Research into the metabolic adaptations of glucose and fatty acid metabolism during the perinatal period and the suckling-weaning transition is summarized.
Background & Aims:
Among substrates available to the colonic mucosa,
n-butyrate from bacterial origin represents a major fuel. The present work investigated possible modifications of energy substrate ...metabolism in colonocytes isolated from germfree rats.
Methods:
Colonocytes isolated from germfree vs. conventional rats were incubated (30 minutes at 37°C) in the presence of
14C-labeled
n-butyrate (10 mmol/L), glucose (5 mmol/L), or glutamine (5 mmol/L).
14CO
2 and metabolites generated were measured. Possible regulatory steps were also investigated.
Results:
Glucose use rate was 25% lower in germfree rat colonocytes due to a reduced glycolytic capacity in these cells. Differences in 6-phosphofructo-1-kinase activity could account for this decrease. In contrast, glutamine use rate was 45% higher, and this was correlated with a higher maximum velocity of glutaminase in these cells. Nevertheless, the capacities to oxidize glucose and glutamine remained unchanged. Although the capacity to use
n-butyrate was maintained in colonocytes of germfree rats, the ketogenic capacity was lower, whereas the capacity to oxidize
n-butyrate was higher. The mitochondrial 3-hydroxy-3-methylglutaryl-coenzyme A synthase protein was identified in the colonic mucosa. Moreover, the messenger RNA and amount of protein were 75% lower in the germfree state.
Conclusions:
The absence of an intestinal microflora induces specific changes in the metabolic capacities of colonocytes.
In newborn-pig hepatocytes, the rate of oleate oxidation is extremely low, despite a very low malonyl-CoA concentration. By contrast, the sensitivity of carnitine palmitoyltransferase (CPT) I to ...malonyl-CoA inhibition is high, as suggested by the very low concentration of malonyl-CoA required for 50% inhibition of CPT I IC(50). The rates of oleate oxidation and ketogenesis are respectively 70 and 80% lower in mitochondria isolated from newborn-pig liver than from starved-adult-rat liver mitochondria. Using polarographic measurements, we showed that the oxidation of oleoyl-CoA and palmitoyl-L-carnitine is very low when the acetyl-CoA produced is channelled into the hydroxymethylglutaryl-CoA (HMG-CoA) pathway by addition of malonate. In contrast, the oxidation of the same substrates is high when the acetyl-CoA produced is directed towards the citric acid cycle by addition of malate. We demonstrate that the limitation of ketogenesis in newborn-pig liver is due to a very low amount and activity of mitochondrial HMG-CoA synthase as compared with rat liver mitochondria, and suggest that this could promote the accumulation of acetyl-CoA and/or beta-oxidation products that in turn would decrease the overall rate of fatty acid oxidation in newborn- and adult-pig livers.
The temporal changes in oleate oxidation, lipogenesis, malonyl-CoA concentration and sensitivity of carnitine palmitoyltransferase I (CPT 1) to malonyl-CoA inhibition were studied in isolated rabbit ...hepatocytes and mitochondria as a function of time after birth of the animal or time in culture after exposure to glucagon, cyclic AMP or insulin. (1) Oleate oxidation was very low during the first 6 h after birth, whereas lipogenesis rate and malonyl-CoA concentration decreased rapidly during this period to reach levels as low as those found in 24-h-old newborns that show active oleate oxidation. (2) The changes in the activity of CPT I and the IC50 (concn. causing 50% inhibition) for malonyl-CoA paralleled those of oleate oxidation. (3) In cultured fetal hepatocytes, the addition of glucagon or cyclic AMP reproduced the changes that occur spontaneously after birth. A 12 h exposure to glucagon or cyclic AMP was sufficient to inhibit lipogenesis totally and to cause a decrease in malonyl-CoA concentration, but a 24 h exposure was required to induce oleate oxidation. (4) The induction of oleate oxidation by glucagon or cyclic AMP is triggered by the fall in the malonyl-CoA sensitivity of CPT I. (5) In cultured hepatocytes from 24 h-old newborns, the addition of insulin inhibits no more than 30% of the high oleate oxidation, whereas it stimulates lipogenesis and increases malonyl-CoA concentration by 4-fold more than in fetal cells (no oleate oxidation). This poor effect of insulin on oleate oxidation seems to be due to the inability of the hormone to increase the sensitivity of CPT I sufficiently. Altogether, these results suggest that the malonyl-CoA sensitivity of CPT I is the major site of regulation during the induction of fatty acid oxidation in the fetal rabbit liver.
The effects of pancreatic hormones and cyclic AMP on the induction of ketogenesis and long-chain fatty acid oxidation were studied in primary cultures of hepatocytes from fetal and newborn rabbits. ...Hepatocytes were cultivated during 4 days in the presence of glucagon (10(-6) M), forskolin (2 x 10(-5) M), dibutyryl cyclic AMP (10(-4) M), 8-bromo cyclic AMP (10(-4) M) or insulin (10(-7) M). Ketogenesis and fatty acid metabolism were measured using 1-14Coleate (0.5 mM). In hepatocytes from fetuses at term, the rate of ketogenesis remained very low during the 4 days of culture. In hepatocytes from 24-h-old newborn, the rate of ketogenesis was high during the first 48 h of culture and then rapidly decreased to reach a low value similar to that measured in cultured hepatocytes from term fetuses. A 48 h exposure to glucagon, forskolin or cyclic AMP derivatives is necessary to induce ketone body production in cultured fetal hepatocytes at a rate similar to that found in cultured hepatocytes from newborn rabbits. In fetal liver cells, the induction of ketogenesis by glucagon or cyclic AMP results from changes in the partitioning of long-chain fatty acid from esterification towards oxidation. Indeed, glucagon, forskolin and cyclic AMP enhance oleate oxidation (basal, 12.7 +/- 1.6; glucagon, 50.0 +/- 5.5; forskolin, 70.6 +/- 5.4; cyclic AMP, 77.5 +/- 3.4% of oleate metabolized) at the expense of oleate esterification. In cultured fetal hepatocytes, the rate of fatty acid oxidation in the presence of cyclic AMP is similar to the rate of oleate oxidation present at the time of plating (85.1 +/- 2.6% of oleate metabolized) in newborn rabbit hepatocytes. In hepatocytes from term fetuses, the presence of insulin antagonizes in a dose-dependent fashion the glucagon-induced oleate oxidation. Neither glucagon nor cyclic AMP affect the activity of carnitine palmitoyltransferase I (CPT I). The malonyl-CoA concentration inducing 50% inhibition of CPT I (IC50) is 14-fold higher in mitochondria isolated from cultured newborn hepatocytes (0.95 microM) compared with fetal hepatocytes (0.07 microM), indicating that the sensitivity of CPT I decreases markedly in the first 24 h after birth. The addition of glucagon or cyclic AMP into cultured fetal hepatocytes decreased by 80% and 90% respectively the sensitivity of CPT I to malonyl-CoA inhibition. In the presence of cyclic AMP, the sensitivity of CPT I to malonyl-CoA inhibition in cultured fetal hepatocytes is very similar to that measured in cultured hepatocytes from 24-h-old newborns.
Fatty acid oxidation and synthesis were studied in isolated hepatocytes from adult rats adapted for 44 days on low-fat, high-carbohydrate (LF), diet or high-fat diets, composed of long-chain (LCT) or ...medium-chain (MCT) triacylglycerols. The rates of 1-14Coctanoate oxidation were almost similar in each group studied, whereas the oxidation of 1-14Coleate was 50% lower in the LF group than in animals adapted to high-fat diets. The rates of oleate oxidation are inversely correlated with the rates of lipogenesis. However, it seems unlikely that malonyl-CoA itself represents the sole mechanism involved in the regulation of oleate oxidation during long-term LCT or MCT feeding, since: (1) despite a 3-fold higher concentration of malonyl-CoA in MCT-fed rats than in LCT-fed ones, the rates of oleate oxidation are similar; (2) when malonyl-CoA concentration is increased after stimulation of lipogenesis (by adding lactate + pyruvate) in MCT-fed rats, to a level comparable with that of the LF group, the rate of oleate oxidation remains 55% higher than that measured under similar conditions in the LF-fed rats; (3) in the LF group, the 90% decrease in malonyl-CoA concentration by 5-(tetradecyloxy)-2-furoic acid is not associated with a stimulation of oleate oxidation. By contrast, the sensitivity of carnitine palmitoyltransferase I (CPT I) to malonyl-CoA is markedly decreased in the LCT- and MCT-fed rats, by 90% and 70% respectively. The relevance of this decrease in the sensitivity of CPT I is discussed.
The aim of the present work was to examine in pigs the effect of a dietary supplementation with the flavor enhancer monosodium glutamate (MSG) on intestinal amino acid metabolism. For this purpose, ...pigs weighing 60 +/- 2 kg received a standard meal twice a day for 1 week, supplemented with either 10 g MSG per meal or, as control experiments, an isonitrogenous amount of glycine together with an equal amount of sodium in the form of NaCl, the animals being their own control in all experiments. At the end of this period, pigs received a MSG or glycine-NaCl-supplemented meal and samples of portal and arterial blood were collected for amino acid analysis in plasma. The results demonstrate after MSG supplementation rapid significant increases in glutamate concentration in the portal and arterial blood plasma after a test meal which resulted in a positive portoarterial difference. In comparison, after glycine-NaCl supplementation, glutamate concentrations were almost identical in portal and arterial plasma. Furthermore, significant increased aspartate concentration in the portal blood plasma was observed after MSG supplementation when compared with control experiments. When enterocytes were isolated at the end of the supplementation period from the jejunum and examined for their metabolic capacities towards L-glutamate and L-glutamine, it was found that metabolism did not differ according to the supplement used, with glutamate and glutamine being oxidized and transaminated at a similar level. It is concluded that the portal hyperglutamatemia observed shortly after the ingestion of a MSG- supplemented meal is likely due to the saturation of the intestinal capacity to metabolize glutamate with no measurable adaptation of the metabolic pathways controlling glutamate metabolism in enterocytes.
Background: The rate of absorption of glucose from carbohydrates is important in several aspects of health. We recently validated a non-invasive technique in pigs, euglycemic hyperinsulinemic clamp ...plus oral carbohydrate loading (OC-Clamp), to quantify the rate of net posthepatic appearance of glucose after ingestion of carbohydrates. Objective: The OC-Clamp procedure was performed in 8 healthy men to compare the net posthepatic appearance of glucose after ingestion of 1 of 3 carbohydrates. Design: Human volunteers underwent the OC-Clamp procedure at an insulin infusion rate of 1.5 mU(.)kg(-1)(.)min(-1) (n = 5). The oral carbohydrate load (1 g/kg) consisted of glucose, cornstarch, or mung bean starch. During the OC-Clamp procedure, the glucose infusion rate decreased during absorption to maintain plasma glucose steady state and the decrease reflected the net posthepatic appearance of glucose. In addition, carbohydrates were loaded without insulin infusion (n = 6) and glycemic indexes were calculated (with glucose as the reference). Results: The mean (+/- SEM) glycemic index of cornstarch was higher (95 +/- 18) than that of mung bean starch (51 +/- 13). In the OC-Clamp experiments, the posthepatic appearance of glucose and cornstarch did not differ significantly and represented 79.4 +/- 5.0% and 72.6 +/- 14.0%, respectively, of the load after complete absorption (within 3 h). In contrast, the net posthepatic appearance of glucose from mung bean starch was significantly lower (35.6 +/- 4.6% of the load, P < 0.001) than that from glucose and cornstarch, even 4.5 h postprandially. Conclusions: The OC-Clamp technique allows a continuous assessment of net posthepatic appearance of glucose after ingestion of carbohydrates and significant discrimination between corn and mung bean starches.
Ketogenesis from endogenous fatty acids or exogenous oleate plus carnitine has been studied in isolated hepatocytes from fetal, newborn, and 70-day-old rabbits. During the first 48 h after birth, ...hepatic triacylglycerol stores decrease by 80%. The hydrolysis of hepatic triacylglycerol stores has been studied in isolated hepatocytes from 24-h-old fasting rabbits by using lysosomal acid lipase inhibitors and lysosomotropic agents. Their addition decreases the rates of ketone body production by 60-70%, suggesting that hepatic triacylglycerol hydrolysis proceeds via an acid lipase located in the lysosomes. Whereas the rates of ketogenesis from endogenous or exogenous fatty acids are very low in isolated hepatocytes from fetal rabbit, an eightfold increase in the rate of ketogenesis occurs between 6 and 24 h after birth; furthermore the hydrolysis of triacylglycerol stores is sufficient to support the ketogenic capacity in the hepatocytes isolated from 24-h-old rabbits. The emergence of ketogenesis in newborn rabbit hepatocytes is triggered by birth-associated factors rather than to an accurate stage of fetal maturation. Fatty acids are mainly oxidized in the mitochondria because peroxisomal oxidation does not exceed 10-15% of the overall beta-oxidation. Isolated hepatocytes incubated with 1-14Coleate exhibit at birth a preferential channeling of fatty acid into esterification (93% of oleate metabolized) rather than into oxidation. Conversely oleate oxidation represents 50% of total oleate metabolized 24 h after birth. Factors involved in this switch on of the partition of oleate into esterification and oxidation during the 1st day after birth are discussed.
PDF
