Mechanism of the preventive effect of sitagliptin upon chances induced by short term fructosa administration in liver glucokinase activity

MASSA ML; FRANCINI F; CASTRO MC; GAGLIARDINO JJ

Lisboa

Congreso; 47th European Association for the Study of Diabtes Annual Meeting; 2011

Background and aims: While the effect of exendin-4 (GLP-1R agonist) and DPP-IV inhibitors upon islet hormones secretion, â-cell mass, appetite and gastric emptying delay has been widely reported, little is known about their effect upon liver glucokinase (GK-l) activity and their physiologic modulators. The aim of this study was to evaluate the effect of exendin-4 and sitagliptin upon fructose (Fr)-induced changes in GK-l activity and the regulatory mechanism of such activity (genic and protein level, cellular compartmentation and interaction with PFK2). Materials and methods: Adult male Wistar rats received (3 weeks) a standard commercial diet, without (C) or with 10% Fr (w/v) in the drinking water (F). C and F were daily treated either with sitagliptin (115 mg/day per rat) (CS and FS) or exendin-4 (0.35 nmol/kg of body weight) (CE and FE). After such treatment, we measured: 1) Serum glucose (G) (enzymatic method), insulin (I) (RIA) and triglyceride (TG) (enzymatic method) levels; 2) GK-l activity in both cytosolic (CF) and nuclear (DNF) fractions (enzyme-coupled photometric assay), 3) GK-l and PFK2 mRNA (qPCR) and 4) protein GK-l and PFK2 concentration (Western blot [Wb]). Results: C CE CS F FE FS G (mM) 4.5±0.1 4.2±0.1 4.5±0.2 4.9±0.2 4.2±0.2 4.7±0.3 TG (mM) 0.47±0.01 0.48±0.06 0.63±0.09 1.14±0.12?? 0.72±0.09* 0.67±0.05** I (ng/ml) 0.30±0.02 0.25±0.03 0.27±0.03 1.09±0.28? 0.30±0.05* 0.31±0.05* Values are means ± SEM (n =20). TG: ?? F vs. C, P < 0.0001; * FE vs. F, P < 0.003; ** FS vs. F, P < 0.001.  I: ? F vs. C, P < 0.02; * FE or FS vs. F, P <0.03. F induced a significat increase of total GK-l activity that was prevented by administration of either exendin-4 or sitagliptin. Most of the GK-l activity was measured in the DNF in C rats but in the CF in F rats. Exendin-4 and sitagliptin administration to F rats prevented such compartmentation changes. GK-l mRNA concentration was significantly higher in F rats and both exendin-4 and sitagliptin administration turn down these increase to values comparable to those measured in C rats. PFK2 mRNA concentration undergo similar changes. GK-l and PFK2 protein expression (Wb) were significantly higher in F rats. While exendin-4 and sitagliptin administration to F did not modify GK-l protein concentration, it induced a significant decrease in PFK2 concentration, suggesting that their effect on GK-l activity would be associated to the latter action. Conclusions: F administration to normal rats induced a) significant increase of serum TG and I levels but not of fasting blood G; administration of either exendin-4 or sitagliptin to F prevented the development of such changes; b) the significant increase in GK-l activity measured in F would result from a combination of an increased protein synthesis, a switch in its cellular compartmentation towards the cytosol, and an increase in PFK2 protein level. Exendin-4/sitagliptin administration prevented the F-induced changes in GK-l activity by affecting PFK2 concentration and GK-l compartmentation rather than GK-l protein level.