Whole blood human transcriptome and virome analysis of ME/CFS patients experiencing PEM following cardiopulmonary exercise testing, 2019, Chiu et al

Andy

Andy

Senior Member (Voting rights)
Abstract
Myalgic encephalomyelitis / chronic fatigue syndrome (ME/CFS) is a syndrome of unknown etiology characterized by profound fatigue exacerbated by physical activity, also known as post-exertional malaise (PEM). Previously, we did not detect evidence of immune dysregulation or virus reactivation outside of PEM periods.

Here we sought to determine whether cardiopulmonary exercise stress testing of ME/CFS patients could trigger such changes. ME/CFS patients (n = 14) and matched sedentary controls (n = 11) were subjected to cardiopulmonary exercise on 2 consecutive days and followed up to 7 days post-exercise, and longitudinal whole blood samples analyzed by RNA-seq. Although ME/CFS patients showed significant worsening of symptoms following exercise versus controls, with 8 of 14 ME/CFS patients showing reduced oxygen consumption (
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) on day 2, transcriptome analysis yielded only 6 differentially expressed gene (DEG) candidates when comparing ME/CFS patients to controls across all time points. None of the DEGs were related to immune signaling, and no DEGs were found in ME/CFS patients before and after exercise. Virome composition (P = 0.746 by chi-square test) and number of viral reads (P = 0.098 by paired t-test) were not significantly associated with PEM. These observations do not support transcriptionally-mediated immune cell dysregulation or viral reactivation in ME/CFS patients during symptomatic PEM episodes.

Open access at https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0212193
 
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It’s so good to see Stevens, Van Ness, and Snell pair up with someone like Charles Chiu.

At first glance, the study design seems excellent. They tested before, during, and after PEM induced by the two-day CPET.

Canadian criteria, all female. N=14.

Used the technique called RNA-seq. Only looked at RNA, not DNA. Whole blood, not cell-free plasma.

Bouquet et al. 2019 said:
We detected sequences from a small number of viruses in the RNA-seq data, including enterovirus A, influenza A virus, anelloviruses/torque teno viruses (TTVs), and human herpesviruses (HHVs) (Table 5).
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The overall virome composition in ME/CFS did not differ significantly from controls (P = 0.746 by chi-square test)
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Reasons for the absence of differential gene expression between ME/CFS patients and controls include
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(3) localization of ME/CFS pathogenicity to a specific tissue (e.g. skeletal muscle or brain tissue) rather than blood
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here, we observed no differences in viral abundance in ME/CFS patients following exercise.
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A limitation of the current study is the small size of the study cohort.
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Null results.

That’s what I’ve gleaned from a first glance...
 
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here, we observed no differences in viral abundance in ME/CFS patients following exercise
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Based on my n=1 experience this doesn't surprise me. I do get very frequent reactivations of HHV1 (based on look & location of blisters, not on lab tests) but they don't occur any more frequently during PEM than at baseline. The only link between my herpes outbreaks and my PEM is the other way round: during a herpes outbreak it takes much less exertion to trigger PEM. Which would pose some practical challenges if anyone ever wanted to study that relationship...
 
Ravn said:
Based on my n=1 experience this doesn't surprise me. I do get very frequent reactivations of HHV1 (based on look & location of blisters, not on lab tests) but they don't occur any more frequently during PEM than at baseline. The only link between my herpes outbreaks and my PEM is the other way round: during a herpes outbreak it takes much less exertion to trigger PEM. Which would pose some practical challenges if anyone ever wanted to study that relationship...
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But healthy people reactivate viruses too on a regular basis, and according to a recent study, astronauts on the international space station do too. Stress does that. PEM is a form of {physical} stress.

It seems clear to me that we should put the virus/infectious hypothesis to rest and focus on mechanism of illness/biomarkers.
 
Even if viruses are involved, it is not likely that it is by them multiplying after exertion. A more likely hypothesis is that they interfere with energy production all the time. Exercising to exhaustion causes damage to the cells and a build up of lactic acid and other toxins.

Healthy people have plenty of excess energy available to mop up all the damage so that within 24 hours they are back to where they started.

People with ME have a very limited amount of energy available for mopping up duties - the Workwell studies show that aerobic respiration is compromised so we rely on the short term anaerobic respiration too much - so that it takes much longer for them to get back to normal.

Their precious energy is now split between repairing yesterday's damage and doing today's task so they will not be able to do so much.

Increasing viral loads or activating the immune system does not seemed to be needed to account for what goes wrong. Just think about someone on minimum wage who has to pay for a school outing, there may just not be enough in the pot for it, without needing any other explanation.

In the long term, there could be consequences of immune activation or viral loads but that would not be found here.
 
Mithriel said:
People with ME have a very limited amount of energy available for mopping up duties - the Workwell studies show that aerobic respiration is compromised so we rely on the short term anaerobic respiration too much - so that it takes much longer for them to get back to normal.
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Yes. Agreed that this seems to explain PEM that arises from physical exertion. However, I'm not sure it does with intellectual exertion like doing math or heavy reading or concentrating. I suppose HR might increase sometimes for this, but many times I think it will not - like in the instance of reading an abstract, no problem, but an entire study, PEM will be an unfortunate by-product, even though I can pretty much guarantee my HR and BP and breathing etc did not change. (This is one of the reasons I think that that whole low carbon tidal pool results from hyperventilating is nonsense, but that's a side bar.)

So if we look at PEM as a discrete phenomenon (albeit part of ME/CFS) then it may be somewhat incongruous to think that different mechanisms are involved for different categories of PEM (physical vs intellectual vs emotional etc). Or not.
 
Milo said:
It seems clear to me that we should put the virus/infectious hypothesis to rest and focus on mechanism of illness/biomarkers.
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Not at all. Although people have talked about viruses ad nauseum for decades, there has been very little true high-quality research to actually look for viruses. This paper appears to be high-quality research, but is just the beginning of the true hunt for viruses.

So we’ve looked in the blood. Next, we’ll look in the CSF after an exercise challenge. Then we have to look in the lymph nodes after an exercise challenge. Then we have plenty more organs in the body to test. It will be a long road to do a real high-quality search for viruses. And, unfortunately, the current technology and lab techniques are woefully inadequate to find viruses, given our modern understanding of the varied complexities of viral persistence.

Mithriel said:
Even if viruses are involved, it is not likely that it is by them multiplying after exertion.
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It’s too early to make such a claim. We are still early in the process, and all of us have to be careful about jumping to conclusions.
 
I can't see how viruses could replicate enough in 24 hours to cause the problems. Exercise would have to let something change so they could thrive then they would have to go back to lower numbers as people recovered. I don't think viral loads work on this sort of timetable. It may be the answer for people who become very sick for months and, as you say, viruses are tricky little beasts we still don't know much about.

My main thought is that this study does not add any evidence that viruses are NOT involved in ME.

Duncan, you make good points. We have to remember that people rarely reach maximal exertion in real life. What the Workwell studies indicated was where things go wrong in our bodies. Basically, they magnified the problem so it could be seen.

It may not be the same for everyone, but I come to a stop for a few seconds then can carry on. As I have become worse I can do very little but that was the pattern even when I was still going to school, it was how I managed.

Intellectual effort will be much the same. Heart rate is useful, but only up to a point.

I hope the disease process of ME will be worked out soon but the anaerobic threshold problems will be part of the solution and help show the way even though they are unlikely to be the final answer.
 
I am revisiting this.

It seems notable because the people assessing the 2-day CPET results were blinded. 8 of 14 ME/CFS patients showing reduced oxygen consumption on day 2. My feeling, without checking, is that this seems a little less clear cut than previous (unblinded) studies. Still there is a real effect.

Virome composition (P = 0.746 by chi-square test) and number of viral reads (P = 0.098 by paired t-test) were not significantly associated with PEM.
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Note that none of the patients had enterovirus A reads (one of the controls did). Table 5 looks like patients had fewer viral reads than controls.

Sooo... what I take away from this is that even with narrow criteria there is no trace of enteroviruses despite an exercise stressor that's enough to provoke measurable PEM in most patients. The enterovirus hypothesis seems defunct.
 
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I wonder what would have been found 3/ 4/ 7 days after.PEM may start on 2nd day but may be nowhere near its peak.
 
strategist said:
I am revisiting this.

Sooo... what I take away from this is that even with narrow criteria there is no trace of enteroviruses despite an exercise stressor that's enough to provoke measurable PEM in most patients. The enterovirus hypothesis seems defunct.
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The reason enteroviruses can become chronic, in heart disease for instance, is that they do not enter the blood stream very often. Other viruses hijack the cell mechanism to make so many copies of themselves that they burst the cell open and spread to other cells. Enteroviruses replicate but live on in the cell. The immune system detects viruses by investigating damaged cells so this is a good way to evade the immune defences.

There is new work which says that enteroviruses use exosomes to invade nerve cells but I am not so sure of the details of that.

The test used for enterovirus was the VP1 test which looked for actual viral particles. Unfortunately this research showed virus in muscles and brain tissue just before CFS was invented and everything went wrong for us.
 
But like they write themselves, blood is not a good place to do transcriptomics (or, I guess it depends what one looks for, I would be more interested in muscle or gut tissue personally, but that's way more invasive).
 
Mithriel said:
The reason enteroviruses can become chronic, in heart disease for instance, is that they do not enter the blood stream very often. Other viruses hijack the cell mechanism to make so many copies of themselves that they burst the cell open and spread to other cells. Enteroviruses replicate but live on in the cell. The immune system detects viruses by investigating damaged cells so this is a good way to evade the immune defences.
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They did get enterovirus reads. In one of the healthy controls.
 
It is not ever in the blood stream just not enough for a negative test to mean it is absent from the body.
 
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