GLUTEN DEPENDENT ACTIVATION OF CD4+ T CELLS BY MHC CLASS II EXPRESSING INTESTINAL EPITHELIAL CELLS

BACKGROUND AND AIM: Intestinal epithelial cells (IECs) are emerging as potential regulators of CD4⁺ T cell responses through MHC class II expression. However, their role as antigen-presenting cells and in the activation of gluten-reactive CD4⁺ T cells has never been proven.

METHODS: We assessed MHC Class II expression in IECs using biopsies from active and treated CeD patients, as well as in gluten-immunized and non-immunized DR3-DQ2.5 transgenic mice that lack murine MHC Class II. Organoid monolayers were developed from DR3-DQ2.5 mice, treated with or without IFN-γ, and epithelial MHC Class II expression was measured by flow cytometry. We studied the functional consequences of MHC Class II expression on organoid monolayers by co-culturing monolayers from DR3.DQ2.5 mice with splenic human (h)CD4⁺ T cells isolated from gluten-immunized double transgenic DR3-DQ2.5-hCD4 mice. Co-cultures were treated with zein (a non-gluten protein), gluten, gluten pre-digested with elastase-producing Pseudomonas aeruginosa PA14, or gluten pre-digested with its lasB mutant. Following 4-days of co-culture, T-cell proliferation, activation, and cytokine release were assessed.

RESULTS: MHC Class II expression was higher in IEC from both active CeD patients and gluten-immunized DR3-DQ2.5 mice than in controls. Organoid monolayers from gluten-immunized mice demonstrated MHC Class II expression, which increased following IFN-γ treatment. Gluten, but not zein, treatment of organoid monolayers cultured with CD4⁺ T cells increased T cell proliferation, expression of activation markers (CD25, CD44, and CD69), and production of proinflammatory cytokines IL-2, IFN-γ, and IL-15. Organoid monolayers exposed to gluten pre-digested by elastase-producing P. aeruginosa PA14 showed increased CD4⁺ T cell proliferation, activation, and cytokine production compared with monolayers exposed to gluten or gluten pre-digested with the lasB mutant.

CONCLUSIONS: Under inflammatory conditions, IECs upregulate MHC Class II, have the capacity to present gluten antigen, and activate gluten-specific CD4⁺ T cells, which is further potentiated by microbial elastase digestion of gluten. IECs may serve to further increase injury to the epithelium caused by gluten-specific CD4⁺ T cells in CeD.