Dolphin
Senior Member (Voting Rights)
Via Dr. Marc-Alexander Fluks
Source: University of Oslo Date: May 2023 URL: https://www.duo.uio.no/bitstream/handle/10852/103812/1/Masterthesis_ElenaYran_May15th2023.pdf
Investigating the potential role of circulatory extracellular vesicles in myalgic encephalomyelitis/chronic fatigue syndrome
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Elena Stovring Yran - Oslo University Hospital, Department of Medical Genetics, University of Oslo, Oslo, Norway. Email: eleyra@ous-hf.no
Abstract
ME/CFS is a debilitating disease thought to affect millions of individuals. Still, the etiology of ME/CFS is unknown, and there are no standard treatments or established biomarkers. Thecurrent symptom-based diagnosis is extensive, and the use of different diagnostic criteria contributes to heterogeneity among patients and may problematize the comparison of findings. Thus, the discovery of a biomarker for ME/CFS is urgent and would benefit both patients and the ME/CFS research field. Extracellular vesicles (EVs) are membrane limited vesicles secreted by all cells to the extracellular environment and can be collected through biofluids. EVs serve many functions, including transferring functional proteins, lipids, and nucleic acids between cells, thus mediating cell-to-cell communication. EV secretion and cargo may reflect disease state and EVs thus pose great potential as source of minimally invasive biomarkers.
The primary aim of this project was to study EVs in plasma from ME/CFS patients and assess the potential of EVs as source of biomarkers for the disease.
Using size exclusion chromatography, EVs were enriched from plasma from ME/CFS patients (n = 20) and healthy controls (n=20). Success of EV isolation was determined in representative patient- and control EV pools (n=5) using western blotting and transmission electron microscopy. Western blot experiments for detection of EV markers CD9, CD63 and TSG101, and albumin, were optimized and confirmed enrichment of EVs and presence of non-EV eluates in the isolated samples. EV enrichment was further validated through observation of intact EVs on transmission electron micrographs, however few CD63-positive EVs were observed. Through analysis of nanoparticle tracking analysis data, the isolated EV population primarily consisted of small EVs (<200 nm). Within this EV population, mean and mode EV size was similar between cohorts, but the EV concentration was significantly elevated in samples from patients compared to controls (p=0.006). However, statistical testsmay have been influenced by high variation within the ME/CFS cohort. Early-stage analysis of tandem mass spectrometry data identified 663 EV associated proteins. The majority of detected proteins overlapped with registered EV proteins, but only few differences could be observed between patient- and control derived samples. However, differential expression was not analyzed.
A biomarker for ME/CFS could not be suggested at this stage of the study, however increased EV concentration suggests abnormality in EV secretion in patients which strengthens their potential as source of biomarkers and further motivates EV research in ME/CFS and related diseases.
Source: University of Oslo Date: May 2023 URL: https://www.duo.uio.no/bitstream/handle/10852/103812/1/Masterthesis_ElenaYran_May15th2023.pdf
Investigating the potential role of circulatory extracellular vesicles in myalgic encephalomyelitis/chronic fatigue syndrome
-------------------------------------------------------------------------
Elena Stovring Yran - Oslo University Hospital, Department of Medical Genetics, University of Oslo, Oslo, Norway. Email: eleyra@ous-hf.no
Abstract
ME/CFS is a debilitating disease thought to affect millions of individuals. Still, the etiology of ME/CFS is unknown, and there are no standard treatments or established biomarkers. Thecurrent symptom-based diagnosis is extensive, and the use of different diagnostic criteria contributes to heterogeneity among patients and may problematize the comparison of findings. Thus, the discovery of a biomarker for ME/CFS is urgent and would benefit both patients and the ME/CFS research field. Extracellular vesicles (EVs) are membrane limited vesicles secreted by all cells to the extracellular environment and can be collected through biofluids. EVs serve many functions, including transferring functional proteins, lipids, and nucleic acids between cells, thus mediating cell-to-cell communication. EV secretion and cargo may reflect disease state and EVs thus pose great potential as source of minimally invasive biomarkers.
The primary aim of this project was to study EVs in plasma from ME/CFS patients and assess the potential of EVs as source of biomarkers for the disease.
Using size exclusion chromatography, EVs were enriched from plasma from ME/CFS patients (n = 20) and healthy controls (n=20). Success of EV isolation was determined in representative patient- and control EV pools (n=5) using western blotting and transmission electron microscopy. Western blot experiments for detection of EV markers CD9, CD63 and TSG101, and albumin, were optimized and confirmed enrichment of EVs and presence of non-EV eluates in the isolated samples. EV enrichment was further validated through observation of intact EVs on transmission electron micrographs, however few CD63-positive EVs were observed. Through analysis of nanoparticle tracking analysis data, the isolated EV population primarily consisted of small EVs (<200 nm). Within this EV population, mean and mode EV size was similar between cohorts, but the EV concentration was significantly elevated in samples from patients compared to controls (p=0.006). However, statistical testsmay have been influenced by high variation within the ME/CFS cohort. Early-stage analysis of tandem mass spectrometry data identified 663 EV associated proteins. The majority of detected proteins overlapped with registered EV proteins, but only few differences could be observed between patient- and control derived samples. However, differential expression was not analyzed.
A biomarker for ME/CFS could not be suggested at this stage of the study, however increased EV concentration suggests abnormality in EV secretion in patients which strengthens their potential as source of biomarkers and further motivates EV research in ME/CFS and related diseases.
