Purdue University Cytometry Laboratories, Department of Basic Medical Sciences, Purdue University, West-Lafayette, IN 47907, USA.
Diabetic Angiopathy is characterized by macrovascular and microvascular abnormalities, which may occur independent of each other. It has been proposed that macrovascular endothelial cells do not function in a similar manner as microvascular endothelial cells from the same vascular bed. The purpose of our study was to compare the function of macrovascular and microvascular endothelial cells from normal and diabetic rats in an identical vascular bed, being the dorsum of the foot.
Endothelial cells were harvested from isolated (micro) vessels by the use of collagen coated 46 µm beads, and purified by fluorescence activated cell sorting. Differences in cultured endothelial cell function were assessed by the use of the following cytometric tools: Superoxide anion and H2O2 production in adherent and non-adherent endothelial cells was evaluated using Hydroethidine and DCFH-DA probes. As modulators of cellular respiration and oxidation allopurinol, oxypurinol, DMSO, Deferoxamine, azide and cyanide were evaluated independently in the same system. There is evidence that nitric oxide synthase activity is reduced in diabetic macrovascular endothelial cells. Non adherent endothelial cells were stimulated with NONO-ate to release NO¥ to be evaluated with DCF an HE probes and the effect of nitric oxide modulators such as L-arginine, L-NAME, and L-NNA was assessed. Comparative data regarding H2O2 , O2- and NO¥ production were collected on the microvascular and macrovascular cells. Functional differences in cultured endothelial cells, while not necessarily physiologic, may reflect abnormalities observed in vivo in pathology such as diabetic angiopathy.