The vascular endothelium is the interface with hyperglycemia in uncontrolled diabetes mellitus and its complications, including nephropathy, myocardial infarction, blindness, and gangrene. To mimic the vascular endothelium in a controlled experimental situation, we use the endothelial cell line (CRL 1998) obtained from The American Type Culture Collection. Cells are grown to confluency in Media 199 containing 10% fetal calf serum and gentamicin (50mg/ml). The growth media is replaced with experimental media containing high glucose (30mM), insulin (20 Units/ml), or heparin (500 Units/ml) either separately or in combinations. Cells are briefly trypsinized, pelleted, embedded, and sectioned with a Diatome diamond knife for TEM or for SEM, grown on 0.45um filters, followed by fixation and air-drying in situ: this approach to culture allows cells to obtain a polar, more natural phenotype than monolayers on glass or plastic.

Incubation in high glucose results in loss of confluency, production of intercellular gaps, and fewer mitotic figures.