大鼠肝脏冷/热缺血再灌注损伤钠钾ATP酶、钙ATP酶及镁ATP酶与肝细胞死亡方式的研究

The effect of cold/warm ischemia reperfusion injury on cell death pattern and the activities of Na^+-K^+ATPase , Mg^(2+)-ATPase and Ca^(2+)ATPase in liver of rats

目的 :研究肝脏冷 /热血再灌注损伤条件下 ,钠钾ATP酶、钙ATPATP酶及镁ATP酶活性与细胞凋亡和胀亡的关系 ,并探讨减少此类损伤的途径。方法 :建立大鼠肝门部分阻断热缺血 1h再灌注模型及大鼠原位肝移植冷缺血 1h再灌注模型 ,分别于再灌注 0h、1h、12h和 72h处死取材 ,测定肝细胞钠钾ATP酶、钙ATP酶及镁ATP酶活性。用形态学及AnnexinV、PI双染法流式细胞仪定量测定早期细胞凋亡和胀亡百分数。结果 :与对照组相比 ,冷 /热缺血 1h(再灌注 0h)后钠钾ATP酶、钙ATP酶及镁ATP酶活性均持续显著下降 ,再灌注 1h达低谷。但热缺血再灌注后 72h钠钾ATP酶活性恢复 ,钙ATP酶及镁ATP酶活性仍未恢复。冷缺血再灌注后 12h钙ATP酶先恢复活性 ,72h钠钾ATP酶、镁ATP酶活性才恢复。细胞死亡方式 :①冷缺血再灌注损伤 ,细胞死亡以凋亡为主 ,并于再灌注后 12h后到顶峰。②热缺血 1h胀亡细胞显著增多 ,再灌注 1h后减少 ,细胞死亡方式转为以凋亡为主。结论 :肝脏冷 /热缺血再灌注损伤均能导致 3种ATP酶活性下降 ,细胞内离子浓度失衡 ,热缺血期ATP酶活性严重下降 ,细胞死亡 ,以胀亡为主。冷缺血期ATP酶轻度下降 ,细胞死亡以凋亡为主。术前或术中给予能量物质 ,对提高钠钾ATP酶、钙ATP酶及镁ATP酶活性 ,减轻炎症反应 ,延长肝门阻断?

Objective:To seek effective way to reduce ischemia / reperfusion(I/R) injury, we investigate the relation between the activities of ion channel such as Na +-K +ATPase, Ca 2+ATPase and Mg 2+ATPase and the percentage of apoptosis hepatocytes and oncosis hepatocytes in rat liver following cold/warm ischemia / reperfusion.Methods:Models of warm ischemia/reperfusion of partial liver vein and hepatic artery of ischemia, cold ischemia/reperfusion of orthotopic liver transplantation were made. The activities of Na +-K +ATPase, Ca 2+ATPase and Mg 2+ATPase were examined at 1h, 12h and 72h following reperfusion, and at 1h of ischemia. And the percentage of apoptosis hepatocytes and oncosis hepatocytes were observed with method of morphology and Annexin V, PI stain.Results:In groups of warm ischemia , the activities of ATPases reduced immediately following 1h of ischemia, felling into the lowest at 1h of reperfusion. And oncosis was the dominant death model of hepatocytes during the ischemia phase of 1h. After reperfusion, the activities of Na +-K +ATPase increased and the death model of hepatocytes conversed into apoptosis, but the activities of Ca 2+ATPase and Mg 2+ATPase were still on low level after 72h of reperfusion. In cold ischemia groups, the activities of ATPases decreased continually, felling into lowest at 1h of reperfusion. And apoptosis was the dominant death model of hepatocytes during the ischemia and reperfusion phase . The activities of ATPase increased after reperfusion, but the recovery of Ca 2+ATPase was faster than that of Na +-K +ATPase and Mg 2+ATPase. Conclusion:Both cold I/R injury and warm R/I injury can make the activities of ATPase decrease, leading to imbalance of ionic concentration in cell, causing oncosis in period of warm ischemia and apoptosis in the period of cold ischemia. It would be useful to improve the activities of Na +-K +ATPase, Ca 2+ATPase and Mg 2+ATPase for reducing the I/R injury, if the energy was addministrated before operation or during operation.