Hepcidin在脑缺血再灌注损伤中对HIF-1α的调控及其机制的研究

The abnormally increased brain iron could distinctly aggravate the process of the neuron damage. The hypoxia induced factor 1α (HIF-1α), a nuclear transcription regulation factor, plays an important role in the low oxygen physiological process. HIF-1α can activate the downstream genes and prevent cells from damage during cerebral ischemia. Researches showed that hepcidin was significantly increased during ischemia-reperfusion, and it participated in the process of brain iron increased. The existing studies had confirmed that the iron could be involved in the regulation of HIF-1α as the hydroxyprolinase cofactor, whether hepcidin could affect the HIF-1α expression by adjusting the change of cellular iron level, and the molecular mechanisms of the regulation has not been clarified at present. Here, we use hepcidin knockout mouse (Hamp-/-), HIF-1α conditional knockout mouse and FPN1 neuron spcific knockout mouse model of MCAO, as well as the different cell types and co-cultrue of the oxygen glucose deprivation(OGD) model in vitro to explore if the molecular mechanism of hepcidin participating in the regulation of HIF-1α. This research will reveal the neurological damage or protection of hepcidin and HIF-1α in the ischemic stroke, and it will provide theoretical guidance for the clinical treatment of ischemic stroke diseases.

脑铁异常增高在脑中风发病过程中对神经细胞损伤有着明显的促进作用，低氧诱导因子HIF-1α是在低氧生理过程中起作用的核转录调控因子，它能够激活下游的基因，具有神经保护的作用。研究表明，铁调素(hepcidin)在脑缺血再灌注过程中明显增高，并参与脑铁增高的过程。已有的研究已经证实，铁作为脯氨酰羟化酶的辅因子参与HIF-1α的调控，那么，hepcidin能否通过调节细胞铁水平的变化进而影响HIF-1α表达，这一调控的分子机制如何，目前尚未阐明。本研究采用hepcidin基因敲除小鼠、神经元中条件性敲除FPN1小鼠，条件性敲除 HIF-1α小鼠，通过制备MCAO模型、不同细胞种类及混合培养的体外糖氧剥夺模型，研究hepcidin对HIF-1α的调控分子机制，揭示hepcidin和HIF-1α在缺血性脑中风模型中，对神经细胞损伤或保护作用，为疾病的治疗提供理论基础。

研究表明，铁调素Hepcidin，作为铁的关键调节因子，在缺血再灌注（ischemia-reperfusion, IR）过程中明显增高，并参与脑铁增高的过程。同时，在IR过程中，缺氧诱导因子HIF-1α（Hypoxia-induciblefactor1α）能够诱导表达，具有一定的神经保护作用。铁作为脯氨酰羟化酶（Prolyl hydroxylase，PHD 2）的辅因子还能够参与HIF-1α活性调节。本项目在IR损伤的过程中，研究hepcidin对HIF-1α表达调控分子机制。我们在制备不同基因敲除小鼠大脑中动脉线栓阻断法（Middle Cerebral Artery Occulation model，MCAO）模型和神经细胞的氧糖剥夺（Oxygen and Glucose Deprivation，OGD）模型的基础上，研究发现：1、随着再灌注时间的增加，梗塞损伤的区域不断增加，在缺血半影区内Hepcidin表达上升，并特异性在星行胶质细胞中表达，而半影区内HIF-1α蛋白却是不断下降的；2、 hepcidin通过降低FPN 1的表达，进一步调节胞内铁水平，增加PHD 2的活性，加剧了HIF-1α的降解，参与神经细胞的损伤；3、全身性敲除Hamp基因的小鼠 (Hamp-/- mice)，表现出了脑铁沉积的现象，进行IR处理后，降低了脑内HIF-1α的表达及抗氧化能力，进而增加了细胞凋亡和脑梗损伤面积情况；4.星形胶质细胞中特异性敲低Hamp基因 (GFAP-shHamp)小鼠能够增加HIF-1α的表达，抑制炎症反应和凋亡通路的激活，来改善IR的损伤。本项目的完成，提出并部分阐明了在IR过程中hepcidin对HIF-1α表达调控的分子机制，为今后缺血性脑中风治疗提供了新的分子靶点，具有重要的理论和现实意义。