TRPM2 contributes to LPS/IFNγ-induced production of nitric oxide via the p38/JNK pathway in microglia.
Microglia are immune cells that maintain brain homeostasis at a resting state by surveying the environment and engulfing debris. However, in some pathological conditions, microglia can produce neurotoxic factors such as pro-inflammatory cytokines and nitric oxide (NO) that lead to neuronal degeneration. Inflammation-induced calcium (Ca(2+)) signaling is thought to underlie this abnormal activation of microglia, but the mechanisms are still obscure. We previously showed that combined application of lipopolysaccharide and interferon γ (LPS/IFNγ) induced-production of NO in microglia from wild-type (WT) mice is significantly reduced in microglia from transient receptor potential melastatin 2 (TRPM2)-knockout (KO) mice. Here, we found that LPS/IFNγ produced a late-onset Ca(2+) signaling in WT microglia, which was abolished by application of the NADPH oxidase inhibitor diphenylene iodonium (DPI) and ML-171. In addition, pharmacological blockade or gene deletion of TRPM2 channel in microglia did not show this Ca(2+) signaling. Furthermore, pharmacological manipulation and Western blotting revealed that Ca(2+) mobilization, the proline-rich tyrosine kinase 2 (Pyk2), p38 mitogen-activated protein kinase (p38 MAPK) and c-Jun NH2-terminal kinase (JNK) contributed to TRPM2-mediated LPS/IFNγ-induced activation, while the extracellular signal-regulated protein kinase (ERK) did not. These results suggest that LPS/IFNγ activates TRPM2-mediated Ca(2+) signaling, which in turn increases downstream p38 MAPK and JNK signaling and results in increased NO production in microglia.
Department of Molecular Pharmacology, Graduate School of Pharmaceutical Sciences, Kyoto University, Kyoto, Japan.
Takahito
T
Miyake
Department of Molecular Pharmacology, Graduate School of Pharmaceutical Sciences, Kyoto University, Kyoto, Japan.
Hisashi
H
Shirakawa
Department of Molecular Pharmacology, Graduate School of Pharmaceutical Sciences, Kyoto University, Kyoto, Japan.
Ayaka
A
Kusano
Department of Molecular Pharmacology, Graduate School of Pharmaceutical Sciences, Kyoto University, Kyoto, Japan.
Shinya
S
Sakimoto
Department of Molecular Pharmacology, Graduate School of Pharmaceutical Sciences, Kyoto University, Kyoto, Japan.
Masakazu
M
Konno
Department of Molecular Pharmacology, Graduate School of Pharmaceutical Sciences, Kyoto University, Kyoto, Japan.
Takayuki
T
Nakagawa
Department of Synthetic Chemistry and Biological Chemistry, Graduate School of Engineering, Kyoto University, Kyoto, Japan.
Yasuo
Y
Mori
Department of Molecular Pharmacology, Graduate School of Pharmaceutical Sciences, Kyoto University, Kyoto, Japan. Electronic address: skaneko@pharm.kyoto-u.ac.jp.
Shuji
S
Kaneko
D000818:Animals; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D051418:Focal Adhesion Kinase 2 / Q000037:antagonists & inhibitors / Q000378:metabolism; D007371:Interferon-gamma / Q000494:pharmacology*; D048031:JNK Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology*; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008825:Miconazole / Q000494:pharmacology; D017628:Microglia / Q000187:drug effects* / Q000378:metabolism; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000378:metabolism; D009569:Nitric Oxide / Q000096:biosynthesis*; D009570:Nitriles; D050053:TRPM Cation Channels / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D020032:Tyrphostins / Q000494:pharmacology; D048051:p38 Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism
D002614:Chelating Agents; D004791:Enzyme Inhibitors; D008070:Lipopolysaccharides; D009570:Nitriles; D050053:TRPM Cation Channels; C544443:TRPM2 protein, mouse; D020032:Tyrphostins; D009569:Nitric Oxide; C001784:SF 6847; D004533:Egtazic Acid; D008825:Miconazole; D007371:Interferon-gamma; D019255:NADPH Oxidases; D051418:Focal Adhesion Kinase 2; C496053:Ptk2b protein, mouse; D048031:JNK Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid; D002118:Calcium
Ca(2+) dynamics; LPS; MAPK; Microglia; Nitric oxide; TRP channel