Generation and functional analysis of human TNF‐α/iNOS‐producing dendritic cells (Tip‐DC)

Abstract Background A unique type of CD11c pos dendritic cells ( DC ) is abundant in inflamed tissue, for example, in chronic inflammatory skin diseases. Due to their remarkable production of tumor necrosis factor ( TNF )‐α and inducible NO synthase ( iNOS ), these cells have been referred to as TNF and iNOS ‐producing DC (Tip‐ DC ). While Tip‐ DC have been mainly characterized in murine models of infection, functional data about their human counterpart are lacking. Objectives We sought to generate human Tip‐ DC in vitro und thus provide a new model for the investigation of their phenotype and function. Methods We generated human Tip‐ DC from monocytic precursor cells of healthy individuals, atopic and psoriatic patients using human serum. Resting and stimulated cells were analyzed by flow cytometry, real‐time PCR , and by ELISA . INOS activity was measured by fluorometric detection of NO . Results Tip‐DC closely resembled their in vivo counterparts by expressing CD 11c, CD 86, and CD 40 while lacking CD 1a, CD 1c, or CD 207/Langerin. Bacterial stimulation of Tip‐ DC from healthy donors, atopic dermatitis, or psoriasis patients resulted in a similar increase in iNOS activity and TNF ‐α production. In kinetic experiments, TNF ‐α, a putative activator of Tip‐ DC , could not induce NOS 2. Upon bacterial stimulation, TNFA , IL6 , IL12B, and IL23A mRNA appeared in a first wave, while IL12A and NOS 2 mRNA were up‐regulated later on but not blocked by anti‐ TNF ‐α agents, implying a biphasic pro‐inflammatory response. Conclusions We developed a new model for the study of human Tip‐ DC and provide the first evidence of their pro‐inflammatory capacity.