Clinically proven mtDNA mutations are not common in those with chronic fatigue syndrome

[Kati]

Clinically proven mtDNA mutations are not common in those with chronic fatigue syndrome

• Elizna M. Schoeman†, Francois H. Van Der Westhuizen†, Elardus Erasmus, Etresia van Dyk, Charlotte V. Y. Knowles, Shereen Al-Ali, Wan-Fai Ng, Robert W. Taylor, Julia L. Newton and Joanna L. Elson

†Contributed equally

Received: 7 October 2016 Accepted: 2 March 2017 Published: 16 March 2017

Open Peer Review reports

Abstract
Background
Chronic Fatigue Syndrome (CFS) is a prevalent debilitating condition that affects approximately 250,000 people in the UK. There is growing interest in the role of mitochondrial function and mitochondrial DNA (mtDNA) variation in CFS. It is now known that fatigue is common and often severe in patients with mitochondrial disease irrespective of their age, gender or mtDNA genotype. More recently, it has been suggested that some CFS patients harbour clinically proven mtDNA mutations.

Methods
MtDNA sequencing of 93 CFS patients from the United Kingdom (UK) and South Africa (RSA) was performed using an Ion Torrent Personal Genome Machine. The sequence data was examined for any evidence of clinically proven mutations, currently; more than 200 clinically proven mtDNA mutations point mutations have been identified.

Results
We report the complete mtDNA sequence of 93 CFS patients from the UK and RSA, without finding evidence of clinically proven mtDNA mutations. This finding demonstrates that clinically proven mtDNA mutations are not a common element in the aetiology of disease in CFS patients. That is patients having a clinically proven mtDNA mutation and subsequently being misdiagnosed with CFS are likely to be rare.

Conclusion
The work supports the assertion that CFS should not be considered to fall within the spectrum of mtDNA disease. However, the current study cannot exclude a role for nuclear genes with a mitochondrial function, nor a role of mtDNA population variants in susceptibility to disease. This study highlights the need for more to be done to understand the pathophysiology of CFS.

Keywords
Chronic fatigue syndrome mtDNA mutations Pathogenicity

 

[Kati]

I think we knew that ME patients in general did not have mitochondrial DNA mutations or inherited mitochondrial disease, so this is a bit redundant in my opinion. i hope this group, if pursueing research for ME will quickly move on to the functional asepct of the mitochondria.

 

[Valentijn]

They're only testing mitochondrial DNA from blood samples, so have proven pretty much nothing. They understand that heteroplasmy means the mutation will primarily appear in the affected tissues, but don't seem bothered about it. If someone has a suspected mitochondrial myopathy, the appropriate way to diagnose it (or rule it out) is by testing the mitochondrial DNA from a muscle biopsy.

I think we knew that ME patients in general did not have mitochondrial DNA mutations or inherited mitochondrial disease ....

Why do you say that? Mitochondrial mutations will typically cause disease in only some people who have them, due to the random manner in which they end up in varying amounts in varying tissues of a fetus. Family members of someone with a severe mitochondrial disease typically have mild and more limited symptoms, or no symptoms at all.

 

[Neunistiva]

Why do you say that?

I think @Kati said it because several ME/CFS researchers, including Dr. Davis said that, while very rarely people with mitochondrial mutations have been misdiagnosed with ME/CFS, they didn't find any mitochondrial mutations in great majority of people with ME/CFS.

Furthermore, Dr. Davis noted:
"ME/CFS patients have a marked decrease in some of the Citric Acid Cycle metabolites while mitochondrial mutations generally cause an increase."

That being said, I think negative scientific results are also very important, and I am glad they published it.

 

[Kati]

They're only testing mitochondrial DNA from blood samples, so have proven pretty much nothing. They understand that heteroplasmy means the mutation will primarily appear in the affected tissues, but don't seem bothered about it. If someone has a suspected mitochondrial myopathy, the appropriate way to diagnose it (or rule it out) is by testing the mitochondrial DNA from a muscle biopsy.

They mentioned in their paper that performing muscle biopsies 'weren't practical'- I guess they would need to prove to the ethics committee this was necessary and worth the risk, and they would need to have sufficient funding to perform this intervention to a sufficient number of patients.

Why do you say that? Mitochondrial mutations will typically cause disease in only some people who have them, due to the random manner in which they end up in varying amounts in varying tissues of a fetus. Family members of someone with a severe mitochondrial disease typically have mild and more limited symptoms, or no symptoms at all.

From personal experience and from mitochondrial specialists. I have had my MT DNA sequenced. The report said it was normal. I am not sure if there were mutations or not, I do not have the sequencing. The specialist said there were a number of us who had symptoms, without proven mito DNA issues. At least 2 of my ME specialists confirmed that usually for ME patients the MT DNA is normal.

 

[Jenny TipsforME]

I noticed quite a few supposedly mitochondria related SNPs in my LiveWello info. I haven't been through them systematically to see whether this is meaningful (e.g. 48% of the population may have the same as me).

 

[Valentijn]

I think @Kati said it because several ME/CFS researchers, including Dr. Davis said that, while very rarely people with mitochondrial mutations have been misdiagnosed with ME/CFS, they didn't find any mitochondrial mutations in great majority of people with ME/CFS.

At least 2 of my ME specialists confirmed that usually for ME patients the MT DNA is normal.

Were they testing blood, or were they testing from a muscle biopsy? Because a negative result from blood is simply meaningless. I don't expect ME researchers or clinicians to know that, but geneticists and mitochondrial disease specialists certainly should.

 

[Deepwater]

Were they testing blood, or were they testing from a muscle biopsy? Because a negative result from blood is simply meaningless. I don't expect ME researchers or clinicians to know that, but geneticists and mitochondrial disease specialists certainly should.

I stand to be corrected by someone with scientific knowledge, but what I've gleaned from recent research is that the mitochondria in themselves indeed seem to be fine, but they can't do their job because of some blocking agent (in the serum?). I happen to know a number of relatives - some quite distant - on the same mtDNA line as myself, and all are well, so I have no trouble believing the illness is not simply due to genetics.
What worries me is that Julia Newton seems to be the UK's/ MEA's go-to expert on mitochondrial research, and if she just stops here and says 'no mitochondrial problem, QED', then the point is likely to be completely missed and we PwME Brits will simply be handed back to the BPS quacks.
I must say I had my worries all along about the scope of the mito research that has been funded in England. I believe the next stage is trying to validate the Myhill-McLaren tests by seeing if muscle biopsies give the same abnormal results. I would love one of the scientists on this forum to set my mind at rest that these two sorts of tests are not geared to picking up different problems.

 

[Kati]

Were they testing blood, or were they testing from a muscle biopsy? Because a negative result from blood is simply meaningless. I don't expect ME researchers or clinicians to know that, but geneticists and mitochondrial disease specialists certainly should.

It was from my muscle biopsy, sent to the US for DNA sequencing and various tests. The DNA seq was negative for the typical genetic abnormalities they find in genetic type of mito diseases. It was in 2012.

I had some out of range results on the tests below- this was on muscle tissus which was weird, but the doctor (mito specialist) didn't seem to be interested into that. The miscroscopy showed my slow twitch muscle were atrophied.

 

[Kati]

What worries me is that Julia Newton seems to be the UK's/ MEA's go-to expert on mitochondrial research, and if she just stops here and says 'no mitochondrial problem, QED', then the point is likely to be completely missed and we PwME Brits will simply be handed back to the BPS quacks.
I must say I had my worries all along about the scope of the mito research that has been funded in England. I believe the next stage is trying to validate the Myhill-McLaren tests by seeing if muscle biopsies give the same abnormal results. I would love one of the scientists on this forum to set my mind at rest that these two sorts of tests are not geared to picking up different problems.

There has been many publications across continents in regards to metabolomics, showing problems with the Citric Acid Cycle and other pathways. Dr Newton would be stalling her career if she chose to disregards those.

UK will have no choice but to wake up to science, because there are very motivated research teams out there willing to dig deeper, find the pathology and find treatments that work.

 

[Valentijn]

That being said, I think negative scientific results are also very important, and I am glad they published it.

I agree that it's important to publish negative results. But the authors are calling this a negative result while it is simply a meaningless result. Even the reviewers of this paper pointed out that blood is the location where you are least likely to find mitochondrial mutations when the patient has a genetic mitochondrial disease.

They mentioned in their paper that performing muscle biopsies 'weren't practical'- I guess they would need to prove to the ethics committee this was necessary and worth the risk, and they would need to have sufficient funding to perform this intervention to a sufficient number of patients.

Or they don't think ME/CFS is serious enough for a muscle biopsy to be warranted, or they weren't willing to risk a positive result. The fact remains that using a blood sample makes their results completely meaningless, and they have absolutely no support for their conclusion that:

This finding demonstrates that clinically proven mtDNA mutations are not a common element in the aetiology of disease in CFS patients.

The mutations would rarely be present in blood for anyone with a genetic mitochondrial disease. They could use exactly the same method to "prove" that people with confirmed mitochondrial disease don't have it after all because it's not in their blood

This paper is either completely pointless, or it's self-sabotaged by design to supposedly rule out another biomedical mechanism and discourage further research into it.

 

[A.B.]

If you read the conclusions and the funding section, it could explain where they are trying to go:

However, to make the breakthough in understanding the genetics of this complex condition a large scale-sequencing project will need to be undertaken to help discover the genes and pathways that differ in those suffering from this complex condition. Data from such a “big data” genomics project will help us understand how this disease is stratified, which in turn should help to guide the way that patients are treated, as well as providing therapeutic targets.

We would like to thanks the following bodies for funding support. The Centre for Human Metabolomics, North-West University The Wellcome trust by means of the Institutional Strategic Support Fund (ISSF) and Action for ME RES/0200/7841 (to JLE). RWT is supported by a Wellcome Trust Strategic Award (096919/Z/11/Z), the MRC Centre for Neuromuscular Diseases (G0601943), the Lily Foundation and the UK NHS Highly Specialised “Rare Mitochondrial Disorders of Adults and Children” Service in Newcastle upon Tyne.

Can you say MEGA?

 

[Valentijn]

Furthermore, Dr. Davis noted:
"ME/CFS patients have a marked decrease in some of the Citric Acid Cycle metabolites while mitochondrial mutations generally cause an increase."

I'm not sure this is accurate, based on OMIM's summary of Complex I Deficiency at least:

Mitochondrial complex I deficiency shows extreme genetic heterogeneity and can be caused by mutation in nuclear-encoded genes or in mitochondrial-encoded genes. There are no obvious genotype-phenotype correlations, and inference of the underlying basis from the clinical or biochemical presentation is difficult, if not impossible (summary by Haack et al., 2012).

OMIM's summaries of individual cases also shows many with different (high or low) levels of varying substances, and some with normal levels. I saw the same thing while reviewing larger research studies. The only consistent lab-test abnormalities for genetic mitochondrial mypothies are in the muscle tissues (fibers and mutations) and the CSF (elevated lactate). MRI brain scans are usually abnormal as well, but less specific to mitochondrial disease.

 

[Kati]

.

Or they don't think ME/CFS is serious enough for a muscle biopsy to be warranted, or they weren't willing to risk a positive result. The fact remains that using a blood sample makes their results completely meaningless, and they have absolutely no support for their conclusion that...

This is where ethics committee would stop well meaning researchers if they feel the intervention on the patient is too invasive for what ails them. It's probably out of despair to prove his theory that Barry Marshall drank a beaker full of H Pylori.

The other kind of committee that stops straight researchers in their track is grant review committees who still believe that ME is all in the head and that medical research is not warranted.

In speaking with researchers in the field, I got a strong feeling that many interventions would be stopped by the ethics committee or by grant reviewers and speaking of which it sounds like it will even get harder to get a ME grant if Trump's budget passes in Congress.

If you live in the US, please phone your Congressman.

 

[trishrhymes]

I'm no expert on inherited mitochondrial disease, but i had understood that we inherit our mitochondria, and hence our mitochondrial DNA from our mothers.

The particular mix of normal and mutated DNA in each cell, including egg cells varies because there are so many mitochondria in each cell, so the proportion carrying a particular mutation will vary.

But all our mitochondrial DNA must have come in that one egg cell from our mother. As embryonic cells differentiate into different tissues, again there is some random variation of proportion of mitochondria carrying a mutation, so tissues may be differentially effected, but basically the DNA is still that inherited from the mother.

So why are you saying testing blood cell mitochondrial DNA is useless? If the aim is to see whether there are people with inherited mitochondrial DNA defects being misdiagnosed as ME, or indeed whether at least some ME is caused by inherited defects, then blood should be no better or worse a source than muscle.

Surely this study, by ruling out inherited mitochondrial disease, is a good start, so long as there is then further investigation into whether there are acquired mutations in particular tissues, or, as seems more likely to me, epigenetic difference in which genes are being switched on or off in our mitochondrial or nuclear DNA.

Please can someone with more genetic knowledge than me pick holes in what I'm saying here. Have I misunderstood something?

 

[Valentijn]

But all our mitochondrial DNA must have come in that one egg cell from our mother. As embryonic cells differentiate into different tissues, again there is some random variation of proportion of mitochondria carrying a mutation, so tissues may be differentially effected, but basically the DNA is still that inherited from the mother.

I had the same misunderstanding until recently. But unlike chromosomal DNA, the mitochondrial DNA is not sourced from a single copy. Instead, there are about 200,000 mitochondria in the mother's egg, which then end up in various fetal cells.

So in a mother (symptomatic or not) who is carrying the mutation, her egg will contain a somewhat random amount of the mutated mitochondria, in addition to the healthy mitochondria. And the fetal cells that those mitochondria end up in will also be pretty random. Then those normal and mutated mitochondria will replicate and one or the other will dominate in the tissues that the mutated mitochondria are present in.

This is called heteroplasmy, and is now recognized as being a fundamental aspect of genetic mitochondrial disease. Due to heteroplasmy, the mutation, and therefore the symptoms, appear randomly in different tissues. This is how family members with exactly the same mutation can end up with different mitochondrial diseases - one might have MELAS affecting the muscles, and another might have just have eye degeneration, or deafness & diabetes, or myoclonus, etc.

If there is too much of the mutation in a fetal cell, and not enough energy production, that cell will probably die. Hence healthy mitochondria can easily dominate at first, and the impact of the mutation is held in check to some degree with each generation. This is the mitochondrial bottleneck.

Due to the bottleneck, even people with the same mutation in the same tissues could end up with completely different levels of severity and age at onset. One person might be symptomatic at birth and die very young, while an immediate relative might only have mild symptoms starting in adulthood, and another relative might have no symptoms at all during her entire life.

So why are you saying testing blood cell mitochondrial DNA is useless? If the aim is to see whether there are people with inherited mitochondrial DNA defects being misdiagnosed as ME, or indeed whether at least some ME is caused by inherited defects, then blood should be no better or worse a source than muscle.

The mutation will only be present in large amounts in the tissue where it is causing problems. So in the case of genetic mitochondrial muscle problems, the mutation would be in the muscles. I'm not sure why it is so rarely found in the blood for any genetic mitochondrial disease, but it's accepted that that is the case, based on the research.

 

[trishrhymes]

Thank you. I basically had the right idea about different levels of mutation in different tissues, just didn't realise this between tissue difference would be so drastic.

I see now why you say it is the mitochondria in the effected tissue that should be tested.

Surely Julia Newton must be aware of this?????

 

[Valentijn]

Surely Julia Newton must be aware of this?????

I doubt it. It's a very specialized part of a broader specialty which Newton is not involved in. She isn't a geneticist. But there are some on the team, and they certainly should know better.

 

[justy]

I haven't read the full paper (brain fog) but to someone who has - did they not find one single case of misdiagnosed mitochondrial disease?

Hmn, but then again, as @Valentijn said you cant find mito genetic defects in the blood - but must do muscle biopsies. Wasn't this what a lot of the hoo-ha was with Dr Myhill's research? people slagging it off left right and centre as it was done on blood - specialists saying those of us who had the testing should realise it was meaningless and the only way we could tell was with a muscle biopsy - and then 9 and a half times out of 10 being refused a muscle biopsy?

@trishrhymes I am 100% certain Julia Newton does know this - and I want to know why they ruled out doing muscle biopsies on this patient group. I have the impression that it is thought to be too invasisve for an illness that is not that serious- this is a pervasive attitude- especially in the UK where we are told over and over again that Rituximab is too serious a drug for us etc etc.

 

[Valentijn]

I haven't read the full paper (brain fog) but to someone who has - did they not find one single case of misdiagnosed mitochondrial disease?

I'm not sure. They have data regarding variants in the supplemental material, but I haven't taken a look. As you say, it doesn't matter much if there isn't, since they aren't using the appropriate methodology to find the mutations.