Involvement of local lamellipodia in endothelial barrier function

Area of Science:

• Cell Biology
• Physiology
• Biophysics

Background:

• Endothelial cells form a barrier regulating permeability.
• Lamellipodia are dynamic actin-rich structures at the cell periphery.
• The role of lamellipodia in endothelial barrier function is not well understood.

Purpose of the Study:

• To investigate the role of local lamellipodia in endothelial barrier integrity.
• To determine the molecular mechanisms regulating lamellipodia dynamics and barrier function.

Main Methods:

• Time-lapse microscopy of endothelial cells expressing fluorescent reporters (GFP-actin, VE-cadherin-GFP).
• Measurement of transendothelial electrical resistance (TER) as an index of barrier function.
• Pharmacological inhibition of lamellipodia formation (blebbistatin) and modulation of signaling pathways (thrombin, S1P, Rac1 inhibitors).
• Assessment of myosin light chain (MLC) phosphorylation and GTPase activation (Rac1, RhoA).

Main Results:

• Thrombin decreased TER and lamellipodia frequency; S1P increased TER and lamellipodia frequency.
• Blebbistatin abolished lamellipodia, reduced TER in cell culture, and increased venule permeability.
• Both thrombin and S1P modulated MLC phosphorylation and RhoA activation.
• Rac1 activation correlated with increased barrier function and lamellipodia activity, while inhibition reduced both.

Conclusions:

• Local lamellipodia, regulated by myosin II and Rac1, are critical for dynamic regulation of endothelial barrier function.
• These findings reveal a novel mechanism for controlling endothelial permeability.