p38α/β和ERK1/2在心肌缺氧预处理信号传递机制中的不同作用
黄轶峰, 龚开政, 张振刚*
扬州大学第二临床学院心血管内科, 扬州225001
摘要: 建立培养乳鼠心肌细胞的缺氧/复氧(A/R)损伤模型和缺氧预处理(APC)模型, 以细胞存活率、细胞内超氧化物趋化酶(SOD)活性、丙二醛(MDA)含量、培养上清液乳酸脱氢酶(LDH)活性作为反映心肌细胞损伤的指标。采用ERK1/2抑制剂PD98059及p38α/β 阻滞剂SB203580干预模型, 并以胶内原位磷酸化法测定ERK1/2和p38 活性, 借以探讨ERK1/2和p38α/β 在缺氧预处理(APC)保护机制中的作用。结果表明, (1)在APC组, 于预处理的缺氧时相给予PD98059, 可以完全消除APC的延迟保护作用; 在A/R组的缺氧时相加入PD98059对细胞损伤无影响。(2)在APC组的预处理缺氧时相给予p38α/β 抑制剂SB203580并不能消除APC的保护作用, 而在A/R组的持续缺氧时相给予SB203580则可显著减轻缺氧对细胞的损伤。(3) ERK1/2和p38总活性测定表明, 缺氧可激活ERK1/2和p38, 它们的活性在缺氧后4 h时达到高峰, 而经过APC处理后, 二者活性高峰提前于3 h时出现, 且峰值显著降低。上述结果提示, 预处理过程中ERK1/2的激活可能是缺氧预处理延迟保护机制中细胞信号传递的重要环节, 预处理阶段p38α/β 的活化不参与APC诱导的延迟保护信号传递过程, p38 的过度激活可能是缺氧/复氧损伤过程中的一个致损伤参与因素, 而预处理抑制随后持续缺氧阶段p38的过度激活可能是其保护机制的一个环节。
关键词: 生理学; 心肌保护; 缺氧; 心肌缺血预处理; 丝裂素活化蛋白激酶p38; 细胞外信号调节蛋白激酶; 大鼠
中图分类号: R540.2
The different
roles of ERK1/2 and p38 MAPKα/β in cell signaling of cardiomyocyte anoxia
preconditioning
HUANG Yi-Feng, GONG Kai-Zheng, ZHANG Zhen-Gang*
Department
of Cardiology, The 2nd Clinical
225001
Abstract: Preconditioning (PC) exhibits earlier and delayed protection. But the mechanism of cellular signaling in delayed protection of PC remains unclear. We explored the roles of ERK1/2 and p38
MAPKα/β in delayed protection of anoxia preconditioning (APC). The anoxia/reoxygenation (A/R) injury and APC models were established in cultured neonatal rat cardiomyocytes. An ERK1/2 inhibitor (PD98059) and a p38α/β blocker (SB203580) were applied and their effects on A/R and APC models were observed. The cellular contents of MDA, SOD, cell viability and LDH release was measured at the end of the study. ERK1/2 and p38 MAPK total activity was measured by “in-gel” myelin basic protein phosphorylation assay at different point during sustained anoxia. The results obtained are as follows: (1) PD98059 (but not SB203580), administered in preconditioning anoxia phase in APC group, abolished completely the delayed protection of APC. (2) SB203580 administered in sustained anoxia phase in A/R group could relieve cell injury induced by anoxia, but not PD98059. (3) The highest activity of ERK1/2 and p38 MAPK induced by anoxia appeared at 4 hour after the beginning of sustained anoxia. APC inhibited the over activation of both ERK1/2 and p38 during the following sustained anoxia. These results suggest that ERK1/2 activation during preconditioning may be an important link of cell signal transduction in the mechanism of APC delayed protection. p38α/β activation in preconditioning stage dose not participate in signaling of APC delayed protection. The excessive activation of p38α/β is possibly a Key factor of mediating cell injury induced by sustained anoxia. The inhibition of p38α/β excessive activation during subsequent sustained anoxia might play a role in delayed protection mechanism of APC.
Key words: physiology; cytopreotection; anoxia; ischemic preconditioning, myocardial; extracellular
signal-regulated kinase; p38 mitogen-activated protein kinases