Oxygen free radicals have been implicated in the pathogenesis of diabetic microangiopathy. The production of superoxide anion (O2-.) by polymorphonuclear leukocytes (PMNs) from 45 insulin-dependent diabetes mellitus patients in the resting state and in response to a soluble stimulus (phorbol myristate acetate) was measured spectrophotometrically and compared with that of 15 age and sex matched controls. The resting superoxide anion production by PMNs from diabetic patients was significantly higher than that of controls (2.17 +/- 1.32 and 1.35 +/- 0.6 nmol/10(5) cells/60 min respectively; p = 0.037). In contrast, PMNs from diabetic patients released significantly lower levels of superoxide anion compared to controls in response to phorbol myristate acetate stimulation (2.33 +/- 2.04 and 3.55 +/- 0.98 nmol/10(5) cells/60 min respectively; p = 0.044). The stimulated superoxide anion production was significantly higher in diabetic patients with retinopathy compared to diabetic patients without retinopathy (2.7 +/- 2.08 and 1.3 +/- 1.6 nmol/10(5) cells/60 min respectively; p = 0.02). Furthermore, stimulated PMNs from diabetic patients with proliferative retinopathy generated superoxide anion at significantly higher rates than did those from diabetics with nonproliferative retinopathy or without retinopathy (3.8 +/- 1.5, 2.08 +/- 2.1 and 1.3 +/- 1.6 nmol/10(5) cells/60 min respectively; p = 0.005). These results suggest that reactive oxygen species produced by PMNs may play a role in the progression of diabetic retinopathy.