Bisphenol A (BPA) and autism continued

"Descriptive analyses indicated that prenatal exposure to maternal BPA [Bisphenol A] concentrations were related to higher levels of anxiety, depression, aggression, and hyperactivity in children. BPA exposure in childhood was associated with higher levels of anxiety, depression, hyperactivity, inattention, and conduct problems."

That was the conclusion reached in the systematic review by Maede Ejaredar and colleagues [1], that provides one of two studies brought to the blogging table today. With the aim of looking at the current collected peer-reviewed literature on the topic of "prenatal and childhood BPA exposure" and offspring/childhood outcomes, the authors suggest that there may indeed be more to see in this area, but with the important requirement for further "Prospective cohort studies" to clarify any relationship.

BPA, by the way, is a chemical of some note in modern society given its quite widespread use in plastics and related materials. Although some agencies have provided current guidance to the effect that BPA is 'safe' and poses no health risk at current levels of exposure, not everyone is so convinced by such sweeping statements about safety.

Indeed to make the point, the findings reported by Meda Kondolot and colleagues [2] add to an existing body of scientific literature suggesting that when it comes to at least 'some' autism, there may be something of an increased biological burden of BPA present. Based on the analysis of 50+ children diagnosed with an autism spectrum disorder (ASD) and an equivalent number of asymptomatic - not autism - controls, a range of metabolites were looked for including "plasma phthalates and BPA" and compounds linked to "oxidant/antioxidant status." The authors reported that: "Plasma BPA levels of children with PDD-NOS [Pervasive Developmental Disorder-Not Otherwise Specified] were significantly higher than both classic autistic children and controls." Combined with some interesting findings potentially reflective of issues with oxidative stress in relation to their participant group, authors speculate that there may be some issues with the metabolism of things like BPA in relation to some autism. I might however also add that the chosen method of analysis of samples used by Kondolot et al - high performance liquid chromatography (HPLC) - is not the most sensitive of methods when used minus it's important detection counterpart, mass spectrometry; particularly when analysing such a complicated medium such as plasma. It implies that further, more technical investigation of samples, is perhaps required.

Being careful not the fall into any 'chemophobic' traps ('chemicals' is a word that has received a bad rap in my opinion), I continue to believe that there is more to see in this area of research. I draw back from any sweeping generalisations that have been put forward with other chemicals in relation to autism (see here for example) because I'm not convinced that all autism is due to BPA or any other single compound. Anyone who follows this blog regularly knows about my fascination with plural autisms (see here) and the fact that autism rarely exists in some sort of diagnostic vacuum (see here). That also takes into account the range of 'chemical exposures' that modern-day life brings.

I would however like to see further investigation on a few aspects: (i) is autism (some autism) associated with an increased exposure risk to certain chemicals? Y'know, the sort of data that is coming out of Vietnam for example (see here); and (ii) are there genetic and/or biological reasons why some people on the autism spectrum have issues with the metabolism of a range of xenobiotics? Under that last category, there are numerous examples in the literature of various compounds being elevated in terms of biological load (see here for example) combined with quite a bit of discussion about 'issues' with methods/systems for removing such compounds from the body (see here). Such findings could be just epiphenomenal to autism but could also represent something rather more central and important [3].

There is quite a bit more science to do in this area.

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[1]  Ejaredar M. et al. Bisphenol A exposure and children’s behavior: A systematic review. Journal of Exposure Science and Environmental Epidemiology. 2016. March 9.

[2] Kondolot M. et al. Plasma Phthalate and Bisphenol A Levels and Oxidant-Antioxidant Status in Autistic Children. Environmental Toxicology and Pharmacology. 2016. March 9.

[3] Kardas F. et al. Increased Serum Phthalates (MEHP, DEHP) and Bisphenol A Concentrations in Children With Autism Spectrum Disorder: The Role of Endocrine Disruptors in Autism Etiopathogenesis. J Child Neurol. 2016 Apr;31(5):629-35.

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Ejaredar M, Lee Y, Roberts DJ, Sauve R, & Dewey D (2016). Bisphenol A exposure and children's behavior: A systematic review. Journal of exposure science & environmental epidemiology PMID: 26956939




Kondolot, M., Ozmert, E., Ascı, A., Erkekoglu, P., Oztop, D., Gumus, H., Kocer-Gumusel, B., & Yurdakok, K. (2016). Plasma Phthalate and Bisphenol A Levels and Oxidant-Antioxidant Status in Autistic Children Environmental Toxicology and Pharmacology DOI: 10.1016/j.etap.2016.03.006

Endocrine disruptors and autism?

"Children with autism spectrum disorder had significantly increased serum MEHP, DEHP, and BPA [mono-(2-ethylhexyl)-phthalate (MEHP), di-(2-ethylhexyl)-phthalate (DEHP), and bisphenol A (BPA)] concentrations."

So said the findings reported by Fatih Kardas and colleagues [1] looking at whether there may be more to see when it comes to phthalate metabolism and autism among other things. For those who might be rolling their eyes at this point, I'll draw your attention to other occasions when similar results have been reported (see here and see here) and the place that the Kardas results seem to share.

This time around nearly 50 children diagnosed with an autism spectrum disorder (ASD) and some 40 asymptomatic controls provided serum samples that were screened for MEHP, DEHP, and BPA. The analytical weapon of choice was high-performance liquid chromatography (HPLC), I assume coupled to something like UV and/or fluorescence detection. The results by group suggested that "endocrine disruptors may have a role in the pathogenesis of autism spectrum disorders" according to the authors. I might add that we have had previous clues that this research was coming to publication [2].

These are interesting results. Whilst the use of HPLC (as a separative method) is not necessarily on its own the most accurate method of assaying for such compounds (that would be coupled to something like mass spectrometry) there are some potentially important things to learn from such findings. Although it would be easy to correlate such results with something like a higher exposure to such compounds in cases of autism, I'm not ready to accept that as an explanation given the frequency with which we all come into contact with them. I'd perhaps favour a hypothesis whereby the ability to metabolise such xenobiotics is differentially affected in at least some cases of autism as per the discussions by Stein and colleagues [3]. Such a metabolic difference could be a genetic issue tied into something like all that chatter about sulphation and glucuronidation and autism down the years (see here). It could also be something a little more functional in terms of how those trillions of wee beasties that call us home (the gut microbiome) might also affect such processes [4] too given the growing interest in the microbiome in relation to autism (see here). At this point I'll also draw your attention to some similarly interesting work on how functional bowel habits and gut microbiota might be linked with some autism in mind [5].

Acknowledging that autism research is still fumbling around a bit when it comes to the precise hows and whys of how environment (non-genetic factors) might fit into autism (see here) and it's relative contribution, I'm a great believer in how the technology we have at our disposal can help. The rise and rise of the various -omics and the concept of a systems biology approach with an autism slant (see here) means that we are getting better at not only analysing genetic and biological parameters but also in putting any potential connections together. Acknowledging that there may be many different types of autism (the autisms) with various different comorbidity patterns accompanying (see here), it is only a matter of time before patterns start to emerge and discussions can move on from simply genetics vs. environment.

Music: The Score - Oh My Love.

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[1] Kardas F. et al. Increased Serum Phthalates (MEHP, DEHP) and Bisphenol A Concentrations in Children With Autism Spectrum Disorder: The Role of Endocrine Disruptors in Autism Etiopathogenesis. J Child Neurol. 2015 Oct 8. pii: 0883073815609150.

[2] Kardas F. et al. P174 – 2732: Increased serum phthalates (MEHP, DEHP) and bisphenol A concentrations in children with autism: The role of endocrin disruptors in autism aetiopathogenesis. Euro J Pediatr Neurology. 2015; 19: Suppl. 1: S142-S143.

[3] Stein TP. et al. Bisphenol A Exposure in Children With Autism Spectrum Disorders. Autism Res. 2015 Jun;8(3):272-83.

[4] Rowland IR. Metabolism of Di-(2-ethylhexyl) phthalate by the contents of the alimentary tract of the rat. Food and Cosmetics Toxicology. 1974; 12: 293-302.

[5] Gabriele S. et al. Slow intestinal transit contributes to elevate urinary p-cresol level in Italian autistic children. Autism Res. 2015. October 6.

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Kardas F, Bayram AK, Demirci E, Akin L, Ozmen S, Kendirci M, Canpolat M, Oztop DB, Narin F, Gumus H, Kumandas S, & Per H (2015). Increased Serum Phthalates (MEHP, DEHP) and Bisphenol A Concentrations in Children With Autism Spectrum Disorder: The Role of Endocrine Disruptors in Autism Etiopathogenesis. Journal of child neurology PMID: 26450281

Baby teeth and autism research

"This report provides evidence that teeth can be useful biomarkers of early life exposure for use in epidemiologic case-control studies seeking to identify differential unbiased exposures during development between those with and without specific disorders such as autism."

That was one of the conclusions reached in the paper by Raymond Palmer and colleagues [1] (open-access available here) who played science tooth fairy with 71 deciduous teeth (baby teeth) provided by children with autism from a "tooth repository consisting of 928 children’s deciduous teeth." As per other reports with autism in mind using these potentially important biological samples [2], there is quite a lot of information potentially available from baby teeth assuming one knows how to handle such samples.

For Palmer et al the analytical methods of choice were "liquid chromatography tandem mass spectrometry and gas chromatography" echoing other autism biomarker research discussed on this blog (see here) and fairly commonly appearing in the peer-reviewed literature [3]. Levels of various compounds were assessed in the pulverised baby teeth samples including "acetaminophen [paracetamol], ARA [arachidonic acid], DEET [diethyl-m-toluamide], TCPy [3,5,6-trichloro-2-pyridinol], IMPy [2-isopropyl-6-methyl-4-pyrimidinol], and MEHP [mono-2-ethylhexyl phthalate]." Just in case you aren't an organic chemist, many of those compounds are metabolites of "pesticides, plastics, or medications" and were studied in light of previous work from this authorship group [4] and the suggestion that some of them might be relevant to autism risk and/or onset (see here for example) or at least serving up an interesting correlation.

Consistent with that previous report, Palmer and colleagues "demonstrated that specific semivolatile organic chemicals relevant to autism etiology can be detected in deciduous teeth." Drawing on information from both US children with autism (a "collection of deciduous teeth through collaborative efforts with the Interactive Autism Network (IAN)") and Mexican children with autism, various results are presented. "Despite demographic differences in the two samples, there were similar rates of detection for all chemicals" was one of the primary findings. So, round about 40% of samples from both geographic groups were detected to have traces of acetaminophen (paracetamol) in them for example. This finding tallied to some degree with parent report about paracetamol use during pregnancy and infancy. Other results can be seen in some of the accompanying tables (see here for example).

Importantly, Palmer and colleagues discuss what their findings do and do not mean. "While we have demonstrated that chemicals relevant to ASD [autism spectrum disorders] can be detected in deciduous teeth and are associated with mothers’ self-reported exposures, our results are limited in generalizability—largely due to the sample consisting entirely of children with ASD." In other words, the sole focus on baby teeth from children with autism does not necessarily mean that any compounds detected 'cause' autism given the snapshot view of their study and the lack of appropriate asymptomatic control samples analysed. Further work is indicated in this area to "include more diverse participants and neurotypical children as controls will allow case/control comparisons."

The use of baby teeth represents an interesting addition to tissue analysis when it comes to autism. Given the availability of such teeth, the timing of their availability and the relatively non-invasive way that such samples can be collected, there are quite a few positives to the use of such as resource in autism research. Combined with other fairly non-invasively collected samples such as urine (see here), saliva samples (see here) and potentially even things like nail clippings [5], there is quite a bit of information potentially available for autism research. That being said, the word 'biomarker' with autism in mind, needs to be rather carefully used in light of things like plurality...

Music: Maroon 5 - Sugar.

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[1] Palmer RF. et al. Organic Compounds Detected in Deciduous Teeth: A Replication Study from Children with Autism in Two Samples. J Environ Public Health. 2015;2015:862414.

[2] Adams JB. et al. Mercury, lead, and zinc in baby teeth of children with autism versus controls. J Toxicol Environ Health A. 2007 Jun;70(12):1046-51.

[3] Wang H. et al. Potential serum biomarkers from a metabolomics study of autism. J Psychiatry Neurosci. 2015 Sep 22;40(5):140009.

[4] Camann DE. et al. Acetaminophen, pesticide, and diethylhexyl phthalate metabolites, anandamide, and fatty acids in deciduous molars: potential biomarkers of perinatal exposure. Journal of Exposure Science and Environmental Epidemiology. 2013; 23: 190–196.

[5] Shu I. et al. Detection of Drugs in Nails: Three Year Experience. J Anal Toxicol. 2015 Oct;39(8):624-8.

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Palmer, R., Heilbrun, L., Camann, D., Yau, A., Schultz, S., Elisco, V., Tapia, B., Garza, N., & Miller, C. (2015). Organic Compounds Detected in Deciduous Teeth: A Replication Study from Children with Autism in Two Samples Journal of Environmental and Public Health, 2015, 1-9 DOI: 10.1155/2015/862414

Environmental exposure and autism continued

In a post not-so-long-ago I talked about the paper by Andrey Rzhetsky and colleagues [1] and their assertion that environment (various facets of environment) might correlate with the increasing numbers of cases of autism being diagnosed. As per what was said on that post, there were lots of media headlines generated about the findings; some balanced and some a little sensational.

Luc Viatour / www.Lucnix.be @ Wikipedia 

One of the main caveats I had with the Rzhetsky study was the reliance on statistical models onwards to their conclusions; so using surrogate markers of exposure and modelling trends but not actually looking at people in any biochemical or genetic sense or importantly, the metabolites of various candidate exposure compounds appearing in biofluids. Today therefore, I'm talking about the paper by Braun and colleagues [2] (open-access here) and some of the accompanying research looking at various measured chemical exposures in relation to autism.

The Braun paper is open-access but here are a few details:

• The aim: "To identify gestational EDC [endocrine disrupting chemical] exposures associated with autistic behaviors" was accomplished by screening 175 pregnant women who were part of the HOME study (looking at "the impact of low-level fetal and early childhood exposures to environmental chemicals on health developmental and behavioral outcomes") for various suspected EDCs in urine and serum mid-pregnancy.
• Said EDCs included "phthalate metabolites", "polychlorinated biphenyls (PCBs)", "brominated flame retardants" (including the PBDEs) and "perfluoroalkyl substances" (see here).
• Keeping the analytical data in mind, mothers then completed the Social Responsiveness Scale (SRS), a standardised instrument which quite accurately seems to perform when it comes to autism screening [3] when offspring were 4-5 years of age.
• Results: "Most of the EDCs were associated with negligible absolute differences in SRS scores", so no smoking gun but... "maternal serum concentrations of trans-nonachlor and PBDE-28 were associated with higher SRS scores". Indeed it's worth pointing out that some of the EDCs actually seemed to negatively correlate with SRS scores as for example: "PBDE-85, PCB-178, β-HCH, and PFOA concentrations were associated with less autistic behaviors".
• That also the authors "adjusted for numerous potential confounders including gestational tobacco smoke exposure, socioeconomic factors, perinatal factors, caregiving environment, maternal IQ, and maternal depressive symptoms" when analysing their results is an important strength of their study.

I was intrigued by the Braun results for several reasons. Bearing in mind this was a study looking at maternal biofluids and in-utero exposures correlating with autistic behaviours (not specifically diagnoses) there are some interesting details requiring follow-up. BDE-28 (one of the congeners of the PBDEs) crop up quite a bit in research terms as per the the findings from Daniels and colleagues [3] looking at human milk samples. As Daniels et al suggest however: "The consequences of exposure to PBDEs are unknown" so I don't think we can draw too many conclusions from that one; indeed as per one of my previous posts on PBDEs and autism, there are a few more studies which could be done. That also another PBDE congener (PBDE-85) was actually associated with lower SRS scores - "consistent with less autistic behaviors" - is another example of just how complicated this area actually is.

Although maternal phthalate metabolites were not seemingly connected to scored offspring autistic behaviours, I'm not quite ready to trash any connection in this area. The paper by Testa and colleagues [4] (open-access here) looking at "primary and secondary metabolites of DEHP [di-(2-ethylhexyl) phthalate] in children with ASD [autism spectrum disorder]" offers some preliminary evidence for a potential role for phthalates exposure and autism. Compounded by the data presented by Stein and colleagues [5] on "a decreased capacity for detoxification via glucuronidation of compounds in the DEHP metabolic pathway" in their cohort with autism, and there are still questions to be answered about this group of compounds. And just in case you are still furrowing your brow at this association, there is the data from Larsson and colleagues [6] (open-access here) to look at, bearing in mind the suggestion that "the use of soft PVC as flooring material may increase the human uptake of phthalates in infants" [7].

Pinpointing one or more single pollutants as being tied into the increasing numbers being diagnosed with autism is always going to be an uphill struggle. Not only do we have very little data on the multitude of 'chemicals' (remember the mis-use of that word) we're all exposed to as part of modern living (or indeed that our parents were exposed to) but examining the synergistic effects of various multiple exposures is going to be problematic at best. As per the Stein research, we've also got to consider how an individual processes or metabolises their pollutant load which must have a bearing on the physiological effect from any exposure. Thinking back also to the recent air pollution - autism work starting to ask about gene x environment interactions, there are multiple dimensions to this work which aren't necessarily going to be answered by just looking at what a person is exposed to.

Oh, and I'll be coming to the paper by Nishijo and colleagues [8] titled "2,3,7,8-Tetrachlorodibenzo-p-dioxin in breast milk increases autistic traits of 3-year-old children in Vietnam" at some point quite soon.

To close, Rock Lobster...

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[1] Rzhetsky A. et al. Environmental and state-level regulatory factors affect the incidence of autism and intellectual disability. PLoS Comput Biol. 2014 Mar 13;10(3):e1003518.

[2] Braun JM. et al. Gestational Exposure to Endocrine-Disrupting Chemicals and Reciprocal Social, Repetitive, and Stereotypic Behaviors in 4- and 5-Year-Old Children: The HOME Study. Environ Health Perspect. 2014 Mar 12.

[3] Norris M. & Lecavalier L. Screening accuracy of Level 2 autism spectrum disorder rating scales. A review of selected instruments. Autism. 2010 Jul;14(4):263-84.

[4] Testa C. et al. Di-(2-ethylhexyl) phthalate and autism spectrum disorders. ASN Neuro. 2012 May 30;4(4):223-9.

[5] Stein TP. et al. Autism and phthalate metabolite glucuronidation. J Autism Dev Disord. 2013 Nov;43(11):2677-85.

[6] Larsson M. et al. Associations between indoor environmental factors and parental-reported autistic spectrum disorders in children 6-8 years of age. Neurotoxicology. 2009 Sep;30(5):822-31.

[7] Carlstedt F. et al. PVC flooring is related to human uptake of phthalates in infants. Indoor Air. 2013 Feb;23(1):32-9.

[8] Nishijo M. et al. 2,3,7,8-Tetrachlorodibenzo-p-dioxin in breast milk increases autistic traits of 3-year-old children in Vietnam. Mol Psychiatry. 2014 Mar 18.

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Braun JM, Kalkbrenner AE, Just AC, Yolton K, Calafat AM, Sjödin A, Hauser R, Webster GM, Chen A, & Lanphear BP (2014). Gestational Exposure to Endocrine-Disrupting Chemicals and Reciprocal Social, Repetitive, and Stereotypic Behaviors in 4- and 5-Year-Old Children: The HOME Study. Environmental health perspectives PMID: 24622245

Urinary phthalate metabolites and autism

I don't know if it is just me but investigations on the potential role of certain environmental factors in relation to at least some cases of autism spectrum conditions seem to be coming in thick and fast recently. I speak for example about the paper suggesting a role for high-fructose corn syrup (HFCS) and other factors; alongside other interesting papers published at the same time as the 'top 10 environmental factors' editorial discussed recently.

Appreciating that there are some gaps in the whole environmental area of investigation with regards to cause-and-effect, mixed in with the question of what might do what and to who, there is some interesting reading in amongst the various studies. This includes discussions around the concept of 'risk' and how autism research seems to be (partially) reinventing itself into a slightly more complicated hypothesis where environment and genes (variably) might play a role in aetiology rather than just genes, genes, genes. I get to say that word again.. 'epigenetics'.

Add then this paper by Testa and colleagues* (full-text) to the list of environmental question marks which was very quietly published recently looking at a possible connection between phthalate excretion and autism.

Phthalates (assuming I have spelled the word correctly) according to the US EPA, are a class of compounds called plasticisers as a result of their ability to make things more 'plastic' in terms of properties such as flexibility and durability. Whilst moves are underway to remove / limit phthalates in the industrial chain, their use is pretty widespread in products as diverse as cosmetics to food packaging, flooring to detergents.

As mentioned, there are moves to reduce the use of phthalates in the production of certain types of product as a result of growing evidence associating their exposure and unwelcome effects such as their endocrine disrupting potential. Just before you start looking around at what products you are surrounded by which might contain phthalates, the answer is probably quite a few; and in terms of exposure to these compounds, generally speaking we all have some of them circulating (or at least traces of them as per the analysis of biofluids like urine). Modern man and woman it seems are probably not going to grace the Visitors table as per an older post.

To the Testa paper:

• Based on the assumption that phthalates might be able to affect aspects of child development (particularly boys), the authors looked at urinary levels of primary and secondary metabolites of di(2-ethylhexyl)phthalate (DEHP) in a small-ish group of Italian children with autism (n=48) compared with an asymptomatic age- and gender-matched group (n=45). I say gender-matched but the authors use the word 'sex comparable' given that the autism group seemed to have a greater ratio of boys than the control group.
• The autism group were all DSM-IV diagnosed with autism and importantly all had ADOS and/or other data as some kind of confirmation of autism.
• First morning, spot urine samples were analysed by a favourite method, mass spectrometry, tandem mass spectrometry, preceded by quite a complicated extraction procedure based on the use of solid-phase extraction (SPE) to clean the sample up. I'm not going to go through the total list of metabolites they looked at but rather pick out the results that were significantly different.
• Results: levels of secondary metabolites, 5-OH-MEHP, 5-oxo-MEHP and MEHP were significantly increased in the autism group as a whole compared with controls. That being said, these metabolites were not detected in every sample from children in the autism group (52% & 46% & 79% respectively) bearing in mind the sensitivity of the assay used and lower limits of detection reported. Further comparison with a separate group of people diagnosed with Rett syndrome (RS) (n=10) - no age range described - suggested that the autism group excreted more secondary metabolites of DEHP (MEHP) than RS participants.
• One secondary metabolite, 5-oxo-MEHP, was suggested to show over 90% specificity when it came to identifying participants with autism.

As far as I can see, the authors are correct in their statement that this is the first time that phthalates have been directly looked for in the biological fluids of people with autism compared to controls. The research literature contains a couple of more speculative papers looking for example at in-door environment and autism (with accompanying reporting here) or reviewing the very limited available evidence on lots of endocrine disrupting compounds in relation to autism but not much else. There is other work in areas such as ADHD for example, again suggesting the possibility of some connection between reported ADHD symptoms and urinary phthalate content. Even executive functions get a look-in with regards to prenatal exposure. But that's your lot.

Where to go from here?

Well, we have to be slightly cautious in making too many assumptions from this current data. Yes, children with autism were better excretors of phthalates and yes, better to the degree that certain metabolites might even serve some identifying feature in comparison to control samples. But, and it is quite a large but(!), this paper does not provide a cause-and-effect role for phthalates in cases of autism. So for example, only urine was looked at and only individual spot samples at that. I could be a little bit pedantic and start asking whether circulating plasma levels of phthalates were any different or asking about levels in other tissues?

The authors did undertake some routine correlational analysis between phthalate metabolite levels and things like CARS scores, finding a positive association between MEHP and CARS scores such that increasing levels of one was correlated to increasing levels of the other. I do find this 'severity' relationship to be interesting although I perhaps would have liked to have seen confirmatory analysis from other tools including the ADOS.

Despite my queries about the results, what this study does offer is another potential target area to include when looking at environmental pollutants and any relationship to autism - add it to the list. Without wishing to seem like I'm too obsessed, I do wonder also about a gut bacterial connection as per other data, very preliminary data, on pesticides and bacteria. Sideways thinking is always advised.

To finish, a spot of New Order. Indeed since I am reminiscing about the Manchester scene, how about some Stone Roses too.

* Testa C. et al. Di(2-ethylhexyl)phthalate and autism spectrum disorders. ASN Neuro. April 2012.
DOI: 10.1042/AN20120015