Endothelial cell monolayer integrity. I. Characterization of dense peripheral band of microfilaments.

Abstract

Although the endothelial cell (EC) cytoskeleton has been studied both in vitro and in vivo, little is known about its role in endothelial integrity. We have previously suggested that specific EC microfilament (MF) structure, which we have termed the "dense peripheral band (DPB)," may play a major role in this process. We have extended our studies to characterize this structure in pig aortic ECs in vitro. During the growth of EC cultures, the DPB appears only when the cultures have attained confluency. Using double fluorescent labeling, we found that alpha-actinin, myosin, and tropomyosin colocalized with the F-actin making up the DPB. Occasional microtubules were present in this region, although there was no preferred association between microtubules and the DPB. Colocalization studies revealed vinculin plaques at the cell-cell interface. Thin MFs extended from the DPB into the cytoplasmic side of these plaques. The DPB was completely disrupted by low dose cytochalasin B within 30 minutes, whereas many central MF bundles were still present at 24 hours. The results of this study suggest that the DPB is a distinct structure in the confluent EC monolayer and is closely associated with the ability of ECs to form and maintain the EC monolayer. The disruption of the DPB as an important initial event in the pathogenesis of vascular diseases such as atherosclerosis is discussed. (Arteriosclerosis 6:212-219, March/April 1986) C urrent theories of atherogenesis suggest that loss of endothelial integrity may be one of the important early events leading to the formation and growth of the atheromatous fibrofatty plaque.1^1 This loss of integrity is due to endothelial cell (EC) denudation from the monolayer 5 " 7 as well as to nondenuding EC dysfunction. 8 ' 9 We have previously proposed that the F-actin cytoskeleton may be important in the maintenance of this integrity. 10The endothelial cytoskeleton has been described in several in vitro 10 " 18 and in vivo 17 " 25 studies in both the normal and injured aortic wall. We have earlier shown the importance of microfilament (MF) bundles, microtubules, and centrosomes in the process of aortic reendothelialization after large 12 " 15 and small 10 denuding injuries made in the confluent in vitro monolayer. The studies suggested that these aortic endothelial structures play an important role in cell adhesion, spreading, and translocation.The F-actin MF bundles in EC are present in two areas of the EC: at the cell periphery as a dense peripheral band (DPB), and more centrally as ventrally located bundles in Received July 19, 1985; revision accepted October 21, 1985. what are generally described as stress fibers. 24 Although much is known about the central MF bundles with respect to structure and function, the EC peripheral actin MF bundles have not been well studied. Our previous in vitro studies have shown that the presence of the DPB was closely associated with the lamellipodia extrusion occurring during rapid closure of single cell wounds in a confluent monolayer.10 In addition...

The role of shear stress in the pathogenesis of atherosclerosis

“…[157][158][159] Actin microfilaments are arranged both centrally and peripherally in endothelial cells to form stress fibers 160 and a dense peripheral band, 161 respectively. Stress fibers are long bundles of actin filaments containing myosin, tropomyosin and alpha-actinin that can contract and exert tension.…”

Thrombospondin-1 Induces Tyrosine Phosphorylation of Adherens Junction Proteins and Regulates an Endothelial Paracellular Pathway

“…Agents that disrupt actin microfilaments increase endothelial permeability, previous F-actin stabilization protects against this increase, and established mediators of permeability induce actin reorganization (Shasby et al, 1982;Bussolino et al, 1987;Goldblum et al, 1993). In EC, F-actin is arranged into both central transcytoplasmic cables and a peripheral band (Wong and Gotlieb, 1986). These microfilaments are linked to two types of adherens junctions, FAs (Clark and Brugge, 1995;Parsons and Parsons, 1997) and the zonula adherens (ZA) (Kemler, 1993;Gumbiner, 1996;Barth et al, 1997).…”

Adenosine diphosphate-ribosylation of G-actin by botulinum C2 toxin increases endothelial permeability in vitro.

“…Circumstantial evidence suggests that enhanced endothelial permeability is related to alterations of the cellular cytoskeleton (6,7,(14)(15)(16). Endothelial cells have been shown to contain an elaborated microfilament system, consisting of actin, myosin, alpha-actinin, tropomyosin, spectrin, and other components (17)(18)(19)(20)(21)(22)(23). Recent studies in skinned endothelial monolayers using N-ethylmaleimide modified myosin subfragment 1 suggest that an interaction of endothelial actin and myosin is important for the regulation of endothelial permeability (24).…”