Human dendritic cells require multiple activation signals for the efficient generation of tumor antigen-specific T lymphocytes

Dendritic cells (DC) are specialized cells of the immune system responsible for the initiation and regulation of both cellular and humoral responses. DC function is highly dependent on their level of maturation. In this study , we postulated that full DC maturation would require a combination of acti vating signals. When cultured monocyte-derived DC received stimulation with CD40 ligand (CD40L) and lipopolysaccharide (LPS) together, the IL-12 secre tion increased 5-60-fold and the IL-10 secretion increased 5-15-fold when c ompared with either stimulation alone. In addition, poly I .C, a double-str anded RNA analog that mimics viral infection, also synergized with CD40L to stimulate DC to secrete high levels of IL-12 and IL-10. Flow cytometry rev ealed an up-regulation in the expression of CD80, CD86 and CD83 following a ctivation with a soluble trimeric form of CD40L (CD40Ls) or LPS. However, n o further up-regulation was observed when both CD40Ls and LPS were used tog ether compared with a single stimulatory signal, suggesting that there was no correlation between the expression of these markers and the level of IL- 12/IL-10 secretion. Finally, specific cytotoxic T lymphocytes (CTL) were ge nerated using DC pulsed with a modified HLA-A2-restricted peptide epitope d erived from the melanoma antigen MART-1. DC activated with a combination of CD40Ls and LPS were more efficient in eliciting MART-specific reactivity c ompared to DC activated with CD40Ls or LPS alone. These results demonstrate that multiple maturational signals have a positive impact on the ability o f DC to secrete IL-12 and IL-10 and more importantly, to generate antigen-s pecific T lymphocytes.