p-cresol and autism: in need of further research

"These results confirm the elevation of urinary p-cresol in a sizable set of small autistic children and spur interest into biomarker roles for p-cresol and p-cresylsulfate in autism".

The peasant dance @ Wikipedia 

That was the primary conclusion from the paper by Gabriele and colleagues [1] looking at "three components of urinary p-cresol, namely p-cresylsulfate, p-cresylglucuronate and free p-cresol" in samples from 33 participants diagnosed with an autism spectrum disorder (ASD) compared with matched asymptomatic controls. The confirmation bit of that quote refers to the fact that members of this authorship group have previously reported on elevated urinary p-cresol in cases of autism [2] which was talked about in the very first proper research-based post on this blog (see here).

Before proceeding, perhaps it might be worth my while going through a few descriptive details. p-cresol (para-cresol) otherwise known as 4-methylphenol is a compound of some note in terms of the various ways and means one arrives at this organic aromatic compound. The solvent toluene is eventually metabolised into p-cresol, as is the amino acid tyrosine in the presence of strains of the anaerobic bacterium Clostridium difficile [3] for example. That being said, there are quite a few other ways in which one might come into contact with this compound.

According to the paper by Vanholder and colleagues [4] there is quite a bit of evidence to suggest that whilst p-cresol and its metabolites are compounds found in some quantity in just about everyone, under certain circumstances, elevations in amount may not be particularly desirable [5] particularly when it comes to renal functions. Indeed, quite a bit of the focus has been on the conjugated derivative p-cresylsulfate (formed through O-sulfonation) when it comes to toxicity [6]. I'll come back to this issue shortly.

A few points on the Gabriele paper might be useful:

• Based on a small participant group comprising 33 participants of various ages on the autism spectrum and 33 sex- and age-matched asymptomatic controls, levels of free p-cresol and it's two metabolites were measured via HPLC with fluorescence detection.
• All metabolites were "significantly elevated" in ASD cases compared to controls.
• "This increase was limited to ASD children ≤8 [less than or equal to 8] years". Also: "Urinary levels of p-cresol and p-cresylsulfate were associated with stereotypic, compulsive/repetitive behaviors (p < 0.05), although not with overall autism severity".

I probably don't need to say it, but when it comes to talk about biomarkers and autism, I do tend to be a little restrained about the promise of any results. Think back to my recent post on organic acids as biomarkers for autism (see here) and just about all the caveats talked about then in terms of heterogeneity and comorbidity come into play here too. That also this and other results from this group are based on HPLC with either UV (ultraviolet) or fluorescence detection could also be considered problematic as a function of the many and varied components found in urine and how without mass spectrometry or NMR, assigning labels to compounds is slightly problematic. Think casomorphins as another example...

Elevated levels of urinary p-cresol are also not a feature of every metabolomic study looking at autism. In their review of all-things p-cresol and autism, Persico & Napolioni [7] talked about how the results from Yap and colleagues [8] reported "blunted and not increased levels of p-cresylsulfate in autistic patients". The Yap study did utilise (1)H NMR spectroscopy and so did not suffer the same analytical shortcomings as the more recent trials. That all being said, I don't want to come down too hard on the latest results from Gabriele and colleagues. They got what they got and now put their results out for further inspection and hopefully, independent verification.

I am also wondering whether the paper by Clayton and colleagues [9] might also be relevant in this case. Dr Clayton, who some might remember from other work talked about on this blog (see here), discussed how "in individuals with high bacterially mediated p-cresol generation, competitive O-sulfonation of p-cresol reduces the effective systemic capacity to sulfonate acetaminophen [paracetamol]". Sulphation capacity when it comes to autism is already something of a research interest (see here) which when added to a growing body of work looking at paracetamol use during pregnancy and possible links to offspring development (see here) might indicate some other interesting investigations to be done. I wonder if perhaps even the sulphation depleting metabolism of something like p-cresol might actually be the more important part of such investigations to autism research?

Music to close, and are you a troublemaker?

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[1] Gabriele S. et al. Urinary p-cresol is elevated in young French children with autism spectrum disorder: a replication study. Biomarkers. 2014 Jul 10:1-8.

[2] Altieri L. et al. Urinary p-cresol is elevated in small children with severe autism spectrum disorder. Biomarkers. 2011 May;16(3):252-60.

[3] Dawson LF. et al. The analysis of para-cresol production and tolerance in Clostridium difficile 027 and 012 strains. BMC Microbiology 2011, 11:86

[4] Vanholder R. et al. p-cresol: a toxin revealing many neglected but relevant aspects of uraemic toxicity. Nephrol Dial Transplant. 1999 Dec;14(12):2813-5.

[5] Liabeuf S. et al. Free p-cresylsulphate is a predictor of mortality in patients at different stages of chronic kidney disease. Nephrol Dial Transplant. 2010 Apr;25(4):1183-91.

[6] Vanholder R. et al. The Uremic Toxicity of Indoxyl Sulfate and p-Cresyl Sulfate: A Systematic Review. J Am Soc Nephrol. 2014 May 8. [Epub ahead of print]

[7] Persico AM. & Napolioni V. Urinary p-cresol in autism spectrum disorder. Neurotoxicol Teratol. 2013 Mar-Apr;36:82-90.

[8] Yap IK. et al. Urinary metabolic phenotyping differentiates children with autism from their unaffected siblings and age-matched controls. J Proteome Res. 2010 Jun 4;9(6):2996-3004.

[9] Clayton TA. et al. Pharmacometabonomic identification of a significant host-microbiome metabolic interaction affecting human drug metabolism. Proc Natl Acad Sci U S A. 2009 Aug 25;106(34):14728-33.



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Gabriele S, Sacco R, Cerullo S, Neri C, Urbani A, Tripi G, Malvy J, Barthelemy C, Bonnet-Brihault F, & Persico AM (2014). Urinary p-cresol is elevated in young French children with autism spectrum disorder: a replication study. Biomarkers : biochemical indicators of exposure, response, and susceptibility to chemicals, 1-8 PMID: 25010144

Autoimmune disease as a risk factor for mood disorder?

Autoimmunity, the process by which the immune system fails to recognise self as self and subsequently targets those self tissues and cells, is something talked about quite a lot on this blog with autism specifically in mind. Part of the very wide and diverse immune-related features which have been discussed with at least some of the autisms in mind, it's not yet altogether clear exactly how and why autoimmunity is linked to behaviour but the association is an interesting one.

Sally? @ Wikipedia  

The paper by Michael Benros and colleagues* expands that autoimmune - behaviour interest and their observations suggesting: "Autoimmune diseases and infections are risk factors for subsequent mood disorder diagnosis". Their conclusion is based on the analysis of a huge patient set from which nearly 100,000 people had a hospital contact for mood disorder. Mood disorder by the way, is an overarching term for quite a few conditions (see here) which as the name suggests, affect mood; be it an unusual 'high' or an unusual 'low'.

"A prior hospital contact because of autoimmune disease increased the risk of a subsequent mood disorder diagnosis by 45%". To me this is a really interesting association. OK I know correlation does not equal causation, but added to the association they made between hospitalisation for infection and subsequent mood disorder (62% increase) and the large participant numbers, there is lots to ponder here.

I've gone on a few times on this blog about the bi-directional link between our physiology and the presentation of more psychiatric or behavioural 'symptoms'. Be it C.diff infection and depression or the skin-brain axis (see here), the connections are starting to be formed and examined. I'm just remembering back to some interesting research on allergic disease and neurodevelopment as also potentially being relevant to this post (see here) with the focus on allergy and its potential developmental effects and indeed the implication of some interesting potential interventions too.

So what could be the mode of action for this autoimmune-infection-mood disorder link? Well, we have some contenders but at the moment nothing concrete. That the immune system might be doing so much more than protecting against disease is already pretty well known about as for example with the rise and rise of research into microglia (see here) and some other areas of interest** particularly with a focus on a very nebulous term: inflammation.

Regular readers might know about my views and opinions on the role of food and the gut to behaviour - no really -  and certainly when it comes to certain autoimmune conditions like coeliac (celiac) disease, we have a body of research suggesting how food, certain elements of food, under the right circumstances might be able to affect behaviour. Think Marios Hadjivassilliou for example.

Even more outlandish, how about some role for those HERVs - human endogenous retroviruses - which seem to be cropping up with lots of conditions in mind (see here) and in particular autoimmune conditions***? Too speculative? Whether any of these areas overlap with the Benros paper is something as yet unknown. But the association posited between autoimmune conditions / infections and subsequent mood disorders is really, really interesting.

To close, with the new Stone Roses inspired movie on the horizon, how about a song about Sally Cinnamon?

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* Benros ME. et al. Autoimmune Diseases and Severe Infections as Risk Factors for Mood Disorders. JAMA Psychiatry. 2013;():1-9. doi:10.1001/jamapsychiatry.2013.1111.

** del Rey A. et al. A cytokine network involving brain-borne IL-1β, IL-1ra, IL-18, IL-6, and TNFα operates during long-term potentiation and learning. Brain, Behavior, and Immunity. 6 June 2013. http://dx.doi.org/10.1016/j.bbi.2013.05.011

*** Dreyfus DH. Autoimmune disease: A role for new anti-viral therapies? Autoimmun Rev. 2011 Dec;11(2):88-97. doi: 10.1016/j.autrev.2011.08.005.

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Benros ME, Waltoft BL, Nordentoft M, Ostergaard SD, Eaton WW, Krogh J, & Mortensen PB (2013). Autoimmune Diseases and Severe Infections as Risk Factors for Mood Disorders: A Nationwide Study. JAMA psychiatry (Chicago, Ill.), 1-9 PMID: 23760347

Depression or antidepressant use linked to C.diff infection?

"There's no mystical energy field that controls my destiny". So said a very sceptical Han Solo.

Regular readers might know that I'm a bit of fan of the whole gut-brain axis; indeed other kinds of axes too. I know that to some it might sound a bit daft that what goes on in our deepest, darkest bowels might actually have some important effects on the operations of the grey-pinkish matter floating around in skull central - and vice-versa -  but nonetheless it interests me. The gastrointestinal (GI) tract is not quite the mystical energy field that Captain Solo was referring to, but make no mistake, we are still very much in the infancy of looking at the connection between the two systems*.

Black dog @ Wikipedia  

I've tended to discuss/speculate on the gut-brain relationship with regards to cases of autism spectrum disorder (ASD) on this blog. In this post I'm branching out to look at the paper by Mary Rogers and colleagues** (open-access) on a potentially new dimension to the gut-brain conversation with depression and Clostridium difficile infection in mind.

The Rogers paper is open-access and has also attracted some media attention as a result (see here and here for the press release). The long-and-short of it was that based on two studies - a sort of scientific BOGOF - looking at the rates of C.diff infection (CDI) in participants with and without depression and the potential effects of antidepressant medication use and hospital-acquired CDI, some interesting correlations were noted. Note that word 'correlations'...

Primary among the findings was the suggestion that the chances of CDI were higher in those presenting with depression: "After adjusting for demographic characteristics, comorbidities and frequency of medical visits, there was a 36% increase in the odds of developing CDI for individuals with major depression compared with those without major depression" (CI: 1-06-1.74, p=0.016). Indeed when it came to the label of "emotional, nervous or psychiatric problems", the CDI risk was found to be even higher (OR: 1.47). Certainly some interesting data, made all the more credible by the fact that the total sample size numbered in the thousands.

When it came to medication use, there were some equally interesting associations (not) made. So for example, laboratory confirmed CDI (via stool testing) seemed not to correlate with the majority of medicines participants were also taking at the time of testing. The exceptions were mirtazapine (OR: 2.14) and fluoxetine (OR: 1.92) which were individually associated with an approximate doubling of CDI risk and also carrying some dose-related associations.

Authors also reported that polypharmacy - if I can use that word with less than 5 meds being taken - might also impact on CDI risk, as per the "significant interaction between mirtazapine and trazodone" where "the odds of a positive C. difficile test were 5.72 times greater" bearing in mind the small participant numbers who were prescribed these two drugs combined. As per the press on this paper: "People who have been prescribed these types of anti-depressants need to keep taking them unless otherwise advised by their physician"; a viewpoint that I can only echo at this stage.

You can perhaps see why I might be interested in this line of research. There is of course the chicken-and-egg question about which came first: microbial changes which place a person at greater risk of CDI and perhaps depression, or depression leading to changes to the gut microbiota and onwards elevated CDI risk. I'm not going to speculate too much on what came first because I dare say the clinical picture is going to be much more complicated than such a simple question. I've talked before about the possibility of a bi-directional relationship between gut bacteria and behaviour (at least in mice) and my viewpoint has changed very little in the intervening years. Bear also in mind that the hows and whys of depression (in all its forms) are likely to be numerous; perhaps even related to our earliest years****

It's interesting that the authors discuss quite a few important overlapping pieces of research in their summary of their findings related to things like the presence of bowel disease in cases of depression*** and that magical word 'inflammation'*****. To quote: "It is possible that there is a lifelong liaison between the gut microbiota and neurologic response to external stimuli" which certainly does seem to link in with at least some of the current research literature.

Alongside the tentative associations made by Rogers et al on the issue of depression and CDI, I find my mind wandering back to the question of whether such an association might be something which could be translated into therapeutic options. Y'know whether treating the CDI actually had any quantifiable impact on the presentation of depression or vice-versa. Indeed whether one of the more unusual methods suggested to help combat CDI - yep, the fecal transplant - might also impact on depression via changes to the gut microbiota as per the very preliminary reports from other conditions such as chronic fatigue syndrome (CFS)? That and possibility that gut bacteria might, just might, be in cahoots with other more barrier-related issues******, makes for some interesting suggestions for further scientific inquiry*******.

To end, y'know I prefer Constantiople over Istanbul....

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* Collins SM. & Bercik P. Gut microbiota: intestinal bacteria influence brain activity in healthy humans. Nature Reviews Gastroenterology and Hepatology. May 2013.

**  Rogers MAM. et al. Depression, antidepressant medications, and risk of Clostridium difficile
infection. BMC Medicine 2013; 11: 121.

*** Graff LA. et al. Depression and anxiety in inflammatory bowel disease: a review of comorbidity and management. Inflamm Bowel Dis. 2009; 15: 1105-1118.

**** Parboosing R. et al. Gestational influenza and bipolar disorder in adult offspring. JAMA Psychiatry. May 2013.

***** Vogelzangs N. et al. Association of depressive disorders, depression characteristics and antidepressant medication with inflammation. Transl Psychiatry. 2012; 2: e79.

****** Maes M. et al. Increased IgA and IgM responses against gut commensals in chronic depression: further evidence for increased bacterial translocation or leaky gut. J Affect Disord. 2012; 141: 55-62.

******* Hughes PA. et al. Immune activation in irritable bowel syndrome: can neuroimmune interactions explain symptoms? Am J Gastroenterol. May 2013

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Rogers, M., Greene, M., Young, V., Saint, S., Langa, K., Kao, J., & Aronoff, D. (2013). Depression, antidepressant medications, and risk of Clostridium difficile infection BMC Medicine, 11 (1) DOI: 10.1186/1741-7015-11-121

A vaccine for autism symptoms? Not exactly

"Vaccine". "Autism".

I'm struggling to think of two words in combination which, in modern times, are any more likely to stir up emotion, debate and even argument. Indeed in these times of measles outbreaks and seemingly daily news reporting on the very, very strong requirement for vaccination to protect against the disease, it is coincidental that two research papers should now land in my inbox which mention both of those words in the title.

Paradise in Zakynthos @ Wikipedia  

The first paper is by Ivan Gentile and colleagues* reporting on seropositivity rates to measles, mumps and rubella in cases of autism spectrum disorder (ASD) following MMR vaccination. The second paper, by Brittany Pequegnat and colleagues**, discusses the concept and early development of a vaccine targeting a specific type of gut bacteria which the authors speculate might have some interesting knock-on effects to some of the signs and symptoms linked to cases of autism.

Although pertinent to the current measles news, I'm not on this occasions heading too far into the Gentile paper. My reasoning is two-fold: (i) the paper is fairly explanatory in that "children with ASD have a similar level and seropositivity rate of antibodies against the MMR vaccine to same-age controls" (bearing in mind the small participant numbers) and, (ii) I'm not really qualified to go into any heavy duty discussions on how this fits into the existing scientific literature on this topic; bearing in mind a similar finding previously published*** and a contrary one****. All I will say is that vaccination saves lives as per this CDC flier.

The Pequegnat paper has received some media attention with headlines like: "Vaccine To Help Autism Symptoms Developed" and "Scientists develop first vaccine to help control autism symptoms". As one might expect, headlines which don't necessarily reflect the actual science reported so far...

The long-and-short of the research is that based on some earlier findings of a specific gut bacterium being discovered in a group of children with autism***** - Clostridium bolteae previously called Clostridium clostridioforme (see here******) but renamed, I think, after Ellen Bolte******* - the authors applied some established know-how to begin formulating a vaccine targeting the surface sugars, polysaccharides, of C.bolteae. If you want to see the human face of the researchers involved, look no further than this article from 2012.

All that talk of this research helping to 'control' autism symptoms is, at the moment, more speculation than fact. As far as I can see, the authors got no further than providing the "first description of a C.bolteae immunogen" following some initial investigation in rabbits. It is therefore a significant jump to say that this vaccine will affect the behavioural presentation of autism. Indeed, no-one really knows if it will impact on any gastrointestinal (GI) symptoms either.

That being said, I am quite interested in their report and the concept that we could artificially stimulate immunity to 'undesirables' such as specific types of gut bacteria. As well as being particularly interested in all things bacteria on this blog (see here and here), a quick trawl of the scientific literature suggests that the future is now as per the paper by Sougioultzis and colleagues******** (open-access) on a C.diff toxoid vaccine. One wonders whether we might also apply similar logic to other bacterial findings related to autism such as that very interesting Sutterella work?

I am also drawn to the polysaccharide bit of the Pequegnat paper and whether or not it is useful to link back to the work of Harumi Jyonouchi and colleagues on the presence of specific polysaccharide antibody deficiency (SPAD) comorbid to some cases of autism. If I am interpreting this correctly, the suggestion is that SPAD interferes with correct antibody formation to polysaccharide coated bacteria which could have implications I assume, for vaccination to/against bacteria like C.bolteae also. I could be wrong though.

I'm gonna stop with this post shortly. There are other things I could say, for example, discussing the method of vaccine delivery (nanoparticle anyone?) including doing away with the big scary needle in favour of something a little more 'ouchless' (microneedles or even inhaled delivery). But that is perhaps fodder for another day.

Oh, and just so you know, on purpose I posted a lovely serene picture from the beautiful island of Zakynthos instead of one of those pictures of big needles complete with crying child which, as other commentators have pointed out, might not necessarily be the best platform on which to discuss the topic of vaccination.

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* Gentile I. et al. Response to Measles-Mumps-Rubella vaccine in children with autism spectrum disorders. In Vivo. 2013; 27: 377-382.

** Pequegnat B. et al. A vaccine and diagnostic target for Clostridium bolteae, an autism-associated bacterium. Vaccine. April 2013.

*** Baird G. et al. Measles vaccination and antibody response in autism spectrum disorders. Arch Dis Child. 2008; 93: 832-837.

**** Singh VK. et al. Abnormal measles-mumps-rubella antibodies and CNS autoimmunity in children with autism. J Biomed Sci. 2002; 9: 359-364.

***** Finegold SM. et al. Gastrointestinal microflora studies in late-onset autism. Clin Infect Dis. 2002; 35 (Supplement 1): S6-S16.

****** Song Y. et al. Real-time PCR quantitation of Clostridia in feces of autistic children. Appl. Environ. Microbiol. 2004; 70: 6459-6465.

******* Bolte ER. Autism and Clostridium tetani. Med Hypotheses. 1998; 51: 133-144.

******** Sougioultzis S. et al. Clostridium difficile toxoid vaccine in recurrent C. difficile–associated diarrhea. Gastroenterology 2005; 128: 764–770.

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Pequegnat B, Sagermann M, Valliani M, Toh M, Chow H, Allen-Vercoe E, & Monteiro MA (2013). A vaccine and diagnostic target for Clostridium bolteae, an autism-associated bacterium. Vaccine PMID: 23602537

Battling Clostridia difficile

Perhaps second only to MRSA, the bacteria Clostridia difficile (C.difficile) is on various 'wanted' posters in many hospitals around the world. I say 'wanted' but in this instance do not refer to a welcome invitation, rather the $10,000 reward variety of wanted. Wanted dead or alive as Bon Jovi once asked? Dead in this case.

Why? Well because C.difficile whilst being present in a proportion of the population, and not seemingly involved in anything untoward in many cases, has the propensity to do some pretty nasty things to a person under the right conditions - usually associated with antibiotic use. One of the most widely presented links is with something called pseudomembranous colitis. I am interested in C.difficile and the colitis angle because of the link to gastrointestinal hyperpermeability (the leaky gut) which has had its speculations for some cases of autism and other conditions.

There is some hope on the horizon for tackling C.difficile as represented by this article* by Savidge and colleagues published in Nature Medicine. Don't be too put off by the title and abstract because the BBC website carries quite a good layman version of the findings. The long and short of it is that the compound S-nitrosoglutathione (GSNO), a nitric oxide donor, may be a key component in inhibiting the ability of the bacteria to enter cells and exert its damaging effect. Tests on mice (sorry!) suggested that GSNO helped survival rates in cases of clostridial infection.

The authors are cautious in their findings and application to treating C.difficile infection. Lots more research is required before this compound goes from peer-reviewed paper to practice not least to make sure additional supplementation is actually safe. I am however intrigued about this compound and its potential effects including its proposed effects on things like nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) which might show more than a passing connection to some cases of autism spectrum conditions. More than that though is the potential to treat C.difficile, which at the moment might (and I do stress 'might') show some connection to autism and some intervention measures previously described.

Your days might be numbered C.difficile infection...

* Savidge TC. et al. Host S-nitrosylation inhibits clostridial small molecule-activated glucosylating toxins. Nature Medicine. August 2011.