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PKCdelta regulates endothelial basal barrier function through modulation of RhoA GTPase activity
Elizabeth O Harrington1, Christopher J Shannon, Nicole Morin
1Pulmonary Vascular Research Laboratory, Providence VA Medical Center, Department of Medicine, Brown Medical School, 151 Research Services, 830 Chalkstone Avenue, Providence, RI 02908, USA. Elizabeth_Harrington@brown.edu
Experimental Cell Research
|June 7, 2005
Summary
Protein kinase C delta (PKCdelta) stabilizes endothelial barrier function by regulating RhoA GTPase activity. This pathway involves focal adhesion kinase and RhoGAP proteins, crucial for maintaining vascular integrity.
Area of Science:
- Cell Biology
- Biochemistry
- Physiology
Background:
- Endothelial barrier integrity is crucial for vascular health.
- Protein kinase C delta (PKCdelta) is implicated in regulating endothelial barrier function.
- RhoA GTPase signaling plays a key role in cell adhesion and cytoskeletal organization.
Purpose of the Study:
- To investigate the signaling pathways by which PKCdelta modulates endothelial barrier function.
- To determine the role of RhoA GTPase and its regulators in PKCdelta-mediated barrier effects.
- To assess the impact of PKCdelta inhibition on endothelial cell structure and function.
Main Methods:
- Utilized rat pulmonary artery endothelial cells (PAEC) and epididymis endothelial cells (FPEC).
- Inhibited PKCdelta using rottlerin and dominant-negative adenovirus (Ad PKCdeltadn).
- Assessed monolayer permeability, stress fibers, focal contacts, and RhoA/FAK activity via immunofluorescence and biochemical assays.
Main Results:
- PKCdelta inhibition (rottlerin or Ad PKCdeltadn) increased endothelial monolayer permeability.
- Inhibition led to reduced stress fibers and focal contacts, correlating with decreased RhoA and focal adhesion kinase (FAK) activity.
- Overexpression of dominant active RhoA protected endothelial cells from barrier disruption, while FAK overexpression had no protective effect.
- PKCdelta was found to interact with p190RhoGAP and p120RasGAP, known modulators of RhoA.
Conclusions:
- PKCdelta enhances endothelial barrier function by promoting focal contact formation and stabilizing microfilaments.
- This regulation is mediated through the RhoA GTPase pathway, influenced by p190RhoGAP and p120RasGAP.
- PKCdelta plays a critical role in maintaining basal endothelial barrier integrity.