Prostaglandins, Leukotrienes and Essential Fatty Acids
Novel plasma phospholipid biomarkers of autism: Mitochondrial dysfunction as a putative causative mechanism
Introduction
Autism is a lifelong disorder of unknown origin. The disorder is characterized by behavioral, developmental, neuropathological and sensory abnormalities [1], and is usually diagnosed between the ages of 2 and 10 with peak prevalence rates observed in children aged 5–8 years [2]. Autism is the most familiar diagnosis among the Autism Spectrum Disorders (ASDs), which encompass multiple conditions involving deficits in social and behavioral interactions, as well as development of normal interpersonal relationships [3]. In addition to autism, Asperger syndrome and pervasive developmental disorder-not otherwise specified (PDD-NOS) are also included in ASDs. Autistic disorder has three core symptomatic domains: deficits in communication, abnormal social interactions and restrictive and/or repetitive interests and behaviors [3]. There is also a significant gender bias in autism, with approximately four times more boys diagnosed than girls [4]. Neuropathological studies in autism have shown increased microglial activation [5], decreased cerebellar Purkinje cell density [6], [7] and abnormal brain swelling, particularly in white matter [8], [9]. Biochemical studies have shown increased oxidative stress [10], [11], [12], [13], [14], [15], abnormal glutathione metabolism [13], decreased melatonin [16] and increased docosahexaenoic acid (DHA) [17] in autistic subjects. Although there is debate as to whether autism has a pre- [8] or post-natal origin [18], it is generally accepted that the symptoms and pathology persist throughout the life of the subject [19]. These studies suggest that there is an underlying and ongoing biochemical abnormality in autism, regardless of its origin.
The following studies were designed to critically examine the hypothesis that the manifestation of the symptoms and pathology of autism is the result of metabolic dysregulation and toxicity, regardless of the initial causal factor or factors of autism. Our first objective was to accurately assess the individual metabolic status of autistic subjects relative to asymptomatic siblings and normal children with no family history of autism. This was accomplished by obtaining three fasting plasma samples from each participant over the course of 1 year so that a true individualized assessment of each subject could be obtained. We included affected/non-affected sibling pairs (high-risk controls), as well as subjects with no family history of autism (low-risk controls). An initial metabolic analysis was performed on all subject samples using a Fourier transform ion cyclotron resonance mass spectrometry (FTMS)-based comprehensive non-targeted metabolomic platform [20], which revealed consistent alterations in the levels of very long chain fatty acid (VLCFA)-containing phosphatidylethanolamines (PtdEtns) and in DHA-containing ethanolamine plasmalogens (PlsEtns). A more thorough investigation of PtdEtn-containing fatty acids ranging from 16 to 40 carbon units and from zero to six double bonds and selected PlsEtn using targeted assays on a triple-quadrupole mass spectrometer was then undertaken. These findings are reported herein and suggest a possible disruption of fatty acid metabolism due to compromised mitochondrial function. Mitochondrial stress was assessed through measurements of reduced glutathione (GSH) and related metabolites. In addition, we investigated and compared the in vitro effects of glutamate toxicity on neuronal, astrocyte and hepatocyte cell cultures to biomarker changes observed in the autistic subjects. Impaired mitochondrial fatty acid oxidation as the underlying cause of elevated plasma levels of VLCFA-containing PtdEtn is hypothesized. In addition, the putative down-stream effects of these metabolic abnormalities are discussed in relation to the known biochemical, neuropathological and epidemiological characteristics of autism.
Section snippets
Clinical information
Subjects participating in the study consisted of children from families enrolled by the Jonty Foundation (Saint-Paul, Minnesota, USA). Informed consent was obtained for all subjects studied. For all subjects a detailed clinical and lifestyle history was collected. All autistic subjects met DSM-IV criteria for autism. No other significant health disorders were present. Controls were either siblings of autistic subjects (9 out of 12 controls) or age-matched children from families with no history
Key metabolic changes in autism
A comprehensive assessment of PtdEtn and PlsEtn levels using triple–quadrupole tandem MS was performed on three plasma draws from 15 autistic subjects and 12 age-matched controls. A full report of these analyses is presented in Supplementary Tables 3A–C. Significant elevations in DHA (22:6)-containing PlsEtn and PtdEtn and VLCFA (26:3 and 28:0)-containing PtdEtn were the most apparent, particularly those containing palmitic acid (16:0) at the sn-1 position. Since 18:3 is the obligate precursor
Discussion
The results presented in this study indicate that chronic mitochondrial stress is pervasive in autism and that elevated levels of fatty acid elongation and desaturation products are useful metabolic biomarkers of both mitochondrial stress and autism. Carnitine supplementation appears to mask these mitochondrial stress biomarkers by shifting the observed elevated species from DHA to arachidonic acid (Fig. 3). Although the underlying cause of this mitochondrial stress in situ, is, as yet,
Acknowledgements
We are greatly indebted to the Jonty Foundation and all the families who participated in the study.
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