In vitro B cell experiments explore the role of CD24, CD38, and energy metabolism in ME/CFS.

Introduction: Disturbances of energy metabolism contribute to the clinical manifestations of myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS). Previously, we found that B cells from ME/CFS patients have an increased expression of CD24, a modulator of many cellular functions including those of cell stress. The relative ability of B cells from ME/CFS patients and healthy controls (HC) to respond to rapid changes in energy demand was compared. Methods: CD24, the ectonucleotidases CD39 and CD73, the NAD-degrading enzyme CD38, and mitochondrial mass (MM) were measured following cross-linking of the B cell receptor and costimulation with either T-cell-dependent or Toll-like-receptor-9-dependent agonists. The levels of metabolites consumed/produced were measured using 1H-NMR spectroscopy and analyzed in relation to cell growth and immunophenotype. Results: Proliferating B cells from patients with ME/CFS showed a lower mitochondrial mass and a significantly increased usage of essential amino acids compared with those from HC, with a significantly delayed loss of CD24 and an increased expression of CD38 following stimulation. Discussion: The immunophenotype results suggested the triggering of a stress response in ME/CFS B cells associated with the increased usage of additional substrates to maintain necessary ATP levels. Disturbances in energy metabolism in ME/CFS B cells were thus confirmed in a dynamic in vitro model, providing the basis for further mechanistic investigations.

CD24 Expression and B Cell Maturation Shows a Novel Link With Energy Metabolism: Potential Implications for Patients With Myalgic Encephalomyelitis/Chronic Fatigue Syndrome.

CD24 expression on pro-B cells plays a role in B cell selection and development in the bone marrow. We previously detected higher CD24 expression and frequency within IgD+ naïve and memory B cells in patients with Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) compared with age-matched healthy controls (HC). Here, we investigated the relationship between CD24 expression and B cell maturation. In vitro stimulation of isolated B cells in response to conventional agonists were used to follow the dynamics of CD24 positivity during proliferation and differentiation (or maturation). The relationship between CD24 expression to cycles of proliferation and metabolism in purified B cells from HC was also investigated using phospho-flow (phosphorylation of AMPK-pAMPK), 1proton nuclear magnetic resonance and Mitotracker Far-red (Mitochondrial mass-MM). In vitro, in the absence of stimulation, there was an increased percentage of CD24+ viable B cells in ME/CFS patients compared to HC (p < 0.05) following 5 days culture. Following stimulation with B cell agonists, percentage of CD24+B cells in both naïve and memory B cell populations decreased. P < 0.01). There was a negative relationship between percentage of CD24+B cells with MM (R2 = 0.76; p < 0.01), which was subsequently lost over sequential cycles of proliferation. There was a significant correlation between CD24 expression on B cells and the usage of glucose and secretion of lactate in vitro. Short term ligation of the B cell receptor with anti-IgM antibody significantly reduced the viability of CD24+ memory B cells compared to those cross-linked by anti-IgD or anti-IgG antibody. A clear difference was found between naïve and memory B cells with respect to CD24 expression and pAMPK, most notably a strong positive association in IgD+IgM+ memory B cells. In vitro findings confirmed dysregulation of CD24-expressing B cells from ME/CFS patients previously suggested by immunophenotype studies of B cells from peripheral blood. CD24-negative B cells underwent productive proliferation whereas CD24+ B cells were either unresponsive or susceptible to cell death upon BCR-engagement alone. We suggest that CD24 expression may reflect variations in energy metabolism on different B cell subsets.