Inhibition of lipopolysaccharide-induced cyclooxygenase-2, tumor necrosis factor-alpha and [Ca2+]i responses in human microglia by the peripheral benzodiazepine receptor ligand PK11195.
The anti-inflammatory actions of the mitochondrial peripheral benzodiazepine receptor (PBR) agonist PK11195 [1-(2-chloro- phenyl)-N-methyl-N-(1-methylpropyl)-3-isoquinoline-carboxamide] were investigated in human microglia. Application of the microglial inflammatory stimulus lipopolysaccharide (LPS, at 100 ng/mL for 3 h), induced enhancement of the expressions of the inducible enzyme, cyclooxygenase-2 (COX-2) and the pro-inflammatory cytokine, tumor necrosis factor-alpha (TNF-alpha). PK11195 (at 50 microm) significantly inhibited the LPS-induced up-regulation of both inflammatory factors; at a lower concentration of PK11195 (2 microm) expression of TNF-alpha, but not COX-2, was reduced. Production of both factors, using immunocytochemistry for COX-2 and ELISA for TNF-alpha, was markedly reduced with 50 microm of PK11195 added to LPS solution. Acute application of LPS induced a transient increase in intracellular Ca2+[Ca2+]i exhibiting both a slow development and recovery in kinetic behavior. This increase in [Ca2+]i consisted primarily of a Ca2+ influx component accompanied by a smaller mobilization from intracellular Ca2+ stores. In the presence of PK11195, the amplitude of the [Ca2+]i response induced by LPS was reduced by 54%. Another mitochondrial agent cyclosporin A (CsA), which also acts at the permeability transition pore (PTP) of mitochondrial membrane but at a site different from the PBR, was ineffective in reducing either the LPS-induced expression of COX-2 and TNF-alpha or the endotoxin increase in [Ca2+]i. These results indicate that the mitochondrial effector PK11195 is a specific and effective agent for inhibiting LPS-induced microglial expressions of COX-2 and TNF-alpha and that modulation of Ca2+-mediated signaling pathways could be involved in the anti-inflammatory actions.
Choi
Hyan B
HB
Department of Pharmacology and Therapeutics, Division of Neurology, Faculty of Medicine, The University of British Columbia, 2176 Health Sciences Mall, Vancouver, British Columbia, Canada V6T 1Z3.
Khoo
C
C
Ryu
Jae K
JK
van Breemen
Edo
E
Kim
Seung U
SU
McLarnon
James G
JG
D001921:Brain / Q000166:cytology / Q000196:embryology; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D051546:Cyclooxygenase 2; D016572:Cyclosporine / Q000494:pharmacology; D004790:Enzyme Induction / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D058785:GABA-A Receptor Agonists; D006801:Humans; D007150:Immunohistochemistry; D007424:Intracellular Fluid / Q000378:metabolism; D007527:Isoenzymes / Q000378:metabolism*; D007546:Isoquinolines / Q000494:pharmacology*; D008024:Ligands; D008070:Lipopolysaccharides / Q000494:pharmacology; D008565:Membrane Proteins; D017628:Microglia / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D011451:Prostaglandin-Endoperoxide Synthases / Q000378:metabolism*; D011963:Receptors, GABA-A / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*; D015854:Up-Regulation / Q000187:drug effects*
D004791:Enzyme Inhibitors; D058785:GABA-A Receptor Agonists; D007527:Isoenzymes; D007546:Isoquinolines; D008024:Ligands; D008070:Lipopolysaccharides; D008565:Membrane Proteins; D011963:Receptors, GABA-A; D014409:Tumor Necrosis Factor-alpha; D016572:Cyclosporine; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; D011451:Prostaglandin-Endoperoxide Synthases; D002118:Calcium; C037850:PK 11195