Abnormalities in primary granule exocytosis in neutrophils from Type I diabetic patients with nephropathy.
Microalbuminuria in Type I diabetes involves a cell membrane abnormality and is associated with a large increase in cardiovascular risk. The hypothesis that the membrane abnormality alters granule exocytosis in neutrophils, which could contribute to the increased incidence of cardiovascular disease, was investigated. PMA-stimulated expression of CD11b and CD69 on neutrophils from normal controls (NC), long-term uncomplicated Type I diabetic control patients (DC) and diabetic nephropathy patients (DN) was determined by fluorescence activated cell scanning. Neutrophils from DN were faster than neutrophils from either NC or DC to exocytose primary granules with CD69 following initial expression of the adhesion molecule CD11b. However, a larger proportion of neutrophils from DN failed to withdraw CD11b from the cell membrane after 90 min incubation. The protein kinase C (PKC) inhibitor, bisindolylmaleimide (BIM), showed that a larger proportion of neutrophils from DN, compared with DC or NC, exocytosed primary granules independent of PKC. The calpain inhibitor, E64d, showed that a larger proportion of neutrophils from both groups of diabetic patients, compared with NC, exocytosed primary granules independent of calpain. Cytoskeletal disruption with cytochalasin D had an effect on CD11b and CD69 exocytosis similar to that of BIM and E64d. The pathways controlling granule exocytosis in neutrophils from diabetic patients are abnormal. A change characteristic of DN causes rapid exocytosis of primary granules, and also causes the adhesion molecule CD11b to persist on an increased proportion of neutrophils. This will make an important contribution to increased vascular damage in these patients.