hCLOCK Causes Rho-Kinase-Mediated Endothelial Dysfunction and NF-κB-Mediated Inflammatory Responses
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Updated: Mar 30, 2026
On-Chip Endothelial Inflammatory Phenotyping
Published on: July 21, 2012
Xiao Tang1, Daqiao Guo1, Changpo Lin1
1Institute of Vascular Surgery, Department of Vascular Surgery, Zhongshan Hospital, Fudan University, Shanghai 200032, China.
Insights
Hypoxia increases human Circadian Locomotor Output Cycle protein Kaput (hCLOCK) expression, leading to oxidative stress and activating RhoA and NF-κB pathways. This suggests hCLOCK drives vascular damage and inflammation in hypoxic conditions.
Area of Science:
- Biochemistry
- Molecular Biology
- Cellular Biology
Background:
- The human Circadian Locomotor Output Cycle protein Kaput (hCLOCK) gene regulates daily rhythms.
- hCLOCK is implicated in various human malignancies through multiple biochemical pathways.
- The study investigates hCLOCK's role in the hypoxia-oxidative stress response.
Purpose of the Study:
- To elucidate the biochemical mechanisms of hCLOCK in the hypoxia-oxidative stress response.
- To determine how hCLOCK influences Rho GTPases and NF-κB signaling under hypoxia.
- To assess the impact of hCLOCK on reactive oxygen species (ROS) production and vascular function.
Main Methods:
- Measured Rho GTPase expression in normoxic and hypoxic conditions.
- Utilized retroviral shRNA, Rho inhibitor, and ROS scavenger to evaluate hCLOCK's effect.
- Assessed hCLOCK, Rho GTPases, NF-κB effectors, in vitro ROS production, and HUVEC tube formation.
Main Results:
- Hypoxia upregulates hCLOCK, inducing ROS production.
- hCLOCK activation leads to the activation of RhoA and NF-κB signaling pathways.
- Inhibition of hCLOCK, RhoA, or ROS attenuates these pathways, with RhoA inhibition not reducing ROS.
Conclusions:
- Hypoxic states trigger vascular oxidative damage and inflammation.
- hCLOCK-mediated ROS production is a key mechanism driving these effects.
- Subsequent activation of RhoA and NF-κB pathways contributes to vascular pathology.
Background:
The human Circadian Locomotor Output Cycle protein Kaput (CLOCK) gene was originally discovered as a regulator of essential human daily rhythms. This seemingly innocuous gene was then found to be associated with a multitude of human malignancies, via several biochemical pathways. We aimed to further investigate the role of hCLOCK in the hypoxia-oxidative stress response system at the biochemical level.
Methods:
Expression levels of Rho GTPases were measured in normoxic and hypoxic states. The effect of hCLOCK on the hypoxic response was evaluated with the use of a retroviral shRNA vector system, a Rho inhibitor, and a ROS scavenger by analyzing expression levels of hCLOCK, Rho GTPases, and NF-κB pathway effectors. Finally, in vitro ROS production and tube formation in HUVECs were assessed.
Results:
Hypoxia induces ROS production via hCLOCK. hCLOCK activates the RhoA and NF-κB signaling pathways. Conversely, inhibition of hCLOCK deactivates these pathways. Furthermore, inhibition of RhoA or decreased levels of ROS attenuate these pathways, but inhibition of RhoA does not lead to decreased levels of ROS. Overall findings show that hypoxia increases the expression of hCLOCK, which leads to ROS production, which then activates the RhoA and NF-κB pathways.
Conclusion:
Our findings suggest that hypoxic states induce vascular oxidative damage and inflammation via hCLOCK-mediated production of ROS, with subsequent activation of the RhoA and NF-κB pathways.
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