,Western blot法检测铁死亡相关蛋白表达。通过STRING数据库预测LPIN1的互作蛋白PPARA,利用Co-IP分析进行验证;JASPAR生物信息学网站预测PPARA与溶质载体蛋白家族47成员A1(solute carrier family 47 member 1,SLC47A1)启动子的结合位点,利用Ch-IP分析进行验证。结果模型组大鼠皮毛悚立,表现出身体持续震颤、动作迟缓、活动能力减弱等PD症状;LPIN1组大鼠运动行为及PD症状较模型组改善/减轻。与假手术组相比,模型组大鼠运动总路程缩短、平均速度降低、步长减小、总静止时间延长、步宽增宽、步态变异率增大,差异具有统计学意义(t=6.816,7.026,26.556,7.454,8.503,7.971,均P<0.05);模型组大鼠脑组织中Fe^(2+)含量升高,GSH含量及TH蛋白表达降低,差异具有统计学意义(t=8.305,13.305,7.709,均P<0.05)。LPIN1组大鼠行为学评估、各指标水平及脑组织病理改变较模型组明Objective To investigate the effect of lipin1(LPIN1)on the progression of Parkinson’s disease(PD)in rats and the possible molecular mechanism of its regulation.Methods The PD rat model was established by injection of 6-Hydroxydopamine hydrobromide(6-OHDA)into the medial forebrain tract of rats,and the LPIN1-overexpressing adenovirus was stably transfected to evaluate the behavioral changes of rats.The content of Fe^(2+)and Glutathione(GSH)and the protein level of tyrosine hydroxylase(TH)in rat brain were detected,and HE staining was used to observe the pathological changes in rat brains.The PD cell model was constructed in vitro,and TH,α-synuclein(α-syn),LPIN1 protein levels and cell viability were detected.LPIN1 small interfering siRNA sequence and overexpression vector and peroxisome proliferator-activated receptor(PPARA)small interfering(siRNA)and overexpression vector were transfected,or ferroptosis inducer erastin was used to treat cell for 24 h,then cells were treated with 6-OHDA for 48h.The levels of Fe^(2+),reactive oxygen species(ROS),malondialdehyde(MDA),GSH and inflammatory factors in cells were detected to evaluate ferroptosis.Cell viability was detected with CCK-8,and the expressions of ferroptosis related proteins were detected with Western blot.The interacting protein PPARA of LPIN1 was predicted by STRING database and verified by Co-IP analysis.The binding site of PPARA to the promoter of solute carrier family 47 member 1(SLC47A1)was predicted by JASPAR bioinformatics and verified by Ch-IP analysis.Results The fur of the rats in the model group was frightened,and PD symptoms such as continuous tremor,slow movement and weakened activity were shown.The motor behavior and PD symptoms of LPIN1 group were improved/alleviated compared with the model group.Compared with the sham operation group,the total distance of the model group was shortened,the average speed was reduced,and the step length was reduced,while the total resting time was prolonged,the step width was widened,and the gait variation ra

关 键 词：帕金森病 脂蛋白1 过氧化物酶增殖物激活受体A 溶质载体蛋白家族47成员A1 铁死亡 

分 类 号：R-332[医药卫生]