Impact of Long-Term Cryopreservation on Blood Immune Cell Markers in Myalgic Encephalomyelitis/Chronic Fatigue Syndrome: Implications for Biomarker Discovery

• Published in: Frontiers in immunology Vol. 11; p. 582330
• Main Authors: Gómez-Mora, Elisabet, Carrillo, Jorge, Urrea, Víctor, Rigau, Josepa, Alegre, José, Cabrera, Cecilia, Oltra, Elisa, Castro-Marrero, Jesús, Blanco, Julià
• Format: Journal Article
• Language: English
• Published: Switzerland Frontiers Media S.A 17.11.2020
• Subjects: Adult; Biomarkers - metabolism; Blood Cells - metabolism; Case-Control Studies; Cells, Cultured; chronic fatigue syndrome; Cohort Studies; Cryopreservation; Fatigue Syndrome, Chronic - diagnosis; Female; freeze-thaw process; Humans; immune biomarkers; Immunology; Immunophenotyping; Killer Cells, Natural - metabolism; Lymphocyte Subsets - metabolism; Male; Middle Aged; myalgic encephalomyelitis; Prospective Studies; T-Lymphocytes - metabolism; chronic fatigue syndrome; myalgic encephalomyelitis; immunophenotyping; cryopreservation; freeze-thaw process; immune biomarkers
• ISSN: 1664-3224; 1664-3224
• DOI: 10.3389/fimmu.2020.582330

Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) is a complex neuroimmune disorder characterized by numerous symptoms of unknown etiology. The ME/CFS immune markers reported so far have failed to generate a clinical consensus, perhaps partly due to the limitations of biospecimen biobanking. To address this issue, we performed a comparative analysis of the impact of long-term biobanking on previously identified immune markers and also explored additional potential immune markers linked to infection in ME/CFS. A correlation analysis of marker cryostability across immune cell subsets based on flow cytometry immunophenotyping of fresh blood and frozen PBMC samples collected from individuals with ME/CFS (n = 18) and matched healthy controls (n = 18) was performed. The functionality of biobanked samples was assessed on the basis of cytokine production assay after stimulation of frozen PBMCs. T cell markers defining Treg subsets and the expression of surface glycoprotein CD56 in T cells and the frequency of the effector CD8 T cells, together with CD57 expression in NK cells, appeared unaltered by biobanking. By contrast, NK cell markers CD25 and CD69 were notably increased, and NKp46 expression markedly reduced, by long-term cryopreservation and thawing. Further exploration of Treg and NK cell subsets failed to identify significant differences between ME/CFS patients and healthy controls in terms of biobanked PBMCs. Our findings show that some of the previously identified immune markers in T and NK cell subsets become unstable after cell biobanking, thus limiting their use in further immunophenotyping studies for ME/CFS. These data are potentially relevant for future multisite intervention studies and cooperative projects for biomarker discovery using ME/CFS biobanked samples. Further studies are needed to develop novel tools for the assessment of biomarker stability in cryopreserved immune cells from people with ME/CFS.