Endothelial Dysfunction - Oxidative Stress, Inflammation and BH4

This post is rather out of sequence, but it draws together several different topics that I have been investigating and introduces another chemical often mentioned in autism research, BH4.  The factor that links them all together is something called Endothelial Dysfunction.

This blog has already established that oxidative stress and neuroinflammation are the key drivers behind autism.  It has not been clear whether the oxidative stress causes the inflammation, vice versa, or perhaps they are self perpetuating.

In my ongoing investigation into the autism comorbidities of asthma and high cholesterol (proxy for cardiovascular disease), I have come across some tantalising fact, such as:-

• Asthma research shows that cigarette smoking gives you oxidative stress and this continues even after stopping smoking.  The oxidative stress reduces the effectiveness of asthma drugs.
• Oxidative stress is a key factor in cardiovascular disease.

• BH4 also known as Tetrahydrobiopterin, THB, trade name Kuvan or sapropterin is an enzymatic cofactor that is the subject of lots of research in cardiovascular disease and even in autism research.  As a drug, BH4 is so expensive that many national health services and insurers will not pay for it.  A pack of 30 pills of 100 mg cost $900.  The typical dose is 10-20mg per kg per day.  The cost per patient was reported to be over $100,000 per year.

The BH4 drug, Kuvan, is used to treat an extremely rare, but debilitating condition called Phenylketonuria, when the body cannot produce its own BH4.  In the UK, the National Health Service will not pay for Kuvan for the very small number of people who suffer Phenylketonuria.


A very interesting chart from the American Heart Association

The following chart was meant for cardiologists, but is extremely relevant if you want to understand autism.  I highlighted the parts in yellow.

In autism, people suffer from oxidative stress and typically exhibit hypercholesterolemia (high cholesterol).  They also suffer from neuroinflammation in the brain.  In my therapy, I use NAC to reduce oxidative stress and the behavioral impact is very marked.  In some autism trials they have given BH4 and the result have been visible, but not dramatic.  From the graphic above it would seem that BH4 is an extremely expensive way to reduce oxidative stress.

The latest BH4 autism trial was funded by the drug producer with 20 mg/kg/day.
 

Tetrahydrobiopterin as a Treatment for Autism Spectrum  Disorders: A Double-Blind, Placebo-Controlled Trial

More interesting for me is to now look into Endothelial Dysfunction and see if that is occurring in autism.

A Cytokine Storm? Mr Spock

I have recently started learning the workings of the human immune system, while 12 year old Ted (“normal” except for a Star Wars obsession) has been discovering Star Trek.  Last weekend we went to the cinema with Adrian “Mole” to see the latest release.  Mr Spock made one interesting observation, regarding what can happen when the interests of the many outweigh the interests of the few; this will be the tittle of a forthcoming post about the fate of Dr Wakefield and his vaccine theory.

Cytokines

Cytokines really do exist, even though they sound like something from science fiction.  They are signalling molecules associated with inflammation.  Several inflammatory cytokines are induced by oxidative stress.  The fact that cytokines themselves trigger the release of other cytokines and also lead to increased oxidant stress, makes them important in chronic inflammation.  In extreme cases, there is a downward spiral of inflammation making it worse and worse.  The Spanish Flue in 1918 and SARS in 2003 are given as examples of such deadly cytokine storms.

The Research

There is a vast amount of research about the role of cytokines in autism and some very good work has been done by Paul Ashwood.  Finally, I have found an Englishman, even though he has gone to live in California, publishing some really high quality and useful research.  It turns out he is a colleague of Dr Wakefield.  Much of Paul Ashwood’s research is not available for free.  This one is:-  The role of immune dysfunction in the pathophysiology of autism

This paper is very readable and shows how a dysfunction of the immune system is without doubt a major part of the autism story. In typical post-Wakefield fashion, nobody wants to stick their necks out and draw usable, if only hypothetical, conclusions; it is easier to just suggest further research.

All the research shows high levels of cytokines in autistic subjects in the brain, spinal fluid, blood and in the gut.  Recent research also shows high levels of cytokines in the siblings of autistic people:- Plasma cytokine profiling insibling pairs discordant for autism spectrum disorder

The researchers comment:-

Thus, the lack of significant differences between sibling pairs discordant for ASD found in our study is in line with the results of previous studies. It is possible that a common immunogenetic background shared by siblings might eventually lead to different clinical outcomes when an environmental stress (for example, prenatal exposure to environmental toxins, viral and bacterial infections, parental microchimerism, etc.) occurs during development.

This last finding was deftly understood by 12 year old Ted, who commented, “Well Dad, you nearly had two autistic children”

Well isn’t he a chip off the old block.

Peter Interpretation

So combining this knowledge with my other readings, drew me to the logical conclusion that the inherited immune dysfunction, combined with the oxidative shock, so well described by Chauhan et al,(in the 400 page book) most likely resulted in a cytokine storm that damaged the brain, and autism resulted.  Due to the feedback loop of the cytokines, the neuroinflammation continues for life.

This then led me to research cytokine storms, to see how the cycle could be stopped and some kind of homeostasis reinstated.  I did not expect to find an answer, but I did.   

First we have to introduce new terms, TNF and TNFR.

TumorNecrosis Factors (TNF)

Tumor necrosis factors (or the TNF family) refer to a group of cytokines whose family can cause cell death or apoptosis.  19 members of the TNF family have so far been identified; the one that caught my eye was OX40L, a cytokine that co-stimulates T cell proliferation and cytokine production.

A tumor necrosis factor receptor (TNFR), or death receptor, is a cytokine receptor that binds TNFs.  The matching TNFR for the TNF OX40L is called OX40 (also known as CD134).

OX40 binds to receptors on T-cells, preventing them from dying and subsequently increasing cytokine production. OX40 has a critical role in the maintenance of an immune response beyond the first few days and onwards to a memory response due to its ability to enhance survival. OX40 also plays a crucial role in both Th1 and Th2 mediated reactions in vivo. T helper cells (type 1 and 2) are white blood cells that play a major role in the immune system
OX40 has been implicated in cytokine storms.

Cause of the Cytokine Storm

When the immune system is fighting pathogens, cytokines signal immune cells such as T-cells and macrophages to travel to the site of infection. In addition, cytokines activate those cells, stimulating them to produce more cytokines.  Normally, this feedback loop is kept in check by the body. However, in some instances, the reaction becomes uncontrolled, and too many immune cells are activated in a single place. The precise reason for this is not entirely understood but may be caused by an exaggerated response when the immune system encounters a new and highly pathogenic invader. Cytokine storms have potential to do significant damage to body tissues and organs.

TNF inhibitors and Cytokine Storms

The cytokine storm is kept going by the TNF cytokines.  So if these cytokines could be inhibited the storm might abate. An existing medication developed for arthritis called a TNF-alpha blocker was proposed as a possible drug. Corticosteroids and NSAIDS (Non-steroidal anti-inflammatory drugs) have been found ineffective.

In 2003 researchers at Imperial College demonstrated the possibility of preventing a cytokine storm by inhibiting or disabling T-cell response. A few days after T cells are activated, they produce OX40, a "survival signal" that keeps activated T-cells working at the site of inflammation during infection with influenza or other pathogens. OX40 binds to receptors on T-cells, preventing them from dying and subsequently increasing cytokine production. A combined protein, OX40- immunoglobulin (OX40-Ig), a human-made fusion protein, prevents OX40 from reaching the T-cell receptors, thus reducing the T-cell response. Experiments in mice have demonstrated that OX40-Ig can reduce the symptoms associated with an immune overreaction while allowing the immune system to fight off the virus successfully. By blocking the OX40 receptor on T-cells, researchers were able to prevent the development of the most serious flu symptoms in these experimental mice.  Sadly, it appears this discovery has been abandoned by the small company that tried to develop it.

And now for the shock …

In 2009 researchers in China found that a statin induced down-regulation of OX40 and OX40L in a concentration-dependent manner.

Simvastatin reduces OX40 and OX40 ligand expression in human peripheral blood mononuclear cells and in patients with atherosclerotic cerebral infarction.

"These findings improve our understanding of the anti-inflammatory and immunomodulatory properties of simvastatin"

Antioxidants have been successfully trialled in cases of Acute Respiratory Distress Syndrome (ARDS), which is another example of cytokine storm.  Organ damage was reduced and there was an improved survival rate.

Conclusion

It would seem that the combination of antioxidant and statin is about as good a combination as is currently possible, to dampen down the remaining effects of a cytokine storm, which is the extreme case of neuroinflammation.

By skill, or luck, this combination is exactly what I am trialling with Monty.

   

By Jupiter! - Satins Part 3

Makes more sense if you have read:-
Statins Part 1
Statins Part 2

 

For most of you Jupiter is the fifth planet from the Sun, or maybe the largest planet in the Solar System.  To a young boy like Monty, Jupiter is a red fire engine, normally driven by Fireman Sam.

If you are a cardiologist you will have heard of the JUPITER trial. (Justification for the Use of Statins in Primary Prevention: An Intervention Trial Evaluating Rosuvastatin trial)

It was a huge study looking into the possible benefits of giving statins to older people with low cholesterol.  All the 17,802 subjects had elevated levels of high-sensitivity C-reactive protein (CRP) levels, which is a marker for cardiac (and neuro) inflammation.  Half were given 20mg of a statin and the other half had a placebo.  The study measured their cholesterol, CRP levels and whether they later had a cardiac incident.  The group with the statin lowered their already okay LDL and triglycerides level and also lowered their CRP level by a thumping 37%.

At the time of study termination (median follow up, 1.9 years; maximal follow-up, 5.0 years), 142 first major cardiovascular events had occurred in the statin group, as compared with 251 in the placebo group.

This was interpreted by the authors as evidence that even older people without elevated cholesterol could benefit from statins to reduce their risk of cardiovascular events.

 

 

JUPITER and autism

What JUPITER tells me is that statins were highly effective at reducing inflammation as measured by CRP.

 

Autism and CRP

Now we just need some data on the level of CRP in Autism.  Thanks to those nice people in Iran we have a study called: - The complementary role of high sensitivity C-reactive protein in the diagnosis and severity assessment of autism.

 They concluded:-

► Inflammatory process can play key role in the pathophysiology of autism.
► Higher levels of hs-CRP are detected in autistic children.

► A correlation exists between hs-CRP level and autism severity.

► Hs-CRP can be considered a complementary diagnostic test for autism.
►These findings affirm the role of inflammation in autism.

I guess because Iran is public enemy number two, nobody took much note of this study, except Paul Whiteley of course.

 
 

Autism & Statins

So it looks pretty likely that statins will reduce CRP in autistic subjects and if statins can do this, they will reduce both the neuroinflammation and, by inference, the severity of autistic behaviours.

 

Peter Research

While in the Astra Zeneca-funded JUPITER study there were 17,802 subjects and five years of research; here in the Peter Research Institute we have one subject and one week of research.

As with my Bumetanide research, I am shocked by the almost immediate effect of the drug.  In terms of lowering cholesterol, statins are supposed to take two weeks to reach full effect.  In terms of reducing neuroinflammation the effect appears to be much faster – very encouraging but, to be honest, quite unexpected.  

Back to Cholesterol & Autism

The important thing is that statins appear to reduce autistic behaviours, at least in my subject; it would however also be nice to fully understand why.  The research shows the presence of dyslipidemia (abnormal amounts of lipids) in boys with autism.

 

Alterations in lipid profile of autistic boys: a case control study

The findings were: - LDL normal, HDL low, Triglycerides high, Total cholesterol normal.  The current benchmark used is that Total Cholesterol divided by HDL should be less than 4.5.  With low HDL and high triglycerides, this could put many autistic subjects in the zone of elevated risk.

Also, be aware of the very rare condition called Smith-Lemli-Opitz Syndrome (SLOS), caused by low levels of cholesterol;  it is explained in this open-access paper:-

Abnormalities of Cholesterol Metabolism in Autism Spectrum Disorders

In one of the studies I read that CRP always drops before the fall in cholesterol.  This would imply that in the case of ASD, the cholesterol issue is just a consequence; it is the precursors that actually matter.  At least to me, that makes a lot of sense.



In case you missed the prequels:

Statins Part 1
Statins Part 2


and now there is Part 4  http://epiphanyasd.blogspot.com/2013/05/tapas-time-statins-part-4.html



 

Statins for Neuroprotection in Autism - Part 2

I suggest you start by reading Part 1.  Click here for Part 1



Choice of Statin
 

Some statins are soluble in fats/lipids (lipophilic) and some are more soluble in water.  In order to cross the blood brain barrier (BBB) to reach the cerebellum and the Purkinje Cell Layer (PCL) a lipophilic statin will be required.  There is a choice of three: - atorvastatin, lovastatin, and simvastatin.  These are also among the most commonly prescribed for cholesterol reduction and so are widely available and inexpensive.

I chose atorvastatin.  Some statins are derived from fungi, but atorvastatin is synthetic.  Lovastatin and simvastatin are pro-drugs, whereas atorvastatin is already in an active form straight out of the box. Absorption of atorvastatin decreases when taken with food.  Due to its long half-life, atorvastatin can be administered at any time of day.

Atorvastatin is approved for use in children as young as 10 and in the US is prescribed to children as young as 5.

Atorvastatin, originally made by Pfizer under name Lipitor, is the best-selling drug in the history of the pharmaceutical industry.  It came off patent recently and so the price has collapsed to a very reasonable level.

In some countries the low dose forms are available over the counter, without a prescription.

 

More Related Research

The research effort into degenerative conditions like Alzheimer’s disease (AD) is far more prolific than into autism.  The closest research to my hypothesis that statins will “perk up the Purkinje cells” is this study:-

Atorvastatin and pitavastatin protect cerebellar Purkinje cells in AD model mice and preserve the cytokines MCP-1 and TNF-α.

  

Fragile X syndrome

Fragile X syndrome is a genetic syndrome that leads to autistic behaviours.  About 5% of the cases defined as autism are due to this genetic flaw.  It also results in certain physical differences, namely:-

• Large, protruding ears (one or both)
• Long face (vertical maxillary excess)
• High-arched palate (related to the above)
• Hyper extensible finger joints
• Hyper extensible ('Double-jointed') thumbs
• Flat feet
• Soft skin
• Hypotonia (low muscle tone)
• single palm crease (crease goes across entire palm)

 At MIT researchers have found that the statin Lovastatin “can correct Fragile X syndrome”.

 

I presume what is actually happening, is that in Fragile X there is also neuroinflammation and this has been reduced by the statin, rather than correcting the syndrome.

  

Retts Syndrome

Retts syndrome is another genetic disorder that causes regression and autism-like behaviours.  It affects mainly girls, because male fetuses with the disorder rarely survive to term.  The prognosis is not good.

Research is underway with statins and currently shows that statins improve symptoms of Rett syndrome in mice.

 

Statins and depression

A large study of patients with heart disease examined the difference between those on statins and those not.  Very interesting was the finding that those on statins had better mental health (i.e. less depression).

Statins: Mechanisms of neuroprotection

A very thorough presentation of the effect of statins and their possible mechanisms along with a review of their use in Alzheimer’s, Parkinson’s, Multiple Sclerosis and strokes, is in the excellent paper:-  Statins: Mechanisms of neuroprotection

 The anti-oxidant effect of statins

A study called The anti-oxidant effect of statins, looks very interesting, but only the abstract is freely available.  Here is the summary:-  

"A number of recent reports have shown that statins may also have important anti-inflammatory effects, in addition to their effects on plasma lipids. Since inflammation is closely linked to the production of reactive oxygen species (ROS), the molecular basis of the observed anti-inflammatory effects of statins may relate to their ability block the production and/or activity of ROS. In this review, we will discuss both the inhibition of ROS generation by statins, through interference with NAD(P)H oxidase expression and activity, and the actions of statins that serve to blunt the damaging effects of these radicals, including effects on antioxidant enzymes, lipid peroxidation, LDL cholesterol oxidation and nitric oxide synthase. These antioxidant effects of statins likely contribute to their clinical efficacy in treating cardiovascular disease as well as other chronic conditions associated with increased oxidative stress in humans."

 

Conclusion

 
Given the minimal side effects, that was more than enough evidence for me to start some primary research of my own. Step one was to try atorvastatin myself. 

My hypothesis is that atorvastatin will reduce autistic behaviours and that the mechanism is the reduction of neuroinflammation in the cerebellum and particularly in the Purkinje Cell Layer (PCL).  I believe that this will be valid regardless of the type of autism. 

The beneficial secondary effect will be reduction in LDL cholesterol, which is typically elevated in cases of autism.

 

Click here for  -  Statins Part 3



 

Praise the Lord and pass the Statins - Part 1

If you are not a native English speaker, you may not have heard the praise “praise the Lord and pass the ammunition”.  It originates from a song written after the Japanese attack on Pearl Harbour in 1942.  A warship’s chaplain puts down his bible and mans a gun firing back at the incoming enemy planes saying, "Praise the Lord and pass the ammunition".

According to Wikipedia, the chaplain was Howell Forgy, was aboard the USS New Orleans.

To hear an original recording click here.

In our case the enemy is neuroinflammation, rather than the Japanese.

 

Deborah Fein and Martha Herbert

There are some very good researchers in the field of Autism and these two ladies are on my list of the best.  It looks like this paper was mainly the work of Ms Fein’s colleagues at the University of Connecticut: - Can children with autism recover? If so, How?

The paper is very readable and not science-heavy at all.

One of the explanations put forward for the rare event of recovery, was the possible reduction in neuroinflammation.  This very much fits in with the conclusions so far on my blog;  reduce neuroinflammation and in particular in the cerebellum.

Now we have a brief time-out to introduce you to our new friends, the Statins.

Source: W. Gibson Wood, Ph.D.  Department of Pharmacology, University of Minnesota

Statins

Statins are a class of drug used to lower cholesterol levels by inhibiting an enzyme which plays a central role in the production of cholesterol in the liver. Increased cholesterol levels have been associated with cardiovascular diseases and statins are therefore used in the prevention of these diseases. Research has found that statins are most effective for treating cardiovascular disease (CVD), with questionable benefit in those without previous CVD, but with elevated cholesterol levels.

Statins act by competitively inhibiting HMG-CoA reductase, the first committed enzyme of the HMG-CoA reductase pathway. Because statins are similar to HMG-CoA on a molecular level, they take the place of HMG-CoA in the enzyme and reduce the rate by which it is able to produce mevalonate, the next molecule in the cascade that eventually produces cholesterol, as well as a number of other compounds. This ultimately reduces cholesterol via several mechanisms.

1.     Inhibiting cholesterol synthesis

By inhibiting HMG-CoA reductase, statins block the pathway for synthesizing cholesterol in the liver. This is significant because most circulating cholesterol comes from internal manufacture rather than the diet. When the liver can no longer produce cholesterol, levels of cholesterol in the blood will fall. Cholesterol synthesis appears to occur mostly at night so statins with short half-lives are usually taken at night to maximize their effect. Studies have shown greater LDL and total cholesterol reductions in the short-acting simvastatin taken at night rather than the morning, but have shown no difference in the long-acting atorvastatin.

2.     Increasing LDL uptake

 

3.    Other effects

Statins exhibit action beyond lipid-lowering activity in the prevention of atherosclerosis. Researchers hypothesize that statins prevent cardiovascular disease via four proposed mechanisms (all subjects of a large body of biomedical research)

1. Improve endothelial function
2. Modulate inflammatory responses
3. Maintain plaque stability
4. Prevent thrombus formation

Statins may even benefit those without high cholesterol. In 2008, the JUPITER study showed fewer strokes, heart attacks, and surgeries even for patients who had no history of high cholesterol or heart disease, but only elevated C-reactive protein levels

 

*****************   Now back to today’s post  *******************

 

Neuroinflammation in the Cerebellum

How hard can it be to find a therapy for neuroinflammation in the cerebellum?  Thanks to Google Scholar, the answer is a few clicks away.

First of all we need to find what other diseases affect the cerebellum or cause inflammation there.  I settled on two completely different cases to investigate:-

1.    Cerebral Malaria 

2.    Traumatic Brain Injury (TBI)

 

Cerebral Malaria (CM) 

First let’s look at what happens in cases of cerebral malaria:-

i) Cognitive sequelae
ii) Speech and language impairment
iii) Epilepsy
iv) Behavior and neuro-psychiatric disorders

Now remember we are looking at malaria, not autism; but this list could just a well be a summary of the effects of autism.

An excellent paper on Cerebral Malaria is: - Cerebral Malaria; Mechanisms Of Brain Injury And Strategies For Improved Neuro-Cognitive Outcome

An emerging area of research is the applications of statins to reduce the neuroinflammation caused by this type of malaria.

Here the secondary action of the statin is important; cholesterol reduction is not relevant.  Here are some highlights:-

·         Cognitive impairment in animals rescued from CM by antiplasmodial drug treatment is abrogated by adjuvant lovastatin administration

·         Lovastatin treatment increases functional capillary density and decreases leukocyte-endothelial interactions

·         Lovastatin protects against blood-brain barrier disruption

·         Lovastatin treatment reduces cytokine levels

·         Lovastatin treatment decreases ROS production

 Read the full report:-  Statins Decrease Neuroinflammation and Prevent Cognitive Impairment after Cerebral Malaria

 

Traumatic Brain Injury (TBI)

It is self-evident that a traumatic brain injury, like a car crash, will lead to neuroinflammation.   The search is on here to find optimal ways to treat this inflammation and achieve an optimal outcome.

 

Here is one paper: - Statins in Traumatic Brain Injury

  

"The use of statins remains a novel therapeutic strategy for TBI. There is robust preclinical data demonstrating the efficacy of statins in acute brain injury models that recapitulate the heterogeneous pathology of clinical TBI. Animal studies have defined mechanisms by which statins may improve outcomes after TBI and should guide statin choice and dosing paradigm for clinical translation."

A more general paper is:- Statins and Brain Dysfunction

and for the older readers:-   Statins: The Role in the Treatment and Prevention of Alzheimer's Neurodegeneration

This should be an interesting paper, but only the abstract is free:-  How do statins control neuroinflammation?

Conclusion

Statins are among the world’s top selling drugs.  With so many people using them, there are of course reported side effects; but as drugs go, the side effects look pretty minimal.  Those at high risk of heart disease, such as those with Type 1 diabetes, are routinely prescribed statins even from a relatively early age.

It has been claimed that autistic people are already at higher risk of heart disease, due to their low level of good cholesterol (HDL) and sometimes higher level of bad cholesterol (LDL). The research is not 100% consistent; but it is very easy to go and check your child's cholesterol.  Holding him still while they draw the blood is another story ....

So it would appear there is one and maybe two very good reasons for autistic people to take statins.


Click below to see Part 2, to decide which statin to choose (there are many).

Statins for Neuroprotection in Autism - Part 2