Kong Ling-Heng, Gu Xiao-Ming, Wu Feng, Jin Zhen-Xiao, Zhou Jing-Jun
Department of Physiology, Fourth Military Medical University, Xi'an, China; Institute of Basic Medical Science, Xi'an Medical College, Xi'an, China.
Department of Physiology, Fourth Military Medical University, Xi'an, China.
Biochem Biophys Res Commun. 2017 Sep 23;491(3):687-692. doi: 10.1016/j.bbrc.2017.07.128. Epub 2017 Jul 25.
Ca/calmodulin-dependent protein kinase II (CaMKII) has been implicated in myocardial ischemia/reperfusion (IR) injury. The aim of this study was to determine the effect of CaMKII on the damage to membrane skeleton proteins, which is an important cause of IR injury. Isolated rat hearts were subjected to 45-min global ischemia/2-h reperfusion. Both KN-62 and KN-93 were used to inhibit CaMKII. Compared with controls, the hearts in the IR group exhibited remarkable myocardial injury area, LDH release, cell apoptosis and contractile dysfunction, along with an increase in the phosphorylation of CaMKII and its substrate phospholamban. Treatment with either KN-62 or KN-93 mitigated both the heart injury and the phosphorylation of CaMKII and phospholamban. The analysis of cell skeleton proteins revealed that IR injury resulted in an increase in the 150-kDa fragments resulting from the degradation of α-fodrin and dystrophin translocating from the sarcolemmal membrane to the cytosol and a decrease in the 220-kDa isoform of ankyrin-B. As expected, Evans blue dye staining showed an increase in membrane permeability or membrane rupture in the IR group. All of these alterations were alleviated by treatment with either KN-62 or KN-93. In addition, both KN-62 and KN-93 blocked the activity and membrane recruitment of calpain, a key protease responsible for destroying cell skeleton proteins during IR injury. In conclusion, our data provide evidence that damage to membrane skeleton proteins via calpain is a destructive downstream event of CaMKII activation in the setting of myocardial IR injury.
钙/钙调蛋白依赖性蛋白激酶II（CaMKII）与心肌缺血/再灌注（IR）损伤有关。本研究的目的是确定CaMKII对膜骨架蛋白损伤的影响，膜骨架蛋白损伤是IR损伤的一个重要原因。将离体大鼠心脏进行45分钟的全心缺血/2小时再灌注。使用KN-62和KN-93抑制CaMKII。与对照组相比，IR组心脏表现出显著的心肌损伤面积、乳酸脱氢酶释放、细胞凋亡和收缩功能障碍，同时CaMKII及其底物受磷蛋白的磷酸化增加。用KN-62或KN-93处理可减轻心脏损伤以及CaMKII和受磷蛋白的磷酸化。细胞骨架蛋白分析显示，IR损伤导致α-血影蛋白降解产生的150 kDa片段增加，肌营养不良蛋白从肌膜转移到细胞质中，并且锚蛋白B的220 kDa异构体减少。正如预期的那样，伊文思蓝染料染色显示IR组膜通透性增加或膜破裂。用KN-62或KN-93处理可减轻所有这些改变。此外，KN-62和KN-93均阻断了钙蛋白酶的活性及其向膜的募集，钙蛋白酶是IR损伤期间负责破坏细胞骨架蛋白的关键蛋白酶。总之，我们的数据提供了证据，表明在心肌IR损伤情况下，通过钙蛋白酶对膜骨架蛋白的损伤是CaMKII激活的一个破坏性下游事件。
