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Noninvasive measurements of glycogen in perfused mouse livers using chemical exchange saturation transfer NMR and comparison to (13)C NMR spectroscopy.

Miller CO, Cao J, Chekmenev EY, Damon BM, Cherrington AD, Gore JC - Anal. Chem. (2015)

Bottom Line: Glycogen measurements from serially acquired CEST Z-spectra of livers were compared with measurements from interleaved natural abundance (13)C NMR spectra.We also observed that the CEST signal from glycogen in liver was significantly less than that observed from identical amounts in solution.Our results demonstrate that CEST provides an accurate, precise, and readily accessible method to noninvasively measure liver glycogen levels and their changes.

View Article: PubMed Central - PubMed

Affiliation: †Merck Research Laboratories, 2000 Galloping Hill Rd., Kenilworth, New Jersey 07033, United States.

ABSTRACT
Liver glycogen represents an important physiological form of energy storage. It plays a key role in the regulation of blood glucose concentrations, and dysregulations in hepatic glycogen metabolism are linked to many diseases including diabetes and insulin resistance. In this work, we develop, optimize, and validate a noninvasive protocol to measure glycogen levels in isolated perfused mouse livers using chemical exchange saturation transfer (CEST) NMR spectroscopy. Model glycogen solutions were used to determine optimal saturation pulse parameters which were then applied to intact perfused mouse livers of varying glycogen content. Glycogen measurements from serially acquired CEST Z-spectra of livers were compared with measurements from interleaved natural abundance (13)C NMR spectra. Experimental data revealed that CEST-based glycogen measurements were highly correlated with (13)C NMR glycogen spectra. Monte Carlo simulations were then used to investigate the inherent (i.e., signal-to-noise-based) errors in the quantification of glycogen with each technique. This revealed that CEST was intrinsically more precise than (13)C NMR, although in practice may be prone to other errors induced by variations in experimental conditions. We also observed that the CEST signal from glycogen in liver was significantly less than that observed from identical amounts in solution. Our results demonstrate that CEST provides an accurate, precise, and readily accessible method to noninvasively measure liver glycogen levels and their changes. Furthermore, this technique can be used to map glycogen distributions via conventional proton magnetic resonance imaging, a capability universally available on clinical and preclinical magnetic resonance imaging (MRI) scanners vs (13)C detection, which is limited to a small fraction of clinical-scale MRI scanners.

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Correlation of CEST vs 13CNMR determined total glycogenin perfused livers under baseline conditions (blue) and followingglucagon addition (red). Note that the slopes in the two plots appearto be similar while the Y-intercept is higher inthe red group due to the release of glucose stimulated by glucagon.
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fig7: Correlation of CEST vs 13CNMR determined total glycogenin perfused livers under baseline conditions (blue) and followingglucagon addition (red). Note that the slopes in the two plots appearto be similar while the Y-intercept is higher inthe red group due to the release of glucose stimulated by glucagon.

Mentions: Figure 7 shows a plot of 13CNMR determined glycogen versus CEST MTRasym AUC0.5–2.5 for data acquired before (blue) and after (red) glucagon. The R2 values were 0.88 ± 0.054 and 0.87 ±0.040; the slope values were 0.0091 ± 0.00078 and 0.0082 ±0.00064, and the Y-intercept values were −0.50± 0.38 and 2.4 ± 0.21, respectively (mean ± SD). Errorbars for the data points in Figure 7 as wellas for the standard deviations for the correlation parameters weredetermined by Monte Carlo simulations (see Methods). We observed a strong linear relationship between 13C NMR determined glycogen and CEST MTRasym AUC0.5–2.5 both before and after glucagon treatment. The slope of the relationshipwas similar in both groups as evidenced by the overlapping standarddeviations. This slope value can be used in future studies as a calibrationfactor between CEST MTRasym AUC0.5–2.5 and total perfused liver glycogen. The fact that the Y-intercept is within two standard deviations of zero (i.e., not statisticallydifferent from zero) in the data obtained before glucagon additiondemonstrates that there are few competing endogenous CEST metabolitesin this spectral region of the liver. The increased Y-intercept value observed after glucagon addition is attributed toglucose release from the liver adding an additional CEST signal duringthis period of the experiment.


Noninvasive measurements of glycogen in perfused mouse livers using chemical exchange saturation transfer NMR and comparison to (13)C NMR spectroscopy.

Miller CO, Cao J, Chekmenev EY, Damon BM, Cherrington AD, Gore JC - Anal. Chem. (2015)

Correlation of CEST vs 13CNMR determined total glycogenin perfused livers under baseline conditions (blue) and followingglucagon addition (red). Note that the slopes in the two plots appearto be similar while the Y-intercept is higher inthe red group due to the release of glucose stimulated by glucagon.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4920106&req=5

fig7: Correlation of CEST vs 13CNMR determined total glycogenin perfused livers under baseline conditions (blue) and followingglucagon addition (red). Note that the slopes in the two plots appearto be similar while the Y-intercept is higher inthe red group due to the release of glucose stimulated by glucagon.
Mentions: Figure 7 shows a plot of 13CNMR determined glycogen versus CEST MTRasym AUC0.5–2.5 for data acquired before (blue) and after (red) glucagon. The R2 values were 0.88 ± 0.054 and 0.87 ±0.040; the slope values were 0.0091 ± 0.00078 and 0.0082 ±0.00064, and the Y-intercept values were −0.50± 0.38 and 2.4 ± 0.21, respectively (mean ± SD). Errorbars for the data points in Figure 7 as wellas for the standard deviations for the correlation parameters weredetermined by Monte Carlo simulations (see Methods). We observed a strong linear relationship between 13C NMR determined glycogen and CEST MTRasym AUC0.5–2.5 both before and after glucagon treatment. The slope of the relationshipwas similar in both groups as evidenced by the overlapping standarddeviations. This slope value can be used in future studies as a calibrationfactor between CEST MTRasym AUC0.5–2.5 and total perfused liver glycogen. The fact that the Y-intercept is within two standard deviations of zero (i.e., not statisticallydifferent from zero) in the data obtained before glucagon additiondemonstrates that there are few competing endogenous CEST metabolitesin this spectral region of the liver. The increased Y-intercept value observed after glucagon addition is attributed toglucose release from the liver adding an additional CEST signal duringthis period of the experiment.

Bottom Line: Glycogen measurements from serially acquired CEST Z-spectra of livers were compared with measurements from interleaved natural abundance (13)C NMR spectra.We also observed that the CEST signal from glycogen in liver was significantly less than that observed from identical amounts in solution.Our results demonstrate that CEST provides an accurate, precise, and readily accessible method to noninvasively measure liver glycogen levels and their changes.

View Article: PubMed Central - PubMed

Affiliation: †Merck Research Laboratories, 2000 Galloping Hill Rd., Kenilworth, New Jersey 07033, United States.

ABSTRACT
Liver glycogen represents an important physiological form of energy storage. It plays a key role in the regulation of blood glucose concentrations, and dysregulations in hepatic glycogen metabolism are linked to many diseases including diabetes and insulin resistance. In this work, we develop, optimize, and validate a noninvasive protocol to measure glycogen levels in isolated perfused mouse livers using chemical exchange saturation transfer (CEST) NMR spectroscopy. Model glycogen solutions were used to determine optimal saturation pulse parameters which were then applied to intact perfused mouse livers of varying glycogen content. Glycogen measurements from serially acquired CEST Z-spectra of livers were compared with measurements from interleaved natural abundance (13)C NMR spectra. Experimental data revealed that CEST-based glycogen measurements were highly correlated with (13)C NMR glycogen spectra. Monte Carlo simulations were then used to investigate the inherent (i.e., signal-to-noise-based) errors in the quantification of glycogen with each technique. This revealed that CEST was intrinsically more precise than (13)C NMR, although in practice may be prone to other errors induced by variations in experimental conditions. We also observed that the CEST signal from glycogen in liver was significantly less than that observed from identical amounts in solution. Our results demonstrate that CEST provides an accurate, precise, and readily accessible method to noninvasively measure liver glycogen levels and their changes. Furthermore, this technique can be used to map glycogen distributions via conventional proton magnetic resonance imaging, a capability universally available on clinical and preclinical magnetic resonance imaging (MRI) scanners vs (13)C detection, which is limited to a small fraction of clinical-scale MRI scanners.

Show MeSH
Related in: MedlinePlus