Endotoxin induced hyperlactatemia and hypoglycemia is linked to decreased mitochondrial phosphoenolpyruvate carboxykinase.
OBJECTIVE
Phosphoenolpyruvate carboxykinase (PEPCK) is the rate limiting enzyme for gluconeogenesis, and plays a key role in recycling lactate for glucose production. It is synthesized as two separate isoforms; cytosolic (PEPCK-C, gene code; PCK1) and mitochondrial (PEPCK-M, gene code; PCK2). Previous studies of gluconeogenesis in endotoxemia have focused solely on PCK1. We investigated the relative roles of the two isoforms in hepatic and renal gluconeogenesis in a rat model of endotoxic shock, and in cultured hepatocytes.


METHODS
Rats were administered lipopolysaccharide (6 mg/kg; LPS) for 6 h. Cultured cells were incubated with lactate (5 mM) with or without tumor necrosis factor alpha (1 - 10 ng/ml). Rat liver and kidney samples as well as cultured cells were subjected to subcellular fractionation to produce mitochondrial and cytosolic fractions for PEPCK activity assay. PCK1 and PCK2 mRNA levels were measured using quantitative RT-PCR.


RESULTS
In rat endotoxemia, hepatic PCK2 mRNA and PEPCK-M enzyme activity decreased by 53% and 38%, compared to sham controls. Hepatic PCK1 mRNA levels increased by 44%, but PEPCK-C enzyme activity remained unchanged. The changes in hepatic PEPCK-M coincided with a marked hypoglycemia and hyperlactatemia as well as elevated plasma interleukin 1 beta (IL1beta). Incubation of cultured hepatocytes with TNF-alpha inhibited lactate-induced increases in glucose production, PCK2 mRNA levels and PEPCK-M enzyme activity but had no effect on PCK1 mRNA levels or PEPCK-C activity.


CONCLUSIONS
These results indicate that decreases in hepatic PEPCK-M play a key role in the manifestation of hyperlactatemia and hypoglycemia in endotoxemia.
Caton
Paul W
PW
William Harvey Research Institute, Bart's and the London, Queen Mary's School of Medicine and Dentistry, London EC1M 6BQ, United Kingdom. p.w.caton@qmul.ac.uk
Nayuni
Nanda K
NK
Murch
Oliver
O
Corder
Roger
R
D000818:Animals; D001794:Blood Pressure / Q000187:drug effects; D004731:Endotoxins / Q000494:pharmacology*; D005947:Glucose / Q000378:metabolism; D006339:Heart Rate / Q000187:drug effects; D007003:Hypoglycemia / Q000097:blood* / Q000139:chemically induced; D053583:Interleukin-1beta / Q000378:metabolism; D007668:Kidney / Q000378:metabolism / Q000473:pathology; D007773:Lactates / Q000097:blood*; D008099:Liver / Q000378:metabolism / Q000473:pathology; D008297:Male; D008928:Mitochondria / Q000187:drug effects / Q000201:enzymology*; D010729:Phosphoenolpyruvate Carboxykinase (GTP) / Q000097:blood*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013347:Subcellular Fractions / Q000187:drug effects / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
D004731:Endotoxins; D053583:Interleukin-1beta; D007773:Lactates; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D010729:Phosphoenolpyruvate Carboxykinase (GTP); D005947:Glucose