Involvement of c-Src in diperoxovanadate-induced endothelial cell barrier dysfunction

Area of Science:

• Cell Biology
• Biochemistry
• Pulmonary Medicine

Background:

• Reactive oxygen species (ROS) from leukocytes compromise endothelial cell (EC) integrity, causing barrier dysfunction and pulmonary edema.
• Mechanisms of ROS-induced EC barrier dysfunction are not fully understood.
• Tyrosine phosphorylation is implicated in cell signaling, but its role in EC barrier function needs clarification.

Purpose of the Study:

• To investigate the role of tyrosine phosphorylation in ROS-induced EC barrier dysfunction.
• To explore the involvement of Src kinase in the disruption of EC barrier function.
• To identify potential cytoskeletal targets regulated by Src in this process.

Main Methods:

• Utilized diperoxovanadate (DPV) as a model agent to induce ROS-like effects in ECs.
• Assessed EC barrier function using dose-response studies.
• Employed tyrosine kinase inhibitors (genistein, PP-2) and Src-specific inhibitors.
• Measured Src activation and its association with cortactin and myosin light chain kinase (MLCK).
• Investigated the effects of Src overexpression (wild-type, constitutively active, dominant-negative mutants).

Main Results:

• DPV disrupted EC barrier function in a dose-dependent manner.
• Tyrosine kinase inhibitors and PP-2 significantly reduced DPV-mediated barrier dysfunction.
• PP-2 attenuated DPV-induced Src activation.
• DPV increased the association of Src with cortactin and MLCK.
• Overexpression of Src (wild-type or active mutant) potentiated DPV effects, while a dominant-negative mutant attenuated them.

Conclusions:

• Src kinase plays a critical role in mediating diperoxovanadate-induced endothelial cell barrier dysfunction.
• Tyrosine phosphorylation, specifically via Src activation, is a key mechanism in ROS-related EC barrier compromise.
• Cortactin and MLCK are identified as potential downstream targets of Src in regulating EC barrier integrity.