Cytokine profiling of extracellular vesicles isolated from plasma in myalgic encephalomyelitis/ chronic fatigue syndrome: a pilot study, by Ludovic Giloteaux, Adam O’Neal, Jesús Castro-Marrero, Susan M Levine, Maureen R Hanson in J Transl Med. 2020 Oct 12;18(1):387 [doi: 10.1186/s12967-020-02560-0]
Research abstract:
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.
Heart rate and blood pressure did not change significantly between 
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he Shortened Fatigue Questionnaire (SFQ) [10] consists of four items (‘I feel tired’, ‘I tire easily‘, ‘I feel fit’ and ‘I feel physically exhausted’; see appendix B). Each item is scored on a 7-point Likert Scale, ranging from 1 ‘yes, that is true’ to 7 ‘no, that is not true’. Scores of items 1, 2 and 4 are reversed and then all item scores are added up which results in a total score varying from 4 to 28. Higher scores reflect a higher level of fatigue.
A total of 60 proteins in the ME/CFS patients were differentially expressed (P < 0.01, Log10 (Fold Change) > 0.2 and < −0.2). Comparison of the PCA selected subgroup of ME/CFS patients (9/11) with controls increased the number of proteins differentially expressed to 99. Of particular relevance to the core symptoms of fatigue and post-exertional malaise experienced in ME/CFS, a proportion of the identified proteins in the ME/CFS groups were involved in mitochondrial function, oxidative phosphorylation, electron transport chain complexes, and redox regulation. A significant number were also involved in previously implicated disturbances in ME/CFS, such as the immune inflammatory response, DNA methylation, apoptosis and proteasome activation.
In the present study, we identified fifty online forum (Reddit) posts, discussing the personal lived experience of Chronic Fatigue Syndrome during lockdown and the global pandemic. These posts were subject to inductive thematic analysis.
Chronic Fatigue Syndrome/Myalgic Encephalomyelits (CFS/ME) is a chronic disease with complex pathophysiology and unknown etiology. It occurs both in children and adolescents, as well as in adults, with equal frequency. The clinical course is characterized by progressive fatigue, a significant reduction in the body’s efficiency, lack of relief despite rest, and numerous accompanying symptoms. Pathognomonic symptom for PE is the increase in fatigue after physical or mental exertion and the persistence of these symptoms for several hours or days.
Adolescents with Chronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME) appear to be more likely to experience anxiety and/or depression using 
Myalgic encephalomyelitis/chronic fatigue syndrome is characterized by persistent and disabling fatigue, exercise intolerance, cognitive difficulty, and musculoskeletal/joint pain. Post–exertional malaise is a worsening of these symptoms after a physical or mental exertion and is considered a central feature of the illness. Scant observations in the available literature provide qualitative assessments of post–exertional malaise in patients with myalgic encephalomyelitis/ chronic fatigue syndrome. To enhance our understanding, a series of outpatient focus groups were convened.
