Mechanism of action of 2,5-anhydro-D-mannitol in hepatocytes. Effects of phosphorylated metabolites on enzymes of carbohydrate metabolism.

Isolated rat hepatocytes convert 2,5-anhydromannitol to 2,5-anhydromannitol-1-P and 2,5-anhydromannitol-1,6-P2. Cellular concentrations of the monophosphate and bisphosphate are proportional to the concentration of 2,5-anhydromannitol and are decreased by gluconeogenic substrates but not by glucose. Rat liver phosphofructokinase-1 phosphorylates 2,5-anhydromannitol-1-P; the rate is less than that for fructose-6-P but is stimulated by fructose-2,6-P2. At 1 mM fructose-6-P, bisphosphate compounds activate rat liver phosphofructokinase-1 in the following order of effectiveness: fructose-2,6-P2 much greater than 2,5-anhydromannitol-1,6-P2 greater than fructose-1,6-P2 greater than 2,5-anhydroglucitol-1,6-P2. High concentrations of fructose-1,6-P2 or 2,5-anhydromannitol-1,6-P2 inhibit phosphofructokinase-1. Rat liver fructose 1,6-bisphosphatase is inhibited competitively by 2,5-anhydromannitol-1,6-P2 and noncompetitively by 2,5-anhydroglucitol-1,6-P2. The AMP inhibition of fructose 1,6-bisphosphatase is potentiated by 2,5-anhydroglucitol-1,6-P2 but not by 2,5-anhydromannitol-1,6-P2. Rat liver pyruvate kinase is stimulated by micromolar concentrations of 2,5-anhydromannitol-1,6-P2; the maximal activation is the same as for fructose-1,6-P2. 2,5-Anhydroglucitol-1,6-P2 is a weak activator. 2,5-Anhydromannitol-1-P stimulates pyruvate kinase more effectively than fructose-1-P. Effects of glucagon on pyruvate kinase are not altered by prior treatment of hepatocytes with 2,5-anhydromannitol. Pyruvate kinase from glucagon-treated hepatocytes has the same activity as the control pyruvate kinase at saturating concentrations of 2,5-anhydromannitol-1,6-P2 but has a decreased affinity for 2,5-anhydromannitol-1,6-P2 and is not stimulated by 2,5-anhydromannitol-1-P. The inhibition of gluconeogenesis and enhancement of glycolysis from gluconeogenic precursors in hepatocytes treated with 2,5-anhydromannitol can be explained by an inhibition of fructose 1,6-bisphosphatase, an activation of pyruvate kinase, and an abolition of the influence of phosphorylation on pyruvate kinase.