At least two sides to every story

This is a blog predominantly about autism research but now and again I have been known to wander. One topic which I wander off into quite a bit is Chronic Fatigue Syndrome (CFS) / Myalgic Encephalomyelitis (ME)  as exemplified by posts like this and this. I wander into CFS/ME because, like autism, here is a spectrum of conditions asking more questions than answering them. One of the more 'heated' areas of questioning in CFS/ME has been around the various twists and turns of the XMRV story.

I'm not going to rehash the same stuff in this post but the main points focus on the almost operatic battle on the question of detecting XMRV: yes I can, no you can't. Yes we can find it in some samples from people with CFS/ME according to this article published in 2009*. No we can't, in this recent paper** which has gone hand in hand with a partial retraction and some discussions on various Internet sites which I am not going to cite in this post.

Up until a few days ago I didn't know all that much about the work coming out of the Whittemore Peterson Institute (WPI) led by Dr Judy Mikovits aside from what I read in cyberspace. I knew a little bit about the backstory: Dr Mikovits asked to leave and the continuing debate about why and what will happen to various research and grants. I know a little bit more now after seeing Dr Mikovits present some of her data and answer some of the questions about this whole story and where it might go next. As in the title of this post, there are always at least two sides to every story. Let me elaborate.

It was a fresh clear Autumnal Friday evening when Dr Mikovits made a presentation at the Education Centre of Sunderland Royal Hospital at the behest of the Sunderland & South Tyneside ME/CFS Support Group. The first impression I got about Dr Mikovits is that she knows her stuff. She is no stranger to research after all with a quite impressive publication record spanning her varied career, covering areas of HIV, product formulation and drug discovery and cancer research. Her work in HIV and retroviruses potentially tied into DNA methylation is perhaps most interesting alongside the now pretty well established link between some viruses and some cancers.

Her presenting style was slick, quick and pretty technical, going through the various stages of the XMRV story and some elaboration of the saga behind the headlines. A few points that I managed to jot down from her presentation:

• Whilst ME/CFS is a focus for the current virology work, she is interested in various other conditions under the umbrella of 'neuro-immune' which potentially share some overlapping features such as cytokine and chemokine abnormalities, inflammation, allergy and intolerances and oxidative stress. Epigenetics (changes in gene expression not involving alterations to the genetic code) was also mentioned. You guessed it; that also includes autism.
• We were given a crash course in human gammaretroviruses (HGRVs). Part of that focused on how these viruses might be involved in all manner of things including potentially interacting with other viruses. One or two interesting primate studies were also discussed including this one by Onlamoon and colleagues*** on disappearing viral and proviral signals in blood after a month of XMRV infection (latency and activation). Work by Makarova and colleagues**** on the low immunogenicity of the virus was also detailed.
• There was some suggestion of a signature 'footprint' cytokine and chemokine profile present in cases of CFS/ME positive for XMRV. This included: IL-8, IL-13, MIP-1alpha and MIP-1beta, TNF-alpha, IL-7, INF-alpha, IL-6, and GM-CSF. I can't readily verify this compound set, the numbers of participants it was based on and whether all were increased or decreased. I would imagine 'increased' given the functions of many being in response to viruses (and bacteria and parasites).
• Mention was also made of nagalase activity being elevated. I remember reading recently about some similar findings of nagalase being reported in cases of autism (here) albeit not formally published on. I'm not going too much further into this area because it is frighteningly outside of my competence and indeed takes CFS/ME and autism into some very unusual territory.
• The 'contamination' debate which is being widely discussed was also presented on. I can't claim to understand everything but I have the words 'plasmid contamination' and 'junction primer' scribbled in my notes if that makes sense to anyone.

The general impression I got, as a non-virologist, about the detection of XMRV and related viruses (as with any virus) is that it is complicated. It's not like saying we can detect this virus or other viruses like we can measure blood glucose levels for example. You need to avoid contamination (as we have seen recently in other areas of forensic science). You need to make sure your are using the right targets to detect the virus. You also need to think about where the virus is and how 'active' it might be (or not) and onwards whether you need to 'provoke' the activation of a virus ('provocation' can be construed in many different ways). In short it is a complicated procedure; much more complicated than I originally thought.

Dr Mikovits accepted that XMRV is probably not going to be the 'be all and end all' of CFS/ME as per the partial retraction of the Science paper. She did however stick to her guns about the potential involvement of other gammaretroviruses as perhaps a source of further investigation.

I don't claim to be an expert in this area of investigation (and speculation). Neither do I offer any view on who is right or wrong in this complicated area. What I do perhaps want to see more of is continued research done in this area as suggested by Lo and colleagues***** and their suggestions of a related polytropic family of viruses, related but not necessarily XMRV, which might be applicable to many more conditions outside of just ME/CFS. Just because the technology and methods might not be suitably honed to detect these complicated viruses, does not mean that they may not be there. The chicken and egg question is what came first: CFS/ME or virus?

As we have seen recently in Alzheimer's disease, there is a lot not known about the viruses we share our little rock with and how we interact with them and them with us.

P.S. I am only reporting what I heard. Don't shoot the messenger.

* Lombardi VC. et al. Detection of an infectious retrovirus, XMRV, in blood cells of patients with chronic fatigue syndrome. Science. 2009. 326: 585-589.

** Simmons G. et al. Failure to confirm XMRV/MLVs in the blood of patients with chronic fatigue syndrome: a multi-laboratory study. Science. 2011. September 2011.

*** Onlamoon N. et al. Infection, viral dissemination, and antibody responses of rhesus macaques exposed to the human gammaretrovirus XMRV. J. Virol. 2011. 85: 4547-57.

**** Makarova N. et al. Antibody responses against xenotropic murine leukemia virus-related virus envelope in a murine model. PLoS ONE. 2011. 6: e18272.

***** Lo SC. et al. Detection of MLV-related virus gene sequences in blood of patients with chronic fatigue syndrome and healthy blood donors. PNAS. 2010. 107: 15874-9.

Caspase the executioner

The very dramatic title to this post pays homage to the caspases and their role in the process of programmed cell death (apoptosis). Their 'executioner' label denotes their role in the cascade effect that ends the life of a cell. If, like me, you watch too many films and documentaries about Medieval England (don't we all?), you might picture the executioner as a hangman, face covered, being paid to go about his duties. Grim stuff indeed.

Enough of all that. Like many things connected to our very complicated human biology, the protease family known as the caspases are also involved in other processes outside of just cell death. One of these processes is inflammation and partially mediating inflammatory functions such as those linked to autoimmunity.

A recent paper by Siniscalco and colleagues* sheds some light on a possible role for the caspases in connection to autism spectrum conditions. There was always going to be some interest for me in this paper given the authorship group. When names like Laura de Magistris (she of the leaky gut) and Alessio Fasano appear, I tend to sit up and take note. Please don't take this as any kind of idol-worshipping or anything like that; merely some admiration for their collected works; sort of winners of an X-Factor for scientists (now there's a novel idea).

Anyway, I mentioned that the caspases are a group of enzymes with quite a few members. Within the family, various caspases have various different duties and show various different effects when things aren't quite as they should be. The crux of the current paper was to demonstrate whether caspase expression (mRNA and protein levels) were different in a small group of children diagnosed with autism compared with controls.

The answer bearing in mind the ever-so-small numbers included for study was yes, levels were different; and different across a few of the caspases. The messenger RNA (mRNA) levels were increased for caspases-1, -2, -4. -5. Caspases-1, -4, and -5 are important because of their connection with inflammation. Important also because the caspases need to work synergistically together in order to have their 'full activity'. When protein levels of the caspases were measured, caspases-3, -7, -12 were found to be elevated in the autistic group. Caspase-3, when activated, for example seems to play a role in neurodegenerative conditions such as Alzheimer's disease (AD) and its connection with the amyloid beta precursor protein (APP). The effect is described as an enhancing one, in that caspase-3 is in the thick of all those tangles and plaques noted in AD. Caspase-12 takes us back to inflammation and caspase-7 back to apoptosis.

Cumulatively, the various caspases examined and reported on, seem to imply inflammation and a role for the immune system in the cases studied. I am a bit baffled by the caspase-3 finding in light of the connection to AD in other work, which perhaps contrasts with what is known about APP and those wretched peptides in relation to autism. Having said that, I do realise that AD is a very complicated condition and the presence of caspase-3 may reflect other processes. So how about a possible link between caspase-3 and type-1 diabetes instead?

This is not the first time that the caspase family have appeared on the autism research landscape. Mady Hornig and colleagues (she of the recent carbohydrate metabolism paper) discussed caspase-1 in relation to apoptosis and neurodevelopmental damage in this paper from a few years back. This paper by Sheikh and colleagues reported caspase-3 as being a more direct marker of apoptosis being elevated in the cerebellum portion of the brains of people with autism. Cerebellum, autism, Eric Courchesne?

The caspases are an interesting family of enzymes, of that there is no doubt. Further large-scale trials are needed to confirm findings and subsequently to start thinking about when, how and why the caspase findings become relevant to autism and what can be done to modify any detrimental effects.

To forget all this talk about executioners, how about Huey and some News?

Siniscalco D. et al. The expression of caspases is enhanced in peripheral blood mononuclear cells of autism spectrum disorder patients. JADD. October 2011.

Vitamins and minerals: lost in translation

Vitamin D, vitamin C and the odd probiotic. That's the sum total of my daily vitamin/bacterial supplementation routine as a nearly-middle aged, borderline healthy man. Why just the three supplements? Well my diet is pretty varied, but I do live pretty far North in a place where sunshine is often in short supply particularly in the Winter months hence the vitamin D supplement just in case. I also want to keep supple and probably don't eat as much fruit as I should hence the vitamin C. My gut is my Temple and just to make sure that it stays like a Temple (quiet and serene) I take the probiotic. I take a relatively small dose of each at particular times every day (chronotherapeutics y'know) and whilst a very subjective observation, can often tell when I've 'gone without' for a while. I can hear the whispers already... hasn't he ever heard of the placebo effect? Er, yes.

Like many millions of other people around the world, vitamin (and mineral) supplementation is part of my daily dietary routine. Why did I start taking them? Well... erm, 'the research told me to Sir'. There is for example some evidence that vitamin D deficiency and supplementation could be associated with lots of different conditions and perhaps even more so for certain groups of people. Same for vitamin C. As for the probiotics, well with my borderline obsession with all things gut bacteria on this and my other blogs should be explanation enough. [Note: I'm not in any way, shape or form advocating any vitamin or mineral or probiotic use. If you need more information, speak to your healthcare provider].

Having said all this I am a firm believer in horses for courses and realise that my supplementation routine might not be for everyone, particularly for those sun-kissed, 5+ fruit and vegetable a day, healthy people out there. So it is with some recent publications which have not been easy on some vitamins and other nutraceutical supplements.  This paper by Mursu and colleagues* has received a lot (a lot!) of media coverage with some pretty sensational headlines like 'Are your vitamins killing you?' and 'We've been wasting a ton of money on vitamins and dietary supplements'. Of course when you look at what the study results actually report, you get a little more of a grounded idea of the findings which basically say that based on self-reported supplement use, there was a variably increased risk of mortality in older women (mean age: 60 years+) when supplementing with either a multi-vitamin or a few individual vitamins and minerals including iron, folic acid and copper. Without wishing to belittle what mortality actually means, some the risk was varied; based on the odds ratios detailed and what they actually translate into in terms of hazard. Some of the risk is described as dose-dependent. In other words, take more and your risk might be increased. Another detail of the study on the potentially 'positive' effects of calcium supplementation on reducing mortality for this age proband seems to have been lost in translation.

Now don't get me wrong, I am all for proper testing for safety and efficacy for everything from pharmaceutics to vitamins and minerals to probiotics even to dietary changes; everything, period. If you put it into your body, you should have the comfort of knowing that it does what it says on the box and does it in a safe a manner as possible including any details on other 'drug' interactions or contra-indications. I do however look at the press that this study seems to have received in some quarters and get the feeling that the findings are being somewhat 'beefed-up' from the original results to suit various purposes. Just me perhaps being a little neurotic.

A second study has emerged which again paints a slightly less rosy picture of the value of supplementation. Findings reported by Clarke and colleagues** suggest that some of the proposed health benefits of broccoli, the isothiocyanates, tend to be less pronounced if taken via supplement rather than the real thing as a function of their lacking an enzyme needed to metabolise them properly normally included in the vegetable.

I am actually much happier with the Clarke study; the way it was carried out and reported on (so far). Basically what it says is that Mother Nature knows what she is doing with our vegetables, fruits, etc. and whilst we all think we are skilled in the science of health and the production of 'synthetic health', we perhaps don't yet know as much as she does about the whole being greater than the sum of its parts. I can imagine a few in the medical and dietetics trades as well as the odd pharmacognosist who will see this as evidence for the message about vitamins and minerals being derived from food not supplements as being all important. That being said I, myself, am unaware of anyone who is regularly taking a broccoli supplement for anything.

I have touched upon the themes of 'association' and 'risk' in other posts on this blog. Association or correlation not being the same as causation works both ways in terms of positive and negative suggested connections. Certainly the study by Mursu and colleagues whilst interesting is no different to any other case. I'm not even going to get started on the self-report angle to the study and the potential pitfalls associated with recollection.

I suppose it all boils down to a few messages:

• treat your vitamins the same way you would your 'regular' medications (dose, side-effects, etc.),
• if you can get your vitamins and minerals from food, do so,
• always question any health claim and whether there are indeed published replicated effects (see this recent Nature blogpost),
• be proactive: research, research, research - don't take anything at face value (particularly sound-bites).
• the hazards of hazard ratios.

[Just in case you need to know, I have no conflicts of interests when writing this post. I don't have my own line of vitamins or anything, don't have any shares in vitamin companies and I am not knowingly on any council or other body with a vitamin promoting agenda].

* Mursu J. et al. Dietary supplements and mortality rate in older women. Archives of Internal Medicine. October 2011.

** Clarke JD. et al. Comparison of isothiocyanate metabolite levels and histone deacetylase activity in human subjects consuming broccoli sprouts or Broccoli Supplement. Journal of Agricultural & Food Chemistry. September 2011.

Progranulin and autism

I have forgotten the number of times that autoimmunity has been discussed in connection to autism spectrum conditions on this blog. I wouldn't say that I am obsessed with the topic; just quietly interested in how the lack of recognition of self, immunologically-speaking, seems to crop up time and time again in at least some cases of autism alongside the subsequent link to inflammation. I offer no opinion on the implications of such findings, and how 'universal' they might be to the autistic spectrum, but they certainly make for some interesting reading. Just in case you need some more discussion on this topic I present this paper (full-text) by Al-Ayadhi and Mostafa* on the the findings of low plasma progranulin levels in children with autism.

As previously mentioned, the amount of research coming out of the Saudi Arabia on autism has been thick and fast this year with some pretty novel associations being suggested. I note that they have even started to look at that most unusually named of compounds, Sonic Hedgehog Protein in relation to oxidative stress (here); fodder for a separate post methinks. The current paper on progranulin levels continues their research theme on the immune system and inflammation where previous observations have linked things like anti-ganglioside M1 auto-antibodies to some cases of autism.

A little bit of description might be useful to start with on what exactly progranulin is and why it might be important. The technical stuff about progranulin is here, but in as few words as possible: the granulins are a family of leukocyte polypeptides, some of which have cell growth modulatory activity, hence their suggested role in lots of things from wound healing to the creation of cancers. This paper suggested that levels of the source protein progranulin, might also serve as a marker for some of the components of metabolic syndrome but that's all I will say on that particular issue for now.

For the purpose of the current study, progranulin was examined in the context of being an anti-inflammatory neurotrophic factor with neuroprotective qualities. A few other details about the study and findings:

• Forty children with autism participated in the trial, all medication-free. Forty control children were also used as comparators matched for age and gender and free of any immunological condition.
• CARS was used to assess the 'level' of autistic behaviours in the experimental group; where levels of plasma progranulin were assessed in all participants.
• Children with autism had significantly lower levels of plasma progranulin (p=0.001) although there did not appear to be any link with symptom severity as judged by CARS scores, age or gender. Sixty-five percent of children with autism had reduced progranulin levels. It is interesting to note the surprising degree of 'homogeneity' of results from the autistic group compared with controls in terms of the dispersion of individual measured levels. 

These are interesting results allowing for the small participant group examined. The authors discuss the possible relevance of their findings and that too makes for some interesting reading particular when you would expect progranulin levels to be elevated if inflammation was present as some of the literature seems to indicate inflammation is in cases of autism. Does this mean that inflammation might not be present? I don't know but then an awful lot of other results must be wrong. Does this mean that genetically there might be 'glitches' in the pathways required to make progranulin in autism in response to inflammation? I don't know either but no data on this measure seems to exist at the moment. Looking at one parameter alone and in isolation does not tend to provide a very complete picture hence the need for further investigations.

As per the authors comments on the current paper, much more research is required in what is a novel area of investigation. Knockout mice model studies, where the gene for progranulin located on chromosome 17q21 (ring any bells?) is absent, have suggested some male-dependent behaviours potentially linked to the serotonergic system, bearing in mind mice are not necessarily the same as people. A small (very small) suggestion that some cases of schizophrenia might also be linked to mutations in the progranulin gene should also be noted. The Saudi authors discuss the possibility of progranulin therapy in cases of autism but I would be hesitant at best to support such a claim at the moment without lots and lots and lots more research being done on safety and the appropriate dosing.

* Al-Ayadhi LY & Mostafa GA. Low plasma progranulin levels in children with autism. Journal of Neuroinflammation. September 2011.

Toxoplasma gondii in the crosshairs

Source: PsychCentral

Today, Monday 10th October 2011, is World Mental Health Day. It's all about raising awareness about mental health issues. It is quite timely that this years theme is 'Investing in mental health' on the back of the quite startling figures recently published about the numbers and cost (personal and financial) of mental ill-health and the relative lack of funding and research on what causes it and what to do about it.

Anyway, I commerate the day with this post on schizophrena and an old friend.

This is not the first time that the topic of Toxoplasma gondii has graced this blog and probably won't be the last time either. What is perhaps so fascinating about this parasitic protozoa is its single-minded drive to survive (and replicate) and, as a result, its ability to affect animal behaviour by whatever mechanism.

Fine if you are a rodent I hear you cry. But when it comes to human beings, much of the recent chatter about T.gondii has been on its speculated links to conditions like psychosis, mood disorders and, in extreme cases, suicide. Presentation of schizophrenia spectrum conditions and the risks associated with T.gondii infection have also figured fairly prominently in the various discussions as a result of studies like this one and this one from Faith Dickerson and colleagues. A recent study by Pedersen and colleagues* published in the American Journal of Psychiatry has confirmed an association between T.gondii infection and risk of schizophrenia. Association is one thing; causation is another. I note that Dr Emily Deans over at Evolutionary Psychiatry has just posted about this study also.

The facts and figures:

• IgG-antibodies specific to T.gondii were measured in over 45,000 women in Denmark giving birth between 1992 and 1995.
• Women were followed up until 2008 for the presence of a schizophrenia spectrum condition.
• A positive association between T.gondii antibody level and risk of schizophrenia was found; with risk varying according to the strength of the antibody response. The overall risk was 1.68, although where antibody levels were highest, that rose to 1.73.

The conclusions from the study was that where antibody levels to T.gondii were highest so the risk of schizophrenia was highest.

There are some interesting points to take from the cumulative research on T.gondii. Not everyone who is infected with this little scoundrel will go on to develop schizophrenia or other conditions. Obviously there are other factors also at work. The immune system is the first place to look; how our bodies handle such infections, viruses, bacteria and how genes and environment might interact either in a protective or facilitative fashion. I touched upon this briefly with my post on the C4B null allele in relation to autism and a few other conditions. I note also this recent publication from Whitmarsh and colleagues** published in the journal Cell Host & Microbe which suggested that knockout mice (is this the right term?) might more frequently succumb to T.gondii infection when specific cytokine signalling molecules are absent; one factor probably among many. It is not as straight-forward as to say this molecule does this and this one does this; more likely a combination of multiple factors coupled with a cascade effect confer protection or not.

So there you have it, more evidence for the influencing factor of environment on our behaviour. Makes you wonder whether we should be looking more closely at some of the treatments for T.gondii and any potential onward effects for psychological symptoms? Or do we already have our treatment measures?

* Pedersen MG. et al. Toxoplasma infection and later development of schizophrenia in mothers. Am J Psychiatry. August 2011.
** Whitmarsh RJ. et al. A critical role for SOCS3 in innate resistance to Toxoplasma gondii. Cell Host & Microbe. September 2011.

Roger Moore's eyebrows, ADHD and coeliac disease

The 'overlap' between somatic conditions and those with a more behavioural or psychological set of characteristics is of great interest to many. When such associations become apparent, and bearing in mind the old mantra 'correlation does not imply causation', it offers a potentially unique window on how body and brain are linked and how affecting one system may affect the other. A previous post discussing the beta-blocker--autism study kinda hinted at the common thread on this blog: a potential anti-hypertensive effect might impact on cognitive organisation/ability (with the caveats that I am not in anyway condoning the use of any pharmaceutic for anything or offering medical advice).

I was therefore suitably interested when this study by Niederhofer (full-text)* cropped up on the PubMed radar on the possible overlap between ADHD-like symptomatology and an over-representation of coeliac (celiac) disease (CD); indeed also what happened to behavioural symptoms when a gluten-free diet was installed. The crux of the study was that in the population studied presenting with ADHD-type symptoms according to Hypescheme (n=67), fifteen percent showed markers for coeliac disease. When I say markers, I am talking about serological testing for anti-gliadin and anti-endomysial antibodies, of which there is still some debate about the best way to diagnose coeliac disease (see NICE guidelines here). Compared with the estimated general population rates of CD, anywhere from 0.1% - 2%+, the 14.9% presented in this study is Roger Moore as James Bond eyebrow lifting (I can't actually believe that there is a Facebook group for this) bearing in mind the small participant group included in the current study. Sir Roger might also lift an eyebrow at the results of the gluten-free trial undertaken following CD diagnosis, where behavioural manifestations seemed to show an improvement when on diet.

Despite the easy-to-pick flaws in the experimental design used, these results comes as little surprise. In previous posts I talked about the possible relationship between coeliac disease and autism and case studies like this one from Stephen Genuis which suggested that adopting a gluten-free diet might have behavioural connotations as well as physical ones, at least for some. The related research from Lidy Pelsser and colleagues on diet and ADHD might also tie in; although one has to caution about the lack of data on the presence of CD in the Pelsser cohort.

What is perhaps more important is what a gluten-free diet might actually be affecting in terms of behaviour. I tried (tried!) to make a case for gluten affecting the attention and hyperactivity comorbid aspects of autism in this post which is perhaps not a million miles away from what the current study is (I think) saying. The logical step in this argument is that any effect from diet in cases of autism might be mediated through such behavioural parameters; so affecting peripheral aspects to the condition potentially might impact on core aspects and the triad, sorry, dyad, of presentation. This is perhaps a theme that needs to be explored with greater assiduity throughout autism intervention research: what is intervention actually targeting?

Outside of such speculations, this study confirms one thing: ADHD like many other developmental conditions is complex and not necessarily rooted entirely in the old noggin. There may be a requirement to screen for coeliac disease when a diagnosis of ADHD is given... too much, yes/no?

To finish, and in tribute to the best James Bond ever (Connery was good, but Moore edged it), nobody does it better... or if you prefer, the Me First and the Gimme Gimme version.. [raise eyebrow and look to camera making some witty remark like 'I think he got the point'].

* Niederhofer H. Association of attention-deficit/hyperactivity disorder and celiac disease: a brief report. Prim Care Companion CNS Disord. 2011: 13(3)