請用此 Handle URI 來引用此文件:
http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/70458完整後設資料紀錄
| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 陳玉怜(Yuh-Lien Chen) | |
| dc.contributor.author | Yi-Chieh Lee | en |
| dc.contributor.author | 李奕潔 | zh_TW |
| dc.date.accessioned | 2021-06-17T04:28:37Z | - |
| dc.date.available | 2023-09-04 | |
| dc.date.copyright | 2018-09-04 | |
| dc.date.issued | 2018 | |
| dc.date.submitted | 2018-08-13 | |
| dc.identifier.citation | Reference
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/70458 | - |
| dc.description.abstract | 近年來心血管疾病成為已開發國家主要死亡原因之一,其中又以心肌梗塞(myocardial infraction,MI) 擁有極高的發病率以及致死率。早期再灌流是治療心肌梗塞最好的方式,但心臟的缺血再灌流(ischemia/reperfusion,IR) 會造成過量的氧化壓力、粒線體失能和細胞凋亡等途徑,導致術後心肌細胞損傷更加嚴重進而使心臟恢復不良。粒線體的動態平衡 (粒線體分裂和粒線體自噬) 在心臟缺血再灌流中扮演著重要的角色,透過創新的藥物研發或小分子干擾進一步減緩再灌流所造成的傷害更是現今急需解決的問題。文獻指出,維他命D3會去調節心血管的功能,但更詳細的生理調節途徑以及對粒線體動態平衡的影響尚未明瞭。在細胞模式的部分,我們利用缺氧再灌氧(hypoxia/reoxygenation,H/R) 的環境誘導H9c2心肌母細胞,模擬動物缺血再灌流的氧化壓力傷害。H9c2心肌母細胞在H/R的環境下,存活率明顯降低。使用TUNEL染色以及觀察細胞凋亡相關蛋白cleaved caspase 3的活化,結果顯示H/R會誘導細胞凋亡。此外,在H/R的情況下,Mff (mitochondrial fission factor) 會調控粒線體分裂相關蛋白p-Drp1 (dynamin related protein 1),由細胞質轉移到粒線體上並促進粒線體的分裂現象。接著利用西方墨點法觀察粒線體自噬相關蛋白LC3BII/I和BNIP3 (BCL2 Interacting Protein 3) 的活化,吖啶橙螢光染色檢測自噬溶小體 (autolysosome) 的多寡以及透過穿透式電子顯微鏡觀察超微結構,顯示在H/R的情況下,粒線體分裂以及粒線體自噬作用皆有上升的現象,而加入維他命D3後會減緩以上的結果。在動物模式 (I/R) 部分,藉由結紮心臟左前降支冠狀動脈(left anterior descending coronary artery,LAD) 造成心肌缺血30分鐘後,再灌流3小時取其心臟組織做觀察。結果顯示在I/R的情況下,無論是細胞凋亡、粒線體分裂或是粒線體自噬皆會有上升的現象,加入維他命D3後也會有減緩傷害的效果。綜合以上的實驗結果,心臟缺血再灌流的情形下,粒線體分裂以及過量的粒線體自噬會促進細胞死亡,而維他命D3可以透過這條途徑減緩細胞凋亡的現象。 | zh_TW |
| dc.description.abstract | Myocardial infarction is a leading cause of morbidity and mortality worldwide. Early reperfusion is the best strategy for the rescue of ischemic myocardium. However, myocardial reperfusion leads to the excessive production of reactive oxygen species, the activation of apoptotic pathways, and the induction of mitochondrial dysfunction, all of which contribute to postischemic cardiomyocyte death and exacerbate cardiac injury. Mitochondrial dynamics (mitochondrial fission and mitophagy) are crucial to the control of cell survival in cardiac I/R injury. Novel pharmacological or molecular interventions mitigating reperfusion injury, adjunctive to current reperfusion therapies, are in great need. The vitamin D3 regulates the cardiovascular function, but its physiological contribution in the reperfused heart, particularly its influence on mitochondrial homeostasis, is unknown. To mimic myocardial I/R injury, we established a hypoxia/reoxygenation (H/R) model in H9C2 cells in vitro. H/R treatment significantly decreased the cell viability of H9c2 cells by MTT assay. H/R condition also increased apoptosis by TUNEL assay and caspase 3 activation. In addition, H/R treatment increased mitochondrial fission demonstrated by the increased expression of dynamin related protein 1 (p-Drp1) and mitochondrial fission factor (mff) as well as mitochondrial translocation of Drp1. H/R condition triggered excessive mitophagy, as demonstrated by the increase of the expression of LC3BII/I and BNIP3 (BCL2 Interacting Protein 3) by western blot, the number of autolysosomes by acridine orange staining and ultrastructure by transmission electron microscopy. In contrast, vitamin D3 reversed these effects. In I/R mouse model, mice were subjected to 30 minutes of ligating the left anterior descending coronary artery, followed by 3 hours of reperfusion (I/R group). The I/R group showed higher apoptotic cells, mitochondrial fission and mitophagy. Vitamin D3 treatment showed decreasing trends in apoptosis, mitochondrial fission and mitophagy. Based on these findings, the I/R could increase mitochondrial fission and excessive mitophagy to induce cell death while vitamin D3 treatment can attenuate the cell apoptosis via these pathways. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-17T04:28:37Z (GMT). No. of bitstreams: 1 ntu-107-R05446006-1.pdf: 8416570 bytes, checksum: f9f4d6e852051c184a8a62919243cc23 (MD5) Previous issue date: 2018 | en |
| dc.description.tableofcontents | 口試委員審定書 I
致謝 II 中文摘要 V Abstract VII 壹、緒論 1 一、 心臟的構造和功能 1 二、 心肌缺血再灌流傷害(myocardial ischemia/reperfusion injury , I/R injury) 2 三、 粒線體 (mitochondria) 4 四、 心肌缺血再灌流傷害與粒線體分裂 (mitochondrial fission) 的關係 5 五、 細胞自噬 (Autophagy) 7 六、 心肌缺血再灌流傷害與粒線體自噬 (mitophagy) 的關係 9 七、 心肌缺血再灌流傷害與細胞凋亡 (apoptosis) 的關係 11 八、 維他命D3 (vitamin D3) 12 九、 研究動機 16 貳、實驗材料 17 一、 儀器設備 17 二、 實驗材料與試劑 18 三、 實驗用溶液配方 22 參、實驗方法 25 一、 心肌母細胞培養 (H9c2 cell culture) 25 二、 細胞活性分析法 (MTT assay) 25 三、 免疫細胞螢光染色 (immunocytofluorescent staining,ICF staining) 26 四、 Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) 27 五、 西方墨點法 (western blotting) 28 六、 吖啶橙螢光染色 (acridine orange staining,AO) 30 七、 穿透式電子顯微鏡 (transmission electron microscope,TEM) 31 八、 活性氧自由基 (ROS) 的測量 31 九、 粒線體產生ATP能力之測定 32 十、 粒線體膜電位 (mitochondrial membrane potential ) 之測定 32 十一、 粒線體分離技術 (mitochondrial isolation) 33 十二、 細胞凋亡偵測技術 (Annexin V/PI) 33 十三、 粒線體即時影像分析 (time-lapse imaging) 34 十四、 小鼠心肌缺血再灌流模式 35 十五、 組織石蠟包埋 36 十六、 蘇木精-伊紅染色 (haematoxylin-eosin staining) 36 十七、 數據統計分析(statistical analysis) 36 肆、實驗結果 37 一、 探討H9c2心肌母細胞在不同時間H/R的處理下粒線體自噬和細胞凋亡之間的關係。 37 二、 探討H9c2心肌母細胞在不同時間H/R的處理下粒線體功能的變化。 38 三、 探討H9c2心肌母細胞在不同時間H/R的處理下粒線體分裂以及粒線體自噬的機制探討。 38 四、 Vitamin D3(25-OH)治療後減少心肌缺氧再灌氧後ROS生成。 39 五、 Vitamin D3(25-OH)治療後改善心肌缺氧再灌氧後的細胞凋亡傷害。 40 六、 Vitamin D3(25-OH)治療後減緩心肌缺氧再灌氧後造成的粒線體損傷。 40 七、 Vitamin D3(25-OH)治療後減緩心肌缺氧再灌氧後對粒線體不正常分裂所造成的傷害。 41 八、 Vitamin D3(25-OH)治療後減緩心肌缺氧再灌氧後造成的過量粒線體自噬。 41 九、 Vitamin D3(25-OH)治療後減緩Drp1轉移到粒線體上的現象以及粒線體上相關蛋白的表現。 42 十、 Vitamin D3(25-OH)改善心臟缺血再灌流後形態上的傷害。 44 十一、 Vitamin D3(25-OH)改善心臟缺血再灌流後粒線體自噬及細胞凋亡的現象。 44 十二、 Vitamin D3(25-OH)可透過Mff調控的粒線體分裂、BNIP3調控的粒線體自噬進而改善心臟缺血再灌流後的細胞凋亡現象。 45 伍、結論與討論 46 陸、參考文獻 52 柒、附圖 66 | |
| dc.language.iso | zh-TW | |
| dc.subject | 粒線體自噬 | zh_TW |
| dc.subject | 心肌缺血再灌流 | zh_TW |
| dc.subject | 粒線體分裂 | zh_TW |
| dc.subject | 維他命D3 | zh_TW |
| dc.subject | 細胞凋亡 | zh_TW |
| dc.subject | 粒線體失活 | zh_TW |
| dc.subject | Mitophagy | en |
| dc.subject | Apoptosis | en |
| dc.subject | Mitochondrial dysfunction | en |
| dc.subject | Mitochondrial fission | en |
| dc.subject | Cardiac ischemia/reperfusion | en |
| dc.subject | Vitamin D3 | en |
| dc.title | 探討維他命D調控粒線體分裂與粒線體自噬對於心肌缺血再灌流所引發細胞凋亡的影響 | zh_TW |
| dc.title | To study the effects of vitamin D on ischemia/reperfusion-induced cardiac injury via mitochondrial fission and mitophagy | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 106-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 王懷詩,吳佳慶,江美治,方嘉佑 | |
| dc.subject.keyword | 心肌缺血再灌流,粒線體自噬,粒線體分裂,維他命D3,細胞凋亡,粒線體失活, | zh_TW |
| dc.subject.keyword | Cardiac ischemia/reperfusion,Mitophagy,Mitochondrial fission,Vitamin D3,Apoptosis,Mitochondrial dysfunction, | en |
| dc.relation.page | 93 | |
| dc.identifier.doi | 10.6342/NTU201802975 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2018-08-13 | |
| dc.contributor.author-college | 醫學院 | zh_TW |
| dc.contributor.author-dept | 解剖學暨細胞生物學研究所 | zh_TW |
| 顯示於系所單位: | 解剖學暨細胞生物學科所 | |
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