N-terminal glycation of cholecystokinin-8 abolishes its insulinotropic action on clonal pancreatic B-cells

Monoglycated cholecystokinin octapeptide (Asp(1)-glucitol CCK-X) was prepar ed under hyperglycaemic reducing conditions and purified by reverse phase-h igh performance liquid chromatography. Electrospray ionisation mass spectro metry and automated Edman degradation demonstrated that CCK-8 was glycated specifically at the amino-terminal Asp(1) residue. Effects of Asp(1)-glucit ol CCK-8 and CCK-8 on insulin secretion were examined using glucose-respons ive clonal BRIN-BD11 cells. In acute (20 min) incubations, 10(-10) mol/l CC K-8 enhanced insulin release by 1.2-1.5-fold at 5.6-11.1 mmol/l glucose. Th e stimulatory effect induced by 10(-10) mom CCK-8 was abolished following g lycation. At 5.6 mmol/l glucose, CCK-8 at concentrations ranging from 10(-1 1) to 10(-7) mol/l induced a significant 1.6-1.9-fold increase in insulin s ecretion. Insulin output in the presence of Asp(1)-glucitol CCK-8 over the concentration range 10(-11)-10(-7) mol/l was decreased by 21-35% compared w ith CCK-8, and its insulinotropic action was effectively abolished. Asp(1)- glucitol CCK-8 at 10(-8) mol/l also completely blocked the stimulatory effe cts of 10(-11)-10(-8) mol/l CCK-8. These data indicate that structural modi fication by glycation at the amino-terminal Asp(1) residue effectively abol ishes and/or antagonises the insulinotropic activity of CCK-8. (C) 1999 Els evier Science B.V. All rights reserved.