Reduced glycolytic reserve in isolated natural killer cells from ME/CFS patients: A preliminary investigation, Nguyen et al, 2018

Indigophoton

Indigophoton

Senior Member (Voting Rights)
Abstract
BACKGROUND:
Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) is medically unexplained post-exertional fatigue associated with significant reduction in natural killer cell (NK) cytotoxicity activity. Cytotoxic activity relies on glycolytic flux and mitochondrial respiration to fulfill energetic cellular demands. While mitochondrial dysfunction has been reported in ME/CFS patients, no previous investigation has examined the bioenergetic profile of isolated NK cells from ME/CFS patients.

OBJECTIVE:
This study was to determine the metabolic function in resting NK cells from ME/CFS patients.

METHOD:
Six ME/CFS patients (aged 50.33±4.95) were age and sex-matched with non-fatigued healthy controls (aged 50.00±5.04). Mitochondrial stress tests measured parameters of mitochondrial function in the NK cells including basal respiration, ATP production, proton leak, maximal respiration, spare respiratory capacity and bioenergetic health index. Glycolytic stress tests measured parameters of glycolytic function such as glycolytic reserve, glycolysis and glycolytic capacity in isolated NK cells from ME/CFS patients and healthy controls using an extracellular flux analyzer, Seahorse XFp.

RESULT:
There was a significant reduction of glycolytic reserve in resting NK cells from ME/CFS patients (0.6±0.07 mpH/ min) compared with healthy control (2.25±1.3 mpH/min). Mitochondrial respiration in resting NK cells did not approach statistical significance between ME/CFS patients and healthy controls.

CONCLUSION:
These findings suggest resting NK cells from ME/CFS patients have reduced ability to increase glycolytic flux to respond to high energetic demands for ATP production. Hence, the reduced glycolytic reserves we have identified in isolated resting isolated NK cells should be further investigated to assist in understanding ME/CFS pathogenesis.
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https://www.ncbi.nlm.nih.gov/pubmed/29981562

The DOI (digital object identifier, aka ID number) for this doesn't seem to work, so I'm not sure anything beyond the abstract has been published yet.
 
Trish said:
Hmm. So they took 6 ME/CFS patients and 6 healthy controls, measured lots of things to do with NK cells and energy, but are only reporting statistical significance for one of those things. So it's very much a tiny preliminary study.
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Yes.

Glycolytic stress tests measured parameters of glycolytic function such as glycolytic reserve, glycolysis and glycolytic capacity in isolated NK cells from ME/CFS patients and healthy controls using an extracellular flux analyzer, Seahorse XFp.
RESULT:
There was a significant reduction of glycolytic reserve in resting NK cells from ME/CFS patients (0.6±0.07 mpH/ min) compared with healthy control (2.25±1.3 mpH/min).
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But that looks like quite a big difference in glycolytic reserve in this sample - the standard deviations suggest discrete results. I think a number of researchers are using the Seahorse tool to research ME. This is something to keep an eye out for as they report their results.

I don't know what a glycolytic reserve is or its relevance, of course.
 
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According to the instruction manual for the Seahorse test used in this paper,
The Agilent Seahorse XF Glycolysis Stress Test is the standard assay for measuring glycolytic function in cells.

By directly measuring the extracellular acidification rate, (ECAR).... The Seahorse XF Glycolysis Stress Test provides a standard and comprehensive method to assess the key parameters of glycolytic flux: Glycolysis, Glycolytic Capacity, Glycolytic Reserve, as well as nonglycolytic acidification.

Glycolysis and oxidative phosphorylation are the two major energy-producing pathways in the cell. Most cells possess the ability to switch between these two pathways, thereby adapting to changes in their environment.

Glucose in the cell is converted to pyruvate (referred to as glycolysis), and then converted to lactate in the cytoplasm, or CO2 and water in the mitochondria.

The conversion of glucose to pyruvate, and subsequently lactate, results in a net production and extrusion of protons into the extracellular medium.... The extrusion of protons results in the acidification of the medium surrounding the cell. The XF instrument directly measures the acidification rate, and reports this as ECAR

• Glycolysis: The process of converting glucose to pyruvate. The XF Glycolysis Stress Test presents the measure of glycolysis as the ECAR rate reached by a given cell after the addition of saturating amounts of glucose.

• Glycolytic capacity: This measurement is the maximum ECAR rate reached by a cell following the addition of oligomycin, effectively shutting down oxidative phosphorylation and driving the cell to use glycolysis to its maximum capacity.

• Glycolytic reserve: This measure indicates the capability of a cell to respond to an energetic demand as well as how close the glycolytic function is to the cell’s theoretical maximum.

• Nonglycolytic acidification: This measures other sources of extracellular acidification that are not attributed to glycolysis.
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So the reduced glycolytic reserve seen in these preliminary results suggests the ME patients' NK cells have a significantly reduced ability to respond to energetic demand.

Eta: I wonder if some of this might possibly intersect with Booth and Myhill's work on mitochondrial ATP production.
 
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So it looks like it's basically saying glycolysis, which is the low energy producing anaerobic first stage of energy production in the cells is going too slowly, so not producing enough substrate for the higher energy producing aerobic stage that happens next in the mitochondria. So the cells can't produce energy fast enough to meet demand. I think...
 
Trish said:
So it looks like it's basically saying glycolysis, which is the low energy producing anaerobic first stage of energy production in the cells is going too slowly, so not producing enough substrate for the higher energy producing aerobic stage that happens next in the mitochondria. So the cells can't produce energy fast enough to meet demand. I think...
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Uncoupling of supply and demand ?
There could be a number of potential mechanisms?
 
Simone said:
With the millions in funding this group has, it’s so frustrating that they often publish such tiny studies.
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Pilot studies are meant to be small. It is designed so they can explore hypothesis without too much financial and human ressource investment. Since they seem to have found something, it will give them power to apply for some grants to either expand on a bigger cohort, or try the same experiment on other cell type, or compare to other similar diseases.

I am thankful that this team is exploring avenues that haven’t been researched through scientific experiments, through hypotheses and they are also teaching health science students about the disease. We need and deserve so much more research.
 
Milo said:
Pilot studies are meant to be small. It is designed so they can explore hypothesis without too much financial and human ressource investment. Since they seem to have found something, it will give them power to apply for some grants to either expand on a bigger cohort, or try the same experiment on other cell type, or compare to other similar diseases.
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The problem is success on a pilot study isn't actually statistically valid to justify further research, it simply shows that such an experimental design is possible.
 
Indigophoton said:
According to the instruction manual for the Seahorse test used in this paper,


So the reduced glycolytic reserve seen in these preliminary results suggests the ME patients' NK cells have a significantly reduced ability to respond to energetic demand.

Eta: I wonder if some of this might possibly intersect with Booth and Myhill's work on mitochondrial ATP production.
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I think there is a definite intersection. They too found an uncoupling of supply and demand; more interestingly, for some they found that ATP was being flung out of the mitochondria faster than it could be replenished ( ostensibly due to substrate issues)- this could be ATP as puringenic signalling, instigating AMPK activation and downregulation ...
 
Jumbled thoughts:

Prof Don Staines presented data, at the Invest in ME Research International ME Conference 2018, indicating problems with calcium regulation. I assumed that they would/had published that data. I assume that the energy problems demonstrated in this paper are proposed to be a consequence of the calcium regulation problem.

This glycolysis study looks similar to other studies i.e. those referred to above (C Tomas etc); therefore, I'm not clear what's new. However, a study in effect confirming problems is still useful i.e. it increases confidence in the finding of energy/metabolic problems.

Ron Davis presented data at the Invest in ME Conference showing that you could potentially use the Seahorse analyser to diagnose ME/CFS.

Interestingly Ron Davis (in one of his talks - December 2017 OMF site?) said that ME/CFS did look like diabetes. Staines, at the Invest in ME Conference, said that TRIP receptors had a role in insulin regulation (diabetes). So possibly the calcium regulation (TRIP) theory Staines etc. are working on would explain the observed metabolic effects observed in ME/CFS.

I would like to see the calcium regulation data resented at the Invest in ME Conference published.
 
Simone said:
With the millions in funding this group has, it’s so frustrating that they often publish such tiny studies.
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I think part of the problem is that this is a university where multiple PhD students need a novel hypothesis to research and report on and that is probably the main focus. I suspect finding answers to ME/CFS is probably a secondary goal.
 
Trish said:
Perhaps the point is that this research group seem to have done lots of small studies and got them published, and got lots of funding, but don't seem to have done any large studies.
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Someone has to write the next grant -- and someone else has to approve it. Yes to the tiny PhD students trying to publish a thing -- and then moving on to the next thing, which may have nothing to do with ME.

Snow Leopard said:
The problem is success on a pilot study isn't actually statistically valid to justify further research, it simply shows that such an experimental design is possible.
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At least theoretically, this is all you're supposed to need to apply for an R-level grant: show the experimental design works out, and show some promising results on a limited sample size.

FMMM1 said:
Interestingly Ron Davis (in one of his talks - December 2017 OMF site?) said that ME/CFS did look like diabetes.
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I don't think Ron would mind if I pointed out that I mentioned if we called it "Type III Diabetes" that's when we'd get some real funding. ;) I was only half-joking.
 
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