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Title: | 藉由李宋小型豬探討粒線體於肥胖心肌病變中之影響 The role of mitochondria involved in obesity cardiomyopathy in the Lee-Sung minipig model |
Authors: | Sin-Jin Li 李欣瑾 |
Advisor: | 陳靜宜(Ching-Yi Chen) |
Co-Advisor: | 丁詩同(Shih-Torng Ding) |
Keyword: | 李宋豬,肥胖心肌病變,粒線體動態平衡,DRP1,心包脂肪,內質網壓力,細胞自嗜, Lee-Sung miniature pigs,obesity cardiomyopathy,mitochondrial dynamics,DRP1,pericardial fat,ER stress,autophagy, |
Publication Year : | 2019 |
Degree: | 博士 |
Abstract: | 根據世界衛生組織調查,心臟疾病盤踞著全球十大死因第一位長達十五年之久。因心臟為一高度氧化代謝之器官,故粒線體對於維持心臟正常功能扮演著重要的角色。近來粒線體動態平衡 (mitochondrial dynamics)常與心血管疾病共同探討。然而,粒線體動態平衡於肥胖心肌病變 (obesity cardiomyopathy)致病機轉中所扮演之角色尚未釐清。於本試驗中,我們建立藉由高脂飼糧之餵飼誘導李宋小型豬產生肥胖誘發之心肌病變。這些肥胖心肌病變豬隻,具有較重之體重、異位脂肪堆積且具代謝症候群之情況。內質網壓力 (endoplasmic reticulum stress)、細胞自嗜 (autophagy)及脂毒性 (lipotoxicity)參與了豬隻肥胖心肌病變之進程。此外,更發現心肌病變豬隻其心臟中之腺苷三磷酸 (ATP)較低。因此,本篇研究進一步藉由體外及體內試驗,以釐清粒線體動態平衡於肥胖心肌病變中之角色。
我們發現肥胖心肌病變豬隻,具心臟氧化壓力、損傷粒線體生合成、動態平衡失衡及誘導粒線體自嗜。為了進一步釐清粒線體動態平衡於高脂飼糧誘發心肌病變中之機制,棕櫚酸處理予H9C2細胞以誘發脂毒性。在體外模式中,棕櫚酸破壞粒線體動態平衡並造成細胞死亡,而抑制粒線體分裂 (fission,抑制DRP1表現)之情況下,於最初可維持粒線體功能並增加細胞存活率。然而,當延長棕櫚酸處理後,不論siDRP1功能存在與否,基礎粒線體氧化功能皆下降。同時,我們也發現,心臟周邊具有較多之心包脂肪堆積 (pericardial adipose tissue)。肥胖豬隻之心包脂肪具有較高量之IL-6及丙二醛 (malondialdehyde)。 為了進一步了解,是否心包脂肪分泌物質對於肥胖心肌病變中粒線體具有局部調節作用,H9C2細胞將給予心包脂肪之條件培養基 (conditioned medium)進行處理。結果顯示,處理心包脂肪條件培養基之組別,其粒線體呼吸作用及ATP產量被抑制,因此造成H9C2之細胞凋亡反應。而粒線體動態平衡或粒線體自嗜作用之相關蛋白質表現皆下降。 綜上所述,此結果指出粒線體動態平衡參與高脂飼糧誘導心肌纖維化之進程。體外模式證實,循環性影響 (棕櫚酸)及局部調節 (心包脂肪分泌物質)皆參與高脂飼糧造成之心肌病變。棕櫚酸及心包脂肪分泌物質兩者皆造成粒線體功能損傷及誘導細胞死亡,而抑制過多粒線體分裂作用於脂毒性初期時,可給予細胞延長壽命之優勢,但當延長棕櫚酸處理時,並無法恢復粒線體功能。儘管有其侷限性,但我們奠定了具潛力的新治療策略的關鍵階段,藉由siDRP1而降低肥胖心肌病變之風險。 According to the World Health Organization, heart diseases remained in the top rank of ten leading causes of death globally in the last 15 years. The heart is a highly oxidative tissue and mitochondria play a critical role in maintaining optimal cardiac function. Recently, mitochondrial dynamics have been connected with cardiovascular diseases (CVD). However, the exact role of mitochondrial dynamics in the pathogenesis of obesity cardiomyopathy (OCM) remains unclear. In present study, we established an OCM minipig model by high-fat diet (HFD) feeding for 6 months. These OCM pigs had a heavier body mass, accumulated more ectopic fat, and exhibited metabolic syndrome. The endoplasmic reticulum stress, autophagy, and lipotoxicity were participated in the cardiac pathological mechanism of OCM pigs. Moreover, a decreasing cardiac ATP production was observed in these OCM pigs. Therefore, the aim of this study was to elucidate the role of mitochondrial dynamics in OCM, in vitro and in vivo. We found that enhanced cardiac oxidative stress, impairment of mitochondrial biogenesis and dynamics, and induced mitophagy were involved in the OCM pigs. To further elucidate the mechanisms of mitochondrial dynamics involved in HFD-induced cardiomyopathy, palmitate was used to induce lipotoxicity in H9C2 cells. In the cell model, palmitate disrupted mitochondrial dynamics and induced cell death, whereas inhibition of mitochondrial fission (DRP1) at the onset of lipotoxicity maintained the mitochondrial function and cell survival. However, there was lower basal mitochondrial oxidative function after prolonged palmitate treatment regardless of the functionality of siDRP1 was present or not. Meanwhile, there was more pericardial adipose tissue (PAT) accumulation around the heart. An elevated content of IL-6 and malondialdehyde was found in the PAT of obese pigs. To examine whether local effect of PAT secretomes regulated the mitochondrial function in OCM, H9C2 cells were treated with PAT-conditioned medium (CM). PAT-CM inhibited basal mitochondrial respiration and ATP production, thus leading to apoptosis of H9C2 cells. The protein expressions of mitochondrial dynamics- and a mitophagy-related protein were suppressed by PAT-CM. In conclusion, the results indicated that mitochondrial dynamics was involved in the progression of HFD-induced cardiac pathogenesis. The model in vitro demonstrated that HFD caused cardiomyopathy via systemic effect (palmitate) and local regulation (PAT secretomes). Both palmitate and PAT secretomes disrupted mitochondrial functions and induced cell death, whereas inhibiting excessive mitochondrial fission at the onset of lipotoxicity provided a prolonged survival advantage, but did not restore mitochondrial function after prolonged lipotoxicity by palmitate. Despite the limitation, we showed a critical stage for potential new therapeutic strategies to reduce the risk of OCM via siDRP1. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/72679 |
DOI: | 10.6342/NTU201902107 |
Fulltext Rights: | 有償授權 |
Appears in Collections: | 動物科學技術學系 |
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