請用此 Handle URI 來引用此文件:
http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/22344
標題: | 後制約訓練對大鼠肝臟缺血再灌流傷害的保護效應與機制 The Protective Effects of Ischemic Postconditioning on Liver Ischemia-reperfusion Injury Among Rats |
作者: | Han-Chen Lin 林含貞 |
指導教授: | 賴逸儒 |
關鍵字: | 缺血,再灌流,粒線體膜通透性,細胞凋亡, ischemia,reperfusion,mitochondrial membrane permeability,apoptosis, |
出版年 : | 2010 |
學位: | 碩士 |
摘要: | 前言:缺血後制約訓練(ischemic postconditioning, iPoC)是改變缺血後的再灌流(reperfusion)程序,以間歇、短暫的方式恢復灌流,而非一般以持續、不間斷的方式恢復灌流,以減少組織所產生的缺血再灌流傷害。本研究用大鼠模式,驗證缺血後制約訓練對肝臟缺血再灌流傷害的保護效果及作用機制。
材料與方法:將雄性Wistar大鼠 (重約160 g-180 g)的肝臟左葉以動脈夾阻斷血流45分鐘(缺血期)後,再移除動脈夾以恢復肝臟灌流240分鐘(再灌流期) ,此為控制(control) 組。缺血後制約訓練(ischemic postconditioning, iPoC)係在恢復灌流前,給予3循環的肝臟短暫的部分缺血再灌流,每循環均包括1分鐘的缺血和1分鐘的再灌流。再灌流240分鐘之後,收集血液樣本以及取下肝臟組織做分析,包括評估血清中GPT值、H&E染色、細胞凋亡評估(TUNEL assay),電子顯微鏡觀察肝臟細胞粒線體結構。實驗分為五個組別,分別是sham組,sham+ATR (Atractyloside)組,control組,iPoC組,iPoC+ATR組。 結果:五組的術前血清GPT無差異,再灌流240分鐘後,iPoC組GPT的上升程度比control組低 (174.0±28.3 U/L v.s 416.3±16.7 U/L,p <0.05);但加入ATR(iPoC+ATR組)後,則GPT的上升程度和control組無差異(557.0±86.7 U/L v.s 416.3±16.7 U/L,p =0.18)。五組的術前TUNEL無差異,再灌流240分鐘後,iPoC組TUNEL-positive的細胞所佔的比例比control組少(44.9±9.9 % v.s 81.1±13.8 %,p<0.05) ,顯示細胞凋亡比例有減少的情形;但加入ATR後,TUNEL-positive的細胞所佔的比例和control組無差異(63.2±4.0 % v.s 81.1±13.8 %,p=0.09)。再灌流240分鐘後,iPoC組細胞質中cytochrome c的表現比control組少(0.1782±0.13 v.s 0.5509±0.14,p<0.05);若是給予ATR處理(iPoC+ATR組),細胞質中cytochrome c的表現和control組沒有差異(0.7562±0.17 v.s 0.5509±0.14,p=0.07)。以上結果顯示缺血後制約訓練可以減少再灌流傷害後GPT的上升程度(細胞傷害) 、細胞凋亡以及粒線體傷害(cytochrome C) ,這些保護效應會因為施予ATR而減少,顯示缺血後制約訓練的保護機制係透過調節粒線體膜通透性減少肝臟的缺血再灌流傷害。 結論: 本研究顯示後制約訓練可以減少由肝臟缺血再灌流所造成的細胞傷害,而且部分是透過調節粒線體膜通透性,減少cytochrome c的釋放來達到保護效果。 Introduction: Ischemic postconditioning (iPoC), a repetitive, brief ischemia- reperfusion maneuver performed at or before the initiation of tissue reperfusion, has been shown to mitigate reperfusion injury in heart and brain. The aim of this study is to investigate the effects of iPoC on liver ischemia- reperfusion injury. Methods: Partial liver ischemia-reperfusion injury is induced by clamping the left lobes of liver for 45 minutes on male Wistar rats (160 g- 180 g). Three cycles of one-minute’s ischemia-reperfusion of the liver, performed by clamping and de-clamping of the liver, are applied before the commencement of reperfusion as the iPoC maneuver. Blood and liver samples are harvested at 240 minutes after reperfusion for end-point assessments which include serum GPT, H&E staining, TUNEL staining, and electron microscopy (EM) study. The results are compared between the sham, sham+ATR(atractyloside), control , postconditioning (iPoC) and postconditioning+ATR (iPoC+ATR) groups. Results: Our data shows that there are no differences in the basal GPT levels of the five groups, but ischemic postconditioning could reduce the elevation of serum GPT level after reperfusion for 240 minutes (174.0±28.3 U/L v.s 416.3±16.7 U/L, p <0.05), and decrease the percentage of apoptotic hepatocytes(44.9±9.9 % v.s 81.1±13.8 %,p<0.05). The co-treatment with iPoC and ATR (iPoC+ATR) could increase the elevation of serum GPT and the number of apoptotic hepatocytes. There are no differences between iPoC+ATR and control groups in serum GPT level after reperfusion for 240 minutes (557.0±86.7 U/L v.s 416.3±16.7 U/L, p=0.18) and the percentage of apoptotic hepatocytes ((63.2±4.0 % v.s 81.1±13.8 %,p=0.09). EM study showed the morphology of mitochondria is more intact after reperfusion when compared to those in control group. Western blot shows increased cytochrome c expression in cytosol portion after reperfusion injury of liver, and postconditioning decreased the expression of cytochrome c after reperfusion. The co-treatment with iPoC and ATR could increase the expression of cytochrome c after reperfusion. Conclusions: This study shows that ischemic postconditioning can attenuate cell deaths after reperfusion injury of liver. The mechanism of protection conferred by postconditioning is related to reduced cytochrome c release, and mediation of mitochondrial permeability transition pore (mPTP). |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/22344 |
全文授權: | 未授權 |
顯示於系所單位: | 解剖學暨細胞生物學科所 |
文件中的檔案:
檔案 | 大小 | 格式 | |
---|---|---|---|
ntu-99-1.pdf 目前未授權公開取用 | 1.31 MB | Adobe PDF |
系統中的文件,除了特別指名其著作權條款之外,均受到著作權保護,並且保留所有的權利。