Regulation of carbohydrate metabolism by 2,5-anhydro-D-mannitol.

Abstract

In hepatocytes isolated from fasted rats, 2,5-anhydromannitol inhibits gluconeogenesis from lactate plus pyruvate and from substrates that enter the gluconeogenic pathway as triose phosphate. This fructose analog has no effect, however, on gluconeogenesis from xylitol, a substrate that enters the pathway primarily as fructose 6-phosphate. The sensitivity of gluconeogenesis to 2,5-anhydromannitol depends on the substrate metabolized; concentrations of 2,5-anhydromannitol required for 50% inhibition increase in the order lactate plus pyruvate < dihydroxyacetone < glycerol < sorbitol < fructose. The inhibition by 2,5-anhydromannitol of gluconeogenesis from dihydroxyacetone is accompanied by an increase in lactate formation and by two distinct crossovers in gluconeogenic-glycolytic metabolite patterns i.e., increases in pyruvate concentrations with decreases in phosphoenolpyruvate and increases in fructose-1,6-bisphosphate concentrations with little change in fructose 6-phosphate. In addition, 2,5-anhydromannitol blocks the ability of glucagon to stimulate gluconeogenesis and inhibit lactate production from dihydroxyacetone. 2,5-Anhydromannitol decreases cellular fructose 2,6-bisphosphate content in hepatocytes; therefore the effects of the fructose analog are not mediated by fructose 2,6-bisphosphate, a naturally occurring allosteric regulator. 2,5-Anhydromannitol also inhibits gluconeogenesis in hepatocytes isolated from fasted diabetic rats, but higher concentrations of the analog are required.2,5-Anhydro-D-mannitol (2,5-AM-ol), an analog of P-D-fructose locked in the furan ring structure, is phosphorylated by fructokinase to form 2,5-AM-ol-l-P (1, 2).Because 2,5-AM-ol is symmetrical, the monophosphate product can be considered an analog of both fructose-i-P and fructose-6-P (3). Because of the stability of its ring structure, 2,5-AMI-ol monophosphate cannot be cleaved bv aldolase in a manner similar to that of fructose-l-P, nor can it act as a substrate for phosphoglucoisomerase and be converted to glucose-6-P in a manner similar to that of fructose-6-P. 2,5-AM-ol monophosphate is a substrate for phosphofructokinase 1 (4, 5). The resulting product, 2,5-AM-ol bisphosphate, is an analog of 8-fructose-1,6-P2 rather than a-fructose-1,6-P2 and, as such, is not hydrolyzed readily by fructose-1,6-bisphosphatase, which prefers the a anomer (6). The bisphosphate compound, thus, should accumulate within the cell. In vitro experiments have shown that 2,5-AM-ol bisphosphate is a competitive inhibitor of fructose-1,6-bisphosphatase (7,8). In view of the described findings, the potential of 2,5-AM-ol to act as a regulator of gluconeogenesis and glycolysis was examined in isolated rat hepatocytes.METHODS AND MATERIALS Synthesis of 2,5-AM-ol. 2,5-AM-ol was prepared as in ref.9 except the crystallization step was omitted. The crude 2,5-AM1-ol, in 5 mM ammonium borate (pH 9), was purified on Dowex-I-X-8 (borate) (10). The unretained material, after deionization and repeated concentration by evaporation of methanol, showe...

Acetaldehyde Stimulation of Net Gluconeogenic Carbon Movement from Applied Malic Acid in Tomato Fruit Pericarp Tissue

“…To ascertain that the arrested label mobilization is caused by the Fru 2,6-P2 inhibitory action, rather than metabolic abnormality, we applied also 2,5-AM-ol. The compound is known to arrest gluconeogenesis in isolated rat hepatocytes by inhibiting the activity of fructose-1,6-bisphosphatase and also of pyruvate kinase (22). In tomato tissue the applied 2,5-AM-ol (Fig.…”

Phosphate and Succinate Use Different Mechanisms to Inhibit Sugar-induced Cell Death in Yeast

“…Instead of using Fru-1,6-P 2 , we used the cell-permeable analog 2,5-AM, which enters cells and is phosphorylated to yield 2,5-AM-ol bisphosphate (30), which accumulates in the cytosol because it is not metabolized. 2,5-AM increased the survival (5.5-fold) of stationary-phase cells in glucose/water and inhibited ROS accumulation and O 2 consumption (Fig.…”

Inhibition by 2,5-anhydromannitol of glycolysis in isolated rat hepatocytes and in Ehrlich ascites cells.

“…Incubation of isolated rat hepatocytes with 2,5-AMol results in a rapid increase in 2,5-AM-ol-1-P and a slower accumulation of 2,5-AM-ol-1,6-P2 (2). Under these conditions, the intracellular concentration of fructose-2,6-P2 (Fru-2,6-P2), a potent activator of phosphofructokinase-1 (PFK-1), is decreased (1), probably because of the accumulation of 2,5-AM-ol-1-P. 2,5-AM-ol-1,6-P2 is a poor substitute for Fru-2,6-P2 as an activator of PFK-1, and at high concentrations it can act as a product inhibitor (2). Since the formation of either of the phosphorylated metabolites of 2,5-AM-ol could cause an inhibition of flux through PFK-1, the effects of 2,5-AM-ol on glycolysis in isolated rat hepatocytes and in Ehrlich ascites tumor cells were examined.…”

“…1). Incubation of isolated rat hepatocytes with 2,5-AMol results in a rapid increase in 2,5-AM-ol-1-P and a slower accumulation of 2,5-AM-ol-1,6-P2 (2). Under these conditions, the intracellular concentration of fructose-2,6-P2 (Fru-2,6-P2), a potent activator of phosphofructokinase-1 (PFK-1), is decreased (1), probably because of the accumulation of 2,5-AM-ol-1-P. 2,5-AM-ol-1,6-P2 is a poor substitute for Fru-2,6-P2 as an activator of PFK-1, and at high concentrations it can act as a product inhibitor (2).…”

“…1). Incubation of isolated rat hepatocytes with 2,5-AMol results in a rapid increase in 2,5-AM-ol-1-P and a slower accumulation of 2,5-AM-ol-1,6-P2 (2).…”