C. Coudray-Lucas

Is α-ketoisocaproyl-glutamine a suitable glutamine precursor to sustain fibroblast growth?

Abstract Background: Glutamine is considered an essential nutrient for cellular growth. Aim: To test the suitability of α-ketoisocaproyl-Gln (Kic-Gin) as a new glutamine (Gin) precursor to sustain human fibroblast growth. Methods: [3H] thymidine uptake into cellular DNA of human fibroblasts. Extracellular and intracellular aminoacid patterns were determined with peptides and acylated compounds. Results: L-alanyl-L-glutamine (used here as a recognized Gin precursor) promoted DNA synthesis, whileN-acetyl-L-glutamine (used here as a negative control since it is known to be a poor Gin precursor) and α-ketoisocaproyl-glutamine had no effect. Alanyl-glutamine progressively gave rise to free glutamine in the growth medium. In contrast, glutaminesupplied in acylated form was poorly available and did not appear in free form in the medium. In addition, only alanyl-glutamine increased intracellular glutamine and glutamate levels. In contrast,Kic-Gln was able to sustain net protein synthesis as judged by total protein content and reduced intracellular levels of most essential amino acids. Conclusion: Kic-Gln appears to be a poor extra-cellular precursor of Gin to sustain cell growth.

Action of ornithine alpha-ketoglutarate, ornithine hydrochloride, and calcium alpha-ketoglutarate on plasma amino acid and hormonal patterns in healthy subjects.

Ornithine alpha-ketoglutarate (OKG) has been useful as an adjuvant of enteral and parenteral nutrition. However, its metabolism and mechanism of action remain unclear although it is known that alpha-ketoglutarate (alpha KG) and ornithine (ORN) follow, in part, common metabolic pathways. Six fasting healthy male subjects underwent three separate oral load tests: (i) they received 10 g of OKG (i.e., 3.6 g of alpha KG and 6.4 g of ORN); (ii) 6.4 g of ORN as ornithine hydrochloride, and (iii) 3.6 g of alpha KG as calcium alpha-ketoglutarate. Blood was drawn 15 times over a five-hour period for measurements of plasma amino acids, alpha KG, insulin, and glucagon. After OKG and ORN administration, plasma ORN peaked at 60-75 min (494 +/- 91 and 541 +/- 85 mumol/L). The increase in plasma alpha KG was very small. OKG, alpha KG, and ORN all increased glutamate concentrations at 60 min (mean: +43%, +68%, +68%, respectively, p less than 0.05 compared to basal values). However, only OKG increased proline and arginine levels at 60 min (mean: +35%, p less than 0.01 and mean: +41%, p less than 0.05). Furthermore, glutamate, proline, and arginine concentrations correlated linearly with ornithine levels at 60 min. Finally, OKG increased insulinemia and glucagonemia (mean: +24% at 15 min, p less than 0.05 and +30% at 60 min, p less than 0.01, respectively). These data provide evidence that the combination of ORN and alpha KG modifies amino acid metabolism in a way which is not observed when they are administered separately. In addition, the OKG-mediated increase in insulin levels probably does not appear to result from a direct action of ORN on pancreatic secretion.

Action of enterally administered ornithine α-ketoglutarate on protein breakdown in skeletal muscle and liver of the burned rat

Several studies concerning burn patients have shown that supplementation of enteral nutrition with ornithine alpha-ketoglutarate (OKG) favorably modifies protein metabolism. Therefore, the effect of OKG administration on muscular and hepatic protein catabolism was evaluated in burned rats. Four groups of six rats were used. Two groups were scalded by immersion of the dorsum in water at 90 degrees C for 10 seconds and then starved for 24 hours. Controlled enteral nutrition was then administered in three boluses daily (Osmolite, 210 kcal/kg/d, 1.2 g N/kg/d); one group was supplemented with OKG (5 g/kg/d, ie, 0.68 g N/kg/d), while the other group received an equivalent amount of nitrogen in the form of glycine. One group of healthy control rats received Osmolite supplemented with glycine and the last group was fed ad libitum. The animals were killed after 2 days of nutrition. Protein catabolism was assessed in vitro by measuring the amount of valine (liver catabolism) and phenylalanine (muscle catabolism) released into the incubation medium of isolated tissues. Tissular and serum glutamine were also assayed. Burn injury induced muscle hypercatabolism without affecting hepatic catabolism. The administration of OKG limited both muscle weight loss and muscle protein hypercatabolism and significantly improved the muscle glutamine pool. These results demonstrate the nitrogen-sparing effect of OKG in muscle in hypercatabolic states.

Ornithine alpha-ketoglutarate counteracts thymus involution and glutamine depletion in endotoxemic rats.

This work studied the action of ornithine a-ketoglutarate (OKG) supplementation in an experimental model of endotoxemia in the rat. Male Wistar rats were injected intraperitoneally with lipopolysaccharide (LPS) from Escherichia coli (0127:B8). They were fasted for 24 h, then refed for 48 h with an enteral diet supplemented with either OKG (66 mg N x kg(-1) x d(-1)) or glycine, isonitrogenous to the OKG group. A control (sham) group was also studied. LPS treatment induced a decrease in thymus and muscle weights compared to controls, and a decrease in glutamine and arginine concentrations in the anterior tibialis muscle. Supplementation with OKG restored thymus weight and muscle arginine level and increased muscle glutamine concentration, when compared to controls. We conclude that OKG counteracts the thymic involution that occurs with endotoxemia, and restores the muscular content of glutamine and arginine, both of which are involved in the regulation of immune function.

Metabolism of ornithine, α-ketoglutarate and arginine in isolated perfused rat liver

: Ornithine (Orn; alpha-ketoglutarate (alpha KG) salt) and arginine (Arg) supplementation of enteral diets has been advocated in the treatment of hypercatabolism of trauma patients, but both compounds are subject to extensive hepatic metabolism. To compare the metabolism of these two compounds and to evaluate the possible influence of the alpha KG moiety, livers were perfused with alpha KG, Orn, ornithine alpha-ketoglutarate (OKG) or Arg (n 6 in each group) for 1 h. Arg uptake was nearly fourfold higher than Orn uptake (690 (SD 162) v. 178 (SD 30) nmol/min per g liver), and Orn uptake was not modified by alpha KG. Orn was totally metabolized by the liver, whereas Arg led to Orn release (408 (SD 159) nmol/min per g liver) and a threefold stimulation of urea production (Arg 1.44 (SD 0.22) v. Orn 0.45 (SD 0.09) mumol/min per g liver). alpha KG alone only increased hepatic aspartate uptake but, when associated with Orn as OKG, it led to an increase in glutamate release and in proline content in the liver and to a decrease in proline uptake. From these findings we conclude that (1) Arg load is extensively metabolized by the liver, inducing urea production, (2) in enteral use, Orn supplementation appears preferable to Arg as it is less ureogenic (as also recently demonstrated in vivo in stressed rats receiving isomolar amounts of Arg and Orn), (3) the liver participates in the Orn-alpha KG metabolic interaction, mostly in proline metabolism, which occurs in the splanchnic area.

Arterio-venous differences in amino acids, glucose, lactate and fatty acids in burn patients; effect of ornithine alpha-ketoglutarate

The study concerns two groups of seven burn patients matched for age, weight and total burn surface. Both groups received conventional enteral nutrition, while one was given a 10 g/day alpha-ketoglutarate ornithine (OKG) supplement. Femoral venous and arterial blood was taken from day 2 to day 13 post-burn in order to determine levels of amino acids, nonesterified fatty acids (NEFA), glucose and lactate. In the control group large negative arterio-venous differences (DeltaA-V) were observed in amino acid and lactate levels whereas they were significantly lower with regard to Hyp, Gly, Lys and Ala in the OKG-treated group. DeltaA-V was near zero for glucose and NEFA in both groups. These results support the view that OKG-therapy limits the output of amino acids in the leg and that glucose and NEFA do not constitute the main fuel in muscle.

Taurine status and response to intravenous taurine supplementation in adults with short-bowel syndrome undergoing long-term parenteral nutrition: a pilot study

Taurine deficiency in patients on long-term parenteral nutrition may be involved in cholestasis. We aimed to assess plasma taurine and tauro-conjugated bile acids in adults with short-bowel syndrome and their response to intravenous taurine. Thirty-two adult patients, who had been on taurine-free parenteral nutrition for a mean of 59 (SE 14) months for short-bowel syndrome, were studied retrospectively. In a second study, a subgroup of ten patients with chronic cholestasis received taurine-enriched (6.0 (SE 0.6) mg/kg per d) parenteral nutrition for 55 (SE 13) months. Post-absorptive plasma taurine and bile acid concentrations were measured and liver function tests routinely sampled. At baseline, plasma taurine was lower in patients with a jejunal length of less than 35 cm (group A, n 16) than in those with a jejunal length of 35 cm or more (group B, n 16): 43 (SE 3) v. 58 (SE 4) micromol/l (P=0.01). The groups were no different in terms of chronic cholestasis (12/16 v.13/16 patients), total bile acids (26 (SE 13) v.14 (SE 5) micromol/l) or the ratio of tauro-conjugated:glyco-conjugated bile acids (5 (SE 2) v.8(SE 4)%, usual range 30-60%). After supplementation, there was an increase in plasma taurine level (63 (SE 8) v. 43 (SE 4), P=0.007) but was no change in either total bile acids or the ratio of tauro-conjugated: glyco-conjugated bile acids. There was a significant decrease in aspartate aminotransferase level. Long-term parenteral nutrition for short-bowel syndrome is associated with an impaired tauro-conjugation of bile acids (enterohepatic pool), irrespective of plasma taurine level (systemic pool) and despite long-term taurine intravenous supplementation.

Influence of enterally administered ornithine α-ketogluta rate on hormonal patterns in burn patients

Abstract Plasma concentrations of glucose, insulin, C-peptide, glucagon, cortisol and hGH were measured in burn patients (mean burn surface area 21 percent) treated or not with ornithine α-ketoglutarate (OKG). An increase in basal values of glucose, insulin, C-peptide and cortisol was demonstrated in both groups, whereas hGH values diminished. OKG modified neither insulin nor hGH values 24 h after its enteral administration nor insulin levels within the first 4h after intake. On the other hand, 60 min after enteral nutrition was restarted the hyperglycaemia observed in untreated subjects was reduced by OKG whereas a hyperinsulinism was observed in both groups. These results suggest that: (i) the anticataboliclanabolic action of OKG in burn patients is not mediated by insulin or hGH, (ii) OKG probably induces an increase in glucose tolerance in burn patients, in whom there is a state of insulin resistance. The mechanism of this action requires further study.

Amino acid metabolism in isolated perfused rat liver

Conflicting evidence concerning hepatic amino acid (AA) metabolism in the isolated perfused rat liver (IPRL) led us to investigate the response of IPRL using perfusates with various AA contents. Perfusion (n = 4) with whole rat blood diluted in Krebs buffer (1:3, v/v) led to acute proteolysis on account of AA deprivation, as shown by the large release of AA (approximately 1400 mumoles in 120 min), especially branched-chain AA (BCAA) (e.g., Leu, 35.4 +/- 10.4 nmole.min-1.g-1 the first hour, 34.3 +/- 5.5 nmole.min-1.g-1 the second hour). In a first attempt to prevent proteolysis, livers (n = 4) were perfused with the previous medium supplemented with AA known for their antiproteolytic activity, at twice their physiological concentrations. Results during the first hour showed uptake of several AA (mainly alanine, glutamine, and proline), reduced release of BCAA (leucine, 12.5 +/- 6.3 nmole.min-1.g-1), and an increase in glucose and urea production. However, during the second hour, because of the use of a recirculating system, progressive AA depletion induced a reappearance of proteolysis. A two-step AA loading technique, i.e., the addition of antiproteolytic AA at the beginning of the perfusion and the addition of a balanced AA mixture at 60 min caused a further decrease in proteolysis during the 2 hr of perfusion (n = 6). Under these conditions, most AA were taken up by the liver with uptake values comparable to those observed in vivo.

Origins of hyperphenylalaninemia in burn patients.

Plasma phenylalanine concentrations were measured on the seventh postburn day in patients with burn surface areas ranging from 1 to 39%. Hepatic function (glutamate-pyruvate transaminase, prothrombin time) and protein breakdown (nitrogen balance, urine 3-methylhistidine/creatinine ratio) were evaluated in parallel. Plasma phenylalanine concentrations in patients with normal hepatic function and those with hepatic failure were similar. A correlation was shown between phenylalaninemia and nitrogen balance for all patients but phenylalaninemia only correlated with the urine 3-methyl-histidine/creatinine ratio in patients with burn surface areas under 20%. Taken as a whole, the data suggest that the determination of plasma phenylalanine concentration is a valuable test to assess the nitrogen balance in burn patients. This test, when performed on day 7, is particularly relevant in patients with normal hepatic function and either slight or moderate burn injury.