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Assessing cellular energy dysfunction in CFS/ME using a commercially available laboratory test, 2019, Morten, Newton et al

Discussion in 'ME/CFS research' started by Sly Saint, Aug 7, 2019.

  1. richie

    richie Senior Member (Voting Rights)

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  2. boolybooly

    boolybooly Senior Member (Voting Rights)

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    I think its a credible response from Dr Myhill and Dr McLaren Howard. In my experience as a patient who had these tests, this response is correct in stating...

    This is correct regarding how it was used in my case, as evidence of low functioning mitochondria to the DWP, while this was the case, not as a diagnostic test for ME CFIDS. Which is supported by the fact that when mito tests normalised for me after quitting nightshades, I was still able to get an ME CFIDS diagnosis based on clear evidence of continuing CFIDS, i.e. the diagnosis was based on clinical presentation, not the test.

    I would also add that Dr Myhill was at all times very ethical over money and I say that as a practiced downshifter, long time obligate rat race revolutionary, green thinker and one time corporate fundraising ethical vetting analyst for OXFAM! Not that people dont make mistakes because we all do and goodness knows OXFAM did, which is why ethical oversight is important.

    So I think its fair to ask searching questions but I do not understand why Tomas et al stated...

    EDIT
    I dont presume to know what is going on here. There may have been more going on with the tests to which I was not a witness, I dont know what I dont know, but the tests were not used as a diagnostic test for the condition in my experience, they were used as a way to investigate mitochondrial function. I fully accept the distinction is a subtle one but it is a distinction which was made in my experience.
     
    Last edited: Aug 17, 2019
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  3. richie

    richie Senior Member (Voting Rights)

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    My hope is that all is not lost in this matter.

    It may be that some practitioners rather than My/Mc presented/used this test as diagnostic.

    Did you improve clinically after removal of nightshades?
    I certainly recall mitochondrial dysfunction as attested in this test as being linked to symptoms and I think this is still the claim of My/Mc.This is not a direct claim of diagnostic validity for ME/CFS but it does imply or assert a positive correlation of bad symptoms and poor mito score and improvement of symptoms after appropriate intervention, alongside better mito function as per test, seems to be a central claim.

    I would like to see issues around timing/procedure sorted and Mc's offer should have been taken up originally imo.

    Then there is the question of clinical results, which is key. Diagnostic invalidity does not in itself render a test useless.
    The same issue has arisen for just about every test that has in any way been presented as diagnostic rather than potentially informative and clinically useful. But we are always demanding and understandably seeking a diagnostic test.......
     
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  4. boolybooly

    boolybooly Senior Member (Voting Rights)

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    @richie something changed, lets put it like that, wasn't all good though.

    I gave up nightshades because they were giving me mouth ulcers. Diagnosed via a food diary and google. I eliminated nightshades reluctantly and the ulcers stopped. Ulcers I thought were probably due to some kind of immunological response.

    Nightshades are believed to assist the immune system by increasing apoptosis aka cell death rates due to the action of solanines and other glycoalkaloids, clearing infected and weakened cells.

    The question of clinical change for me is thorny. I still have PEM meaning I cant do much, my virus attacks got more frequent probably due to reduced cell death not clearing infected cells and I got more muscle pain possibly due to improved calcium retention in mitochondria, ironically.

    On the other hand the deathly malaise I used to get sometimes between virus attacks, which I believed were associated with my very highest cell free DNA scores, improved. This may also have been linked to improved mito function, no way to know for sure.

    Giving up nightshades changed things but it was swings and roundabouts and coincided with normalised mito scores with this test. But what this really means is open to question as I still have PEM, virus recurrence and neurological hypersensitivity etc.

    e.g. hypothetical scenario. If the test removes ME mediating plasma factor/s from the cell environment then the nightshade free mito score improvement may have reflected a change in the way my cells react to the test, due to some affect related to nightshades. JMH had commented on my mitochondria translocator protein tests before stopping nightshades, that the TL site appeared to be blocked by large proteins possibly immune complexes. If that was a nightshade related complex (related to the reaction causing ulcers), eliminating nightshades may have changed the way my cells react to the test, in particular the rapidity with which my cells respond to the presence or absence of the hypothetical ME mediating plasma factor/s. i.e. Possibly the protein complex made my cells less permeable and slower to let go of the ME state once the plasma factor was no longer present. Once the protein complexes were gone then the way the test apparently isolates cells and removes plasma factors by design may have made my cells bounce back from the ME CFIDS state in vitro thus masking their real condition in vivo.

    I am curious whether this test is separating cells from plasma and thereby obscuring mito dysfunction for other people and whether this has anything to do with the problems with replication.
     
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  5. FMMM1

    FMMM1 Senior Member (Voting Rights)

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    Karl Morten, in a presentation in New Zealand (2018? - you tube + thread on this site?) emphasised that samples are changing from the moment they are taken e.g. glucose is being consumed --. At the time I thought OK you've made your point Karl, move on; however, this paper sheds new light on his comments --. Karl Morten's group, and Julia Newton's group, have worked together e.g. they published this paper on a potential diagnostic test [https://ora.ox.ac.uk/objects/uuid:56069cef-320f-43ad-b970-20c1fdced6e4].
    Evidently they were concerned, and justifiably so, about this flawed test being offered for sale to vulnerable people ---

    This raises a a wider point; in the case of Lyme disease, European Union (EU Commission) funding was made available to fund the development of a diagnostic test and treatments. I recall Members of the European Parliament lobbying the Commission (bit that has the money) that vulnerable people were travelling to other countries to try to get a diagnosis --. Maybe we should highlight this scandal as a reason for the UK/European-- governments to fund the development of a diagnostic test and treatments @EspeMor . There's currently a petition, to the European Parliament [Petition No 0204/2019], requesting funding for the development of a diagnostic test and treatments - only EU citizens can sign the petition. There's a thread on this site regarding the petition [https://www.s4me.info/threads/eu-pe...or-funding-for-me-research.10363/#post-193653].

    Another thing that caught my attention, in this paper, is this statement:
    "Other tests of energetic dysfunction could be developed using the seahorse extracellular flux assay but more research is required as to the meaning of the results in the aetiology of CFS/ME before a test using this approach should be developed."
    My understanding is that Karl Morten's view is the same i.e. "more research is required---before a test -- should be developed".
    Ron Davis (from memory) said that a test which shows you are ill, i.e. not normal/healthy, may be of benefit [check out his presentation at the IiME Conference]. Also, there are potential treatments which could be evaluated e.g. copaxone & SS-31. On balance, I'm with Ron on this i.e. I'd like to see a test even if it we do not understand the underlying disease mechanism - risky(?) yes.
    Labels like "cancer" have turned out not to be single diseases but multiple diseases --- MS (and others like Parkinson's?) could well be spectrum's ---- ME, who knows what it will eventually be related to.

    To summarise - I'm with Ron i.e. in favour of the development of a diagnostic test now --- also, check out the EU petition and consider lobbying your politician (painful as I know).
     
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  6. Adrian

    Adrian Administrator Staff Member

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    I'm not convinced by the line they draw it could be more like clusters with severe, less severe (and healthy) (where 4 is an overlap) and this could represent the difficulty in getting and sending blood samples.
     
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  7. Adrian

    Adrian Administrator Staff Member

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    From their response
    https://www.drmyhill.co.uk/wiki/Res...vailable_laboratory_test'_by_Cara_Tomas_et_al

    This worries me in terms of the suggestion that the test needs to be done 4 times due to a high error rate. It leads to questions about which result they take or how they combine them. It also suggests some serious work needs to be done on testing the reliability and codifying the protocol in detail.

    It seems they have done some testing on timings. One thing I wondered about was this statement "Provided the samples were kept in the original ‘vacutainer’ tubes and not subjected to extremes of temperature there were only minor changes in test results up to the 48 hour point." . The first thought was what are extremes of temperature and what happens to mail (in summer, winter etc) or is there an effect if it is flown (as I think some cargo planes have unpressurized stores).

    But the comment on in the original tube was also interesting. Does for example moving tubes represent some sort of shaking which would be less than in the post? Or why else in the original container. I guess it is a vacume - looking at the name - hence perhaps its about oxidization and contamination?
     
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  8. Simon M

    Simon M Senior Member (Voting Rights)

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    I have now read the Tomas paper and the Myhill/McLaren Howard response, and think it would help to focus on some of the detail.

    Overall, the findings in the new paper appear to be pretty good evidence that the MES test isn't reliable, because it can't separate patients and controls. However, it's worth looking at both the use of frozen and fresh samples, and also the significance of processing delays.

    Main result

    The translocator components of the MES test couldn't be replicated because reagents listed in the protocol are no longer available. That left 3 elements.

    For neutrophils, there was no significant difference (fig 1) between patients and controls on any of the three remaining tests. One, for ADP to ATP efficiency, came close (p = 0.054), but there was extensive overlap between the effeciency levels patients and healthy controls (see graph). So that means it isn't clinically useful.

    Tomas-v-Myhill,-main-result.jpg

    The study is quite small, with 10 patients and 13 controls but, as the paper says, "abnormalities in CFS/ME patients should be reproducible even in small samples given the current use of the test for diagnostic purpose".

    The situation for PBMC's (white blood cells, or leucocytes) was very similar. In any case, although the MES test has switched from neutrophhils to PBMCs, there is no published data to show that this gives an effective test.


    The impact of delayed processing


    The authors were concerned that delaying analysis of blood samples for 24 hours or more would affect test results. This is because the status of the cells would change (eg neutrophil activation), cells would deteriorate and ADP/ATP-relevant metabolite levels would change. The study found substantial changes to the neutrophil fraction within 24 hours and dramatic changes to glucose levels (see graph).

    Fig-5-glucose-tomas.jpg

    Although McLaren Howard said that blood sample were tested the following day, the new study shows there would have been substantial changes in that time.

    Critically, all the neutrophil results and almost all the results with PBMCs were significantly different 24 hours on from blood draw (see fig 6).

    By contrast, McLaren Howard said that he "explored heparinised whole blood storage times in relation to patients and controls. Samples were processed within minutes of venepuncture and retested 6, 12, 24, 48 and 72 hours later. Provided the samples were kept in the original "vacutainer" and not subject to extremes of temperature there were only minor changes in test results up to be 48-hour point."

    Interesting point from @Adrian that blood sent through the post might experience considerable changes in temperature as well as a lot of agitation.

    The new paper says blood was either processed immediately or processed after 24 hours, suggesting that the blood was left in vacutainers - but this wasn't explicitly stated.

    Use of both fresh and frozen samples

    The new study deviates from the published protocol in one significant way, using frozen-then-thawed samples as well as fresh ones. "The use of fresh and cryopreserved samples was born out of necessity as samples could not be collected in a short enough time frame to conduct the experiments at the same time for all CFS/ME samples and controls". So that all samples are effectively processed within a couple of hours.

    However, the authors showed (supplementary data) that there was no significant difference in MES results between fresh and frozen samples. You can see the same pattern in Fig 1 (eg the first graph above) where frozen samples are indicated by solid markers and fresh samples by open ones.

    McLaren Howard reports a very different experience with freezing/thawing samples.

    "That is not in accord with my findings or those of several scientists from other labs who have spent time in my lab exploring these tests."

    That raises some doubt, but Tomas et al have published data to support their claim which really carries more weight. But perhaps this area needs exploring.

    Perhaps, once a little water has flowed under the bridge, the two parties can sit down and discuss how they got different results. But as things stand I think there hs to be considerable doubt over the orginal findings and continued use of the test.

    Footnote

    Within his response, McLaren Howard seems to argue that their results mus be correct because they explain fatigue problems on inceased energy demand in ME/CFS patients. "As almost all the energy for such activity is ATP-derived, one would expect to see the kind the differences that we so frequently find in our test." And the new results make it "very difficult" to explain the same fatigue. Strange to use an assumed specific biological abnormality to justify your own result and cast doubt on a null result.
     
    Last edited: Aug 20, 2019
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  9. Adrian

    Adrian Administrator Staff Member

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    This did make me wonder if the test that McLaren Howard is doing is in some way different but perhaps not well documented within his protocol that Tomas tried to follow.
     
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  10. Hoopoe

    Hoopoe Senior Member (Voting Rights)

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    It's a bit like the PACE authors who justified outcome switching by saying it made the results align with previous studies and their clinical experience (or something along those lines).

    It would make a lot of sense to me if there was some energy generation problem in ME/CFS but accepting false findings won't help us. There could still be an energy generation problem, just one that needs a different test to become visible.
     
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  11. Jonathan Edwards

    Jonathan Edwards Senior Member (Voting Rights)

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    I agree. This comment appears to reveal significant naivety about how you do scientific experiments.

    My own view is that it is extraordinarily difficult to see how the McLaren Howard results tie in with the clinical pattern of ME. I agree with Mike Murphy that ME does not seem like a primary problem of energy production at all.
     
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  12. Robert 1973

    Robert 1973 Senior Member (Voting Rights)

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    I wonder if McLaren Howard could request 20 samples from the ME biobank – 10 ME patients and 10 healthy controls – to see if his test can differentiate between the two. If he is right it should be able to. If Thomas et al are right it won’t.
     
  13. adambeyoncelowe

    adambeyoncelowe Senior Member (Voting Rights)

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    Good idea!
     
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  14. Andy

    Andy Committee Member

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    The question would be who would pay for them though.
     
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  15. FMMM1

    FMMM1 Senior Member (Voting Rights)

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    What jumps out at me from this is that the "MES test does not have the reliability and reproducibility required of a diagnostic test" & "The differences observed by the Myhill group may be down to differences in sample processing time between cohorts".
    So the test was unreliable; I assume that this could be due to the poor sampling protocol i.e. not processing samples within the required time (5 hours?).
    The fact that they initially used the wrong sample type (neutrophils rather than PBMCs) is also worrying.
    I think this emphasises why we need a properly validated diagnostic test delivered by the NIH.

    Haven't read the paper by Karl Morten et al in full.
     
  16. Milo

    Milo Senior Member (Voting Rights)

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    I wonder how much Myhill charged for this test, and comment that patients from around the world sent their samples in a desperate plea to provide biological proof that they were sick, out of necessity to show their doctors or to provide their insurers.

    It goes on to say that people must use caution when sending their money and blood/ body fluid samples to a private labs. Has their methods been validated and reproduced by an independent lab? Has the stability of the sample been tested for the sample to make its way in the mail?

    It is horribly hard to accept that something is wrong with us and yet there are no approved blood test that has been approved by the international community. And i still think about my negative XMRV results from 2009...
     
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  17. Annamaria

    Annamaria Senior Member (Voting Rights)

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    As I understand it, Dr Myhill didn't charge for the test; she charged for a letter interpreting the results; the test fee was passed on to Dr McLaren Howard.
     
  18. wigglethemouse

    wigglethemouse Senior Member (Voting Rights)

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    I think McL/H prefers fresh samples......
     
  19. Milo

    Milo Senior Member (Voting Rights)

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    Yes, I stand corrected. However the gist of it is that someone benefits from this test, directly or indirectly. For the physicians that use for-profit labs, it is their responsibility to ensure that it is a reliable lab, that quality control is assured and that the particular test has been validated.
     
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  20. Amw66

    Amw66 Senior Member (Voting Rights)

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    The ATP profiles test was based on an existing test used in labs , but which was not available clinically.

    Do we know how this predessessor differs?
     

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