Glycolipids from the gut symbiont Bacteroides fragilis are agonists for natural killer T cells and induce their regulatory differentiation

Natural Killer T (NKT) cells are a lipid-antigen reactive, CD1d-restricted T cell subset with diverse functional properties that contribute to inflammatory and regulatory immune responses. The most studied lipid antigen target for these cells is alpha-galactosylceramide (GC). The commensal organism Bacteroides fragilis (B. fragilis) produces several forms of GC, but conflicting information exists about the influence of these lipids on NKT cells. Herein, we report the total synthesis of a major form of alpha-GC from B. fragilis (Bf alpha-GC), and several analogues thereof. We confirm the T cell receptor (TCR)-mediated recognition of these glycolipids by mouse and human NKT cells. Despite the natural structure of Bf alpha-GC containing lipid branching that limits potency, we demonstrate that Bf alpha-GC drives mouse NKT cells to proliferate and differentiate into producers of the immunoregulatory cytokine, interleukin-10 (IL-10). These Bf alpha-GC-experienced NKT cells display regulatory function by inhibiting the expansion of naïve NKT cells upon subsequent exposure to this antigen. Moreover, this regulatory activity impacts more than just NKT cells, as demonstrated by the NKT cell-mediated inhibition of antigen-stimulated mucosal-associated invariant T (MAIT) cells. These findings reveal that B. fragilis-derived NKT cell agonists may have broad immunoregulatory activity, providing insight into the mechanisms influencing immune tolerance to commensal bacteria and highlighting a potential means to manipulate NKT cell function for therapeutic benefit.