Urinary volatile metabolites of amygdala-kindled mice reveal novel biomarkers associated with temporal lobe epilepsy

Abstract

Epilepsy is a chronic neurological disorder affecting mammals, including humans. Uncontrolled epilepsy is associated with poor quality of life, accidents, and sudden death. In particular, temporal lobe epilepsy (TLE) is the most common type of pharmacoresistant epilepsy, which easily gets out of control in human adults. The aim of this study was to profile urinary volatile organic compounds (VOCs) in a mouse model of TLE using solid-phase microextraction (SPME) gas chromatography mass spectrometry (GC-MS). Thirteen urinary VOCs exhibited differential abundance between epileptic and control mice, and the corresponding areas under the receiver operating characteristic (ROC) curve were greater than 0.8. Principal component analysis (PCA) based on these 13 VOCs separated epileptic from sham operated-mice, suggesting that all these 13 VOCs are epilepsy biomarkers. Promax rotation and dendrogram analysis concordantly separated the 13 VOCs into three groups. Stepwise linear discriminant analysis extracted methanethiol; disulfide, dimethyl; and 2-butanone as predictors. Based on known metabolic systems, the results suggest that TLE induced by amygdala stimulation could affect both endogenous metabolites and the gut flora. Future work will elucidate the physiological meaning of the VOCs as end-products of metabolic networks and assess the impact of the metabolic background involved in development of TLE.

Review on Volatolomic Studies as a Frontier Approach in Animal Research

“…[ 47 ] Until analysis, the collected urine was stored in vials at –20 °C, [ 47,51 ] –80 °C, [ 50,52,53 ] or in a nitrogen gas tank. [ 54 ] In all studies, urine VOCs were extracted with an SPME fiber from the headspace formed inside sealed vials heated at 37 °C for 30 min, at 45 °C for 50 or 60 min, or at 60 °C for 10 or 30 min. The reported urine amounts and vial volumes were 200 µL urine in 2 mL vials, 25 µL urine in 4 mL vials, and 75 µL urine in 10 mL vials, respectively.…”

“…Mice urine was analyzed for two other induced neurological disorders. For temporal lobe epilepsy, there were found 13 possible biomarkers (Table 6—Section E), [ 54 ] while for anxiety and depression there were identified eight possible biomarkers (Table 6—Section F). [ 53 ] Nevertheless, none of these biomarkers were shared by both diseases.…”

Liquid Biopsies in Epilepsy: Biomarkers for Etiology, Diagnosis, Prognosis, and Therapeutics

“…These nucleic acids are often contained within exosomes released into the bloodstream from blood cells, the brain, and the spine [43]. Advances in sequencing technology have allowed higherlevel molecular profiling of blood, CSF, and more recently urine and saliva, to investigate nucleic acids within epilepsy outside of the exosome [44][45][46]. While evidence on the utility of circulating nucleic acids in epilepsy is underdeveloped in comparison to other diseases, we review the current landscape below.…”

Urinary volatilome analysis in a mouse model of anxiety and depression

“…The fibers were preconditioned at 250˚C for 30 min and inserted into the headspace of the vial after the vial was equilibrated for 1 min at 45˚C. The volatile compounds in the headspace were extracted using SPME fibers at 45˚C for 60 min as previously described [35,36].…”

“…To separate the desorbed volatiles, InertCap PureWAX with ProGuard and T.L. column (60 m + 10 m proguard line and 2 m transfer line, 0.25 mm internal diameter, 0.5 μm film thick; GL science, Tokyo, Japan) was used as previously described by Fujita et al [36]. The oven temperature was programmed as follows: held at 40˚C for 10 min, ramped to 240˚C at 5˚C/min, and held for 10 min.…”