Cofilin mediates ATP depletion-induced endothelial cell actin alterations

Abstract

Ischemia and sepsis lead to endothelial cell damage, resulting in compromised microvascular flow in many organs. Much remains to be determined regarding the intracellular structural events that lead to endothelial cell dysfunction. To investigate potential actin cytoskeletal-related mechanisms, ATP depletion was induced in mouse pancreatic microvascular endothelial cells (MS1). Fluorescent imaging and biochemical studies demonstrated a rapid and progressive increase in F-actin along with a decrease in G-actin at 60 min. Confocal microscopic analysis showed ATP depletion resulted in destruction of actin stress fibers and accumulation of F-actin aggregates. We hypothesized these actin alterations were secondary to dephosphorylation/activation of actin-depolymerizing factor (ADF)/cofilin proteins. Cofilin, the predominant isoform expressed in MS1 cells, was rapidly dephosphorylated/activated during ATP depletion. To directly investigate the role of cofilin activation on the actin cytoskeleton during ischemia, MS1 cells were infected with adenoviruses containing the cDNAs for wild-type Xenopus laevis ADF/cofilin green fluorescent protein [XAC(wt)-GFP], GFP, and the constitutively active and inactive isoforms XAC(S3A)-GFP and XAC(S3E)-GFP. The rate and extent of cortical actin destruction and actin aggregate formation were increased in ATP-depleted XAC(wt)-GFP- and XAC(S3A)-GFP-expressing cells, whereas increased actin stress fibers were observed in XAC(S3E)-GFP-expressing cells. To investigate the upstream signaling pathway of ADF/cofilin, LIM kinase 1-GFP (LIMK1-GFP) was expressed in MS1 cells. Cells expressing LIMK1-GFP protein had higher levels of phosphorylated ADF/cofilin, increased stress fibers, and delayed F-actin cytoskeleton destruction during ATP depletion. These results strongly support the importance of cofilin regulation in ischemia-induced endothelial cell actin cytoskeleton alterations leading to cell damage and microvascular dysfunction.

A Stromal Cell-Derived Factor 1α Analogue Improves Endothelial Cell Function in Lipopolysaccharide-Induced Acute Respiratory Distress Syndrome

“…Genomic ablation of miR-126 has been shown to increase vascular permeability (16,17,19,21). MicroRNA-126 targets PIK3R2 and Spred-1 leading to stabilization of vascular endothelial (VE)-cadherin via AKT and ERK 1/2 signaling cascades (22,23). CTCE injections result in significantly increased plasma miR-126 levels in CLP-induced sepsis suggesting a potential mechanism by which it reduces endothelial permeability (12).…”

Roles of BMP6/7 in Actin Dynamics in Amyloid β-Induced Neurotoxicity

“…Actin has two forms found in neurons: monomer form-globular actin (G-actin) and polymer form-filamentous actin (F-actin) (Suurna et al, 2006). F-actin forms a two-stranded asymmetrical helical structure which allows for different binding specificities at each end.…”

“…The binding of ADF/cofilin to G-actin leads to sequestering actin monomers, while the binding of ADF/cofilin to F-actin leads to severing actin polymers (Jang et al, 2005). ADF/cofilin prefers binding the ADP form of F-actin, inducing a twist in the filament that will destabilize interaction between actin monomers and enhances the dissociation of actin at the pointed end of F-actin (Minamide et al, 2000;Suurna et al, 2006). Filament severing by ADF/cofilin is a "Janus-faced" process, since the process can cause either an increase or a decrease in polymerized actin.…”

“…Rapid reassembly and elongation of actin filaments can be seen during cell recovery in the early period of neurodegenerative stimulation. First, less F-actin destruction is observed following 30 min of ATP depletion, then the concentration of F-actin increases at 60 minutes of ATP depletion, leading to promote actin assembly and form a very fine actin network (Suurna et al, 2006). Incubating Aβ with primary cultured cortical neurons for 60 minutes also induces actin polymerization, which can form stress fibers and exert a protective role (Song et al, 2002) (Figure 1).…”

“…There are very complex molecule signals involving the regulation of LIMK1 and SSH. Previous experiments have demonstrated that CDC42/Rac considerably overlaps with early cytoskeleton abnormalities which were observed in AD brain sections, the defects in P21-activated kinase (PAK) activity can lead to actin pathology during development of AD, and high levels of LIMK1 coexist with delayed F-actin cytoskeleton destruction in transient ATP depletion (Mendoza-Naranjo et al, 2007;Suurna et al, 2006;Heredia et al, 2006). Thus, it can be assumed that reactive CDC42/Rac-PAK-LIMK1 pathway is promoted to counteract over hyper-dephosphorylation of ADF/cofilin and rescue cytoskeleton damage.…”