甲一型腎上腺受體阻斷劑與新型強心配糖體對心臟之作用

Abstract

高血壓是國人常見的心血管疾病之一，並常導致一些併發症如冠狀動脈心臟疾病、中風、心室肥大及心衰竭等。而在治療方面，有多種藥物用於治療高血壓，其中包括了甲一型腎上腺受體阻斷劑。 在心血管系統中，甲一型腎上腺受體不但參與正常心臟生理調控血管張力及血壓功能之調節，也在心肌肥大或衰竭及心律不整的病理機轉中扮演相關角色。甲一型腎上腺受體的阻斷劑包含 prazosin、doxazosin、bunazosin以及terazosin等，其廣泛用於降血壓及良性攝護腺肥大症之治療。然而近年來卻有許多臨床上的證據顯示使用doxazosin會增加心血管疾病的風險。其中最具信服力的就屬Antihypertensive and Lipid-Lowering Treatment to Prevent Heart Attack Trial (ALLHAT)這一項臨床試驗，其設計目的為比較四種降血壓藥物對於高血壓病人造成致命性冠狀動脈心臟疾病及非致命性心肌梗塞的機率，結果發現長期使用doxazosin的病人得到這些心血管疾病尤其是鬱血性心衰竭的機率比使用cholorthalidone的病人要高出許多。此外，Vasodilator Heart Failure Trial（V-HeFT）這項臨床研究也發現，使用prazosin比使用其他血管放鬆劑的心衰竭病人具有較高的發病率與死亡率。 由於這些臨床的研究結果，引起了我們想要瞭解甲一型腎上腺受體拮抗劑在冠狀動脈心臟病上所扮演之角色。因此本論文第一部份欲探討甲一型腎上腺受體阻斷劑對於心臟冠狀動脈再灌流損傷的影響。我們在離體大鼠心臟利用Langendorff灌流方式比較四種甲一型腎上腺受體拮抗劑prazosin、 doxazosin、bunazosin、及 terazosin對於離體心臟缺血-再灌流損傷之影響。事先給予此四種甲一型腎上腺受體拮抗劑30分鐘後，將其左冠狀動脈結紮造成心肌局部缺氧30分鐘再鬆開重新恢復血流兩個小時，並測量心肌梗塞區域。結果發現prazosin及doxazosin在心臟重新恢復血流兩個小時後顯著增加了心肌梗塞區域、CK-MB及LDH，然而bunazosin則相反的減少了心肌梗塞區域及與細胞損傷相關之生化因子。另一方面，terazosin對此則無任何顯著影響。儘管此四種甲一型腎上腺受體拮抗劑對於缺血-再灌流的心肌梗塞區域有不同的影響，但對於TUNEL染色結果及caspase-3蛋白質表現量的影響卻有限。因此我們認為四種甲一型腎上腺受體拮抗劑對於缺血-再灌流後心肌梗塞區域之變化來自於其對心肌細胞壞死（necrosis）的影響，而非改變心肌細胞的自主凋亡（apoptosis）。 在第一部份的實驗中，我們意外發現在離體大鼠心臟利用Langendorff灌流方式，事先給予10μmicroM doxazosin後有將近一半的機率會引起心臟在心電圖上的PR時期的延長，造成類似房室結傳導障礙 (AV block) 之偶發性心律不整，進而完全抑制竇性自主節律。然而，在事先給予同樣濃度bunazosin、prazosin及terazosin之離體心臟則不會有相同情形。因此，為了釐清doxazosin造成心律不整的原因，在本論文第二部份中我們利用全細胞膜鉗定法（whole-cell patch clamp）的方式比較doxazosin及bunazosin兩者對於心肌細胞的電生理基本特性。結果顯示，doxazosin在濃度為10μM doxazosin後有將近一半的機率會引起心臟在心電圖上的PR時期的延長，造成類似房室結傳導障礙 (AV block) 之偶發性心律不整，進而完全抑制竇性自主節律。然而，在事先給予同樣濃度bunazosin、prazosin及terazosin之離體心臟則不會有相同情形。因此，為了釐清doxazosin造成心律不整的原因，在本論文第二部份中我們利用全細胞膜鉗定法（whole-cell patch clamp）的方式比較doxazosin及bunazosin兩者對於心肌細胞的電生理基本特性。結果顯示，doxazosin在濃度為10μM的時候會抑制心肌細胞的鈉電流（INa）、L型鈣電流（ICa,L）、瞬時外流鉀電流（Ito）及穩態鉀電流（Iss），但不改變內向整流鉀電流（IK1），而bunazosin對這些電流的影響則僅對L型鈣電流（ICa,L）有約百分之三十之抑制作用。此外，doxazosin也顯著地將鈉電流穩定狀態不活化曲線向左飄移，表示其降低了鈉離子通道的可用率。另一方面，doxazosin延長了單一心肌細胞的動作電位長度並且降低了動作電位強度及去極化速度，而bunazosin則沒有顯著影響動作電位。由以上研究我們推測，doxazosin所引起之心律不整最可能起因於其對房室節組織各部位之鈣離子電流抑制程度不均，進而引起房室節回路傳導。 綜合以上在缺血-再灌流離體心臟及單一心肌細胞電生理特性之結果顯示，長期使用doxazosin的病人比使用其他甲一型腎上腺受體拮抗劑的病人具有較高之心臟疾病風險。根據我們的研究，這四種甲一型腎上腺受體拮抗劑在同一濃度時對於冠狀動脈血流的作用類似，且對於缺血-再灌流之心肌梗塞區域的影響具有濃度相關性，因此我們認為在本實驗模式下，造成doxazosin與其他甲一型腎上腺受體拮抗劑作用不同的原因並非透過其對甲一型腎上腺受體作用之不同，而是另有作用點。然而其中詳細的作用機轉仍須之後更多的實驗證據進一步釐清。 本論文的第三部份則是針對高血壓之併發性心衰竭研發新型的治療藥物，利用各種實驗比較新型強心配糖體AT-11與ouabain之效用。結果證實AT-11無論在離體心臟肌肉或活體天竺鼠上的強心作用及安全性都比ouabain高，且AT-11對於心跳速率的影響也較大。然而，AT-11較高安全性之原因尚待更進一步的研究，其可能與AT-11對於副交感神經之活化及對交感神經之抑制有關。

Hypertension is one of the most common cardiovascular diseases. It results in many complications such as coronary heart disease, stroke, hypertrophy, and heart failure. There are many classes of drugs used in the treatment of hypertension including alpha1-adrenoceptors antagonists. In the cardiovascular system, alpha1-adrenoceptors play important roles not only in the normal physiological regulation of vascular tone and blood pressure, but also in the pathogenesis of cardiac hypertrophy, heart failure, and arrhythmias. Alpha1- adrenoceptors antagonists such as prazosin and its congeners including doxazosin, bunazosin, and terazosin are widely used as antihypertensive and anti-BPH agents. However, there are many clinical observations in recent years showing that using doxazosin may result in higher risk of cardiovascular accidents. The most convincing evidence is the large clinical study, Antihypertensive and Lipid-Lowering Treatment to Prevent Heart Attack Trial (ALLHAT) which was designed to determine whether the incidence of the primary outcome differed among treatments with four types of antihypertensive drugs in hypertensive patients. The conclusion of ALLHAT is that compared with chlorthalidone, long-term use of doxazosin significantly increased the risk of combined cardiovascular disease events, especially congenital heart failure. Therefore, the findings in ALLHAT arouse our interests in the role of alpha1-adrenergic blockers on coronary artery disease. In the first part of this dissertation, we tried to determine the effects of alpha1-adrenoceptors blockers on occlusion-reperfusion injury. Langendorff-perfused rat hearts were pretreated with prazosin, doxazosin, bunazosin, and terazosin and then the left main coronary artery was occluded. After 30 min occlusion, the hearts were reperfusefd for 2 hrs. Two of the compounds studied, prazosin and doxazosin, apparently increased infarct size, CK-MB, and LDH activities after reperfusion. In contrast, bunazosin decreased and terazosin unchanged the infarct size. Although infarct sizes were differently affected by these four blockers, TUNEL-positive nuclei and caspase-3 protein expressions of all groups were not significantly different. Therefore, we supposed that the different effects of these four agents on infarct size came from the difference in necrosis rather than apoptosis. Simultaneously, we also found doxazosin prolonged PR interval and induced occasional arrhythmia followed by complete inhibition of sinus rhythm, whereas bunazosin, prazosin, and terazosin did not. Therefore, in the second part of this dissertation, we continued to compare the electrophysiological profiles between doxazosin and bunazosin. The results of voltage-clamp study showed that doxazosin inhibited INa, ICa,L, Ito, and Iss without changing IK1 but bunazosin only decreased ICa,L by 30% of basal value. Doxazosin also caused markedly negative shift of the INa inactivation curve which reduced INa availability. Besides, doxazosin prolonged action potential duration in association with the decreased action potential amplitude and upstroke velocity, whereas bunazosin did not. As the results shown, we hypothesize that doxazosin-induced arrhythmia may result from the heterogeneous or different level of ICa,L blockade of AV nodal tissue. In conclusion, this present study suggests that bunazosin is safer than doxazosin for long-term treatment in view of the effects on occlusion-reperfusion rat hearts and electrophysiological effect of single cardiomyocyte. According to our data, all four alpha1-adrenoceptor antagonists were found to have similar initial increase of coronary flow, and concentration-dependently affected the infarct size of the reperfused heart. Therefore, we suggest that the different effects between these four agents could be alpha1-adrenoceptor independent and not due to their different potency on the site of action. The precise underlying mechanisms are still needed to be further determined. In the third part of this dissertation, we compared the effects of cardiac glycoside ouabain with a novel agent AT-11. We found that both inotropic effect and safety of AT-11 were better than ouabain in isolated muscle strips and live guinea pigs. The heart rate reduction was more in AT-11. The reason why AT-11 have larger safety index remained to be further determined.