氣體傳遞物質硫化氫對於阿茲海默氏症的神經保護功效

Abstract

目前阿茲海默氏症發病率持續上升，又缺乏有效治療方法的現況已造成全球醫療嚴重的挑戰。近期有研究指出硫化氫對許多神經退化性疾病如阿茲海默氏症具有抗氧化、抗細胞凋亡，以及抗發炎的效果。然而，硫化氫、類澱粉蛋白，以及tau蛋白之間的關聯性尚未闡明。在此研究中，我們將以oligomeric Aβ42處理的海馬迴初級神經元培養系統與給予壓力的阿茲海默氏症基因轉殖鼠探討硫化氫對於阿茲海默氏症的作用效果與機轉。首先，在建立海馬迴初級神經元培養系統中確認培養到第9天時，海馬迴神經元之樹突開始延展及分支;另外也發現2 μM AraC能抑制星型膠細胞之生長且對神經元造成之傷害性較小。在確認了海馬迴初級神經元培養系統之條件後，我們接著進行1 μM oligomeric Aβ42對細胞處理0，0.5，1，6及12小時後之影響，發現1 μM oligomeric Aβ42培養1小時後會造成細胞存活率下降、LDH釋放量增加、成熟神經元數目減少、神經突數目與長度減少及突觸密度下降等傷害，所以以1μM oligomeric Aβ42培養1小時為一進行藥物評估之適合條件。另外，不同劑量的硫化氫鈉（硫化氫供應物）與oligomeric Aβ42以前處理、共同處理及後處理三種方式加入海馬迴初級神經元培養系統內，我們發現50 μM硫化氫鈉前處理或共同處理oligomeric Aβ42及10 μM硫化氫鈉後處理可以保護海馬迴初級神經元培養的神經細胞型態及突觸密度。在動物實驗上發現電刺激所產生的壓力對於6個月大的阿茲海默氏症基因轉殖鼠僅造成有限之傷害。因此，此種電刺激之強度及給予方式對於6個月大的阿茲海默氏症基因轉殖鼠可能不足以產生嚴重之壓力。另外，硫化氫鈉之給予可以避免認知功能之退化可能是經由PERK/eIF2α途徑所產生的抗氧化與抗發炎反應於電刺激的阿茲海默氏症基因轉殖鼠。。因此，硫化氫對於阿茲海默氏症或許是一個具有潛力的治療策略之選擇。The worldwide incidence of Alzheimer's disease (AD) is increasing and creating an unsustainable healthcare challenge due to a lack of effective treatment options. Recent evidence shows that H2S could exert antioxidant, anti-apoptotic, and anti-inflammatory effects against neurodegenerative disease such as AD. However, the correlation among H2S, Aβ and tau protein in AD are not been fully elucidated. In this study, the effects and molecular mechanisms of H2S in AD were investigated in hippocampal primary neuronal culture and APP/PS1/Tau triple transgenic (3×Tg-AD) mice with inescapable foot-shock stress. At first, we established the hippocampal primary neuronal culture as an in vitro platform for a primary study. The neuronal processes were extended and branched on 9 days in vitro (DIV). In addition, the medium with 2 μM cytosine arabinoside (AraC) inhibited the population of astrocyte associated with limited neuronal damage. We then evaluated the effects of 1 μM oligomeric Aβ42 on hippocampal primary neurons with cultivating timing from 0.5 to 12 hr. We found that 1 hr treatment induced total cell loss and lactate dehydrogenase (LDH) released, and mature neuronal number, neuritic length, processes, branches, and synaptic density decreased. Therefore, treatment of oligomeric Aβ42 for 1 hr is an optimized timing for evaluation of drug treatment. Different concentrations of NaHS (an H2S donor) were applied to oligomeric Aβ42-treated hippocampal primary neuronal culture in pre-, co-, or post-treatment manners. We further found that NaHS pre-/co-treatment (50 μM) and post-treatment (10 μM) could effectively attenuate oligomeric Aβ42-induced toxicity. We then evaluate the in vivo effect of NaHS on the 3×Tg-AD mice associated with inescapable foot-shock stress. The 3×Tg-AD mice (6 month-old) with stress were administrated with either NaHS or vehicle, and subjected to a series of behavioral evaluation (light-dark transition test, open field test, elevated plus maze, and Morris water maze). We found that inescapable foot-shock stress only induced limited impact in 3×Tg-AD mice. However, the administration of NaHS prevented the cognitive decline associated with anti-inflammation and anti-oxidation via PERK/eIF2α pathway in 3×Tg-AD mice with mild stress. Therefore, H2S is a potential therapeutic strategy to prevent the cognitive dysfunction for AD.