Long-term exposure to high glucose up-regulates VCAM-induced endothelial cell adhesiveness to PBMC.
BACKGROUND
The changes induced on endothelial cells by a long-term exposure to high glucose, a situation that mimics the hyperglycemia of diabetics, have not yet been determined. We compared short- and long-term effects of elevated glucose on macrovascular and microvascular endothelial cells.


METHODS
Endothelial cells were grown in high-glucose media for 24 hours and for 8 weeks. Cell proliferation was evaluated by cell counting, apoptosis and expression of adhesion molecules by flow cytometry; nitric oxide (NO) by measuring the concentration of nitrite/nitrate in the cell supernatant; alpha 2(IV) collagen mRNA and protein by reverse transcriptase-polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. The adhesion of peripheral blood mononuclear cells (PBMCs) to endothelial cells was evaluated by adhesion assay. In some experiments, endothelial cells were preincubated with anti-vascular cell adhesion molecule-1 (VCAM-1) and anti-receptor for advanced glycation end product (RAGE) blocking antibodies.


RESULTS
At 24 hours, but not at 8 weeks, high glucose increased endothelial cell proliferation and apoptosis. High glucose did not modify NO synthesis at 24 hours and 8 weeks. Collagen production and expression were increased only after eight weeks. VCAM-1 but not intercellular adhesion molecule-1 was up-regulated after 8 weeks, a change not observed after 24 hours. The adhesion of PBMCs was significantly increased at eight weeks and was completely abrogated by anti--VCAM-1 and by anti-RAGE antibodies. After 24 hours, there was a modest increase of PBMC adhesion that was not blunted by anti-RAGE antibodies.


CONCLUSIONS
Increased adhesion of PBMCs, caused by up-regulation of VCAM-1 with a mechanism involving advanced glycation end product (AGE) adducts, and augmented collagen deposition are critical effects of long-term high glucose on endothelial cells, and may eventually promote the atherosclerotic process.
Esposito
C
C
Unit of Nephrology, Dialysis and Transplantation, IRCCS Policlinico San Matteo, University of Pavia, Pavia, Italy. ciroe@pop.systemy.it
Fasoli
G
G
Plati
A R
AR
Bellotti
N
N
Conte
M M
MM
Cornacchia
F
F
Foschi
A
A
Mazzullo
T
T
Semeraro
L
L
Dal Canton
A
A
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002417:Cattle; D002448:Cell Adhesion / Q000187:drug effects / Q000502:physiology; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D003094:Collagen / Q000096:biosynthesis; D003925:Diabetic Angiopathies / Q000209:etiology; D004730:Endothelium, Vascular / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D005947:Glucose / Q000008:administration & dosage / Q000494:pharmacology*; D006801:Humans; D066298:In Vitro Techniques; D007963:Leukocytes, Mononuclear / Q000166:cytology; D009569:Nitric Oxide / Q000096:biosynthesis; D015854:Up-Regulation / Q000187:drug effects; D019010:Vascular Cell Adhesion Molecule-1 / Q000378:metabolism*
D019010:Vascular Cell Adhesion Molecule-1; D009569:Nitric Oxide; D003094:Collagen; D005947:Glucose