Successful Milk Oral Immunotherapy Promotes Generation of Casein-Specific CD137+ FOXP3+ Regulatory T Cells Detectable in Peripheral Blood.

Background: Oral immunotherapy (OIT) is an emerging treatment for cow's milk protein (CMP) allergy in children. The mechanisms driving tolerance following OIT are not well understood. Regulatory T cells (TREG) cells are key inhibitors of allergic responses and promoters of allergen-specific tolerance. In an exploratory study, we sought to detect induction of allergen-specific TREG in a cohort of subjects undergoing OIT. Methods: Pediatric patients with a history of allergic reaction to cow's milk and a positive Skin Pick Test (SPT) and/or CMP-specific IgE >0.35 kU, as well as a positive oral challenge to CMP underwent OIT with escalating doses of milk and were followed for up to 6 months. At specific milestones during the dose escalation and maintenance phases, casein-specific CD4+ T cells were expanded from patient blood by culturing unfractionated PBMCs with casein in vitro. The CD4+ T cell phenotypes were quantified by flow cytometry. Results: Our culture system induced activated casein-specific FOXP3+Helios+ TREG cells and FOXP3- TEFF cells, discriminated by expression of CD137 (4-1BB) and CD154 (CD40L) respectively. The frequency of casein-specific TREG cells increased significantly with escalating doses of milk during OIT while casein-specific TEFF cell frequencies remained constant. Moreover, expanded casein-specific TREG cells expressed higher levels of FOXP3 compared to polyclonal TREG cells, suggesting a more robust TREG phenotype. The induction of casein-specific TREG cells increased with successful CMP desensitization and correlated with increased frequencies of casein-specific Th1 cells among OIT subjects. The level of casein-specific TREG cells negatively correlated with the time required to reach the maintenance phase of desensitization. Conclusions: Overall, effective CMP-OIT successfully promoted the expansion of casein-specific, functionally-stable FOXP3+ TREG cells while mitigating Th2 responses in children receiving OIT. Our exploratory study proposes that an in vitro TREG response to casein may correlate with the time to reach maintenance in CMP-OIT.

Signaling Through gp130 Compromises Suppressive Function in Human FOXP3+ Regulatory T Cells.

The CD4+FOXP3+ regulatory T cell (Treg) subset is an indispensable mediator of immune tolerance. While high and stable expression of the transcription factor FOXP3 is considered a hallmark feature of Treg cells, our previous studies have demonstrated that the human FOXP3+ subset is functionally heterogeneous, whereby a sizeable proportion of FOXP3+ cells in healthy individuals have a diminished capacity to suppress the proliferation and cytokine production of responder cells. Notably, these non-suppressive cells are indistinguishable from suppressive Treg cells using conventional markers of human Treg. Here we investigate potential factors that underlie loss of suppressive function in human Treg cells. We show that high expression of the IL-6 family cytokine receptor subunit gp130 identifies Treg cells with reduced suppressive capacity ex vivo and in primary FOXP3+ clones. We further show that two gp130-signaling cytokines, IL-6 and IL-27, impair the suppressive capacity of human Treg cells. Finally, we show that gp130 signaling reduces the expression of the transcription factor Helios, whose expression is essential for stable Treg function. These results highlight the role of gp130 in regulating human Treg function, and suggest that modulation of gp130 signaling may serve as a potential avenue for the therapeutic manipulation of human Treg function.