Induction of prostaglandin E2 synthesis and microsomal prostaglandin E synthase-1 expression in murine microglia by glioma-derived soluble factors. Laboratory investigation.
OBJECTIVE
Microglia are one of the members of monocyte/macrophage lineage in the central nervous system (CNS) and exist as ramified microglia in a normal resting state, but they are activated by various stimuli, such as tumors. Activated microglia induce immune responses in the CNS, but the precise functions of microglia in glioma microenvironments are not clear. It has been reported that glioma cells produce prostaglandin (PG)E2, which promotes the growth of tumor cells and possesses immunosuppressive activity. The authors previously reported that PGE2 production by peritoneal macrophages was enhanced by glioma-derived soluble factors, which induce an immunosuppressive state. In this study, they investigated PGE2 production by microglia treated with glioma cells and assessed the role of microglia in glioma microenvironments in the mouse.


METHODS
Microglia and peritoneal macrophages were cultured in vitro with or without lipopolysaccharide, and tumor necrosis factor (TNF) and PGE2 in the culture supernatant were measured using L929 bioassay and enzyme immunoassay. The expression of mRNA was measured using reverse transcriptase polymerase chain reaction, and the protein expression was assayed with Western blotting. In some experiments glioma cells and conditioned glioma medium were added to the microglia cultures.


RESULTS
Glioma cells studied in this report did not produce a significant amount of PGE2. However, the coculture of microglia with glioma cells or conditioned glioma medium led to the production of a large amount of PGE2. The enhancement of PGE2 production by microglia was more significant than that by peritoneal macrophages. The expression of cyclooxygenase (COX)-2 and particularly the expression of microsomal PGE synthase (mPGES)-1 (a terminal enzyme of the arachidonate cascade) in microglia were enhanced by conditioned glioma medium. The enhancement of mPGES-1 expression in microglia was more significant than that in peritoneal macrophages. The production of TNF was suppressed when culturing microglia with conditioned glioma medium, but this suppression was abrogated by the addition of a COX inhibitor (NS-398) and a PGE2 receptor (EP4) antagonist. Furthermore, TNF production was not suppressed in microglia from mPGES-1-deficient mice.


CONCLUSIONS
These results indicate that PGE2 production by microglia is enhanced by conditioned glioma medium, which induces an immunosuppressive state in the CNS. Therefore, the manipulation of microglia, from the standpoint of PGE2, provides investigators with an important strategy to induce an effective antiglioma immune response.
Nakano
Yoshiteru
Y
Department of Neurosurgery, School of Medicine, University of Occupational and Environmental Health, Kitakyushu, Japan.
Kuroda
Etsushi
E
Kito
Tomohiro
T
Uematsu
Satoshi
S
Akira
Shizuo
S
Yokota
Akira
A
Nishizawa
Shigeru
S
Yamashita
Uki
U
D000818:Animals; D015153:Blotting, Western; D002478:Cells, Cultured; D018920:Coculture Techniques; D017077:Culture Media, Conditioned; D051546:Cyclooxygenase 2 / Q000032:analysis; D052246:Cyclooxygenase 2 Inhibitors / Q000494:pharmacology; D015232:Dinoprostone / Q000032:analysis*; D005260:Female; D005910:Glioma / Q000378:metabolism*; D019746:Intramolecular Oxidoreductases / Q000032:analysis*; D008070:Lipopolysaccharides / Q000494:pharmacology; D017737:Macrophages, Peritoneal / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D008815:Mice, Inbred Strains; D017628:Microglia / Q000378:metabolism*; D008861:Microsomes / Q000201:enzymology*; D009578:Nitrobenzenes / Q000494:pharmacology; D000071876:Prostaglandin-E Synthases; D012333:RNA, Messenger / Q000032:analysis; D018078:Receptors, Prostaglandin E / Q000037:antagonists & inhibitors; D058309:Receptors, Prostaglandin E, EP4 Subtype; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013449:Sulfonamides / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis
D017077:Culture Media, Conditioned; D052246:Cyclooxygenase 2 Inhibitors; D008070:Lipopolysaccharides; D009578:Nitrobenzenes; C545651:Ptger4 protein, mouse; D012333:RNA, Messenger; D018078:Receptors, Prostaglandin E; D058309:Receptors, Prostaglandin E, EP4 Subtype; D013449:Sulfonamides; D014409:Tumor Necrosis Factor-alpha; C080955:N-(2-cyclohexyloxy-4-nitrophenyl)methanesulfonamide; D051546:Cyclooxygenase 2; D019746:Intramolecular Oxidoreductases; D000071876:Prostaglandin-E Synthases; C000605865:Ptges protein, mouse; D015232:Dinoprostone