Abnormalities of AMPK Activation and Glucose Uptake in Cultured Skeletal Muscle Cells from Individuals with Chronic Fatigue Syndrome, 2015, Brown+

[jnmaciuch]

Abnormalities of AMPK Activation and Glucose Uptake in Cultured Skeletal Muscle Cells from Individuals with Chronic Fatigue Syndrome (2015)

Audrey E. Brown, David E. Jones, Mark Walker, Julia L. Newton

Background
Post exertional muscle fatigue is a key feature in Chronic Fatigue Syndrome (CFS). Abnormalities of skeletal muscle function have been identified in some but not all patients with CFS. To try to limit potential confounders that might contribute to this clinical heterogeneity, we developed a novel in vitro system that allows comparison of AMP kinase (AMPK) activation and metabolic responses to exercise in cultured skeletal muscle cells from CFS patients and control subjects.

Methods
Skeletal muscle cell cultures were established from 10 subjects with CFS and 7 age-matched controls, subjected to electrical pulse stimulation (EPS) for up to 24h and examined for changes associated with exercise.

Results
In the basal state, CFS cultures showed increased myogenin expression but decreased IL6 secretion during differentiation compared with control cultures. Control cultures subjected to 16h EPS showed a significant increase in both AMPK phosphorylation and glucose uptake compared with unstimulated cells. In contrast, CFS cultures showed no increase in AMPK phosphorylation or glucose uptake after 16h EPS. However, glucose uptake remained responsive to insulin in the CFS cells pointing to an exercise-related defect. IL6 secretion in response to EPS was significantly reduced in CFS compared with control cultures at all time points measured.

Conclusion
EPS is an effective model for eliciting muscle contraction and the metabolic changes associated with exercise in cultured skeletal muscle cells. We found four main differences in cultured skeletal muscle cells from subjects with CFS; increased myogenin expression in the basal state, impaired activation of AMPK, impaired stimulation of glucose uptake and diminished release of IL6. The retention of these differences in cultured muscle cells from CFS subjects points to a genetic/epigenetic mechanism, and provides a system to identify novel therapeutic targets.

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[Jonathan Edwards]

Post exertional muscle fatigue is a key feature in Chronic Fatigue Syndrome (CFS).

That first sentence is odd both in that PEM is considered a specific feature of ME/CFS but was not for 'CFS' as previously known and in that this isn't PEM. Do people with ME/CFS have post-exertional muscle fatigue?

And in cultures, genetic or long erm epigenetic differences might persist but they are unlikely to reflect post-exertional changes.

 

[jnmaciuch]

I saw there wasn't already a thread on this paper. Since I mentioned it in a comment here, I wanted to discuss it a bit. Main limitations are small sample size and Fukuda criteria.

However, I recently came back to this study and had to admit that despite these limitations, I think these findings are actually quite important (as well as other findings from Newton/Walker using these same muscle cultures). As they say in the abstract, using muscle cultures enables you to get rid of a lot of possible confounders that tend to plague -omics studies or studies which directly assess blood/tissue samples.

I do have some critiques, particularly in Fig 2A. Left is control, right is ME/CFS (this wasn't labeled on the chart or in the legend, just mentioned in the text). I would have liked to see comparisons of these levels between ME/CFS and control, not just comparisons between timepoints.

 

[jnmaciuch]

And in cultures, genetic or long erm epigenetic differences might persist but they are unlikely to reflect post-exertional changes.

I think you're right [edit: that it doesn't reflect post exertional changes], but actually I think the benefit of this study is showing those long term changes rather than post-exertional changes. The options are genetic or epigenetic (as you said), or a positive feedback loop that is able to be maintained only in muscle cells. Despite the limitations I think those differences, whatever they may be, are rather important as they suggest a persistent change which can be maintained even in cultured cells that becomes relevant during activity.

[Edit: this difference is not PEM itself, but could absolutely be the basis for triggering PEM or at least concurrent with the trigger]

 

[jnmaciuch]

Do people with ME/CFS have post-exertional muscle fatigue?

I absolutely do, others have mentioned as well

 

[Utsikt]

I absolutely do, others have mentioned as well

I do as well. When I exercised for more than a year, the new physio was shocked when I still had severe muscle fatigue, pain and DOMS towards the end. Normally, you would expect that to stop being an issue after a few months at most.

In hindsight, it should have been a clue for that exercise maybe wasn’t the best idea for me.

 

[Jonathan Edwards]

I absolutely do, others have mentioned as well

So what exactly is 'muscle fatigue'? I can understand fatiguability - which is post-exertional weakness. I can understand muscle pain. But I am unclear what 'muscle fatigue' would be.

 

[jnmaciuch]

So what exactly is 'muscle fatigue'? I can understand fatiguability - which is post-exertional weakness. I can understand muscle pain. But I am unclear what 'muscle fatigue' would be.

Weakness would be the best way to describe it I suppose. Feels like what is often attributed to lactate buildup in anaerobic exercises. Inability to continue reaching maximal capacity, takes much more effort to give the same output.

 

[Simon M]

I saw there wasn't already a thread on this paper. Since I mentioned it in a comment here, I wanted to discuss it a bit. Main limitations are small sample size and Fukuda criteria.

However, I recently came back to this study and had to admit that despite these limitations, I think these findings are actually quite important (as well as other findings from Newton/Walker using these same muscle cultures). As they say in the abstract, using muscle cultures enables you to get rid of a lot of possible confounders that tend to plague -omics studies or studies which directly assess blood/tissue samples.

I do have some critiques, particularly in Fig 2A. Left is control, right is ME/CFS (this wasn't labeled on the chart or in the legend, just mentioned in the text). I would have liked to see comparisons of these levels between ME/CFS and control, not just comparisons between timepoints.

This appeared to be a promising line of inquiry when it came out (I'm pretty sure Julia Newton presented about this work at the conference). But either it wasn't followed up, or perhaps it was and proved a dead end, with results not published. That has often been the way of things.

 

[Kitty]

So what exactly is 'muscle fatigue'?

For me it often takes the form of heaviness. As if someone's turned gravity up a few notches, so everything takes twice the effort to do.

 

[jnmaciuch]

This appeared to be a promising line of inquiry when it came out (I'm pretty sure Julia Newton presented about this work at the conference). But either it wasn't followed up, or perhaps it was and proved a dead end, with results not published. That has often been the way of things.

The AMPK-related findings seemed to be corroborated in other work by this group, in 2018 and 2020. It looks like since then, both first authors probably graduated and moved on. Mark Walker seems to be the last author with the specific wet lab expertise that was probably overseeing these studies, though his focus is diabetes not ME/CFS.

Given the later findings it seems more like it could have been promising, but they just didn't have people interested in continuing this work. I've emailed Julia Newton to see if there's any chance at all that they still have these muscle cultures but it's probably a long shot.

 

[jnmaciuch]

Overall my thoughts are that this study does have limitations, but there is something important here which points to either an epigenetic change or some positive feedback loop maintained by muscle cells. I'd be very interested in finding out exactly what that change is.

 

[Simon M]

The AMPK-related findings seemed to be corroborated in other work by this group, in 2018 and 2020. It looks like since then, both first authors probably graduated and moved on. Mark Walker seems to be the last author with the specific wet lab expertise that was probably overseeing these studies, though his focus is diabetes not ME/CFS.

Given the later findings it seems more like it could have been promising, but they just didn't have people interested in continuing this work. I've emailed Julia Newton to see if there's any chance at all that they still have these muscle cultures but it's probably a long shot.

I’ll be interested to hear what Julie Newton says. It was also PhD, she using some kind of 3-D Muscle cell culturing technique, with electrical stimulation, designed to give more realistic results. But that found very little and the PhD author decided it showed that CBT was the answer (despite presenting their evidence this was the case).

What I don’t understand is, if you’re on a really hot lead for a poorly understood illness that affects so many people, why wouldn’t you try harder to keep it going, even if it’s by trying to interest other research groups?

 

[Utsikt]

So what exactly is 'muscle fatigue'? I can understand fatiguability - which is post-exertional weakness. I can understand muscle pain. But I am unclear what 'muscle fatigue' would be.

I experience many different things.

1. My muscles tire out more easily (even before I got deconditioned)
2. I experience severe muscle pain, stiffness and soreness
3. It get’s harder to get my muscles to react, like the signals are delayed or tuned down
4. It feels like my muscles are under many G
5. I get both visible and invisible tremors and shaking

I don’t know what would qualify as «muscle fatigue».

 

[jnmaciuch]

It was also PhD, she using some kind of 3-D Muscle cell culturing technique, with electrical stimulation, designed to give more realistic results. But that found very little and the PhD author decided it showed that CBT was the answer (despite presenting their evidence this was the case).

Do you happen to have a link for that? I hadn't seen this come up in my googling but perhaps I just wasn't using the right search terms.

What I don’t understand is, if you’re on a really hot lead for a poorly understood illness that affects so many people, why wouldn’t you try harder to keep it going, even if it’s by trying to interest other research groups?

I can understand it to an extent, even if seeing it in others makes me extremely disappointed. There's a lot of structural difficulty that makes studying ME/CFS much harder than other diseases--I've only been earnestly trying to get people [edit: both inside and outside of the field] interested in helping me for a few months so far and it's been like pulling teeth the entire time.

Is that a good excuse for the field as a whole? Absolutely not. Is it going to stop me? No. But I can see how it might just exhaust someone who doesn't have personal ties providing sustained motivation through all of that.

 

[Utsikt]

Is that a good excuse for the field as a whole? Absolutely not. Is it going to stop me? No. But I can see how it might just exhaust someone who doesn't have personal ties providing sustained motivation through all of that.

I think there might be plenty of other strong motivators than just personal ties, though. People start businesses all the time because they just want to - it’s rarely something as personal for them as having ME/CFS or close family with it. They just have a desire to achieve that particular thing.

I completely understand that it must be extremely frustrating to work uphill, and that it is financially insecure and unstable. But that’s also how many other professions are, so I don’t buy that it’s such an insurmountable barrier in academia. The contrast is probably that it’s possible to do academia without that struggle by going safe (at least to some degree), so some people are not in academia with an intention to overcome those struggles.

 

[Holinger]

So what exactly is 'muscle fatigue'? I can understand fatiguability - which is post-exertional weakness. I can understand muscle pain. But I am unclear what 'muscle fatigue' would be.

I would describe it as an awareness of feeling in the muscles. A ‘lactic’ type feeling with a bit of pain but mainly a heavy-ness fatigue feeling. They have done muscle biopsies in Australia on us people but no lactic acid was found during a crash event.

 

[jnmaciuch]

I completely understand that it must be extremely frustrating to work uphill, and that it is financially insecure and unstable. But that’s also how many other professions are, so I don’t buy that it’s such an insurmountable barrier in academia. The contrast is probably that it’s possible to do academia without that struggle by going safe (at least to some degree), so some people are not in academia with an intention to overcome those struggles.

Not saying it's insurmountable for an individual to overcome those struggles. But the hard part is convincing others to be interested, since these projects are rarely possible as the work of just one person. You may be able to achieve that with a handful of students or paid staff members, as it seems like Newton did for a few years. But I've seen labs have to shut down for this reason even if the PI themselves was extremely motivated, despite showing up to every recruitment event and reaching out to students and putting up job listings.

I can't emphasize enough that simply being highly motivated, reaching out to tons of people, and making a great pitch is not enough to get others interested if there isn't already some interest in your niche topic. At least in a business you can just pay someone to help if you know money is going to be coming in--but even though salaried staff scientists are a thing, it's a different ball game in academia. It becomes a matter of pure luck rather than a matter of effort or caring enough.

 

[Utsikt]

It becomes a matter of pure luck rather than a matter of effort or caring enough.

That’s business for you as well. Companies go bankrupt if one large contract doesn’t pay for an extended period of time for any reasons, even if they are otherwise profitable. Funding is scarce and expensive.

20 % of new companies fail within one year. 45 % within five. 66 % within ten. Some are simply not fit for survival because they have a bad product, but so many good ideas never survive because of other aspects of running a business.

So as an economist, I struggle to understand why there is so much talk about how difficult it is to survive in academia, because it sounds just like what everyone in my world experience. And as a former consultant, I’m used to spending a large proportion of the time working on proposals, offers, etc. to have something to do the next months. I’d estimate that you’re lucky if you can bill >2/3 of your hours to a client, so 1/3 in the long run is spent on admin, internal stuff (required by regulations, etc.) or trying to get more work. And unless you’re working in one of the truly massive companies, your job is eventually at risk.

Not saying that it makes it any easier to be in academia, just that the conversation (mostly elsewhere) seems kind of detached from what everyone else struggles with as well.

 

[jnmaciuch]

Not saying that it makes it any easier to be in academia, just that the conversation (mostly elsewhere) seems kind of detached from what everyone else struggles with as well.

I agree, I don't think academics necessarily have it harder than anyone else and that's not the discussion I'm trying to invoke here. More specifically trying to answer the question of why you see so many seemingly promising findings never get followed up on in ME/CFS research. That's where I think the difficulty in attracting the interest of others has to be invoked. IMO it's the more plausible explanation than assuming that the group did keep trying to keep investigating in earnest but the findings just didn't hold up.

Also you wouldn't believe how allergic many academics are to looking back through older literature to see if this exact situation happened and there's an interesting thread that hasn't been fully explored.