The ability of urinary metabolomics to detect meaningful, tissue-specific, biological effects (i.e., toxicity, disease) is compounded by high background variability. We hypothesize that sensitivity can be enhanced by imposing a tissue-targeted metabolic stressor.
We tested whether the sensitivity of metabolomics to assess kidney function is improved under the diuretic stress of furosemide.
To mildly compromise kidney, rats were given a sub-acute dose of d-serine. Then at 24 h postdose, we administered vehicle solution (control) or the diuretic drug, furosemide, and conducted NMR-based urinary metabolomics.
Principal Components and OPLS discriminant analyses showed no effects on urinary profiles in rats receiving d-serine alone. However, the effects of d-serine were observable under furosemide-induced stress, as urinary profiles classified separately from rats receiving furosemide alone or vehicle-treated controls (p < 0.001). Furthermore, this profile was uniquely different from a co-treatment effect observed following co-administration of d-serine + furosemide. We identified 24 metabolites to classify the effects of furosemide in normal rats vs. d-serine-compromised rats. Most notably, a furosemide-induced increase in urinary excretion of α-ketoglutarate, creatinine, trigonelline, and tryptophan in control rats, was significantly reduced in d-serine exposed rats (p < 0.05). Interestingly, increased tryptophan metabolism has been shown to correlate with the severity of kidney transplant failure and chronic kidney disease.
We attribute these effects to differences in kidney function, which were only detectable under the stress imposed by furosemide. This technique may extend to other organ systems and may provide improved sensitivity for assessment of tissue function or early detection of disease.
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We “spiked” a urine sample with furosemide at a concentration equivalent to that which would be expected assuming that the entire furosemide dose was excreted into urine within 24 h postdose (~0.3 mM). The observable furosemide signals were very weak, and in experimental animal samples the signals were barely detectable and did not interfere with our analyses, as none were chosen as salient features for class discrimination. Detection of furosemide metabolites was ignored since >80% is known to be excreted as the parent compound.
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This work was supported by a grant from the Ohio Third Frontier, Research Challenge Fund (NVR), and funds provided by the Air Force Research Laboratory, 711th Human Performance Wing, Human Effectiveness Directorate, Wright-Patterson Air Force Base, OH (DRM).
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All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. All procedures performed in studies involving animals were in accordance with the ethical standards of the institution at which the studies were conducted.
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Sibomana, I., DelRaso, N.J., Mattie, D. et al. Furosemide enhances the sensitivity of urinary metabolomics for assessment of kidney function. Metabolomics 13, 24 (2017). https://doi.org/10.1007/s11306-017-1162-6