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Updated: Jul 18, 2026
Evaluating Cell Death Signaling by Immunofluorescence in a Rat Model of Ischemic Stroke
Published on: January 3, 2025
Down-regulation Cdc42 attenuates neuronal apoptosis through inhibiting MLK3/JNK3 cascade during ischemic reperfusion
Jie Zhao1, Dong-Sheng Pei, Quan-Guang Zhang
1Research Center for Biochemistry and Molecular Biology, Jiangsu Key Laboratory of Brain Disease Bioinformation, Xuzhou Medical College, Xuzhou, China.
Abstract:
JNK signaling pathway is activated and involved in the selective neuronal death in the hippocampal CA1 subfield following cerebral ischemia. However, little is known about upstream partner controlling the pathway. Here we reported that ischemia/reperfusion significantly elevated Cdc42 activity, enhanced assembly of the Cdc42-MLK3 complex and activation of JNK pathway. Most importantly, knock-down endogenous Cdc42 selectively suppressed the MLK3/MKK7/JNK3 cascade, and subsequently blocked the phosphorylation of c-Jun and FasL expression. Meanwhile, Bcl-2 was inactivated and the release of cytochrome c was diminished. These alterations eventually perturbed the caspase-3 activation as well as post-ischemic neuronal cell death. Taken together, our findings strongly suggest that Cdc42 serves as an upstream activator and modulates JNK-mediated apoptosis machinery in vivo, which ultimately results in neuronal apoptosis via nuclear and non-nuclear pathways. Thus, Cdc42 may be a potential therapeutic target in ischemic brain injury.
Insights
Cdc42 (cell division control protein 42) activates the JNK pathway, promoting neuronal death after ischemic stroke. Inhibiting Cdc42 reduced this cell death, suggesting it as a therapeutic target for brain injury.
Area of Science:
- Neuroscience
- Molecular Biology
- Cell Signaling
Background:
- The c-Jun N-terminal kinase (JNK) signaling pathway is implicated in selective neuronal death in the hippocampus following cerebral ischemia.
- The upstream regulators controlling JNK pathway activation in this context remain largely unknown.
Purpose of the Study:
- To investigate the role of Cdc42 (cell division control protein 42) as an upstream activator of the JNK pathway in post-ischemic neuronal death.
- To elucidate the molecular mechanisms by which Cdc42 influences JNK-mediated apoptosis in the brain.
Main Methods:
- Assessment of Cdc42 activity and its complex formation with MLK3 (Mixed Lineage Kinase 3) after ischemia/reperfusion.
- Utilizing knockdown of endogenous Cdc42 to analyze its effect on the MLK3/MKK7/JNK3 cascade.
- Evaluation of downstream targets including c-Jun phosphorylation, FasL expression, Bcl-2 inactivation, cytochrome c release, and caspase-3 activation.
Main Results:
- Ischemia/reperfusion significantly increased Cdc42 activity and the formation of the Cdc42-MLK3 complex, leading to JNK pathway activation.
- Knockdown of Cdc42 selectively inhibited the MLK3/MKK7/JNK3 cascade, reducing c-Jun phosphorylation and FasL expression.
- Cdc42 inhibition diminished Bcl-2 inactivation, decreased cytochrome c release, and attenuated caspase-3 activation and neuronal cell death.
Conclusions:
- Cdc42 acts as a crucial upstream activator modulating the JNK-mediated apoptosis pathway in vivo.
- Cdc42 influences neuronal apoptosis through both nuclear and non-nuclear pathways following ischemic brain injury.
- Cdc42 presents a potential therapeutic target for mitigating ischemic brain injury and neuronal loss.
