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請用此 Handle URI 來引用此文件: http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/84026
完整後設資料紀錄
DC 欄位值語言
dc.contributor.advisor林琬琬(Wan-Wan Lin)
dc.contributor.authorChun-Hung Leeen
dc.contributor.author李駿宏zh_TW
dc.date.accessioned2023-03-19T21:28:22Z-
dc.date.copyright2022-04-26
dc.date.issued2022
dc.date.submitted2022-04-20
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dc.identifier.urihttp://tdr.lib.ntu.edu.tw/jspui/handle/123456789/84026-
dc.description.abstract第三誘餌受體,別名腫瘤壞死因子受體超家族成員6b,是一個多效性的可溶因子,藉由誘餌及非誘餌兩種不同方式調節細胞的功能。過去在癌症及發炎性疾病的研究中,第三誘餌受體展示其抗細胞凋亡與抗發炎的作用。類鐸受體(TLRs)屬於模式辨認受體,參與啟動發炎反應以對抗病原體。目前關於第三誘餌受體在TLR4引起的發炎反應中是否扮演角色仍不是很清楚。在這篇研究中,我們發現在骨髓源性巨噬細胞,第三誘餌受體並不影響脂多醣引起的COX-2、iNOS、NLRP3 及 pro-IL-1 蛋白表現。先前報導指出脂多醣刺激巨噬細胞會形成一種稱為aggresome-like induced structures (ALIS) 的結構。ALIS含有泛素化蛋白的聚集,屬於一種壓力引起的反應,參與第一型主要組織相容性複合體的抗原呈現。我們的結果顯示,第三誘餌受體會藉由抑制活性氧化物及p38 MAPK的磷酸化來減少脂多醣所引起的ALIS生成。除了脂多醣外,我們還發現第三誘餌受體也會抑制ZnPP、 bafilomycin A1與MG132引起的泛素化蛋白。已知ALIS會受到自噬作用的調控,我們發現rapamycin會減少脂多醣引起的ALIS生成,而在WT與DcR3表現的巨噬細胞中,bafilomycin A1會增加相同程度的LC3-II累積。不過,DcR3不改變脂多醣引起的p62與HO-1表現。整體而言,儘管第三誘餌受體不影響脂多醣引起的發炎反應,我們的結果顯示第三誘餌受體在ALIS的生成中扮演全新的角色。zh_TW
dc.description.abstractDecoy receptor 3 (DcR3), also known as tumor necrosis factor receptor (TNFR) superfamily member 6b (TNFRSF6B), is a pleiotropic factor which modulates cell functions via decoy and non-decoy actions. DcR3 has been shown its anti-apoptotic and anti-inflammatory effects on cancers and inflammatory diseases. Toll-like receptors (TLRs), which are the members of pattern recognition receptors (PRRs), trigger inflammatory response against invading pathogens. However, it remains unclear whether DcR3 plays a role in TLR4-induced inflammatory response. In this study, we demonstrated that in bone marrow-derived macrophages (BMDMs) DcR3 did not affect lipopolysaccharide (LPS)-induced COX-2, iNOS, NLRP3 and pro-IL-1 protein expression. It has been reported that aggresome-like induced structures (ALIS) are formed in response to LPS in macrophages. ALIS, which containing ubiquitinated protein aggregates, are stress-induced response involved in MHC class I antigen presentation. Our findings illustrated that DcR3 can decrease LPS-induced ALIS formation via inhibiting cellular ROS production and p38 MAPK phosphorylation. In addition to LPS, ZnPP-, bafilomycin A1- and MG132-induced ubiquitinated protein expression were blocked by DcR3. Confirming the notion that ALIS is controlled by autophagy, we found that rapamycin can decrease LPS-induced ALIS formation, while bafilomycin A1 can increase LC3-II accumulation with similar extent in WT and DcR3 macrophages. Nevertheless, DcR3 expression does not change p62 and HO-1 expression induced by LPS. Taken together, despite the fact that DcR3 does not alter LPS-induced inflammatory response, our data suggest novel roles of DcR3 in ALIS formation.en
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dc.description.tableofcontents口試委員會審定書 i 誌謝 ii 中文摘要 iii Abstract iv Abbreviations vi Chapter 1. Introduction 1 1.1 Decoy receptor 3 (DcR3) and its decoy and non-decoy actions 1 1.1.1 Structure of DcR3 1 1.1.2 Decoy function of DcR3 2 1.1.3 Non-decoy function of DcR3 3 1.1.4 DcR3 and cancer 4 1.1.5 DcR3 and inflammation 5 1.1.6 DcR3 and toll-like receptors (TLRs) 6 1.2 Innate immune system 6 1.2.1 Innate immunity and pattern recognition receptors (PRRs) 6 1.2.2 Toll-like receptors (TLRs) 8 1.3 Aggresome-like induced structures: a specific cellular stress response in control of protein homeostasis and antigen presentation 10 1.3.1 Cellular stress response 10 1.3.2 Protein homeostasis 10 1.3.3 Aggresome-like induced structures 11 1.3.4 The function of ALIS 12 1.3.5 The regulation of ALIS 13 Specific Aim 15 Chapter 2. Materials and Methods 16 2.1 Reagents and antibodies 16 2.2 Mice and ethics statement 17 2.3 Enzyme-linked immunosorbent assay (ELISA) 17 2.4 Cell culture of bone marrow-derived macrophages 18 2.5 Immunoblotting 19 2.6 Measurement of cellular reactive oxygen species 20 2.7 Immunofluorescence 21 2.8 Statistical analysis 22 Chapter 3. Results 23 3.1 DcR3 does not alter LPS-induced inflammatory responses in BMDMs 23 3.2 ALIS forms in BMDMs in response to LPS 23 3.3 DcR3 decreases LPS-induced ALIS formation 24 3.4 ALIS formation requires ROS and p38 activation 25 3.5 DcR3 attenuates resting state and LPS-induced cellular ROS production and inhibits LPS-induced p38 phosphorylation 25 3.6 DcR3 does not alter LPS-induced HO-1 expression 26 3.7 Hemin and ZnPP do not affect LPS-induced ALIS formation in BMDMs, and ZnPP-induced ubiquitinated protein expression is blocked by DcR3 27 3.8 Bafilomycin A1- and MG132-induced ubiquitinated protein expression are blocked by DcR3 28 3.9 Rapamycin inhibits LPS-induced ALIS formation in macrophages, while DcR3 does not affect bafilomycin A1-induced LC3-II accumulation 29 Chapter 4. Discussion 31 References 39 Figures and Legends 56 Figure 1. DcR3 does not alter LPS-induced inflammatory responses in BMDMs 56 Figure 2. LPS induces ALIS formation in BMDMs 58 Figure 3. DcR3 expression reduces ubiquitinated protein and puncta in BMDMs after LPS stimulation 59 Figure 4. ROS production and p38 activation contribute to LPS-induced ALIS formation 61 Figure 5. DcR3 attenuates the resting and LPS-induced cellular ROS level and inhibits LPS-induced p38 phosphorylation 63 Figure 6. DcR3 does not alter LPS-induced HO-1 expression 65 Figure 7. Hemin and ZnPP do not affect ALIS formation in LPS-stimulated BMDMs, and ZnPP-induced ubiquitinated protein expression is blocked by DcR3 66 Figure 8. Bafilomycin A1- and MG132-induced ubiquitinated protein expression are blocked by DcR3 68 Figure 9. Rapamycin inhibits LPS-induced ALIS formation in macrophages, while DcR3 does not affect bafilomycin A1-induced LC3-II accumulation 69
dc.language.isoen
dc.subject泛素化zh_TW
dc.subject發炎反應zh_TW
dc.subject脂多醣zh_TW
dc.subject巨噬細胞zh_TW
dc.subject第三誘餌受體zh_TW
dc.subjectALISzh_TW
dc.subjectBMDMen
dc.subjectubiquitinationen
dc.subjectALISen
dc.subjectlipopolysaccharideen
dc.subjectinflammationen
dc.subjectDcR3en
dc.title探討DcR3參與脂多醣在巨噬細胞誘導免疫反應及ALIS之角色zh_TW
dc.titleThe Roles of DcR3 in the Regulation of LPS-Induced Immune Response and ALIS in Macrophagesen
dc.typeThesis
dc.date.schoolyear110-2
dc.description.degree碩士
dc.contributor.oralexamcommittee謝世良(Shie-Liang Hsieh),徐立中(Li-Chung Hsu),張永祺(Yung-Chi Chang)
dc.subject.keyword巨噬細胞,第三誘餌受體,ALIS,泛素化,脂多醣,發炎反應,zh_TW
dc.subject.keywordBMDM,DcR3,ALIS,ubiquitination,lipopolysaccharide,inflammation,en
dc.relation.page70
dc.identifier.doi10.6342/NTU202200703
dc.rights.note未授權
dc.date.accepted2022-04-20
dc.contributor.author-college醫學院zh_TW
dc.contributor.author-dept藥理學研究所zh_TW
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