Background. In our previous studies, we found that intraglomerular depositi
on of fibrin and its metabolites was related to glomerular sclerosis and re
duced renal function. It has been reported that both overlying and underlyi
ng fibrin may induce specific morphological changes of cultured endothelial
cells from large blood vessels. The dependency of these morphological chan
ges on the integrin alpha(v)beta(3)-arginyl-glycyl-aspartyl-serine (RGDS) i
nteraction is still controversial. We hypothesized that glomerular endothel
ial cells (GECs) stimulated by fibrin might undergo morphological changes t
hrough an integrin alpha(v)beta(3)-RGDS interaction.
Methods. In vitro studies were performed to examine the growing status of G
ECs stimulated by overlying and underlying fibrin gels in the presence or a
bsence of the following: 50 mu g/ml anti-alpha(v)beta(3) integrin monoclona
l antibody 23C6 or nonimmune mouse IgG, 1 mg/ml synthetic RGDS or arginyl-g
lycyl-glycyl-serine (RGGS) peptide, 10 mg/ml sodium heparin, 100 mu g/ml cy
cloheximide, and 10 mu M actinomycin D. Fast protein liquid chromatography
(FPLC)-purified fibrinogen and the third to fifth passages of human GECs we
re also used in this study.
Results. GECs developed capillary tube structure after 60 hours of culturin
g on fibrin gels, and GECs cultured on gelatin-coated plates displayed a mo
nolayer of cobblestone-like cells in the presence or absence of 23C6 and sy
nthetic RGDS peptide. Fibrin-induced capillary tube formation was promoted
by 23C6 and inhibited by RGDS peptide, cycloheximide, and actinomycin D. Di
sorganization of the GEC monolayer was induced by overlying fibrin, but was
not induced by overlying agarose gels and glass cover slips or culturing i
n fibrinogen, 0.05 NIH U/ml thrombin, fibrin supernatants, as well as in fi
brin degradation products. Disorganization of GEC monolayer can be induced
by both des-AA-fibrin and des-AABB-fibrin and was unaffected by heparin. Fu
rthermore, both 23C6 and synthetic RGDS peptide prevented disorganization o
f GECs induced by overlying fibrin, whereas nonimmune mouse IgG, synthetic
RGGS peptide, cycloheximide, and actinomycin D had no similar effect.
Conclusions. GECs cultured on fibrin gels may develop capillary structure s
pontaneously, and GECs covered by fibrin gels may undergo disorganization.
Our data suggest that these GEC morphological changes are mediated by an in
tegrin alpha(v)beta(3)-RGDS interaction.