Laura Maggi1, Gianni Montaini1, Alessio Mazzoni1, Beatrice Rossettini1, Manuela Capone1, Maria Caterina Rossi1, Veronica Santarlasci1, Francesco Liotta2, Oliviero Rossi3, Oreste Gallo4, Raffaele De Palma5, Enrico Maggi2, Lorenzo Cosmi2, Sergio Romagnani1, Francesco Annunziato6. 1. Department of Experimental and Clinical Medicine and DENOTHE Center, Florence, Italy. 2. Department of Experimental and Clinical Medicine and DENOTHE Center, Florence, Italy; Regenerative Medicine Unit and Immunology and Cellular Therapy Unit of Azienda Ospedaliera Careggi, Florence, Italy. 3. Regenerative Medicine Unit and Immunology and Cellular Therapy Unit of Azienda Ospedaliera Careggi, Florence, Italy. 4. Department of Surgery and Translational Medicine, University of Florence, Florence, Italy. 5. Department of Clinical & Experimental Medicine, Second University of Naples and Center for Biomolecular Studies Supporting Human Health, Second University of Naples, Naples, Italy. 6. Department of Experimental and Clinical Medicine and DENOTHE Center, Florence, Italy; Regenerative Medicine Unit and Immunology and Cellular Therapy Unit of Azienda Ospedaliera Careggi, Florence, Italy. Electronic address: francesco.annunziato@unifi.it.
Abstract
BACKGROUND: Protection against helminths consists of adaptive responses by TH2 cells and innate responses by group 2 innate lymphoid cells (ILC2s), with these latter being well characterized in mice but less so in human subjects. OBJECTIVE: We sought to characterize human circulating ILC2s and compare their functional profile with that of autologous TH2 cells. METHODS: Circulating ILC2s and TH2 cells were isolated by means of fluorescence-activated cell sorting and magnetic cell sorting and expanded in vitro. ILC2s were then stimulated with phorbol 12-myristate 13-acetate plus ionomycin, IL-25 plus IL-33 (IL-25/IL-33), or a mixture of Toll-like receptor ligands to evaluate their ability to produce cytokines, express CD154, and induce IgE production by autologous B cells. Cytokines and transcription factor gene methylation were assessed. RESULTS: ILC2s expressed GATA-3, retinoic acid orphan receptor (RORC) 2, and RORα; were able to produce IL-5, IL-13, and IL-4; and, accordingly, were characterized by demethylation of IL4, IL13, IL5, GATA3, and RORC2, whereas the IFNG, IFNG promoter, and TBX21 regions of interest were methylated. ILC2s expressed TLR1, TLR4, and TLR6, and TLR stimulation induced IL-5 and IL-13 production. Moreover, ILC2s expressed CD154 in response to phorbol 12-myristate 13-acetate plus ionomycin, IL-25/IL-33, or a mixture of TLR ligands. Stimulated ILC2s also induced IgM, IgG, IgA, and IgE production by B cells. Finally, circulating ILC2s from atopic patients were not different in numbers and frequency but expressed higher IL-4 levels than those from nonatopic subjects. CONCLUSION: This study provides the first evidence that human ILC2s can express CD154 and stimulate the production of IgE by B lymphocytes through IL-25/IL-33 stimulation or TLR triggering.
BACKGROUND: Protection against helminths consists of adaptive responses by TH2 cells and innate responses by group 2 innate lymphoid cells (ILC2s), with these latter being well characterized in mice but less so in human subjects. OBJECTIVE: We sought to characterize human circulating ILC2s and compare their functional profile with that of autologous TH2 cells. METHODS: Circulating ILC2s and TH2 cells were isolated by means of fluorescence-activated cell sorting and magnetic cell sorting and expanded in vitro. ILC2s were then stimulated with phorbol 12-myristate 13-acetate plus ionomycin, IL-25 plus IL-33 (IL-25/IL-33), or a mixture of Toll-like receptor ligands to evaluate their ability to produce cytokines, express CD154, and induce IgE production by autologous B cells. Cytokines and transcription factor gene methylation were assessed. RESULTS: ILC2s expressed GATA-3, retinoic acid orphan receptor (RORC) 2, and RORα; were able to produce IL-5, IL-13, and IL-4; and, accordingly, were characterized by demethylation of IL4, IL13, IL5, GATA3, and RORC2, whereas the IFNG, IFNG promoter, and TBX21 regions of interest were methylated. ILC2s expressed TLR1, TLR4, and TLR6, and TLR stimulation induced IL-5 and IL-13 production. Moreover, ILC2s expressed CD154 in response to phorbol 12-myristate 13-acetate plus ionomycin, IL-25/IL-33, or a mixture of TLR ligands. Stimulated ILC2s also induced IgM, IgG, IgA, and IgE production by B cells. Finally, circulating ILC2s from atopic patients were not different in numbers and frequency but expressed higher IL-4 levels than those from nonatopic subjects. CONCLUSION: This study provides the first evidence that human ILC2s can express CD154 and stimulate the production of IgE by B lymphocytes through IL-25/IL-33 stimulation or TLR triggering.
Authors: Alina Neunkirchner; Bernhard Kratzer; Cordula Köhler; Ursula Smole; Lukas F Mager; Klaus G Schmetterer; Doris Trapin; Victoria Leb-Reichl; Edward Rosloniec; Ronald Naumann; Lukas Kenner; Beatrice Jahn-Schmid; Barbara Bohle; Rudolf Valenta; Winfried F Pickl Journal: EBioMedicine Date: 2018-04-05 Impact factor: 8.143