EBV/HHV-6A dUTPases contribute to myalgic encephalomyelitis/chronic fatigue syndrome pathophysiology by enhancing TFH cell differentiation and extrafollicular activities
Brandon S. Cox, … , William P. Lafuse, Maria Eugenia Ariza
Brandon S. Cox, … , William P. Lafuse, Maria Eugenia Ariza
Published April 28, 2022
Citation Information: JCI Insight. 2022;7(11):e158193. https://doi.org/10.1172/jci.insight.158193.
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Research Article Infectious disease

EBV/HHV-6A dUTPases contribute to myalgic encephalomyelitis/chronic fatigue syndrome pathophysiology by enhancing TFH cell differentiation and extrafollicular activities

Abstract

Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a chronic, debilitating, multisystem illness of unknown etiology for which no cure and no diagnostic tests are available. Despite increasing evidence implicating EBV and human herpesvirus 6A (HHV-6A) as potential causative infectious agents in a subset of patients with ME/CFS, few mechanistic studies address a causal relationship. In this study we examined a large ME/CFS cohort and controls and demonstrated a significant increase in activin A and IL-21 serum levels, which correlated with seropositivity for antibodies against the EBV and HHV-6 protein deoxyuridine triphosphate nucleotidohydrolase (dUTPases) but no increase in CXCL13. These cytokines are critical for T follicular helper (TFH) cell differentiation and for the generation of high-affinity antibodies and long-lived plasma cells. Notably, ME/CFS serum was sufficient to drive TFH cell differentiation via an activin A–dependent mechanism. The lack of simultaneous CXCL13 increase with IL-21 indicates impaired TFH function in ME/CFS. In vitro studies revealed that virus dUTPases strongly induced activin A secretion while in vivo, EBV dUTPase induced the formation of splenic marginal zone B and invariant NKTFH cells. Together, our data indicate abnormal germinal center (GC) activity in participants with ME/CFS and highlight a mechanism by which EBV and HHV6 dUTPases may alter GC and extrafollicular antibody responses.

Authors

Brandon S. Cox, Khaled Alharshawi, Irene Mena-Palomo, William P. Lafuse, Maria Eugenia Ariza

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Figure 3

ME/CFS sera–induced TFH cell differentiation of human naive CD4+ T cells is mediated by activin A.

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ME/CFS sera–induced TFH cell differentiation of human naive CD4+ T cells...
For human activin blocking experiments, dUTPase-derived or control DC-conditioned media (25% vol/vol) as well as ME/CFS or control sera (2.5% vol/vol) were preincubated with follistatin-315 (FST, 1 μg/mL) or vehicle (DMSO) for 1 hours, added to freshly isolated naive CD4+ T cells, and incubated for 3 days at 37°C. On day 4, supernatants and cells were collected for analysis of IL-21 levels in the presence or absence of FST by ELISA and TFH differentiation blockade by FACS, respectively. Activin A + IL-12 treatment in the presence of FST was used as a positive control for TFH differentiation blockade, and bead-treated cells were used as a negative control. Activin A + IL-12 treatment was used as a positive control for inducing TFH cell differentiation. (A) Representative flow cytometry plots indicating the frequency of PD-1+CXCR5+ double-positive TFH cells (upper right quadrants) and (B) quantitative bar graph of percentage of PD-1+CXCR5+ cells following human naive CD4+ T cell stimulation for 3 days as described above. (C) ELISA of IL-21 levels in supernatants of naive CD4+ T cells differentiated as described in A in the presence or absence of FST. Data represent a minimum of 3 experiments, with mean ± SEM. (B and C) n = 8–24, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 of treatment groups versus negative control or control sera by 1-way ANOVA Kruskal-Wallis multiple comparisons test with Dunn’s correction.