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A Flow Cytometry-based Assay for Measuring Mitochondrial Membrane Potential in Cardiac Myocytes After Hypoxia/Reoxygenation
Published on: July 13, 2018
Synaptotagmin-1 attenuates myocardial programmed necrosis and ischemia/reperfusion injury through the mitochondrial
Teng Sun1, Jialei Li2, Shuang Wang2
1Key Laboratory of Cellular Physiology at Shanxi Medical University, Ministry of Education, and the Department of Physiology, School of Basic Medicine, Shanxi Medical University, Taiyuan, China. tengsun@sxmu.edu.cn.
Insights
Synaptotagmin-1 (Syt1) protects the heart from damage by preventing programmed cell death (necroptosis). This study reveals Syt1
Area of Science:
- Cardiology
- Molecular Biology
- Cell Death Research
Background:
- Programmed necrosis, or necroptosis, significantly contributes to cardiac disorders like myocardial infarction and heart failure.
- The precise mechanisms of myocardial necroptosis, particularly mitochondria-dependent pathways, remain incompletely understood.
- Synaptotagmin-1 (Syt1), a calcium sensor, is expressed in cardiomyocytes, but its role in cardiac pathology is unclear.
Purpose of the Study:
- To investigate the function and molecular mechanisms of Synaptotagmin-1 (Syt1) in myocardial necroptosis.
- To elucidate the role of Syt1 in cardiac disorders such as ischemia/reperfusion (I/R) injury.
- To identify the regulatory pathways controlling Syt1 expression and its impact on cardiac cell death.
Main Methods:
- Investigated Syt1 expression in mouse models of cardiac injury (I/R) and in vitro models (H2O2-challenged and hypoxia/reoxygenation-damaged cardiomyocytes).
- Examined the effects of Syt1 overexpression on myocardial necroptosis, fibrosis, and cardiac function in I/R mice.
- Utilized co-immunoprecipitation and ubiquitination assays to explore the interaction between Syt1, Parkin, and CypD.
- Analyzed the regulatory relationship between miR-193b-3p and Syt1 using molecular biology techniques.
Main Results:
- Syt1 expression was significantly downregulated in I/R injured heart tissues and stressed cardiomyocytes.
- Enforced Syt1 expression attenuated myocardial necroptosis, reduced interstitial fibrosis, and improved cardiac function post-I/R.
- Syt1 interacts with Parkin, promoting Parkin-mediated CypD ubiquitination, inhibiting mitochondrial membrane permeability transition pore (mPTP) opening, and suppressing necroptosis.
- miR-193b-3p was identified as a negative regulator of Syt1, influencing cardiomyocyte necrosis and mPTP opening.
Conclusions:
- Revealed a novel regulatory pathway involving miR-193b-3p, Syt1, Parkin, and CypD in myocardial necroptosis.
- Syt1 plays a protective role against cardiac cell death by modulating the mitochondrial pathway.
- This pathway presents potential therapeutic targets for heart protection strategies.
Abstract:
Programmed necrosis/necroptosis greatly contributes to the pathogenesis of cardiac disorders including myocardial infarction, ischemia/reperfusion (I/R) injury and heart failure. However, the fundamental mechanism underlying myocardial necroptosis, especially the mitochondria-dependent death pathway, is poorly understood. Synaptotagmin-1 (Syt1), a Ca2+ sensor, is originally identified in nervous system and mediates synchronous neurotransmitter release. The later findings of Syt1 expressions in many non-neuronal tissues including muscles suggest that Syt1 may exert important functions beyond regulation of neurotransmitter release. Syt1 is highly expressed in cardiomyocytes and has been used as an extracellular molecular probe for SPECT imaging of cardiac cell death in acute myocardial infarction. However, whether Syt1 functions in the pathogenesis of cardiac disorders and what is the molecular etiology have not yet been clarified. We showed here that Syt1 expression was significantly down-regulated in mice I/R injured heart tissues, H2O2-challenged cardiomyocytes and hypoxia/reoxygenation (H/R)-damaged cardiomyocytes. Enforced expression of Syt1 significantly inhibited myocardial necrotic cell death and interstitial fibrosis, and improved cardiac function in mice subjected to I/R operation. In exploring the underlying mechanisms, we found that Syt1 interacted with Parkin and promoted Parkin-catalyzed CypD ubiquitination, thus inhibited mitochondrial membrane permeability transition pore (mPTP) opening and ultimately suppressed cardiomyocyte necrosis. We further found that Syt1 expression was negatively regulated by miR-193b-3p. MiR-193b-3p regulated cardiomyocyte necrosis and mPTP opening by targeting Syt1. Our present work revealed a novel regulatory model of myocardial necrosis composed of miR-193b-3p, Syt1, Parkin, and CypD, which may provide potential therapeutic targets and strategies for heart protection.
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