Dendritic cells matured in the presence of the lycopodium alkaloid annotine direct T cell responses toward a Th2/Treg phenotype

Abstract

Annotine is a lycopodane-type alkaloid isolated from the Icelandic club moss Lycopodium annotinum ssp. alpestre. Annotine does not inhibit acetylcholinesterase, as some other lycopodium alkaloids do, and other bioactivities have not been reported. The aim of this study was to determine the effects of annotine on maturation of dendritic cells (DCs) and their ability to activate allogeneic CD4+ T cells. Human monocyte-derived DCs were matured in the absence or presence of annotine at a concentration of 1, 10 or 100 μg/ml. The effect of the annotine on maturation of the DCs was determined by measuring concentration of cytokines in culture supernatant by ELISA and expression of surface molecules by flow cytometry. DCs matured in the absence or presence of annotine at 100 µg/ml were also co-cultured with allogeneic CD4+ T cells and concentration of cytokines in supernatants determined by ELISA and expression of surface molecules by flow cytometry. When cultured alone, DCs matured in the presence of annotine secreted less of the pro-inflammatory cytokines IL-6 and IL-23 and had a tendency toward less secretion of IL-12p40 than DCs matured in the absence of annotine. However, when DCs were matured in the presence of annotine and then co-cultured with allogeneic CD4+ T cells they secreted more IL-12p40 and had a tendency toward secreting more IL-6 than DCs matured in the absence of annotine and then co-cultured with T cells. Allogeneic CD4+ T cells co-cultured with DCs matured in the presence of annotine secreted more IL-13 than T cells co-cultured with DCs matured in the absence of annotine, but stimulating the DCs in the presence of annotine did not affect T cell secretion of IFN-γ and IL-17. There was also more IL-10 in co-cultures of T cells and DCs matured in the presence of annotine than in co-cultures of T cells and DCs matured in the absence of annotine. These results show that annotine increases the ability of DCs to direct the differentiation of allogeneic CD4+ T cells toward a Th2/Treg phenotype, which may be of interest in the development of new treatments for Th1- and/or Th17-mediated inflammatory diseases.