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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 陳玉怜 | zh_TW |
| dc.contributor.advisor | Yuh-Lien Chen | en |
| dc.contributor.author | 李紫琳 | zh_TW |
| dc.contributor.author | Tzu-Lin Lee | en |
| dc.date.accessioned | 2025-02-20T16:29:04Z | - |
| dc.date.available | 2025-02-21 | - |
| dc.date.copyright | 2025-02-20 | - |
| dc.date.issued | 2024 | - |
| dc.date.submitted | 2024-10-21 | - |
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/96675 | - |
| dc.description.abstract | 流行病學研究顯示，細顆粒物（particulate matter, PM）暴露與心血管疾病之間存在密切關聯。然而，目前尚不清楚PM是否會加重心肌缺血/再灌流（ischemia/reperfusion, I/R）損傷，以及相關的機制。本研究探討PM暴露對I/R誘導的心臟損傷的影響，並分析脂肪幹細胞來源的外泌體（adipose stem cell-derived exosome, ADSC-Exo）中高含量的miR-221/222的潛在作用。使用miR-221/222基因剔除（KO）和miR-221/222過表達基因（TG）小鼠模型，進行10 mg/kg PM的氣管內注射處理。24小時後，小鼠接受左冠狀動脈結紮30分鐘後，進行3小時的再灌注（I/R）。在體外實驗，H9c2細胞前處理PM（50 µg/mL）並於1% O2缺氧環境下培養6小時，然後在正常氧氣環境培養12小時（hypoxia/reoxygenation, H/R）。結果顯示，PM加重了I/R（或H/R）引起的心臟損傷，通過增加活性氧化物（reactive oxygen species, ROS）和引發粒線體功能障礙，導致粒線體分裂相關蛋白（Drp1和MFF）及粒線體自噬相關蛋白（BNIP3和LC3B）的表達增加。ADSC-Exo或miR-221/222 mimics的治療顯著減少了PM+I/R誘導的心臟損傷。值得注意的是，ADSC-Exo中的miR-221/222直接調節BNIP3、LC3B和PUMA並減少其表達，最終降低細胞的粒線體自噬和細胞凋亡反應。本研究發現ADSC-Exo通過miR-221/222/BNIP3/LC3B/PUMA途徑調控粒線體自噬和細胞凋亡，顯著減少了PM+I/R所引起的心臟損傷，揭示ADSC-Exo在緩解PM加重心肌I/R損傷中的潛在治療價值。 | zh_TW |
| dc.description.abstract | Studies in epidemiology have highlighted a significant link between exposure to fine particulate matter (PM) and an increased risk of cardiovascular disease. However, whether PM exacerbates myocardial ischemia/reperfusion (I/R) injury remains unclear, and the specific mechanisms involved in this process have not yet been fully elucidated. Our previous study has shown that adipose stem cell-derived exosomes (ADSC-Exo) contain high levels of miR-221/222. This study investigated the effects of PM exacerbates mitochondrial dysfunction and increases the processes of mitophagy and cell apoptosis in I/R settings, while demonstrating that miR-221/222 in ADSC-Exo may help alleviate these detrimental effects. Wild-type, miR-221/222 knockout (KO), and miR-221/222 overexpressing transgenic (TG) mice were administered 10 mg/kg PM through intratracheal injection. After 24 h, the mice underwent left coronary artery ligation for 30 min, followed by 3 h of reperfusion to model I/R injury. We also established an H9c2 cell model under PM (50 µg/mL) and 1% O2 for 6 h, then reoxygenated for 12 h (hypoxia-reoxygenation, H/R). PM exposure exacerbated cardiac injury induced by I/R or H/R by elevating reactive oxygen species (ROS) levels and disrupting mitochondrial function. This dysfunction was associated with an upregulation of mitochondrial fission proteins (Drp1 and MFF) and mitophagy-related proteins (BNIP3 and LC3B), both in in vivo and in vitro experimental models. Administration of ADSC-Exo or miR-221/222 mimics significantly alleviated cardiac injury induced by PM and I/R. Notably, ADSC-Exo, which are enriched with miR-221/222, directly target and suppress the expression of mitophagy and apoptosis regulators such as BNIP3, LC3B, and PUMA. This suppression reduces excessive mitophagy and apoptosis, thereby mitigating cardiac injury. The present data showed that ADSC-Exo treatment regulated mitophagy and cell apoptosis through the miR-221/222/BNIP3/LC3B/PUMA pathway and significantly reduced the cardiac damage by PM+I/R. The present study revealed the novel therapeutic potential of ADSC-Exo in alleviating PM-induced exacerbation of myocardial I/R injury. | en |
| dc.description.provenance | Submitted by admin ntu (admin@lib.ntu.edu.tw) on 2025-02-20T16:29:04Z No. of bitstreams: 0 | en |
| dc.description.provenance | Made available in DSpace on 2025-02-20T16:29:04Z (GMT). No. of bitstreams: 0 | en |
| dc.description.tableofcontents | 目次
口試委員會審定書 i 誌謝 ii 中文摘要 iii Abstract iv Chapter 1. Introduction 1 1.1 Structure and function of the heart 1 1.2 Treatment approaches for myocardial infarction 2 1.2.1 Pharmacological therapy 2 1.2.2 Percutaneous coronary intervention (PCI) 3 1.2.3 Coronary artery bypass grafting (CABG) 4 1.3 Ischemia/Reperfusion injury in cardiovascular disease 4 1.4 Particulate matter (PM) exposure and I/R 5 1.5 The role of reactive oxygen species (ROS) in the interaction of PM exposure and the progression of I/R injury 6 1.6 The role of mitochondrial dynamics and mitophagy in PM exposure with I/R injury 7 1.7 Adipose-derived stem cells (ADSCs)-derived exosomes in tissue repair 9 1.8 The role of miR-221/222 in regulating cardiac apoptosis and mitophagy 11 1.9 Aim 13 Chapter 2. Materials and Methods 14 2.1 Animal experimental procedures 14 2.2 Cell experimental procedures 20 2.3 Measurement of apoptosis 23 2.4 Analysis of ROS productions 26 2.5 Analysis of autolysosomes 28 2.6 Assessment of mitochondrial functions 29 2.7 MicroRNA expression analysis 34 2.8 Statistical analysis 36 Chapter 3. Results 37 3.1 Assessment of cardiac function and injury in PM with I/R treated mice 37 3.2 Histological and apoptotic analysis of PM with I/R treated mice 38 3.3 ROS production, mitochondrial dysfunction, and structural changes in PM with I/R treated mice 38 3.4 Examination of PM and H/R-induced exacerbation of apoptosis 40 3.5 PM enhances H9c2 cell apoptosis by inducing ROS under H/R conditions 41 3.6 PM aggravates apoptosis in H/R-treated H9c2 cells by disrupting mitochondrial function and promoting mitophagy 42 3.7 MiR-221/222-enriched ADSC-Exo alleviates PM+H/R-induced mitophagy and apoptosis 44 3.8 MiR-221/222 regulates apoptosis in PM+H/R-treated cells through the AMPK/p27 pathway 48 3.9 ADSC-Exo and miR-221/222 mimics alleviate mitochondrial dysfunction and apoptosis in PM+H/R-treated H9c2 cells 49 3.10 ADSC-Exo alleviates PM+I/R-induced cardiac injury through miR-221/222 in WT and miR-221/222 KO mice 52 3.11 BNIP3 and LC3B knockdown improves cardiac function and reduces apoptosis in miR-221/222 KO mice exposed to PM+I/R injury 53 3.12 Impact of miR-221/222 overexpression in TG mice on PM+I/R-induced mitophagy, apoptosis and cardiac injury 55 Chapter 4. Discussion 57 4.1. Differences between in vivo and in vitro studies 68 Chapter 5. Conclusion 70 Chapter 6. Limitations and Future works 71 Chapter 7. References 73 Chapter 8. Figures 95 Figure 1. PM reduces cardiac function and increases apoptosis in WT mice during I/R 96 Figure 2. Impact of PM exposure and I/R on cardiac structure and apoptosis in mice. 98 Figure 3. Analysis of intracellular and mitochondrial ROS levels in PM+I/R treated WT mice. 100 Figure 4. Analysis of ultrastructural morphology and protein expression in PM+I/R treated WT mice. 102 Figure 5. PM exposure significantly exacerbates H/R-induced damage in H9c2. 104 Figure 6. Analysis of ROS production in PM+H/R treated cells using fluorescent staining and flow cytometry. 106 Figure 7. MitoTEMPO and NAC mitigate PM+H/R-induced mitochondrial ROS production. 108 Figure 8. MitoTEMPO and NAC alleviate PM+H/R-induced cell apoptosis. 110 Figure 9. Impact of PM and H/R on mitochondrial function in H9c2 cells. 112 Figure 10. Assessment of mitochondrial membrane potential, mitochondrial morphology, and fission protein expression. 114 Figure 11. Analysis of protein expression related to mitophagy and mitochondrial dynamics in PM+H/R-treated H9c2 cells. 116 Figure 12. Autolysosome formation and mitophagy in H9c2 cells treated with PM+H/R. 118 Figure 13. MiR-221/222 expression analysis and interaction with target genes in PM+H/R-treated cells. 120 Figure 14. Regulation of BNIP3 and LC3B expression by miR-221/222 in PM+H/R-treated H9c2 cells. 122 Figure 15. Evaluation of ADSC-Exo on OCR and cell viability in PM+H/R treated H9c2 cells. 124 Figure 16. Apoptosis and protein expression analysis in PM+H/R-treated H9c2 cells following ADSC-Exo and miR-221/222 modulation. 126 Figure 17. Effects of BNIP3, LC3B, and PUMA downregulation on apoptosis in PM+H/R-treated H9c2 cells. 128 Figure 18. Impact of siBNIP3 and siLC3B on PM+H/R-induced apoptosis-related proteins. 130 Figure 19. Effect of PUMA downregulation and miR-221/222 overexpression on apoptosis proteins in PM+H/R-treated H9c2 cells. 132 Figure 20. Involvement of the AKT pathway in PM+H/R-induced apoptosis and the mitigating effects of ADSC-Exo and miR-221/222 mimics. 134 Figure 21. Assessment of the AMPK/p27 pathway in ADSC-Exo-mediated reduction of PM+H/R-induced cell apoptosis. 136 Figure 22. Impact of ADSC-Exo, miR-221/222 mimics, and MitoTEMPO on mitochondrial ROS and function in PM+H/R-treated H9c2 cells. 138 Figure 23. Impact of ADSC-Exo on mitochondrial length and fission protein expression in PM+H/R-treated H9c2 cells. 140 Figure 24. Evaluation of mitophagy following treatment with ADSC-Exo in PM+H/R-treated H9c2 cells. 142 Figure 25. Effects of ADSC-Exo on apoptosis in PM+H/R-treated H9c2 cells. 144 Figure 26. ADSC-Exo decreases PM+I/R-induced cardiac damage in WT, miR-221/222-KO mice. 146 Figure 27. Assessment of mitochondrial and cellular ROS levels in WT and miR-221/222 KO mice using MitoSOX Red and DHE staining. 148 Figure 28. Effect of BNIP3 and LC3B expression in WT and miR-221/222 KO mice under PM+I/R. 150 Figure 29. Knockdown of BNIP3 and LC3B expression improves cardiac function and reduces apoptosis-related proteins. 152 Figure 30. Evaluation of apoptosis in WT and miR-221/222 KO mice treated with ADSC-Exo in PM+I/R. 154 Figure 31. Comparison of cardiac function and ROS in miR-221/222 overexpression TG mice and WT mice following PM and I/R treatment. 156 Figure 32. Assessment of mitophagy- and apoptosis in miR-221/222 overexpression TG mice and WT mice under PM+I/R treatment. 158 Appendix 159 Figure A1. MiR-221/222-expression and characterization of ADSC-Exo. 159 Figure A2. Comparison of ADSC-Exo from WT and miR-221/222 KO mice. 160 Figure A3. Graphical abstract. 163 Prior Publications 164 表次 Table 1. List of primary and secondary antibodies used in Immunohistochemistry. 20 Table 2. List of antibodies used in western blotting. 25 Table 3. List of antibodies used for immunofluorescence staining. 34 | - |
| dc.language.iso | en | - |
| dc.subject | 粒線體自噬 | zh_TW |
| dc.subject | 顆粒物 | zh_TW |
| dc.subject | miR-221/222 | zh_TW |
| dc.subject | 缺血/再灌流損傷 | zh_TW |
| dc.subject | ADSC-Exosome | zh_TW |
| dc.subject | particulate matter | en |
| dc.subject | ADSC-Exosome | en |
| dc.subject | ischemia/reperfusion injury | en |
| dc.subject | miR-221/222 | en |
| dc.subject | mitophagy | en |
| dc.title | 研究脂肪幹細胞外泌體在懸浮微粒和缺血/再灌流引發的心臟損傷中對粒線體功能和細胞凋亡的影響 | zh_TW |
| dc.title | To study the effects of exosomes from adipose-derived stem cells on mitochondrial function and apoptosis in particulate matter and ischemia/reperfusion-induced cardiac injury | en |
| dc.type | Thesis | - |
| dc.date.schoolyear | 113-1 | - |
| dc.description.degree | 博士 | - |
| dc.contributor.oralexamcommittee | 莫凡毅;吳佳慶;蔡亭芬;陳瀅;王淑慧 | zh_TW |
| dc.contributor.oralexamcommittee | Fan-E Mo;Chia-Ching Wu;Ting-Fen Tsai;Ying Chen;Shu-Huei Wang | en |
| dc.subject.keyword | ADSC-Exosome,缺血/再灌流損傷,miR-221/222,粒線體自噬,顆粒物, | zh_TW |
| dc.subject.keyword | ADSC-Exosome,ischemia/reperfusion injury,miR-221/222,mitophagy,particulate matter, | en |
| dc.relation.page | 164 | - |
| dc.identifier.doi | 10.6342/NTU202404484 | - |
| dc.rights.note | 同意授權(限校園內公開) | - |
| dc.date.accepted | 2024-10-22 | - |
| dc.contributor.author-college | 醫學院 | - |
| dc.contributor.author-dept | 解剖學暨細胞生物學研究所 | - |
| dc.date.embargo-lift | 2025-02-21 | - |
| 顯示於系所單位： | 解剖學暨細胞生物學科所 | |
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