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Abstract
Prostaglandins (PGs) protect a variety of gastrointestinal cells against injury induced
by ethanol and other noxious agents. This investigation attempted to discern the mechanism
of cytoprotection as it relates to the relationship between actin and PGs in IEC-6
cells (a rat intestinal epithelial cell line). IEC-6 cells were incubated in Dulbecco's
modified Eagle's medium ± 16,16-dimethyl prostaglandin E2 (dmPG, 2.6 μM) for 15 min and subsequently incubated in medium containing 1, 2.5,
5, 7.5, and 10% ethanol (EtOH). Cells were then processed for immunocytochemistry
using FITC-phalloidin in order to stain the actin cytoskeleton, and cell viability
was determined by trypan blue exclusion. Quantitative Western immunoblotting of fractioned
G-actin (nonpolymerized; S1) and F-actin (polymerized; S2) was also carried out. EtOH
concentrations equal to and greater than 5% led to the collapse of the actin cytoskeleton
as depicted by extensive disorganization and fragmentation. In addition, these same
EtOH concentrations significantly decreased the S2 fraction and increased the S1 pool
of actin. Preincubation with dmPG prevented collapse of the actin cytoskeleton, significantly
increased the S2 polymerized fraction as determined by quantitative immunoblotting,
and increased cell viability in EtOH-treated cultures. Prior incubation with cytochalasin
D, an actin disruptive agent, not only reduced cell viability but also prevented the
cytoprotective effects of dmPG. Phalloidin, an actin stabilizing agent, had effects
similar to that of dmPG as demonstrated by stability of the actin cytoskeleton and
increased cellular viability. Such findings indicate that PGs are important in the
organization and stability of actin under in vitro conditions. These effects on actin may play an essential role in the mechanism of
PG-induced cytoprotection.
Keywords
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© 2000 Academic Press. Published by Elsevier Inc. All rights reserved.
