1(a), LTC4 increased in a dose-dependent manner, the expression of MHC class II on immature DCs was more significant at 10−8 m, so the trials were conducted using this concentration. Then, considering that LTC4 is released during inflammatory responses,17,30 we studied the effect of LTC4 (10−8 m) on the phenotype of immature DCs and LPS-stimulated DCs. Interestingly, after selleck kinase inhibitor 18 hr of culture, LTC4 strongly inhibited the expression of CD86 and CD40 molecules (Fig. 1b,c,f) when DCs were activated with 1 μg/ml LPS, whereas the lipid mediator

had no effect on immature DCs. However, in the case of the class II molecules, LTC4 had antagonistic effects depending on the activation status of DCs, increasing its expression in immature DCs and inhibiting in LPS-treated DCs (Fig. 1d,f). As shown in Fig. 1(g), although MHC class II decreased its expression in LPS-activated DCs, LTC4 had the ability to prime T lymphocytes, because it induced a low but significant increase selleck screening library in the allostimulatory response mediated by activated DCs. This effect was also observed in immature DCs, which correlates with the increased expression of class II molecules by LTC4. Immature DCs are specialized to

sense the microenvironment and when stress or infection are detected they incorporate the antigen through phagocytosis or endocytosis.28,29,31,32 We aimed to determine whether LTC4 was able to affect the antigen uptake of immature and activated DCs. To this end, cells were treated or not with LPS (1 μg/ml) for 30 min at 37°, then DCs were incubated without or with 10−8 m LTC4 for 30 min at 37°. Finally, cells were washed and incubated in the presence of Zy (10 particles/DC) coupled to FITC for 30 min at room temperature or DX-FITC (100 μg/ml) for 40 min at 37°. The phagocytosis controls were supplied by DCs treated with cytochalasin B, a disruptor of actin microfilaments, 33 previous to their incubation with Zy-FITC. For DX endocytosis, the control of reaction was provided by DCs incubated with the antigen at 4°, because this is a temperature-dependent phenomenon. In

addition, we analysed the uptake of HRP. For this, after treatment with LTC4 (0·01 μm) of both DCs and LPS-stimulated DCs, these were cultured with 150 μg/ml HRP for 40 min at 37°. Subsequently, cells were washed Endonuclease several times with cold PBS and permeabilized by addition of 0·5% Triton X-100 in PBS for 30 min at room temperature. The control was provided by DCs treated with HRP but not permeabilized. Finally, the enzymatic activity was measured in supernatants of reaction by addition of the substrate [alpha-phenylendiamine (OPD)] and read at 492 nm. In Fig. 2(a), we demonstrated that LTC4 increased the phagocytosis of Zy-FITC by immature DCs but had no effect in LPS-activated DCs. In contrast, as shown in Fig. 2(b,c), uptake of DX and HRP was increased by LTC4 in both immature and LPS-stimulated DCs.