Hyperpolarized Dihydroxyacetone Is a Sensitive Probe of Hepatic Gluconeogenic State

Abstract

Type II diabetes and pre-diabetes are widely prevalent among adults. Elevated serum glucose levels are commonly treated by targeting hepatic gluconeogenesis for downregulation. However, direct measurement of hepatic gluconeogenic capacity is accomplished only via tracer metabolism approaches that rely on multiple assumptions, and are clinically intractable due to expense and time needed for the studies. We previously introduced hyperpolarized (HP) [2-13C]dihydroxyacetone (DHA) as a sensitive detector of gluconeogenic potential, and showed that feeding and fasting produced robust changes in the ratio of detected hexoses (6C) to trioses (3C) in the perfused liver. To confirm that this ratio is robust in the setting of treatment and hormonal control, we used ex vivo perfused mouse livers from BLKS mice (glucagon treated and metformin treated), and db/db mice. We confirm that the ratio of signal intensities of 6C to 3C in 13C nuclear magnetic resonance spectra post HP DHA administration is sensitive to hepatic gluconeogenic state. This method is directly applicable in vivo and can be implemented with existing technologies without the need for substantial modifications.

Enrichment of hepatic glycogen and plasma glucose from H₂18O informs gluconeogenic and indirect pathway fluxes in naturally feeding mice

“…The supernatant was then transferred to 0.5‐ml Reacti‐therm V‐vials and thoroughly dried. For derivatization of glucose to the aldonitrile pentapropionate, the dried extract was reconstituted in 50 μl of hydroxylamine hydrochloride in pyridine (20 mg/ml) and agitated with a micro stir bar at 90°C for 1.5 h. One hundred microliters of propionic anhydride was then added to each vial and agitated for 30 min at 70°C 20 . After derivatization, the samples were dried under N 2 and reconstituted in 100 μl of ethyl acetate.…”

“…The supernatant was then transferred to 0.5-ml Reacti-therm V-vials and thoroughly dried. For derivatization of glucose to the aldonitrile pentapropionate, the dried extract was reconstituted in 50 μl of hydroxylamine hydrochloride in pyridine (20 mg/ml) and agitated with a micro stir bar at 90 C for 1.5 h. One hundred microliters of propionic anhydride was then added to each vial and agitated for 30 min at 70 C. 20 ual isotopologues were identified and quantified by area using their known m/z ions. The isotopic 18 O ratio for a given fragment ion was calculated as the ratio of the intensity of each mass isotopologue to the sum of all mass isotopologues.…”

Detecting de novo Hepatic Ketogenesis Using Hyperpolarized [2-13C] Pyruvate

“…Experiments involving C57/BL6N mice were handled in compliance with University of Florida Institutional Animal Care and Use Committee approved protocol (#201909320). Liver perfusions were carried out as described in the literature (Moreno et al, 2014;Ragavan et al, 2021). Briefly, male mice (10-13 weeks old) in the fed state were anesthetized using isoflurane followed by an intraperitoneal injection of heparin.…”

“…DNP can produce "hyperpolarized" (HP) signal enhancements for 13 C of as much as 40,000 times depending upon the field strength of the detection magnet (Ardenkjaer-Larsen et al, 2003). Hyperpolarization has enabled studies of metabolism in cell culture (Harrison et al, 2012;Sriram et al, 2015), perfused organs including heart and liver (Moreno et al, 2010(Moreno et al, , 2014Jin et al, 2016;Ragavan et al, 2021), and in vivo (Cunningham et al, 2016;Chung et al, 2019).…”

Non-Invasive Analysis of Human Liver Metabolism by Magnetic Resonance Spectroscopy

“…Thus, the transition from fasting to feeding, where hepatic carbohydrate metabolic fluxes must undergo acute rearrangements in order to maintain whole body glucose homeostasis-and is, therefore, the most critical and testing phase for glucoregulation-is little understood. The ability to observe fast real-time alterations in hepatic sugar phosphates and other metabolites following administration of tracers, such as [2-13 C]dihydroxyacetone [95][96][97], [1-13 C]pyruvate [98][99][100], and [1-13 C]gluconolactone [101], promises to be invaluable for unveiling the redirection of hepatic carbohydrate fluxes during the fasted to fed transition. Moreover, the direct observation of hepatic metabolites overcomes another important limitation of gluconeogenic tracer enrichment of blood glucose: the inability to resolve gluconeogenic activity of the liver from that of other tissues such as the kidney and intestine.…”