Facial phenotypes of autism continued

Quite a few years back, I talked on this blog about research suggesting that the structure of faces might be an important research area when it comes to at least some autism (see here). As per that blogging occasion, I cautioned that one has to be a little careful in this area of science so as not to make too many sweeping generalisations from any findings, but that there may some important *associations* to be noted. Such associations may, for example, be particularly important in the context of various genetic syndromes manifesting specific facial features as well as expressing autism or autistic traits.

In recent times I've noticed a couple of articles emerging on this topic from the University of Western Australia under the guidance of Prof. Andrew Whitehouse. Research from Prof. Whitehouse has been discussed before on this blog (see here and see here for examples) and I have to say I'm quite a fan of some of his, and his teams, efforts.

This time around I'm blogging about the papers by Maryam Boutrus and colleagues [1] and Diana Weiting Tan and colleagues [2] who talked about applying "a hypothesis-driven approach" to the study of facial phenotypes when it comes to autism and then offered up one such hypothesis: "comparing the facial masculinity/femininity of boys and girls with ASD [autism spectrum disorder] to that of typically-developing children" in the context of elevated prenatal testosterone exposure being *associated* with autism (albeit probably not universally so). I yet again hasten to add that the term 'typically-developing' is the authors' choice not mine.

The Tan paper talks about 3D imaging of faces from "a normative sample of 48 boys and 53 girls" which built up a kind of composite image to differentiate boy faces from girl faces. Some 21 facial landmarks were initially used but eventually this was boiled down into 11 selected facial features: "three linear distances (alar-base width, nose height and upper lip height) and eight geodesic distances (outer-canthal width, forehead width, forehead height, right upper cheek height, nasal tip protrusion, nose height, upper lip height, and nasal bridge length)."

The second part of the study then applied these 11 parameters deemed useful for differentiating "the faces of typically-developing boys and girls" and used them to compare faces of children diagnosed with ASD compared with those with no diagnosis - "54 autistic and 54 control boys... and also for 20 autistic and 60 control girls." This involved the application of "an overall facial masculinity/femininity index" providing a scale scoring between extreme masculinity and extreme femininity.

Results: "autistic boys had significantly lower gender scores for their faces (i.e., more masculine) when compared to the control boys." Also: "For girls, ANOVA showed that gender scores were significantly lower (i.e., less feminine) for the ASD group compared to the control group." These findings were accompanied by some equally interesting observations in relation to how the gender context of facial features seemed to tie into aspects of autistic presentation. Specifically: "increased facial masculinity in the ASD group correlated with more social-communication difficulties based on the Social Affect score derived from the Autism Diagnostic Observation Scale-Generic (ADOS-G)." No such relationship was reported in relation to the other part of the diagnostic dyad currently used to diagnose autism - Restricted and Repetitive Behaviours (RRBs).

Obviously, more needs to be done in this area. The sample size for both parts of the study reported by Tan et al is preliminary at best and replication is most definitely the name of the research game. I have to say that outside of the sexing/gendering of faces with autism in mind, I'm particularly interested in how facial masculinity might be linked to greater issues with social affect. One wonders whether this might for example, extend to the more general population and indeed, sub-clinical signs and symptoms of autism such as that noted in the broader autism phenotype (BAP)?

To close, although unrelated to the material covered today, facial phenotyping is starting to ask some other interesting questions too...

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[1] Boutrus M. et al. Investigating facial phenotype in autism spectrum conditions: The importance of a hypothesis driven approach. Autism Res. 2017 Aug 17.

[2] Tan DW. et al. Hypermasculinised facial morphology in boys and girls with Autism Spectrum Disorder and its association with symptomatology. Sci Rep. 2017; 7: 9348.

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On the under-studied populations within the autism spectrum

I don't typically post on a Sunday, but given that today - 2nd April - is World Autism Awareness Day I've decided to make an exception. The theme of today's post is based around the notion that the autism spectrum is truly wide and heterogeneous, and although this is fairly universally acknowledged, the current peer-reviewed research literature on autism is perhaps not yet so accepting. So...

Consistent with the idea that scientific research seems to go through cycles of themes/interests, the commentary paper by Bhismadev Chakrabarti [1] (open-access) continues an important theme talking about the representativeness of autism research (see here). Specifically how: "Research on the autistic phenotype has focused mostly on higher functioning individuals on the spectrum, neglecting those on the lower end."

OK, first things first. The idea of 'functioning' in relation to the autism spectrum is something that some people (including myself) find a little problematic. Yes, I know what it is trying to describe in terms of ability levels, adaptive skills and the level of support seemingly required as examples. But like many things when it comes to the autism spectrum, the [sweeping] generalisation that high-functioning autism automatically means 'can function' autism and low-functioning autism conversely means 'can't' doesn't really do justice to the complexity underneath such categorisations. I say all that acknowledging that no simple, viable alternative currently exists to replace 'functioning' at the present time.

Chakrabarti takes the reader through the issues of research representativeness based on the findings reported by Jack & Pelphrey [2] and their research review of neuroimaging studies in relation to autism. They concluded that: "There is a paucity of neuroimaging research on ASD [autism spectrum disorder] + ID [intellectual disability], ASD + MV [minimally verbal], and ASD + R [developmental regression], and what findings do exist are often contradictory, or so sparse as to be ungeneralizable."

I'm gonna pull out a couple of key points raised by Chakrabarti that are worthy of lots more research and clinical inspection.

First: "Should we be thinking of these different populations (MV, R and ID) as distinct subgroups within ASD?" Set within the context of 'the plural autisms' (see here) and how autism as a singular label seems to have very little usefulness as a research starting point (see here), it strikes me that Chakrabarti's suggestion of 'phenotypic dimensions' is quite a good one. The fact that developmental regression gets a look-in is also quite important (see here and see here) (no, not every single case of autism was present before or at birth/early infancy).

Second, on the question of 'neuroimaging phenotypes' akin to some of the parameters set out in the RDoC alternative to DSM (see here) I think we have to wait and see. From what we already know about neuroimaging results when it comes to the autism spectrum as a whole, there is no one 'brain area' seemingly linked to all diagnoses of autism (see here) as things currently stand, bearing in mind the limitations of the technology currently used. I don't doubt however that specific groups of people on the autism spectrum might be more likely to show definite collective brain pathology (see here) particularly where certain over-represented comorbidity might complete the clinical picture. The current state of findings in this area also has implications for the use of problematic terms such as 'neurotypical' to denote not-autism (I personally have no idea what neurotypical looks like on a brain scan nor in terms of development, behaviour, maturation or comorbidity).

I'm hoping that papers/commentaries such as the one from Chakrabarti are a call to action when it comes to making autism research 'work' for everyone on the autism spectrum. That and acknowledging that the existing - skewed - research base might be missing some important details when it comes to the very wide and very heterogeneous autisms...

To close and without getting too political, I want to link to a piece that was published in the Huffington Post this week (see here) discussing the idea of 'celebrating' world autism awareness day. It's something that I've seen quite a lot of these past years. Reiterating that the autism spectrum is indeed wide and heterogeneous, I found the article to very moving particularly the writer's notion that: "What I will do is celebrate my son for who he is... But I won’t celebrate the struggles we call autism." Appreciating that autism as a label is 'identity' for some on the spectrum, such sentiments reaffirm the requirement to ensure that all voices on the autism spectrum are heard, and that 'celebration' is reserved for people and their achievements, not their labels...

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[1] Chakrabarti B. Commentary: Critical considerations for studying low-functioning autism. J Child Psychol Psychiatry. 2017 Apr;58(4):436-438.

[2] Jack A. & A Pelphrey K. Annual Research Review: Understudied populations within the autism spectrum - current trends and future directions in neuroimaging research. J Child Psychol Psychiatry. 2017 Apr;58(4):411-435.

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Chakrabarti B (2017). Commentary: Critical considerations for studying low-functioning autism. Journal of child psychology and psychiatry, and allied disciplines, 58 (4), 436-438 PMID: 28346760

Subgroups in autism (without intellectual disability)

"Children with ASD [autism spectrum disorder] without ID [intellectual disability] could be differentiated into Moderate and Severe Social Impairment subgroups when core ASD symptoms were more closely examined."

So said the findings reported by Felicity Klopper and colleagues [1] looking at an important part of the autism research scene related to the 'plurality' of the term autism and the seemingly vast range of presentations included under the label. Reliant on data obtained from "the ‘gold standard’ ASD diagnostic instruments" (including the ADOS and ADI), researchers looked at the "presence of phenotypic subgroups" in their cohort.

As per the opening sentence to this post, there were some differences to be seen in the cohort, and in particular, how social interaction issues might be a key part of any differentiation. The authors talk about how social interaction issue differences seemed to tie into other core behavioural features such as communication and the presence of restricted/repetitive behaviours. They concluded: "both categorical and dimensional approaches may be useful in classifying ASD, with neither alone being adequate."

It is not necessarily new news that the label of autism is good for diagnosis but seemingly says little about the range of presentation included under the heading (see here for example). Indeed, in these days of ESSENCE I might forward the view that even the label autism might be part of a wider heterogeneous presentation (see here) and one should further expand those subgroup notions at the label as well as symptom level. The focus on overt behaviour (as assessed by those gold-standard instruments) in the Klopper study is but one part of looking at such 'heterogeneity' (see here for example) as the authors argue that: "The dissociated profiles of ASD features could represent different underlying neurobiological mechanisms for each subgroup." At least one of the authors on the Klopper paper probably, more than most, realises that fact (see here).

There are other key areas to this focus on the presentation of autism that also need to be factored in: sex differences and comorbidity profiles. Specifically, the growing realisation that girls and boys on the autism spectrum probably show subtle differences in presentation (see here) and, minus any sweeping generalisations, should be considered in future studies in this area. Oh, and keep in mind that those diagnosed with autism with an intellectual disability (ID) could also be 'sub-grouped' according to symptom presentation too with similar caveats. The question is: how many sub-groups of autism will we eventually end up with?

Music, and because Spring has Sprung... In Bloom.

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[1] Klopper F. et al. A cluster analysis exploration of autism spectrum disorder subgroups in children without intellectual disability. Research in Autism Spectrum Disorders. 2017; 36: 66-78.

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Felicity Klopper, Renee Testa, Christos Pantelis, & Efstratios Skafidas (2017). A cluster analysis exploration of autism spectrum disorder subgroups in children without intellectual disability Research in Autism Spectrum Disorders : 10.1016/j.rasd.2017.01.006

On biological markers and "subphenotypes" of autism

I don't want to keep you today. Just long enough to draw your attention to the paper by Jones and colleagues [1] regarding "ongoing efforts toward identification of early biological markers specific to subphenotypes of ASD [autism spectrum disorder]."

The potential biomarkers in question this time around were the cytokines/chemokines - those various signalling molecules that seem to have more than a few connections to important processes like inflammation - and how their profile ("mid-gestational serum profiles") might show some interesting links to different phenotypes of autism. Similar sentiments are already present in the peer-reviewed literature (see here) I might add.

Comparing 22 different cytokines and chemokines from mothers' samples across various groupings (ASD, developmental delay, 'not-autism' controls) and importantly including a distinction between autism diagnosed with intellectual (learning) disability (ASD+ID) and autism on its own (ASD-noID), researchers reported some interesting trends. Not least was the suggestion of an "immunologic distinction between mothers of children with ASD+ID from mothers of children with ASD-noID" including elevations in something called interleukin-6 (IL-6) (see here).

Bearing in mind a continued focus on immune function in relation to [some] autism (see here) taking into account the very wide heterogeneity present across the spectrum (autisms people, autisms), the requirement for yet further study in this area is immense. With all that's starting to be known about the various presentations of immune function in autism (see here and see here for example) and beyond (see here), I find it surprising that so little research has so far been translated from 'bench-to-bedside' in terms of screening and intervention when immune-related issues are found. Yes, dogma about what autism(s) is and isn't still pervades the scientific literature and beyond (in many areas!), but surely routine immune panel screening could be looked at being rolled out for example?

Just one more thing to add about the Jones paper. The eagle-eyed among you might already have noted the involvement of the MIND Institute in the authorship group bearing in mind their long (long) history in autism research (see here). But did you also pick up a certain Robert Yolken from the 'Stanley Division of Developmental Neurovirology' as also being present? Yes, one and the same researcher, who along with various other members of the this facility, are making some real research waves when it comes to the interplay between behaviour, immune function and infection (see here and see here for examples).

Methinks this could be the start of a beautiful (and hopefully rewarding) research relationship [2]...

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[1] Jones KL. et al. Autism with intellectual disability is associated with increased levels of maternal cytokines and chemokines during gestation. Molecular Psychiatry. 2016. May 24.

[2] Grether JK. et al. Prenatal and Newborn Immunoglobulin Levels from Mother-Child Pairs and Risk of Autism Spectrum Disorders. Front Neurosci. 2016 May 18;10:218.

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Jones KL, Croen LA, Yoshida CK, Heuer L, Hansen R, Zerbo O, DeLorenze GN, Kharrazi M, Yolken R, Ashwood P, & Van de Water J (2016). Autism with intellectual disability is associated with increased levels of maternal cytokines and chemokines during gestation. Molecular psychiatry PMID: 27217154

Pregnancy paracetamol use and the 'hyperactive behavioral phenotype' of autism

"Prenatal use of acetaminophen was associated with an increased risk of ASD [autism spectrum disorder] accompanied by hyperkinetic symptoms..., but not with other ASD cases."

I was rather interested to read that conclusion presented in the study by Zeyan Liew and colleagues [1] talking about how acetaminophen (or paracetamol as it is known here in Blighty) use during pregnancy might have some rather important connections to offspring outcomes specifically with autism and hyperkinetic symptoms in mind.

Interested not only because I've discussed other research talking about how we might need to be a little more cautious about how OTC pain relief such as paracetamol is used in certain circumstances with potential offspring outcomes in mind (see here and see here) but also because it makes reference to a quite specific effect to the "hyperactive behavioral phenotype" of autism. That hyperkinetic behavioural phenotype by the way, plugs into a growing body of peer-reviewed evidence talking about the over-representation of ADHD (attention-deficit hyperactivity disorder) symptoms in relation to [some] autism (see here).

Liew et al "followed 64,322 children and mothers enrolled in the Danish National Birth Cohort (DNBC; 1996–2002) for average 12.7 years to investigate whether acetaminophen use in pregnancy is associated with increased risk of ASD in the offspring." Details about paracetamol use during pregnancy were "collected prospectively from three computer-assisted telephone interviews" and offspring outcomes with labels like autism were compared. This follows a similar methodological template from some of this research team (see here).

Some 1.6% of children included in the analysis were eventually diagnosed with as ASD. Roughly a third of that 1.6% of children were also diagnosed with hyperkinetic disorders (which seems to tally with other independent data analysis [2]). Perhaps unsurprisingly, paracetamol use during pregnancy was pretty widespread in the cohort (~50%) but, despite this, authors detailed something of a potentially important relationship between pregnancy paracetamol use and offspring "ASD accompanied by hyperkinetic symptoms." That there also appeared to be a relationship between length of use of paracetamol and risk of offspring autism with hyperkinetic symptoms, seems to strengthen details of a possible association.

Accepting that this was a study plotting pregnancy medication use and offspring outcomes and hence findings need to be treated with a degree of caution (as with other studies on other medicines), set within the other research talking about paracetamol use during pregnancy I'm coming around the idea that more investigation is required in this area [3]. I could, once again, start talking about how pregnancy paracetamol use might link into childhood asthma symptoms [4] (albeit with some cautions attached [5]) and what that might mean for the behavioural 'associations' noted in some cases of asthma (see here) as a possible angle for future research. I would however temper such musings within the context of autism (see here); although the preliminary suggestion that prenatal exposure to certain drugs used to treat asthma might also affect risk of offspring autism is also interesting [6].

I do like that Liew and colleagues talked about the idea of a specific behavioural phenotype potentially linked to pregnancy paracetamol exposure as providing a guide for more targeted investigation and perhaps to some degree bypassing the need for the sweeping generalisations about 'all autism' (we've had enough of those down the years). I believe there is quite a bit of research traction in this area with the notion of the autisms (see here) in mind.

Music: Elvis Presley - If I Can Dream.

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[1] Liew Z. et al. Maternal use of acetaminophen during pregnancy and risk of autism spectrum disorders in childhood. Autism Research. 2015. Dec 21.

[2] Berenguer-Forner C. et al. Comorbidity of autism spectrum disorder and attention deficit with hyperactivity. A review study. Rev Neurol. 2015 Feb 25;60 Suppl 1:S37-43.

[3] de Fays L. et al. Use of paracetamol during pregnancy and child neurological development. Dev Med Child Neurol. 2015 Aug;57(8):718-24.

[4] Eyers S. et al. Paracetamol in pregnancy and the risk of wheezing in offspring: a systematic review and meta-analysis. Clin Exp Allergy. 2011 Apr;41(4):482-9.

[5] Cheelo M. et al. Paracetamol exposure in pregnancy and early childhood and development of childhood asthma: a systematic review and meta-analysis. Arch Dis Child. 2015 Jan;100(1):81-9.

[6] Gidaya NB. et al. In utero Exposure to β-2-Adrenergic Receptor Agonist Drugs and Risk for Autism Spectrum Disorders. Pediatrics. 2016. 6 Jan.

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Liew Z, Ritz B, Virk J, & Olsen J (2015). Maternal use of acetaminophen during pregnancy and risk of autism spectrum disorders in childhood: A Danish national birth cohort study. Autism research : official journal of the International Society for Autism Research PMID: 26688372

Immune endophenotypes in paediatric autism

Today I'm serving up the paper by Milo Careaga and colleagues [1] for your blogging delight, who concluded that: "Children with ASD [autism spectrum disorder] may be phenotypically characterized based upon their immune profile." Further that there may be: "several possible immune subphenotypes within the ASD population that correlate with more severe behavioral impairments."

With many thanks to Natasa for the paper, participants - 50 boys with a median age of 3.2 years diagnosed with an ASD and enrolled "through the Autism Phenome Project (APP) study" and who were free of any "major immune modifying medications" - provided a blood sample. A similar process was employed for a smaller group of typically developing (asymptomatic) control group (n=16). Said blood sample went through various processes to harvest peripheral blood mononuclear cells (PBMC) which were then 'stimulated' to provoke an immune reaction via "either lipopolysaccharide (LPS) or phytohaemagglutinin (PHA)." Various cytokines were then assayed for in the stimulated PBMC and results analysed according to immune responses and behavioural outcomes.

Results: as per the opening paragraph, there was potentially something to see in the findings added to a more general role for cytokines in relation to autism [2]. Those children with ASD who presented with a more 'pro-inflammatory' cytokine profile in their stimulated blood results "showed more impaired developmental and behavioral scores, as well as increased problems with sleep and aggression." That pro-inflammatory cluster by the way (n=22) tended to show significantly increased production of cytokines such as IL-6 for example, than those children with autism (n=28) "who displayed a less robust response to LPS." Ergo, perhaps more to see and certainly more investigations required in these days of plural autisms.

I was taken by one particular sentence included in the conclusion of the Careaga paper: "Although immune abnormalities were first described in ASD over forty years ago, no consensus has been reached as to what constitutes clinically significant immune dysfunction in ASD." As per quite a bit of autism research, sweeping generalisations about this, that or t'other 'causing' autism or being part and parcel of autism have been a big contributor to the noticeable lack of progress on knowledge about autism and where required and wanted, what can be done to ameliorate the more disabling aspects including that related to comorbidity (see here). Realisation that 'autism' is probably better described as providing an umbrella term for various different conditions on a genetic and molecular level is making some headway these days (see here) including that linked to immune function (see here for example). This might have important implications for intervention (see here) as per other recent results that I'll be musing over soon.

As part of a broader realisation that immune function and psychiatry probably show a lot more connections than many people first realised (see here), I think we are seeing a shift in knowledge here. As per the Careaga results, the idea that there may be distinct clusters within the presentation of autism linked to immune function, opens up a whole new world of more 'targeted' inspection and intervention which, added to other similar phenotype work (see here), is probably an important direction for autism research...

Music: Duran Duran - Pressure Off feat. Janelle Monáe and Nile Rodgers.

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[1] Careaga M. et al. Immune endophenotypes in children with autism spectrum disorder. Biological Psychiatry. 2015. 10 Sept.

[2] Krakowiak P. et al. Neonatal Cytokine Profiles Associated with Autism Spectrum Disorder. Biol Psychiatry. 2015 Aug 14. pii: S0006-3223(15)00655-1.

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Milo Careaga, Sally Rogers, Robin L. Hansen, David G. Amaral, Judy Van de Water, & Paul Ashwood (2015). Immune endophenotypes in children with autism spectrum disorder Biological Psychiatry : 10.1016/j.biopsych.2015.08.036

New name: Systemic Exertion Intolerance Disease?

The name is: Systemic Exertion Intolerance Disease (SEID) (which as one of my very observant Facebook friends pointed out is DIES backwards).

A very quick post to direct you to the public release of the findings from the US Institute of Medicine (IoM) looking at the name and current criteria used to diagnose Chronic Fatigue Syndrome / Myalgic Encephalomyelitis (CFS/ME) (see here). The proposed diagnostic criteria for CFS/ME, sorry SEID can be viewed here.

Some of the background to these findings can be seen here and some of the media about the new IoM recommendations can be seen here and here.

Many with either personal or professional experience of CFS/ME will know all about the issue of diagnosis and CFS/ME, and outside of the struggle to recognise that this is a very real condition which can severely impact on a person's life, how confusing the diagnostic criteria can be. This has also had various implications on both research and practice [1].

The IoM was charged with looking at the existing ways and means that CFS/ME was defined and "recommend clinical diagnostic criteria for the disorder to address the needs of health providers, patients, and their caregivers" (see here). To that end, their guidance now reflects these efforts although at the current time I'm having some difficulty in locating how they tackled the idea of "distinguish[ing] between disease subgroups" which I think should have represented quite a big leap forward in these days of plural labels and heterogeneity in symptoms and comorbidity (see here). At this point I might refer you to a paper I wrote that touched upon this a few years back [2]. I am however happy to see that pediatric ME/CFS or SEID has been mentioned in their various guidance in light of the growing realisation that the condition can manifest in children and young people too (see here).

Obviously it's going to take time for these findings to find their way from guidance to practice and beyond so I don't think massive changes will be immediately forthcoming. One would hope that alongside the growing interest in the biology/genetics of CFS/ME (acknowledging how the condition can also impact on psychology too) that moves towards the development and testing of biological therapeutics in particular, might be accelerated (see here).

To close: Breaking Bad and Say My Name... 'sorta Greg'.

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[1] Morris G. & Maes M. Case definitions and diagnostic criteria for Myalgic Encephalomyelitis and Chronic fatigue Syndrome: from clinical-consensus to evidence-based case definitions. Neuro Endocrinol Lett. 2013;34(3):185-99.

[2] Whiteley P. et al. Correlates of Overlapping Fatigue Syndromes. Journal of Nutritional and Environmental Medicine. 2004; 14: 247-259.

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Morris G, & Maes M (2013). Case definitions and diagnostic criteria for Myalgic Encephalomyelitis and Chronic fatigue Syndrome: from clinical-consensus to evidence-based case definitions. Neuro endocrinology letters, 34 (3), 185-99 PMID: 23685416

Maternal smoking during pregnancy and offspring autism: no measurable association but...

Oh. Yes sir. How doth the little
bumblebee improve each...

"We found no evidence to support a measurable association between maternal prenatal smoking and ASD [autism spectrum disorder] in offspring."

That was the conclusion reached in the meta-analysis published by Brittany Rosen and colleagues [1] looking at the collected peer-review literature examining any correlation between maternal tobacco smoking during pregnancy and risk of offspring receipt of a diagnosis of autism or ASD. Based on a review of 15 studies in this area, researchers concluded that there was very little to see when it came to any association: "summary OR [odds ratio] 1.02, 95 % CI [confidence intervals] 0.93–1.12".

I think most people nowadays have probably seen or heard of the messages about the potential dangers of smoking during pregnancy and the benefits of abstaining/quitting to both mother and unborn child. That's not to say however that every mum-to-be has understood the risks as per the findings from Cnattingius [2]. Indeed without being judgemental, even at rare visits to my own local hospital, I have seen heavily pregnant women puffing away outside the maternity unit and can't help wondering why, particularly when so many alternatives or quitting aids (with caveats) are available these days.

Insofar as the various investigations into smoking with offspring autism in mind, there is quite a bank of research in this area. That's not to say that the evidence is all one-way when it comes to smoking during pregnancy and offspring risk of autism as per studies like the one from Phuong Lien Tran and colleagues [3] (open-access) who concluded that smoking during the whole of pregnancy might have a modest impact on autism risk, at least in Finland. The findings reported by Visser and colleagues [4] on smoking during pregnancy appearing to "contribute more to broadly defined (PDD-NOS) than to narrowly defined ASD (AD)" are also worthwhile including in these days of phenotypes and plural autisms. Indeed, the report from Amy Kalkbrenner and colleagues [5] (open-access) continues the theme: "The possibility of an association with a higher-functioning ASD subgroup was suggested, and warrants further study."

On the basis of these studies and others [6], I'd be minded to suggest that whilst the Rosen findings are reassuring for the more general concept of risk (i.e. autism overall) the message about quitting smoking before and during pregnancy as potentially affecting offspring autism risk, is not yet settled when it comes to specific types of autism or specific places on the autism spectrum being implicated. I say this not to add to any further burden about this or that 'causing autism' as per examinations on things like maternal infection during pregnancy (see here) or C-sections (see here) or maternal diabetes (see here) for example, but rather using science as a potential informer in raising awareness of this possible outcome. Indeed, if one is to assume that the [preliminary] evidence on air pollution and genetics mixing might also impact on autism risk (see here), the question of whether direct inhalation of several thousand pollutants can impact on foetal outcomes becomes rather more compelling including whether risk of certain comorbidity appearing alongside autism might also be influenced [7].

Oh, and father's tobacco habits might also be important [8]...

Music, and something quiet from Henry Rollins (not)...

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[1] Rosen BN. et al. Maternal Smoking and Autism Spectrum Disorder: A Meta-analysis. Journal of Autism and Developmental Disorders. 2014. November 29.

[2] Cnattingius S. The epidemiology of smoking during pregnancy: smoking prevalence, maternal characteristics, and pregnancy outcomes. Nicotine Tob Res. 2004 Apr;6 Suppl 2:S125-40.

[3] Tran PL. et al. Smoking during pregnancy and risk of autism spectrum disorder in a Finnish National Birth Cohort. Paediatr Perinat Epidemiol. 2013 May;27(3):266-74.

[4] Visser JC. et al. Narrowly versus broadly defined autism spectrum disorders: differences in pre- and perinatal risk factors. J Autism Dev Disord. 2013 Jul;43(7):1505-16.

[5] Kalkbrenner AE. et al. Maternal smoking during pregnancy and the prevalence of autism spectrum disorders, using data from the autism and developmental disabilities monitoring network. Environ Health Perspect. 2012 Jul;120(7):1042-8.

[6] Habek D. & Kovačević M. Adverse pregnancy outcomes and long-term morbidity after early fetal hypokinesia in maternal smoking pregnancies. Arch Gynecol Obstet. 2011 Mar;283(3):491-5.

[7] Kovess V. et al. Maternal smoking and offspring inattention and hyperactivity: results from a cross-national European survey. Eur Child Adolesc Psychiatry. 2014 Nov 21.

[8] Laubenthal J. et al. Cigarette smoke-induced transgenerational alterations in genome stability in cord blood of human F1 offspring. FASEB J. 2012 Oct;26(10):3946-56.

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Rosen BN, Lee BK, Lee NL, Yang Y, & Burstyn I (2014). Maternal Smoking and Autism Spectrum Disorder: A Meta-analysis. Journal of autism and developmental disorders PMID: 25432101

Evidence for an autoimmune aetiology of epilepsy?

With a title like that, I was bound to post about the paper by Mei-Sing Ong and colleagues [1] and their suggestion that: "Epilepsy and autoimmune disease frequently co-occur; patients with either condition should undergo surveillance for the other".

Capo di Noli @ Wikipedia 

United under the umbrella that is autism spectrum comorbidity (even possible phenotypes), I've talked quite a bit on this blog about epilepsy / seizure disorders (see here) and autoimmune conditions (see here), and how coexistence alongside the presentation of autism may be a potential route towards some overlapping or shared genetic or biochemical pathways being involved. It is indeed a coincidence that one of the authors on the Ong paper - Isaac Kohane - is one and the same with some autism [comorbidity] research in mind.

The Ong paper details the results of a a survey of health insurance claims in the US where authors "examined the relationship between epilepsy and 12 autoimmune diseases: type 1 diabetes mellitus, psoriasis, rheumatoid arthritis, Graves disease, Hashimoto thyroiditis, Crohn disease, ulcerative colitis, systemic lupus erythematosus, antiphospholipid syndrome, Sjögren syndrome, myasthenia gravis, and celiac disease". They reported that the risk of epilepsy was higher amongst those people with an autoimmune condition - all 12 of the autoimmune conditions looked at - and particularly when it came to risk for children. More chatter about this study can be found here.

Looking through many of those autoimmune conditions examined by Ong, the first thing that struck me was that many of those conditions have been talked about in research circles in the context of autism. Bowel conditions such as ulcerative colitis and Crohn's disease have certainly seen their fair share of discussion (see here) alongside the possibility of other bowel disease presentation in cases (see here). Coeliac (celiac) disease and autism... well, don't get me started. Antiphospholipid syndrome, or at least anti-phospholipid antibodies, have also been talked about in the context of autism too (see here). I could go on and on and on; reiterating that a diagnosis of autism is seemingly protective of nothing when it comes to other health issues, and quite a few of those health issues tend to fall into the autoimmune domain.

The next thing that struck me about that list of autoimmune conditions and indeed, how they might link into something like epilepsy, was the issue of food, and in particular gluten. Now, I'm not trying to make any sweeping generalisations here or anything like that, but outside of the classical relationship between coeliac disease and gluten, there is quite a bit of emerging evidence that gluten may be implicated in so much more including some overlap with autism. Don't believe me? Well, take type 1 diabetes as one example and papers like the one by Marietta and colleagues [2] on gluten modulating the incidence of type 1 diabetes (at least in mice). The paper by Sildorf and colleagues [3] even went as far as reporting on how a gluten-free diet was associated with clinical remission without insulin therapy for one boy with type 1 diabetes, bearing in mind no clinical or medical advice is given or intended from me on this issue. At this point I might also drop in a note about the various research being done on epilepsy, some epilepsy, and the use of a ketogenic diet (high fat, low carbohydrates which I assume impacts on gluten intake) as potentially being relevant here too.

Alessio Fasano, who has been mentioned on this blog before, summarised the possibility of a connection between gluten (gliadin) and type 1 diabetes quite nicely in one of his papers [4]. Indeed, following on from that paper and the notion that "loss of intestinal barrier function is necessary to develop autoimmunity", the question emerges: does the association between autoimmune diseases and epilepsy also suggest that gut permeability (a.k.a leaky gut) might be a feature of some cases of epilepsy? I don't want to get too bogged down with answering that question aside from reiterating that following on from the observations of Ong et al if it was eventually confirmed that there may be an autoimmune component to at least some cases of epilepsy, autism and particularly those cases of autism accompanied by a diagnosis of epilepsy, may very well represent one of the next stages in the evolution of this research topic.

Music to close. It's been a while since I've linked to an Elvis song, so here's Viva Las Vegas.

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[1] Ong MS. et al. Population-Level Evidence for an Autoimmune Etiology of Epilepsy. JAMA Neurol. 2014 Mar 31.

[2] Marietta EV. et al. Low incidence of spontaneous type 1 diabetes in non-obese diabetic mice raised on gluten-free diets is associated with changes in the intestinal microbiome. PLoS One. 2013 Nov 13;8(11):e78687.

[3] Sildorf SM. et al. Remission without insulin therapy on gluten-free diet in a 6-year old boy with type 1 diabetes mellitus. BMJ Case Rep. 2012 Jun 21;2012.

[4] Visser J. et al. Tight junctions, intestinal permeability, and autoimmunity: celiac disease and type 1 diabetes paradigms. Ann N Y Acad Sci. 2009 May;1165:195-205.

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Ong MS, Kohane IS, Cai T, Gorman MP, & Mandl KD (2014). Population-Level Evidence for an Autoimmune Etiology of Epilepsy. JAMA neurology PMID: 24687183

Regression and autism continued (yet again)

I noticed a few weeks back that the topic of regression - developmental regression - with autism in mind resurfaced during a few media reports (see here and here). That last report by Jennifer Richler, who knows more than most about the research around regression and autism [1] particularly took my attention, and the implication that perhaps we need a little more investigation into this area with a focus on developmental trajectories and the small details which might reflect onset of the regressive phenotype(s) and indeed, any pre-regressive presentation. Whether also a history of developmental regression might affect age at diagnosis might be another question to be asked in light of the study findings from Mishaal and colleagues [2].

Journeys... @ Wikipedia 

It is therefore timely that the paper by Kern and colleagues [3] (open-access here) also appeared in my inbox recently talking about regression in relation to autism based on parental reports. The quote: "82 children (60.7%) were reported to have R[egressed]" was interesting because this seemed to be quite a high figure bearing in mind the participant group size (N=135).

My previous post on the topic of regression and autism highlighted the meta-analysis paper by Barger and colleagues [4] which at most set parent-reported regression in autism at around 40% of cases, bearing in mind how one goes about defining regression. Indeed, the recent paper by Goin-Kochel and colleagues [5] similarly found the rate to be around the 40% mark (36.9% overall) in their cohort of over 2000 children. Allowing for the assumption that rates of regression in cases of autism are a static entity and not subject to changes over time, the disparity between the figures is interesting.

I'm not on this occasion going to go through the Kern paper in excruciating detail because it is open-access. It is not altogether dissimilar from the earlier Richler paper in that parental report forms the crux of the observations and factors such as gastrointestinal (GI) issues were also included in the research mix. Where the two results separate is on a few main points: (a) Richler and colleagues included a control group; Kern et al did not, (b) Richler talks about: "no evidence that onset of autistic symptoms or of regression was related to measles-mumps-rubella vaccination"; Kern suggests: "The majority of parents reported that the regression was preceded by or was associated with vaccinations (57.3%) or another medically related event (11.0%)", and (c) Richler talks about children "who lost skills" had "more gastrointestinal symptoms than children with ASD and no regression"; Kern by contrast reports: "no significant relationship between the children’s age, gender, race, severity, or GI symptoms, and their membership in the D[elayed], DR [delayed and later regressed], or R[egressed] groups". I don't want to head too far into the discussions surrounding point (b) but will draw your attention to the paper by Woo and colleagues [6], who looking at data from the US VAERS (Vaccine Adverse Event Reporting System) observed that "The proportion of VAERS cases of autism with regression was greater than that reported in population-based studies, based on the subset of VAERS cases with medical record confirmation". I wonder if this might have something to do with the high rate of regression reported by Kerns et al also?

I was particularly interested in the differing results reported as a consequence of the presence of GI factors comorbid to core autism presentation with regression in mind. I note in the paper by Valicenti-McDermott and colleagues [7], they reported that "children with language regression more frequently exhibited an abnormal stool pattern" assuming that one equates abnormal stool pattern[s] as being the same as GI symptoms; well, functional GI symptoms at least. The paper from Mady Hornig and colleagues [8] (open-access here) adds to this sentiment with their observation that "Autism with GI disturbances is associated with elevated rates of regression in language or other skills and may represent an endophenotype distinct from other ASD". All of this kinda puts a new slant on the recent papers confirming an over-representation of GI issues in cases of autism (see here).

I'm intrigued by the notion that combined regression and GI issues might be a distinguishing endophenotype (sub-group) in the growing plurality of autism. Autism research is still feeling its way through the concept of regression outside of something like Heller's syndrome / CDD (and anti-NMDA receptor encephalitis and other viral infections in mind) and how such reports fit into the grand scheme of how and why autism comes about. As per some recent chatter on the early [behavioural] identification of autism, regression occurring in cases of autism is also a potential fly in the ointment for establishing very early indicators of autism, further compounded by the start-stop that accompanies child development. And then there is the issue of racial differences in the reported rates of regression in autism... already covered by some media just to complicate matters further.

More to do methinks.

To close, although Europe is braced for that annual get-together that is the Eurovision Song Contest tomorrow (Saturday 10th May 2014) I'm not gonna post to any of the past or present songs. Instead, some very motivational lyrics from Kiss. Thank God.

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[1] Richler J. et al. Is there a 'regressive phenotype' of Autism Spectrum Disorder associated with the measles-mumps-rubella vaccine? A CPEA Study. J Autism Dev Disord. 2006 Apr;36(3):299-316.

[2] Mishaal RA. et al. Age of autism spectrum disorder diagnosis is associated with child's variables and parental experience. Res Autism Spect Disord. 2014; 8: 873-880.

[3] Kern JK. et al. Evaluation of regression in autism spectrum disorder based on parental reports. N Am J Med Sci. 2014 Jan;6(1):41-7.

[4] Barger BD. et al. Prevalence and onset of regression within autism spectrum disorders: a meta-analytic review. J Autism Dev Disord. 2013 Apr;43(4):817-28.

[5] Goin-Kochel R. et al. Developmental regression among children with autism spectrum disorder: Onset, duration, and effects on functional outcomes. Res Autism Spec Disord. 2014; 8: 890-898.

[6] Woo EJ. et al. Developmental regression and autism reported to the Vaccine Adverse Event Reporting System. Autism. 2007 Jul;11(4):301-10.

[7] Valicenti-McDermott MD. et al. Gastrointestinal symptoms in children with an autism spectrum disorder and language regression. Pediatr Neurol. 2008 Dec;39(6):392-8.

[8] Hornig M. et al. Lack of association between measles virus vaccine and autism with enteropathy: a case-control study. PLoS One. 2008 Sep 4;3(9):e3140.

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Kern JK, Geier DA, & Geier MR (2014). Evaluation of regression in autism spectrum disorder based on parental reports. North American journal of medical sciences, 6 (1), 41-7 PMID: 24678477

Oxidative stress inducing mitochondrial dysfunction in autism?

Consider this post a sort of accompaniment to my recent chatter about the paper by Suzanne Goh and colleagues on brain lactate levels and mitochondrial dysfunction as a neurobiological subtype of autism (see here). Read on...

I've probably mentioned it before but outside of some light reading around the topic, mitochondria (plural) and mitochondrial dysfunction still remains a source of bafflement to me. Yes, I think I know the basics as per my learner post (see here) but it's the kinda of topic where I need to keep reading and re-reading sentences before any sense comes of it.

Bord de mer II @ Wikipedia 

It is therefore with a degree of trepidation that I approach the paper by Shannon Rose and colleagues [1] (open-access here) and their conclusion: "a significant subgroup of AD [autistic disorder] children may have alterations in mitochondrial function, which could render them more vulnerable to a pro-oxidant microenvironment as well as intrinsic and extrinsic sources of ROS [reactive oxygen species] such as immune activation and pro-oxidant environmental toxins". I might add that reports of mitochondrial dysfunction in cases of autism are seemingly increasing in the peer-reviewed literature [2].

OK, in very small steps, a few things to talk about here outside of my previous links to describing mitochondria shown above. ROS - reactive oxygen species - describes as the name suggests, molecules which contain oxygen (the important stuff that most creatures including us humans rely on for living). ROS are both a by-product of living (endogenous production) and can produced via other external sources (exogenous ROS). Endogenous production of ROS involves mitochondria, the so-called power plants in cells. Oxygen and various simple sugars are used to create ATP (adenosine triphosphate) as part of the process of oxidative phosphorylation. ROS are a by-product of this reaction which under certain circumstances can result in cell damage.

I'll also draw your attention to the concept of oxidative stress and it's relevance to this story. Basically, the body has ways to ensure that ROS production doesn't get out of hand via various cellular antioxidants. Regular readers of this blog might have already comes across some discussions on one of those antioxidants - glutathione - and the various findings in relation to autism (see here). As long as a happy balance is struck between appropriate levels of ROS and the scavenging availability of antioxidants, everything should tick along just fine. If however, ROS start to get the upper hand as described by the term oxidative stress, all sorts of effects can occur such as lipid peroxidation and oxidative damage to DNA. Ergo, balance is an important concept.

Back to the Rose paper then, which relied on something called lymphoblastoid cell lines (LCLs) as a means of measuring something called reserve capacity: "a measure of the ability of the mitochondria to respond to physiological stress" in cases of autism. LCLs came from the AGRE or the NIMH and researchers looked at mitochondrial reserve capacity "before and after exposure to reactive oxygen species (ROS)". Actually that wasn't the only research done on these LCLs, as the effect of N-acetylcysteine (NAC), "a glutathione precursor", pretreatment was also reported in the paper. The agent of choice for representing ROS in the study was something called DMNQ by the way.

Results, bearing in that I'm not going to go into the details of all the results because it's all there in open-access form for your interpretation:

• First and foremost: "LCLs derived from children with AD exhibit significant abnormalities in mitochondrial respiration after exposure to increasing levels of ROS". I temper those words with the authors suggestion that getting on for about half of the LCLs looked at (44%) seemed to drive the results obtained for the autism group. This might imply that in amongst all the talk about plural autisms, a sizeable subgroup of people on the autism spectrum may demonstrate such issues.
• "NAC rescues the atypical mitochondrial respiratory response". So, that pretreatment of the LCLs with NAC, specifically those 44% (10/22) who showed an abnormal adaptive response to ROS, seemed to improve mitochondrial respiration. This effect was not significantly present in the remaining LCLs from participants with autism.
• Glutathione (GSH) levels were also checked in cells (intracellular free GSH and also reduced glutathione - GSSG). As probably would be expected based on the other research literature in this area [3] lower glutathione levels were reported in the LCLs from participants with autism and a higher GSSG compared with controls. Perhaps not unexpectedly: "Pretreatment with NAC increased intracellular GSH and the GSH/GSSG ratio and reduced GSSG" in the LCLs from participants with autism.
• There are various other findings reported but the authors summarise by saying: "we demonstrate a new type of mitochondrial disorder that may affect a significant subgroup of AD children and provide insight into the interactions between systems that have been independently demonstrated to be abnormal in ASD [autism spectrum disorder]".

I know that I have gone on a little bit in this post because of the often quite technical jargon which is included in a study like this, so for that I apologise. My amateur status when it comes to talking about mitochondria and autism has probably not helped matters but I hope you can seem some glimmers of what the important results were derived from this relatively small-scale study. And yes, replication - independent replication - is an absolute must for this area of research bearing in mind facets of these results overlap with previous research by some of the same authors [4] and also here [5]. I might also add that other groups have started to talk about similar processes being involved in some autism too [6].

That all being said, I think you can see how important this work might eventually turn out to be when it comes to the area of mitochondrial dysfunction and autism, and indeed the possibility of links with other important areas of work such as the glutathione and NAC stories. Once again, I'm waiting attentively to see other published results in this area... including further replication of even more results from Napoli and colleagues [7] further discussed here.

And just in case my explanation of this area of work still leaves you baffled, I'll hand over to the experts and their recent review paper [8] ...

[Update: 16 March 2015: a clarification has been published over the paper from Rose and colleagues. Overlapping results had been mentioned...]

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[1] Rose S. et al. Oxidative stress induces mitochondrial dysfunction in a subset of autistic lymphoblastoid cell lines. Transl Psychiatry. 2014 Apr 1;4:e377.

[2] Giulivi C. et al. Mitochondrial Dysfunction in Autism. JAMA. 2010; 304(21): 2389–2396.

[3] Main PAE. et al. The potential role of the antioxidant and detoxification properties of glutathione in autism spectrum disorders: a systematic review and meta-analysis. Nutr Metab (Lond). 2012; 9: 35.

[4] James SJ. et al. Cellular and mitochondrial glutathione redox imbalance in lymphoblastoid cells derived from children with autism. FASEB J. 2009 Aug;23(8):2374-83.

[5] Rose S. et al. Oxidative stress induces mitochondrial dysfunction in a subset of autism lymphoblastoid cell lines in a well-matched case control cohort. PLoS One. 2014 Jan 8;9(1):e85436.

[6] Napoli E. et al. Evidence of reactive oxygen species-mediated damage to mitochondrial DNA in children with typical autism. Mol Autism. 2013 Jan 25;4(1):2.

[7] Napoli E. et al. Deficits in Bioenergetics and Impaired Immune Response in Granulocytes From Children With Autism. Pediatrics. 2014. April 21.

[8] Rossignol DA. & Frye RE. Evidence linking oxidative stress, mitochondrial dysfunction, and inflammation in the brain of individuals with autism. Front. Physiol. 2014. April 22.

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Rose, S., Frye, R., Slattery, J., Wynne, R., Tippett, M., Melnyk, S., & James, S. (2014). Oxidative stress induces mitochondrial dysfunction in a subset of autistic lymphoblastoid cell lines Translational Psychiatry, 4 (4) DOI: 10.1038/tp.2014.15

Asthma as a risk factor for autism?

I start with a quote from the paper by Po-Hsin Tsai and colleagues [1]: "This prospective study indicated a temporal relation between asthma and subsequent ASD [autism spectrum disorder] diagnosis, supporting the immune hypothesis of ASD pathogenesis". Based on a familiar collaboration residing in Taiwan (see here and see here for some discussions on previous data), yet more evidence is emerging of a correlation between immune function, physiology and behavioural / developmental conditions.

Chen Cheng-po @ Wikipedia 

I say more evidence because as per the links above, I've posted about other research from this group looking at the comorbidity of allergic and autoimmune diseases in cases of autism [2] and more recently evidence linking early life asthma with a heightened risk of subsequent attention-deficit hyperactivity disorder (ADHD) [3].

So here goes with yet more data from this group based on their analysis of the National Health Insurance Research Database (NHIRD) in Taiwan. Natasa also receives a thank you for bringing the full-text of this paper to my [virtual] desk.

• An asthma cohort was plucked from the NHIRD based on the random selection of 1 million persons on the database. Those aged between 0-3 years and with a physician diagnosis of asthma (ICD diagnosis) (n=2134) were age- and sex-matched with 4 control patients (1:4 ratio) with no asthma and no recorded history of psychiatric disorder (n=8536). Researchers then looked at the frequency of "Psychiatrists diagnosed ASD" (ICD-9 autism) up to some 8 years later. Other variables were also recorded for participants including other atopic/allergic diseases and level of urbanisation.
• Results: well, as per the headline: "The asthmatic group exhibited higher incidence rates of ASD (1.3% vs 0.7%, P=.007)". As perhaps one would expect, levels of other allergic diseases were also more frequently reported for the asthma group. Also, the asthma group were slightly more likely to be living in urban areas than controls, although urbanisation was not reported to significantly increase the risk of ASD (see a related post here).
• Interestingly, the authors also suggest that the presence of asthma "influences the manifestation pattern of ASD" based on the fact that asthmatics tended to be diagnosed with ASD later than those who were diagnosed with ASD in the control group (although this difference was not statistically significant: 5.6 years experimental group vs. 4.5 years control group).

As the authors point out, there is some noticeable methodological strength in their findings based on the database used, the numbers of participants included and the prospective nature of their inquiry. They do however caution that a cause-and-effect relationship between asthma and autism is not proved from their data. This same sentiment could also be applied to the recent attention given to the study by D’Onofrio and colleagues [4] on that age-old (pardon the pun) issue of older dads and autism risk.

I've talked about asthma and autism previously on this blog (see here) and how there may indeed be some overlapping features when it comes to the two conditions. I also hark back to some of the work by Kevin Becker (see this post) and his speculations on autism, asthma, inflammation and the Hygiene hypothesis [5] (open-access) which may well have been borne out to some degree by the Tsai data. If you want to get even more speculative, you might also have a look at my past post talking about the opinions of Moises Velasquez-Manoff and the immune roots of autism (see here)... [sorry for the sweeping generalisation by the way].

The only other things I'll add to the authors speculations about the 'mast cell activation' hypothesis (see here) as being linked to the findings is the possibility of a couple of other variables being potentially related too. So, how about medication being a common factor as per the preliminary work throwing prenatal paracetamol (acetaminophen) into the research limelight? Or, with my sunshine vitamin interest in mind, how about vitamin D as being something to tie the two conditions together [6]? And then there's the bank of evidence talking about maternal infection being linked to offspring asthma [7] which invites further study of a growing area of interest with autism in mind. Finally, there is all the chatter about environmental exposures and asthma [8] (open-access) some of which, without making a sweeping generalisation about the definition of 'environmental exposure', may overlap with some autism discussions [9]. Quite a few lines of potential further investigation methinks.

Outside of the need for quite a lot more study on this association (importantly, replication with other geographical datasets) including disentangling the very complicated issue of immune function and autism, there is a potentially important message to take from the Tsai data. Although perhaps only a small increased risk where early asthma is present, one might think about some additional screening for conditions like autism or ADHD just in case?

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[1] Tsai PH. et al. Increased risk of autism spectrum disorder among early life asthma patients: An 8-year nationwide population-based prospective study. Research in Autism Spectrum Disorders. 2014; 8: 381-386.

[2] Chen MH. et al. Comorbidity of allergic and autoimmune diseases in patients with autism spectrum disorder: A nationwide population-based study. Research in Autism Spectrum Disorders. 2013; 7: 205-212.

[3] Chen MH. et al. Asthma and attention-deficit/hyperactivity disorder: a nationwide population-based prospective cohort study. J Child Psychol Psychiatry. 2013 Nov;54(11):1208-14.

[4] D’Onofrio BM. et al. Paternal Age at Childbearing and Offspring Psychiatric and Academic Morbidity. JAMA Psychiatry. 2014. February 26.

[5] Becker KG. Autism, Asthma, Inflammation, and the Hygiene Hypothesis. Med Hypotheses. 2007; 69(4): 731–740.

[6] Gupta A. et al. Vitamin D and asthma in children. Paediatr Respir Rev. 2012 Dec;13(4):236-43

[7] Illi S. et al. Perinatal influences on the development of asthma and atopy in childhood. Annals of Allergy, Asthma & Immunology. 2014; 112: 132-139.

[8] Matsui EC. Environmental exposures and asthma morbidity in children living in urban neighborhoods. Allergy 2014; DOI: 10.1111/all.12361.

[9] Grandjean P. & Landrigan PJ. Neurobehavioural effects of developmental toxicity. Lancet Neurol. 2014 Mar;13(3):330-338.

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Po-Hsin Tsai, Mu-Hong Chen, Tung-Ping Su, Ying-Sheue Chen, Ju-Wei Hsu, Kai-Lin Huang, Wen-Han Chang, Tzeng-Ji Chen, & Ya-Mei Bai (2014). Increased risk of autism spectrum disorder among early life asthma patients: An 8-year nationwide population-based prospective study. Research in Autism Spectrum Disorders DOI: 10.1016/j.rasd.2013.12.022