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 me dia 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: alpha2(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. By 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 hut 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 upregulation 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.