In this study, we found that the ratio of proinsulin to total immunoreactive insulin was much higher in 22 patients with Type 2 (non-insulin-dependent) diabetes mellitus than in 28 non-diabetic control subjects of similar age and adiposity (32±3 vs 15±1%, p<0.001). In addition, the arginine-induced acute proinsulin response to total immunoreactive insulin response ratio was greater in diabetic patients (n=10) than in control subjects (n=9) (8±2 vs 2±0.5%, p=0.009), suggesting that increased islet secretion per se accounted for the increased ratio of proinsulin to immunoreactive insulin. One explanation for these findings is that increased demand for insulin in the presence of islet dysfunction leads to a greater proportion of proinsulin secreted from the B cell. We tested this hypothesis by comparing proinsulin secretion before and during dexamethasone-induced insulin resistance in diabetic patients and control subjects. Dexamethasone treatment (6 mg/day for 3 days) raised the proinsulin to immunoreactive insulin ratio in control subjects from 13±2 to 21±2% (p<0.0001) and in diabetic patients from 29±5 to 52±7% (p<0.001). Dexamethasone also raised the ratio of the acute proinsulin response to the acute immunoreactive insulin response in control subjects from 2±0.5 to 5±2% (p=0.01) and in diabetic patients from 8±2 to 14±4% (p=NS), suggesting that the dexamethasone-induced increment in the basal ratio of proinsulin to immunoreactive insulin was also due to increased secretion. We conclude that: (1) The basal proinsulin to immunoreactive insulin ratio is increased in obese Type 2 diabetic patients. (2) An increase in tissue demand for insulin leads to a rise in the proinsulin to immunoreactive insulin ratio, which is exaggerated in Type 2 diabetic patients. (3) The increased proinsulin to immunoreactive insulin ratio in these diabetic patients in the basal state and in diabetic patients and control subjects during experimental insulin resistance is probably due to increased B-cell secretion of proinsulin.