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Updated: Aug 25, 2026
Improved Rodent Model of Myocardial Ischemia and Reperfusion Injury
Published on: March 7, 2022
ROS-mediated ERK activation in delayed protection from anoxic preconditioning in neonatal rat cardiomyocytes
Kai-zheng Gong1, Zhen-gang Zhang, Ai-hua Li
1Department of Cardiology, First People's Hospital of Yangzhou, Yangzhou 225001, China. yungkzh@126.com
Background:
The activation of extracellular signal-regulated kinase1/2 (ERK1/2) has been shown to be important signaling pathway in the ischemic preconditioning (IPC) response. Recently, some studies suggest a key role for the mitochondrial ATP-sensitive potassium channel (mKATP) as both a trigger and an end effector of acute and delayed protection of IPC. Hence, this study was undertaken to elucidate the relationship between mKATP and ERK1/2 in the delayed protection mechanism of anoxic preconditioning (APC).
Methods:
An APC model was established using cultured neonatal rat cardiomyocytes. Pharmacological agents [diazoxide, 5-hydroxydecanoate (5-HD), 2-mercaptopropionylglycine (MPG), and PD98059] were used to modulate mKATP and ERK1/2 activation. Cellular injury was evaluated by measuring cellular superoxide dismutase (SOD) activity, cell viability, and lactate dehydrogenase (LDH) release. The generation of cellular reactive oxygen species (ROS) and the activation of ERK1/2 were determined at different time points starting from the beginning of preconditioning with anoxia or diazoxide (an mKATP opener).
Results:
Cell viability and SOD activity in the APC [(81.9 +/- 11.4)%, (13.6 +/- 3.7) U/L] and diazoxide [(79.2 +/- 12.4)%, (16.5 +/- 4.6) U/L] groups were significantly higher than in the anoxia/reoxygenation (A/R) [(42.2 +/- 7.3)%, (8.8 +/- 2.8) U/L] group (all P < 0.01). LDH activity in the APC group [(101.9 +/- 18.9) U/L] and diazoxide group [(97.5 +/- 17.7) U/L] was significantly lower than in the A/R group [(250.5 +/- 43.6) U/L] (all P < 0.01). Both APC and diazoxide simultaneously facilitated intracellular ROS generation and rapid ERK1/2 activation. But the effects of APC and diazoxide were remarkedly attenuated by 5-HP (an mKATP blocker) and by MPG (a free radical scavenger). In addition, the ERK1/2 inhibitor PD98059 also abolished the cellular protective effects induced by diazoxide.
Conclusion:
mKATP may mediate ERK1/2 activation during anoxia preconditioning by generating ROS, which then triggers the delayed protection of APC in rat cardiomyocytes.
Insights
Mitochondrial ATP-sensitive potassium channels (mKATP) may activate extracellular signal-regulated kinase1/2 (ERK1/2) via reactive oxygen species (ROS) to provide delayed protection in anoxic preconditioning (APC). This mechanism is crucial for cellular protection in rat cardiomyocytes.
Area of Science:
- Cardiology
- Cellular Biology
- Biochemistry
Background:
- Extracellular signal-regulated kinase1/2 (ERK1/2) is a key signaling pathway in ischemic preconditioning (IPC).
- Mitochondrial ATP-sensitive potassium channels (mKATP) are implicated as both triggers and effectors in IPC's protective mechanisms.
- The precise relationship between mKATP and ERK1/2 in anoxic preconditioning (APC) requires further elucidation.
Purpose of the Study:
- To investigate the role of mitochondrial ATP-sensitive potassium channels (mKATP) in the delayed protective effects of anoxic preconditioning (APC).
- To determine the relationship between mKATP and extracellular signal-regulated kinase1/2 (ERK1/2) activation in APC.
- To explore the involvement of reactive oxygen species (ROS) in this signaling cascade.
Main Methods:
- An anoxic preconditioning (APC) model was established using cultured neonatal rat cardiomyocytes.
- Pharmacological agents were used to modulate mKATP (diazoxide, 5-hydroxydecanoate) and ERK1/2 (PD98059) activity.
- Cellular injury markers (SOD, cell viability, LDH) and intracellular ROS generation were measured.
Main Results:
- APC and diazoxide (mKATP opener) significantly improved cell viability and SOD activity while reducing LDH release compared to anoxia/reoxygenation (A/R).
- Both APC and diazoxide promoted ROS generation and ERK1/2 activation.
- These protective effects were attenuated by mKATP blockers (5-HD) and ROS scavengers (MPG), and abolished by an ERK1/2 inhibitor (PD98059).
Conclusions:
- Mitochondrial ATP-sensitive potassium channels (mKATP) play a critical role in mediating ERK1/2 activation during anoxic preconditioning (APC).
- Reactive oxygen species (ROS) generation is a key intermediate step in the mKATP-dependent activation of ERK1/2.
- This pathway contributes to the delayed cellular protection observed in APC of rat cardiomyocytes.
