Investigation of cytokine responses and variations in the expression of beta-defensin-3 of Caco-2 human colon epidermal adenocarsinoma and THP-1 human leukemia monocyte cell lines in response to Toxoplasma gondii under various conditions

Mononuclear phogocytic cells and epithelial cells are effective during the initiation and regulation of the innate immune response. They have an active role in mucosal immune response both mechanically and by interaction with other cells with cytokine release. Defensins are microbicidal peptides that are expressed in various cells and are thought to be effective in the first line defense against pathogens. IL-12 and IL-10, showing proinflamatory and antiinflamatory activities, respectively, are actors of the cellular immunity and limit the infection of the host without causing immunopathology. The aim of this study was to observe the differences in the release of IL-12 and IL-10 and the expression of human beta-defensin-3 (hBD-3) inCaco-2 (human colon epidermal adenocarcinoma cell) and THP-1 (human leukemia monocytic cell) cell lines cultured alone or in co-culture, by the stimulation of Toxoplasma gondii tachyzoites either in direct contact with the cells or separated by an insert filter from the cells. Twenty-four hours after the addition of RH strain tachyzoites to the cells, the supernatants were collected from the experiment wells, and commercial ELISA kits (Invitrogen) were used according to the manufacturers instructions to measure IL-12 and IL-10 levels. HBD-3 expression of cells collected from the experiment wells afterfour and 24 hourswere analyzed by using real time PCR. For this procedure, complementary c-DNA was obtained (Transcriptor High Fidelity cDNA Synthesis Kit, Roche Diagnostics, Germany)after the extraction of RNA with a commercial kit (High pure RNA isolation kit, Roche Diagnostics, Germany). IL-12 was higher than IL-10 in all experiment wells. IL-12 was induced more in the co-culture wells where Caco-2 and THP-1 cells were challenged together, than the wells in which the cells infected with T.gondii tachyzoites alone. No differences in respect to cytokine response were observed between the cells with which tachyzoites were in contact and the cells which were separated from the parasites with an insert. In hBD-3 experiments, Caco-2 and THP-1 cells interacted in co-culture wells when infected with tachyzoites and displayed a higher level of hBD-3 expression than the condition when they were infected alone. This study showed that, IL-12 release and hBD-3 expression, which play a role in innate immunity, are greater when various antigens of T.gondii interacted with stimulated mononuclear cell and epithelial cells.