Exploring the mechanism of berberine-mediated Tfh cell immunosuppression

Abstract

Background: Our previous research revealed a novel function of berberine (BBR), a clinically relevant plant-derived alkaloid, as a suppressor of follicular T helper (T_fh) cell proliferation in secondary lymphoid organs of BBR-treated mice that underwent immunization for collagen-induced arthritis (CIA) in DBA1/J mice. Due to the importance of T_fh cell and B cell interactions in the generation of T cell-dependent humoral responses, the suppression of T_fh cell activity may have implications for the general safety of BBR as a prophylactic dietary supplement, and its potential use in antibody-driven autoimmune and hypersensitivity disorders.

Purpose: This research aims to characterize BBR's impact on the activation, differentiation, and proliferation of T_fh cells by examining the expression of key extracellular signaling molecules, as well as the activity of intracellular signaling molecules involved in the Ca²⁺-calcineurin-NFAT pathway and STAT3 phosphorylation, following activation.

Study design: In vitro experimental study using primary tissues.

Methods: To explore the direct effects of BBR on the proliferation and differentiation of Tfh cells, isolated naïve CD4⁺ T cells (>95% pure) were activated and differentiated into pre- T_fh cells in the presence or absence of BBR. The resulting T_fh cell populations and the expression of the key extracellular molecules CXCR5, ICOS, and PD-1 were measured. In addition, we examined the impact of BBR treatment on the activity of key intracellular signaling molecules involved in T_fh cell activation and differentiation following TCR ligation and/or CD28 signaling (p-ZAP-70, p-Lck, p-PLCγ1, NFATc1 and intracellular calcium, Ca²⁺, concentrations), as well as IL-6 signaling (p-STAT3).

Results: Treatment with BBR significantly reduced the expression of both CXCR5 (p < 0.01) and ICOS (p < 0.005), but not PD-1, and reduced the percentage of T_fh cells within the total CD4⁺ T cell population. BBR treatment also led to a reduction in intracellular Ca²⁺ flux, activation of p-STAT3, and IL-21 production.

Conclusion: Our observations provide insight into the mechanism of BBR-mediated T_fh cell suppression and suggest that BBR treatment can directly inhibit T_fh cell activity, perhaps through interfering with cytokine receptor or downstream signaling.

Keywords: CXCR5; IL-21; STAT3; T cell; T follicular helper cell; berberine.