Patti A. Quant

Hepatic Mitochondrial 3-Hydroxy-3-Methylglutaryl-Coenzyme A Synthase Deficiency

There are at least two isoenzymes of 3-hydroxy-3-methylglutaryl (HMG)-CoA synthase (EC 4.1.3.5) located in the mitochondrial matrix and the cytoplasm of hepatocytes, respectively. The mitochondrial enzyme is necessary for the synthesis of ketone bodies, which are important fuels during fasting. We report a child with a deficiency of this isoenzyme. He presented at 16 mo with hypoglycemia. There was no rise in ketone bodies during fasting or after a long chain fat load but there was a small rise after a leucine load. Measurement of β-oxidation flux in fibroblasts was normal. Using antibodies specific for mitochondrial HMG-CoA synthase, no immunoreactive material could be detected on Western blotting. Total HMG-CoA synthase activity in liver homogenate was only slightly lower than in control samples. Presumably, as there was no mitochondrial HMG-CoA synthase enzyme protein, this activity arose from the cytoplasmic or other (e.g. peroxisomal) isoenzymes. With avoidance of fasting, our patient has had no problems since presentation and is developing normally at 4 y of age.

Neonatal oxidative liver metabolism: effects of hydrogen peroxide, a putative mediator of septic damage.

BACKGROUND/PURPOSEnSurgical neonates are at risk for sepsis and liver dysfunction. These complications are more common in preterm neonates and in those who receive total parenteral nutrition. Elevated levels of reactive oxygen species (eg, hydrogen peroxide) have been reported in these at-risk patients and may be the mediators of liver impairment via their effect on oxidative energy metabolism. The aim of this study was to test the hypothesis that elevated levels of hydrogen peroxide (H2O2) impair neonatal liver oxidative energy metabolism.nnnMETHODSnAn in vitro model to test this hypothesis was developed in hepatocytes isolated from neonatal (11-day to 15-day) rats. The cells, respiring on palmitate (0.5 mmol/L in 2% bovine serum albumin), were exposed to H2O2. Oxygen consumption was measured polarographically. In experiment A, H2O2 was added to the cell preparation at different concentrations (0.5 mmol/L, 1 mmol/L, 1.5 mmol/L, 2 mmol/L) to assess the effect on oxygen consumption. In experiment B, H2O2 (2 mmol/L) was added to hepatocytes in the presence of inhibitors of mitochondrial respiration to define the site of action of H2O2. In experiment C, electron microscopy was performed on hepatocytes after incubation with 1 mmol/L and 2 mmol/L of H2O2.nnnRESULTSnIn experiment A, H2O2 significantly reduced hepatocyte oxygen consumption at 1.5 and 2 mmol/L. In experiment B, in the presence of inhibitors of mitochondrial respiration, myxothiazol (inhibitor of substrate oxidation), and oligomycin (inhibitor of adenosine triphosphate (ATP) synthase), no further inhibition by H2O2 occurred, indicating that the effect of H2O2 was intramitochondrial and affecting the synthesis of ATP. In experiment C, microscopic alterations of mitochondria were noticed exclusively in hepatocytes incubated with 2 mmol/L H2O2.nnnCONCLUSIONSnResults of this study demonstrate that H2O2 impairs neonatal liver oxidative metabolism. H2O2 probably directly inhibits ATP synthase. The authors hypothesize that H2O2 may play a role in the biochemical pathogenesis of liver dysfunction associated with sepsis. Identification of the precise target site of H2O2 may be valuable in directing therapy in septic neonates.

Analgesic doses of fentanyl impair oxidative metabolism of neonatal hepatocytes

BACKGROUND/PURPOSEnStudies in human surgical neonates have shown that intraoperative fentanyl analgesia results in greater fall in perioperative body core temperature compared with morphine analgesia. The aim of the study was to compare in a neonatal animal model the biochemical effect of fentanyl and morphine on hepatocyte oxidative metabolism.nnnMETHODSnHepatocytes were isolated from suckling rats and the oxygen consumption from palmitate was measured polarographically. In experiment A, fentanyl and morphine within the respective analgesic serum ranges were added to hepatocytes to assess the effect on oxygen consumption. In experiment B, fentanyl was added to hepatocytes in the presence of inhibitors of mitochondrial respiration to investigate its site of action. In experiment C, hepatocytes were incubated with either fentanyl or morphine, centrifuged, and then examined ultrastructurally by electron microscopy.nnnRESULTSnIn experiment A, fentanyl inhibited oxygen consumption by up to 40% (P < .01). Morphine inhibited oxygen consumption to a maximum of 25% (P < .01). In experiment B, in the presence of oligomycin, fentanyl increased the inhibition of oxygen consumption; however, in the presence of myxothiazol, no further inhibition by fentanyl occurred. In experiment C, mild ultrastructural alterations to hepatocytes were observed after incubation with fentanyl but not with morphine.nnnCONCLUSIONSnThis study demonstrates that therapeutic doses of two commonly used analgesic drugs impair neonatal hepatic oxidative metabolism. Fentanyl exerts a greater effect than morphine by diminishing liver oxygen consumption by up to 40%. The inhibitory effect of fentanyl occurs directly on the mitochondrial respiratory chain, either on substrate oxidation or on the thermogenic proton leak. The findings of this study are relevant to the perioperative management of surgical neonates.

Current Views in Fatty Acid Oxidation and Ketogenesis

Preface & Dedication P.A. Quant, S. Eaton. Historical overview. Cambridge, colleagues, carnitine and ketogenesis ... P.K. Tubbs. Carnitine and carnitine acyltransferases. 1. Biogenesis of the rat liver mitochondrial carnitine palmitoyltransferase I I. Cohen, et al. 2. Subcellular distribution of mitochondrial carnitine palmitoyltransferase I in rat liver: Evidence for a distinctive N-terminal structure of the microsomal but not the peroxisomal enzyme V.A. Zammit,et al. 3. Topology of hepatic mitochondrial carnitine palmitoyltransferase I K. Kashfi, G.A. Cook. 4. Possible involvement of cytoskeletal components in the control of hepatic carnitine palmitoyltransferase I activity G. Velasco, et al. 5. Regulation of beta-oxidation and CPT I activity by 3-thia fatty acids in cultured rat hepatocytes J. Sleboda, J. Bremer. 6. Carnitine acyltransferases and associated transport systems in the endoplasmic reticulum N.M. Broadway, et al. 7. Reciprocal enzymatic interference of carnitine palmitoyl transferase I and glycerol-3-phosphate acyltransferase in purified liver mitochondria F. Beauseigneur, et al. 8. Control of human muscle carnitine palmitoyltransferase I gene transcription by peroxisome proliferator-activated receptor C. Mascaro, et al. 9. Kinetic investigation of carnitine palmitoyltransferases in homogenates of human skeletal muscle using L-aminocarnitine and malonyl-CoA K. Hertel, et al. 10. Processing of carnitine octanoyltransferase pre-mRNAs by cis- and trans-splicing C. Caudevilla, et al. 11. Selective modulation of carnitine long-chain acyltransferase activities: kinetics, inhibitors andactive sites of COT and CPT II R.R. Ramsay, R.D. Gandour. 12. Confocal laser scanning microscopy of human skin fibroblasts showing transient expression of a green fluorescent carnitine palmitoyltransferase 1 fusion protein F.R. van der Leij, et al. 13. Carnitine biosynthesis: purification of gamma-butyrobetaine from rat liver F. Vaz, et al. Mitochondrial fatty acid oxidation and ketogenesis. 14. Hypolipidemic 3-thia fatty acids: Fatty acid oxidation and ketogenesis in rat liver under proliferation of mitochondria and peroxisomes R.K. Berge, et al. 15. Molecular mechanisms of fatty acid b-oxidation enzyme catalysis S.-Y. Yang, X.-Y. He. 16. Control of mitochondrial b-oxidation at the level of [NAD+]/[NADH] and CoA acylation S. Eaton, et al. 17. Production and export of acyl-carnitine esters by neonatal rat hepatocytes S. Eaton, et al. 18. Tissue specific differences in intramitochondrial control of b-oxidation S. Eaton, K. Bartlett. 19. Endotoxin-induced changes in very-low-density lipoprotein and myocardial utilisation of triacylglycerol from abnormal VLDL in the rat D.G. Hole, et al. 20. Effect of valproic acid on the expression of acyl-CoA dehydrogenases in various tissues M. Nagao, et al. 21. Formation of a human electron transferring flavoprotein: medium chain acyl CoA dehydrogenase complex - preliminary evidence from crosslinking studies A. Parker, P.C. Engel. 22. Cloning and regulation of peroxisome proliferator-induced acyl-CoA thioestereases from mouse liver M. Hunt, et al. 23. Metabolic effects of 3-thia fatty acids in cancer cells K.J. Tronstad,

A new mouse mutant, skijumper

Low blood sugar levels are a well-known cause of severe illness and often death in newborn humans, especially those that are small for age. Few of the causes of neonatal hypoglycemia are known, and many remain to be found. We describe a novel mouse mutant, skijumper (skimp), in which pups, despite feeding well, have low levels of glucose and develop opisthotonos, followed by death typically within a few days after birth. Genetic mapping studies have localized the lesion to a approximately 1 cM interval on mouse Chromosome (Chr) 7 between D7Mit318 and D7Mit93. We have carried out extensive analysis to define the phenotype and its likely cause. In addition to low blood glucose, affected skijumper mice have lowglycogen and ketone levels. Mass spectrometric analysis of blood samples has excluded major defects in amino acid metabolism. Initial biochemical analyses suggested a defect in ketogenesis as one possible cause of this phenotype. However, measurements of levels and activities of carnitine, carnitine palmitoyl transferases, and other enzymes involved in ketogenesis, along with studies of mitochondrial structure and function, did not demonstrate significant differences between skijumper, unaffected littermates, and control wild-type mice. These results indicate that abnormal enzyme activity in known pathways does not appear to be the primary biochemical lesion in skijumper. The skijumper may be a new valuable model for studying and understanding one type of neonatal morbidity and death.