Abstract C294: Involvement of iNOS/NO and ROS in a reciprocal interaction of glioma and microglia cells via TNF-α and MCP-1 production.

Abstract

Abstract The tumor microenvironment plays a critical role in glioblastoma invasion and progression, and microglia is one of the most abundant immune cells in the tumor. Using transwell co-culture methods with glioma cells (U87 and C6) and microglia cells BV-2, increased migration of glioma cells U87 and C6 by microglia BV2 cells or NO donor sodium nitro prusside (SNP) accompanied by increased NO production was observed, and BV2 co-culture-induced migration of U87 and C6 was blocked by adding a nitric oxide synthase (NOS) inhibitor N-nitro L-arginine methyl ester (NAME). The conditioned mediums (CM) derived from both glioma cells U87 and C6 (U87-CM and C6-CM) were prepared, CMs from glioma cells significantly induced iNOS protein and NO production cells in according with an occurrence of activated macrophages and without alternations in cell viability and cell cycle progression in microglia BV2. ERK inhibitor U0126 and JNK inhibitor SP600125 suppressed U87-CM and C6-CM-induced iNOS/NO production with respectively blocking phosphorylated ERK (pERK) and JNK (pJNK) protein expression stimulated by U87-CM and C6-CM. A significant increase in intracellular peroxide by U87-CM and C6-CM was detected via DCHF-DA assay, and vitamin C (Vit C) and N-acetyl cysteine (NAC)-reduced intracellular peroxide levels with inhibition of iNOS/NO production and pERK/pJNK protein stimulated by U87-CM and C6-CM were identified in BV-2 cells. Contribution of ROS, pERK, and pJNK to the migration of glioma cells was further demonstrated in the transwell co-culture system of glioma U87 and C6 with microglia BV-2 cells. Furthermore, TNF-α, MCP-1, CSF-1, and TGF-β were applied, and TNF-α and MCP-1 induced iNOS protein expression in time and concentration-dependent manners, and expression of TNF-α and MCP-1 mRNA in U87 and C6 cells were detected by RT-PCR. Neutralization of TNF-α or MCP-1 in U87-CM and C6-CM using TNF-α or MCP-1 antibody inhibited iNOS protein expression, and increased intracellular peroxide by TNF-α or MCP-1 was identified in BV2 cells. The reciprocal activation of glioma cells and microglia via ROS-dependent iNOS/NO elevation at least in part mediated by TNF-α and MCP-1 is elucidated. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):C294. Citation Format: Yen-chou Chen. Involvement of iNOS/NO and ROS in a reciprocal interaction of glioma and microglia cells via TNF-α and MCP-1 production. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr C294.