超音波逆散射總成影像分析心肌組織的臨床意義與細胞生物學應用

Ultrasonic Integrated Backscatter Imaging Characterizing Myocardium:The Clinical Implications and Applications for Cellular Biology

林隆君(Lung-Chun Lin)

指導教授 ： 李源德 ； 謝豐舟

國立臺灣大學/醫學院/臨床醫學研究所/博士(2005年)

https://doi.org/10.6342/NTU.2005.02045

摘要

在冠狀動脈疾病患者中，暈厥心肌和冬眠心肌均表示心肌存活但其收縮功能減弱。暈厥心肌為在解除瞬間缺血情況之後，心肌仍持續一段時間表現較差的收縮力，但自發性逐漸恢復收縮力的病生理學機制。冬眠心肌則表示心肌藉由減退收縮力與能源需求以配合不足的灌流，進而適應慢性局部缺血並得以藉此存活的觀念。如果改善局部缺血，心肌亦可回復其收縮功能。兩者可同時並存。另一方面，近年來研究顯示，冬眠心肌可能經由反覆發作之暈厥心肌造成；另一方面，冬眠心肌並不為一穩定適應狀態。持續灌注不足可能導致心肌細胞內細胞生理的變化，包括收縮纖維的退分化，終至細胞凋亡等不可逆之結果。
心肌組織藉由心肌細胞間低傳導阻抗的細胞間「隙連結通道」形成電氣生理上的協同體，並藉此使得心肌細胞的收縮同步化。在缺血性心臟病的病理機制中，除了個別細胞內的病理變化外，細胞間隙連結通道的再塑形也佔有重要地位。然而，由於適用的活體研究模式很少，也不易於細胞培養中達成，評估細胞間隙連結通道的變化與所引起的機械功能改變相當困難。以超音波逆散射總成影像來描述心肌組織的特性已經證實可以於各種不同心肌病理上顯示心肌的物理性質並度量心肌壁內收縮力。超音波行進於心肌組織中，因心肌收縮與舒張時改變了心肌細胞內容物與細胞外間質纖維結構之音波阻抗反差，造成超音波逆散射能量總成隨心搏週期而變動的曲線；非同步收縮之心肌細胞減低了音波阻抗反差變化的大小而干擾了此心搏週期變動曲線的變動量。因此，吾等可視超音波逆散射總成影像為一種可以研究細胞間聯結功能與所相關的收縮機械協同性之工具。
吾等自從1996年引進能夠收集並處理超音波聲頻原始資料以產生超音波逆散射總成影像的原型機後，隨即針對各種不同心臟疾病研究其心肌超音波逆散射總成的變化。最初吾等以急性心肌梗塞患者為研究對象，證實以超音波逆散射總成影像分析心肌組織所得心搏週期相關曲線之相位權值後超音波逆散射總成變化量可以用來偵測冠狀動脈疾病，以相位角偏移度多寡可以偵測梗塞相關冠狀動脈的再穿通血流TIMI程度。而以超音波逆散射總成影像分析梗塞區域的心肌組織，其相位權值後超音波逆散射總成變化量會隨著心肌存活度不同及是否具有殘存之冠狀動脈狹窄而不同，例如，存活心肌經冠狀動脈介入性治療殘餘血管狹窄後相位權值超音波逆散射變化也增加。配合同步之鉈201心肌灌注掃描及藥物催迫式心臟超音波檢查，於穩定心絞痛慢性冠狀動脈疾病患者中，吾等之研究亦證實心肌潛存缺氧及存活情形均影響超音波逆散射總成心搏週期之循環變動曲線，而權值後振幅的大小可以預測收縮不良處心肌於血管再穿通治療後之恢復情形。
於心肌缺血性病理學中，心肌組織因進行性的細胞損失及逐漸產生的結構變性，以及於心肌細胞間亦因非同步電氣激活收縮而影響壁內收縮力。因此，吾等以慢性冠狀動脈心臟病患者為對象，以心肌因冠狀動脈疾病造成潛存缺氧，接著出現心室壁收縮運動異常之冬眠心肌，進展致心肌受損而降低收縮力儲留為缺氧性心肌病變系列進行之研究模式；以冠狀動脈血管成形術為介入性變因，探討缺氧性心肌病變中各階段，以及改善冠狀動脈血管管徑狹窄及出現再狹窄時，相對應造成心肌組織超音波逆散射總成能量心搏週期變動曲線之改變。結果顯示，雖然於基本休息狀態時，心室壁運動狀似正常，冠狀動脈阻塞所致之心肌潛存缺氧已使得超音波逆散射總成心搏週期變動量減少，而此變化可於冠狀動脈血管成形術成功擴張血管狹窄後二十四小時內恢復。於冬眠心肌中，改善冠狀動脈血管狹窄會同時迅速改善逆散射總成心搏週期變化量，但逆散射總成心搏週期能量曲線之谷底偏移度（相位偏移）與心室壁收縮運動的改善同步但均較延遲出現。冠狀動脈再狹窄造成超音波逆散射總成心搏週期變動量之改變回復至術前缺氧狀態。無收縮力儲留之非存活心肌則不因冠狀動脈血管成形術而改善其超音波逆散射總成心搏週期循環變動。根據吾等臨床研究結果，心肌超音波逆散射總成心搏週期循環變動曲線代表心肌壁內收縮性，而且可以用來評估缺血性心臟病於各種病程中收縮機制之變化。
於臨床及人體組織研究顯示，潛存缺氧的心肌及冬眠心肌中，細胞間「隙連結通道」出現再塑形作用，佔致病機制中的重要地位。吾等嘗試於活體動物實驗中，以超音波逆散射總成心搏週期變動量評估細胞間隙連結通道的變化與所引起收縮機械性的功能變化。過去的觀察研究顯示，高膽固醇飼料餵食之實驗兔，其離體心臟之心肌收縮力變差；而其心室傳導一方面出現電氣生理去極化之不協調性，另一方面也易誘發出心室顫動。此種同時出現電氣及機械性質功能改變之實驗動物表現型，意味著心肌組織細胞間聯結功能可能發生了變化。吾等以高膽固醇飼料餵食之實驗兔為研究模式，探討超音波逆散射總成心搏週期變動曲線與細胞間隙連結通道功能之關聯。結果顯示，高膽固醇飼料餵食組細胞間隙連結通道蛋白Cx43表現量隨餵食時間延長而遞減，改以正常飼料餵食後表現量又恢復。免疫螢光共軛焦顯微鏡影像亦顯示隨著被餵食富含膽固醇飼料的週數增加，實驗兔心肌中細胞間隙連結通道蛋白Cx43的螢光呈色逐漸減少，且由原本分佈於心肌細胞長軸兩端之交接盤結構中再分佈至細胞側緣。在改以正常飼料餵食18週後螢光呈色數量與分佈又恢復。以半定量之反轉錄聚合脢連鎖反應顯示，經高膽固醇飼料餵食的實驗兔心肌細胞隙連結通道蛋白Cx43之訊息者核醣核酸隨著餵食的實驗週數進行而逐漸增加。代表此等蛋白表現量減少乃基於轉錄後機制，與心臟細胞c-Jun NH2-terminal kinases (JNK)之活化增加相關。於心肌收縮功能表現的方面，高膽固醇飼料餵食組在餵食6週後，實驗兔心肌之超音波逆散射總成心搏週期變動量與僧帽瓣外緣收縮期尖峰速度均減少，修正後Tei參數所代表之心肌機械表現變差，餵食9及12週後，此種收縮功能的減弱更加顯著，恢復正常飼料餵食18週後，可以看到超音波逆散射總成心搏週期變動量和修正後Tei參數得到改善。
吾等研究的意義可分為三方面。首先，吾等驗證心肌超音波逆散射總成心搏週期變動曲線代表了心肌壁內收縮性，而且可以用為評估缺血性心臟病於各種病程中收縮機制之變化。其次，吾等應用此超音波技術，使得針對細胞間乃至組織功能變化之研究可於活體動物研究模型中進行。於現今基因轉植技術、後基因體及蛋白體醫學發展的時代，此種針對功能性或表現型的研究愈顯得重要。最後，吾等研究指出超音波聲頻原始資料可提供關於組織機械性質相當豐富的資訊，配合電腦運算處理技術的進步，如組織追蹤影像之發展，吾人得以更加利用影像技術探索生命科學之奧秘。

關鍵字

收縮力 ； 超音波逆散射總成影像 ； 隙連結通道

並列摘要

In patients with coronary artery disease, postischemic dysfunction (stunned myocardium) and chronic hypoperfusion (hibernating myocardium) are two major pathophysiologic states in which viable myocardium demonstrates reduced contractility. It has long been recognized that the working myocardium behaves as an electrical syncytium. Ultrasonic tissue characterization (UTC) with integrated backscatters (IBS) has shown to be a promising approach in detecting various forms of myocardial pathology by delineating the physical properties and intramural contractility. Normal myocardium exhibits cardiac cycle-dependent variation of integrated backscatter that will be blunted in amplitude and have temporal shift in the nadir by experimental ischemia. Reperfusing the acutely infarcted myocardium will restore the cyclic variation before the wall motion recovery that implicates the stunned myocardium. In the setting of hibernating myocardium, some investigations have declared that UTC measures the intramural contractility that is relatively independent of resting wall motion and parallels contractile reserve. Rose et al. validated the simulation model and proposed that, with the microscopic elastic wave theory, the size and interdistance of scatters (myocytes), the impedance mismatching between the myocytes' intracellular contents and the extracellular collagen networks as well as the ultrasonic insonification angle determine the acoustic contrast responsible for the scattering and generate the cardiac cycle-dependent variation.
After introducing the prototype ultrasound machine capable of collecting and processing the raw data of radiofrequency to generate the IBS imaging since 1996, we have been studying the alteration of myocardial ultrasonic backscatters in various cardiac diseases, including detecting multivessel coronary artery disease and TIMI 3 flow of infarct-related artery, evaluating the viability and residual ischemia in the setting of acute myocardial infarction. For the patients having coronary stable angina, we have validated that the results of UTC are influenced by myocardial viability and ischemic burden, indicating the intramural contractility. The inhomogeneous intramural contraction and the degeneration of myocardial structure followed by the loss of viability, implicating the ischemic progress in myocardial pathology, may influence the alterations of UTC by relieving the coronary stenosis. We performed a study to delineate the alterations of myocardial UTC relating to the coronary revascularization and restenosis in patients having chronic coronary heart disease, staging by the development of wall motion abnormality and the absence of contractile reserve and comparable to our previous investigations in cases of acute myocardial infarction, and found that (1) although the baseline wall motion was normal, the myocardium with coronary obstruction might develop intramural contractile dysfunction shown by abnormal modulation in the IBS power curve; (2) relieving the coronary stenosis in viable myocardium would restore its cyclic variations of IBS; (3) the nonviable myocardium had prominent deviation and small weighted amplitude irrelevant to the coronary patency.
We have observed that hypercholesterolemia can induce myocardial electrical remodeling, which was associated with prolonged action potential duration, longer QTc intervals, increased repolarization dispersion, and increased vulnerability to ventricular fibrillation. The dietary hypercholesterolemia has been also shown to induce contractile dysfunction independent of vascular disease, characterized by a decrease in the maximum rate of shortening and relaxation. We also showed that diet-induced hypercholesterolemia in rabbits caused down-regulation of myocardial Cx43 protein and redistribution of Cx43 gap junction, up-regulation of Cx43 transcripts, and activation of JNK. The hypercholesterolemic rabbit hearts, in vivo demonstrated to have systolic dysfunction by using the CVI, mitral ring systolic velocity, and modified Tei index, and suggest that the remodeling of cardiomyocyte gap junctions may contribute to the impaired systolic function. Because gap junctions play a pivotal role in the coordinated excitation of the heart, their remodeling by hypercholesterolemia may partly explore the beneficial effects of lipid-lowering therapy in heart diseases.
There are three important implications by our investigations. First of all, we have validated the clinical application of UTC to access intramural contractile events in various conditions of ischemic heart disease including distinguishing the stunned myocardium, detecting the early successful reperfusion of infarct-related artery and multivessel coronary artery disease for patients with acute myocardial infarction, as well as risk stratification and interventional strategy for the jeopardized infarct myocardium. In the setting of chronic coronary artery disease, the UTC, influenced by myocardial viability and ischemic burden, can be used to predict the functional recovery of dyssynergic myocardium after revascularization and detected coronary restenosis. Secondly, we applied the UTC as a tool for in vivo functional assessment of animal model and found the correlation between remodeling of gap junction and impairment of myocardial contractility in rabbits subjecting to a cholesterol-enriched diet. The study of intercellular contractile synchronism, beyond the intracellular mechanisms that were studied in cultured cells, may be demonstrated via the imaging modality, opening a new horizon in delineating the cellular interaction functionally in tissue preparations, isolated perfused organs or living animals. This progress will be accompanied with the advanced genetic manipulation such as transgenic models or controlled expression in the functional proteonomic era. Finally, there is much progress in the process of radiofrequency raw signals after the fantastically growing capability of computer technique. Our studies has proved the IBS data informative, which will be enriched under the high-frame rate imaging acquisition and serve as the base of speckle recognition and tracking to construct the multi-dimensional myocardial strain imaging instead of those derived from Doppler ultrasound.

並列關鍵字

contractility ； integrated backscatters imaging ； gap junction

參考文獻

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