Biomedical subjects

Craig R Malloy

Publications and source records attributed to Craig R Malloy.

24 records · Page 2Linked to original sources

PubMed2003

Sources of plasma glucose by automated Bayesian analysis of 2H NMR spectra.

Sources of blood glucose can be determined after oral ingestion of (2)H(2)O followed by isolation of plasma glucose and measurement of the relative (2)H enrichments in select positions within the glucose molecule. Typically, (2)H enrichments are obtained by mass spectrometry but (2)H NMR offers an alternative. Here it is demonstrated that the entire analysis may be automated by Bayesian analysis of a (2)H free induction decay signal of monoacetone glucose to obtain a direct readout of the relative contributions of glycogenolysis, glycerol, and phosphoenol pyruvate to plasma glucose production. Furthermore, Markov Chain Monte Carlo (MCMC) simulations of the posterior probability density provide uncertainties in all metabolic parameters from a single patient, thereby allowing comparisons in glucose metabolism from one individual to another. The combined MCMC Bayesian methodology is operationally simple and requires little intervention from the operator.

Autoanalysis↗

PubMed2003

Measuring in-vivo metabolism using nuclear magnetic resonance.

PURPOSE OF REVIEW: This review introduces physiologists and clinical investigators to an ever-widening array of nuclear magnetic resonance applications. In particular, it highlights a multiple tracer technique that provides a comprehensive picture of metabolic processes within human liver. RECENT FINDINGS: Magnetic resonance spectroscopy is an important technique for studying metabolism in the brain, liver, heart and skeletal muscle. One fundamental advantage is that the studies are inherently noninvasive, so time-dependent information can be obtained. For example, 31P nuclear magnetic resonance investigations indicate that greater maximal oxygen uptake and oxidative capacity in trained athletes can be partially attributed to adaptations enhancing the rates at which phosphocreatine and inorganic phosphate recover during stress. In-vivo measurements of lipids and glycogen by 1H and 13C spectroscopy demonstrate that accumulation of intracellular lipids and impaired rates of glycogen synthesis contribute to insulin resistance and type 2 diabetes mellitus. Similar techniques can be used to analyze blood and urine samples obtained during administration of 2H or 13C tracers to yield information that cannot be easily obtained by mass spectrometry. Additional information available from nuclear magnetic resonance yields a comprehensive picture of liver metabolic pathways from a single clinical study. SUMMARY: A variety of magnetic resonance spectroscopy protocols have been validated and exploited for clinical studies, but relatively few investigators are comfortable with technical aspects of these protocols and utilize them for clinical research. Increased interaction between spectroscopists and other investigators is needed if the potential of nuclear magnetic resonance for studying in-vivo metabolism is to be fully realized.

Carbon Isotopes↗

PubMed2003

Mechanisms by which liver-specific PEPCK knockout mice preserve euglycemia during starvation.

Liver-specific PEPCK knockout mice, which are viable despite markedly abnormal lipid metabolism, exhibit mild hyperglycemia in response to fasting. We used isotopic tracer methods, biochemical measurements, and nuclear magnetic resonance spectroscopy to show that in mice lacking hepatic PEPCK, 1) whole-body glucose turnover is only slightly decreased; 2) whole-body gluconeogenesis from phosphoenolpyruvate, but not from glycerol, is moderately decreased; 3) tricarboxylic acid cycle activity is globally increased, even though pyruvate cycling and anaplerosis are decreased; 4) the liver is unable to synthesize glucose from lactate/pyruvate and produces only a minimal amount of glucose; and 5) glycogen synthesis in both the liver and muscle is impaired. Thus, although mice without hepatic PEPCK have markedly impaired hepatic gluconeogenesis, they are able to maintain a near-normal blood glucose concentration while fasting by increasing extrahepatic gluconeogenesis coupled with diminishing whole-body glucose utilization.

Animals↗

PubMed2002

13C isotopomer analysis of glutamate by tandem mass spectrometry.

Tandem mass spectrometry allows a compound to be isolated from the rest of the sample and dissociated into smaller fragments. We show here that fragmentation of glutamate mass isotopomers yields additional mass spectral data that significantly improve the analysis of metabolic fluxes compared to full-scan mass spectrometry. In order to validate the technique, tandem and full-scan mass spectrometry were used along with (13)C NMR to analyze glutamate from rat hearts perfused with three substrate mixtures (5 mM glucose plus 5 mM [2-(13)C]acetate, 5 mM [1-(13)C]glucose plus 5 U/L insulin, and 5 mM glucose plus 1 mM [3-(13)C]pyruvate). Analysis by tandem mass spectrometry showed that the enriched substrate contributed 98 +/- 2, 53 +/- 2, and 84 +/- 7%, respectively, of acetyl-coenzyme A while the rate of anaplerotic substrate entry was 7 +/- 3, 25 +/- 8, and 16 +/- 8%. Similar results were obtained with (13)C NMR data, while values from full-scan data had higher error. We believe that this is the first use of tandem mass spectrometry to determine pathway flux using (13)C-enriched substrates. Although analysis of the citric acid cycle by NMR is simpler (and more intuitive), tandem mass spectrometry has the potential to combine high sensitivity with the high information yield previously available only by NMR.

Animals↗

PubMed2002

Hepatic gluconeogenesis and Krebs cycle fluxes in a CCl4 model of acute liver failure.

Acute liver failure was induced in rats by CCl4 administration and its effects on the hepatic Krebs cycle and gluconeogenic fluxes were evaluated in situ by 13C NMR isotopomer analysis of hepatic glucose following infusion of [U-13C]propionate. In fed animals, CCl4 injury caused a significant increase in relative gluconeogenic flux from 0.80+/-0.10 to 1.34 +/-0.24 times the flux through citrate synthase (p<0.01). In 24-h fasted animals, CCl4-injury also significantly increased relative gluconeogenic flux from 1.36+/-0.16 to 1.80+/-0.22 times the flux through citrate synthase (p<0.01). Recycling of PEP via pyruvate and oxaloacetate was extensive under all conditions and was not significantly altered by CCl4 injury. CCl4 injury significantly reduced hepatic glucose output by 26% (42.8+/-7.3 vs 58.1+/-2.4 micromol/kg/min, p=0.005), which was attributed to a 26% decrease in absolute gluconeogenic flux from PEP (85.6+/-14.6 vs 116+/-4.8 micromol/kg/min, p<0.01). These changes were accompanied by a 47% reduction in absolute citrate synthase flux (90.6+/-8.0 to 47.6+/-8.0 micromol/kg/min, p<0.005), indicating that oxidative Krebs cycle flux was more susceptible to CCl4 injury. The reduction in absolute fluxes indicate a significant loss of hepatic metabolic capacity, while the significant increases in relative gluconeogenic fluxes suggest a reorganization of metabolic activity towards preserving hepatic glucose output.

Alanine Transaminase↗

PubMed2002

Effects of hypothermia on myocardial substrate selection.

BACKGROUND: Hypothermia lowers the metabolic rate and increases ischemic tolerance but the effects of temperature on myocardial substrate selection are not well defined. METHODS: Isolated rat hearts were perfused with physiologic concentrations of 13C labeled lactate, pyruvate, acetoacetate, mixed long-chain fatty acids, and glucose. Hearts were cooled over 5 to 10 minutes to one of four target temperatures (37 degrees, 32 degrees, 27 degrees, or 17 degrees C), then perfused for an additional 30 minutes, freeze-clamped, and extracted. 13C NMR spectra were obtained and substrate oxidation patterns were determined by isotopomer analysis. RESULTS: Although hearts in all groups were supplied with identical substrates, the percentage of acetyl-CoA oxidized within the citric acid cycle that arose from fatty acids decreased significantly from 53.8% +/- 0.8% in the 37 degrees C group to 33.1% +/- 3.3% in the 17 degrees C group. Lactate or pyruvate utilization increased from 3.3% +/- 0.5% to 25.7% +/- 3.6%, respectively (p < 0.05 by one-way ANOVA). CONCLUSIONS: These data suggest that moderate hypothermia suppresses fatty acid oxidation and deep hypothermia significantly increases utilization of lactate and pyruvate. These effects may result from relative inhibition of catabolism of complex molecules such as fatty acids, or stimulation of pyruvate dehydrogenase. These effects on substrate metabolism may play a role in myocardial protection afforded by hypothermia.

Animals↗