A new hyperpolarized 13C ketone body probe reveals an increase in acetoacetate utilization in the diabetic rat heart

Figures

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  Figure 1. Diabetic characteristics of GK rats. The change in blood glucose levels within 2 hour duration upon (a) intraperitoneal injection of oral glucose (IpGTT) and upon (b) intraperitoneal injection of insulin (IpITT). Data are means ± SD (controls n = 5, GK n = 5). *P < 0.05 vs. controls.

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  Figure 2. 13C MR spectrum of 4 mM hyperpolarized [3-13C]acetoacetate. Upon injection of 80 mM hyperpolarized [3-13C]acetoacetate in vivo, the expected [3-13C]acetoacetate concentration in the blood is ~4 mM. The [3-13C]acetoacetate is seen at 209.9 ppm. The natural abundant [1-13C]acetoacetate appears at 174.6 ppm, and an impurity peak is visible at 181.0 ppm.

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  Figure 3. Detection of ketone metabolic products in the heart using hyperpolarized 13C MRS. (a) Metabolic fate of hyperpolarized [3-13C]acetoacetate. Metabolites in grey can be detected by in vivo 13C MRS in the heart. (b) Dynamic in vivo cardiac 13C MR spectra over 1 minute period after [3-13C]acetoacetate injection, with a time resolution of ~2 s. The spectra are truncated at 170 ppm. (c) A representative cardiac 13C MR spectrum, summed over 60 seconds (30 spectra) upon [3-13C]acetoacetate arrival, with [1-13C] urea used as an external reference. β-OHB: β-hydroxybutyrate, BDH1: 3-hydroxybutyrate dehydrogenase-1, SCOT: succinyl-CoA:3-ketoacid-coenzyme A transferase, ACAT: acetoacetyl-CoA thiolase, CAT: carnitine-acylcarnitine translocase, CS: citrate synthase, GDH: glutamate dehydrogenase, TCA: tricarboxylic acid.

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  Figure 4. Myocardial ketone body utilization in GK rats. The detection of [3-13C]acetoacetate, [1-13C]acetoacetate, [5-13C]glutamate, and [1-13C]acetylcarnitine over 2 minutes upon [3-13C]acetoacetate injection in (a) controls and (b) GK rats. (c) Representative cardiac 13C MR spectra from a control and a GK rat. The quantification of (d) [5-13C]glutamate + [1-13C]acetylcarnitine, (e) [5-13C]glutamate, (f) [1-13C]acetylcarnitine, and (g) [3-13C]β-OHB. Metabolic conversion rates for (h) [3-13C]acetoacetate to [5-13C]glutamate exchange and (i) [3-13C]acetoacetate to [1-13C]acetylcarnitine exchange. Data are means ± SD (except for (a) and (b): mean ± SEM), and normalized to [3-13C]acetoacetate (controls n = 10, GK n = 9; except for (i) GK n = 8). AcAc: acetoacetate, acc: acetylcarnitine, cit: citrate, glu: glutamate. *P < 0.05, **P < 0.01, ***P < 0.001 vs. controls.

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  Figure 5. Increased ketone body utilization in GK rats was associated with increased SCOT activity. (a) SCOT activity. Correlation between (b) SCOT activity and ([5-13C]glutamate + [1-13C]acetylcarnitine), and between (c) SCOT activity and [5-13C]glutamate. Data are means ± SD (controls n = 10, GK n = 9, except for the correlation n = 8). One outlier was not included in the correlation, see Supplementary Fig. S4 for complete data. Acc: acetylcarnitine, glu: glutamate. *P < 0.05 vs. controls.

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  Figure 6. GK rats exhibited cardiac hypertrophy and dysfunction. (a) LV mass, (b) diastolic wall thickness, (c) heart mass index (post-mortem), (d) end diastolic volume, (e) end systolic volume, (f) stroke volume, (g) heart rate, (h) cardiac output, and (i) ejection fraction. Data are means ± SD (controls n = 10, GK n = 9). **P < 0.01, ***P < 0.001 vs. controls.

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  Figure 7. Total of ketone body utilization products correlated with cardiac hypertrophy and function. Correlation between (a) ([5-13C]glutamate + [1-13C]acetylcarnitine) and cardiac function, and (b) between ([5-13C]glutamate + [1-13C]acetylcarnitine) and heart mass index. Controls n = 10, GK n = 8 (one outlier was not included, see Supplementary Fig. S4 for complete data). Acc: acetylcarnitine, glu: glutamate.