Background/Purpose:

Dendritic cells (DCs) play a central role in regulating immune responses and govern T cell priming and polarization, mediating immunity and tolerance. Recent progress uncovered the Wnt signaling pathway is crucial for the immune balance and focuses on DCs as a direct target for their immunoregulatory role. In this study, we investigated the effect of the canonical and non-canonical Wnt signaling pathways regulate the differentiation and function of human DCs.

Methods:

Human peripheral blood CD14+ monocytes were cultured for 6 days with GM-CSF/IL-4 and then treated with LPS, Wnt3a, Wnt5a, GSK-3b inhibitor (SB216763) or silencing b-catenin for 24 h. Immunophenotypic characterizations of DCs were analyzed by flow cytometry. The supernatants were collected and cytokines (IL-17, IL-23, IL-10, TNF-¦Á, IFN-¦Ã) were assessed. Human peripheral blood and active rheumatoid arthritis patients’ synovial fluid CD3+ monocytes were isolated for mixed lymphocyte reaction. Proteins were extracted and incubated with antibodies against the total or phosphorylated forms of b-catenin, GSK-3b.

Results:

Our results showed that regulating the canonical and non-canonical Wnt signaling altered the maturation status and function of already differentiated DCs. The differentiation of DCs resulted in inhibiting activity of GSK-3b and stabilizing its substrate, b-catenin. GSK-3b inhibitor induced a full maturation phenotype of DCs that expressed mostly CD14. These DCs produced low levels of IL-17, IL-23 but high level of IL-10. Consequently, these DCs have a reduced capacity to stimulate Th17 differentiation and increased Th2 and Treg differentiation. The addition of Wnt3a or Wnt5a both increased CD1a+DCs but reduced CD83, CD80, CD86, CD40 and HLA-DR expression. Wnt3a-DCs produce scant amounts of cytokines and have a reduced capacity to stimulate all T cell subsets differentiation. Wnt5a-DCs produced low level of IL-17, TNF-¦Á while increased IL-23, IL-10 secretion and have an increased capacity to stimulate Th2 and Treg differentiation. b-catenin silencing reduced CD83 expression on LPS-induced DCs maturation and IL-10 secretion, without significant effect on capacity of DCs to stimulate T cells differentiation.

Conclusion:

Wnt signaling pathway regulates multi-unconventional differentiation and function of human DCs. GSK-3b inhibitor could induce terminal differentiation DCs into unconventional phenotypic maturation of DCs with tolerogenic features. Silencing of b-catenin partly impair LPS induced maturation of DCs. Exogenous Wnt3a inhibited maturation and function of DCs while Wnt5a inhibited maturation of DCs with regulatory features.