Quantitation of gluconeogenesis in humans

Quantitation of Gluconeogenesis in Humans by Karin Ekberg, Department of Clinical Physiology, Karolinska Hospital and Institute,S-171 76 Stockholm, Sweden Several methods have been introduced for estimating gluconeogenesis, but noneprovides a direct measurement without assumptions that condition the accuracy ofthe estimate. The aim of this work has been to develop a new method for quantitativestudies of gluconeogenesis, applicable in humans. We postulated that by using (U-13C3)glyceroland application of mass isotopomer distribution analysis (MIDA) to the enrichmentsof 13C incorporated into blood glucose the contribution of gluconeogenesis to wholebody glucose production could be determined. However, when applied in healthy subjectsfasted for 60 h, the estimated contribution was only 50-60% when close to 100% ofthe glucose production was expected to derive from gluconeogenesis. The values obtainedwere incompatible with glucose being formed from a single pool of triose phosphatein the liver, which was a condition for the validity of the method. Thus, we concludedthat the MIDA technique based on administration of 13C-labeled glycerol does notprovide a valid quantitation of gluconeogenesis in vivo. Gluconeogenesis can be estimated a few hours after ingestion of a dose of deuteratedwater that can be safely administered to humans (body water is enriched to 0.5%).The method is based on the incorporation of deuterium into the hydrogen bound tocarbon 5 of glucose (C-5) formed by gluconeogenesis and to carbon 2 (C-2) of allnewly formed glucose molecules (i.e formed after the ingestion of deuterated water).The contribution of gluconeogenesis to whole body glucose production thus equalsthe ratio of enrichments at C-5 to C-2. Simultaneously, glucose production can beestimated by isotope dilution technique using (6,6-2H2)glucose infusion. Together,the methods provide a quantitative measurement of gluconeogenesis. The applicabilityof the techniques rests on the conversion of the tracer hydrogens and their carboninto formaldehyde and its further polymerization to hexamethylenetetramine (HMT).Thereby, an accurate assessment of the enrichments is facilitated although body waterhas only been enriched to 0.5%. Moreover, the glucose rate of appearance can be determinedwithout influence from the ingested deuterated water. Applied in healthy subjects in the overnight fasted state, approximately equalproportions of glucose production derived from gluconeogenesis and glycogenolysis.During continued fasting the contribution from gluconeogenesis increased gradually(54% at 14 h and 62% at 22 h), but in absolute terms it stayed constant between 14h and 22 h of fasting (5.5 µmol/kg/min). Thus, the fall in glucose productionobserved between 14 and 22 h of fasting, from 10.2 to 8.6 lµmol/kg/min, couldbe attributed to a decrease in the rate of glycogenolysis After 42 h of fasting almostall glucose produced (94%) derived from gluconeogenesls. In the overnight fasted state, gluconeogenesis, as reported above, may be equatedwith hepatic gluconeogenesis. This was evident from the application of catheterizationtechnique in healthy subjects together with labeled glucose infusion for estimationacross both splanchnic and renal tissues: in the 12 h fasted state, hepatic glucoseproduction (the sum of net splanchnic output and glucose uptake to the splanchnicbed) accounted for 100% of whole body glucose production as measured by the isotopedilution technique. No net renal production of glucose and no or only a minimal fractionalextraction of labeled glucose were found. In the 60 h fasted state, hepatic glucoseproduction accounted for 80% of whole body glucose production. No renal extractionof glucose but a significant net renal glucose production were found. Thus the renalcontribution to glucose production may represent 10-25% after 60 h of fasting. Key words: Gluconeogenesis, mass isotopomer distribution analysis, deuteratedwater, hexamethylenetetramine, splanchnic and renal glucose production ISBN 91 -628-2810-X Stockholm 199