Invest in ME Research International ME Conference 2018

Karl Johan Tronstad started by saying that the past two days had been very constructive and it is a really different atmosphere compared to normal scientific meetings.

His topic is cellular energetics and their approach to studies in ME/CFS. Dr Tronstad works very closely with Drs Fluge and Mella and their translational research team at Haukeland University Hospital.
Samples are collected from patients attending the clinic, stored in a biobank and then the researchers try to find chemical changes in these samples that can be used in their laboratory experiments. Until now they have been working on metabolomics (metabolic profiling of the samples), some gene regulation and they also have a study where they try to identify ME in families by looking for gene variants that can be important for the disease.
They also use fibroblasts and targeted studies trying to find mechanisms and maybe biomarkers.

The team has been funded by the Kavli Trust, patients and their families and the Norwegian ME Association. Last year they received funding from the Norwegian Research Council and that project is trying to find out if an initial insult leads to defective energy metabolism.

The project has four goals
1/ Strengthening and extending the ME/CFS biobank
2/ Metabolic profiling of patient samples
3/ Identification of risk genes (based on the ME family studies using exome sequencing)
4/ Mechanisms and biomarkers that can be taken further

When you study a patient sample you get a snapshot of that point in time so one has to be careful of interpretation and to avoid this they do many different analyses. They are looking for changes in fuels that are circulating (sugars, carbohydrates, proteins that are degraded to amino acids and also lipids). They are also looking at specific factors that can be a source for an abnormal response.
In 2016, they published a paper that showed a change in the pattern of 20 amino acids and this change was linked to the central axis in energy metabolism. In a normal cell there are many back up pathways if one pathway is blocked so you may not see very much but the team thought that this change in amino acid pattern could mean something important. They are studying the meaning of this blockage and the cell’s ability to adapt to such a blockage. They also asked whether there is something in the patient’s blood that is causing the metabolic change. They took healthy, normal muscle cells and exposed them to sera from patients and healthy controls. The take home message was that there was a change in oxygen consumption (important for mitochondrial function) and also in lactate production. There is something in the patients’ sera that changes muscle cell metabolism but they do not know what it is and it can be quite complex.

They are now also using radioactivity labelling to try to see what is happening to the cellular fuels (not in the glycolysis but mitochondria). They label various substrates with radioactive markers and then measure CO2 coming out of the cells. They are not only looking at the nutrient flow but factors that may regulate this and to do this they try to learn from other conditions such as cancer, aging, neurodegeneration, diabetes etc. They try to do targeted approaches to find out which factors might be involved as these pathways can be affected by many things such as fasting, vigorous exercise, starvation and so on.

So far they have fibroblasts (skin cells) from one ME affected family and in this way can look out for mutations that may be the cause of the disease. They are measuring oxygen consumption of these fibroblasts under different nutrient conditions and they think they can see some differences in the cells’ ability to adapt to different conditions. These cells are not optimal cells to look at metabolic changes but they are cells that are easiest to obtain and can be stored in the laboratory for some time. Until now, they have worked with PBMCs that are easy to obtain but they cannot be maintained in the laboratory and fresh samples would be needed often to do such a study. They also use some commercially available muscle cells. There was a cartoon illustrating the team’s concept. The energy output needs to be balanced by fuel going in. This is being mediated by cell metabolism and kept quite constant in variable conditions. Cells cannot do more than their energy levels permit. Threats to this system can be such as reduced level of fuel, energy leak, metabolic obstruction, disturbed control and these may be overlapping and all may contribute.
The energy demand will also depend on different conditions (rest, work recovery). If there is a cellular obstruction as in the Tronstad team’s model then the cells need to use an alternative fuel source that may not be ideal.
This may lead to metabolic disruption and weakening of the system. The cells try to do damage control and maybe the system is not really resting. If you add increased energy requirements to such a system you may be overcharging the system and this is not sustainable. They are looking for a cellular marker for post exertional malaise.
They try to mimic and target these different conditions using laboratory cell models (Hela for example). They will investigate these mechanisms and target them with different kind of manipulations (pharmacological, nutritional and also genetic). They also aim to get samples from patients so that they can use a translational approach in the laboratory.
The way they can stress cells in a culture is to put them under nutrient deficient conditions, they can make restrictions for the total energy supply, they can control oxygen level but they can also use targeted methodologies to modulate specific metabolic programmes (pharmacologically or genetically) and they can also add specific inhibitors.

The conclusion is that there is suboptimal energy metabolism and that creates a weakness that may not be visible under every condition. They think these mechanisms are reversible and they are working on ways to rectify this.

Question Have the team looked at mitochondrial DNA in siblings of ME patients as mitochondria are inherited from the mother?

Answer If you are looking at mechanisms it is quite a complicated way to go as the mitochondria only code for 13 proteins that belong to the respiratory system. If there were a genetic defect you would see it in different means.

Answer by Maureen Hanson: They published a paper on mitochondrial DNA after sequencing 200 patients and 200 controls but they did not find any differences. They did find some differences that correlated with severity of certain symptoms but not having a disease or not.

Question: Any thoughts on mitochondrial supplementation?

Answer

It is difficult to come up with any recommendation. Based on the team’s review there is no reason for them to advice on anything regarding mitochondrial targeted treatment.

Don Staines talked about the emerging TRP pathology the NCNED is studying in Queensland, Australia. They are interested in a group of nonspecific ion channels called the TRPM3 ion channels. These are important in bringing in calcium into the cells. These are critical in several biochemical functions (mitochondrial, respiratory etc.) and the talk concentrated on TRPM3 expression in NK cells and the gastrointestinal tract.

There has been 30 years of research into NK cells in ME/CFS and the NCNED team use NK cells as the model but Dr Staines stated that what happens in NK cells probably happens in all the other cells that carry these ion channels as well. They have previously found reduction in NK cell lysis and this leads to immune dysfunction, impaired function of the NK cells. As they cannot access tissue in the brain and heart for example they use fresh blood samples instead. Frozen blood cannot be used for this research.
TRPM3 has more isoforms than any other TRP. One of the subtypes is non-functional and contaminates other TRP channels nearby.

When people say ME/CFS is a neurological disease one can say yes and no as all organ systems are likely to be affected by ME/CFS.
The NCNED group’s hypothesis is
1/ There is a significant link between TRP dysfunction and CFS/ME
2/ There is no relationship between TRP dysfunction and CFS/ME

Research question 1 They identified genetic difference in TRPM3 in CFS/ME compared to healthy controls and to understand these differences they need to examine TRPM3 expression on the cell surface
1/ Can they identify novel TRPM3 receptors on the surface of isolated NK cells from healthy controls?
2/ Is there a difference in TRPM3 expression between CFS/ME and healthy controls?

Next step is to examine what is the effect. Do calcium levels in isolated NK cells differ between patients and healthy controls?
In bright NK cells there is decreased expression of the ion channel, decreased transmission of calcium, changes in biochemistry and impaired lysis of the target cell.
Can the levels of calcium coming in and out of the cells be influenced pharmacologically? In vitro tests have been done using pregnenolone sulphate which increases the number of receptors but nothing else changes. Calcium levels are still low in the cytosol and endoplasmic reticulum. It made no difference.
They did additional tests by using drugs that block loss of calcium from the cells and this seemed to work. It demonstrated (in vitro) amenability to pharmacological intervention showing that ME/CFS is potentially reversible.
As TRPM3 receptors are located in the white matter of the CNS (brain and spinal cord), eye, trigeminal and ciliary ganglia, pancreas, gastrointestinal tract, cardiovascular system and is co-location with mACh receptors (e.g. lipid rafts) they may correlate with various symptoms ME/CFS patients’ experience.

Research question 2: Tissue Accessibility?
Until now, the work has been done in blood due to inaccessibility of tissue samples but they have now been able to get gut tissue. The TRPM3 expression is sparse in ME/CFS patients gut tissue compared to healthy controls.

Research question 3 involved muscarinic receptors. Can they identify and characterise them in the gastrointestinal tract of patients and healthy controls? Are there inflammatory changes in the gastrointestinal biopsies between ME/CFS and healthy controls?
mAChM3 is reduced in ME/CFS vs healthy control and IL-alpha is reduced in ME/CFS vs healthy control suggesting it is not the primary mediating pathology.

Conclusion: Impaired expression and function of TRPM3 ion channels affecting calcium signalling/stores suggests the primary pathology of ME/CFS.

Question:
Do you have a biomarker?

Answer (Don Staines): This is always a difficult big/question to answer. The SNPs would be a beginning of a biomarker but there is a caveat that the results may be different in a different population.
Question:
What is the next stage?

Answer:
The damaged cells are amenable to pharmacological intervention and the task is now to start isolating various drugs and drug derivatives to look at.

Question:
Do you anticipate there being differences between cells from a single person so that there might be combination of chemicals that worked in some cells but might not work in others?

Answer:
Changes in the genes are different. We are all different. The test mentioned involved 20 to 30 patients. You take one person and their SNP profile may be different from the other person’s so you cannot say one drug fits all. What is tricky here is the concept of personalised medicine where you have to diagnose each case as it comes from a profile of SNPs and ion channels and there may be subtle differences in drugs and drug derivatives. Therefore, you may have to develop a cocktail approach to each patient.

Remembering the influence of Anne Örtegren.

The lecture started with a tribute to Anne Örtegren.

Professor Jonas Blomberg opened the lecture by giving insight into the contact he had with Anne. It started five years ago.
Anne was like a good fairy guiding Professor Blomberg into the ME field.
Anne was very considerate, very knowledgeable and also a kind person. That was a very good introduction to ME research, patient advocacy and collaboration between researchers and patients which was new to Professor Blomberg.
He liked this collaborative approach. Within the Swedish and international networks emails were exchanged.
Anne was always helpful and sent good suggestions and found new research that the researchers should know about.
Then the emails that used to come rather frequently, stopped.
Later Professor Blomberg understood why.
Anne had intense and intractable pain and suffering.
It was understandable that she could not continue.
We should remember Anne as a very good proponent of the ME cause.
http://www.investinme .org/IIMEC13-news-180201.shtml

Professor Theoharis Theoharides

Everything Dr Theoharides was going to talk about could be found on his website www.mastcellmaster.com.

Mast cells are involved in allergic reactions and that is the tip of the iceberg. Why mast cells? Dr Theoharides referred to Anne’s symptoms of hyper reactivity and burning skin. Whether we call this “allergy” or “atopy” or “mast cell activation syndrome” we owe it to Anne to investigate the connection she described and find suitable treatments and a cure.

Atopy is caused by a zillion things and seems to attract ME and the more allergies and sensitivities one has the more likely the symptom severity is being affected or having the chance of developing the disease. There was a paper published recently to look at precursors for mast cells where they measured telltale signs on the surface of the precursors and they found more mast cell precursors in ME patients. Of course, they do not know where these cells go and what they do.

Mast cells are made in the bone marrow and travel via the blood but do not circulate. They home into tissues.

There are many different things that present as allergies such as angioneurotic edema, asthma, atopic dermatitis, food allergy, food intolerance, idiopathic urticaria and so on. All of these involve mast cells but only allergies are literally allergies yet mast cells not only participate in all these other diseases but are activated by other things other than allergies. Atopic diseases can be diagnosed using blood (IgE,IgG₄) but the most proper way is to collect 24h urine (kept cold) sample where they can measure breakdown molecules of two metabolites histamine and prostaglandin₂.
90% of patients that show up in clinic have shiny circles under their eyes and if you scratch the surface of their skin (under the arm or back for example) they get a red welt within a minute. This means the mast cells on their skin respond to pressure and not to allergens. This is a quick tell- tale clinical sign that might allow one to start thinking something is going on.

Many years ago, long before becoming involved in ME, Dr Theoharides and team started looking at other diseases and they showed that mast cells are involved in interstitial cystitis, fibromyalgia, autism but that does not mean that mast cells do exactly what they do in allergy. They can do a number of things depending on the trigger and circumstance. They can be triggered by numerous factors such as environmental factors, pathogens, peptides from the brain, growth factors etc. Mast cell looks like a soccer ball and it can explode and release its contents of which we mostly know histamine and then we use antihistamines to address the histamine after it has been released. We really need a good way of blocking this occurring.

What interested Dr Theoharides in this field that many patients with mastocytosis or mast cells that were super activated had cognitive problems, especially brain fog. Depending on what the mast cells release, they can affect every organ in the body, including the brain. Since the publication of this information in New England Journal of Medicine there has been a new insurance accepted diagnosis called mast cell activation syndrome.

There are two important factors. You do not need to have too many mast cells or allergies necessarily but the mast cells are activated by everything under the sun and the diagnosis includes neurological problems for the first time. All of these patients complain of brain fog as well as symptoms (flushing, headaches, hives, myalgias etc.) seen in ME patients.

Mast cells are not isolated. They talk to macrophages, T-cells and they have receptors on the surface that can be activated by bacteria, viruses, fungi, mold etc. They respond to heavy metals and herbicides.

Why do we think mast cells are involved in ME?
The hypothesis is based on the work on mast cells in the brain. If you take a cross section of blood vessels in the hypothalamus that regulate homeostasis in the body you can see red blood cells, endothelial cells, the pericytes that make up the blood brain barrier and the mast cells “hug” the blood brain barrier. Dr Theoharides and his team believe based on their work in autism that mast cells are involved in a look- a-like inflammatory response. Why hypothalamus? Because strangely it has as many mast cells per unit volume as one’s skin yet the brain does not get allergic reactions. So obviously these cells are doing something there and the more we can identify what they do the better for a number of diseases. We know that homeostasis is derailed in ME patients.

Many conditions worsen by stress, and we know that major stressors (mother’s stressors) during pregnancy makes the baby more prone to allergies. The more atopic conditions or tendencies the child has the more likely the child develops autism. This was tested in a mouse model where the mouse was stressed by being put into a confined space. Stress hormones called corticotropin releasing hormones (CRH) trigger the release of mast cells that open up the blood brain barrier. Together with neurotensin mast cells induce the dilation of blood vessels. When the blood brain barrier is opened up all kinds of toxins and potential pathogens get in whether from the environment or the gut.

Things get worse as when neurotensin sees the mast cells as it makes them grow receptors so they can respond to CRH and this becomes a self-amplifying process that cannot be stopped unless the mast cells are stopped. It may be useful to look at multiple triggers in combination rather than single triggers as smaller amounts of many triggers can do the damage. The team have measured biomarkers in Fibromyalgia and have shown that CRH and substance P were significantly increased in Fibromyalgia patients and they are known to stimulate mast cells to release chemokines. They are currently comparing ME patients to Fibromyalgia patients.

In summary so far:
Mast cells can explode in severe allergic reaction in which case histamine and tryptase are being released. A number of cytokines and pro inflammatory molecules are also being released without undergoing degranulation.
They also studied mitochondria and discovered that mitochondrial DNA together with CRH increases allergic responses. Mast cells are involved not only in allergic reactions but inflammation as well. All inflammation is not the same. Systemic sepsis is inflammation and it is called cytokine storm. If you have encephalomyelitis it tends to be generalised in the brain. If you have meningitis it is the lining of the brain and if it is in myelin that insulates the nerves it is Multiple Sclerosis (MS).

So what about ME?
Dr Theoharide’s take is that it is focal inflammation in the hypothalamic area and if we are talking about myalgic encephalomyelitis it should be myalgic encephalomyelitis in the hypothalamus and not necessarily the whole brain.
Dr Theoharides then talked about toxins that are in the environment ( seafood related toxins, spider bites, mold).

How to measure inflammation? Dr Theoharides measures both pro-inflammatory and anti- inflammatory marker molecules because the range gives more information than individual molecules alone.

How does inflammation occur in the brain?
It can be local or it can be materials coming in the blood and affecting the hypothalamus. The team’s hypothesis is that the mast cells activate microglia that are defenders of the brain. There are no white blood cells in the brain and if white blood cells get into the brain they
get recognised as foreign and get attacked and then you get MS. Microglia are like spiders with a web and neurons grow on the web and if microglia detect dangers they start to multiply and try to confine that area and get rid of it.

If something is coming from the periphery, how does it get to the brain?
Blood brain barrier might be disrupted and that is how it goes in but there is another way they can go in. There are tiny vesicles that are released from cells and they are called microvesicles or exosomes and it turns out they carry cargo to various places and they are so small that they get through the blood brain barrier. A study showing an increased number of microvesicles in the brain of autistic patients compared to healthy controls has been done and they are in the process of doing the same in ME.
They also wanted to know what happens to microglia if they are exposed to microvesicles.

They cultured human microglia in the laboratory and isolated microvesicles from an autistic patient and measured the main inflammatory cytokine IL1 beta. IL1 beta was raised but they did not know what the cargo was but thought whether it might be mitochondrial DNA so they measured mitochondrial DNA in the microvesicles and it was more than in controls.

Does it mean we have a biomarker?
It gives the team something to move forward with.

Danger signals in the pathogenesis of ME were listed as cachexins, electromagnetic waves, heavy metals, herbicides, inflammatory cytokines, mitochondrial DNA, mycotoxins, neuroendocrine disruptors, neuropeptides, reactive oxygen species, venoms and xenobiotics. The connectivity is unknown. Can they block it?

The team have been studying a class of molecules called flavonoids that are very good antioxidants and anti-inflammatories and they have shown that they block mast cells and microglia. Luteolin in particular was mentioned.
They had done a study on mice trying to induce fatigue by giving the mice poly (I:C) that mimics viral infection and then making the mice swim in cold water and they measured inflammatory molecules in the serum and in the brain after giving the mice normal chow or chow enriched with flavonoids.
The inflammatory molecules dropped down to zero when mice where given chow enriched with flavonoids.

In summary:
Mast cells talk to microglia. Microglia contribute to the symptoms of ME. For better or worse flavonoids could be used to treat brain related inflammation.
One needs to be sure other similar diseases are excluded.
They might be co morbid and some might be treatable. These are Chronic Inflammatory Response Syndrome (CIRS), Fibromyalgia Syndrome, Ehlers- Danlos Syndrome (EDS), Gulf War Syndrome, Interstitial Cystitis / Bladder Pain Syndrome (IC/BPS), Mast Cell Activation Syndrome (MCAS), Post-Lyme Syndrome and Post Traumatic Stress Disorder (PTSD).

Treatment Approaches- Exclude/Avoid
Exclude: Hypophysitis and pitutary adenoma, Mast Cell Activation Syndrome
Avoid: allergens, colourings, deodorants, perfumes, electromagnetic waves, flame retardants, heat, histamine rich foods (ripe avocados, ripe tomatoes, cheese, sardines, spices), plastics, preservatives and stress.

Treatment approaches: Brain Gain (a combination of supplements) for cognitive functioning, Caprylic Acid for mold exposure, antihistamines such as Cyproheptadine which increases appetite and is good for migraine prevention, Doxepin, Ibuprofen (anti-inflammatory), Propranolol (beta blocker).
Antacid is always added for stomach protection

Question: Would you measure tryptase?

Answer: Unless one has full blown mastocytosis tryptase is not elevated and it is not part of mastocytosis diagnosis. It is a good quick test and if it is negative then one can look for metabolitis in the urine.

Question: Do cannabidoids help with bone pain?

Answer: An audience member said they found CBD oil helped with pain. Dr Theoharides said he had no experience of it but it seems to work for epilepsy.
Word of caution for product quality in that FDA had found some CBD oil had been produced from plants that had been stored wrongly and been moldy so the oil contained toxins.
The same goes for all flavonoids and dietary supplements as the purity is not guaranteed.

Mady Hornig’s presentation concentrated on trying to parse the heterogeneity of ME/CFS in terms of diagnosis and treatment.

Dr Hornig thinks of immune related brain disorders being based on the following three elements: genes, environment and timing and thinking even prenatally there could be some event that influences the person’s development over the course of their lifetime. Examples of disorders were given such as autism and ADHD in early life, MS and ME/CFS in middle life and Parkinson’s and Alzheimer’s in later life.

The programme at Columbia university relating to biomarker discovery has been wide from RNA based to protein based and DNA studies. They have tried to utilise all of these to make meaningful subsets.
Characteristics of the disease relate to length of illness, onset type, age at onset, disease severity, atypical features and comorbidities. Up to 70% of patients have comorbidities.
Allergy/Atopic disorder studies include asthma, seasonal rhinitis, atopic dermatitis, sinusitis, hives, eczema, hay fever and interstitial cystitis.

Neurological impairment studies involve excessive brain fog, perceptual impairment, attentional deficits and memory dysfunction.

Gastrointestinal dysfunction studies not only consist of microbiome but they are also looking into celiac disease, inflammatory bowel disease and irritable bowel syndrome.

Dr Hornig described the group’s previous immune marker studies into early disease versus late disease, where they described the immune exhaustion paradigm. These studies suggested there were meaningful subsets. She then went on to describe their more recent study on spinal fluid samples. They showed that there is a difference between patients with a classical presentation of the illness (viral onset) versus those with atypical based either over the course of the illness or presentation and who went on to develop other severe conditions later on such as cancer, MS or gulf war illness. IL7 and IL17 were low in the atypical group of patients. These patients have sparse inter-cytokine networks. The results were found before these individuals developed some of the other conditions mentioned and they fitted the ME/CFS diagnosis at onset.

We may think that classical ME/CFS involves increased microglial activation but based on this study the classical ME/CFS group had signs of less microglial activity.
They have also been thinking of allergic/atopic disorders as a subtype. What they found was increased pain and gastrointestinal disturbances and higher levels of endocrine disturbances were predictive of the allergic group.
They were also interested in severe cognitive dysfunction. There has been very little work done on cognitive sensory dysfunction. They’ll use cluster analysis to try to define cognitive dysfunction and ultimately see how this correlates with peripheral immune dysfunction and they hope to have spinal fluid samples from other cohorts to look at the central immune regulation.

They also looked at gastrointestinal dysfunction. The gut brain axis is going to be important in regulating normal health as well as disease. They used stool microbiome doing metagenomics and also predicted bacterial metabolomics in 50 cases and 50 controls. They then used topological data analysis to see how the data clustered together. They saw differences in patients with and without IBS in their metabolic profiles as well as their bacterial profiles. They saw particularly high levels of deethylatrazine (a herbicide common in ground water throughout the US) in all of the groups. It has endocrine disruptor effect. Vitamin B6 pathways were elevated in the non-IBS group. Patients were also grouped based on BMI.

They want to separate biomarkers which are stable versus fluctuate over time and which of them are disease associated versus state associated.
They are interested in the degradation of the herbicide deethylatrazine as a potent endocrine disruptor.

They have also done plasma metabolomics to see which metabolites may have come out of the gut or been transported by the short chain fatty acids into the blood stream of these patients. The cytokines and metabolites can affect the blood brain barrier.
Diet is very important as there some substances such as tryptophan that are important both in production of serotonin as well as downstream synthesis of melatonin which influences the circadian rhythm. Healthy CNS and immune function depend on balance.
Autoimmune disturbances may result from failed uptake of dietary antioxidant precursors in terminal ileum. Certain microbes may also lead to a more pro-inflammatory effect of Th17 and counter reduction of Treg cells.
They use deep data mining techniques in their biomarker discovery project. They want to look at both environmental exposures as well as genetic variants.

Question How does your allergy-testing correlate with standard allergy tests such as IgG and skin tests?
Answer Point taken. Patients’ self-reports were being used and objective measures have not been looked at yet except in that cytokines are allergy associated. Funding needed to do studies with proper measures.

Markku Partinen started by confirming the WHO classification of ME being neurological. He said unfortunately from his experience as a neurologist it does not guarantee that neurologists will see ME patients. He said his hypothesis was similar to the ones presented by Mady Hornig and Theoharis Theoharides.
He explained disease being made up of factors such as genome that explains 25% and 75% being explained by ambiome and microbiome. Ambiome is the environment and everything contained in it.

Sleep is necessary for life. 100% of rats die within a month if they are not allowed to sleep. Billions of dollars have been spent on the exercise industry but exercise is not vital for life whereas nutrition and sleep are.
He then explained the orexin system that is the most important system for sleep. It is in the hypothalamus regulated by the central circadian clock. Last year the Nobel price was given to three researchers who demonstrated that each cell in the human body has a peripheral clock. This system is also connected to the autonomic nervous system, the endocrine system, emotional regulation and energy balance. Sleep is described by a flip-flop theory. After a time awake sleep follows and in sleep disorders there is no flip-flop between these two states but the balance is broken.

Orexin cells are glucose sensing cells.
When glucose is down orexin cells become activated resulting in more alertness. When glucose level goes up orexin goes down and one becomes sleepy. When ME patients eat carbohydrates they feel worse. A high carbohydrate meal makes one sleepy and fatigued. If ME patients are at the limit of hypoglycaemia, almost acidic, they usually feel better.

Orexin cells are also salt sensing cells.
When salt goes up orexin goes up and this leads to wakefulness and increased blood pressure.
When salt goes down orexin goes down leading to sleepiness and lowered blood pressure. ME patients often lack salt which causes low blood pressure and increased heart rate.

Position, immobilisation syndrome.
There are a lot of studies of ME and exercise but very few studies on position. Experience from studies in the elderly shows that exercise is not important but position is. If a person is horizontal for a long time salt goes down and these patients will develop hyponatremia.
Just by elevating the top half of the bed a little leads to sodium going up. This is something that should be investigated in ME patients in a controlled way.
It is a vicious cycle for ME patients that cannot remain upright leading to salt going down.

Why is sleep necessary?
Sleep is necessary for the energy of the brain and brain clearance. It also has a role in immune function and memory function. The amount of sleep needed is individual (between 6 and 9 hours) any time between 9pm and 9am.

“Astrocyte-Neuron-Lactate-Shuttle”.
Glucose penetrates the blood brain barrier. Most of it will go to the astrocytes to be stored as glycogen. Another place for stored glycogen is the liver. By glycolysis lactate is formed. When a person is awake neurons use lactate as fuel. Some of the previous talks discussed evidence of increased lactate in the brain of ME/CFS patients and also increased lactate when measured in the periphery.
Dr Partinen stated that it is normal that lactate is accumulated in the astrocytes during deep sleep and this is very good and essential for good neurological function when people are awake. Another important function is the glutamate –glutamine cycle. Without working glutamate cycle the lactate shuttle is not working. If people are not active in the daytime they are not producing energy in the night time.

Clearance of metabolites.
In the deep slow wave sleep the brain’s interstitial fluid is increased by more than 60%. Glymphatic (glial-lymphatic) flow helps clear accumulated substances in the deep slow wave sleep phase. Possible reasons for unrefreshing sleep are behaviourally induced insufficient sleep, medications (bentzodiazepenes), mental hyperactivity and circadian rhythm disorder. Sleep disorders must be ruled out before giving a patient an ME/CFS diagnosis but sleep disorders may sometimes be co-morbid.

Autonomic nervous system
There are two parts, the sympathetic and the parasympathetic system. The sympato-vagal balance regulates the immunological homeostasis. If, theoretically, there is a very low vagal tone (low parasympathetic tone) the risk for inflammation increases. On the contrary if the vagal system is stimulated tolerance is increased.
In a simplistic way by lowering the parasympathetic tone immunity decreases and inflammation increases. To improve vagal tone needs to be increased by vagal stimulation or the sympathetic load needs to be lowered by increasing fitness to a sufficient level but not too fit. PEM after exercise in ME is partly due to this system. Important things are relaxation and good sleep and sometimes beta blockers can be used.

Sympathicotonia
Many ME patients are sympathicotonic which means that their sympathetic tone is dominant. This is a good thing and many top athletes are like this.
They perform well under pressure but get emotional when pressure is off. Many ME patients have been sporty or mentally very active so they belong to this group and then something happens and they become unwell.

In sleep studies blood pressure can be measured at every heartbeat. During normal sleep blood pressure goes down and vagal tone increases to induce relaxation. In insomnia there is no lowering of blood pressure and during the whole night the system is sympathicotonic.
This is exactly the same way that dogs sleep. They sleep but they are constantly alert to any noises around them. They are never in a deep slow wave sleep. ME patients and insomniacs do not go into deep slow wave sleep and this leads to the lymphatic system not working and there is no good energy formation. There have not been many polysomnographic (PSG) studies in ME but the few studies that have been done show that subjective sleep quality is poor but there are no significant measurements in cortical differences.

Partinen’s group decided to measure the autonomic tone in their PSG sleep study. In normal healthy people parasympathetic tone increases but their study showed that the parasympathetic tone in ME patients decreased. They do not know the reason why this happens but there is clearly something wrong with the autonomic control relative to sleep. In this small study ME patients were compared to tired controls but when they compare ME patients to normal healthy controls the difference is much higher (study underway).

The team’s hypothesis is that ME patients have abnormal deep N3 -sleep. Parasympathetic tone is decreasing instead of increasing. This leads to dysfunctioning of the glymphatic system and lack of brain energy.

In 1934 there was a publication by Lapage that used the term “Autophilia” and Dr Partinen theorises based on this paper that due to reasons such as genetics, microbiome, pregnancy, infancy some people develop autophilia.
These are the persons that develop autopathias (allergies, atopy etc.), spasmophilia, neurological symptoms and psychiatric symptoms (e.g., anxiety is a problem in the autonomic nervous system). In ME one can find all these symptoms. In childhood these patients tend to be sensitive, flushing easily, experience fainting, travel sickness and nosebleeds. They are often lean and catabolic and have lower resistance to infections. As adults they are emotional, sensitive, excitable, mentally active, creative, culturally active and sporty.

In conclusion: There is abnormal N3-sleep that causes lack of brain energy and problems with glymphatic flow. Autophilia is a predisposing factor and restoring sleep will help in regaining homeostasis. Avoid medications but use sleep hygiene, CBTi (insomnia CBT), nutrition and perhaps H1 antagonists in very small doses (e.g., doxepin) and also vagal stimulation (no studies on this yet).

Question What are your thoughts on “owl” patients whose sleep is reversed? Answer It is a complex problem because normally people should sleep when it is dark and be awake when it is light. There are some patients who feel better if they keep their reversed sleep pattern. Whatever we look at (hormones or white blood cells) there are circadian rhythms and they vary. There are no studies asking whether patients feel better at a certain time of the day and what is their worst time of the day.

Question You mentioned vagal stimulation. There are suggestions such as gargling or singing very loudly can help vagal stimulation. The person asking the question found singing loudly improved their sleep.

Answer That is possible and vagal stimulation is now used in intractable epilepsy and they may even put electrodes inside the brain but there are now some external stimulators as well. Studies should be done to test this properly.

Question There is a saying: “Time spent sleeping is never time wasted” mainly because sleeping is healing. Are forceful sleep hygiene measures good for patients?

Answer Theoretically the best time to sleep is somewhere between 10pm to 9-10am. If one can choose then it is better to go to sleep earlier and wake up earlier.
The important thing is to have at least 6h of sleep. Interestingly they have followed Parkinson’s disease patients over several years and their hypothesis was that if these patients had less sleep there would have higher mortality but the result was the opposite.
Patients who slept less than 6h had lower mortality.
They looked at all kinds of variables and found that patients that were optimistic despite their many problems had better outcomes. ME is a serious disease but the mortality is not very high. Is it so that despite severe symptoms there is still internal hope.

Question The more exercise I do during the day the longer I sleep and then I am more likely to feel more ill the next day, with bad headaches. There is a paradox in that sleeping more makes one feel worse. Conclusion is to sleep less and do less exercise to avoid increased headaches.

Answer This is a touchy area because theoretically if you do a lot of exercise you sleep well until you overdo it and develop overtraining syndrome. In ME one important thing is the position when sleeping. Elevation of the head is important. If you sleep too long mental function gets worse and that is why sleep deprivation is used as treatment in serious depression. Sleeping too much is worse than sleeping too little. The hours needed to sleep vary individually.

James Baraniuk presented his “Year End” report. They have an exercise model that they have used for ten years. They do an extensive set of questionnaires (about 50) and these are done online so that the patient can stop any time they want to and carry on later. The physical and history part is now 50 pages long and it can take up to four days to complete. It is not something a GP can do in eight minutes.

Dr Baraniuk is trying to work with the common data elements to try to develop the key questions that should be asked in every study so that the patient populations from all around the world can be standardised.

They also do allergy skin testing and the rate of positivity is the same in ME/CFS as in the control group. They have now done this in their fourth cohort. The levels of antibodies to allergy IgE are also equivalent between the two groups. Dr Baraniuk does not believe there is an increase in true allergic disease but there is an increase in sensitivity.

After talking to the participants they do an MRI and this includes an MRI during the cognitive test so that they can see if there are differences in the regions of the brain that are activated while the participants are doing the memory test. After this participants do an exercise test that is similar to the one described by Dr Hanson except that Dr Baraniuk’s participants do a submaximal test (at 70% of the person’s predicted maximum heart rate) which lasts 30 minutes as opposed to 10-12 minutes as described by Dr Hanson.
The participants can increase their heart rate up to 85% of their predicted maximum that is the same as cardiac stress test. They do not have to do 30mins. If they want to stop earlier they can stop and that would essentially be the person’s maximum.
Dr Baraniuk finds that his research participants’ day one and day two tests are highly reproducible.

After the second day exercise the participants do their second MRI to see if there are areas of the brain that have changed while they do their thinking and are those changes caused by the exercise and whether this is a model for exertional exhaustion. Finally, everyone who is agreeable gets a lumbar puncture. These participants are kept overnight to reduce the risk of spinal headaches.

The talk then went into discussing hypotheses including depression, pain and tenderness, exercise, postural tachycardia which is not the same as POTS, orthostatic intolerance, MRIs and some results from the CSF studies.

Depression
The depression scale used is a Center for Epidemiological Studies- Depression (CESD) questionnaire involving 20 questions with a scale of 0 to 3. The maximum score is 60 and if one’s score is 16/60 or more one is classed as depressed. This has been shown in 28 studies involving over 22,000 people. The problem is the scale has a 30% false positive rate.
One can do a factor analysis as the questionnaire can be broken down to four factors that are grouped together from the 20 items such as somatic, depressed, anhedonia (lack of pleasure) and interpersonal. The somatic factors are fatigue, sleep, cognition, effort, bother, talking less and appetite change. These four factors have been known for decades but nobody has ever bothered to score them in any patient population.
Dr Baraniuk’s group did this and discovered that using this scale 24% of the US population is at risk of depression and 54% of ME/CFS patients are at risk of depression but this score is biased as it is caused by high somatic domain scores (sleep, cognition etc.). It is the same thing for Gulf War Illness. Depression in ME/CFS discounted based on this evidence.

Pain
Pain is not in the SEID criteria because it does not differentiate CFS from other diseases such as Fibromyalgia. Dr Baraniuk says he has a big problem with how these diseases are defined and to bypass that he wanted to know about tenderness. Does 4kg pressure hurt?
The Baraniuk team uses a spring-loaded gauge to measure pain and the person being measured is in charge during the procedure. They use this on all of the 18 fibromyalgia pressure points and they can then calculate a score for average pain. The score for healthy people is about 7kg of pressure. The CFS group peaks at 4kg. The same thing happens when patients breath in formaldehyde or any other chemical, are exposed to bright lights or loud sounds.
The threshold is broken. An ambient background noise can become a loud and overblown when you have CFS. It is not imaginary but a very real broken switch. It is a hypothesis.

Submaximal exercise
When they did this they were expecting to see similar results to those of the VO2max stress tests. They did not see any difference in day one and day two results. They were identical and overlapped.
The test is up to 30 minutes but people can stop earlier. If people can only do 2 minutes on the first day they can only do 2 minutes on day two as well so it is highly reproducible and turns out to be useful as far as the MRIs are concerned. It also means that if there is a decrease in VO2 in the maximal test there is something magical happening after the Baraniuk participants stop.
We need to know how many people in the VO2max test actually have decreased performance, how much is that decrease and we need to know the sensitivity and specificity particularly if this is going to be used as diagnostic for CFS and certainly if it is being used for disability purposes.
Result: No difference in performance between day one and day two submaximal exercise tests.

Postural tachycardia
In previous research on Gulf War Illness (presented at IiME by Rakib Rayhan) they found 1/3rd of patients developed POTS after day 1 of submaximal exercise. Lying down they would have their normal heart rate and before exercise after they stood up from lying down position their heart rate would increase by about 12 beats.
After exercise 1/3rd would have an increase of at least 30 beats when they stood up and this is a definition used for POTS. This was an unexpected result. They got funding for a verification study in GWI that has now been finished but has been rejected for publication and that is what happens for verification studies.

They found that the START (Stress Test Activated Reversible Tachycardia) group had brain stem atrophy and this is similar to what Richard Kwiatek (Australia) reported in CFS. They then did the same study in their CFS group and report similar findings to GWI START group.
Dr Baraniuk encouraged all exercise study groups to do lying to standing heart rate measurements before and after exercise to disprove his findings.
To find out which groups had POTS they took dizziness scores lying down after exercise and all three (CFS with no heart rate change, CFS START and CFS POTS ) groups had similar scores. After standing up all three groups had again similar scores and this made

Dr Baraniuk think what orthostatic intolerance really is. Is it a difference in sensitivity coming from the vestibular system that one is feeling dizzy and light headed even when lying down and it has nothing to do with one’s heart rate or autonomic system.
Result: Exercise does not worsen orthostatic intolerance.

MRI
There are some changes but it is not that big. They can conclude that those groups that have changes in their heart rate have worse brain function after exercise.

Cerebrospinal fluid & biomarkers
miRNA levels were higher in people who did not exercise and they went down after exercise.
It is not a robust enough diagnostic marker for CFS but it has led to some ideas about what might be going wrong. If the miRNA level goes down one has more messenger RNA and one can make more protein and that protein will have more function. It looks like exercise in CFS leads to increase in some particular enzymes such as Insulin Llike Growth Factor 1 receptor, Transforming Growth Factor Beta Receptor 1 which are very important for proliferation of brain cells.
They have also looked at where these are found and they are in the microglia and choroid plexus which is a vascular region where most of the cerebral spinal fluid is being created. This gives some hints as to where dysfunction might be found after exercise challenge.
Conclusion: CFS is not depression, CFS is somatic, tenderness is increased, there is cardiac effect, MRI effect and CSF changes.

Question The effects of activity may not be visible the day after exercise/activity but later on.

Answer One thing the Baraniuk group is looking at is how much exercise is needed to cause a delay in the response. A curious thing is that sometimes a small effort leads to an immediate collapse and a bigger effect may have a longer delay.

Question A patient with POTS went to get tilt table testing and managed to get their doctor to insert a Hickman line. They now start the day with a litre of saline and that has improved the patient’s POTS. Does anyone else have the same experience? It seems a drastic measure.

Answer It is drastic. Dr Baraniuk said he has had one patient that had a central line and would infuse two litres and then go shopping and come back and do another two litres and go to sleep and they felt good. Also the patients that get saline after their spinal tap brighten up.

Dr Baraniuk recommends Gatorade to his patients as it is impossible to get insurance companies to pay for saline infusions.

Ron Davis talked about the Stanford group’s latest discoveries and future plans. All their work has been supported so far by the Open Medicine Foundation (OMF).
They have a scientific advisory board and many of them were in attendance at the colloquium and conference (Maureen Hanson, Jonas Bergquist, Øystein Fluge, Ron Tompkins and Wenzhong Xiao). OMF have just funded Drs Tompkins and Xiao to set up clinical research at Harvard University.

One thing that has been clear to Davis’s team for some time is that they need an assay for a biomarker as so many patients (including Dr Davis’s son) have been told that there is nothing wrong with them because all routine tests come back normal. Dr Davis’s son is now severely ill. He cannot talk or read and Dr Davis still takes blood samples from him regularly and they run the same tests on him as doctors do and he is perfectly normal. The problem is that the routine tests are no good.
The focus of the team is now to prove that the patients are not healthy and they are being compared to healthy controls to see if there is a clear difference. That is task number one.
The next task is to find out if they can distinguish ME/CFS from another disease. This is very difficult to do as for one thing one needs to do all the other diseases.
Dr Davis is not sure this is necessary. What is necessary is to work with physicians to discuss with them what they need to know for a diagnostic test of this type or can they diagnose patients using some other criteria, by just talking to the patient.

Seahorse Instrument
The Stanford group has done some standard testing using the Seahorse instrument. They get best results by using isolated T-cells from patients and stimulating those cells and then measuring the energy production. The results have been reproducible.
If they do not stimulate the T-cells the results are more variable and it is also dependent on the time the patients have last eaten.
It is difficult for the patient not to be allowed to eat before they come in for testing. This test is also very expensive.

Nanoneedle
Another tool the Stanford group has developed is the so-called Nanoneedle. This tool measures electrical impedance that is a very sensitive electrical measurement. They make 400 measurements/second. Once they have finished taking a sample they have made 1 billion measurements. This is much more useful than medical devices that make just one measurement.
The Nanoneedle can be made commercially and if made in large numbers they will not be expensive. The Stanford group could make them for $1 to $5. They have now also figured out how to clean them so they do not have to remake them.
The measurement is done using a blood sample with red blood cells removed and then only a drop of blood is needed. A healthy person’s measurement looks flat on a graph and it is the same for CFS.
They reasoned that CFS patients’ cells need to be challenged so they added sodium chloride salt to make the cells work harder and then their impedance increases compared to the healthy controls. This same test can be used to see which antibiotic works on cultured bacteria and it can also be used on tumours. Physiological change will change the impedance. It is a cheap way of doing testing and the results can be obtained real-time. This might be useful as a diagnostic tool.

The team has also used the Nanoneedle to test how drugs work on the cells and they have found a number of drugs that seem to be able to get rid of this impedance effect. It is unknown if these drugs turn out to be useful or not.
Dr Davis showed an example of one of their experiments to show there was a very big difference between patients and controls, something that has only 1 in 10 million probability of occurring by chance.

They have done a few experiments to understand what works in this assay and this is called “The Plasma Swap Experiment”. The initial thought was that it is in the cell so they tested that doing a plasma swap. They took plasma from CFS patients and put it on healthy cells and vice versa.
They find that the signal they get tracks with the plasma and not the cell. That even suggests that the cells are healthy but there is something in the plasma that is causing this effect.
They do not know what it is yet but it is something that needs to be figured out as it is going through the entire body and may be causing some of the effects. If they can figure out what it is and what size it is they can try to figure out how to remove it and that could be treatment.

Microfluidic Device
Another device that the team have tested is a microfluidic device and they are collaborating with Professor Anand Ramasubramanian on this. This device measures the deformability of red blood cells. The idea is that red cells have to go through a device that is a bit larger than a lot of the capillaries in the human body.
If cells are not deformable they get stuck when they go in and when they reform they go slower through the device. The amount of elongation of the cell can be measured also. This is easy to do. The device needs refinement to make it smaller but they do see difference already in that the red cell entry is slower, transit velocity is slower and the elongation capacity is less in ME/CFS.
This does not currently work as a diagnostic tool as there is a lot of variance. This is an inexpensive and simple test. They will use Atomic Force Microscopy to measure deformability accurately.

Biomarker-Free Sorting
Another device the Stanford team has come up is a Biomarker-Free Sorting of Rare Cells from Blood.
It is a device that measures magnetic levitation. This means that they can suspend something in magnetic fluid if they put a magnet on it. This allows cells to be suspended very quickly.
Blood is put in the device and when the magnet is put on the cells will quickly go into density in a long tube. This device was developed for cancer to separate tumour cells from the white blood cells. The device can separate lots of different cells. Each run on this device costs only 5 cents. CFS patient’s results on white blood cells showed variation and density and some of the results were near normal and it turns out the patient had bacterial infection at the time of better results. The bacterial infection made the patient feel better not antibiotics. There are other patients who report similar saying fever makes them feel better.
Dr Davis thinks it is the bacterial infection that makes them feel better not the fever. In general the patients’ cells are light.
The team want to look at what happens in bacterial infection to make patients feel better.
Portable, mobile health platforms that use smartphones as computers allow rapid sample preparation, blood cell separation and count.

Biomarker Bake Off
Next task is going to be a biomarker Bake Off. This means all the samples coming in are being run on all these devices (nanoneedle, microfluidic blood flow, magnetic levitation, Seahorse and metabolomics).
Robert Naviaux found some metabolomic markers so they are comparing their findings to his finding. This will give five different instruments to look at the consistency. The reason for doing this is that there might be some exceptions and things do not work quite right but the other diagnostic tool did work right and is measuring something different. So in fact one might need to use two different instruments. They do not want many false negatives.

They also need to look at other diseases and how well the tests perform in those. That is when they need the help of physicians because it is possible that other diseases will show the same behaviour. The biggest goal is to show that the patients clearly are not healthy.

A Big Data Approach to Understanding ME/CFS
This project is funded by OMF. It involves 20 very severely ill patients. The project cost about $2million. The patients are bedbound or at least house bound and could not attend clinics.
They have also done a study of 15 families where there is more than one affected family member including unaffected members. It is unfortunately not uncommon to have more than one family member affected. There is a reasonable chance that there is something genetic going on in these families. The data is not yet available.

Hypothesis
Most people think that the way to do research is that you create a hypothesis and then you do a test to try to rule out that hypothesis. That is actually wrong according to Dr Davis. He says NIH has got it wrong. That is not a scientific method but that is what the NIH requires if you write a grant proposal. The scientific method is observation and then hypothesis.
Without observations random hypotheses are being formed based on nothing and we will never get there. What you want is an observation and that observation leads to an idea and that is the hypothesis. You are testing the hypothesis based on something that has been observed. That is how all science is done.
Why they say that when you write a grant you test a hypothesis and inevitably you do new observations and that is the basis of the next experiment and that can keep going on and on. What happens when you start a new disease? You have to make observations and then you can generate hypotheses.
Dr Nath’s talk showed that this is exactly what he is doing. He is making lots and lots of observations. There is no hypothesis there. So the idea is to create as much data as they can and that is what they are doing and that is why it is taking so long to even talk about it. It is a massive undertaking to analyse the data.

Findings so far
They have found that CFS is negatively correlated with depression. They also tested cell free DNA for traces of viruses that patients tell they have or had when they first became ill. Even if the organism is not in the blood their DNA will be.
They tested multiple regions of the viruses using a multiplex assay. The result was there were no viral infections that were different from healthy controls or that the healthy controls had more viruses that ME/CFS patients.
Dr Davis said that the result makes him suspect that patients have something else that feels like a viral infection. Inflammation for example can make people feel as if they had a viral infection. Most of the viruses they tested for do not do anything by themselves.
It is the body itself that does the signalling to tell that there is something wrong. The reason Dr Davis was stressing this point was to tell that if patients do not have a viral infection they should not be taking antivirals.
It is not healthy. The reason being that antivirals generally target the synthesis of the DNA from the virus and it works because they are very primitive DNA preliminaries. They can be inhibited without inhibiting the human preliminaries but the problem is that mitochondrial preliminaries are primitive also and some of the antivirals can inhibit mitochondrial preliminaries.
Given the fact that there is some evidence of lack of mitochondrial activity in ME/CFS it is probably not a good thing to take antivirals. Dr Davis thinks patients should not take anything they do not need.

New Probing
Dr Davis and his team are also doing new probing as they have found that ME/CFS patients’ gene expression is similar to the expression from trypanosome (Sleeping sickness). The symptoms for Sleeping sickness resemble those of ME/CFS. One notable symptom is sleep reversal and that is common in ME/CFS too. This discovery led to the decision to develop multiple DNA probes against all known trypanosomes and parasites.

The West African trypanosome is very easy to detect in the blood but the East African one is very difficult to find. They also hope to make probes against RNA viruses that is not easy as RNA is not stable in the blood. This approach would allow them to look at a lot of infecting organisms. There are a lot of possibilities and maybe it is a new virus involved in ME/CFS and there was the excitement with XMRV. They have done a new microbe discovery project once and that came out empty.

Heavy Metals
Some patients have told Dr Davis they have heavy metal contamination and they need to detox from the heavy metals. The team have done urine analysis of all the severely ill patients and have found that they do not have heavy metals. However, they found that these patients are low in essential metals.
They have a hypothesis of this. One is that patients are low in essential metals as they are detoxing and removing essential metals. The way the urine tests were done did not detect mercury so the team took some new patients to test this by doing a hair analysis.
They found that high mercury is associated with low selenium (antioxidant) and the patients that had higher mercury levels had got it through eating a lot of fish. A little bit of a surprise was a patient from Finland who had a fair amount of uranium which is probably environmental and they do not know the medical consequences of this.

Gene Expression
The gene expression study in the severely ill patients showed a lot of changes in gene expression in the immune system and that means there is a lot of immunology going on and the team are not surprised by that. These same patients have high cytokines as well.

The team also want to look at the autoimmune system and what they are looking for is T-cell activation. They are now adding single cell analysis to this project. T-cells eliminate cells that are infected with any pathogen. If the T-cells find something foreign in the body they amplify and this is called T-cell expansion.
They have found T-cell expansion in ME/CFS and that means T-cells are recognising something. Dr Davis’s team have looked for a foreign agent but have not found anything. The T-cells could also be seeing self and that would be an autoimmune disease.
It could also be something else that is triggering the immune system and then it would not be an autoimmune disease.

Whole Genome Sequencing
Dr Davis’s team have also done whole genome sequencing. There are some genes that are different. Some of them are involved in NK-cell recognition. They are now going to do HLA and KIR sequencing. They continue to explore DNAH11, FAM20C, KIRs and NRXN1 (neurexin, a very large gene expressed in the brain).
They also have found a lot of metabolic changes and you see a lot when you look at different small molecules in the blood. They see activation of the immune system, a lot of genetic and metabolic changes. They have also done the microbiome.

Future Work
Looking at metabolic traps (combining metabolite and genomic data taking a systems biology approach). The idea is that due to genetics and inflammation that gets started the pathways are not working as they should due to mutations. If this is true it is probably easy to fix according to Dr Davis. This has not been tried before. It should be inexpensive and should only take a few days to get the patient out of this “metabolic trap”. This research is not aimed at getting a publication out but trying to figure out what it is. Dr Davis wants everyone to start thinking this way to help find a solution. Dr Davis is optimistic.

Question: There have been a number of reports that cholesterol is high in many ME patients even though they are non- obese. Cholesterol could change in the discourse of the blood due to temperature (from 37 to 34 degrees) changes. Could that change affect the impedance measurements in your instrument?

Answer: The disease affects a lot of lipids, metabolism and that is why the white cells are light and that is why a lot of the metabolites are lipid containing. They do not know why this is but one thing they see is a fair amount of mutations in fat metabolism in the patients compared to healthy controls. If one cannot metabolise fats they accumulate and then you get feedback systems and it affects all sorts of other fats as well. You ought to get rid of them so you may not be able to oxidise them in the mitochondria and you may have to oxidise them in ER.

Question continued: What might be a good control group that would mimic ME patients. Perhaps a group of people with high cholesterol levels?

Answer: People with high cholesterol would probably be genetically determined and it is probably very narrow in terms of the lipid problems they have. It also looks like a lot of the glucose is shifted into the sorbitol pathway which then goes to fructose and then to fatty acid synthesis so your body is even making fat. There is no understanding of this at the moment but there is something that is off in the lipids in the patients.

Question relating to improvement under fever conditions that has been also observed in autism (speaking during fever but not otherwise) and the other question relating to red cell deformability. In sickle cells there are certain processes that are oxygen dependent and some that are not. Do you or can you control oxygen sensing in your system?

Answer: There is a close relationship to autism. The metabolites look very similar. There is obviously a difference in onset time. Bob Naviaux has treated some autistic children with Suramin that blocks the purinergic receptor and he got a great deal of improvement. Dr Davis has been trying to get hold of Suramin but he cannot. However, a new manufacturing plant is being set up in the US and it may be available by the end of the year. The nice thing is that Suramin is not very toxic and curiously, it is also used for trypanosomes.

Question: Have you got plans of trialling any of the drugs tested on the impedance device on patients?

Answer: We are trying drugs that are FDA approved but as we are not physicians we cannot so we partner with other physicians and that would be something we are going to discuss with Ron Tompkins. We found one drug that seems to improve the ATP production and that is a drug called Adavan. It helps but only for a few hours and it is used for crisis management only.
There was a general question related to what was needed most to make progress. Dr Davis said money to pay for staff salaries for at least two years ahead. There is a lot of head hunting going on and if people do not know about their future salaries for longer than 6 months ahead they move on.