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  • In summary we found that LTD modulates brain edema CysLT


    In summary, we found that LTD4 modulates SU 9516 edema; CysLT1 receptor mediates vasogenic edema while CysLT2 receptor may mediate cytotoxic edema via up-regulating AQP4 expression. These findings indicate the distinct roles of CysLT1 and CysLT2 receptors in the pathophysiological processes in the brain.
    Cysteinyl leukotrienes (CysLTs, including LTC, LTD, and LTE), 5-lipoxygenase metabolites of arachidonic acid, are potent inflammatory mediators . The actions of CysLTs are mediated by activating their receptors, cysteinyl leukotriene receptor 1 (CysLT) and receptor 2 (CysLT) . In peripheral tissues, CysLT and CysLT receptors modulate at least four responses: vascular and smooth muscle cell function, immune, inflammation, and tissue repair . In the central nervous system, the roles of CysLTs and 5-lipoxygenase in cerebral ischemic injury have been indicated by a line of evidence , , , , , , ; however, whether and how CysLT and CysLT receptors mediate the ischemic neuronal injury remains unknown. Recently, we found that the expressions of CysLT and CysLT receptors are induced in the neuron-appearing cells in human brain specimens from patients with traumatic brain injury and brain tumors , . We also reported that CysLT receptor antagonists, pranlukast and montelukast, protect against ischemic brain injury in rats and mice in vivo , , , . These findings indicate that CysLT and CysLT receptors may modulate brain injuries, including ischemic injury. However, since we have not found any protective effects of CysLT receptor antagonists on the in vitro ischemic-like injury in the primary cultured neurons (unpublished observations), the roles of CysLT and CysLT receptors in neuronal injury need to be further studied. Both CysLT and CysLT receptors are Gαq protein coupled receptors; but they may play distinct roles. For example, CysLT receptor plays an inhibiting role while CysLT receptor plays a facilitating role in bleomycin-induced pulmonary fibrosis in the receptor gene-knockout mice , . Whether the two receptors also play distinct roles in ischemic brain/neuron injury is unknown. To explore the possibly distinct roles, we investigated the effects of CysLT and CysLT receptor gene over-expression and antagonists on oxygen glucose deprivation (OGD)-induced in vitro ischemic injury in a neural cell line, rat pheochromocytoma cells (PC12 cells) , , . Materials and methods Cell culture and receptor gene transfection. PC12 cells were purchased from the Institute of Cell Biology, Chinese Academy of Sciences, Shanghai, China. The cDNA for mouse CysLT1 or CysLT2 receptor (mCysLT1 and mCysLT2, subcloned into pcDNA3.0) was kindly gifted by Professor C.D. Funk (University of Pennsylvania, USA). The pcDNA3.0 null vector was purchased from Invitrogen (Carlsbad, California, USA). The receptor cDNA expressing vectors and the null vector were lineared by PvuI and transfected into PC12 cells using Lipofectamine 2000 (Invitrogen, Carlsbad, California, USA) according to the manufacturer’s instructions. The permanently transfected PC12 cells were selected with 500μg/ml G418 in DMEM supplemented with 10% FBS, 100U/ml penicillin and 100μg/ml streptomycin. Single-cell subclones were isolated and plated at low density in 24-well plates, so that only a few clones grew per plate and one clone grew per well. Cells were grown for over 2 months in the selection media. The transfected PC12 cells were defined PC12/WT (transfected with null pcDNA3.0), PC12/mCysLT1 (pcDNA3.0/mCysLT1), and PC12/mCysLT2 (pcDNA3.0/mCysLT2) cells. Before experiment, the cells were photographed with a digital camera (Nikon Coolpix 4500, Japan) under a microscope (Nikon Eclipse TS100, Japan), and their sizes were measured with ImageTool software (University of Texas Health Science Center, San Antonio, USA).