Activating transcription factor 3 (ATF3)在囓齒類動物缺血性腦中風的角色及其神經保護機制

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Title:	Activating transcription factor 3 (ATF3)在囓齒類動物缺血性腦中風的角色及其神經保護機制 Role of activating transcription factor 3 (ATF3) in the ischemic stroke and its neuroprotective mechanisms in rodents
Authors:	黃千瑜 Huang, Chien-Yu
Contributors:	何元順林天南
Keywords:	中風;神經保護;ATF3;CTMP;p-Akt stroke;neuroprotection;ATF3;CTMP;p-Akt
Date:	2014-05-26
Issue Date:	2019-09-16 13:03:31 (UTC+8)
Abstract:	中風是造成全世界失智、行動不便和死亡的主要原因之一，然而受限於有限的治療藥物及其使用時機，使中風至今仍是一個棘手的疾病。Activating transcription factor 3 (ATF3)隸屬於含有basic-region leucine zipper (bZIP) motif 蛋白結構的ATF/cyclic AMP response element-binding (ATF/CREB) 轉錄因子家族，過去的研究指出，當大鼠大腦遭受缺血再灌流(MCAO)之傷害時，ATF3的mRNA會被短暫地誘發表現，若預先以AAV-ATF3大量表現ATF3，則可減少小鼠永久性腦缺血所造成的腦損傷。然而因為先前的研究對ATF3在缺血性中風的相關保護機制仍然不清楚。故本論文的目的主要探討ATF3在缺血性中風的保護作用，並分析其調控的訊息傳遞路徑。 ATF/CREB家族至少有7個成員，在大鼠中大腦動脈阻斷(MCAO)模式中，ATF3被誘發的表現變化最為明顯，若預先或在MCAO傷害後，給予大鼠Ad-ATF3則可以減少大腦損傷；同樣，預先大量表現ATF3也會減少OGD stress所造成的神經細凋亡。反之將ATF3基因剔除(ATF3 KO)或靜默(ATF3-siRNA)則會加重缺血性造成的神經細胞死亡及大腦損傷，然而，若進一步將Ad-ATF3送入ATF3基因剔除之神經細胞或小鼠，則會減少此一損傷情形。接下來我們發現，ATF3在大腦遭受缺血再灌流傷害的保護機制，是透過直接抑制下游基因CTMP (Akt抑制因子)的轉錄而來。大量表現ATF3會直接結合在CTMP的啟動子上並抑制其轉錄而增加p-Akt(473)表現；反之在ATF3基因剔除神經細胞或小鼠，則會有較低的p-Akt(473)但較高的CTMP蛋白表現，而且ATF3和CTMP被OGD stress所誘發的表現則有此消彼長之現象。在MCAO後再投予CTMP-siRNA則可增加p-Akt(473)、降低腦損傷範圍以及改善大鼠行為表現。且在靜默ATF3情況下，給予CTMP-siRNA則可以減少神經細胞死亡。由以上結果證明ATF3在缺血性中風的神經保護機制是透過抑制CTMP而來，而本論文所解開之訊息傳遞路徑-ATF3-CTMP-p-Akt(473)或許可提供一個新的治療標的供未來進一步研究。 Stroke is a common cause of dementia, disability and death worldwide. However, due to the limited therapeutic options, the disease is difficult to treat. Activating transcription factor 3 (ATF3) belongs to the ATF/cyclic AMP response element-binding (ATF/CREB) super family of transcription factors, which is characterized by a basic-region leucine zipper (bZIP) motif. ATF3 mRNA transiently increased after brain ischemia-reperfusion. Furthermore, over-expression of ATF3 in vivo via recombinant adeno-associated virus reduces brain damage following a permanent cerebral ischemic insult in mice. However, the pathophysiological role of ATF3 after brain ischemia remains unknowed. In the present study, we investigated the neuro-protective role of ATF3 and its underlying mechanisms following transient ischemic insult. There are at least 7 members of the ATF/CREB family. However, ATF3 mRNA is the most highly regulated member following 60-min MCAO (Middle Cerebral Artery Occlusion) model, and expression of ATF3 peaks 12-hr after reperfusion. Interestingly, treating rats with Ad-ATF3 before or immediately after ischemic insult reduces brain damage. Neuronal cell death was attenuated by ATF3 gene transfer and KO or silenced by ATF3 siRNA in primary neurons after exposure to an OGD challenge worsened neuronal cell death. We then used a loss-of-function approach to demonstrate that ATF3 knockout mice had larger infarct volumes and neurological deficits upon transient ischemic insult. This worsened effect was abolished when knockout mice received adenovirus gene transfer of ATF3. Next, we found that the protective effect of ATF3 mediates its effects through suppressing a downstream gene, CTMP (an endogenous inhibitor of Akt). Our data demonstrate that the expression pattern of ATF3 and CTMP is temporally negatively correlated, and ATF3 increases p-Akt(473) by directly suppressing CTMP. Moreover lower p-Akt(473) is coincident with high levels of CTMP in ATF3 KO mice or ATF3 KO primary neurons after ischemia-reperfusion. Furthermore, we also demonstrate that silencing CTMP increases p-Akt(473) and decreases neural cell death and infarct volume upon transient ischemic insult. Finally, our results show that ATF3-CTMP-p-Akt(473) signaling is a potential therapeutic target for ischemic stroke in future studies.
Description:	博士指導教授-何元順共同指導教授-林天南委員-王寧委員-王家儀委員-李文森委員-李怡萱委員-施純明
Data Type:	thesis
Appears in Collections:	[Graduate Institute of Medical Sciences] Dissertation/Thesis

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