The itaconate shunt hypothesis

[Simon M]

Re the discussion of glutamine metabolism, glucose, fats (etc) in prior work: indeed, nothing is proven yet. Good news is that people are doing follow-up studies to validate these theories (including with different methods or with limitations of prior studies in mind)..., one is that I am also working with Chris to look at these metabolic questions in cells.

Very good to hear there are more studies in the pipeline.

Am I right in thinking that other metabolomics studies (Lipkin, Hansen, maybe Unutmaz, Naviaux) didn't find increased use of glutamine/ate?
Though I note this:

rom memory proteomics research ( Hanson,? ) showed a difference between metabolism of male and females re TCA deficiencies.

Females were more likely to sub protein ( amino acids) for glucose as fuel. Males more likely to sub fats.

 

[Sasha]

Is this the sort of hypothesis that the DecodeME GWAS study could rule in or out?

 

[Jonathan Edwards]

I don't get the impression that the hypothesis is well enough formulated as yet to be testable by anything much. If it was I suspect it would be most easily testable by simple MRI spectroscopy of tissues before and after exercise. If the problem is really a shortage of energy supply from a diverted metabolic pathway then different levels of metabolites would show up in a very predictable way.

DecodeME would only be relevant if the hypothesis stated what possible genetic predisposing factors might occur such that an immune response might lead to an unusual diversion.

 

[Kitty]

I suspect it would be most easily testable by simple MRI spectroscopy of tissues before and after exercise. If the problem is really a shortage of energy supply from a diverted metabolic pathway then different levels of metabolites would show up in a very predictable way.

Would this be likely to hold, no matter what type of metabolic diversion were in place?

(I suspect life's not that simple, but it sounds so much like what we need that I thought I'd wonder about it aloud anyway...)

 

[Jonathan Edwards]

Would this be likely to hold, no matter what type of metabolic diversion were in place?

I cannot answer that for sure. However, in the 1980s I worked in the next lab to people who did MR spectroscopy and they showed plots with peaks for pretty much any metabolite you might be interested in as far as I remember.

If cells are really running out of ATP or pyruvate or whatever it ought to be possible to show some shifts in at least some of these peaks. And of course if you have different proportional shifts in a range of metabolites you should be able to work out roughly where the blockage is.

I suspect the reason why it is not used more in ME is that it has been tried and nothing found. Some recent stuff looked at lactate in brain ventricles, so you can do lactate. Even that did not seem to be followed up.

 

[Midnattsol]

I cannot answer that for sure. However, in the 1980s I worked in the next lab to people who did MR spectroscopy and they showed plots with peaks for pretty much any metabolite you might be interested in as far as I remember.

If cells are really running out of ATP or pyruvate or whatever it ought to be possible to show some shifts in at least some of these peaks. And of course if you have different proportional shifts in a range of metabolites you should be able to work out roughly where the blockage is.

I suspect the reason why it is not used more in ME is that it has been tried and nothing found. Some recent stuff looked at lactate in brain ventricles, so you can do lactate. Even that did not seem to be followed up.

Finding a peak doesn't mean one is able to identify the metabolite that belongs to the peak (talking metabolomics).

 

[Hoopoe]

I've been going swimming almost every day lately and there is a trend of finding it increasingly difficult over time. I keep having to reduce the time and intensity and have to spend more time horizontally during the rest of the day. This is similar to other periods in my life so it's probably not due to confounders like say the heat wave. There has to be some way to measure what is changing in my body that is causing this. My heart rate doesn't seem to have increased, if anything it's slightly lower. This seems consistent with some degree of cardiovascular training occurring but for whatever reason I'm getting weaker.

PS: regular swimming was an attempt to see if I could recreate my substantial improvement that occurred last summer but this year it's just not happening it seems.

 

[roller*]

I wonder what the theorized impaired use of glucose and fat at the mitochondrial level, compensated for by increased use of amino acids, would imply on the macro level.

Not sure, but I guess fatty liver could be one effect.

https://www.healthrising.org/blog/2...gregor-metabolism-chronic-fatigue-glycolysis/[/URL]
His (Neil McGregor) results suggest that difficulty turning glucose and fats into energy is causing people with this disease to turn to their last and worst option – breaking down their own muscles to provide the substrates the energy production process needs.

As their tissues are broken down, the metabolites needed to replenish them are being flushed into the urine, resulting in a chronically depleted system. McGregor believes this chronic depletion of essential metabolites plays a crucial role in ME/CFS and results in numerous issues, including post-exertional malaise, gut malabsorption, connective tissue problems, inflammation and more.

Because creatinine is an important factor in the production of energy in the muscle, brain and cardiac tissue, the low creatinine levels translated into low energy levels.

would this be a typical "wasting disease"?

my blood-creatinine values (these values paralyzed ppl usually have):
2009 = 0.68 mg/dL (Range 0.60 - 1.10)
2022 = 0.64 mg/dL (Range 0.67-1.17) DOWN!!

I have been on a (at least once) weekly extraordinary strengeous exercise since october 2021.
The very minimum is 9 hours in a row with heavy equipment.
i think i got (slooooowlly...) some "muscles" which are still invisible.

could this "wasting theory" explain, that the additional exercise-creatinine is used for more energy-production?
i do have strikingly more energy.

but it took months und it would be of course still from the "wrong cycle".

and the liver values are still as bad as they were.
there is potassium above the range (Hyperkalemia), which indicates cardiac issues, ie. not sufficient muscle to keep heart rate up

 

[Kitty]

I cannot answer that for sure. However, in the 1980s I worked in the next lab to people who did MR spectroscopy and they showed plots with peaks for pretty much any metabolite you might be interested in as far as I remember.

I suspect the reason why it is not used more in ME is that it has been tried and nothing found. Some recent stuff looked at lactate in brain ventricles, so you can do lactate. Even that did not seem to be followed up.

Thank you, that's interesting. I suppose it partly comes down to money—it doesn't sound like a particularly cheap technique, and it might be more difficult than it looks at first sight to get the timing right.

Finding a peak doesn't mean one is able to identify the metabolite that belongs to the peak (talking metabolomics).

No, but perhaps if you could show something odd was going on, it might provide enough of an evidence base to get more research funded. And you'd be able to discuss it with people who're finding oddnesses in other conditions, which might help you direct your investigation. It's the sort of thing that needs to happen to move us forward.

 

[Jonathan Edwards]

Finding a peak doesn't mean one is able to identify the metabolite that belongs to the peak (talking metabolomics).

Well, when I was shown these peaks each one was clearly identified as lactate or glutamate or pyruvate or whatever. There was lots of them. As far as I know each molecule has a particular signature that is known beforehand.

 

[Jonathan Edwards]

Thank you, that's interesting. I suppose it partly comes down to money—it doesn't sound like a particularly cheap technique, and it might be more difficult than it looks at first sight to get the timing right.

I doubt it has much to do with money. These are machines that have been available since maybe the 1960s, at least since 1980. They do not require the complicated set up of MR imaging although they need a powerful magnet. My guess would be that once you have a machine studies cost rather little.It may be that to get results relating to effects of exercise you need to make use of glucose labelled with a carbon isotope to follow the metabolism through. I forget. But I don't think cost is an issue.

I just think that it must be that when people with ME were studied nothing showed up - which makes it hard to sustain any theory that puts symptoms down to failure of metabolic pathways I think.

 

[Hoopoe]

A study by Morten and his Polish collaborators found a decrease in central energy metabolism in patients with ME/CFS when comparing pre and post exercise program timepoints. There was no control group so this was not published.

This study of exercise for long covid also seems to be reporting something similar: "a decrease of oxidative metabolism index of 6.89 standard units."

https://www.s4me.info/threads/can-a...-long-covid-outcomes-2022-lobanov-et-al.28618

 

[Midnattsol]

Well, when I was shown these peaks each one was clearly identified as lactate or glutamate or pyruvate or whatever. There was lots of them. As far as I know each molecule has a particular signature that is known beforehand.

While many molecules have easily identifiable signatures/peaks this is not true for all, and even for those that have these easily identifiable peaks noise in the data can make it difficult. Look at untargeted metabolomics studies and there will e a number of unknown metabolites. For targetted metabolomics an issue is that different labs include different metabolites so comparing across studies is not possible since not all studies will include the same metabolites. While I'd be surprised to not see glutamate, I would not be surprised if lactate or pyruvate wasn't looked at, but there might be differences between fields I guess.

 

[Hutan]

Yes, we've talked about this problem with mass spectroscopy elsewhere on the forum. Here's something from a 2010 paper. I'm sure that there has been progress since then, but there are still problems:

The identification of compounds from mass spectrometry (MS) data is still seen as a major bottleneck in the interpretation of MS data. This is particularly the case for the identification of small compounds such as metabolites, where until recently little progress has been made.

The identification without reference spectra is even more challenging, because it requires approaches to interpret tandem mass spectra with regard to the molecular structure.

Edit to add - here's a 2019 paper that is a relatively up to date survey of mass spectroscopy for metabolite identification:
Mass spectrometry-based metabolomics in health and medical science: a systematic review

The acquisition speed, accuracy, sensitivity and coverage of MS still require further improvement. New processes for the accurate identification of metabolites, rapid acquisition methods, and novel detection methods with high derivative efficiency should be developed to achieve large-scale coverage of metabolites and to facilitate data processing. Although metabolomics can identify a great quantity of metabolites, researchers usually do not identify metabolites that are not found in databases or websites. The exploration of unknown metabolites is a real challenge and research direction in metabolomics. In addition, some standards are difficult to obtain, and the existence of cis- and trans-isomers can also lead to inaccurate identification of metabolites. Thus, it is necessary to speed up the construction of standard product databases and information sharing.

Current MS-based metabolomic analysis mostly adopts relative quantification or qualitative analysis with fewer examples of quantitative analysis. The accurate concentration of metabolites in vivo has an important influence on the interpretation of metabolite function. How to simultaneously quantify multiple metabolites is the short board of current metabolomics? The ideal state is to use stable isotope-assisted metabolomics to track reaction substrates and determine the role of metabolites in metabolic pathways.94 Furthermore, the diversity of many enzyme substrates, in addition to the complexity of metabolic networks, make it difficult to influence the content of a single metabolite without affecting others.

 

[Midnattsol]

No, but perhaps if you could show something odd was going on, it might provide enough of an evidence base to get more research funded. And you'd be able to discuss it with people who're finding oddnesses in other conditions, which might help you direct your investigation. It's the sort of thing that needs to happen to move us forward.

You would not necessarily get something odd, or anything that can be discussed besides "there are many metabolites we can't identify or quantify". When data is shared it might be possible for others to help identify/make sense of the peaks, but it is simply not always possible (and data sharing is not the standard it should be! A lot of metabolomics studies still only report whatever the researchers found interesting, like ten or twenty out of a set of several 100s. Had the data on the other metabolites also been available in an easy way it would be much easier to collaborate.

 

[Jonathan Edwards]

I was talking about magnetic resonance spectroscopy rather than mass spectroscopy.

 

[Jonathan Edwards]

While many molecules have easily identifiable signatures/peaks this is not true for all,

Sure, but if the claim is that symptoms are due to failure of major respiratory pathways preventing energy usage then surely some easily identified molecules would show shifts. You might not be able to track exactly why but if there isn't even a shift in the some common pathway elements it is hard to sustain a theory based on energy blockade. I am not familiar with the detail but I am sceptical of fancy 'trap' theories that do not seem to take into account the need to show something pretty simple at the end of the day - not enough of some common energy pathway product.

 

[Hutan]

From that 2019 paper I mentioned above re NMR:

The two main analytical techniques for metabolic analysis are 1H nuclear magnetic resonance (NMR) and MS. Both of these techniques can analyze a large number of small molecules coexisting in complex samples, including metabolite identification and quantification. NMR spectroscopy can directly identify and quantitatively analyze numerous analytes. Although the reproducibility of NMR is better than that of MS-based techniques, the sensitivity of NMR is lower and a larger sample size is required. In the field of medical science, clinical samples are particularly valuable; thus MS-based metabolomic technology is widely used in clinical research.5

NMR does seem to be much better for identification of novel compounds. It sounds, though, that you need a lot more of a metabolite (10 to 100 x what MS will work with) for NMR to find it. For anyone just learning, this 2019 paper seems good;
NMR Spectroscopy for Metabolomics Research

I am sceptical of fancy 'trap' theories that do not seem to take into account the need to show something pretty simple at the end of the day - not enough of some common energy pathway product.

You may well be right. It's hard to remember what exactly has been been done so far, and the potential problems with each study. I just know that we've seen a lot of research that has been, in many ways, poor or technically limited. So I wouldn't necessarily assume there's nothing left to find.

Yesterday I had to go for a long walk, and the impact of that on my leg muscles, the inability to function smoothly, and huge loss of power, and the pain in the night makes me think there must surely be some metabolite to find that would explain why that happens.

 

[Jonathan Edwards]

NMR does seem to be much better for identification of novel compounds. It sounds, though, that you need a lot more of a metabolite (10 to 100 x what MS will work with) for NMR to find it. For anyone just learning, this 2019 paper seems good;

Yes, but if you do NMR on a whole leg you have plenty of metabolite! You can do NMR on arms and legs without disturbing the patient at all - other than asking them to put a leg in a ring magnet.

 

[SNT Gatchaman]

For MR spectroscopy in clinical imaging, we typically target a voxel. For metabolic diseases this is usually standardised to the left lentiform nucleus, but looks like right thalamus in the example below. Lactate, NAA, creatine and choline are the typical metabolites of interest in the brain. In the normal example below there's no lactate peak (at 1.3 ppm).

Glutamine/glutamate and GABA are both 2.2-2.4 ppm so may not be discriminated.

Usual peaks

• lipids: 1.3 ppm
• lactate: 1.33 ppm
• alanine: 1.48 ppm
• N-acetylaspartate (NAA): 2.0 ppm
• glutamine/glutamate: 2.2-2.4 ppm
• GABA: 2.2-2.4 ppm
• 2-hydroxyglutarate: 2.25 ppm 6
• citrate: resonates 2.6 ppm
• creatine: 3.0 ppm
• choline: 3.2 ppm
• myo-inositol: 3.5 ppm
• water: 4.7 ppm

Less common peaks

• propylene glycol: 1.14 ppm
• ethanol: 1.16 ppm
• acetate: 1.9 ppm
• acetone: 2.22 ppm
• acetoacetate: 2.29 ppm
• succinate: 2.4 ppm
• methylsulfonylmethane: 3.15 ppm
• scyllo-inositol: 3.36 ppm
• taurine: 3.4 ppm
• glucose: 3.43 ppm and 3.8 ppm
• mannitol: 3.78 ppm
• creatine (second peak): 3.95 ppm 10
• lactate quartet: 4.11 ppm