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  • In summary in the present study


    In summary, in the present study we found that CysLT1 receptor attenuates while CysLT2 receptor facilitates OGD-induced PC12 cell death as confirmed by receptor over-expression and antagonism. However, we cannot explain why their roles are distinct and why the cell size is increased in the CysLT2 receptor over-expressing PC12 cells, so much more should be investigated to clarify the implications and the properties of signal transduction of the two receptors in ischemic neuronal injury.
    Basophils are central to IgE-mediated inflammation, are found at increased levels in tissues of allergic subjects, and are further increased after allergen challenge., , Cross-linking high-affinity IgE receptors with LY2606368 HCl synthesis leads to generation of leukotriene (LT) C, which is converted to LTD and then to LTE., The cysteinyl leukotrienes (cysLTs) LTC, LTD, and LTE are increased in the airways of asthmatic subjects, with the ability to induce bronchoconstriction, vascular permeability, and mucus secretion., , , , CysLTs are well established as mediators of allergic rhinitis and asthma on the basis of the efficacy of antileukotriene therapy,, , and they might play a role in the accumulation of inflammatory cells to allergic tissue. Eosinophils and CD34 cells, which express cysLT receptors, are found at increased numbers in allergic tissue and have been shown to migrate in response to LTD., , Furthermore, cysLTs have been shown to play a role in survival of eosinophils and proliferation of eosinophil-basophil progenitor cells. Basophil numbers have been shown to increase in the airway after inhalation of LTE, supporting the hypothesis that cysLTs might have a direct effect on basophil accumulation in the airways. CysLT receptors are expressed on a variety of peripheral blood leukocytes, but of interest, they are also expressed on cells that synthesize LTC, such as eosinophils and mast cells. The expression of cysLT receptors has not been evaluated on basophils, and efforts to do this have been hampered by the limited number of basophils that can be purified from peripheral blood. Because cysLT1 receptors are expressed on the shared eosinophil and basophil progenitor cells, CD34 cells, and both cysLT1 and cysLT2 receptors are present on mature eosinophils, we speculated that basophils might also express cysLT receptors. Basophils were therefore evaluated for cysLT1 and cysLT2 message, protein, and functional responses to LTD. Methods
    Discussion We have shown that basophils express mRNA for cysLT1 and cysLT2 and cell-surface receptor by means of flow cytometry. Our data obtained from Western blotting experiments are not as clear. Probing with the available polyclonal antibodies revealed bands at approximately 52 and 58 kd in eosinophil lysates, which was not the expected molecular weight. Different glycosylation or dimerization of receptors by mature leukocytes might explain our observation of higher-molecular-weight leukotriene receptors than previously reported. The calculated molecular mass based on the amino acid sequence of cysLT1 is reported to be 38.5 kd,30, 31 and the monomeric recombinant protein is reported to be 42 kd, although these authors also reported larger amounts of dimerized and oligomerized receptors in the presence of denaturing gel detergents. An earlier study with a different polyclonal antibody to cysLT1 has reported a single band at approximately 45 kd in CD34+ hematopoietic progenitors and a number of cell lines. Using the polyclonal antibodies in the current study, we would have expected to find a band for cysLT1 near 50 kd and a band for cysLT2 between 53 and 55 kd (Cayman Chemical, personal communication). However, in both basophil and eosinophil lysates, we observed a 58-kd band when probing for cysLT1 and cysLT2. That we observed a reduction in the intensity of the band at 58 kd with cysLT2 blocking peptide suggests this band is indeed the cysLT2 receptor. There was no reduction in the intensity of the 58-kd band when a cysLT1 blocking peptide was applied, suggesting this band is not the cysLT1 receptor. These Western blotting results for cysLT2 are consistent with the flow cytometric data in basophils, together demonstrating that cysLT2 protein is detectable on the cell surface and in cell lysates. However, protein for cysLT1 is only detectable by means of flow cytometry with the available cysLT1 antibody.