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题名: 血红素调节的eIF2α激酶(Hri)对抗重金属血液毒性及维持铁代谢平衡的作用机理
作者: 王小岩
学位类别: 博士
答辩日期: 2014-11
授予单位: 中国科学院研究生院
授予地点: 北京
导师: 刘思金
关键词: ; 血液毒性 ; Hri ; 红细胞 ; 铁代谢 ; Ferroportin,Hri ; Erythroid cells injury ; Red blood cell ; Ferroportin
其他题名: Heme-regulated eIF2α kinase (Hri) plays a protective role against stresses induced by heavy metals and in orchestrating Iron metabolism
学位专业: 环境科学
中文摘要:     铁是组成生物体最重要的元素之一。人体铁代谢与红系造血系统之间的关系十分紧密。然而遗传缺陷、疾病、环境因素等可导致铁代谢异常,并损伤造血系统。目前,针对铅等重金属对血液系统的损伤,人体中是否存在对抗机制仍是未知。
    铅对红系细胞的影响主要体现在抑制血红素的合成和直接作用于成熟红细胞并引发溶血。红系细胞中血红素缺乏可激活血红素调节的eIF2α激酶(Hri),导致eIF2α磷酸化,终止蛋白合成。我们前期研究发现,Hri在红系细胞对抗砷和镉引发的氧化应激中起到保护作用。在本课题中,我们通过体内实验发现铅暴露下,Hri基因敲除小鼠(Ko)出现了典型的溶血性贫血。体内和体外研究均表明,Hri缺失情况下,铅可以导致红细胞前体分化受阻,细胞凋亡增加,即导致无效的红细胞生成。此外在动物实验中我们发现,铅暴露也促进了Ko小鼠肝脏Hepcidin表达,导致铁代谢异常,抑制红细胞生成。这些结果证实了,在铅引发的红系细胞毒性中Hri起到了至关重要的作用。
    在对成熟红细胞的研究中我们发现,Hri缺失的红细胞膜骨架存在缺陷,导致成熟红细胞生存能力减弱、供氧能力不足。Hri缺失的小鼠则表现出溶血风险增加,运动能力下降,低氧耐受能力降低,从而明确了Hri基因在小鼠成熟红细胞的功能中的重要作用。在对分子机制的探索中我们还揭示了Hri介导的Atf4信号通路在红系细胞膜骨架基因Gsn表达中的控制作用。提示着Hri-Atf4信号通路可能成为治疗溶血性贫血的新的靶点。
    此外,我们对于机体铁代谢中的关键基因Ferroportin(Fpn)在红系细胞中的功能和调节方式进行了初步研究。结果显示Fpn缺失可以导致红系前体细胞分化滞后,对抗外界压力的能力降低。通过对Fpn1a和Fpn1b的分析我们发现,Fpn1a在红系分化过程中受其启动子区域甲基化水平调节,并通过实验证明,红系细胞分化过程中可能存在一定程度的活跃的去甲基化过程,从而降低了Fpn1a基因启动子的甲基化程度,调高Fpn1a在细胞内的RNA水平。总的来说,我们发现了Fpn在红系细胞中的重要作用,同时对其在转录水平的调节进行了初步探索。
    综上所述,我们证实了Hri基因在红系细胞对抗铅损伤中的保护作用,并首次发现该基因缺失对成熟红细胞功能的影响。同时,在对Fpn的研究中,明确了Fpn在红系前体细胞中的重要功能,同时对红细分化过程中铁代谢基因的调节方式有的新的认识。研究铁代谢相关基因在血液系统中的作用及其调节机制,对于深入探索血液系统发生机制有着重要的科学意义,对于治疗和预防血液系统疾病也有一定的临床应用价值。
英文摘要:     Chronic exposure of heavy metal can induce animia. Our previous studies recognized a protective role of heme-regulated eIF2α kinase (Hri) against oxidative stress in erythroid cells exerted by arsenic and cadmium. In the current study, we looked into erythropoiesis upon Pb(NO3)2 exposure with the in vivo mouse model and ex vivo cultured E14.5 fetal liver (FL) cells. Diagnostic of hemolytic anemia were observed in Hri knockout (Ko) mice only, upon low-dose Pb administration. In vivo and ex vivo results manifested that expanded erythroid precursors experienced blocked differentiation and enhanced apoptosis, leading to ineffective erythropoiesis for Hri-null erythroid precursors under Pb exposure. Additionally, Pb treatment also promoted hepcidin expression and consequentially increased splenic iron storage, resulting in restrained iron availability for erythropoiesis. Our findings therefore deciphered a crucial role of Hri in protecting erythroid cells against Pb-induced toxicity.
    The lack of Hri can result in the abnormality of cell membrane of mature RBCs which resulted in the poor survival of RBCs and insufficient oxygen transportation under lead or hypoxia exposure. Here, we decipher a novel role of Hri mediated Atf4 signaling in governing membrane gene expression in erythrocytes. This signaling was involved in the RBC cell membrane assembling which played a critical role in the erythrocytes survival. Our study also indicated that Hri-Atf4 signaling pathway should be a novel target for hemolytic anemia therapeutics. Elevated eIF2αP and Atf4 after drug treatment should be the feasibility to enhance the stability of cell membrane of erythrocytes, which may be important in reducing the risk of hemolytic anemia under stress.
    In addition, we conducted a preliminary study in the function and regulation mode of Ferroportin (Fpn) in erythroid cells. The results showed that, deletion of Fpn can blocked erythroid precursor cells differentiation, and reduced the capacity against external pressure. We also found that Fpn1a expression was regulated by the methylation level of its promoter region during erythroid differentiation. There may be a certain degree of active demethylation process during erythroid cell differentiation, thereby reducing the methylation level of Fpn1a gene promoter, and induced the transcription of Fpn1a.
    In conclusion, we demonstrated that Hri plays a protective role under lead exposure in erythroid cells. And we find that Hri is important for the cell viability and functions of mature red blood cells. the study of iron metabolism related genes in erythroid cells has an important science significance of exploring the pathogenesis of blood system, as well as certain clinical value for the treatment and prevention of diseases of erythroid cells.
内容类型: 学位论文
URI标识: http://ir.rcees.ac.cn/handle/311016/34445
Appears in Collections:环境化学与生态毒理学国家重点实验室_学位论文

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王小岩. 血红素调节的eIF2α激酶(Hri)对抗重金属血液毒性及维持铁代谢平衡的作用机理[D]. 北京. 中国科学院研究生院. 2014.
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