Fig. 4

Hypoxia-induced increase in ins-PRS-protein binding activity is counteracted by hydrogen peroxide but not affected by nitric oxide. The binding activity was measured by EMSA. The ³²P-labelled insulin mRNA-probe, ins-PRS, was incubated with rat islets extract, from islets incubated for 20 min in normoxic or hypoxic (1% O₂) 2.8 mM glucose medium, with or without hydrogen peroxide (50 µM) or the nitric oxide donor DETA NONOate (2.5 mM). The reactions were performed with (11 mM) or without DTT and separated with nondenaturating polyacrylamide gel electrophoresis. (A) Ins-PRS binding activity during each condition was expressed as ratio between the shifted ins-PRS-protein complex without and with DTT. Results are mean ±SEM for six observations. Results were compared by Student paired t-test. *p < 0.05 when comparing versus corresponding control. (B) Representative autoradiograph showing EMSA analysis of ins-PRS binding activity. The white control bar in Figure 3A represents the ratio between lanes 1 and 2 in Figure 3B, and the black control bar represents the ratio between lanes 3 and 4 in Figure 3B, and so forth.