Cellular bioenergetics is impaired in patients with chronic fatigue syndrome

Demepivo

Dolores Abernathy
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Just going through it now but my first (parochial) thought is, wow, this is BRITISH research. Actual British research that doesn’t involve false illness beliefs, beat-em-harder exercise techniques, or expensive magic circles devised by tarot practitioners.

The tide is turning.
Fortunately Julia Newton has been doing good science for more than a decade. She is well regarded by the MRC & Newcastle Hospitals where she is a senior person.

As one person said, "Julia Newton has left the CMRC to spend more time on ME Research"
 

alex3619

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My guess is that I can't see that it would be a good diagnostic test for now.
It does not even come close to being a diagnostic test. I called it a confirmatory test. If its not confirmed, you probably don't have ME. You might have a particular CFS subtype, that is a separate issue. Its not about diagnosing ME, its about removing non ME cases.

One of the huge difficulties we face is that we can reliably get good test sensitivity now, but not specificity. It takes a lot more research to assess specificity, and maybe even a mechanism is required.


If we take the measurements for which there is the biggest difference between CFS and Controls, say Fig 3.D or Fig 3.E (Maximal Respiration and Reserve Capacity, respectively), whilst there's a highly statistically significant difference between the groups, there is still a reasonable amount of overlap between the two.
That is one interpretation. As I see it, if you focus on fresh samples, for basal respiration, maximal respiration, and reserve capacity, there is a clear separation between a cluster of four and the rest of the CFS patients, though this is less obvious with basal respiration. Most patients cluster together, and clearly are below the controls.

My interpretation of this is we are possibly seeing either two or more subtypes, or the same subtypes on a good day. If you remove the clear cluster at the top then the very close cluster below is totally separable. This needs more investigation. It has no clinical use right now, but is very much a valid research question.


Edit: though I should note the overlap could be due to the diagnostic criteria used (Fukuda) picking up people with a different aetiology for their fatigue. All part of the fun of trying to research heterogeneous conditions that may be more than one underlying illness.
This is possible, but they might all have the same type of illness and you could still see these kinds of results. ME appears to be dynamic, and on a good day a mild patient might test normal. I think we need to develop specific challenge testing for ME, then test a large cohort.

The Seahorse machine is getting to be popular for ME/CFS. The very small sample of Seahorse data I saw at NIH very broadly agrees with this study - CFS patients did not ramp up energy production in response to demand as well as controls, by a big margin. The NIH study is using white blood cells too, but the scientist running the machine really wants to get her hands on muscle cells.
What I hope to see is that PBMC results closely correlate with muscle results. If so then we can dispense with biopsies and use blood data. Its nice to know that the NIH is finding something that is along the same lines. I greatly look forward to seeing their findings published or released.

PS Even better is if we can find a good correlation between PBMC results and repeat CPET testing.
 

alex3619

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If Mark Davis’ work is correct regarding CD8+ T cell expansion in ME/CFS, then the OXPHOS impairment they're seeing in this study may simply be the relevant T cells having undergone activation and consequently a switch to aerobic glycolysis away from OXPHOS for expansion/proliferation purposes - i.e., the usual process that T cells normally use when activated.
Yes. Further investigation is needed to exclude, or confirm, this possibility. However this will only alter the interpretation a little as controls and patients are separable. It does matter though whether this is a general tissue finding or a T cell or even B cell finding.
 

Marco

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Patients having a good day, or a case of misdiagnosis?
Could be. Although I'm generally not a fan of ever more strict criteria in all circumstances it would have been useful to know if these measures correlated with functionality.

It does also highlight the problems of making straight group comparisons without publishing the raw data. In other circumstances this could easily have been a null result.
 

Demepivo

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*ICYMI Julia Newton gave a talk at September CMRC conference talking about various studies she is involved in including the metabolomic one with Karl Morten of Oxford University. (who follows on from her in the talk).

*Her cohorts are well defined & exclude patients with depression and other mental health co-morbidities.

 

Londinium

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Fortunately Julia Newton has been doing good science for more than a decade. She is well regarded by the MRC & Newcastle Hospitals where she is a senior person.
Agreed, I was being a little unfair. My impression had been that Julia Newton had done quite a bit of epidemiological stuff (e.g. 40+% of ME/CFS referrals having other dignosable conditions) and the biomedical stuff had so far not turned up much, compared to this study. (Not that negative results aren't important).

It does not even come close to being a diagnostic test. I called it a confirmatory test. If its not confirmed, you probably don't have ME. You might have a particular CFS subtype, that is a separate issue. Its not about diagnosing ME, its about removing non ME cases.
I think a similar problem applies though: it's not even confirmatory. If you test 'high' (i.e. more in line with controls) we can't conclude you don't have ME because, as you correctly note further on, we still don't know whether variation between results is due to temporal variance and whether some ME patients may only test 'low' on a bad day but fine on a good day. We need longitudinal testing to confirm. I'm wary of saying 'you don't have ME you must have some other form of CFS' based on one blood test given how little we currently know.

That is one interpretation. As I see it, if you focus on fresh samples, for basal respiration, maximal respiration, and reserve capacity, there is a clear separation between a cluster of four and the rest of the CFS patients, though this is less obvious with basal respiration. Most patients cluster together, and clearly are below the controls.
I'm cautious on looking at the fresh samples, because there were only three data points for the controls. Thus I don't think we can draw any conclusions on the specificity of fresh CFS vs fresh controls as we may be getting a very incomplete view of the latter. The spread within the fresh CFS group is certainly interesting and warrants further investigation, but could be down to the remitting/relapsing nature of ME in many cases. As always, more research needed :)

What I hope to see is that PBMC results closely correlate with muscle results.
Totally agree. Ideally (money no object) I'd like to see the same team try:
  • Testing of other cell types to try to isolate whether this is a wider metabolic issue vs a immune cell issue
  • Longitudinal testing, plus correlation analysis with symptoms;
  • Use of sedentary controls;
  • More fresh controls;
  • Test in and out of serum.
I think (at work so no time to check) the study team are already planning to do quite a few of these.
 

Demepivo

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Agreed, I was being a little unfair. My impression had been that Julia Newton had done quite a bit of epidemiological stuff (e.g. 40+% of ME/CFS referrals having other dignosable conditions) and the biomedical stuff had so far not turned up much, compared to this study. (Not that negative results aren't important).
*Yes, her intitial work was on autonomic dysfunction/orthostatic intolerance and she hasdocumented that in patients.

*Other work has looked at misdiagnosis of patients

*More recently the metabolomic stuff with Karl Morten of Oxford University

*Agreed that not all of her studies have turned up positive results but that has been the case for many researchers. New techniques & equipment are helping.
 

JamBob

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If they only test against healthy controls, how can we know that the results aren't just an effect of inactivity. I wish they would include an extra inactive control group when they do these kind of studies.
 

Tunguska

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It's impossible to compare the various studies out because I don't think any of them detail the phenotype of the WBCs they're studying and this does matter when it comes to metabolic function. I hope that from here we can move away from using WBCs for this type of research because it introduces confusion and seemingly conflicting results. It's time to start looking at organ and muscle cells.
That's the #1 thing (in queue) I'd like to see. Most of these things being posted are nice but mostly redundant one way or another.
 

Jesse2233

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My mind automatically goes to treatment possibilities even though I know this study is preliminary. Might a white blood cell reset using a modality such as photopheresis be appropriate?
 

Scarecrow

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Agreed, I was being a little unfair. My impression had been that Julia Newton had done quite a bit of epidemiological stuff (e.g. 40+% of ME/CFS referrals having other dignosable conditions) and the biomedical stuff had so far not turned up much, compared to this study. (Not that negative results aren't important).
*Yes, her intitial work was on autonomic dysfunction/orthostatic intolerance and she hasdocumented that in patients.

*Other work has looked at misdiagnosis of patients

*More recently the metabolomic stuff with Karl Morten of Oxford University

*Agreed that not all of her studies have turned up positive results but that has been the case for many researchers. New techniques & equipment are helping.
Not forgetting the in vitro muscle cell study:
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0122982
Nice summary from MERUK.

@halcyon @Murph
Thought you might be interested in the earlier Newton study using muscle cells. As expected, fewer samples than in this PBMC study.
 

alex3619

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My mind automatically goes to treatment possibilities even though I know this study is preliminary. Might a white blood cell reset using a modality such as photopheresis be appropriate?
First I would want to know if this was just white cells, or was in other tissues as well. However if the main issue is immune signalling, which we do not know, then treating just immune cells might be enough. Do you have a link to a good paper on photopheresis?
 

alex3619

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If you test 'high' (i.e. more in line with controls) we can't conclude you don't have ME because, as you correctly note further on, we still don't know whether variation between results is due to temporal variance and whether some ME patients may only test 'low' on a bad day but fine on a good day. We need longitudinal testing to confirm. I'm wary of saying 'you don't have ME you must have some other form of CFS' based on one blood test given how little we currently know.
In the context I am discussing this the problem is irrelevant. I made it clear I am not talking about clinical practice. I think a more distinct subgroup could be found if those who show no such problem are removed. Even if some of those with core ME are removed its not a problem for a research cohort.

Clinically its a different picture, but we are not seeing enough research to justify any clinical use as yet.

It is clear this research needs a lot more work. A larger study, more fresh samples, and so on. A longitudinal study would be great, but they are so very rare in ME, there is not sufficient funding usually available, or at least few apply for it. It should still be on the wish list though.

To me the importance is it offers a potential blood test. This would definitely require some kind of challenge, such as with glucose or salt, or substances that impede or enhance various enzymes under investigation. I think this shows potential for further studies, and comparisons with repeat CPET and metabolomics data, and even cellular impedence, might correlate. We need to know if they do. We also need to know which other tissues, of those that can be safely biopsied, this can also be found in.

I am aware of other findings of low metabolic rate, but much of it is not published. So very much our clinicians learn is not published. This line of investigation, with Seahorse etc., gives a cellular snapshot of energy metabolism. Combine that with metabolomics and we might learn a lot more.

Of course if a larger study, or one by a different group, shows different results, then that will put this in a different perspective. We need those studies.
 

FMMM1

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It does not even come close to being a diagnostic test. I called it a confirmatory test. If its not confirmed, you probably don't have ME. You might have a particular CFS subtype, that is a separate issue. Its not about diagnosing ME, its about removing non ME cases.

One of the huge difficulties we face is that we can reliably get good test sensitivity now, but not specificity. It takes a lot more research to assess specificity, and maybe even a mechanism is required.
Thanks Alex.

Take a look at Ron Davis's talk on the recent OMF Symposium re his impedance chip. From memory one of the things that reversed the block in metabolism was suramin (an anti-purinergic signaling drug). Perhaps testing using suramin would increase specificity. He's probably looked at who bunch of other drugs etc re impedance chip.

If I understand your comments correctly i.e. that this would aid research (in selecting a uniform test group) then I agree. Surely if you have a heterogeneous population then the data is more difficult to interpret.

One of the plus points of a diagnostic test, based on this device (Seahorse) or some other, is that it may put more pressure on those who allocate research funding. E.g. the European Union Horizon 2020 pot is 80 billion euros ish and I can't think of a single thing they've funded for ME/CFS (please point stuff out if you can). Mind you I can't think of much they've funded for fibro or lymes either.
 

alex3619

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I wonder how/if the results would differ if the CFS patient PBMCs were first placed in the serum of a healthy individual.
That is an experiment I would love to see.
Take a look at Ron Davis's talk on the recent OMF Symposium re his impedance chip
This is promising technology, but we still do not know what it is doing. My guess is that Seahorse testing would be one of the things he is using to find out. I mentioned a salt challenge specifically because that is what Ron Davis is testing.
 
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this study may be confirming that a population of T cells is inappropriately activated - rather than there being some bodywide OXPHOS dysfunction that is also occurring in other cells like muscle, liver, etc (although deranged cytokine signaling from the activated T cells can potentially cause mitochondrial dysfunction in those other cells).
I wanted to say this as well. It's very tempting to look at results like this, and go "Look, this proves we can't produce ATP as well as healthy people! This is why we're tired!", but the key thing a lot of people looking at studies like that are forgetting is cell types. The human body is made of a multitude of different types and suptypes of cells, that do very different things, express different proteins, and have many other differing properties. And yes, the energy metabolizm of most cells consists of the same molecular machinery - glycolysis, pyruvate decarboxylation, beta-oxidation, krebs cycle, electron transport chain. But the regulation of those processes differs from cell to cell. Every time you hear about a discovered intracellular dysfunction, you have to ask the question "okay, what cells exactly?"

A lot of those studies, even Ron's work, use PBMCs. Peripheral Blood Mononuclear Cells are a middle layer of blood, separated by centrifugation. It contains T-cells, B-cells, NK cells, monocytes, and sometimes trace basophils. In other words, immune system cells. Our PBMCs having its energy metabolism impaired doesn't directly explain us feeling fatigue, or not being able to tolerate exercise. Such findings in muscle cells could maybe, kinda explain it (I don't think fully though). Interestingly enough, Fluge and Mella did, in fact, perform those same Seahorse measurements in cultured muscle cells exposed to ME/CFS serum. And the results were the opposite - oxygen consumption was actually increased, not decreased. So I would be careful with assuming that those results here, or any similar studies, can be extrapolated to the whole body. Those results are indicative of an *immune* alteration, if anything. It can be a dysfuntion of the immune cells, a different cell type ratio than the controls, or just immune activation. We don't know.

It's nice the UK is coming around to doing actual research in ME/CFS. But this doesn't really tell us anything new about the illness mechanism, in my opinion.

What I would like to see more of, is actual cell sorting in those studies. If the measurements of those things varied a lot across PBMC types, it could tell us more about what the immune system is doing, and if they didn't, that would actually be indicative of a body-wide cellular metabolism impairment.

This is also why I was excited when Ron said he wanted to use pluripotent stem cells, because that way you can test all the cell types you want (with limitations, but still).
 
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alex3619

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I do think cell sorting should be tried, but I am unsure that it will help. Most of my caveats are technical, so if anyone knows a lot about cell sorting then comment would be good, though you might also need to know a lot about seahorse technology.

First, such cell fractions have further cell fractions within them. I don't think this is a major issue, but it might mean a series of studies is needed.

Second, when you do cell sorting you might change cell chemistry. Various controls would be needed, including cells replaced in patient serum.

Third, when you cell sort you decrease quantity. A lot. So at what quantity will seahorse technology become unreliable?

For me I would first want to know if skin cells or muscle cells have similar issues. Then I would look at cell subfractions if that is viable. However if muscle cells are normal the big question has been answered, and subfraction testing would be about finding more clues. If muscle cells are not normal then we can put subfraction testing off for a while as its no longer so important.
 
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