- 學位論文
骨骼肌遠端缺血前置處理對心肌之保護--自由基之啟動者角色
Skeletal Remote Ischemic Preconditioning in Myocardial Protection--free radical as a trigger
摘要
缺血性前置處理(IPC)在各種動物實驗研究探討已超過10年，而且除了心臟也在以外器的官發現此現象。器官內之“遠端前置處理”(RPC)的概念也形成，可用來保護心肌。而後，器官內遠端前置處理概念也推展到器官間遠端前置處理。骨骼肌遠端前置處理似乎是一個臨床上可行之心肌保護方法，但是仍有些爭論存在，而且沒有進一步研究來探討其作用機轉。此研究之目的便是在探討骨骼肌遠端前置處理對心肌梗塞之作用與自由基所扮演的角色。 首先，骨骼肌前置處理之設計是對大鼠之單側股動脈行四次十分鐘缺血與十分鐘之再灌流。實驗分成四組:第一組(I):模擬組，第二組(Ⅱ):只做骨骼肌前置處理，第三組(Ⅲ):只有心肌梗塞，第四組(IV): 骨骼肌前置處理，2小時後再行心肌梗塞。以化學冷光分析儀來分析在行骨骼肌前置處理後，自由基的變化，發現在四次缺血/再灌流所產生之自由基最明顯，2小時後最高，4小時後便下降。如果在骨骼肌前置處理期間，給予MPG之注射，則發現自由基增加的現象會消失。此結果可以幫助往後實驗釐清自由基角色上的判讀。 我們首先以一種特殊染色來區別梗塞區域與存活區域，由此來做解剖學上之研究。其梗塞面積(infarct size)在第四組明顯比第三組少(24.7±8.8% in IV vs 51.4±9.1% in III, p < 0.01)。且此現象可以被MPG前處理所抑制(49.2±6.3% in MPG+III vs. 50.1±8.2% in MPG+IV, p > 0.05)。我們測定心肌酵素之釋放來校正之，發現CK-MB與TnI在第四組的釋放量比第三組來的少;同時，其保護效果也可以被MPG所拮抗。用西方漬墨法來探討熱休克蛋白質及抗氧化酵素在心肌中之變化，發現在第Ⅱ、Ⅲ、IV組可以看到熱休克蛋白質及抗氧化酵素(Mn-SOD與GPx)的增加。不過，使用MPG前處理，只看到熱休克蛋白質的下降，而抗氧化酵素則沒有變化。此外，由危險區域與非危險區域蛋白質之分佈，骨骼肌前置處理可能經由神經系統傳遞信息。 大鼠骨骼肌前置處理可以在梗塞模式產生心肌保護作用，其作用會經自由基啟動，而且和熱休克蛋白質與抗氧化酵素有關。
並列摘要
Ischemic preconditioning (IPC) has been widely explored in various experimental models for more than 10 years and it has been also observed in several organs other than the heart. The concept of intraorgan “remote preconditioning” (RPC) has been previously advocated in heart to reduce the infarct size. The concept of intraorgan RPC had beem extended to the interorgan RPC. Skeletal RPC seemed to be a good way for myocardial protection, but some controversial data still exists and no further mechanism was investigated. The aim of this thesis was to investigate the effect of skeletal ischemia/reperfusion on myocardial infarction and the role of free radicals. First, skeletal RPC was designed in rats by repeated 4-cycle 10-min ischemia-reperfusion of femoral artery. Four experimental groups were included: I, sham group; II, RPC only; III, infarction only; IV, incorporating both RPC and infarction. Chemiluminescence study showed significant elevation of free radical in groups II and IV, and pretreated mercaptopropionyl-glycine (MPG), a free radical scavenger, could abolish the production of free radicals. The result would help to delinate the role of free radicals in RPC. The infarct size was significantly reduced in group IV (24.7±8.8%) compared to group III (51.4±9.1%) (p < 0.001), and this effect was abolished by MPG pretreatment (49.2±6.3% in MPG+III vs. 50.1±8.2% in MPG+IV, p > 0.05). Cardiac enzymes also revealed significant decrease in group IV compared to group III, and the protective effect could be abolished by MPG. Western blotting of heat shock protein (HSP) revealed that consistent elevation of HSP 25 and 70 in group II, III and IV, and the elevation can be abrogated by MPG pretreatment. The expression of the antioxidant enzymes, Mn-superoxidase dismutase and glutathione peroxidase, in the area of risk were consistently elevated in groups II, III, and IV similar to HSP. However, the MPG pretreatment could not decrease the expression of the antioxidant enzymes as HSP. The skeletal RPC in rats can produce a protective effect in an infarction model that may be triggered through free radical pathway, and this protective effect was associated with heat shock protein and antioxidant enzymes.