A metabolomics approach to investigate urine levels of neurotransmitters and related metabolites in autistic children

Significance 

Autism spectrum disorder (ASD) is a complex syndrome associated with stereotyped and repetitive patterns of behavior as well as communicative deficits. In the US, autism was considered an uncommon disorder about 20 years ago, as its incidence was 1 in 10,000 births. However, now its incidence in the US has reached a staggering 1 in 59 births. Currently, clinical observation using the criteria of the fifth edition of Diagnostic and Statistical Manual of Mental Disorders is used for the diagnosis of autism. It is believed that the availability of biochemical biomarkers in biological fluids that can diagnose autism at a younger age devoid of invasive methods will be helpful to clinicians in providing earlier and more reliable diagnoses. There have been recent studies that have linked alterations in the gut microbiota to ASD, although the mechanisms were not fully understood. Several investigations using metabolomics have been performed in the last decade to investigate the global biochemical metabolisms of autistic individuals so as to identify the alterations of specific metabolic pathways. The importance of finding biochemical biomarkers in biological fluids that can diagnose autism at a younger age, without invasive methods, could help clinicians to provide earlier and more reliable diagnoses. In particular, the usefulness of urine analysis, which is an easy-to-take sample for young kids it is as a fast reliable screening test in autistic children during the first few years of life.

Now a new study, Italian researchers: Dr. Federica Gevi, Antonio Belardo and led by Professor Lello Zolla from the University of Tuscia used advanced metabolomics approach to investigate the neurotransmitter levels in urine samples of autistic children and analysed the altered metabolic pathway involved in their production. Their findings showed alterations in the Tryptophan metabolism, in Vitamins B6, B9 and B12 absorption and in the level of metabolites involved in neurotransmitter production that correlated with a peculiar intestinal microbiome in autistic subjects. The last research article is now published in the Journal Biochimica et Biophysica Acta – Molecular Basis of Disease.

The research team explored ASD-specific urinary metabolomics patterns in 40 ASD children and 40 matched controls with the use of untargeted metabolomics through Ultra High Performance Liquid Chromatography mass spectrometry and then used XCMS Metlin software for data interpretation. They also observed that that 222 statistically significant metabolites were identified by the XCMS software to be up-regulated or down-regulated in autistic urine samples.

Specifically, the authors observed that in autistic children, there were higher levels of dopamine. The metabolomic analysis conducted also showed that the accumulation of dopamine in the urine of autistic subjects increased with the increase of 4-cresol. The levels of ascorbic acid, which is required by 4-cresol to inhibit dopamine β-hydroxylase was also found to be elevated.

They found that in autistic children, dopamine was not biotransformed into noradrenaline but prevalently into its metabolite homovanillic acid. Accordingly, the first noradrenaline metabolite, 3-methoxy-4hydroxyphenethyleneglycol and the last metabolite of noradrenaline degradation, vanillylmandelic acid were both found to be lower in autistic children. Furthermore, they also noticed that in autistic children, there were lower levels of the active form of vitamin B6, pyridoxal phosphate, which is an essential cofactor for the biotransformation of glutamate into GABA.  Consequently, higher levels of glutamate as well as lower concentrations of GABA were also recorded in autistic children.

4-Cresol is produced specifically by Clostridium difficile, which many authors have observed in the intestine of ASD children, therefore, their study has clarified the potential role of the gut microbiota in the regulation of neurological and neurodevelopmental disorders through the “microbiota-gutbrain axis”.  In fact, gut bacteria may also produce their own neurotransmitters, which simulating those of the host and may compromise host neural pathways, such as serotonin, GABA, noradrenaline and dopamine, as showed in this paper.

Reference

Gevi F, Belardo A, Zolla L. A metabolomics approach to investigate urine levels of neurotransmitters and related metabolites in autistic children. Biochim Biophys Acta Mol Basis Dis. 2020;1866(10):165859.

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