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Updated: Aug 29, 2025
Evaluating Cell Death Signaling by Immunofluorescence in a Rat Model of Ischemic Stroke
Published on: January 3, 2025
Sam50 exerts neuroprotection by maintaining the mitochondrial structure during experimental cerebral
Xulong Yin1,2, Jiahe Wang2,3, Siyuan Yang2,3
1Department of Neurology, The First Affiliated Hospital of Soochow University, Suzhou, China.
Aim:
To investigate the role of Sam50, a barrel protein on the surface of the mitochondrial outer membrane, in cerebral ischemia-reperfusion (I/R) injury and its underlying mechanisms.
Methods:
A middle cerebral artery occlusion/reperfusion (MCAO/R) model in adult male Sprague-Dawley rats was established in vivo, and cultured neurons were exposed to oxygen-glucose deprivation/reoxygenation (OGD/R) to simulate I/R injury in vitro. Lentiviral vector encoding Sam50 or Sam50 shRNA was constructed and administered to rats by intracerebroventricular injection to overexpress and knockdown Sam50, respectively.
Results:
First, after MCAO/R induction, the mitochondrial structure was damaged, and Sam50 protein levels were increased responsively both in vivo and in vitro. Then, it was found that Sam50 overexpression could reduce infarction size, inhibit neuronal cell death, improve neurobehavioral disability, protect mitochondrial structure integrity, and ameliorate mitochondrial dysfunction, which was induced by I/R injury both in vivo and in vitro. However, Sam50 downregulation showed the opposite results and aggravated I/R injury by inducing neuronal cell death, neurobehavioral disability, and mitochondrial dysfunction. Moreover, we found that the interaction between Sam50 and Mic19 was broken off after OGD/R, showing that the Sam50-Mic19-Mic60 axis was breakage in neurons, which would be a reason for mitochondrial structure and function abnormalities induced by I/R injury.
Conclusion:
Sam50 played a vital role in the protection of neurons and mitochondria in cerebral I/R injury, which could be a novel target for mitochondrial protection and ameliorating I/R injury.
Insights
Sam50 protein protects against brain damage from stroke (cerebral ischemia-reperfusion injury). Overexpressing Sam50 reduces injury, while reducing it worsens outcomes, highlighting Sam50 as a therapeutic target.
Area of Science:
- Neuroscience
- Mitochondrial Biology
- Ischemia-Reperfusion Injury
Background:
- Cerebral ischemia-reperfusion (I/R) injury significantly impacts neurological outcomes.
- Mitochondrial dysfunction is a key factor in I/R-induced neuronal damage.
- The role of outer mitochondrial membrane proteins in I/R injury requires further elucidation.
Purpose of the Study:
- To investigate the function of Sam50, an outer mitochondrial membrane protein, in cerebral I/R injury.
- To explore the underlying molecular mechanisms by which Sam50 influences I/R pathophysiology.
- To assess Sam50 as a potential therapeutic target for mitigating I/R damage.
Main Methods:
- Established in vivo (middle cerebral artery occlusion/reperfusion in rats) and in vitro (oxygen-glucose deprivation/reoxygenation in cultured neurons) models of cerebral I/R injury.
- Utilized lentiviral vectors for Sam50 overexpression and knockdown in vivo and in vitro.
- Assessed infarct size, neuronal cell death, neurobehavioral deficits, and mitochondrial structure/function.
Main Results:
- Sam50 protein levels increased in response to I/R injury, with damaged mitochondrial structure.
- Sam50 overexpression reduced infarct size, neuronal death, and neurobehavioral deficits, while preserving mitochondrial integrity and function.
- Sam50 knockdown exacerbated I/R injury, increasing neuronal death and mitochondrial dysfunction.
- The Sam50-Mic19-Mic60 axis interaction was disrupted by I/R, contributing to mitochondrial abnormalities.
Conclusions:
- Sam50 plays a crucial protective role in neurons and mitochondria during cerebral I/R injury.
- Modulating Sam50 levels offers a promising therapeutic strategy for neuroprotection and reducing I/R damage.
- Targeting the Sam50-Mic19-Mic60 axis may represent a novel approach for treating stroke-related brain injury.
