J. Chen et al., Delayed branching of endothelial capillary-like cords in glycated collagenI is mediated by early induction of PAI-1, AM J P-REN, 281(1), 2001, pp. F71-F80
Development of micro- and macrovascular disease in diabetes mellitus (DM) w
arrants a thorough investigation into the repertoire of endothelial cell (E
C) responses to diabetic environmental cues. Using human umbilical vein EC
(HUVEC) cultured in three-dimensional (3-D) native collagen I (NC) or glyca
ted collagen I (GC), we observed capillary cord formation that showed a sig
nificant reduction in branching when cells were cultured in GC. To gain ins
ight into the molecular determinants of this phenomenon, HUVEC subjected to
GC vs. NC were studied using a PCR-selected subtraction approach. Nine dif
ferent genes were identified as up- or downregulated in response to GC; amo
ng those, plasminogen activator inhibitor-1 (PAI-1) mRNA was found to be up
regulated by GC. Western blot analysis of HUVEC cultured on GC showed an in
crease in PAI-1 expression. The addition of a neutralizing anti-PAI-1 antib
ody to HUVEC cultured in GC restored the branching pattern of formed capill
ary cords. In contrast, supplementation of culture medium with the constitu
tively active PAI-1 reproduced defective branching patterns in HUVEC cultur
ed in NC. Ex vivo capillary sprouting in GC was unaffected in PAI-1 knockou
t mice but was inhibited in wild-type mice. This difference persisted in di
abetic mice. In conclusion, the PCR-selected subtraction technique identifi
ed PAI-1 as one of the genes characterizing an early response of HUVEC to t
he diabetic-like interstitial environment modeled by GC and responsible for
the defective branching of endothelial cells. We propose that an upregulat
ion of PAI-1 is causatively linked to the defective formation of capillary
networks during wound healing and eventual vascular dropout characteristic
of diabetic nephropathy.