Extracellular signal-regulated kinase 5 (ERK5) has been reported to regulate endothelial integrity and protect from vascular dysfunction under laminar flow. Previously reported research indicates that under laminar flow ERK5 is activated with production of atheroprotective molecules. However, the characterization of ERK5 activation and levels under different flow patterns has not been investigated.
Confluent HUVECs were serum-starved then seeded on glass slides. HUVECs incubated in 1% FBS were exposed to continuous laminar flow (CLF), to-and-fro flow (TFF), or pulsatile forward flow (PFF) in a parallel plate flow chamber. At the end of experimentation, cell lysates were immunoblotted with antibodies to phospho-ERK5 and total ERK5. ERK5 activation was assessed by the levels of phosphorylated ERK5. The densitometric mean ± SEM is calculated and analyzed by ANOVA. p < 0.05 is considered significant.
Levels of ERK5 decreased with all flow conditions with the largest decrease in TFF flow condition. TFF and CLF exhibited sustained ERK5 phosphorylation in HUVECs stimulated for up to 4 hours. PFF had transient phosphorylation of ERK5 at 2 hours, which then became undetectable at 4 hours of exposure to flow. Also, TFF and CLF both showed decreased levels at 4 hours, suggesting a decrease in activation for these flow conditions.
Exposure of HUVEC to different types of shear stress results in varying patterns of activation of ERK5. Activation of ERK5 with TFF suggests a role in the pathogenesis of atherosclerosis and vascular remodeling under disturbed flow conditions.