Insulin and high glucose modulation of phosphatase and reductase enzymes in the human erythrocytes: a comparative analysis in normal and diabetic states

The ability of insulin to influence activities of various protein kinases a nd protein phosphatases. that are thought to mediate insulin action, are li mited in patients with insulin resistance. Because numerous responses to in sulin are affected, we undertook studies to determine whether protein tyros ine phosphatases (PTPs) activities are altered in patients with diabetes sy ndrome. In order to evaluate abnormal PTP activities, we done a comparative study using erythrocytes from normal and diabetic patients. We determined the activity of the cytosolic acid PTP in basal and insulin-dependent state s. Mean basal PTP activities, were found to be significantly higher in diab etics than in normal subjects (type 1 diabetics: 0.36 +/- 0.01 vs 0.28 +/- 0.01 mmol p-nitrophenolate/h per g hemoglobin (Hb), P < 0.001; type 2 diabe tics: 0.35 +/- 0.01 vs 0.28 +/- 0.01 mmol p-nitrophenolate/h per g Hb, P < 0.001). Insulin, at concentrations above physiological levels (1 mIU/ml), i nhibited the PTP activities in erythrocytes from normal subjects (- 15 +/- 4.1%, P < 0.01). Insulin could also modulate glycolysis, probably as a cons equence of receptor tyrosine kinase activation, inducing phosphorylation of protein band 3 and hence the release of glycolytic enzymes. We have previo usly reported that a reductase enzyme in human erythrocytes is dependent on glycolysis being significantly activated (+ 28 +/- 3.1%, P < 0.001) by hig h insulin levels (1 mIU/ml). Mean basal reductase activities were found to be significantly lower in diabetics than in normal subjects (type 1 diabeti cs: 0.77 +/- 0.03 vs 0.97 +/- 0.02 mmol ferrocyanide/20 min per 1 cells, P < 0.001; type 2 diabetics: 0.77 +/- 0.04 vs 0.97 +/- 0.02 mmol ferrocyanide /20 min per 1 cells, P < 0.001), indicating altered erythrocyte metabolism in the diabetic patients. High glucose levels were used to mimic hyperglyce mia condition, using erythrocytes from normal subjects. At 30 mM glucose, e rythrocytic phosphatase activity was stimulated (+ 32 +/- 4.2%, P < 0.0001) , although no effect was observed on the reductase enzyme at the same gluco se levels. Results indicated that diabetic disorders appear to be associate d with quantitative alterations of erythrocyte acid phosphatase activity an d other enzymes that depend on the glycolytic rate (reductase). The overall data suggest that erythrocyte acid phosphatase may have a role in the modu lation of glycolytic rates through the control of insulin receptor phosphor ylation. (C) 2000 Elsevier Science Ireland Ltd. All rights reserved.