2017-08-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/645598摘要：原發性皮質醛酮症是一個可以治癒的常見高血壓疾病:它是由於因腎上腺有腫瘤或不正常增生而分泌過多之皮質醛酮所造成。其盛行率目前約占全部高血壓族群的5-10%。原發性皮質醛酮症較原發性高血壓的患者有更高的心血管疾病的罹病率及心房顫動的發生率，而這現象與血壓影響無關；此外，原發性皮質醛酮症患者較原發性高血壓的患者有較厲害的左心室肥厚、左心室舒張功能異常及心肌纖維化之現象，同時血管方面，也會造成血管內膜增厚及血管硬化的情形。鑑於其日漸重要的角色，我們的研究團隊(TAIPAI)在2005 年成立，目的是集合各科的專長多方面的研究此一重要的課題。目前登錄的個案已超過1000 人，為亞洲第一。每年約有50 位新診斷的原發性皮質醛酮症病人，為目前國內可最適合從事原發性皮質醛酮症的研究團體之一。我們根據臨床資料分析發現，原發性皮質醛酮症的血流導介的血管擴張功能(flow-mediateddilatation)及硝基甘油媒介的擴張(nitroglycerin-mediated vascular dilation)均出現問題。血流導介的血管擴張功能失常代表內皮細胞功能異常，原發性皮質醛酮症患者的內皮細胞功能異常已為人所熟知；然而，硝基甘油媒介的擴張失常則代表血管平滑肌的結構及放鬆功能產生了問題。在放鬆功能方面，NO之所以造成平滑肌細胞放鬆，sarcoplasmic reticulum Ca (superscript 2+)-ATPase (SERCA)所媒介的鈣離子回收佔了十分重要的角色。我們的研究成果，發現了皮質醛酮會藉由抑制mitochondrial transcriptionfactor A (TFAM) 及 transcription factor B2, mitochondrial (TFB2M)導致人類血管平滑肌細胞HumanAortic Smooth Muscle Cells (HAOSMC)的SERCA 表現下降(發表於JCEM,2015)。由於我們也發現到皮質醛酮會抑制心肌粒線體的數量，而粒線體的數量與功能和血管放鬆功能也相當有關；因此，皮質醛酮導致人類血管平滑肌異常的機轉中粒線體很有可能也扮演極重要角色。然而，皮質醛酮是否影響血管平滑肌或心肌的粒線體，目前尚無明確資料。在我們初步資料中，皮質醛酮會影響心肌的粒線體的數量及電子傳遞鏈的蛋白表現。此外，皮質醛酮也同時會影響血管平滑肌細胞之粒線體的數量。在本研究中我們將以細胞與動物試驗探討皮質醛酮影響血管平滑肌細胞之粒線體的活性、能量代謝反應的角色與機制進一步探討粒線體治療之應用。本研究之研究策略與方法為:第一年:探討皮質醛酮對血管平滑肌細胞之粒線體的數量及代謝調控的分子機制。主要將以商業購買之初代培養的人類血管平滑肌細胞HAOSMC 進行機制的研究；在給予不同之皮質醛酮相關受體(MR orGR)或訊息傳遞抑制劑的情況下，運用海馬生物能量測定儀(Seahorse XFe extracellular analyzer)比較皮質醛酮對粒線體代謝變化情形，包含ATP 產率; ROS 產生;耗氧率;粒線體代謝途徑(糖解作用、脂肪酸代謝)等之變化。第二年:探討皮質醛酮過度表現的情況下，活體內單位血管平滑肌細胞之粒線體的數目在時間上的變化。主要將以動物模式探討在不同釋放濃度之皮質醛酮皮下釋放錠劑與不同的作用時間之情況下，以粒線體螢光染色與電顯確認血管平滑肌組織細胞之粒線體的數目與型態的表現；並以qRT-PCR 量化粒線體及粒線體相關細胞壓力蛋白質之變化。第三年:探討粒線體治療對皮質醛酮過度表現情況下之血管平滑肌組織的效應。將以皮質醛酮過度表現之動物模式，測試增加粒線體數量;增加ATP;增加抗ROS 能力等粒線體治療方式以qRT-PCR 量化粒線體;及偵測粒線體相關細胞壓力蛋白質之變化之分子機制檢視治療的潛力。原發性皮質醛酮症是重要的臨床問題，期望本研究能釐清粒線體之能量代謝反應在皮質醛酮過度表現情況下，影響血管平滑肌組織的病理角色，並由粒線體治療方式，發展出可能的預防與治療的策略。<br> Abstract: Primary aldosteronism (PA) is a common, curable hypertensive disease with a prevalence of 5-10%in hypertensive population. Compared to essential hypertensive patients, PA patients have higherprevalence of cardiovascular disease and arrhythmia. Therefore, PA becomes a more and more importantissue in recent years. In vascular system, PA patients also have more severe degree of increasedintima-media thickness and arterial stiffness.Due to the important role of PA, our research team (TAIPAI) was assembled in 2005 to do themulti-discipline research on this important issue. There are more than 1000 cases in our data registrywhich is the number 1 in Asia. About 50 PA patients are new-diagnosed or referred to our team each year.TAIPAI is one of the most suitable team to do PA research.According our clinical data, PA patients have impaired vascular function such as flow-mediateddilatation (FMD) and nitroglycerin-mediated vascular dilation (NMD). Impairment of FMD means theendothelium dysfunction, which is already well known in PA patients. However, impairment of NMDrepresents the impairment of vascular relaxation function which is strongly associated with sarcoplasmicreticulum Ca (superscript 2+)-ATPase (SERCA) mediated ca reuptake. In our recent study, we foundaldosterone induced Human Aortic Smooth Muscle Cells SERCA expression decrease viadownregulating mitochondrial transcription factor A and B2. We also found aldosterone decrease theamount of mitochondria in myocardium. The mitochondria amount is also closely associated withvascular relaxation function. Therefore, mitochondria may also play an important role inaldosterone-induced vascular dysfunction. In our preliminary data, aldosterone decrease the amount ofmitochondria in myocardium, and expression of electrical chain protein. Therefore, there is a highpossibility that aldosterone also influences mitochondria amount ad activity in vascular smooth musclecell. In the current study, we will investigate the influence of aldosterone on mitochondria activity,metabolism, and possible utility of mitochondria therapy.First year: Using Human vascular smooth muscle cell model to investigate the detail mechanism ofaldosterone-induced mitochondria change. We will also use Seahorse XFe extracellular analyzer tomeasure the metabolic function of mitochondria, including ATP production rate, ROS production rate,mitochondria metabolic pathway.Second year: Using aldosterone infusion animal model to investigate the influence of aldosteroneon mitochondria in vascular smooth muscle cell. We will use confocal fluoroscopy andelectromicroscopy to confirm the expression of mitochondria amount and morphology. We will alsoquantify the mitochondria protein by qRT-PCR.Third year: We will try to use mitochondria therapy to rescue the influence of aldosterone onmitochondria. We will try several methods including increase the mitochondria amount, increase ATP,increased anti-ROS.