Silencing alanine transaminase 2 in diabetic liver attenuates hyperglycemia by reducing gluconeogenesis from amino acids

Hepatic gluconeogenesis from amino acids contributes significantly to diabetic hyperglycemia, but the molecular mechanisms involved are incompletely understood. Alanine transaminases (ALT1 and ALT2) catalyze the interconversion of alanine and pyruvate, which is required for gluconeogenesis from alanine. We found that ALT2 was overexpressed in liver of diet-induced obese and db/db mice and that the expression of the gene encoding ALT2 (GPT2) was downregulated following bariatric surgery in people with obesity. The increased hepatic expression of Gpt2 in db/db liver was mediated by activating transcription factor 4; an endoplasmic reticulum stress-activated transcription factor. Hepatocyte-specific knockout of Gpt2 attenuated incorporation of 13C-alanine into newly synthesized glucose by hepatocytes. In vivo Gpt2 knockdown or knockout in liver had no effect on glucose concentrations in lean mice, but Gpt2 suppression alleviated hyperglycemia in db/db mice. These data suggest that ALT2 plays a significant role in hepatic gluconeogenesis from amino acids in diabetes.

Branched-chain amino acid supplementation during bed rest: effect on recovery

Bed rest is associated with a loss of protein from the weight-bearing muscle. The objectives of this study are to determine whether increasing dietary branched-chain amino acids (BCAAs) during bed rest improves the anabolic response after bed rest. The study consisted of a 1-day ambulatory period, 14 days of bed rest, and a 4-day recovery period. During bed rest, dietary intake was supplemented with either 30 mmol/day each of glycine, serine, and alanine ( group 1) or with 30 mmol/day each of the three BCAAs ( group 2). Whole body protein synthesis was determined with U-15N-labeled amino acids, muscle, and selected plasma protein synthesis withl-[2H5]phenylalanine. Total glucose production and gluconeogenesis from alanine were determined with l-[U-13C3]alanine and [6,6-2H2]glucose. During bed rest, nitrogen (N) retention was greater with BCAA feeding (56 ± 6 vs. 26 ± 12 mg N · kg−1 · day−1, P < 0.05). There was no effect of BCAA supplementation on either whole body, muscle, or plasma protein synthesis or the rate of 3-MeH excretion. Muscle tissue free amino acid concentrations were increased during bed rest with BCAA (0.214 ± 0.066 vs. 0.088 ± 0.12 nmol/mg protein, P < 0.05). Total glucose production and gluconeogenesis from alanine were unchanged with bed rest but were significantly reduced ( P < 0.05) with the BCAA group in the recovery phase. In conclusion, the improved N retention during bed rest is due, at least in part, to accretion of amino acids in the tissue free amino acid pools. The amount accreted is not enough to impact protein kinetics in the recovery phase but does improve N retention by providing additional essential amino acids in the early recovery phase.

Effects of ATP infusion on glucose turnover and gluconeogenesis in patients with advanced non-small-cell lung cancer

Cancer cachexia is associated with elevated lipolysis, proteolysis and gluconeogenesis. ATP infusion has been found to significantly inhibit loss of body weight, fat mass and fat-free mass in patients with advanced lung cancer. The present study was aimed at exploring the effects of ATP on whole-body glucose turnover, alanine turnover and gluconeogenesis from alanine. Twelve patients with advanced non-small-cell lung cancer (NSCLC) were studied 1 week before and during 22–24 h of continuous ATP infusion. After an overnight fast, turnover rates of glucose and alanine, and gluconeogenesis from alanine, were determined using primed constant infusions of [6,6-2H2]glucose and [3-13C]alanine. Thirteen NSCLC patients and eleven healthy subjects were studied as control groups without ATP infusion. During high-dose ATP infusion (75 µg·min-1·kg-1), glucose turnover was 0.62±0.07 mmol·h-1·kg-1, compared with 0.44±0.13 mmol·h-1·kg-1 at baseline (P = 0.04). For gluconeogenesis a similar, but non-significant, trend was observed [baseline, 0.30±0.16 mmol·h-1·kg-1; during ATP, 0.37±0.13 mmol·h-1·kg-1 (P = 0.08)]. At lower ATP doses (37–50 µg·min-1·kg-1) these effects were not detected. The relative increase in glucose turnover during ATP infusion compared with baseline showed a significant correlation with the ATP dose (r = 0.58, P = 0.02). No change in alanine turnover was observed at any ATP dose. The results of this study indicate an increase in glucose turnover during high-dose ATP infusion compared with baseline levels. During high-dose ATP infusion, glucose turnover was similar to that during low-dose ATP infusion and to that in control NSCLC patients. Between ATP infusions, however, glucose turnover in patients treated with high-dose ATP was significantly lower than that in the low-dose and control NSCLC patients (P = 0.04 and P = 0.03 respectively), and similar to that in healthy subjects. This would suggest that repeated high-dose ATP infusions may inhibit glucose turnover between infusion periods.

Hepatic sugar phosphate levels reflect gluconeogenesis in lung cancer: simultaneous turnover measurements and 31P magnetic resonance spectroscopy in vivo

Stable-isotope tracers were used to assess whether levels of phosphomonoesters (PME) and phosphodiesters (PDE) in the livers of lung cancer patients, as observed by 31P magnetic resonance (MR) spectroscopy, reflect elevated whole-body glucose turnover and gluconeogenesis from alanine. Patients with advanced non-small-cell lung cancer without liver metastases (n = 24; weight loss 0–24%) and healthy control subjects (n = 13) were studied after an overnight fast. 31P MR spectra of the liver in vivo were obtained, and glucose turnover and gluconeogenesis from alanine were determined simultaneously using primed-constant infusions of [6,6-2H2]glucose and [3-13C]alanine. Liver PME concentrations were 6% higher in lung cancer patients compared with controls (not significant); PME levels in patients with ⩾ 5% weight loss were significantly higher than in patients with < 5% weight loss (P < 0.01). PDE levels did not differ between the groups. In lung cancer patients, whole-body glucose production was 19% higher (not significant) and gluconeogenesis from alanine was 42% higher (P < 0.05) compared with healthy subjects; turnover rates in lung cancer patients with ⩾ 5% weight loss were significantly elevated compared with both patients with < 5% weight loss and healthy subjects (P < 0.05). PME levels were significantly correlated with glucose turnover and gluconeogenesis from alanine in lung cancer patients (r = 0.48 and r = 0.48 respectively; P < 0.05). In conclusion, elevated PME levels in lung cancer patients appear to reflect increased glucose flux and gluconeogenesis from alanine. These results are consistent with the hypothesis that elevated PME levels are due to contributions from gluconeogenic intermediates.

Mechanisms of increased gluconeogenesis from alanine in rat isolated hepatocytes after endurance training

This work aimed at further investigating the mechanisms by which liver gluconeogenic capacity from alanine is improved after training in rats, with an isolated hepatocyte model. Compared with controls in hepatocytes from trained rats incubated with gluconeogenic precursors (20 mM), the glucogenic flux ( J glucose) was increased by 64% from alanine (vs. 21% for glycerol, 18% for lactate-pyruvate 10:1, and 10% for dihydroxyacetone). Maximal intracellular alanine accumulation capacity was also increased by 50%. Further experiments conducted on perifused hepatocytes showed that the putative adaptation at the level of the phospho enolpyruvate-pyruvate cycle, which could be involved in the increased J glucose from lactate-pyruvate, was not involved in the increased J glucose from alanine after training. For alanine concentration higher than ∼1 mM, an increased flux through alanine aminotransferase appeared responsible for the increased J glucose. This could, in turn, depend on an increased supply of cytosolic 2-oxoglutarate because of the higher mitochondrial respiration observed in hepatocytes from trained rats and the activation of the malate-aspartate shuttle. At lower alanine concentration, the increase in J glucose appeared to be entirely due to the improved transport capacity.