Abstract
AIM: Cysteinyl leukotriene receptor 1 (CysLT(1) receptor) is located in epithelial cells, and translocates from the plasma membrane to the nucleus in a ligand-dependent manner. Here, we investigated whether CysLT(1) receptors translocated to the nucleus in endothelial cells after ischemic insult in vitro and whether it was involved in ischemic injury to endothelial cells. METHODS: EA.hy926 cell line, derived from human umbilical vein endothelial cells, was subjected to oxygen-glucose deprivation (OGD). The expression and distribution of CysLT(1) receptors were detected by immunofluorescent staining, immunogold labeling and immunoblotting analyses. Cell viability was evaluated using MTT reduction assay. Necrosis and apoptosis were determined by double fluorescent staining with propidium iodide and Hoechst 33342. RESULTS: CysLT(1) receptors were primarily distributed in the cytoplasm and nucleus in EA.hy926 cells, and few was found in the cell membrane. OGD induced the translocation of CysLT(1) receptors from the cytoplasm to the nucleus in a time-depen dent manner, with a peak reached at 6 h. OGD-induced nuclear translocation of CysLT(1) receptors was inhibited by pretreatment with the CysLT(1) receptor antagonist pranlukast (10 μmol/L), or by preincubation with NLS-pep, a peptide corresponding to the nuclear localization sequence of CysLT(1) receptor (10 μg/mL). However, zileuton, an inhibitor of 5-lipoxygenase that was a key enzyme in cysteinyl leukotriene generation, did not inhibit the nuclear translocation of CysLT(1) receptors. Moreover, preincubation with NLS-pep (0.4 μg/mL) significantly ameliorated OGD-induced cell viability reduction and necrosis. CONCLUSION: CysLT(1) receptors in endothelial cells translocate to the nucleus in a ligand-independent manner after ischemic insult in vitro, and it is involved in the ischemic injury.