12 research outputs found
Untargeted Metabolomics Analysis Reveals a Link between ETHE1-Mediated Disruptive Redox State and Altered Metabolic Regulation
Defects in the gene encoding the
persulfide dioxygenase ETHE1 are
known to cause the severe inherited metabolic disorder ethylmalonic
encephalopathy (EE). In spite of known clinical characteristics, the
molecular mechanisms underlying the ETHE1 deficiency are still obscure.
Herein, to further analyze the molecular phenotype of the disease,
we applied an untargeted metabolomics approach on cultivated fibroblasts
of EE patients for pinpointing alterations in metabolite levels. Metabolites,
as direct signatures of biochemical functions, can decipher biochemical
pathways involved in the cellular phenotype of patient cells. Using
liquid chromatography–mass spectrometry-based untargeted metabolomics,
we identified 18 metabolites that have altered levels in fibroblasts
from EE patients. Our data demonstrate disrupted redox state in EE
patient cells, which is reflected by significantly decreased level
of reduced glutathione. Furthermore, the down-regulation of several
intermediate metabolites such as the redox cofactors NAD<sup>+</sup> and NADH as well as Krebs cycle intermediates revealed clear alteration
in metabolic regulation. Pantothenic acid and several amino acids
exhibited decreased levels, whereas the β-citrylglutamate with
a putative role in brain development had an increased level in the
EE patient cells. These observations indicate the severe impact of
ETHE1 deficiency on cellular physiology and redox state, meanwhile
suggesting targets for experimental studies on novel treatment options
for the devastating metabolic disorder
A Metabolomics Study of Retrospective Forensic Data from Whole Blood Samples of Humans Exposed to 3,4-Methylenedioxymethamphetamine: A New Approach for Identifying Drug Metabolites and Changes in Metabolism Related to Drug Consumption
The
illicit drug 3,4-methylenedioxymethamphetamine (MDMA) has profound
physiological cerebral, cardiac, and hepatic effects that are reflected
in the blood. Screening of blood for MDMA and other narcotics are
routinely performed in forensics analysis using ultra-performance
liquid chromatography with high-resolution time-of-flight mass spectrometry
(UPLC–HR-TOFMS). The aim of this study was to investigate whether
such UPLC–HR-TOFMS data collected over a two-year period could
be used for untargeted metabolomics to determine MDMA metabolites
as well as endogenous changes related to drug response and toxicology.
Whole blood samples from living Danish drivers’ positive for
MDMA in different concentrations were compared to negative control
samples using various statistical methods. The untargeted identification
of known MDMA metabolites was used to validate the methods. The results
further revealed changes of several acylcarnitines, adenosine monophosphate,
adenosine, inosine, thiomorpholine 3-carboxylate, tryptophan, <i>S</i>-adenosyl-l-homocysteine (SAH), and lysophospatidylcholine
(lysoPC) species in response to MDMA. These endogenous metabolites
could be implicated in an increased energy demand and mechanisms related
to the serotonergic syndrome as well as drug induced neurotoxicity.
The findings showed that it was possible to extract meaningful results
from retrospective UPLC–HR-TOFMS screening data for metabolic
profiling in relation to drug metabolism, endogenous physiological
effects, and toxicology
Infarct size and correlation with blood glucose.
<p><b>A.</b> Infarct size/area-at-risk ratio (IS/AAR). Black bars: Control, White bars: ZDF. <b>B.</b> Correlations between preoperative fasting blood glucose and IS/AAR at prediabetes, onset and late type 2 diabetes mellitus. Mean ± SEM. Circles: ZDF 6 weeks, Squares: ZDF 12 weeks, Triangles: ZDF 24 weeks.</p
Myocardial glycogen concentrations.
<p>Myocardial concentrations of total-glycogen in vivo (<b>A</b>) and after ischemia and 30 min reperfusion (<b>B</b>) in control and ZDF rats. Mean ± SEM. Black bars: Control, White bars: ZDF. <b>IR</b>: ischemia-reperfusion. p<0.0001 vs. age-matched control and ZDF 6 weeks.</p
Study design and experimental protocol.
<p>Overview of groups, number of animals (<b>N</b>) and experimental protocols. <b>KH-buffer:</b> Krebs-Henseleit buffer.</p
Expression of proteins involved in the malate-aspartate shuttle.
<p>Fold change in expression of proteins participating in the malate-aspartate shuttle. Controls are set to 0 at all ages. Bars indicate fold change in ZDF hearts compared to control hearts. <b>A:</b> Transporter proteins. <b>B:</b> Enzymes. C: Representative western blots. There were no significant changes in expression at 6 and 12 weeks, while citrin and EAAT1 were significantly, and aralar non-significantly, downregulated at 24 weeks of age. Mean ± SEM. <b>EAAT</b>: Excitatory Amino Acid Transporter. <b>ASAT:</b> Aspartate Amino Acid Transaminase. <b>MDH:</b> Malate Dehydrogenase. *p<0.05.</p
Animal characteristics.
<p>Mean±SEM.</p>*<p>p<0.05 compared to age-matched controls.</p>†<p>p<0.05 compared to 6 weeks control/ZDF.</p
Interstitial concentrations of metabolites.
<p>Interstitial concentrations of glycolytic - and citric acid cycle metabolites measured by myocardial microdialysis during ischemia-reperfusion in control (closed squares) and ZDF (open squares) at 12 weeks. Mean ± SEM. <b>A:</b> Lactate, <b>B:</b> Pyruvate, <b>C:</b> Succinate, <b>D:</b> Fumarate, <b>E:</b> Malate.</p
Glucose metabolism prior to ischemia-reperfusion.
<p>Glucose uptake – glycolysis- and glucose oxidation rates during stabilization. Mean ± SEM. Black bars: Control, White bars: ZDF.</p
Glucose metabolism during reperfusion.
<p><b>A</b>: Glycolysis and <b>B:</b> Glucose oxidation rates during reperfusion at 6, 12 and 24 weeks. Mean ± SEM. Closed symbols: Control, Open symbols: ZDF. P-values indicate differences between control and ZDF from 0–5 and 10–30 minutes of reperfusion, respectively.</p