A regulatory defect of constitutive no-synthase in islet endothelial cells correlates with probability of disease manifestation in BBdp rats.

Abstract

Type I (insulin-dependent) diabetes mellitus is characterised by leucocyte infiltration of pancreatic islets and a progressive destruction of insulin-producing beta cells. As endothelial nitric oxide production is known to regulate adhesion molecule expression and leucocyte permeation, we examined the activity and expression of the constitutive nitric oxide synthase (ecNOS) of islet endothelial cells from prediabetic BBdp rats. Cultures of aortic endothelial cells and islet capillary endothelial cells were established from young normoglycaemic BBdp rats, Wistar rats and diabetes-resistant BBdr rats, all matched for age. Nitrite and citrulline production was measured in all culture supernatants as indicators for ecNOS activities. Expression of ecNOS mRNA was assessed by reverse transcription-polymerase chain reaction. In contrast to those of the aorta, the Wistar rat islet derived endothelial cells exhibited a strong positive correlation of ecNOS activity with the culture medium glucose concentration but none of the BB rat-derived islet endothelial cells showed a similar glucose-responsiveness. Furthermore, at physiological as well as at increased glucose concentrations islet endothelia from all BBdp rats exhibited a considerable decrease in ecNOS activity by a factor of 3 to 6, indicating a specific dysfunction which is also found for the inducible nitric oxide synthase activity after cytokine challenge but effects were less (2.5 to 3 times) dramatic. In contrast, aorta endothelia from all rats exhibited identical ecNOS activities and no glucose responsiveness. We also found a correlation between ecNOS activities and ecNOS-mRNA expression and can exclude the involvement of the inducible isoform. A reproducible and highly significant dysfunction of islet ecNOS expression and activity in young normoglycaemic BBdp rats, which strongly correlates with the probability for disease manifestation is shown.