Abstract
摘要：近年來，致力於改善急救現場效率的研究大大改善心肺復甦術的急救成功率。然而，在心跳停止的急救過程中，急救所產生的傷害以及所引起一連串的連鎖反應，會造成急救成功後心臟血管及腦部功能的惡化，引發心跳停止後症候群(post-cardiac arrest syndrome)，導致短期內導致多重器官衰竭及死亡。目前，在恢復自主循環後的給予攝氏32~34℃為時12到24小時的低溫治療已被證實可以改善心室顫動的到院前心跳停止病患的神經學預後與存活率。雖然，對於低溫治療的可能作用機轉與訊息傳遞路徑之研究蓬勃發展，然而這些研究卻發現這些作用機轉與訊息傳遞路徑數目眾多而複雜。微型核糖核酸(microRNA)在生物體調控基因表現或抑制基因表現，進而影響細胞發育、增生、分化、凋零等相關蛋白的表現。研究顯示許多疾病都與微型核糖核酸的改變有關。是否能夠藉由低溫治療所造成的微型核糖核酸與其特定蛋白質表現的改變，來進一步了解低溫治療的作用機轉，是本研究的目標之一。另外，微型核糖核酸變化能否作為預測心跳停止病患預後之可能生物血清標記(biomarkers)，也值得吾人更深入的研究與探討。本研究團隊已於第一年的研究計畫內，利用已建立之心室顫動心跳停止的動物研究模型，來研究復甦急救成功後，實驗動物在分別接受復甦後低溫治療與常溫治療的情況之下的大腦、心肌組織、白血球及血漿中微型核糖核酸的表現情形。再分別利用神經細胞與心肌細胞細胞株(cell line)與原代培養(primary culture) 進行低溫與常溫的缺氧再灌流傷害，然後使用Real-time PCR 將微型核糖核酸的表現做定量之分析。比對大腦組織與神經細胞及心肌組織與心肌細胞的微型核糖核酸的表現，本研究團隊分別選定了數個目標微型核糖核酸，將進一步將細胞內的目標微型核糖核酸分別進行轉殖(transfection)與抑制(knock down)，再進行缺氧再灌流傷害，藉由細胞的存活與蛋白質的表現來評估目標微型核糖核酸在低溫治療中所扮演的角色。此外，本研究團隊亦將根據細胞研究的成果來進行成鼠的目標微型核糖核酸抑制試驗，然後誘發心室顫動心跳停止再施以急救，評估目標微型核糖核酸在活體生物中是否有一致的作用，並研究與低溫治療相關的微型核糖核酸其上下游之訊息傳遞路徑。本研究團隊亦將藉由研究大腦與心肌組織的組織學傷害範圍以及特點、與包含微型核糖核酸與大腦與心肌組織特異性之血清標記來評估大腦與心肌受損程度及預後，進一步探討及確認微型核糖核酸在活體動物體內參與低溫保護作用的重要性與影響範圍。最後，本研究團隊收集臨床上心跳停止經急救復甦成功之後低溫與常溫治療病患之血液檢體，加以檢測根據動物實驗研究結果所選定之微型核糖核酸與其特定蛋白質的表現情形。這些選定之微型核糖核酸與其特定蛋白質可做為進一步復甦後低溫研究之標的。本研究團隊亦將利用已建立之收集心跳停止及心肺復甦急救患者之資料庫模式，收集心跳停止經心肺復甦急救患者之臨床資料與血液檢體，測定包含目標微型核糖核酸與其特定蛋白質等之多項重要血清生物標記，並進一步分析這些標記與患者各項預後的關連性，作為將來應用並改善此類患者臨床醫療及照護的基礎，以提升復甦醫療之水準。
Abstract: In recent years, the researches devoted in improving the efficiency of resuscitation at the sceneimprove the successful rate of cardiopulmonary resuscitation (CPR). However, theischemia/reperfusion injury during CPR and the following vicious circle deteriorate the cardiovascularand cerebral functions, and resulted in post-cardiac arrest syndrome and multiple organ failure, andthus death. Hypothermia of 32~34℃ for 12-24 hours after successful resuscitation has been proved toimprove the neurological outcome and survival in ventricular fibrillation (VF) cardiac arrest. Manyresearches studying therapeutic hypothermia find the complicated mechanisms and pathways involvedin hypothermia therapy.The microRNA affects the development, proliferation, differentiation and apoptosis of cell bymodifying or inhibiting the gene expression. Several studies demonstrate that many diseases areassociated with the microRNA abnormality. One of the objects in the current study is to evaluate themechanisms of therapeutic hypothermia by investigating the changes of microRNAs and associatedprotein during cooling. Besides, whether the microRNA can be used as a prognostic biomarker inpredicting outcomes in cardiac arrest survivors also needs further studies.In the first year of the study, we have extracted and analyzed the microRNAs from the brain, heart,white blood cell and plasma of animals respectively receiving normothermia and hypothermia afterVF cardiac arrest. We also applied ischemia-reperfusion injury to the cell line and primary culture ofneuron and cardiomyocyte under normothermia and hypothermia, respectively, and then analyzed themicroRNA of the cells by using real-time PCR. After comparing the microRNA expression betweenthe neuron and brain tissue and between the cardiomyocyte and myocardium, several target microRNAhave been chosen to undergo transfection and knock down experiments of cells. The cells aftertransfection and knock down experiments will be applied ischemia-reperfusion injury. The role ofmicroRNA in therapeutic hypothermia will be evaluated by analyzing the survival of cell andassociated protein expression. Besides, we will also knock down the target microRNA in adult ratbased on the results of cell culture, and then induced VF cardiac arrest and applied resuscitation. Therole of microRNA in therapeutic hypothermia and the reciprocal upstream and downstream pathwayswill be investigated. The histological characteristics and specific biomarkers of brain and heart willalso be examined to evaluate the damage.Finally, we will collect the clinical data and blood from cardiac arrest survivors with establishedformat, and measure the important biomarkers including microRNAs selected based on the animalstudy. The association between the biomarkers and the outcomes of the cardiac arrest survivors will beanalyzed to provide the important information in monitoring the therapy effect and predicting theoutcomes in cardiac arrest.
Keyword(s)
心跳停止
低溫治療
微型核糖核酸
生物血清標記
大腦
心肌組織
白血球
血漿
Cardiac arrest
therapeutic hypothermia
microRNA
biomarker
brain
heart
white
blood cell
plasma
