T Cell Intrinsic Responses to TGFβ Regulates Allergen Sensitization By Modulating Type 2 T Follicular Helper Cell Development

Immunoglobulin E (IgE) is a potent mediator of allergic diseases, but the mechanisms that regulate IgE responses to innocuous environmental and food antigens remain unclear. Patients with Loeys-Dietz syndrome (LDS) who have mutations in genes encoding the TGFβ receptor are predisposed to IgE-mediated disorders. Here, using patient samples and a mouse model, we demonstrate that LDS mutations lead to reduced canonical TGFβ signaling, elevated total and allergen-specific IgE, increased type 2 follicular helper T cells (Tfh2), and exaggerated germinal center activity that was not prevented by the presence of wild type T regulatory cells. T cell intrinsic defects in LDS mice resulted in spontaneous sensitization to orally administered OVA, and inhibition of mammalian target of rapamycin (mTOR) prevented the exaggerated Tfh and IgE responses to OVA. Thus, TGFβ limits human and mouse Tfh2 cell development via the phosphatidylinositol-3-OH kinase gamma (PI3Kg)/AKT/mTOR pathway, and disruption of this pathway promotes allergic inflammation. The M318R allele previously identified in LDS patients was knocked into the mouse Tgfbr1 locus resulting in heterozygous TgfbrIWT/mut mice. Eight week old mice were orally gavaged once a week for 4 weeks with 5mg crude peanut extract and 10g Cholera Toxin from Vibrio cholera (Sigma) in 200uL water. One week after the last gavage, mice were bled from the submandibular cheek vein. OTII cells were purified from lymph nodes and spleen from OTII+/+RAG-/- mice that were WT or TgfbrIWT/mut, CD4+ bead purified (Miltenyi Biotec), and injected i.v. into CD45.1 WT mice. Hosts were fed water with 1.5% endotoxin-free OVA (Sigma Aldrich) for 7 days post-injection with normal mouse chow. In the rapamycin treatment experiments, mice were orally gavaged daily with vehicle or 125g rapamycin dissolved in DMSO and Kolliphor (Sigma) in a 3:1 ratio and then dissolved in water in a 1:25 ratio, on days 3-7. On day 8, the mesenteric lymph node and Peyer’s patches were collected, and smashed through 70M filters to make single-cell suspensions.