Going to give some quick responses as I try to finalise a paper today:
The biggest finding to date from this team has been that there is a Complex V deficiency in the mitochondria of lymphoblasts derived from patient B cells - published in 2019. 160 patient samples have now been analysed.
Also important are substrate provision changes and elevated respiratory capacity (also mTORC1 activity in what may be a subset) but I didn't have the time for this nuance in the presentation. re: C5 deficiency - my interpretation is less a deficiency and more "less proportional usage of". It could be a deficiency but then I wonder why absolute or basal respiration are not changed on average and why electron flow is apparently unchanged. And if it were, we don't have clear evidence of an inborn complex 5 error and this would manifest earlier in life and with different consequences, looking at genetic C5 disease. So how would it manifest? Perhaps by a regulatory issue via an inhibitory factor. (I think we are pursuing this)
One new observation is that orthostatic intolerance inversely correlates with spare respiratory capacity. The worse the OI, the less spare respiratory capacity in lymphoblasts.
this came up in the original paper as well. I didn't want to show much unpublished data. I don't know what the meaning of this is, if any since they are b cell-derived. Which is why I kept the speculation minimal.
Research into lipids led to a 2024 paper showing less lipid droplets of larger size in patients vs healthy control cells.
small cohort with fibroblast primary lines
https://advanced.onlinelibrary.wiley.com/doi/full/10.1002/adsr.202300178
They have also grown fibroblasts from patient skin samples and this does not show the respiration issue that is present in immune cells. They have compared lymphoblasts and fibroblasts derived from the same patients to confirm this.
hopefully in pub soon. large part of my reasoning for currently not expecting there to be a systemic mito issue. I think it is an immunometabolic issue that involves (whether cause or consequence) abnormal substrate usage and mitochondrial function. It is all so tightly entangled. It would fit with so many different models for the clinical picture, eg: including JE's hypothesis paper. This isn't to say that there's nothing wrong in the fibro lines, we did see the preliminary lipid droplet difference and there is other stuff we're doing, but yes in seahorse nothing stood out.
The follow on work on lymphoblasts is analyzing the "things" that are different between ME/CFS and healthy control lymphoblasts and manipulating them one by one to try and find if there is a "master" regulator causing the Complex V deficiency.
not only necessarily the cause of the C5 thing, but establishing causal relationships between any parts of the phenotype that we can. Factors where the disturbances we've seen can be causal for the most parts of the picture become of increasing interest for closer investigation or as upstream targets. We are also manipulating by multiple approaches / in multiple model systems where possible
My impression the first time I heard Daniel speak was that he is a great communicator, a nice person and is clearly genuinely invested in contributing to finding the mechanism underlying ME/CFS. This presentation reinforces that impression. Thanks for the years of effort, and sticking with ME/CFS after completing your PhD Daniel.
thank you, I can seriously promise you that it is genuine. I go to sleep and wake up thinking about ME/CFS biology.
1. Why do you refer to the syndrome as ME? Wouldn't ME/CFS be better in terms of having a consistent way of referring to the syndrome?
I was asked by the organisers to do so
2. You mentioned that the degree of increase in the spare respiratory capacity and decrease in the proportional use of Complex V is correlated with orthostatic intolerance. How did you measure orthostatic intolerance? You say that this has been measured in 160 people so far - has that been published, how many controls? How clean is the separation? Have you done measurements on the same person on good days and bad days?
Don Lewis' old standing test with scoring mechanisms investigated here by Alice and Brett
https://pmc.ncbi.nlm.nih.gov/articles/PMC5898049/. Our clinical phenotyping for studies moving forward will be more standardised and broad (as it has been in the current mason study w 100 participants) but this was one way to benefit from the older CFS discovery samples (it's one of the useful bits of clinical data we have for those people that's common to the newer samples)
3. Do you think the H+ are getting used by something other than Complex V? You point to the carntitine/acylcarnitine carrier a production of NADH. Can you say more about that? (There is an interesting chart at about 9 minutes.)
Yes this would be the straightforward interpretation - "used somewhere else" and we see the other proportional subcomponents of basal respiration are elevated (proton leak and nonrespiratory oxygen consumption). Narrowing that down specifically could be a series of PhD projects in itself. We have not tested anything directly so I won't speculate too much
My thinking is moving to: how might particular immunometabolic abnormalities drive or result from abnormal response to signal. The paper I am trying to finish today has identified a gene of interest that I am trying to wrap my head around.
I find it fascinating that lymphoblasts used in the work derive from B cells given the fact that Chris Armstrong & Mensah (Jo Cambridge lab) found energy issues in young B cells that caused changes in maturation. And now we have the Edwards, Cambridge, Cliff hypothesis of "junk" antibodies derived from B cells could be affecting macrophages. Could there a causative link between all three?
Yes and Chris/Jo/Fane's B cell paper is concordant with what we have seen. And yes it is easily possible that metabolic changes are consequential from or contribute to the dysfunction proposed in these particular hypotheses by they or Jonathan or whoever else. I am open to and interested in all of them and think it is all part of one picture. However the big question marks are that the environment is in culture conditions and that the cells I have used are transformed which causes fairly sweeping epigenetic changes. The good news is that we are doing epigenetic profiling of these lines to see what has persisted into culture and survived the transformation which will hopefully help us relate what we are seeing backwards. But we are also moving into other types of sample as well.
This is from the transcript at the end of the webinar
If I was even half as coherent as that transcript suggests I am shocked, but yes what I was simply trying to get at there is that a) we don't know cause from effect yet and b) there is no clear evidence of an underlying genetic mitochondrial defect so speculating based on the most straightforward alternatives makes some sense (in this case what came to mind as an example was a homeostatic shift arising from possibly subtle, simultaneous changes in genetic background or
environment)
Anyway one of my biggest goals for this year is to nail down plausible,
specific links between all of these elements based on what is already known (and to identify the gaps) to form the basis for a) ME/CFS-specific hypothesis testing b) basic science that I am becoming particularly interested in filling in, that will benefit ME/CFS in turn. A large part of this may come from a gene of interest that I have identified using multi-omics in a draft paper (the one I am trying to finish today).
