Detecting liver injury non-invasively using hyperpolarized ¹³C MRI

In this issue of Liver International, Moon et al. (1) investigate the potential utility of hyperpolarized (HP) ¹³C magnetic resonance imaging (MRI) for detecting hepatic ischemia reperfusion injury (IRI), an important complication of liver transplantation. Significantly elevated hepatic conversion of intravenously injected HP [1-¹³C]pyruvate to both [1-¹³C]lactate and [1-¹³C]alanine was detected following experimental hepatic IRI in rats, as compared with sham-operated controls. Analogous metabolic changes have also previously been detected using HP ¹³C MRI in rodent models of IRI in both heart (2) and kidneys (3). The new results reported here in liver, taken together with prior work in the field, indicate strong potential for application of this approach to the diagnosis and monitoring of liver injury.

HP ¹³C MRI (Figure) is a new medical imaging modality that can uniquely track both uptake and metabolic conversion of stable isotope ¹³C-labeled small molecules. The method of dissolution dynamic nuclear polarization (DNP) (4) readily enhances the polarization of ¹³C nuclei by four to five orders of magnitude beyond the limits of clinical MRI magnets. In this approach, the ¹³C probe is first pre-polarized in a separate magnet at very low temperatures (~1 K) by microwave irradiation in the presence of an electron paramagnetic agent (EPA, typically a trityl radical), and subsequently rapidly dissolved into a liquid contrast agent. Unlike other medical imaging contrast agents, the spatial distributions of both the injected probe as well as its individual downstream metabolites can be discriminated (5) based on fine differences in their resonance frequencies (chemical shifts), which are dependent on molecular structure. However, in contrast with the tiny ¹H polarization of water that is the basis for conventional MRI, HP magnetization is short-lived and not readily regenerated. The short half-life (t_1/2) of the hyperpolarized state, whose specific value is also determined largely by molecular structure, requires rapid transit to the MRI scanner and rapid metabolic processing in vivo. By convention, T₁ exponential decay rate constants are reported (t_1/2=0.69*T₁), which are on the order of one minute or less for probes of interest.

Figure.

Illustration of the process of hyperpolarized (HP) ¹³C magnetic resonance imaging (MRI) via the method of dissolution dynamic nuclear polarization (DNP). After pre-polarization and rapid dissolution (left), the HP [1-¹³C]pyruvate solution is rapidly transported to the MRI scanner and injected into the subject. ¹³C MRI data is collected (right) showing not only the distribution of the substrate (pyr) but also its local conversion to [1-¹³C]lactate (lac) and [1-¹³C]alanine (ala) via key biochemical reactions.

[1-¹³C]pyruvate (5) is the most promising HP ¹³C probe by far, based on several favorable properties with respect to HP ¹³C technology, including high polarization, slow rate of demagnetization, and rapid metabolic processing to interesting biochemical fates in vivo. Among its biochemical pathways, rapid enzymatic reduction of HP [1-¹³C]pyruvate to [1-¹³C]lactate (catalyzed via lactate dehydrogenase or LDH), transamination to [1-¹³C]alanine (via alanine transaminase or ALT), and in some tissues decarboxylation to [¹³C]bicarbonate (via pyruvate dehydrogenase or PDH), are readily detected and imaged in vivo. Building on strong preclinical results from multiple groups, HP [1-¹³C]pyruvate MRI was translated into humans for the first time in a proof-of-concept study of patients with prostate cancer (6). Further human studies targeting applications in imaging of cancer and cardiovascular disease (7) are currently underway at several sites, with >20 new commercial dissolution DNP polarizers suitable for human use (i.e. equipped with sterile fluid paths) (8) recently installed or pending installation worldwide.

Liver injury is also emerging as another particularly promising target for HP ¹³C MRI, based on associated metabolic changes within the liver, and/or the effect of infiltration by glycolytic inflammatory cells. Seminal studies on HP [1-¹³C]pyruvate in perfused rat liver were conducted by Merritt et al. (9), who demonstrated capability of this new molecular imaging technology to investigate vital anaplerotic and decarboxylative fluxes of pyruvate, as well as its interconversions to lactate and alanine. In addition to the present work investigating the effect of IRI, prior studies have also documented similar changes in the conversion of HP [1-¹³C]pyruvate after liver injury induced by hepatotoxic agents carbon tetrachloride (10) and 1,3-dichloro-2-propanol (11) (the latter being earlier work from the same group that authored this new article), which chemically induce inflammation and oxidative stress in the liver. These findings suggest utility of the HP ¹³C MRI approach for monitoring liver injury associated with non-alcoholic fatty liver disease (NAFLD) and transition to non-alcoholic steatohepatitis (NASH), an important clinical dilemma that is not adequately addressed by existing diagnostic approaches (12). If identified at an early stage, patients with NASH could benefit from therapies to mitigate the harmful effects of oxidative stress (e.g. vitamin E supplementation) and/or improve their insulin sensitivity (e.g. glitazones), the traditional “two hits”, in addition to several other experimental treatments that are rapidly being developed by pharmaceutical companies to combat this growing epidemic. As a frequent contributing factor, the influence of insulin resistance must also be considered here. Diabetes (both insulin resistant or deficient) independently causes changes in hepatic HP lactate and alanine signals (13), which may be additive with changes due to liver injury. A promising approach for potentially improving the specificity of detected hepatic signal changes is the application of a liver-targeted paramagnetic contrast agent (e.g. gadoxetate) to selectively quench signal arising from normal hepatocytes (14), as compared to infiltrative or transformed cell types.

Other HP ¹³C MRI probes could also provide further detail about the specific nature of liver injury, when deployed separately or even simultaneously with [1-¹³C]pyruvate as a multi-agent diagnostic cocktail (15). For example, assessment of hepatic energy charge based on phosphorylation of HP [2-¹³C]dihydroxyacetone (DHA) (16) to [2-¹³C]glycerol-3-phosphate (G3P) complements the redox-based assessment offered by HP [1-¹³C]pyruvate. This is significant because a progressive derangement of normal hepatic energy metabolism, centered on mitochondria, is known to underlie progression of NAFLD to NASH. Hepatic conversion of HP [2-¹³C]DHA to [2-¹³C]G3P was recently shown to be markedly attenuated by an ATP-depleting fructose challenge in rats (17). Additionally, conversion of HP [1,4-¹³C₂]fumarate to [1,4-¹³C₂]malate has been used to detect hepatic cellular necrosis following transcatheter arterial embolization (TAE) of human orthotopic liver tumors implanted in rats (18). Only following the compromise of plasma membranes in association with necrosis can fumarate (a dicarboxylate with relatively low membrane permeability) be converted enzymatically to malate on the timescale of the HP ¹³C experiment. Finally, it is also possible to monitor oxidative stress directly based on conversion of HP [1-¹³C]dehydroascorbate (also abbreviated DHA) (19) to [1-¹³C]vitamin C, in a reaction whose reducing power is believed to be supplied principally by reduced glutathione (GSH). This approach has been applied to detect liver injury in a mouse model of steatohepatitis (methionine-choline deficient diet) (20).

In summary, the exciting new results of Moon et al. in liver IRI add to a growing body of evidence showing great potential value of new HP ¹³C MRI technology as a new means for non-invasive diagnosis and monitoring of liver injury. This line of research is progressing quickly, with application to human studies of liver injury likely in the coming years.

Abbreviations

HP

hyperpolarized

MRI

magnetic resonance imaging

IRI

ischemia reperfusion injury

DNP

dynamic nuclear polarization

EPA

electron paramagnetic agent

LDH

lactate dehydrogenase

ALT

alanine transaminase

PDH

pyruvate dehydrogenase

NAFLD

non-alcoholic fatty liver disease

NASH

non-alcoholic steatohepatitis

DHA

dihydroxyacetone [disambiguation 1]

G3P

glycerol-3-phosphate

TAE

transcatheter arterial embolization

DHA

dehydroascorbate [disambiguation 2]

GSH

reduced glutathione