Maximum activities of some enzymes of glycolysis, the tricarboxylic acid cycle and ketone-body and glutamine utilization pathways in lymphocytes of the rat

Abstract

1. The maximum activity of hexokinase in lymphocytes is similar to that of 6-phosphofructokinase, but considerably greater than that of phosphorylase, suggesting that glucose rather than glycogen is the major carbohydrate fuel for these cells. Starvation increased slightly the activities of some of the glycolytic enzymes. A local immunological challenge in vivo (a graft-versus-host reaction) increased the activities of hexokinase, 6-phosphofructokinase, pyruvate kinase and lactate dehydrogenase, confirming the importance of the glycolytic pathway in cell division. 2. The activities of the ketone-body-utilizing enzymes were lower than those of hexokinase or 6-phosphofructokinase, unlike in muscle and brain, and were not affected by starvation. It is suggested that the ketone bodies will not provide a quantitatively important alternative fuel to glucose in lymphocytes. 3. Of the enzymes of the tricarboxylic acid cycle whose activities were measured, that of oxoglutarate dehydrogenase was the lowest, yet its activity (about 4.0mumol/min per g dry wt. at 37 degrees C) was considerably greater than the flux through the cycle (0.5mumol/min per g calculated from oxygen consumption by incubated lymphocytes). The activity was decreased by starvation, but that of citrate synthase was increased by the local immunological challenge in vivo. It is suggested that the rate of the cycle would increase towards the capacity indicated by oxoglutarate dehydrogenase in proliferating lymphocytes. 4. Enzymes possibly involved in the pathway of glutamine oxidation were measured in lymphocytes, which suggests that an aminotransferase reaction(s) (probably aspartate aminotransferase) is important in the conversion of glutamate into oxoglutarate rather than glutamate dehydrogenase, and that the maximum activity of glutaminase is markedly in excess of the rate of glutamine utilization by incubated lymphocytes. The activity of glutaminase is increased by both starvation and the local immunological challenge in vivo. This last finding suggests that metabolism of glutamine via glutaminase is important in proliferating lymphocytes.

Fuel utilization in colonocytes of the rat

“…The activity of hexokinase in colonic mucosa is similar to that reported for intestinal mucosa (Hanson & Carrington, 1981;Budohoski et al, 1982) and lymphocytes (Ardawi & Newsholme, 1982). The activity of 6-phosphofructokinase is similar to that of hexokinase, supporting the view that glucose is the major carbohydrate that is used by this tissue.…”

“…lymphocytes, thymocytes, tumour cells), these epithelial cells are also characterized by a high rate of glycolysis in either the fed or the starved state (Watford et al, 1979;Hume et al, 1978;Ardawi & Newsholme, 1983a). Experiments with incubated lymphocytes suggested that most of the energy requirements in either the fed or the starved state were provided from the utilization of fatty acids, glutamine and glucose, whereas ketone bodies appeared to be much less important (Ardawi & Newsholme, 1982, 1983a,b, 1984a,b, 1985. Experiments with incubated colonocytes suggests that most of the energy requirements were provided from the oxidation of fatty acids, ketone bodies and/or glutamine (Roediger, 1982).…”

“…4.1 ,tmol/min per g dry wt., or about 25 % of the maximum activity of oxoglutarate dehydrogenase when calculated at 37°C (Table 3). If the maximum activity of oxoglutarate dehydrogenase indicates the maximum capacity of the cycle in colonocytes, this finding suggests that the rate of the cycle could be markedly increased in other conditions, perhaps during proliferation (as suggested for lymphocytes; see Ardawi & Newsholme, 1982). Adenine nucleotide concentrations These were measured in colonic mucosa or isolated colonocytes as soon as possible after scraping or preparation and then at various times after incubation in the presence of normal incubation medium containing 10 mM-glucose.…”

“…The extraction media for all enzymes studied were as described previously (see Ardawi & Newsholme, 1982), except for fructose-bisphosphatase, which was extracted in a medium as described by Budohoski et al (1982). Assay of enzyme activities Enzyme activities were measured as described in the cited references: hexokinase ; glucose-6-phosphatase (Lackner et al, 1984); 6-phosphofructokinase (Opie & Newsholme, 1967); fructose-bisphosphatase 3-oxoacid CoA-transferase, acetoacetyl-CoA thiolase and D-3-hydroxybutyrate dehydrogenase (Williamson et al, 1971); glutaminase (Curthoys & Lowry, 1973); glutamate dehydrogenase ; alanine aminotransferase and aspartate aminotransferase (Sugden & Newsholme, 1975).…”

“…However, the pathway for the utilization and the metabolic interrelationship between these fuels in colonocytes have not been studied. Hence, to provide further information about metabolism ofthese fuels in colonocytes, the maximum catalytic activities of key enzymes in the pathways of glucose, glutamine and ketone-body metabolism were measured in colonic mucosa in a similar manner to that done for lymphoid tissues (Ardawi & Newsholme, 1982). The rates of utilization and the metabolic fate of glucose, glutamine, acetoacetate and n-butyrate in incubated colonocytes have been investigated by measurement of the intermediates that accumulate on incubation of these cells with the various fuels.…”