Aluminium in brain tissue in autism: a first?

"These are some of the highest values for aluminium in human brain tissue yet recorded and one has to question why, for example, the aluminium content of the occipital lobe of a 15 year old boy would be 8.74 (11.59) μg/g dry wt.?"

That was one of the statements/questions posed in the paper by Matthew Mold and colleagues [1] covering yet another potential 'hot potato' topic in the area of autism research and science (see here). Indeed, the idea mentioned by Mold et al that: "Animal models of ASD [autism spectrum disorder] continue to support a connection with aluminium and to aluminium adjuvants used in human vaccinations in particular" is likely to met with furrowed brows in quite a few quarters given the recent goings-on with regards to some of the 'mouse' work in this area (see here).

But as far as I can see, the Mold data is a stand-alone piece of work that sought to cover a 'missing piece' in relation to how "no previous reports of aluminium in brain tissue from donors who died with a diagnosis of ASD" have been published in the peer-reviewed domain. In that respect, I'm covering it on this blog, hypothesis-free in terms of the possible wheres-and-hows...

Relying on one of the most personal and precious resources in autism research - brain tissue - "from the Oxford Brain Bank ", researchers examined brain tissue samples from various parts of the brain donated by families of "5 individuals with ADI-R confirmed (Autism Diagnostic Interview-Revised) ASD." As far as I can make out, we aren't told an awful lot about the 5 people studied aside from their diagnosis, sex/gender and age range. We don't for example, know about any comorbidities (autism rarely appears in some sort of diagnostic vacuum) or indeed, the reason(s) why they died.

Brain tissues were prepared and treated ready for analysis of aluminium content via "transversely heated graphite furnace atomic absorption spectrometry (TH GFAAS) using matrix-matched standards and an established analytical programme alongside previously validated quality assurance data." From what I gather, this is an accepted (albeit quite old) method for analysis of various trace metals in biological fluids including aluminium [2]. It's also worthwhile bearing in mind the publication record of some of the authors in the area of aluminium (see here) including research pertinent [3] to the Camelford 'incident'. Some people here in Blighty might remember Camelford, where the accidental addition of some 20 tonnes of aluminium sulphate into the local drinking water led to it being described as "Britain's worst mass poisoning event".

Alongside assaying for aluminium content in brain samples, researchers also examined brain sections to see where aluminium congregated in samples via the use of fluorescence microscopy. This involved the selective staining of cells for aluminium content and onward looking at where and what type of cell showed such staining.

Results: "The brains of all 5 individuals had at least one tissue with a pathologically-significant content of aluminium." Further: "The brains of 4 individuals had at least one tissue with an aluminium content ≥5.00 μg/g dry wt. while 3 of these had at least one tissue with an aluminium content ≥10.00 μg/g dry wt." I don't know enough about aluminium and what should or shouldn't be there, but I believe the authors hark back to some of their previous work to try and define "loose categories of brain aluminium content." In terms of the aluminium cell staining work, some interesting findings are reported pertinent to how "aluminium somehow had crossed the blood-brain barrier and was taken up by a native cell namely the microglial cell." Other authors had previously speculated on this scenario [4].

It's worthwhile mentioning/reiterating some of the limitations of this work before any grand sweeping judgements or generalisations are made. Not only was this a very, very small study with no specific control samples available during this particular investigation, but the amount of tissue available to researchers was limited. One cannot also rule out issues such as sample degradation being a factor (see here) given the lack of specific details on the samples being investigated. As I said, brain samples are one of the most precious resources available to autism research.

There is however a need for much more research in this area, particularly when going back to the opening statement/question of this post in terms of amount of aluminium detected and where it seemed to congregate in the brain samples analysed. Indeed, with all the chatter about microglia and inflammation in relation to [some] autism down the years (see here for example), it's looking increasingly *interesting* that aluminium might play some kind of role, for some at least. Added to independent findings suggesting that the blood-brain barrier should also be target for further study for some (see here) and observations that upwards of about 15% of individuals with an ASD might present with elevated levels of aluminium in other tissue types (see here) and future work is indicated on a larger scale minus hype and/or generalisations. I might also suggest that further research be directed on the basis of other organs potentially affected by aluminium concentrations such as the kidneys..

I'm also minded to point out that following other work on other metals in the context of autism (see here), I'm still siding with the idea that various genetic and biological mechanisms involved in the *removal* of metals may be 'atypical' in relation to at least some autism. If that is the case, there could be several different ways to potentially intervene [5]...

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[1] Mold M. et al. Aluminium in brain tissue in autism. Journal of Trace Elements in Medicine and Biology. 2017. Nov 26.

[2] Bradley C. & Leung FY. Aluminum determined in plasma and urine by atomic absorption spectroscopy with a transversely heated graphite atomizer furnace. Clin Chem. 1994 Mar;40(3):431-4.

[3] King A. et al. Unusual neuropathological features and increased brain aluminium in a resident of Camelford, UK. Neuropathol Appl Neurobiol. 2017 Oct;43(6):537-541.

[4] Morris G. et al. The putative role of environmental aluminium in the development of chronic neuropathology in adults and children. How strong is the evidence and what could be the mechanisms involved? Metab Brain Dis. 2017 Oct;32(5):1335-1355.

[5] Yu L. et al. Lactobacillus plantarum CCFM639 can prevent aluminium-induced neural injuries and abnormal behaviour in mice. Journal of Functional Foods. 2017; 30: 142-150.

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"New form of autism found"

"New form of autism found" went one of the headlines reporting on the paper by Dora C. Tărlungeanu and colleagues [1] and findings that "elucidate a neurological syndrome defined by SLC7A5 mutations and support an essential role for the BCAA [branched-chain amino acids] in human brain function." This work continues a rather important research story talking about how one 'type' of autism might have some important roots in relation to the branched-chain amino acids and their metabolism (see here and see here for more information).

So, mice were the focus on the paper by Tărlungeanu et al (including one Gaia Novarino on the authorship list) and an extension of the idea that the BCAAs may play an important role in some autism in these days of the plural 'autisms' (see here). SLC7A5 represents a gene that codes for a protein involved in the transport of BCAAs into the brain among other things. Researchers studied mice who were genetically 'edited' to present with a "deletion of Slc7a5 from the endothelial cells of the BBB [blood-brain barrier]." In effect, the area of the body where SLC7A5 serves those important transport duties, a hold-my-hand partner was missing resulting in lower brain levels of the BCAAS.

Researchers noted a few important things in those SLC7A5-missing mice; not least in relation to their mouse behaviour(s) and how bearing in mind mice are mice not people, they seemed to present with behavioural issues not a million miles away from that noted in relation to autism. 'Social interaction' was as I understand it, something potentially affected in those SLC7A5-missing mice. Further: "we identified several patients with autistic traits and motor delay carrying deleterious homozygous mutations in the SLC7A5 gene" suggesting that their results might stretch to people too.

And then something else that might eventually be important: "we demonstrate that BCAA intracerebroventricular administration ameliorates abnormal behaviors in adult mutant mice." Intracerebroventricular administration basically means an injection straight into the brain. After a few weeks of such injections, researchers noted that mouse behaviours began to change coincidental to the direct administration of those BCAAs.

This is interesting research. I know that not everyone on the autism spectrum presents with issues with the BCAAs (as far as we know). But in these days of increasing plurality when it comes to autism coupled to the rise and rise of study on the various inborn errors of metabolism in relation to autism (see here), this could be pertinent to at least one type of autism. I also appreciate that 'brain injections' of something like BCAAs are not exactly a desirable option for anyone so there is still some work to do in terms of how to correct any central BCAA deficiency if and when identified. Talk about a possible relationship between the BBB and autism in the Tărlungeanu paper also continues a theme (see here) where this important barrier separating the brain from the other contents of the body (and indeed, the outside world) might represent something potentially quite important to autism (see here) and indeed, with 'transporters' also in mind (see here).

Much more research is implied.

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[1] Tărlungeanu DC. et al. Impaired Amino Acid Transport at the Blood Brain Barrier Is a Cause of Autism Spectrum Disorder. Cell. 2016. Dec 1.

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Tărlungeanu, D., Deliu, E., Dotter, C., Kara, M., Janiesch, P., Scalise, M., Galluccio, M., Tesulov, M., Morelli, E., Sonmez, F., Bilguvar, K., Ohgaki, R., Kanai, Y., Johansen, A., Esharif, S., Ben-Omran, T., Topcu, M., Schlessinger, A., Indiveri, C., Duncan, K., Caglayan, A., Gunel, M., Gleeson, J., & Novarino, G. (2016). Impaired Amino Acid Transport at the Blood Brain Barrier Is a Cause of Autism Spectrum Disorder Cell, 167 (6), 1481-2147483647 DOI: 10.1016/j.cell.2016.11.013

Gut barrier integrity meets blood-brain barrier integrity with autism in mind

"In the ASD [autism spectrum disorder] brain, there is an altered expression of genes associated with BBB [blood-brain barrier] integrity coupled with increased neuroinflammation and possibly impaired gut barrier integrity."

Although pretty enthused to see research linking names like Anna Sapone, Tim Buie and Alessio Fasano in the recent paper published by Maria Fiorentino and colleagues [1] (open-access), I was slightly less impressed with the use of the term 'the ASD brain' in their paper potentially joining two concepts that I've been quite interested in down my research years: gut barrier and blood-brain barrier function in the context of autism. Yes, I accept that those most precious of resources, donated brains from the deceased, represented some of the 'material' under scientific scrutiny, but if science has learned anything about autism down the years, it is that sweeping generalisations such as terms like 'the autism brain' don't reflect what the existing research tells us about the heterogeneity under the label. I might just as well use the term 'blogger brain' to denote some of my activities, but such a label tells you nothing about me aside from my pastime.

After that little rant, the paper from Fiorentino is an interesting one in that the goal was to "investigate whether an altered BBB and gut permeability is part of the pathophysiology of ASD." To do this, tissue from both brain and gastrointestinal (GI) tract donated by a small number of deceased and non-deceased participants who were diagnosed with autism, schizophrenia or nothing related (not-autism controls) were analysed "for gene and protein expression profiles." This work was undertaken on the basis of "the interconnectivity of the gut–brain axis, [that] suggests that inappropriate antigen trafficking through an impaired intestinal barrier, followed by passage of antigens or activated immune complexes through a permissive blood–brain barrier (BBB), can be part of the chain of events leading to neuroinflammation and thereby subsequent disease." I might add that the use of the word 'disease' in that sentence is, I think, aiming to describe the physiological effects of 'leaky barriers' not the diagnosis of autism. It is unfortunate however that 'disease' still continues to be banded around in the context of autism [2].

I think it's important to stress that the Fiorentino study was in effect two studies: one that looked at brain samples from one participant group who had died, and one that looked at GI samples from those who were still living (at the time of sample collection) and who presented with "GI symptoms undergoing esophagogastroduodenoscopy (EGD) for clinically indicated reasons." This was not a study where biological samples - brain and gut - came from the same person but rather a mash-up. Keep that in mind for now. The sorts of genes that were focused in on were those "associated with the formation, integrity, and function of the BBB and neuroinflammation" and included the claudins and something called MMP-9 and MMP-2 that have been discussed previously on this blog (see here) with leaky barriers in mind. The key words are 'barrier integrity' when it comes to the list of compounds that were under inspection.

Results: well it was good to see the authors list details of each of the participants from which tissue were used in their study. Brain tissue from the deceased with autism for example, is subject to quite a few factors that can influence the outcome of any results obtained; not least whether specific comorbidity accompanied their autism diagnosis and the nature of their death. Indeed, looking through the various case report numbers, I'm struck by how young many participants, particularly those diagnosed with autism, were at the time of their death. This ties into other discussions and debates (see here).

"Our molecular analysis of the BBB integrity and function shows an altered BBB in the ASD subjects evaluated." This was evidenced by elevations in the gene expression of MMP-9 and its proposed connection to disturbances of BBB integrity. Further: "Of the four claudins (i.e., CLDN-1, -3, -5 and -12) that to date are thought to be incorporated in the BBB... we found that two were significantly more expressed in the ASD brain as compared in HC [healthy controls]." Once again I might suggest the term 'healthy controls' is not an inappropriate one when it comes to determining not-autism or not-schizophrenia.

Then to analysis of those [independent] gut biopsy samples: "results, showing increased expression levels of pore-forming (66% of the ASD samples) and decreased levels of barrier-forming (75% of the ASD samples) TJ [tight junction] components in the duodenal samples, suggest an impaired gut barrier and serve as a proof of concept to support the hypothesis of a gut–brain axis dysfunction in a subgroup of ASD patients." So, those compounds linked to making the gut barrier more 'leaky' were seemingly increased in expression, and those linked to making the gut barrier less 'leaky' were reduced in quite a few of the samples from those diagnosed with autism. Mmm...

There is quite a bit more science included in the Fiorentino study but I think I've gone on long enough in this post. Suffice to say that the whole gut-brain axis thing with autism in mind gets a boost but more work is indicated, not least with larger sample groups and perhaps combining tissues from gut and brain from the same person. I would also like to see a little more done on this topic with some 'interventions' in mind, based on the other autism research that potentially links the authors (see here). Drawing for example, on a paper written by Prof Fasano titled: 'Zonulin, regulation of tight junctions, and autoimmune diseases' [3] suggesting that "gliadin, a storage protein present in wheat and that triggers celiac disease in genetically susceptible individuals, also affect the intestinal barrier function by releasing zonulin" one might see how far from being a set-in-stone state of affairs, dietary changes for some on the autism spectrum, might actually set in motion a host of biological changes pertinent to this area of work. And such changes might not be just confined to accepted gluten-related conditions either...

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[1] Fiorentino M. et al. Blood–brain barrier and intestinal epithelial barrier alterations in autism spectrum disorders. Molecular Autism. 2016; 7:49.

[2] Simms MD. When Autistic Behavior Suggests a Disease Other than Classic Autism. Pediatr Clin North Am. 2017 Feb;64(1):127-138.

[3] Fasano A. Zonulin, regulation of tight junctions, and autoimmune diseases. Annals of the New York Academy of Sciences. 2012;1258(1):25-33.

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Fiorentino, M., Sapone, A., Senger, S., Camhi, S., Kadzielski, S., Buie, T., Kelly, D., Cascella, N., & Fasano, A. (2016). Blood–brain barrier and intestinal epithelial barrier alterations in autism spectrum disorders Molecular Autism, 7 (1) DOI: 10.1186/s13229-016-0110-z

More suramin and autism [mouse] findings

The headline: 'Century-old drug reverses signs of autism in mice' brought the paper by Jane Naviaux and colleagues [1] (open-access) to my attention and some slightly familiar work (see here) on the use of suramin in a mouse model of autism, or rather a mouse model of maternal immune activation. Indeed, I seem to remember that the previous study by this group [2] courted similar publicity, with some familiar headlines...

Not lecturing... @ Wikipedia 

The latest offering from Naviaux et al is not so dissimilar from their previous work on that mouse model of autism, this time testing "the hypothesis that the behavioral manifestations of the MIA [maternal immune activation] model are a consequence of pathological persistence of the evolutionarily conserved CDR [cell danger response]... and that the CDR is maintained by dysregulated purine metabolism and secondary abnormalities in purinergic signaling". The press release about the study can be viewed here.

What this all translates into is that a state of immune arousal above and beyond what would be typically expected during pregnancy is somehow impacting on offspring development pertinent to an elevated risk of conditions like autism or schizophrenia. There is quite a bit of literature on this topic from the autism perspective (see here) and based on animal models other than just rodents (see here). The cell danger response (CDR) described by Naviaux (Robert that is) [3] represents "the evolutionarily conserved metabolic response that protects cells and hosts from harm". Indeed that last reference [3] contains just about every biological link known to autism science at the current time... including mitochondria. The idea is that triggering immune activation activates "a conserved cellular response to stress" called the CDR and the proposed master regulator of the CDR is purinergic signalling [4] - "purine nucleotides and nucleosides as extracellular messengers". ATP (adenosine triphosphate) as well as being quite an important molecular fuel source, is suggested to be one of the nucleotides which "can bind to cell surface receptors and act as signaling molecules and neuromodulators that are important in inflammation.. neurotransmission.. and many other biological processes".

A few basic points about the Naviaux study:

• Mice were the lucky volunteers for this study, and again the 'good breeder' that is the C57BL/6J variety. As per their previous trial, pregnant female mice were given something to artificially stimulate their immune system and "initiate the MIA model" or a saline control and then their offspring were the study focus.
• Suramin or saline (as a control) was then administered to 6-month old offspring mice and thereafter "behaviors were evaluated". Suramin levels were also examined, as were a broad range of metabolites as part of some metabolomic analysis based on the use of triple quad mass spectrometry.
• Results: "MIA animals showed social deficits from an early age". Nothing too novel there bearing in mind previous observations in this area of research. But... "Single-dose APT [antipurinergic therapy] with suramin completely reversed the social abnormalities in 6.5-month-old adults". Social behaviour by the way, was quantified as "time spent interacting with a novel ("stranger") mouse".
• The benefit of suramin also lasted for quite a while: "a small residual benefit to social behavior was still detectable" even after 5 weeks following the intervention. There is some discussion by the authors about this effect; noted to be potentially "due to the development of metabolic memory and/or somatic epigenetic DNA changes that lasted longer than the physical presence of the drug".
• Then the biochemistry. Suramin seemed to by-pass that very important gateway, the blood-brain barrier (BBB) and end up in the brainstem following some analysis of sacrificed offspring mice: "consistent with the notion that nuclei in brainstem, or their projection targets in distant sites of the brain, may mediate the dramatic behavioral effects of acute and chronic APT in this model".
• And more: "Comprehensive metabolomic analysis revealed disturbances in several other metabolic pathways relevant to children with ASDs. These included disturbances in microbiome, phospholipid, cholesterol/sterol, sphingolipid, glycolytic and bile salt metabolism". This bearing in mind that only "male animals that had been behaviorally evaluated were tested". Purine metabolism and the gut microbiome seemed to be quite important to the author's results. At this point I'll refer you back to the work by Elaine Hsiao and colleagues... 
• Moreover: "The top, non-microbiome-associated metabolite was quinolinic acid... which was decreased in the MIA model". Quinolinic acid implies the involvement of one of those aromatic amino acids, tryptophan, which is an autism research favourite [5]. More than that is the literature on the intersecting kynurenine pathway and how that might relate to a condition like schizophrenia (see here).
• The authors caution that their results are (a) mouse based and (b) "suramin is a poor drug choice for chronic use because of potentially toxic side effects that can occur with prolonged treatment". That being said, they do suggest that "new drugs might be given only once, or intermittently, during sensitive windows to unblock metabolism, restore more normal neural network function, improve resilience and plasticity, and permit improved development in response to behavioral and interdisciplinary therapies, and to natural play". Just in case you'd like an alternative reading of this study, have a look at this write-up too.

I do apologise for all the quotes taken from the Naviaux paper and used in this post, but when the authors say it better than I could, why would I try and complicate things any further? Indeed, the more I read the paper by Naviaux and colleagues, the more I see what a potential gem it actually is. I say this based on the pretty comprehensive way that the authors went about looking at the MIA model and the effect of their intervention, crossing behavioural (mouse behavioural) and biochemical fields. Mention of the words 'microbiome' and 'metabolome' are also guaranteed to perk my interest. All that enthusiasm is however tempered dependent on the work being replicated and not just in a mouse model either.

A few closing comments are all that are required. Looking at the list of biochemical pathways altered in the MIA model of autism (see Table 1 here) we see lots of familiar names too numerous to mention. Outside of looking at the effect of suramin on these pathways, I'm minded to suggest that other interventions might also benefit from some similar inspection of their effects on said pathways. Even as one commentator has already noted: "[The findings] are valuable, but the main problem is that they rely on a model of immune infection, not a genuine model of autism... They should have tested one of the classical models of autism... meaning genetic models of autism — to see whether suramin indeed corrects autistic behaviors".

I've also not really gone into the detail when it comes to the cell danger response (CDR) in this post because it is a complicated area and seems to tie into various issues which are slightly beyond my level of expertise. I would like to bring in the letter by Theoharides [6] at this point however, and his discussion linking "extracellular mitochondrial material" to mast cell activation (see here) on the back of the previous study by this authorship group, as another area crying out for further investigation.

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[1] Naviaux JC. et al. Reversal of autism-like behaviors and metabolism in adult mice with single-dose antipurinergic therapy. Translational Psychiatry. 2014; 4: e400; doi:10.1038/tp.2014.33

[2] Naviaux RK. et al. Antipurinergic therapy corrects the autism-like features in the poly(IC) mouse model. PLoS One. 2013;8(3):e57380.

[3] Naviaux RK. Metabolic features of the cell danger response. Mitochondrion. 2014 May;16:7-17.

[4] Burnstock G. Pathophysiology and therapeutic potential of purinergic signaling. Pharmacol Rev. 2006 Mar;58(1):58-86.

[5] Boccuto L. et al. Decreased tryptophan metabolism in patients with autism spectrum disorders. Mol Autism. 2013 Jun 3;4(1):16.

[6] Theoharides TC. Extracellular Mitochondrial ATP, Suramin, and Autism? Clinical Therapeutics. 2013; 35: 1454-1456.

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Naviaux JC, Schuchbauer MA, Li K, Wang L, Risbrough VB, Powell SB, & Naviaux RK (2014). Reversal of autism-like behaviors and metabolism in adult mice with single-dose antipurinergic therapy. Translational psychiatry, 4 PMID: 24937094

MMP-9 and symptom severity of ADHD

"The statistical regression analysis revealed a correlation between increased serum MMP-9 levels and severity of symptoms in the ADHD". That was the sentence that caught my eye taken from the paper by Halina Kadziela-Olech and colleagues [1] (open-access here) looking at serum matrix metalloproteinase-9 (MMP-9) levels in children with hyperkinetic disorder (HKD) [2] "a severe form of a syndrome which is referred to in DSM-IV... and the American literature as attention deficit hyperactivity disorder (ADHD)".

Hamlet @ Wikipedia 

MMP-9 is something that has already been talked about on this blog with autism in mind (see here) but basically refers to a protease which is involved in degrading proteins of the extracellular matrix [3] and other duties. As Kadziela-Olech et al  point out, there is some preliminary interest in MMP-9 with reference to "neuropsychiatric disorders" particularly with schizophrenia in mind [4]. Interestingly also is that quite a bit of the conversation about MMP-9 and its potential effects have focused on the blood-brain barrier [5] under certain circumstances, which is something that I've always been more than a little interested in (see here) particularly transport across this important barrier.

Anyhow, back to the paper in question:

• Analysis was conducted on 37 boys with HKD/ADHD all properly assessed, as were cognitive functions (via the WISC-R). Blood samples were donated and MMP-9 activity assessed by ELISA (see here).
• Results: bearing in mind there was no asymptomatic control group, it's pretty pointless for me to talk about the serum levels of detected MMP-9 in cases overall. Of more relevance were the various associations made between levels of MMP-9 and presented symptoms: "The MMP-9 levels were significantly associated with symptoms severity of HKD and of ADHD".
• In particular, MMP-9 levels seemed to correlate best with the impulsivity symptom domain when it came to HKD.

In the paper discussion, the authors do go through some of the possible ways that MMP-9 might relate to ADHD/HKD. Part of those discussions focus on blood-brain barrier (BBB) permeability and whether some of the reported risk factors for ADHD may affect MMP-9 expression onwards to a sort of 'leaky' BBB (sounds very familiar). I note also mention of minocycline as a MMP-9 inhibitor (see here) which may very well tie back into some of the research work being done with autism / Fragile X syndrome in mind, particularly given the overlap being reported between diagnoses like autism and ADHD. Some light reading around the topic of MMP-9 also reveals that there may be lots of other functions/relationships to had for this protease. With my cherry-picking hat on, I note the paper by Rodrigues and colleagues [6] talking about inflammation and in particular: "MMP-9 expression in the colon causes alterations in the fecal microbiome" given my interest in all-things gut microbiomics.

Ultimately however, quite a bit more work needs to be done in this area before anyone gets too carried away. Bearing in mind the Kadziela-Olech study was a snapshot study and as I said, without relevant control groups, there are some limitations on what can be said about the findings. Looking also at the plots correlating behavioural presentation and MMP-9 levels it's quite obvious that the relationship between the two variables is not altogether straight-forward despite what the statistics might suggest.

But don't let that deflect from their findings and the need for more research in this area...

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[1] Kadziela-Olech H. et al. Serum matrix metalloproteinase-9 levels and severity of symptoms in boys with attention deficit hyperactivity disorder ADHD/hyperkinetic disorder HKD. Eur Child Adolesc Psychiatry. 2014 Mar 17.

[2] Cameron M. & Hill P. Hyperkinetic Disorder: Assessment and Treatment. Adv Psychiatr Treat. 1996; 2: 94-102.

[3] Birkedal-Hansen H. et al. Matrix metalloproteinases: a review. Crit Rev Oral Biol Med. 1993;4(2):197-250

[4] Yamamori H. et al. Plasma levels of mature brain-derived neurotrophic factor (BDNF) and matrix metalloproteinase-9 (MMP-9) in treatment-resistant schizophrenia treated with clozapine. Neurosci Lett. 2013 Nov 27;556:37-41.

[5] Li YJ. et al. Disruption of the blood-brain barrier after generalized tonic-clonic seizures correlates with cerebrospinal fluid MMP-9 levels. J Neuroinflammation. 2013 Jul 5;10:80.

[6] Rodrigues DM. et al. Matrix metalloproteinase 9 contributes to gut microbe homeostasis in a model of infectious colitis. BMC Microbiol. 2012 Jun 13;12:105.

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Kadziela-Olech H, Cichocki P, Chwiesko J, Konstantynowicz J, & Braszko JJ (2014). Serum matrix metalloproteinase-9 levels and severity of symptoms in boys with attention deficit hyperactivity disorder ADHD/hyperkinetic disorder HKD. European child & adolescent psychiatry PMID: 24633733

Matrix metalloproteinases and autism

Peptidases. The enzymes that split apart peptide bonds, have always been of some interest to me. Not only resulting from my continued interest in peptides as being one potential suggestion why a gluten- and/or casein-free diet might affect symptom presentation in some cases of autism but also because of their relationship to the building blocks of functional life, the amino acids, which are really starting to make some waves in autism research.

Just split @ Wikipedia  

With all this in mind, it was perhaps inevitable that I would arrive at a post on a specific class of peptidases, the matrix metalloproteinases (MMPs) and some very preliminary suggestions of potential involvement in cases of autism and beyond. Please note that not all peptidases work on the same types of peptides, just in case you wondered.

It was the paper by Abdallah and colleagues* which spurred me into writing this post, and their suggestion that analysis of amniotic fluid samples indicated that levels of MMP-9 were elevated in cases later diagnosed with an autism spectrum disorder (ASD). The fact that they also mentioned BDNF was a bonus interest.

Perhaps I should back up a little and provide a quick overview of the MMPs. So with my Twitter limit: zinc-dependent, embryonic development, removal of extracellular matrix, immune function and inflammation, synaptic plasticity**, etc. There are a number of very good review articles on the MMPs including this one by Birkedal-Hansen and colleagues*** (open-access).

The specific MMP discussed by Abdallah et al, MMP-9, has been the topic of quite a lot of research interest over the years. The degradation of collagen is an obvious starting point for all the MMPs, hence a link with connective tissue disorders such as rheumatoid arthritis**** (open-access). It is however with the brain and various neuropsychiatric conditions in mind, that I find myself drawn to the potential effects of MMP-9.

Domenici and colleagues***** (open-access) for example, reported on the possibility that elevated plasma MMP-9 levels may (alongside other compounds) be a biomarker for depression in their cohort. Indeed the genetics of MMP-9 have been similarly linked to bipolar and related disorders. There remains however the chicken-and-egg situation of which came first, MMP-9 or symptoms, not really answered by studies such as this one by Rybakowski and colleagues****** relying on MMP-9 levels to predict staging of bipolar disorder.

Outside of the Abdallah study, there is not presently a great wealth of research done on the MMPs with autism in mind. Accepting the fact that Abdallah was looking in amniotic fluids samples and therefore not functional levels per se, I did turn up this paper by Siller and Broadie******* (open-access) looking at MMPs in Fragile X syndrome (FXS). Regular readers might remember a few papers discussed on this blog in recent times with FXS as the target condition (see here and here) presenting with autistic features. Siller and Broadie asked whether administration of the antibiotic minocycline might actually serve to inhibit MMP (activity or formation?). The answer: it might as per other work in this area but please note that this is not to be construed as medical advice.

I was likewise interested to read the paper by Jang and colleagues******** on how everyone's favourite sleeping aid, melatonin, might also have some important effects on MMP-9 levels, at least in a rat model of stroke. That and the possibility(!) that this relationship might also involve blood-brain barrier (BBB) disruption really got the grey matter working. Once again a pharmacological effect that you won't see on the drug packaging insert.

We have yet to see all there potentially is about MMPs with autism in mind. With all the current interest in excitotoxicity in cases of autism linked to things like glutamate and compounded by the whole microglia story, I'm sure it is a topic that will gain in research popularity and perhaps even open up new avenues for intervention********* for specific people and/or groups should perturbed levels be found and replicated. Given also the possibility of a viral link to elevations in MMP9********** one has to assume that environment may also play a hand in some presentations and potentially tie into those models of maternal infection as being a risk factor for offspring autism?

To finish, the King at his best...

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* Abdallah MW. et al. Amniotic fluid MMP-9 and neurotrophins in autism spectrum disorders: an exploratory study. Autism Res. September 2012.

** Huntley GW. Synaptic circuit remodelling by matrix metalloproteinases in health and disease. Nature Reviews Neuroscience. 2012; 13: 743-757.

*** Birkedal-Hansen H. et al. Matrix metalloproteinases: a review. Crit Rev Oral Biol Med. 1993 ;4: 197-250.

**** Distler JHW. et al. The induction of matrix metalloproteinase and cytokine expression in synovial fibroblasts stimulated with immune cell microparticles. PNAS. 2005; 102: 2892-2897.

***** Domenici E. et al. Plasma protein biomarkers for depression and schizophrenia by multi analyte profiling of case-control collections. PLoS One. 2010; 5: e9166.

****** Rybakowski JK. et al. Increased serum matrix metalloproteinase-9 (MMP-9) levels in young patients during bipolar depression. J Affect Disord. August 2012.

******* Siller SS. & Broadie K. Matrix metalloproteinases and minocycline: therapeutic avenues for fragile X syndrome. Neural Plast. 2012: 124548

******** Jang JW. et al. Melatonin reduced the elevated matrix metalloproteinase-9 level in a rat photothrombotic stroke model. J Neurol Sci. 2012. pii: S0022-510X(12)00524-2

********* Leonardo CC. & Pennypacker KR. Neuroinflammation and MMPs: potential therapeutic targets in neonatal hypoxic-ischemic injury. J Neuroinflammation. 2009; 6: 13.

********** Kolb SA. et al. Matrix metalloproteinases and tissue inhibitors of metalloproteinases in viral meningitis: upregulation of MMP-9 and TIMP-1 in cerebrospinal fluid. J Neuroimmunol. 1998; 84: 143-150.

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Abdallah MW, Pearce BD, Larsen N, Greaves-Lord K, Nørgaard-Pedersen B, Hougaard DM, Mortensen EL, & Grove J (2012). Amniotic Fluid MMP-9 and Neurotrophins in Autism Spectrum Disorders: An Exploratory Study. Autism research : official journal of the International Society for Autism Research PMID: 23008271

P-glycoprotein: lock on transporters

During a post a while back, I did promise that I would discuss an interesting compound called P-glycoprotein in relation to the blood-brain barrier and its helping, transporting hand. Well here is it is. A recent article titled "P-glycoprotein: Why this drug transporter may be clinically important" attracted my attention in timely fashion discussing drug interactions which, as coincidence would have it, is something that I have also been reading about recently.

This paper by van Assema and colleagues* cropped up on my Twitter radar recently. In it, the authors describe how problems with a pretty important transporter system, P-glycoprotein (otherwise known as ATP-binding cassette sub-family B member 1, ABCB1) might put some fundamental observations in Alzheimer's disease (AD) into perspective.

I don't want to rehash the whole AD story because it has been summarised in other posts. The watchwords are: beta-amyloid peptide fragment, reactivity, and plaques. Dr/Prof. van Assema and team carried out some investigations on P-glycoprotein function in a small number of patients with AD compared with controls. They reported lower P-glycoprotein functioning and concluding that the ability to transport things out of the brain through the blood-brain barrier might be problematic at least in some cases of AD. 

This is not the first time that P-glycoprotein has been implicated in AD. This paper and its accompanying commentary hinted at similar things in a mouse model looking at transporting those pesky beta-amyloid peptides away and reducing the accumulation. I note that there is some growing interest in this and related areas following the news that AD might have some interesting spreading capability. 

As per my previous post on amyloid precursor protein, AD is not autism. Indeed, the evidence seems quite the contrary when it comes to looking at both the precursor protein and plasma levels of the nasty peptide fragment/s in relation to autism.

Having said that P-glycoprotein has been looked at in cases of autism; specifically to account for the variable effectiveness of some medications given to manage peripheral issues such as hyperactivity. This paper by McCracken and colleagues** suggested that individual responses to the drug guanfacine might be influenced quite strongly by certain genetic differences in the p-glycoprotein gene. So the presence of a certain mutation (SNP) might account for lower levels of P-glycoprotein, which in turn means less 'hold my hand' transport of the drug across the blood-brain barrier to exert an effect. 

Acknowledging that the blood-brain barrier is a highly complex barrier and that we need to tread cautiously in terms of what we allow in and out across this barrier, there is a whole new world to be explored here perhaps rivaling that other barrier of great interest in relation to autism and a few other things. 

* van Assema et al. Blood–brain barrier P-glycoprotein function in Alzheimer's disease. Brain. November 2011.

** McCracken JT. et al. Possible influence of variant of the P-glycoprotein gene (MDR1/ABCB1) on clinical response to guanfacine in children with pervasive developmental disorders and hyperactivity. Journal of Child & Adolescent Psychopharmacology. February 2010.

Even Superman had problems with lead

Whilst the Man of Steel was all but impervious to bullets and explosions, he was not without his weaknesses. Kryptonite was his primary problem, but when it came to using that X-ray vision of his, another obstacle stood in his way, lead.

Lead (Pb or plumbum) has popped up here and there in a few recent posts on this blog. It was one of the factors, the reduction of which, was put forward to partially account for the drop in crime levels in the US in this post; a factor no-one can rule out in relation to the risk of being diagnosed with ADHD in this post; and also exemplified as the archetypal behaviour-changer in this post on detox. Yes sir, lead might have quite a bit to answer for.

Lead exposure is a well-known effector of health. For children with their developing brain in particular, lead exposure is not good news and can affect both physical and cognitive development. It is perhaps with these quite important effects in mind, that lead exposure has been looked at with reference to quite a few childhood conditions, including those with a developmental aspect to them such as ADHD.

The possibility that lead exposure either pre-, peri- or post-natally might be implicated in the risk of being diagnosed with an autism spectrum condition has also been the topic of some research. Studies such as this one, this one and this one have all suggested some (varying) association between lead exposure and autism, either on the basis of treatment figures or based on biological measures of detected amounts. The literature has been joined by this recent paper suggesting chronic lead toxicity in a number of children with autism resident in Saudi Arabia.

I have blogged about the Saudi research group involved in this new study already, based on their excellent publication record this year in autism research which rivals that of the MIND Institute and Prof. Jim Adams. In this latest trial, for which I only have the abstract so far, it appears that they are signalling post-natal lead toxicity as, quote ".. could represent a causative factor in the pathogenesis of autism" on the basis of some very high levels of the lead ion being detected in plasma. Don't ask me how they made the detection because I don't have the full-text paper yet, but I would hazard a guess that based on their results they might have used our old friend ICP Mass Spectrometry or possibly via atomic absorption.

I don't want to go too overboard on this latest study yet. It is interesting to note that lead poisoning in the early years can affect various brain functions including synaptogenesis and the correct functioning of the blood-brain barrier. Both of which have been implicated, to varying degrees in cases of autism and related developmental conditions. Having said that, autism does not seem to have a monopoly on these issues outside of other conditions; whether such mechanisms could so fundamentally affect a person as to 'lead' (pardon the wordplay) to autism or autistic symptoms I don't know.

There are various potential implications from any association between lead and autism. Risk issues such as the environment, the prevalence of pica in autism and the various suggestions about the use of chelating agents to remove bioaccumulated lead all might figure in any discussions. What I would perhaps like to see initially is some guidance on ruling out lead poisoning in any and all cases of developmental diagnoses as a routine measure, bearing in mind what is known about the effects of lead and the developing brain.

Don't let me in: the blood-brain barrier

Several of the posts on this blog include the concept of biological barriers and what potentially might be the effects of problems with hyperporosity (leakiness) of such barriers. This is particularly true of the gastrointestinal barrier in relation to things like coeliac disease and inflammatory bowel disease. Autism also seems to figure to varying degrees in such discussions.

The gastrointestinal barrier may well be important for lots of different reasons; predominantly stopping things getting out - fragments of food proteins, peptides, eosinophils, etc. which could conceivably have various somatic effects (and possibly a few psychological ones too).

In this post I want to talk about a barrier which is primarily charged with stopping things getting in - the blood-brain barrier (BBB). I am not going to just recite the various descriptions of the BBB. Suffice to say that it is a partly chemical and partly physical barrier which protects perhaps our most complex organ. The scientific literature extensively discusses the BBB with reference to just about everything - structure, development, disease, etc. With regards to autism, there is also a fair amount of material to analyse.

One might assume some potential relationship between the BBB and some cases of autism given the effect of conditions such as encephalitis and meningitis on barrier function potentially connected to autism onset tied into such infections. The fact that seizures and epileptic syndromes can also affect/be affected by the barrier might also be of some interest given the presence of this fairly common co-morbidity.

Quite a bit of the early autism-BBB material is taken up on peptide-hormone chemistry and in particular the opioid-excess theory of autism. There is still a degree of speculation on this theory with reference to the BBB (not so much the gut membrane), and not everyone is as much a fan as I am of the potential for the theory, but that is perhaps fodder for another post. There are also subtle hints of immune effects to things like cows milk protein potentially tied into 'alterations in the blood-brain barrier' in autism. Although the precise mechanism has yet to be elucidated, inflammation seems to be key, and cytokines might have quite a significant role to play.

Other research has suggested a connection between aberrant amino acid transport across the BBB and autism. Levels of serum tryptophan, for example, have been suggested to be perturbed in some cases of autism with possible implications for levels available for subsequent brain 5-HT synthesis given also the enzyme kinetics linked to the presence of other amino acids.

Our old friend tetrahydrobiopterin (BH4) has also been mentioned with the BBB and autism in mind.
This is an interesting study on the possibility that genetic differences in one or more of the proteins which potentially aid transport of drugs across the BBB might (partially) account for differences in the effectiveness of some medications used for autistic symptoms. I do wonder if similar studies might also shed some light on the different experiences of medication used in autism.

What seems to be missing from the research on the BBB and autism is data on the integrity of the BBB. There are quite a few ways to measure such permeability based on the various drug transport studies around including use of Evans Blue and other contrast agents coupled with MRI.

I assume that research will eventually get round to looking further at the BBB in relation to autism given the current focus on brain structure and autism. I will perhaps come back to the BBB at a later date given its potential importance for lots of things related to autism and beyond.