2012-08-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/646223摘要：雖然心肺復甦急救治療已發展了四十年，突發性心跳停止患者的整體存活率依然偏低，對於此類非預期性死亡的患者，其所造成家庭及社會的影響相當的大，因此如何改善接受心肺復甦急救患者預後的研究，也受到越來越多的重視。心跳停止患者經初步急救成功後，患者的血行動力指數仍呈現相當不穩定的狀態，同時在急救過程中的代謝生理階段所產生的全身性缺血-再灌流的傷害還是持續的進行，這些傷害會引發一連串的惡性循環並加速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)，進而在短期內導致多重器官衰竭及死亡。因此了解心跳停止及心肺復甦急救所引發心肌功能失調的機轉，並從而找出改善急救後患者心臟功能的方式及開發出潛在的治療標的，便成為最重要的課題。近來發現粒線體功能的失常在缺血再灌流的傷害中，佔了非常重要的角色。在缺血再灌流的過程中，細胞內外環境酸鹼值的改變及鈣離子的進出，造成細胞及粒線體的鈣離子過度負載，並導致粒線體通透性轉移孔 (mitochondria permeability transition pore, mPTP)開啟，引發細胞進入壞死及凋亡的狀態。環孢靈(cyclosporine)為近來被發現有抑制mPTP開啟的作用，在心肌局部缺血傷害的動物實驗中，於再灌流時施予cyclosporine，初步發現可減少心肌梗塞區域，產生顯著的保護作用。心跳停止及後續發生之心跳停止症候群，基本上其機轉為全面性的缺血再灌流傷害，其與冠狀動脈阻塞所帶來之局部傷害，其範圍及傷害時間有所不同，而且對細胞及粒線體帶來的傷害及其嚴重程度也未知。Cyclosporine 此直接的mPTP opening抑制劑，對於心跳停止症候群的治療是否有效，而其最佳的有效劑量、及投予時間點等重要議題目前也未明，本計劃之目的即是透過先前建立的心肺復甦急救模式，來探討心跳停止及後續發生之心跳停止症候群對粒線體產生的傷害及cyclosporine治療的效果，並找出最適宜的治療時間點、劑量及相關機轉，以作為將來應用在臨床治療的基礎。本計畫之初步研究結果顯示cyclosporine對於心肌細胞低氧-再給氧的傷害有保護作用，其機轉與粒線體膜電位的維持有直接的關係。在吾人建立的動物模式中發現，心肌細胞中掌管粒線體的呼吸功能的電子傳遞鏈酵素的活性在心跳停止及復甦急救後都減低，以電子顯微鏡觀察，也發現粒線體的內膜會呈現不規則的變化及腫脹的現象，且粒線體mPTP開啟的程度較未發生過心跳停止的控制組明顯且嚴重，代表了心跳停止及復甦急救對粒線體的功能確實產生顯著的影響。進一步應用動物模式檢視cyclosporine是否可改善整體預後及心臟功能。初步發現在開始CPR之前以靜脈注射的方式投予cyclosporine 10mg/kg，其左心室功能及三天存活率較控制組為佳。在本研究計畫中，第一年及第二年的部分吾人將確立窒息引發心跳停止及復甦急救的動物研究模式，來完整的探討cyclosporine治療對心臟功能及整體預後的效果，並嘗試找出其最佳的劑量及投藥時間點之therapeutic window。同時進一步分離粒線體出來，評估心跳停止及復甦急救後，粒線體的mPTP的完整性、電子傳遞鏈Complex I、II、IV的呼吸功能、biogenesis及ATP的產生，粒線體內的過氧化物壓力及蛋白酵素氧化傷害的程度，是否受到影響，並評估cyclosporine的治療究竟是減緩粒線體內哪些部分的傷害，從而找出並確立決定粒線體功能完整性的相關步驟及可能之治療標的，以釐清cyclosporine改善心跳停止後心肌功能失調之機轉，並做為發展未來相關臨床治療的基礎。第三年則是在確認Cyclosporine對心跳停止及復甦急救的治療效果後，我們進一步透過合併如低溫治療等已知有效但不同機轉的治療方式，來嘗試更進一步改善心跳停止症候群的預後。並研究合併治療的狀況下，對整體心肌損傷的保護作用及是否有新的機轉產生，以做為將來發展創新治療的基礎。<br> Abstract: The overall survival is still low for the sudden cardiac arrest patients although the modern cardiopulmonary resuscitation has been developed for more than 40 years. The impacts of sudden cardiac arrest for the patients and their families are huge. After initial successful resuscitation, the hemodynamic status is still unstable in the cardiac arrest patients, the global ischemia-reperfusion injuries keep going in the post-cardiac arrest stage. These injuries will lead to post-cardiac arrest syndrome with multi-organ failure and mortality. Mitochondria dysfunction has been found to play a critical role in ischemia-reperfusion injuries. The intra-cellular acidosis and calcium overloading during reperfusion stage leads to mitochondria injuries, opening of mitochondria permeability transition pores (mPTP) and cell death. It has been found recently that cyclosporine can inhibit mPTP opening in the localized myocardial ischemia-reperfusion injuries. The extent and duration of cardiac arrest and post-cardiac arrest injuries are different from localized myocardial injuries. The severity of mitochondria damage after cardiac arrest injuries and the response to cyclosporine therapy is unclear. The optimal dosage and timing of administrating the cyclosporine are also needed to be elucidated. The aims of the project are to study the mitochondria injuries induced by cardiac arrest and post-cardiac arrest syndrome, the therapeutic effects and optimal treatment protocols of cyclosporine. These results could be the basis for the clinical application in the future.In our prior study, we found the cyclosporine had cardiomyocytes protection effects against hypoxia-reoxygenation injuries through the trans-mitochondria membrane potential stabilizing effects in in vitro study. In animal model, we found the mitochondria respiratory function and electron transport chain complexes activity became decreased, the mitochondria inner membrane became swelling and dys-regulated and the mPTP opening became more significant after cardiac arrest injuries. Our preliminary results showed the cardiac function and 3-day survival were better in the cyclosporine treatment group (10 mg/kg administrated just before starting CPR). In the project, we plan to evaluate and confirm the effects of cyclosporine on the cardiac arrest and post-cardiac arrest injuries. Therapeutic window, including the optimal dosage and timing will be studied. Then we will isolate the mitochondria from the myocardium and to check the mPTP integrity and opening, the mitochondria respiratory function, biogenesis and ATP generation, the oxidative stress within mitochondria in the 1st and 2nd year of the project. The targets responded to the cyclosporine treatment will be evaluated to elucidate the critical pathway for cell survival and cardioprotection. These results can be the basis for the clinical application in the future. In the 3rd year, we will try to combine the cyclosporine treatment with the current available and effective therapy, such as hypothermia therapy for cardiac arrest to improve the cardiac function and overall outcomes further. Whether new mitochondria-related protective effects or mechanisms exist in the combination therapy will be checked.心跳停止心跳停止後症候群粒線體粒線體通透性轉移孔環孢靈Cardiac arrestPost-cardiac arrest syndromeMitochondriaMitochondria permeability transition poreCyclosporine