Characterization and production in vitro of murine dendritic cells.
In the last five years a series of protocols has been developed to generate dendritic cells in high quantities. In our country, research in this area of the immunology has been initiated with the intention of using them in therapeutic approaches, in particular in developing experimental protocols for immune therapy against cancer. These studies require to be validated and deeply studied using experimental models, particularly, the murine model. In this project, the primary goal was generate murine dendritic cells derived from stem cells from the bone marrow, using for this issue an experimental protocol standardized by Lutz and col. (1999) and modified in this study. Then, for the phenotypic characterization of dendritic cells, surface markers such as; CD11c, CD86 and H2Kb were analyzed, through quantification by flow cytometry. The final goal of this work was to standardize a method for obtaining murine dendritic cells with the stable phenotypic characteristics, to be use in the immunization of mice with antigenic peptides for the study of biological mechanisms in the tumor immunology.
The results obtained at different days of cell culture (6, 8 and 10 days), showed a high presence of molecule integrin CD 11c and the coestimulatory molecule of T lymphocytes, CD86, at days 10 of culture, which would indicate a estate of phenotypic maturity of analyzed dendritic cells. In addition, it observed a high expression of Mayor Histocompatibility complex class I molecules (H2Kb), in analyzed dendritic cells, indicating that those cells may be able to present antigen and induce a specific T lymphocytes response in vivo. Finally, a double cellular staining was performed to detect the population that co-expressed the molecules CD11c and CD86 and which we defined as dendritic cells. The result showed that 55.27% of produced cells should correspond to mature dendritic cells. These studies will allow the initiation of project about T lymphocytes in vivo induction through the immunization with in vitro established dendritic cells.