An Isolated Complex V Inefficiency and Dysregulated Mitochondrial Function in Immortalized Lymphocytes from ME/CFS Patients Missailidis et al. 2019

The results are certainly eye-catching. I've not yet read the full paper (and am hoping to avoid having to) . I wonder if those good people who made the effort could answer a few questions.

1. How do the Seahorse results from this study fit in with previous Seahorse results? I am aware that while previous studies have found differences between patients and controls, there have been a lack of consistency in findings. Related to that, could the complex V issue explain the previous Seahorse results?


2. The generic issue question for all studies finding mitochondrial problems: how does this fit with the relatively normal day-one maximal exercise test results? The deficits don't appear until the second mximal exercise test 24 hours later.


3. And what about cell imortalisation? That seems to me like a big step. Do the authors discuss how to check if the differences are also apparent in more real-life situations?

 

Good point.

 

Meaning that researchers should focus on tissue cells and not blood cells?

 

Sudden onset is not really for the start of ME but for the infection which then becomes ME. Before CFS it was believed that you got a common infection, often enteroviral, but developed an abnormal consequence of that disease. That is the way polio works and other enteroviruses are very closely related.

Gradual onset was possibly a subclinical infection so you did not realise you had it but the ME developed as a consequence anyway.

In sudden and gradual onset ME the ME symptoms can gradually become worse.

 

This is a reasonable question, but what is often overlooked in the descriptions of ME is that there is a different way that overexertion becomes apparent.

I have a typical 3 day delay after I do too much. That is when I begin to get sore throats, swollen glands and be unable to do very much. But day to day, I get a problem where I am doing something then I have to stop, wait a minute or so then I can go back. Lots of little things like not being able to say a sentence, or lift a cup without a quick rest.

I do not know what it means for this research or CPET testing though.

 

The CPET is only showing us a small part of what is occurring. The studies on cells are not showing what happens when a CPET is done.

The CPET must have immediate effects (or it could not have any delayed ones), we just have found them yet.

 

Perhaps compensatory mechanisms manage to even things out on day one, and then they fail by the time day two comes around. Or maybe it is indeed something different, and the second day results show a failure of sleep to clear away the improper signalling so that the effects are cumulative the next day.

 

Maybe we're looking at a series of compensatory mechanism at different levels. Fisher seems to have found one at individual cell level, maybe being activated rapidly, perhaps within minutes. There could be others maybe at brain level that take longer to become active, maybe more at typical PEM timeframes. It's still too early to make sense of the whole picture.

 

What a great comment, so many interesting ideas, but over my head unfortunately. What I did wonder though; ur saying the genetic makeup of stem cells/early immune cells can permanently change because of environmental (in a broad sense) factors?

And regarding what u said about some irreversible signal; what cells/systems have the authority(?) to override bone marrow development like that? This was new to me

Hope my questions make sense

 

Just thought I would post this again. Paul Fisher really does an excellent job explaining this work in the video of this thread. Well worth a watch. [Some slides were blacked out pre-publication].
https://www.s4me.info/threads/video...nsatory-changes-in-me-cfs-patient-cells.9177/

Direct link to video : https://mecfsconference.org.au/videos/paulfisher/

Not sure if you read the bit where they describe how they exposed lymphocytes to EBV virus to create the lymphoblasts. So they were doing something at this point. I'm wondering if this is why they saw the clear mTORC1 difference in these particular cells which might not be present in other cells doing nothing. i.e. you only see a clear difference when cells are doing something. I think in Maureen Hanson's study she had to activate T-Cells to see a difference on the Seahorse. In her case the T-cells needed to be doing something to see a difference.

It's so frustrating isn't it. It took me a whole day to read the paper, a bit at a time. Wish I was well enough to go back and help answer your questions. Sucks so bad. I'm sure I speak for many when I say I wish you were well enough to write a "Janet and John"/Ladybird version of the paper on your blog ........

 

@Jonathan Edwards Have you heard any other researchers notice lymphocytes dying faster in ME patients? This is what @Hutan posted from the paper. If this is the case what would the health implications be of faster recycling of lymphocytes or would this likely be a cell culture effect not seen in the body?

 

I don't believe that they do. However they have used the same process of analysis in other diseases previously and the mitochondrial dysfunction is different here

As @Hutan posted Lymphocyte basal OCR is difficult to measure as the level is approaching the measurement limit of the instrument used. Lymphoblasts OCR is 100x or more, so good repeatable measurements are possible.


It makes me mad to think Paul Fisher has wanted to work on ME for 10 years and only recently found a way in [from video]. His first work on ME finds this!

 

Has anyone checked their ATP gene results (I've only got the 23andme) and put them in Enlis.

or does anyone know if any of the genes have been found in research to be significant for ME?

 

@Andy - is Paul Fisher, or anyone else from this team, on your interview list?

It would be very interesting to hear how they view some of the issues brought up in this thread, especially:

1) the pros and cons of using of immortalised lymphocytes rather than other (non-blood) cell types; do we have the technology to test for Complex V problems in other cell types and, if so, would doing such a test be a good confirmation of Complex V problems (CVP) in ME?

2) how their findings fit with the 'thing in the blood' findings - are they mutually exclusive?

Thought experiment:

If we view the 'thing in the blood' (X) to be a factor that causes cells to need to produce more energy, there wouldn't be any contradiction with parts of the nano-needle findings:

ME cells (with CVP) in ME plasma (with X) need to produce more energy but can't. That fits.​

ME cells (with CVP) in healthy plasma (without X) don't need to produce more energy so they look normal despite their CVP. That also fits.​

Healthy cells (without CVP) in healthy plasma (without X) are fine. Fits.​

But this bit of the nano-needle findings is more difficult to explain:

Healthy cells (without CVP) in ME plasma (X) are acting ill. Yet their complex V should be just fine and so they shouldn't have any trouble producing the extra energy. So there's an apparent contradiction here.​

Could we design an experiment where we immortalise healthy cells and test some of them for Complex V to get a baseline. Then we take some and place them into ME plasma for a while to expose them to X, then clean them up again so no factor X remains, and then test those cells for CVP.

If the result is that the previously healthy cells now have a newly acquired CVP, that would indicate X may be causing it and it's permanent. It would resolve the apparent contradiction between the CVP and X models.

If the result is that the cells do not have any CVP the contradiction remains and the head-scratching continues.

Does any of this make sense?

 

He's somebody I intend to reach out to but I haven't yet.

 

This is the problem. It seems unlikely that other cells/systems can do that. The genetic material should not be changed. The effect should be epigenetic, but in general epigenetic changes involved in cell maturation and function do not survive transformation into immortalised lines.

 

I am unsure exactly what immortalisation of cells means ( guessing that they don't die off and keep dividing?)

Epigenetics ( in mice ) have reported mutations as being heritable - would such mutations not be preserved in immortalisation ?

 

Yes, good question, it would be useful for people to have a handle on this.

Immune cells / white blood cells in general derive from bone marrow stem cells that can decide to follow one of several maturation paths. They can become B lymphocytes or macrophages or even red cells. To follow a path the cell has to rearrange methyl groups stuck to the DNA (and probably other mechanisms) to allow some packs of genes to be read and others not. DNA is a bit like a washing machine manual in 20 different languages. The methyl groups are like sellotape stuck around all the languages except your own so you can only read that.

So the DNA stays the same throughout but a B lymphocyte has sellotape stuck around all the genes for doing what T cells and neutrophils etc. do. (With epigenetic traits carried across generations in mice it is the same - there is no mutation as such.)

Maturation along a path through methylation is programmed so that the cell goes through a series of life stages and then dies - just as the whole body is. It works a bit like clicking through the pages of a website where you cannot go back. So almost all white blood cells isolated in the lab will die within days. During that time you can get them to mature e.g. from naive B cell to antibody secreting plasma cell, but they then die and generally do not divide. (Cells like fibroblasts are rather different, they can divide about 30 times in culture and only then die.)

EBV has evolved DNA that has a specific ability to get into B cells and to override the methylation control signals in the nucleus so that the cell effectively becomes like a stem cell or tumour cell in that it can keep dividing for ever. It remains committed to being a B cell, so is not actually a stem cell, but it no longer has any of the normal B cell maturation control instructions. It is as if you were sent a new manual for the washing machine in you language which said put sellotape around the whole of the old manual, open the back of the machine and press the button saying 'do not press' and your machine will repeat washing cycles continuously for ever.

The issue that both Simon and I are concerned about is that if you override epigenetic control in this way you tend to lose any memory of previous epigenetic signals. (Although not all because the B cell remembers to be a B cell. In a high grade lymphoma cancerous B cells even forget that they are B cells - they are just berserk cells without a name.) The puzzle would be to think what in ME might induce an epigenetic shift in B cells that would still be detectable once you had screwed up maturation control with EBV.

Such a persisting epigenetic shift is not impossible. Say you had set your washing machine to 'coloureds, quick wash' then pressing the button at the back might give you endless cycles of coloured quick wash. However, immortalisation has dramatic effects on cell metabolism because it switches on active synthesis of all sorts of things and it would be a bit surprising if a previous signal affecting activity levels stayed detectable.

 

Cancer has always seemed to me to be like embryiogenesis ( apologies if I have just invented a word) .

If something can signal an obsolete process to start up, perhaps it is similar in ME? Perhaps it is a hijacked process either preserved but not generally active, or an ancient bit of code activated by something equally as ancient?

Apologies for taking this thread completely off subject and probably making absolutely no sense. Lack of sleep has a lot to answer for.

 

Thank you @Jonathan Edwards . I always find your explanations helpful and entertaining