MARCKS protein mediates hydrogen peroxide regulation of endothelial permeability

Abstract

Impairment of endothelial barrier function is implicated in many vascular and inflammatory disorders. One prevalent mechanism of endothelial dysfunction is an increase in reactive oxygen species under oxidative stress. Previous reports have demonstrated that hydrogen peroxide (H 2 O 2 ), a highly stable reactive oxygen species that modulates physiological signaling pathways, also enhances endothelial permeability, but the mechanism of this effect is unknown. Here, we identify the actin-binding protein myristoylated alanine-rich C-kinase substrate (MARCKS) as a key mediator of the H 2 O 2 -induced permeability change in bovine aortic endothelial cells. MARCKS knockdown and H 2 O 2 treatment alter the architecture of the actin cytoskeleton in endothelial cells, and H 2 O 2 induces the phosphorylation and translocation of MARCKS from the cell membrane to the cytosol. Using pharmacological inhibitors and small interference RNA constructs directed against specific proteins, we uncover a signaling cascade from Rac1 to Abl1, phospholipase Cγ1, and PKCδ that is triggered by H 2 O 2 and leads to MARCKS phosphorylation. Our findings establish a distinct role for MARCKS in the regulation of H 2 O 2 -induced permeability change in endothelial cells, and suggest potential new therapeutic targets for the treatment of disorders involving oxidative stress and altered endothelial permeability.

Endothelial Na + /H + exchanger NHE1 participates in redox-sensitive leukocyte recruitment triggered by methylglyoxal

“…Cariporide may, in addition, counteract inflammation by abating cytokine production [82]. MG was previously shown to trigger increased microvascular permeability [21], an effect mediated by oxidative stress [21,83]. Our data reveal that NHE1 inhibition attenuated MG-triggered microvascular hyperpermeability.…”

“…However, whether NHE1 activation directly regulates microvascular barrier function is not known. It is possible that reduction in MG-triggered microvascular hyperpermeability after cariporide treatment results from the suppression of NHE1 activation which regulates both oxidative stress and transendothelial migration of leukocytes, key elements that are vital to the microvascular permeability regulation [83,84]. By the same token, leukocyte recruitment is also triggered by oxidative stress [21,85] and our data suggests that inhibition of MG-induced leukocyte recruitment by cariporide involves a redox-sensitive mechanism.…”

Reactive Oxygen Species in Inflammation and Tissue Injury

“…Oxidation of actin monomers (G-actin) by H 2 O 2 alters its polymerization ability as well as its interaction with actin regulatory proteins filamin and a-actinin (77). H 2 O 2 is known to induce actin polymerization at doses ranging from 0.2 to 5 mM with an effective dose (ED50) at 1 mM H 2 O 2 (198,324,411). The cysteine and methionine residues in G-actin are particularly susceptible to oxidation.…”

Endothelial Angiogenesis and Barrier Function in Response to Thrombin Require Ca2+ Influx through the Na+/Ca2+ Exchanger

“…8). ROS are well established to cause barrier dysfunction (57), and thrombin is reported to increase ROS production in ECs primarily via Nox2 (33). Moreover, Ca 2ϩ signaling and Nox2 activity have been implicated in endothelial hyperpermeability in models of liposaccharide-induced vascular inflammation (53) and lung ischemia-reperfusion injury (58) in mice.…”