Cytokine profiling of extracellular vesicles isolated from plasma in ME/CFS: a pilot study, 2020, Giloteaux, Levine, Hanson et al

Cytokine profiling of extracellular vesicles isolated from plasma in myalgic encephalomyelitis/chronic fatigue syndrome: a pilot study
Ludovic Giloteaux, Adam O'Neal, Jesús Castro-Marrero, Susan M. Levine & Maureen R. Hanson

Background
Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) is a debilitating disease of unknown etiology lasting for a minimum of 6 months but usually for many years, with features including fatigue, cognitive impairment, myalgias, post-exertional malaise, and immune system dysfunction. Dysregulation of cytokine signaling could give rise to many of these symptoms. Cytokines are present in both plasma and extracellular vesicles, but little investigation of EVs in ME/CFS has been reported. Therefore, we aimed to characterize the content of extracellular vesicles (EVs) isolated from plasma (including circulating cytokine/chemokine profiling) from individuals with ME/CFS and healthy controls.


Methods
We included 35 ME/CFS patients and 35 controls matched for age, sex and BMI. EVs were enriched from plasma by using a polymer-based precipitation method and characterized by Nanoparticle Tracking Analysis (NTA), Transmission Electron Microscopy (TEM) and immunoblotting. A 45-plex immunoassay was used to determine cytokine levels in both plasma and isolated EVs from a subset of 19 patients and controls. Linear regression, principal component analysis and inter-cytokine correlations were analyzed.


Results
ME/CFS individuals had significantly higher levels of EVs that ranged from 30 to 130 nm in size as compared to controls, but the mean size for total extracellular vesicles did not differ between groups. The enrichment of typical EV markers CD63, CD81, TSG101 and HSP70 was confirmed by Western blot analysis and the morphology assessed by TEM showed a homogeneous population of vesicles in both groups. Comparison of cytokine concentrations in plasma and isolated EVs of cases and controls yielded no significant differences. Cytokine-cytokine correlations in plasma revealed a significant higher number of interactions in ME/CFS cases along with 13 inverse correlations that were mainly driven by the Interferon gamma-induced protein 10 (IP-10), whereas in the plasma of controls, no inverse relationships were found across any of the cytokines. Network analysis in EVs from controls showed 2.5 times more significant inter-cytokine interactions than in the ME/CFS group, and both groups presented a unique negative association.

Conclusions

Elevated levels of 30-130 nm EVs were found in plasma from ME/CFS patients and inter-cytokine correlations revealed unusual regulatory relationships among cytokines in the ME/CFS group that were different from the control group in both plasma and EVs. These disturbances in cytokine networks are further evidence of immune dysregulation in ME/CFS.

 

I had a look at the website for the journal the above paper came from and found this page which might be of interest to people.

https://translational-medicine.biomedcentral.com/articles/sections/illnesses-of-unknown-etiology

 

https://twitter.com/user/status/1315704656975269889

 

So now the next question would be what characterizes EVs of 30-130 nm.

There will definitely be no shortage of preliminary research to build upon if and when the money finally happens.

Research that open up new questions is always the most interesting. It just sucks that the normal way of doing things doesn't apply to us, but definitely once things get serious there will be a very fertile field to till and plant on.

 

What I like most about this #mecfs paper is researchers doing a pilot study in order to improve the quality of their final studies. Will be excited to see how this work pans out in the pre/post exercise studies to come.

Note that the cytokine study only had a sample size of 19 patients/controls.

Pity about the lack of difference in cytokine concentrations.

I do wonder if the cytokines – cytokine correlations will replicate in larger samples, but let’s see. And I am hopeful that testing before and after exercise will provide insight.

 

But, as Simon points out above, cytokines of only 19 and 19 were analysed.

 

I have the impression the authors are hoping that a more focused study could reveal greater differences and turn some of the findings that were not statistically significant after correcting for multiple comparisons into significant ones.

This sounds like it could be the most abnormal finding in the study.

What is an inter-cytokine correlation exactly?

 

I wonder what factors determine the size distribution of EVs, and if any other conditions show a distribution like the one they found.

I did find this overview of EVs, but it seems a bit "above my pay grade" on first glance.

Overview of Extracellular Vesicles, Their Origin, Composition, Purpose, and Methods for Exosome Isolation and Analysis