Unlike Ts2, Ts6 did not induce LTB4 and PGE2 production during the initial cell activation, Selleckchem RAD001 but induced distinct amounts throughout the activation time course. Ts6 induced an upregulation on these mediators after 24 h and the rate of PGE2/LTB4 production remained constant throughout all the previous time points (Fig. 4B). Taking into consideration our findings that revealed leukocyte recruitment following the Ts2
or Ts6 injection, we investigated the role of potent leukocyte chemoattractants known as LTs (Faccioli et al., 1991; Herschman, 1996; Medeiros et al., 1999). For this purpose, we pre-treated mice with MK-886 to inhibit LTs synthesis (Ford-Hutchinson et al., 1980) and observed reduced cell numbers after Ts2 or Ts6 injection. We also employed mice that were unable to produce LTs (5-LO−/−) and injected them with Ts2 or Ts6. These mice demonstrated decreased cell numbers compared to WT animals. In addition, LTB4 was increased in the peritoneal fluid of mice exposed to Ts2 or Ts6
in comparison to mice injected with PBS (control). Taken together, these results showed that LTs, predominantly CYC202 mw represented by LTB4, are necessary to promote cellular migration following Ts2 or Ts6 inoculation. Taking into consideration that prostanoids are also involved in cell recruitment, we explored the involvement of cyclooxygenase (COX)-derived PGs in the cell increase observed in our results. For that purpose, we pre-treated mice with a COX-2 inhibitor celecoxib (Warner et al., 1999). Celecoxib-treated mice had a significantly diminished cellular migration, indicating that PGs (-)-p-Bromotetramisole Oxalate could be involved in this process.
Moreover, we also demonstrated a significant PGE2 increase in the peritoneal fluid of mice exposed to Ts2 or Ts6 compared with the PBS control. It is known that the secretion of lipid mediators can be associated with an influx of neutrophils and an increase in inflammatory cytokines (Medeiros et al., 1999; Fernandes et al., 2007; Bagga et al., 2003). Taken together, these results demonstrated that the influx of cells to the peritoneal cavity induced by Ts2 or Ts6 is partially dependent on LTs and PGs. Finally, we immunophenotyped the cells recruited to the peritoneal cavity after Ts2 or Ts6 injection. We observed that the cells were positive for GR1, F4/80, CD3, CD4 and CD8 markers after the Ts2 or Ts6 injection. These are the common surface markers used to characterize neutrophils (GR1), macrophages (F4/80+), CD4 (CD3+/CD4+) and CD8 (CD3+/CD8+) lymphocytes (Ramalingam et al., 2003; Pillai et al., 2009). Thus, this result reinforced the observation that neutrophils are increased in mice injected with the toxins and showed that the detected mononuclear cells are mainly macrophages and lymphocytes. As expected, after treatment with MK-886 or celecoxib, the percentage of cells expressing surface markers to GR1+ decreased.