br Materials and methods br Results br Discussion We

Materials and methods
Results
Discussion We found that CysLT2 receptors were constitutively expressed in bronchial smooth muscle cells, and that CysLT2 receptor-expressing leukocytes, mainly consisting of macrophage/dendritic cells and eosinophils, infiltrated into the lung after multiple antigen challenges in sensitized mice. In addition, CysLT2 receptor activation may be functionally involved in allergic airway infiltration of eosinophils and mononuclear cells. To our knowledge, this is the first report demonstrating the existence of CysLT2 receptors in the histology of murine lung. Compared with CysLT2 receptors, CysLT1 receptor expression in the lung was not pronounced, even in bronchial smooth muscle. We used two different commercially available primary dpp-4 inhibitor against CysLT1 receptors, but found only a limited number of antibody-positive areas. Low expression of CysLT1 receptors, even in airway smooth muscle, is consistent with the fact that mouse airway smooth muscle only weakly responds to CysLTs [35]. It is impractical to directly compare expression between CysLT2 and CysLT1 receptors because different primary antibodies were used for detection. However, CysLT2 receptor expression in the murine lung was clearly detected in bronchial smooth muscle and in leukocytes infiltrating into the lung, implying that CysLT2 receptors exert functional roles in mouse lung pathophysiology. The CysLT2 receptor antagonist BayCysLT2RA significantly suppressed multiple antigen challenge-induced infiltration of eosinophils and mononuclear cells into the lung, indicating that CysLT2 receptor activation is involved in leukocyte migration. In contrast to the present finding, Barret et al. [21] reported that antigen-induced eosinophilic pulmonary inflammation was markedly augmented in CysLT2 receptor-deficient mice. In addition, negative regulation of CysLT2 receptors in Th2-type pulmonary inflammation was induced by inhibiting both CysLT1 receptor signaling and expression [21]. An in vitro study suggested that CysLT2 receptors can negatively regulate CysLT1 receptor expression and LTD4-induced mitogenic responses in mast cells [36]. The difference between our results and the literature [21] may be because of different experimental tools, CysLT2 receptor antagonists, and CysLT2 receptor-deficient mice. We have confirmed that BayCysLT2RA lacked effects on other receptors associated with smooth muscle constriction or relaxation, such as adrenergic α2A, α2B and β2 receptors, histamine H1 and H2 receptors, and endothelin ETA and ETB receptors (unpublished data). Regarding pharmakokinetics, when BayCysLT2RA was i.v. administered at 0.3 and 1 mg/kg, t1/2 was approximately 0.3 h (unpublished data). Collectively, the effect of BayCysLT2RA is specific for CysLT2 receptors, but not so long-lasting. This situation of lowered function of CysLT2 receptors under the treatment with BayCysLT2RA should be different from that in CysLT2 receptor deficient mice, in which CysLT2 receptors congenitally defected. In gene knock out mice, however, other compensatory changes due to the gene modification should also be considered. On the other hand, inhibition by CysLT2 receptor antagonist was not dose-dependent from 0.03–0.3 mg/kg. Increasing the BayCysLT2RA dose to 1 and 3 mg/kg, did not further inhibit infiltration of eosinophils and mononuclear cells (data not shown). The absence of dose-dependency may be related to the regulatory functions of CysLT2 receptors as reported by others [21], [36], but we have not examined the effect of BayCysLT2RA on CysLT1 receptor expression.