Hyperpolarized 13 C magnetic resonance evaluation of renal ischemia reperfusion injury in a murine model.

Acute kidney injury (AKI) is a major risk factor for the development of chronic kidney disease (CKD). Persistent oxidative stress and mitochondrial dysfunction are implicated across diverse forms of AKI and in the transition to CKD. In this study, we applied hyperpolarized (HP) 13 C dehydroascorbate (DHA) and 13 C pyruvate magnetic resonance spectroscopy (MRS) to investigate the renal redox capacity and mitochondrial pyruvate dehydrogenase (PDH) activity, respectively, in a murine model of AKI at baseline and 7 days after unilateral ischemia reperfusion injury (IRI). Compared with the contralateral sham-operated kidneys, the kidneys subjected to IRI showed a significant decrease in the HP 13 C vitamin C/(vitamin C + DHA) ratio, consistent with a decrease in redox capacity. The kidneys subjected to IRI also showed a significant decrease in the HP 13 C bicarbonate/pyruvate ratio, consistent with impaired PDH activity. The IRI kidneys showed a significantly higher HP 13 C lactate/pyruvate ratio at day 7 compared with baseline, although the 13 C lactate/pyruvate ratio was not significantly different between the IRI and contralateral sham-operated kidneys at day 7. Arterial spin labeling magnetic resonance imaging (MRI) demonstrated significantly reduced perfusion in the IRI kidneys. Renal tissue analysis showed corresponding increased reactive oxygen species (ROS) and reduced PDH activity in the IRI kidneys. Our results show the feasibility of HP 13 C MRS for the non-invasive assessment of oxidative stress and mitochondrial PDH activity following renal IRI.

Molecular detection of inflammation in cell models using hyperpolarized 13C-pyruvate.

The detection and treatment monitoring of inflammatory states remain challenging in part due to the multifactorial mechanisms of immune activation and spectrum of clinical manifestations. Currently, diagnostic strategies tend to be subjective and limited quantitative tools exist to monitor optimal treatment strategies. Pro-inflammatory M1 polarized macrophages exhibit a distinct metabolic glycolytic phenotype compared to the continuum of M2 polarization states. In the present study, the distinct metabolic phenotypes of resting and activated macrophages were successfully characterized and quantified using hyperpolarized carbon-13 (13C) labeled pyruvate and its metabolic products, i.e. lactate, as a biomarker of resting, disease and treated states. Methods: Mouse macrophage J774A.1 cells were used as a model system in an NMR compatible bioreactor to facilitate dynamic hyperpolarized 13C measurements. The glycolytic metabolism of the cells in the quiescent or resting state were compared with macrophages stimulated by lipopolysaccharide, a classical M1 activator using hyperpolarized 13C labeled pyruvate. Additionally, the activated macrophages were also treated with a non-steroidal anti-inflammatory drug to assess the changes in hyperpolarized lactate signal. The hyperpolarized lactate signals were then correlated using biochemical and molecular assays. Results: We first validated our model system of inflammatory cells by the hallmarks of M1 polarization using steady state metabolic profiling with high resolution NMR in conjunction with nitric oxide Greiss assay, enzyme activity, and mRNA expression. Subsequently, we clearly showed that the cutting edge technology of hyperpolarized 13C NMR can be used to detect elevated lactate levels in M1 polarized macrophages in comparison to control and non-steroidal anti-inflammatory drug treated M2 states. Conclusion: Hyperpolarized 13C lactate has the potential to serve as a biomarker to non-invasively detect and quantify pro-inflammatory state of immune regulatory cells and its response to therapy.