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Reactive Oxygen Species‐Dependent Nitric Oxide Production in Reciprocal Interactions of Glioma and Microglial Cells
Author(s) -
Shen ShingChuan,
Wu MingShun,
Lin HuiYi,
Yang LiangYo,
Chen YiHsuan,
Chen YenChou
Publication year - 2014
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/jcp.24659
Subject(s) - nitric oxide , reactive oxygen species , glioma , chemistry , oxygen , reciprocal , microbiology and biotechnology , biology , biochemistry , cancer research , organic chemistry , linguistics , philosophy
Conditioned mediums (CMs) from glioma cells U87, GBM‐8401, and C6 significantly induced iNOS protein and NO production by microglial cells BV‐2 but without altering the cell viability or cell‐cycle progression of BV2 microglia. Significant increases in intracellular peroxide by U87‐CM and C6‐CM were detected by a DCHF‐DA assay, and vitamin (Vit) C and N‐acetyl cysteine (NAC)‐reduced intracellular peroxide levels elicited by CMs lead to inhibition of iNOS/NO production The extracellular signal‐regulated kinase (ERK) inhibitor, U0126, and c‐Jun N‐terminal kinase (JNK) inhibitor, SP600125, suppressed U87‐CM‐ and C6‐CM‐induced iNOS/NO production by respectively blocking phosphorylated ERK (pERK) and JNK (pJNK) protein expressions stimulated by U87‐CM and C6‐CM. Increased migration of U87 and C6 glioma cells by a co‐culture with BV‐2 microglial cells or adding the nitric oxide (NO) donor, sodium nitroprusside (SNP) was observed, and that was blocked by adding an NO synthase (NOS) inhibitor, N‐nitro L‐arginine methyl ester (NAME). Contributions of ROS, pERK, and pJNK to the migration of glioma cells was further demonstrated in a transwell coculture system of U87 and C6 gliomas with BV‐2 microglial cells. Furthermore, expressions of tumor necrosis factor (TNF)‐α and monocyte chemoattractant protein (MCP)‐1 messenger (m)RNA in U87 and C6 cells were detected by an RT‐PCR, and TNF‐α and MCP‐1 induced iNOS protein expression in time‐ and concentration‐dependent manners. Neutralization of TNF‐α or MCP‐1 in U87‐CM and C6‐CM using a TNF‐α or MCP‐1 antibody inhibited iNOS protein expression, and increased intracellular peroxide by TNF‐α or MCP‐1 was identified in BV‐2 cells. The reciprocal activation of glioma cells and microglia via ROS‐dependent iNOS/NO elevation at least partially mediated by TNF‐α and MCP‐1 is elucidated. J. Cell. Physiol. 229: 2015–2026, 2014. © 2014 Wiley Periodicals, Inc.