Figure 6. Effect of transfection of dn-PKC-α on insulin secretion.

A, detection on PKC-α expression in cultured pancreatic islets with or without Adex-dn-PKC-α transfection. Islets sections were double-stained with antibodies against PKC-α (red) and insulin (green). The Adex-dn-PKC-α expression (A3) detected was apparently larger than the endogenous PKC-α (A1) both in the periphery and in the core while there is no apparent changes in insulin expression in the control islet (A2, the same islet as in A1) and Adex-dn-PKC-α-expressing islet (A4, the same islet as in A3). Magnification × 200. The figure is representative of 40 islets detected from three independent experiments. B, detection of PKC-α in islet.The expression of PKC-α was determined by immunoblotting in control islets, and islets transfected with Adex-LacZ or Adex-dn-PKC-α. The results are representative of two experiments. C, 10 min insulin secretion from dn-PKC-α transfected islets. Islets were transfected with Adex-LacZ (open column) or Adex-dn-PKC-α (filled column) or without adenovirus transfection (stippled column). After 24 h incubation, the medium was changed to normal extracellular solution containing 3 mm glucose (G3) or 40 mm KCl (K40) or 20 mm glucose (G20). Values are the means ± s.e.m. for 10 experiments. D, 1 h insulin secretion from dn-PKC-α transfected islets. Islets were transfected with Adex-LacZ (open column) or Adex-dn-PKC-α (filled column) or without adenovirus transfection (stippled column). Islets were then incubated for 1 h with 3 mm glucose (G3), 40 mm KCl (K40) or 20 mm glucose (G20). Values are the means ± s.e.m. for 10 experiments. E, effect of dn-PKC-α on MARCKS phosphorylation. Islets transfected by dn-PKC-α or LacZ were incubated for 10 min in the presence or absence of 500 nm PMA. MARCKS phosphorylation (p-MARCKS) was determined by immunoblotting. The figure is a representative immunoblot from two independent experiments.*P < 0.05 versus Adex-LacZ, **P < 0.01 versus Adex-LacZ.