Phoenix Rising Team submitted a new blog post:
Dr. Kerr, I presume?
Clark Ellis brings us a rare interview with British researcher Dr. Jonathan Kerr who is now living in Colombia.
Dr. Jonathan Kerr is regarded as one of Britain’s foremost myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) researchers, and an expert in microbiology, inflammation and genetics.Formerly of St. George’s University of London, he is now working at the Escuela de Medicina y Ciencias de la Salud, Universidad del Rosario, Bogota, Colombia.
Dr. Kerr kindly agreed to be interviewed, with topics including his recent publication, which identified SNPs associated with ME/CFS, his prior research of the disease, diet and its role in health, disease definitions, and why he dropped off the ME/CFS scene for a little while.
Genetics Work
Clark:
Before we go into detail on your latest findings, let’s review your prior work which is the foundation for your latest publication. Your first paper on gene expression in ME/CFS came in 2005, where you reported differential gene expression in ME/CFS and this attracted significant interest and was even covered by the BBC.
Building on that work, in 2008 you reported that you had identified 88 genes which were being differentially expressed, compared to healthy controls. And importantly, in that paper, having identified these genes, you then checked a second group of patients to see if they showed the same pattern of gene expression, which they did.
What’s more, clustering analysis of those genes suggested seven distinct subtypes within the ME/CFS patients, and these subgroupings exhibited markedly different symptoms and severity of illness, suggesting that a gene-based method may be useful for clinically meaningful subtyping of ME/CFS.
This work looked highly promising. There was talk of trialling interferon-beta and one of the anti-tumour necrosis factor-alpha drugs. Patients, advocates and researchers began to get excited.
But then you reported, in 2011, that you had put these gene signatures to a blinded test of new samples and unfortunately they did not robustly identify the ME/CFS patients in the sample. Although it worked for around two-thirds of samples, there was also a high rate of misclassification.
Science is like that, and too often people aren’t brave enough to report the negative result. I feel a bit gutted about it though – you must have been disappointed yourself?
Dr. Kerr:
An inherent difficulty in gene expression studies is that their results are expressed as relative values (relative to the levels of gene expression in a comparison group), and this is further "relativised" to expression of cellular control genes.
As compared with studies of DNA mutations, or single nucleotide polymorphisms (SNPs) in which results are absolute (homozygous A, homozygous B or heterozygous, for each locus), results of gene expression studies are quite messy, although that is not to detract from their utility in elucidating pathogenetic mechanisms.
The study using genes to predict the class of new samples, which found that only about two-thirds of samples could be classified correctly, was a test of whether these gene expression values could be used as a diagnostic test, and they clearly can not be successfully used in this way.
A large confounding factor in any study of ME/CFS is the existence of subtypes, and we have clear evidence of the existence of these subtypes in the actual gene expression values of this study, as well as the clinical symptoms, showing that the already "messy" gene data is made even more so by the fact that these ME/CFS patients were a heterogeneous group.
The third issue was the sample sizes used in our studies, which were small, due to the expensive nature of the methods used. So, it seems to me a success that a subset of these genes could have had a success rate of two-thirds given the problems I have outlined.
But, this does not detract from the insight gained in our gene studies, towards an understanding of the pathogenesis of ME/CFS, and this is an important distinction. Understanding of pathogenesis was always the goal of these gene expression studies.
We undertook separate studies using SELDI-TOF [a] and SNPs in order to develop a diagnostic test, as these methods are much more suited to that goal, as they are much less relative.
(The SELDI-TOF study did not yield useful biomarkers, and the SNP study has just been published, which we’ll discuss in more detail in a moment).
Clark:
You stated, back in 2007, the importance of ensuring that once a gene signature was identified, it could be proven to be specific to CFS and not also present in other diseases. Three years later you reported the same signature in patients with Q-fever-associated ME/CFS, while other control groups showed different signatures.
Do you think the gene signature was specific to ME/CFS or would you have liked to test it against other, more distinct, diseases?
Dr. Kerr:
We also tested these genes in well-defined patients with endogenous depression, and found that the gene signature was quite different from that in ME/CFS. Endogenous depression is the most important group to test in this regard, and the fact that this group has a different signature, is further evidence against a psychological basis for ME/CFS.
The fact that Q-fever-triggered ME/CFS has the same signature as idiopathic ME/CFS is to be expected, given the fact that the trigger of the disease occurred at least 6 months prior to the diagnosis of ME/CFS.
In the case of idiopathic ME/CFS, various triggers are involved, and by definition, there is little knowledge of which were involved in a given patient at the time of diagnosis. In the few cases of parvovirus B19-associated ME/CFS that we tested, a typical idiopathic ME/CFS profile of gene expression was also found.
New Paper
Clark:
Although the gene-sets studied failed to work out as a robust diagnostic tool, it was still of interest that each gene on its own performed better than a random predictor ... This suggested that these genes were still relevant to the disease, and that a revised approach would be needed to improve the diagnostic potential of this method.
You mentioned already that gene expression values are relative, and that SNP testing is absolute. That’s what led to your latest study, right?
Dr. Kerr:
That´s correct. As I have explained above, it is extremely difficult to develop a diagnostic test using gene expression data, and much more feasible to do so using SNPs, simply by the nature of the science in each case.
Clark:
Your latest study then, published in the Journal of Clinical Pathology, and funded by ME Research UK, studied 108 ME/CFS patients, 17 people with endogenous depression, and 68 healthy controls.
You looked at 504 SNPs and reported that 21 SNPs had a significant and unexpected distribution across ME/CFS, depression and normal groups. And that 10 of these 21 SNPs’ alleles are associated with ME/CFS when compared against healthy controls.
How did you choose which SNPs to investigate?
Dr. Kerr:
The starting point was the 88 human genes that we found to be differentially expressed in ME/CFS using the Affymetrix U133+2 comprehensive gene array [a], and which were confirmed using real-time PCR.
We studied up to 8 SNPs per ME/CFS-associated gene among these 88 genes for which a validated PCR test was available. This approach resulted in a total of 504 SNPs.
Clark:
Although 21 alleles out of 504 SNPs is perhaps a lower than expected return, given that these were selected from genes already associated with ME/CFS, you think this is down to the heterogeneous nature of ME/CFS?
Dr. Kerr:
Yes, I believe that the heterogeneity of ME/CFS is one of the most difficult problems that we face in this research, because all ME/CFS clinicians would agree that such subtypes exist and are important for understanding of pathogenesis and management, but an effective means to subclassify patients has yet to be found.
So, each study potentially suffers from this issue. But the eventual benefits of arriving at a meaningful method of patient subtyping will be enormous.
We have tried to address it by looking at each of 8 subtypes in this SNP study, despite the fact that in doing so, the number of patients in each group is reduced. But nonetheless, we found 148 SNPs that had significant differences in prevalence across each of the 8 subtypes.
The distribution of particular SNP alleles across particular ME/CFS subtypes is interesting, and shows that each subtype is associated with particular SNP alleles, and that these SNP alleles are grouped within particular genes for each ME/CFS subtype. Such that, for example, SNP alleles within the gene ACTR3 are associated with subtype F, while SNP alleles within the gene AKAP10 are associated with subtype G. (Supplementary Table)
Clark:
You also established that 4 of the 21 alleles associated with ME/CFS are believed to influence gene expression, and 3 of these occur across all the disease subgroups. This seems like it could be pretty significant as it ties all your genetics research together. Can you expand on this and tell us what you think?
Dr. Kerr:
We looked for SNPs that occurred in the binding sites of probes used in the Affymetrix U133+2 array, and only 4 occurred within probe sequences which were important in our previous gene array study (Kerr et al, 2008).
Basically, we found this reassuring that only 4 SNPs have the potential to modify the results of gene array testing in the context of ME/CFS (reassuring that our gene expression signature was still valid). In addition, these transcription factors have numerous binding sites in the promoter regions of these ME/CFS-associated genes.
So it seems unlikely that these SNPs would result in significant differences in the results of gene expression analysis, particularly as real-time PCR was used as well as microarrays, and because each of these utilised different sequences within each individual gene.
Clark:
Looking at the p-values (table 2) in your statistical analysis of the SNPs (table 2) they’re pretty low, comfortably below the threshold of 0.05 that you set for significance. You’ve mentioned already the disadvantage of having relatively small sample numbers, and the inherent problems with disease heterogeneity. What’s the risk of these results being statistical anomalies?
Dr. Kerr:
A good question. According to the statistical tests, it is most unlikely that we are wrongly accepting these 21 SNPs as associated in some way with ME/CFS. But, until such associations are reproduced independently, it would not be wise to infer too much.
But, having said that, these SNPs were selected for testing in the first place as we found their respective transcripts to be differentially expressed using 2 methods and in 2 studies (some were tested in 3 studies). So, the fact that some of these SNPs are associated at all with CFS appears to be quite interesting. But, as always, more work is required!
Clark:
If your findings were replicated, do you think these SNPs could be used to produce a diagnostic test for ME/CFS, able to robustly differentiate patients from controls?
Dr. Kerr:
It is possible, although I have not undertaken this study as yet. If it is possible to obtain funding for this study, then it would be fairly straightforward to test that hypothesis.
Disease Definitions
Clark:
Your study participants met the Fukuda criteria with some additional exclusions applied. Given the difficulties with the heterogeneous nature of ME/CFS, as already discussed, do you think the Fukuda criteria are up to the task and what’s your view on the IOM and P2P processes going on at the moment which are very unpopular with patients?
Dr. Kerr:
These new initiatives appear to represent a further dumbing down of the research response to CFS, and is consistent with what has happened in the UK. I believe this is revealed by the fact that non-experts are included in panels, and in P2P, panels consist entirely of non-experts.
Non-experts will have different and varied priorities which are unlikely to be linked to CFS. The case definition used in P2P is too broad and does not recognise heterogeneity and subtypes, which we know are key to the study of CFS.
The NIH states that the panel is unbiased, but the truth is that everyone is biased, and I would have thought that was the point, in that those present should actually CARE about the process and the final outcome for ME/CFS patients. Otherwise, they will be considering the material WITHOUT a focus on the biological understanding and treatment of patients with ME/CFS.
Parvovirus
Clark:
Although ME/CFS is considered to be a disease of unknown aetiology, several pathogens are known to trigger the disease. These include enteroviruses, Epstein-Barr virus, Giardia lamblia, Q-fever, and Parvovirus B19. This is where I understand you first became interested in ME/CFS while researching parvovirus, where you showed that some patients went on to develop the condition after the infection?
Dr. Kerr:
I believe this approach is hugely valuable because these infections represent models of pathogenesis of ME/CFS, in which the time course of infection and specific immune response can be studied, in order to understand which factors are important in deciding whether a patient controls the infection and recovers, or progresses to development of ME/CFS.
In addition, these models are important because most of these infections have specific treatments which have been shown in several studies to benefit patients with ME/CFS, where the microbial trigger can be identified."
Diet
Clark:
What many readers may not know, is that food and, it’s relationship with health, is a topic you are passionate about. Many people with ME/CFS have gut dysfunction and the gut microbiota is a relatively new and promising area of research into the disease. How did you become interested in this area?
Dr. Kerr:
My interest in food and its effect on health began when I discovered the incredible work of Weston Price, who was a dentist from Canada in the 1930s, who discovered that diet was key to dental and physical health.
Basically, a modern processed food diet is responsible for dental caries, physical degeneration and disease and even susceptibility to tuberculosis. In addition, congenital abnormalities such as cleft palate / lip were associated with a modern processed food diet of the mother during pregnancy [1, 2, 3].
Weston Price travelled the world to study the diet and health of isolated tribes, and found that, in general, people who avoided eating modern processed food, such as the tribes, had excellent overall health and physical development and an absence of dental decay.
Clark:
So what processed foods should be avoided?
Dr. Kerr:
Modern processed foods are easily remembered in four main groups:
GrainsFirst, poorly processed grains (wheat, corn, beans, oats, etc). In the distant past, prior to eating, grains were soaked in water for 48 hours; partly to remove toxins (from beans, for example) and partly to remove so-called ´anti-nutrients´, which are normal contents of grains, but which prevent the proper absorption of the nutrients contained in the grains.
In addition, soaking removes lectins from the grains which result in the micro-perforation of the gut wall, causing leaky gut. 80 years ago, the Quaker oat box advised soaking for 48 hours, but this advice has now been removed.
Another problem for the grains, is genetic modification (GM). In large parts of the world, including the USA, wheat, rice and corn are GM. This is not good for health and is actually bad. GM foods have been shown to be less nutritious and even harmful. When given the choice, laboratory animals routinely avoid eating GM foods of different types.
A particular problem is the identification of GM foods in the supermarket, because in some areas (e.g., USA), proper labelling is unfortunately not mandatory, and so bread, cereal, cake, for example, may be GM in the absence of a GM label.
Pasteurised DairySecond, pasteurisation of milk and cream. We have been trained to be terrified of the bacteria which occur in milk, and also of the possible presence of mycobacteria and brucella species, and this is how pasteurisation was sold to the public.
Milk proteins are large and complex and are extremely good for health and gaining and maintaining weight, and for proper immune function. Unpasteurised cream is especially good for health, with extremely high levels of vitamins A and D, and a lot of beneficial saturated fat (see below).
However, pasteurisation fragments these proteins and destroys their benefits. Attention to the health and hygiene of cows, along with testing for the presence of pathogens, virtually excludes the potential for human infection with these pathogens.
Although it is illegal to sell raw milk in many parts of the world, a personal relationship with a farmer can provide the necessary access. The Weston Price Foundation website provides information on how to buy raw milk, some of which is organic, in the UK and the USA. Apart from the health benefits, it is really delicious, as food once was!
Modern Vegetable OilsThird, polyunsaturated vegetable oils. These have been marketed as the healthy alternative to the "damaging" saturated fats, such as butter, lard, tallow, etc. But they are not healthy, because they contain free electrons which are donated to our tissues and result in cellular oxidation and damage. These "healthy" oils have been heated to high temperatures in processing thus destroying virtually all omega-3 components, and even worse, may contain chemicals used for extraction (e.g., hexane).
The studies which supposedly prove that saturated fat is damaging and linked with heart disease and other degenerative disease, do not actually prove this at all. And, recently, it is becoming widely recognised that there is no evidence that saturated fats are bad for health! In fact, it is the opposite. They are good for health in a multiplicity of ways. Our bodies are actually made of saturated fat, and so to eat more of what we are made of, makes perfect sense.
The bad press that cholesterol has received is in a similar category. Once the pariah of food, it is now recognised to be anti-oxidant, heart protective and supremely good for health. Cholesterol is used in cell walls, and is critical for brain development and efficient brain functioning.
We have been told that 4 eggs per day is much too much cholesterol to be healthy, and cholesterol screening purports to be helpful in this regard. However, increased dietary cholesterol actually decreases blood cholesterol, the body fluid of choice for cholesterol monitoring.
One hundred years ago in the USA, there was no modern processed food, and there was virtually no heart disease. Both modern processed food and heart disease are new phenomena, but we are supposed to believe that it is the old-fashioned diet that is to blame. This is ludicrous. The truth is the opposite.
Refined SugarAnd fourth, refined sugar and fructose. Ingestion of sugar results in very high insulin levels as the body attempts to utilise the sugar and control the level of circulating glucose, and this is damaging in that continual ingestion of sugar leads to a reduced cellular responsiveness to insulin (or insulin resistance).
This insulin resistance is the basis of type 2 diabetes mellitus (90% of diabetes mellitus) and leads to increased insulin secretion and more extreme insulin resistance. The incidence of type 2 diabetes has skyrocketed in the last 60 years, and it is due to ingestion of sugar.
It has been shown that in intensive care patients, those who have high average glucose levels during their stay, have a much higher rate of complications than those whose glucose is well controlled.
Diabetic patients, and especially those whose glucose is poorly controlled, suffer lack of energy and rapid aging, and the problem is poor control of glucose, with high levels of insulin, which results in these problems.
Another very damaging sugar is fructose, and especially high-fructose corn syrup (HFCS) which is used in many processed foods and drinks, and is even worse than glucose, because there is no metabolic mechanism for the utilisation of fructose in our bodies.
It is important to mention here that sugar substitutes, such as saccharin, aspartame, and the others, are also toxic for health, in that they decimate the gut flora in terms of numbers of bacteria, and also the diversity of bacterial species. This is very damaging and adversely affects gut function, and overall body functioning, as a healthy gut flora is very important for every system.
Junk Food – all four bad food groups in one
All junk food consists of 2 or more of these 4 bad food groups. For example, cookies have poorly processed wheat (maybe GM wheat), polyunsaturated vegetable oil, and sugar. Pizza bases are usually made of wheat (maybe GM wheat) and polyunsaturated vegetable oil. French fries are cooked in polyunsaturated vegetable oils (although they used to be cooked in saturated fats, such as tallow (beneficial fat extracted from the fatty tissue around the cow´s kidney) .. even McDonald's used tallow for their french fries in the 1960s).
Unless you know the butcher, it is no longer possible to buy the fatty tissue from which tallow is made. White bread is made from poorly processed, refined and bleached wheat (maybe GM wheat). Some examples of bleaching agents used for this are oxide of nitrogen, chlorine, chloride, nitrosyl and benzoyl peroxide. These are either harmful in themselves or result in production of other toxins, which are then ingested.
Back to an old-fashioned dietWeston Price documented in multiple different tribes that a healthy diet consisted of high levels of saturated fat (up to 80% of the diet by calories), meat, pork, chicken, fish, eggs, unpasteurised milk & cream, butter, lard, vegetables, a little fruit, properly processed grains (and in many cases fermented grains, which are great for gut health), and other fermented products (for example, kefir and sauerkraut). And, no sugar, sugar substitutes, or other processed foods.
Those who try this old-fashioned diet, will be amazed at how good it tastes, like the food made by our grandmothers, and will realise health benefits that have formerly been elusive, and may even cure chronic diseases, which were not known previous to the introduction of the modern diet.
Clark:
I think many readers will be open to the idea of a more natural diet, and its potential health benefits. But isn’t it a little late once you’ve got a chronic disease like ME/CFS?
Dr. Kerr:
Absolutely not, in my opinion. We may have setbacks, but if we don´t properly address problems that can be identified, things will surely get worse. ME/CFS can be cured and that has been shown. Diet is critically important, and it is easy to see why. We eat food several times per day, and it is the balance of effects, good and bad, of whatever we eat at each meal, which determines whether the food is an overall benefit or detriment.
Until people try this old-fashioned diet, they will never know how good it is, and how good it can make you feel. We have been trained to ignore food quality; a quick bite and then back to work. But, it has only been that way recently, and it is neither an enjoyable nor healthful situation.
Clark:
Because of sufferers limited resources, eating a healthy meal can be very challenging. What easy things do you think people can do to improve what they eat within these constraints?
Dr. Kerr:
It all starts with education about food, and once you have that, you can exclude the bad things (improperly processed grains, pasteurised milk, polyunsaturated vegetable oil and sugar) and at the same time include some good things (eggs, saturated animal fat, organic vegetables).
One very big problem is that most people believe we need to get energy from sugar and carbohydrate. Energy from carbohydrate is not as efficient or long-lasting as energy from good fats, such as eggs, fatty meat, pork and bacon.
Lack of time is not the problem, it is lack of understanding, because a lot of good foods are actually widely available, but we don´t realise which are good and which are bad.
I am witness to the health benefits of this diet after a lifetime of eating what is now ubiquitously accepted as normal food. Good food is more difficult to obtain and prepare, but it is certainly possible to do so with little difference in cost, a huge increase in food enjoyment, and most importantly, a huge increase in bodily health.
The most important thing required to eat well is an education in the area of food. And, the simplest way to obtain that is to go to Dr. Mercola's excellent and extensive website (www.mercola.com) and follow your interest.
If you try this diet, I guarantee that you will not be disappointed, as it consists of foods that we all enjoy. In fact, this real old-fashioned food is the real medicine, both as a preventative and as a treatment.
What Next?
Clark:
Patients have always appreciated your strong and committed work, and your past colleagues speak admirably of you. The availability of funding for ME/CFS has already been an issue and continues to be; even Professor Lipkin, who you would think would have no trouble landing research funding for his microbiome study, was turned down.
From the outside at least, your ME/CFS research seemed to come to a bit of a halt a few years ago and came with the news that you had left your position at St George’s hospital, London. Did these inherent problems with research funding in ME/CFS make it difficult to carry out the research you wanted to? What happened?
Dr. Kerr:
Yes, it is difficult to obtain funding for any research, and even more difficult to obtain it for ME/CFS research.
After the death of David Tyrrell, the CFS Research Foundation made it a priority to join with the MRC in funding CFS research of a combined biological and psychological nature.
Around the same time, Judy Mikovits and I were awarded $2 million from NIH to study biological aspects of CFS in the UK and USA. I resigned from this grant because of the increasing controversy surrounding XMRV, and the fact that my lab could not detect XMRV in any CFS patient.
Judy's lab found it in a majority of patients, including samples which we found to be repeatedly negative, using the identical PCR primers, while she was actually offering the same test privately.
With these two grant incomes gone, there was insufficient funding to sustain the research.
Clark:
You’re now working at the Universidad del Rosario, Bogota, Colombia - are you still involved with research into the disease and if so then what do we have to look forward to?
Dr. Kerr:
I plan to look at several CFS-associated genes in more detail, to continue the Parvovirus B19 work and to explore ways in which the role of diet can be studied with regard to immune function/ dysfunction.
Clark:
Thank you, Dr. Kerr.
Clark Ellis writes about ME/CFS research and is writing a fiction novel/screenplay with themes related to ME/CFS. You can follow him on twitter to hear about his work, ME/CFS research, fundraising and advocacy.
FURTHER INFORMATION AND REFERENCES:
ME/CFS SNP paper
Shimosako N, Kerr JR.
Use of single-nucleotide polymorphisms (SNPs) to distinguish gene expression subtypes of chronic fatigue syndrome/myalgic encephalomyelitis (ME/CFS). J Clin Pathol
2014 Sep 19. pii: jclinpath-2014-202597. doi: 10.1136/jclinpath-2014-202597. [Epub]
1. Folic acid supplements and risk of facial clefts: national population based case-control study
2. Maternal dietary intake of vitamin A and risk of orofacial clefts: a population-based case-control study in Norway.
3. Maternal Western dietary patterns and the risk of developing a cleft lip with or without a cleft palate
Book of Weston Price, ´Nutrition and Physical Degeneration´ (available free as PDF)
Book & reviews available on Amazon
Wikipedia page on Weston Price
The Weston Price Foundation website
Dr Mercola´s website
Saturated fat, by Dr Mercola
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Continue reading the Original Blog Post
Dr. Kerr, I presume?
Clark Ellis brings us a rare interview with British researcher Dr. Jonathan Kerr who is now living in Colombia.
Dr. Jonathan Kerr is regarded as one of Britain’s foremost myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) researchers, and an expert in microbiology, inflammation and genetics.Formerly of St. George’s University of London, he is now working at the Escuela de Medicina y Ciencias de la Salud, Universidad del Rosario, Bogota, Colombia.
Dr. Kerr kindly agreed to be interviewed, with topics including his recent publication, which identified SNPs associated with ME/CFS, his prior research of the disease, diet and its role in health, disease definitions, and why he dropped off the ME/CFS scene for a little while.
Genetics Work
Clark:
Before we go into detail on your latest findings, let’s review your prior work which is the foundation for your latest publication. Your first paper on gene expression in ME/CFS came in 2005, where you reported differential gene expression in ME/CFS and this attracted significant interest and was even covered by the BBC.
Building on that work, in 2008 you reported that you had identified 88 genes which were being differentially expressed, compared to healthy controls. And importantly, in that paper, having identified these genes, you then checked a second group of patients to see if they showed the same pattern of gene expression, which they did.
What’s more, clustering analysis of those genes suggested seven distinct subtypes within the ME/CFS patients, and these subgroupings exhibited markedly different symptoms and severity of illness, suggesting that a gene-based method may be useful for clinically meaningful subtyping of ME/CFS.
This work looked highly promising. There was talk of trialling interferon-beta and one of the anti-tumour necrosis factor-alpha drugs. Patients, advocates and researchers began to get excited.
But then you reported, in 2011, that you had put these gene signatures to a blinded test of new samples and unfortunately they did not robustly identify the ME/CFS patients in the sample. Although it worked for around two-thirds of samples, there was also a high rate of misclassification.
Science is like that, and too often people aren’t brave enough to report the negative result. I feel a bit gutted about it though – you must have been disappointed yourself?
Dr. Kerr:
An inherent difficulty in gene expression studies is that their results are expressed as relative values (relative to the levels of gene expression in a comparison group), and this is further "relativised" to expression of cellular control genes.
As compared with studies of DNA mutations, or single nucleotide polymorphisms (SNPs) in which results are absolute (homozygous A, homozygous B or heterozygous, for each locus), results of gene expression studies are quite messy, although that is not to detract from their utility in elucidating pathogenetic mechanisms.
The study using genes to predict the class of new samples, which found that only about two-thirds of samples could be classified correctly, was a test of whether these gene expression values could be used as a diagnostic test, and they clearly can not be successfully used in this way.
A large confounding factor in any study of ME/CFS is the existence of subtypes, and we have clear evidence of the existence of these subtypes in the actual gene expression values of this study, as well as the clinical symptoms, showing that the already "messy" gene data is made even more so by the fact that these ME/CFS patients were a heterogeneous group.
The third issue was the sample sizes used in our studies, which were small, due to the expensive nature of the methods used. So, it seems to me a success that a subset of these genes could have had a success rate of two-thirds given the problems I have outlined.
But, this does not detract from the insight gained in our gene studies, towards an understanding of the pathogenesis of ME/CFS, and this is an important distinction. Understanding of pathogenesis was always the goal of these gene expression studies.
We undertook separate studies using SELDI-TOF [a] and SNPs in order to develop a diagnostic test, as these methods are much more suited to that goal, as they are much less relative.
(The SELDI-TOF study did not yield useful biomarkers, and the SNP study has just been published, which we’ll discuss in more detail in a moment).
Clark:
You stated, back in 2007, the importance of ensuring that once a gene signature was identified, it could be proven to be specific to CFS and not also present in other diseases. Three years later you reported the same signature in patients with Q-fever-associated ME/CFS, while other control groups showed different signatures.
Do you think the gene signature was specific to ME/CFS or would you have liked to test it against other, more distinct, diseases?
Dr. Kerr:
We also tested these genes in well-defined patients with endogenous depression, and found that the gene signature was quite different from that in ME/CFS. Endogenous depression is the most important group to test in this regard, and the fact that this group has a different signature, is further evidence against a psychological basis for ME/CFS.
The fact that Q-fever-triggered ME/CFS has the same signature as idiopathic ME/CFS is to be expected, given the fact that the trigger of the disease occurred at least 6 months prior to the diagnosis of ME/CFS.
In the case of idiopathic ME/CFS, various triggers are involved, and by definition, there is little knowledge of which were involved in a given patient at the time of diagnosis. In the few cases of parvovirus B19-associated ME/CFS that we tested, a typical idiopathic ME/CFS profile of gene expression was also found.
New Paper
Clark:
Although the gene-sets studied failed to work out as a robust diagnostic tool, it was still of interest that each gene on its own performed better than a random predictor ... This suggested that these genes were still relevant to the disease, and that a revised approach would be needed to improve the diagnostic potential of this method.
You mentioned already that gene expression values are relative, and that SNP testing is absolute. That’s what led to your latest study, right?
Dr. Kerr:
That´s correct. As I have explained above, it is extremely difficult to develop a diagnostic test using gene expression data, and much more feasible to do so using SNPs, simply by the nature of the science in each case.
Clark:
Your latest study then, published in the Journal of Clinical Pathology, and funded by ME Research UK, studied 108 ME/CFS patients, 17 people with endogenous depression, and 68 healthy controls.
You looked at 504 SNPs and reported that 21 SNPs had a significant and unexpected distribution across ME/CFS, depression and normal groups. And that 10 of these 21 SNPs’ alleles are associated with ME/CFS when compared against healthy controls.
How did you choose which SNPs to investigate?
Dr. Kerr:
The starting point was the 88 human genes that we found to be differentially expressed in ME/CFS using the Affymetrix U133+2 comprehensive gene array [a], and which were confirmed using real-time PCR.
We studied up to 8 SNPs per ME/CFS-associated gene among these 88 genes for which a validated PCR test was available. This approach resulted in a total of 504 SNPs.
Clark:
Although 21 alleles out of 504 SNPs is perhaps a lower than expected return, given that these were selected from genes already associated with ME/CFS, you think this is down to the heterogeneous nature of ME/CFS?
Dr. Kerr:
Yes, I believe that the heterogeneity of ME/CFS is one of the most difficult problems that we face in this research, because all ME/CFS clinicians would agree that such subtypes exist and are important for understanding of pathogenesis and management, but an effective means to subclassify patients has yet to be found.
So, each study potentially suffers from this issue. But the eventual benefits of arriving at a meaningful method of patient subtyping will be enormous.
We have tried to address it by looking at each of 8 subtypes in this SNP study, despite the fact that in doing so, the number of patients in each group is reduced. But nonetheless, we found 148 SNPs that had significant differences in prevalence across each of the 8 subtypes.
The distribution of particular SNP alleles across particular ME/CFS subtypes is interesting, and shows that each subtype is associated with particular SNP alleles, and that these SNP alleles are grouped within particular genes for each ME/CFS subtype. Such that, for example, SNP alleles within the gene ACTR3 are associated with subtype F, while SNP alleles within the gene AKAP10 are associated with subtype G. (Supplementary Table)
Clark:
You also established that 4 of the 21 alleles associated with ME/CFS are believed to influence gene expression, and 3 of these occur across all the disease subgroups. This seems like it could be pretty significant as it ties all your genetics research together. Can you expand on this and tell us what you think?
Dr. Kerr:
We looked for SNPs that occurred in the binding sites of probes used in the Affymetrix U133+2 array, and only 4 occurred within probe sequences which were important in our previous gene array study (Kerr et al, 2008).
Basically, we found this reassuring that only 4 SNPs have the potential to modify the results of gene array testing in the context of ME/CFS (reassuring that our gene expression signature was still valid). In addition, these transcription factors have numerous binding sites in the promoter regions of these ME/CFS-associated genes.
So it seems unlikely that these SNPs would result in significant differences in the results of gene expression analysis, particularly as real-time PCR was used as well as microarrays, and because each of these utilised different sequences within each individual gene.
Clark:
Looking at the p-values (table 2) in your statistical analysis of the SNPs (table 2) they’re pretty low, comfortably below the threshold of 0.05 that you set for significance. You’ve mentioned already the disadvantage of having relatively small sample numbers, and the inherent problems with disease heterogeneity. What’s the risk of these results being statistical anomalies?
Dr. Kerr:
A good question. According to the statistical tests, it is most unlikely that we are wrongly accepting these 21 SNPs as associated in some way with ME/CFS. But, until such associations are reproduced independently, it would not be wise to infer too much.
But, having said that, these SNPs were selected for testing in the first place as we found their respective transcripts to be differentially expressed using 2 methods and in 2 studies (some were tested in 3 studies). So, the fact that some of these SNPs are associated at all with CFS appears to be quite interesting. But, as always, more work is required!
Clark:
If your findings were replicated, do you think these SNPs could be used to produce a diagnostic test for ME/CFS, able to robustly differentiate patients from controls?
Dr. Kerr:
It is possible, although I have not undertaken this study as yet. If it is possible to obtain funding for this study, then it would be fairly straightforward to test that hypothesis.
Disease Definitions
Clark:
Your study participants met the Fukuda criteria with some additional exclusions applied. Given the difficulties with the heterogeneous nature of ME/CFS, as already discussed, do you think the Fukuda criteria are up to the task and what’s your view on the IOM and P2P processes going on at the moment which are very unpopular with patients?
Dr. Kerr:
These new initiatives appear to represent a further dumbing down of the research response to CFS, and is consistent with what has happened in the UK. I believe this is revealed by the fact that non-experts are included in panels, and in P2P, panels consist entirely of non-experts.
Non-experts will have different and varied priorities which are unlikely to be linked to CFS. The case definition used in P2P is too broad and does not recognise heterogeneity and subtypes, which we know are key to the study of CFS.
The NIH states that the panel is unbiased, but the truth is that everyone is biased, and I would have thought that was the point, in that those present should actually CARE about the process and the final outcome for ME/CFS patients. Otherwise, they will be considering the material WITHOUT a focus on the biological understanding and treatment of patients with ME/CFS.
Parvovirus
Clark:
Although ME/CFS is considered to be a disease of unknown aetiology, several pathogens are known to trigger the disease. These include enteroviruses, Epstein-Barr virus, Giardia lamblia, Q-fever, and Parvovirus B19. This is where I understand you first became interested in ME/CFS while researching parvovirus, where you showed that some patients went on to develop the condition after the infection?
Dr. Kerr:
I believe this approach is hugely valuable because these infections represent models of pathogenesis of ME/CFS, in which the time course of infection and specific immune response can be studied, in order to understand which factors are important in deciding whether a patient controls the infection and recovers, or progresses to development of ME/CFS.
In addition, these models are important because most of these infections have specific treatments which have been shown in several studies to benefit patients with ME/CFS, where the microbial trigger can be identified."
Diet
Clark:
What many readers may not know, is that food and, it’s relationship with health, is a topic you are passionate about. Many people with ME/CFS have gut dysfunction and the gut microbiota is a relatively new and promising area of research into the disease. How did you become interested in this area?
Dr. Kerr:
My interest in food and its effect on health began when I discovered the incredible work of Weston Price, who was a dentist from Canada in the 1930s, who discovered that diet was key to dental and physical health.
Basically, a modern processed food diet is responsible for dental caries, physical degeneration and disease and even susceptibility to tuberculosis. In addition, congenital abnormalities such as cleft palate / lip were associated with a modern processed food diet of the mother during pregnancy [1, 2, 3].
Weston Price travelled the world to study the diet and health of isolated tribes, and found that, in general, people who avoided eating modern processed food, such as the tribes, had excellent overall health and physical development and an absence of dental decay.
Clark:
So what processed foods should be avoided?
Dr. Kerr:
Modern processed foods are easily remembered in four main groups:
GrainsFirst, poorly processed grains (wheat, corn, beans, oats, etc). In the distant past, prior to eating, grains were soaked in water for 48 hours; partly to remove toxins (from beans, for example) and partly to remove so-called ´anti-nutrients´, which are normal contents of grains, but which prevent the proper absorption of the nutrients contained in the grains.
In addition, soaking removes lectins from the grains which result in the micro-perforation of the gut wall, causing leaky gut. 80 years ago, the Quaker oat box advised soaking for 48 hours, but this advice has now been removed.
Another problem for the grains, is genetic modification (GM). In large parts of the world, including the USA, wheat, rice and corn are GM. This is not good for health and is actually bad. GM foods have been shown to be less nutritious and even harmful. When given the choice, laboratory animals routinely avoid eating GM foods of different types.
A particular problem is the identification of GM foods in the supermarket, because in some areas (e.g., USA), proper labelling is unfortunately not mandatory, and so bread, cereal, cake, for example, may be GM in the absence of a GM label.
Pasteurised DairySecond, pasteurisation of milk and cream. We have been trained to be terrified of the bacteria which occur in milk, and also of the possible presence of mycobacteria and brucella species, and this is how pasteurisation was sold to the public.
Milk proteins are large and complex and are extremely good for health and gaining and maintaining weight, and for proper immune function. Unpasteurised cream is especially good for health, with extremely high levels of vitamins A and D, and a lot of beneficial saturated fat (see below).
However, pasteurisation fragments these proteins and destroys their benefits. Attention to the health and hygiene of cows, along with testing for the presence of pathogens, virtually excludes the potential for human infection with these pathogens.
Although it is illegal to sell raw milk in many parts of the world, a personal relationship with a farmer can provide the necessary access. The Weston Price Foundation website provides information on how to buy raw milk, some of which is organic, in the UK and the USA. Apart from the health benefits, it is really delicious, as food once was!
Modern Vegetable OilsThird, polyunsaturated vegetable oils. These have been marketed as the healthy alternative to the "damaging" saturated fats, such as butter, lard, tallow, etc. But they are not healthy, because they contain free electrons which are donated to our tissues and result in cellular oxidation and damage. These "healthy" oils have been heated to high temperatures in processing thus destroying virtually all omega-3 components, and even worse, may contain chemicals used for extraction (e.g., hexane).
The studies which supposedly prove that saturated fat is damaging and linked with heart disease and other degenerative disease, do not actually prove this at all. And, recently, it is becoming widely recognised that there is no evidence that saturated fats are bad for health! In fact, it is the opposite. They are good for health in a multiplicity of ways. Our bodies are actually made of saturated fat, and so to eat more of what we are made of, makes perfect sense.
The bad press that cholesterol has received is in a similar category. Once the pariah of food, it is now recognised to be anti-oxidant, heart protective and supremely good for health. Cholesterol is used in cell walls, and is critical for brain development and efficient brain functioning.
We have been told that 4 eggs per day is much too much cholesterol to be healthy, and cholesterol screening purports to be helpful in this regard. However, increased dietary cholesterol actually decreases blood cholesterol, the body fluid of choice for cholesterol monitoring.
One hundred years ago in the USA, there was no modern processed food, and there was virtually no heart disease. Both modern processed food and heart disease are new phenomena, but we are supposed to believe that it is the old-fashioned diet that is to blame. This is ludicrous. The truth is the opposite.
Refined SugarAnd fourth, refined sugar and fructose. Ingestion of sugar results in very high insulin levels as the body attempts to utilise the sugar and control the level of circulating glucose, and this is damaging in that continual ingestion of sugar leads to a reduced cellular responsiveness to insulin (or insulin resistance).
This insulin resistance is the basis of type 2 diabetes mellitus (90% of diabetes mellitus) and leads to increased insulin secretion and more extreme insulin resistance. The incidence of type 2 diabetes has skyrocketed in the last 60 years, and it is due to ingestion of sugar.
It has been shown that in intensive care patients, those who have high average glucose levels during their stay, have a much higher rate of complications than those whose glucose is well controlled.
Diabetic patients, and especially those whose glucose is poorly controlled, suffer lack of energy and rapid aging, and the problem is poor control of glucose, with high levels of insulin, which results in these problems.
Another very damaging sugar is fructose, and especially high-fructose corn syrup (HFCS) which is used in many processed foods and drinks, and is even worse than glucose, because there is no metabolic mechanism for the utilisation of fructose in our bodies.
It is important to mention here that sugar substitutes, such as saccharin, aspartame, and the others, are also toxic for health, in that they decimate the gut flora in terms of numbers of bacteria, and also the diversity of bacterial species. This is very damaging and adversely affects gut function, and overall body functioning, as a healthy gut flora is very important for every system.
Junk Food – all four bad food groups in one
All junk food consists of 2 or more of these 4 bad food groups. For example, cookies have poorly processed wheat (maybe GM wheat), polyunsaturated vegetable oil, and sugar. Pizza bases are usually made of wheat (maybe GM wheat) and polyunsaturated vegetable oil. French fries are cooked in polyunsaturated vegetable oils (although they used to be cooked in saturated fats, such as tallow (beneficial fat extracted from the fatty tissue around the cow´s kidney) .. even McDonald's used tallow for their french fries in the 1960s).
Unless you know the butcher, it is no longer possible to buy the fatty tissue from which tallow is made. White bread is made from poorly processed, refined and bleached wheat (maybe GM wheat). Some examples of bleaching agents used for this are oxide of nitrogen, chlorine, chloride, nitrosyl and benzoyl peroxide. These are either harmful in themselves or result in production of other toxins, which are then ingested.
Back to an old-fashioned dietWeston Price documented in multiple different tribes that a healthy diet consisted of high levels of saturated fat (up to 80% of the diet by calories), meat, pork, chicken, fish, eggs, unpasteurised milk & cream, butter, lard, vegetables, a little fruit, properly processed grains (and in many cases fermented grains, which are great for gut health), and other fermented products (for example, kefir and sauerkraut). And, no sugar, sugar substitutes, or other processed foods.
Those who try this old-fashioned diet, will be amazed at how good it tastes, like the food made by our grandmothers, and will realise health benefits that have formerly been elusive, and may even cure chronic diseases, which were not known previous to the introduction of the modern diet.
Clark:
I think many readers will be open to the idea of a more natural diet, and its potential health benefits. But isn’t it a little late once you’ve got a chronic disease like ME/CFS?
Dr. Kerr:
Absolutely not, in my opinion. We may have setbacks, but if we don´t properly address problems that can be identified, things will surely get worse. ME/CFS can be cured and that has been shown. Diet is critically important, and it is easy to see why. We eat food several times per day, and it is the balance of effects, good and bad, of whatever we eat at each meal, which determines whether the food is an overall benefit or detriment.
Until people try this old-fashioned diet, they will never know how good it is, and how good it can make you feel. We have been trained to ignore food quality; a quick bite and then back to work. But, it has only been that way recently, and it is neither an enjoyable nor healthful situation.
Clark:
Because of sufferers limited resources, eating a healthy meal can be very challenging. What easy things do you think people can do to improve what they eat within these constraints?
Dr. Kerr:
It all starts with education about food, and once you have that, you can exclude the bad things (improperly processed grains, pasteurised milk, polyunsaturated vegetable oil and sugar) and at the same time include some good things (eggs, saturated animal fat, organic vegetables).
One very big problem is that most people believe we need to get energy from sugar and carbohydrate. Energy from carbohydrate is not as efficient or long-lasting as energy from good fats, such as eggs, fatty meat, pork and bacon.
Lack of time is not the problem, it is lack of understanding, because a lot of good foods are actually widely available, but we don´t realise which are good and which are bad.
I am witness to the health benefits of this diet after a lifetime of eating what is now ubiquitously accepted as normal food. Good food is more difficult to obtain and prepare, but it is certainly possible to do so with little difference in cost, a huge increase in food enjoyment, and most importantly, a huge increase in bodily health.
The most important thing required to eat well is an education in the area of food. And, the simplest way to obtain that is to go to Dr. Mercola's excellent and extensive website (www.mercola.com) and follow your interest.
If you try this diet, I guarantee that you will not be disappointed, as it consists of foods that we all enjoy. In fact, this real old-fashioned food is the real medicine, both as a preventative and as a treatment.
What Next?
Clark:
Patients have always appreciated your strong and committed work, and your past colleagues speak admirably of you. The availability of funding for ME/CFS has already been an issue and continues to be; even Professor Lipkin, who you would think would have no trouble landing research funding for his microbiome study, was turned down.
From the outside at least, your ME/CFS research seemed to come to a bit of a halt a few years ago and came with the news that you had left your position at St George’s hospital, London. Did these inherent problems with research funding in ME/CFS make it difficult to carry out the research you wanted to? What happened?
Dr. Kerr:
Yes, it is difficult to obtain funding for any research, and even more difficult to obtain it for ME/CFS research.
After the death of David Tyrrell, the CFS Research Foundation made it a priority to join with the MRC in funding CFS research of a combined biological and psychological nature.
Around the same time, Judy Mikovits and I were awarded $2 million from NIH to study biological aspects of CFS in the UK and USA. I resigned from this grant because of the increasing controversy surrounding XMRV, and the fact that my lab could not detect XMRV in any CFS patient.
Judy's lab found it in a majority of patients, including samples which we found to be repeatedly negative, using the identical PCR primers, while she was actually offering the same test privately.
With these two grant incomes gone, there was insufficient funding to sustain the research.
Clark:
You’re now working at the Universidad del Rosario, Bogota, Colombia - are you still involved with research into the disease and if so then what do we have to look forward to?
Dr. Kerr:
I plan to look at several CFS-associated genes in more detail, to continue the Parvovirus B19 work and to explore ways in which the role of diet can be studied with regard to immune function/ dysfunction.
Clark:
Thank you, Dr. Kerr.
Clark Ellis writes about ME/CFS research and is writing a fiction novel/screenplay with themes related to ME/CFS. You can follow him on twitter to hear about his work, ME/CFS research, fundraising and advocacy.
FURTHER INFORMATION AND REFERENCES:
ME/CFS SNP paper
Shimosako N, Kerr JR.
Use of single-nucleotide polymorphisms (SNPs) to distinguish gene expression subtypes of chronic fatigue syndrome/myalgic encephalomyelitis (ME/CFS). J Clin Pathol
2014 Sep 19. pii: jclinpath-2014-202597. doi: 10.1136/jclinpath-2014-202597. [Epub]
1. Folic acid supplements and risk of facial clefts: national population based case-control study
2. Maternal dietary intake of vitamin A and risk of orofacial clefts: a population-based case-control study in Norway.
3. Maternal Western dietary patterns and the risk of developing a cleft lip with or without a cleft palate
Book of Weston Price, ´Nutrition and Physical Degeneration´ (available free as PDF)
Book & reviews available on Amazon
Wikipedia page on Weston Price
The Weston Price Foundation website
Dr Mercola´s website
Saturated fat, by Dr Mercola
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