Antioxidative Effect Of Montelukast And HAMI3379, Cysteinyl Leukotriene Receptor Antagonists, On Ischemic Injury In Rat Cortical Neurons In Vitro Preliminary Research Of Cysteinvl Leukotriene Receptor Expression In Microglial Cells In Vitro

Objective:To investigate the antioxidative effects of two antagonists for cysteinyl leukotriene receptors (CysLT1R and CysLT1R) in ischemic injury of rat cortical neurons.Methods:Rat cortical neurons in vitro were pretreated with the CysLT1R antagonist montelukast and the CysLT1R antagonist HAMI3379, then they were exposed to oxygen-glucose deprivation/recovery (OGD/R) or H2O2. Reactive oxygen species (ROS), mitochondrial membrane potential (MMP) depolarization, neuronal viability and lactate dehydrogenase (LDH) release were determined to evaluate drug effects. Meanwhile, RNA interference was used to inhibit the expression of CysLT1R and CysLT2R, and the effects were observed.Results:ROS production in neurons was significantly increased after1-h OGD, which reached a peak at0.5h and lasted about1.5h after recovery. Montelukast and HAMI3379at0.01～1μM moderately decreased OGD/R-increased ROS production (P <0.05). Montelukast mildly attenuated OGD/R-induced MMP depolarization (P<0.05), but HAMI3379had no such an effect. Exogenous H2O2reduced neuronal viability and increased LDH release, namely inducing neuronal injury. Montelukast and HAMI3379at0.1～1μM moderately attenuated H2O2-induced neuronal injury (P<0.05). However, both CysLT1R siRNA and CysLT2R shRNA did not affect the responses mentioned above.Conclusion:In ischemic neuronal injury, montelukast and HAMI3379exert a moderate antioxidative effect, and this effect may be receptor-independent. Objective:To preliminarily investigate the cysteinyl leukotriene receptor expression in microglial cells in vitro.Methods:From the brain of neonatal rat born within24h, microglia cells were isolated and purified to perform primary cultures. The expression of the cysteinyl leukotrienes receptors CysLT1R and CysLT2R as well as GPR17on microglial cells was observed by immunofluorescent staining, and the colocalization of these three receptors was also investigated. Moreover, the inflammation inducer LPS and the agonist NMLTC4were used to induce localization changes of the receptors.Results:In the purified microglial cells, the expression of the cysteinyl leukotrienes receptors CysLT1R, CysLT2R and GPR17was detected by immunofluorescent staining. Their colocalization, namely CysLT1R/CysLT2R, CysLT1R/GPR17and CysLT2R/GPR17, was determined. The inflammation inducer LPS induced translocation of CysLT2R into the cytoplasma with a concentration phenomenon; the CysLT2R selective agonist NMLTC4had a similar but less obvious effect.Conclusion:The cysteinyl leukotriene receptors CysLT1R, CysLT2R and GPR17express in primary microglial cells, and their colocalization has been observed. The inflammation inducer LPS can induce CysLT2R endocytosis and concentration, while the same effect of the CysLT2R selective agonist NMLTC4should to be further investigated.