2016-08-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/644210摘要：長壽基因Sirt1 是因其可延長生物體的壽命而得名。它可經由其本身具去甲基酵素的功能來對生物體做基因外修飾。奇妙的是它也是某些植物性荷爾蒙的作用的標的，這些植物性荷爾蒙因其具選擇性促進(在骨骼)或擷抗(在乳房)女性荷爾蒙受體的特性，故現被嘗試用來治療停經後骨質疏鬆症。婦女停經後由於缺乏女性荷爾蒙所以容易引起骨質流失，然而直接補充女性荷爾蒙現在已知會增加乳癌的風險，因此具選擇性女性荷爾蒙功能的植物性荷爾蒙在這方面的研發遂變為重要。我們實驗室一直以來都在從事成骨細胞及其前驅細胞─間葉幹細胞─應用在骨質疏鬆症的研究(J Biol Chem 2007; J Biol Chem 2010; Cell Transplant 2011; Biomaterials 2012;Biomaterials 2013; Oncotarget 2015)。我們實驗室之前曾經闡明Resveratrol 這種植物性荷爾蒙，可以在人類間葉幹細胞去調控骨化最主要的轉譯因子Runx2 (J Bone Miner Res2011, 迄今已被引用超過50 次)。之前有文獻指出，氧化反應物(ROS)--常見於細胞老化時--的積聚會減少間葉幹細胞的骨化。有趣的是，我們最實驗室近發現在間葉幹細胞中長壽基因Sirt1 可以直接和Runx2 作用，然它的表現量會被ROS 影響，而且長壽基因Sirt1的下游分子Foxo3a (它同時也是強力的抗氧化分子) ，在高濃度的ROS 下會被甲基化，而長壽基因Sirt1 其本身具去甲基酵素的功能。這些現象似乎暗示著：在氧化反應物(ROS)或老化減少間葉幹細胞的骨化過程中，長壽基因Sirt1 的Sirt1/Foxo3a/Runx2 訊息傳導路徑可以扮演反向的調控角色。因此，我們希望提出下列的研究計劃，來探討長壽基因Sirt1 在間葉幹細胞自我再生及骨化上所扮演的角色。我們的研究目標包括：目標一：闡明氧化反應物(ROS) 在間葉幹細胞分化及自我再生上的影響，及長壽基因Sirt1 在其中的調控。目標二：剖析在間葉幹細胞的骨化過程中，長壽基因Sirt1 對Foxo3a 及Runx2 的作用機轉。目標三：以Sirt1 轉殖小鼠的動物模式，驗證在ROS 減少間葉幹細胞骨化造成骨鬆時，長壽基因Sirt1 的Sirt1/Foxo3a/Runx2 訊息傳導路徑確可扮演反向的調控角色。希望經由這個研究的進行，能讓我們進一步了解在氧化反應物減少間葉幹細胞的骨化過程中，長壽基因Sirt1 所扮演的調控角色及其作用機轉，以提供治療治骨質疏鬆可能的藥物標的。<br> Abstract: First discovered as a deactylase important in epigenetic regulation, the longevity gene Sirt1 hassince been shown to extend the lifespan of many organisms. It is also a target of somephytoestrogens, which are plant-derived compounds with selective estrogenic properties (i.e.agonistic for bone but antagonistic for breast) which are relevant for treatment of osteoporosis(OP). OP is especially critical to postmenopausal women, since the loss of estrogen at menopauseleads to a rapid drop in bone density. However, direct estrogen replacement increases the risk of breastcancer; thus, phytoestrogens remain an important area of research for combating postmenopausal OP.Our laboratory’s overall goal is to understand mechanisms of osteo-differentiation in osteoblastsand mesenchymal stem cells (MSCs)—the progenitors of osteoblasts—for therapeutic application ofpostmenopausal OP (J Biol Chem 2007; J Biol Chem 2010; Cell Transplant 2011; Biomaterials 2012;Biomaterials 2013; Oncotarget 2015). We have previously elucidated the molecular mechanismsby which the phytoestrogen resveratrol transcriptionally modulate Runx2—the mastertranscription factor for osteogenesis (J Bone Miner Res 2011, cited >50 times). Previous studieshave demonstrate that reactive oxygen species (ROS), which is elevated in aging and with cellularsenescence, can decrease MSC osteo-differentiation. Interestingly, our recent findings show thatSirt1 could directly interact with Runx2, yet its expression level in MSCs is modulated by ROS.Moreover, its downstream modulator Foxo3a—a prominent anti-oxidative molecule—is acetylatedunder high levels of ROS and Sirt1 functions as a deacetylase. Thus, our current findings suggestthat the Sirt1/Foxo3a/Runx2 axis may be a pathway to target for reversal of ROS/age-relateddecreases to MSC osteo-differentiation.In this Proposal, therefore, we hope to explore the role of the longevity gene Sirt1 on MSCself-renewal and osteo-differentiaton. Our aims are to:Aim 1: Elucidate ROS-mediated changes to all MSC lineages and MSC self-renewal, and effectsof Sirt1 modulationAim 2: Dissect the mechanisms of (a) Sirt1 deactylase actions on Foxo3a and (b) Sirt1 directinteractions with Runx2 during MSC osteo-differentiationAim 3: In vivo validation of modulating Sirt1/Foxo3a/Runx2 axis in ROS-induced osteoporosisusing Sirt1 transgenic miceWe hope in this Proposal to clearly understand the molecular interactions in theSirt1/Foxo3a/Runx2 axis and ROS/age-related perturbations during MSC osteo-differentiation, andthus lead to discovery of druggable anabolic targets for use towards OP.骨質疏鬆症長壽基因Sirt1間葉幹細胞氧化反應物去甲基酵素osteoporosisSirt1mesenchymal stem cellsreactive oxygen species (ROS)deacetylase