2015-08-012024-05-17https://scholars.lib.ntu.edu.tw/handle/123456789/686048摘要：在此一研究計畫中我們主要是研究新陳代謝與發炎反應與免疫調控之間的關係，因此我們將建立目前已經知道具有免疫調控能力的間葉幹細胞(mesenchymal stem cells)來進行相關的研究。我們將研究代謝性發炎反應與脂肪細胞分化和間葉幹細胞的訊息傳導調控，並進一步了解飽和脂肪酸是否會影響到間葉幹細胞的分化能力和免疫調控的活性。此外，我們也將建立高脂肪飲食誘發的小鼠肥胖模式來研究在這些小鼠體內的間葉幹細胞的活性和調控能力。我們也想進一步研發出相關的可能方法來抑制與代謝性發炎的訊息傳導，來達到讓間葉幹細胞有更好免疫調控能力的方法。第一年：我們將研究與脂肪細胞相關的發炎反應對間葉幹細胞在分化和免疫調控能力上的影響，因此在第一年的計畫中我們要研究間葉幹細胞在不同濃度和種類的脂肪酸培養下是否對其分化成脂肪細胞有所影響(Aim#1)或是會促進間葉幹細胞較容易成為發炎狀態(Aim#2)。所以我們要研究含有Palmiticacid 和stearic acid 的飽和脂肪酸培養基內對間葉幹細胞分化成脂肪細胞的影響。此外，我們也將進一步研究在這些狀態下培養出來的間葉幹細胞是否與adipokines 和adiponectin 的分泌有關。我們也將進一步研究這些經過這些飽和脂肪酸培養基處理過的間葉幹細胞是否會影響到其免疫調控的能力也將進一步進行研究。第二年：在第二年的計畫中，我們將利用高油脂飲食來誘導小鼠的肥胖模式，再進一步分離BM-MSCs和AD-MSCs 來進行研究(Aim#3)和代謝性發炎反應在TLR-4 和NLRP-3 發炎體在這些MSCs 上的影響(Aim#4)。這些分離出來的MSCs 將進一步研究其分化成脂肪細胞、骨細胞和軟骨細胞的能力，同時也會測定這些MSCs 抑制T 細胞增殖反應的能力。當然，我們也將評估MSCs 與TLR-4 和NLRP-3 發炎體訊息傳導的影響。第三年：在第三年的計畫中我們將研究是否能夠針對前兩年的研究結果找出可能的方法來改變這些訊息傳導的方法，尤其是在肥胖小鼠(Aim#5)。所以我們將研究抑制這些與脂肪細胞分化相關的訊息傳導，是否能夠讓間葉幹細胞較不會發展成脂肪細胞而較能夠保有免疫調節的能力。尤其是利用我們之前建立的肥胖小鼠的動物模式來了解這些相關的訊息傳導抑制後是否在這些具有發炎傾向的小鼠中的間葉幹細胞能夠保有較好的免疫調控能力。這樣，未來便有機會將這些資訊應用到臨床上。這幾年有愈來愈多的研究顯示新陳代謝與發炎反應在疾病的發生扮演了一個重要的角色，所以建立一個研究的模式來研究代謝性發炎反應與脂肪細胞分化和間葉幹細胞免疫調控能力之間的關係將可以讓我們對新陳代謝與發炎疾病和免疫調控之間的相關性有更清楚的了解。我們也相信一旦建立此一系統後將有助於我們研究代謝性發炎反應與間葉幹細胞和其免疫調控能力之間的關係，並利用此一研究系統研發出更有效的方法來促進間葉幹細胞的免疫調控能力。<br> Abstract: In this project, we aim to study the effect of metabolism-related inflammation on the developmentand immunomodulatory activities of mesenchymal stem cells (MSCs). We aim to investigate the effectof high level of fatty acids on the differentiation ability and also immune regulatory activity. Further, wealso like to study the genes involved in the adipogenesis and their role in the development and immuneregulatory functions of mesenchymal stem cells. We like to clarify the relationship betweenmetinflammation-related signals and the immunoregulatory activity of mesenchymal stem cells.First year: Investigate whether adipocyte-induced metaflammation will drive BM-MSC lineage commitmentto favor adipogenesis (Aim #1) and will promote MSCs to a pro-inflammatory state in vitro (Aim #2).To simulate an obese environment rich in SFA, we will treat differentiated 3T3-L1 preadipocytes with SFAmixture containing palmitic and stearic acids. Hypertrophied 3T3-L1 with larger lipid droplets by prolongedcell culture will also be used. Levels of adipokines including MCP-1, TNF-α, IL-1β, and adiponectin in thisobesity-mimic culture system will be quantified. The isolated MSCs will be assayed for their differentiationability and immune regulatory activity.Second year: Elucidate the immunoregulatory effects of BM-MSCs and AD-MSCs derived from high-fatdiet induced obese murine (Aim #3) and examine the effects of metaflammation on TLR4 and NLRP3inflammsome signaling pathways in MSCs (Aim #4).To explore the MSC function under the influence of obesity-associated metaflammation in vivo, we willestablish a murine model of diet-induced obesity. Both BM-MSCs and AD-MSCs will be isolated andexpanded from lean and obese mice. The isolated MSCs will be (i) tested for trilineage differentiationpotential into adipocytes, osteocytes and chondrocytes; (ii) examined for suppressive potency onCD3/CD28-activacted T cell proliferation; (iii) assessed for activation of components of TLR4 (TLR4,NF-κB) and NLRP3 inflammasome (NLRP3, ASC, and caspase-1) signaling.Third year: Develop therapeutic strategy to restore the impaired immunosuppressive function of MSCs in thecontext of metaflammation in obese mice (Aim #5).To confirm the clinical significance of the experimental findings achieved in the second year, we willinvestigate effects of supplement with galecin-3, neutralization of TLR4, inhibition of NLRP3 inflammasome,or treatment with short-chain fatty acids in protecting MSCs from impaired function in obese mice. Thepotential efficacy of these strategies will be tested in the obesity-mimic culture system first. Afterwards, themost effective method will be further investigated in the model of high-fat induced obese mice.In this project, we like to study the effect of metinflammation on the development and immune regulatoryactivity of mesenchymal stem cells. We will establish both in vitro and in vivo models to study themetinflammation-related genes and signals in the adipogenesis and immune regulatory activity ofmesenchymal stem cells. We also like to explore possible strategies to develop the mesenchymal stem cellswith higher immune regulatory activity.