Who are the pleasant authors, what is the title of the study, and what year was it published?
Authors: S Jill James, Paul Cutler, Stepan Melnyk, Stefanie Jernigan, Laurette Janak, David W Gaylor, and James A Neubrander
Title: Metabolic biomarkers of increased oxidative stress and impaired
methylation capacity in children with autism. AmJ Clin Nutr 2004;
What is the study about?
This study is about the metabolic profile of children with autism and investigating if specific targeted nutritional interventions could improve the metabolic profile. The study compared the concentrations of plasma metabolites in the methionine cycle and transsulfuration pathway between autistic and control children, which could suggest a metabolic imbalance in children with autism, marked by increased oxidative stress and impaired methylation capacity.
What previous research did the authors cite on this topic?
Autistic individuals have a high prevalence of gastrointestinal disease and dysbiosis (a 2003 study), autoimmune disease (another 2003 study), and mental retardation (a study in 2002).
A significant role for genetics in the etiology of autism disorder is supported by a high concordance of autism between monozygotic twins and increased risks among siblings of affected children, as well as the association of autistic symptoms with several heritable genetic diseases (a third 2003 study; a study in 1998). Autism has been reported as a comorbid condition associated with Rett syndrome, fragile X, phenylketonuria, adenylosuccinate lyase deficiency, dihydropyrimidine dehydrogenase deficiency, and 5′-nucleotidase hyperactivity, although these genetic diseases account for less than 10% of autism cases (a fourth and fifth 2003 study; one in 1996; another in 1984; and one study in 2000).
The current study was prompted by the serendipitous observation in a previous study that the metabolic profiles of dizygotic twins —one with Down syndrome and one with autism— were virtually identical with respect to methionine cycle and transsulfuration metabolites (a study in 1982).
The methionine cycle is a process in the body that involves the conversion of methionine, an essential amino acid, into other important molecules. One of the key reactions in this cycle is the transfer of a methyl group, which is a chemical structure consisting of one carbon and three hydrogen atoms, from a molecule called 5-methyltetrahydrofolate to homocysteine. This reaction is dependent on vitamin B-12. The regenerated methionine can then be activated to form S-adenosylmethionine (SAM), which serves as a primary supplier of methyl groups for various cellular reactions.
SAM plays a crucial role in the methylation process, which involves adding a methyl group to DNA, RNA, proteins, phospholipids, and neurotransmitters. This methylation is important for regulating gene expression, protein function, and overall cellular activity. As a result of the methyl transfer from SAM, a molecule called S-adenosylhomocysteine (SAH) is produced.
SAH can be converted back to homocysteine and adenosine through the action of an enzyme called SAH hydrolase (SAHH). Homocysteine can be further metabolized in two ways. It can be remethylated back into methionine or irreversibly removed from the methionine cycle by an enzyme called cystathionine β-synthase (CBS). When homocysteine is removed from the cycle, it leads to a decrease in the production of SAM for methylation reactions and a decrease in the synthesis of cysteine and glutathione, which are important for antioxidant activity in the body.
In simpler terms, the methionine cycle is a process that converts methionine into other important molecules in the body. It involves the transfer of a methyl group, which is necessary for various cellular functions. This cycle is crucial for proper gene regulation, protein function, and overall cell activity. Disruptions in the methionine cycle can lead to decreased methylation activity and reduced antioxidant capacity.
What were the methods?
The metabolic study included a total of 20 autistic children and 33 control children. Among the autistic children, all were white, 14 were boys, 6 were girls, and 19 were diagnosed with regressive autism, while 1 had infantile autism. Prior to the study, 16 autistic children were already taking a multivitamin and mineral supplement containing 400 µg of folic acid and 3 µg of vitamin B-12. None of the autistic children were using prescribed medications that could impact methionine metabolism, such as valproic acid or anticonvulsants. The study did not administer a quantifiable diet questionnaire, so specific dietary differences within and between the groups could not be determined.
The control subjects in the study were healthy white US children without a history of chronic disease or autism who had previously participated in a baseline study involving children with Down syndrome. The control children were taking over-the-counter vitamin supplements and were not using medications known to interfere with methionine metabolism.
Methionine, S-adenosylmethionine (SAM), S-adenosylhomocysteine (SAH), SAM:SAH, Adenosine, Homocysteine, Cystathionine, Cysteine, total glutathione (tGSH), Oxidized glutathione, tGSH:GSSG.
These are metabolic biomarkers that reflect the function of the one-carbon metabolism and transsulfuration pathways, as well as oxidative stress and antioxidant capacity. These pathways have important roles in various physiological processes, including DNA synthesis and methylation, neurotransmitter synthesis, and redox balance.
In this study, the researchers obtained USP-grade folinic acid from Douglas Laboratories and Thorne Research, Inc. The folinic acid was administered twice a day as an oral powder in juice, with a dosage of 800 micrograms. They also acquired betaine (trimethylglycine) of USP grade from Life Extension Foundation, which was given twice a day as an oral powder in juice, with a dosage of 1000 micrograms. The researchers obtained USP methylcobalamin from Hopewell Pharmaceuticals and Unique Pharmaceuticals as an injectable liquid, which was administered subcutaneously at a dosage of 75 micrograms per kilogram of body weight, twice a week.
The metabolic study consisted of 3 parts. In the first component, baseline concentrations of plasma metabolites in the methionine cycle and transsulfuration pathway were measured in 20 autistic children and compared with plasma concentrations in 33 control children to establish whether the metabolic profile of the autistic children differed significantly from that of the control children.
In the second component, based on the observed abnormalities in plasma metabolites, a subset of 8 autistic children were given oral supplements of 800 μg folinic acid and 1000 mg betaine (anhydrous trimethylglycine) twice a day in an attempt to improve the metabolic profile; this is referred to as intervention 1. After 3 months on this regimen, blood samples were again taken and the metabolite concentrations were compared with baseline concentrations of each metabolite.
In the third component, the same subset of 8 children were given an injectable form of methylcobalamin (75 μg/kg) twice a week in addition to the oral folinic acid and betaine for an additional month; this is referred to as intervention 2. Each child served as his or her own control for the intervention study.
What were the findings?
The concentrations of metabolites in the methionine cycle and transsulfuration pathway differed significantly between autistic children and control children. In the methionine cycle, autistic children had lower plasma levels of methionine, S-adenosylmethionine (SAM), and homocysteine, while S-adenosylhomocysteine (SAH) and adenosine concentrations were higher compared to control children (Table 1). The SAM to SAH ratio was approximately 50% lower in autistic children compared to control children. Additionally, autistic children showed significant reductions in plasma cystathionine and cysteine concentrations, indicative of decreased CBS-mediated transsulfuration (Table 1). The lower mean plasma cysteine concentration was accompanied by an increase in oxidized glutathione (GSSG) levels by almost twofold and a 70% decrease in the ratio of total glutathione (tGSH) to GSSG (tGSH:GSSG).
The results showed that compared to their baseline levels, the concentrations of methionine, SAM, homocysteine, cystathionine, cysteine, and total glutathione (tGSH) in the plasma of these children increased. Additionally, the ratios of SAM to SAH and tGSH to GSSG also improved. Furthermore, the initially elevated levels of SAH and adenosine decreased with the supplementation. These changes brought the concentrations of these metabolites closer to those observed in the control children, indicating an improvement in their metabolic profiles.
Although the intervention successfully normalized most of the methionine cycle metabolites to levels seen in the control group, the concentrations of tGSH, GSSG, and the ratio of tGSH to GSSG were still not fully restored to normal levels. Despite this, the intervention showed significant improvement in these aspects.
The results showed that the addition of methylcobalamin did not significantly change the concentrations of methionine, SAM, SAH, or homocysteine compared to the changes observed with the intervention of folinic acid and betaine alone.
However, compared to the previous intervention, the addition of methylcobalamin further decreased the concentrations of adenosine and oxidized glutathione (GSSG). Additionally, it further increased the concentrations of methionine, cysteine, total glutathione (tGSH), and the ratios of SAM to SAH and tGSH to GSSG.
In simpler terms, the second intervention involving the addition of methylcobalamin did not have a significant effect on the levels of methionine, SAM, SAH, or homocysteine compared to the first intervention. However, it further decreased adenosine and oxidized glutathione levels, while increasing methionine, cysteine, total glutathione, and certain ratios related to these metabolites.
What are the implications of the study?
An increased vulnerability to oxidative stress and a decreased capacity for methylation may contribute to the development and clinical manifestation of autism. The observed imbalance in methionine and homocysteine metabolism in autistic children, characterized by reduced methionine, SAM, and homocysteine levels, along with increased adenosine and SAH levels, suggests a disruption in methionine synthase activity and decreased SAH hydrolysis. These alterations impact cellular methylation capacity and antioxidant activity, indicating a potential link to the development of autism.
The disrupted metabolic pattern in the transsulfuration pathway further supports the association between autism and oxidative stress. Lower levels of cystathionine, cysteine, and tGSH, along with an increase in oxidized glutathione (GSSG) and a decreased ratio of tGSH to GSSG, indicate chronic oxidative stress in autistic children. Oxidative stress affects enzymes within the methionine cycle and decreases CBS activity, leading to reduced cysteine synthesis and lower glutathione levels. These metabolic changes highlight the increased vulnerability of autistic children to oxidative stress.
The underlying causes of the observed oxidative stress and metabolic imbalances in autistic children are not fully understood. However, decreased adenosine deaminase activity and genetic variations associated with low adenosine deaminase activity may contribute to increased adenosine levels, which inhibit glutathione synthesis and exacerbate the imbalance. Genetic predisposition to environmental factors promoting oxidative stress could also play a role in the metabolic abnormalities observed in autism.
Targeted nutritional interventions have shown promise in addressing these metabolic imbalances. Supplementation with folinic acid and betaine successfully normalized the concentrations of metabolites within the methionine cycle and improved metabolites in the transsulfuration pathway. The addition of injectable methylcobalamin further enhanced methionine concentrations and positively affected adenosine, SAH, and glutathione levels, potentially alleviating oxidative stress. These interventions offer hope for improving the metabolic profiles and antioxidant activity in individuals with autism.
While specific improvements in speech and cognition were observed subjectively, further research is needed to quantitatively measure these outcomes. Additionally, dietary differences and genetic variations may influence the metabolic abnormalities seen in autism. Understanding the underlying metabolic mechanisms and their connection to genetic factors requires continued investigation. Nevertheless, the ability to correct metabolic imbalances through targeted nutritional interventions offers potential avenues for treating certain aspects of autism and improving the well-being of affected individuals.
What other research within the library is this study related to?
The study by El-Ansary et al. (2020) found that there was a complete separation of autistic and control participants using nine biomarkers, which provides insight into the possible pathophysiology of the autism, particularly in relation to oxidative stress, energy metabolism, mitochondrial dysfunction, and apoptosis.
The study by Al-Yafee et al. (2011) found multiple significant differences in the autism group on several metabolic biomarkers related to sulfur-dependent detoxification mechanisms, which are involved in the detoxification of neurotoxins and oxidative stress.
Can I read the full study somewhere?