Activation of nuclear factor-kappaB in C6 rat glioma cells after transfection with glia maturation factor.

Abstract

The 17-kDa endogenous brain protein glia maturation factor (GMF) was transfected into C6 rat glioma cells using a replication-defective human adenovirus vector. The cells overexpressed GMF but did not secrete the protein into the medium. Transfection with GMF led to the activation of the transcription factor nuclear factor-kappaB (NF-kappaB), as evidenced by electrophoretic mobility shift assay of the nuclear extract, using a double-stranded oligonucleotide probe containing the consensus binding sequence for NF-kappaB. The specificity of binding was demonstrated by competition with unlabeled probe and by the nonbinding of the mutant probe. Binding was detectable as early as 3 h after transfection, peaked at 6 and 12 h, and gradually declined thereafter. The observed NF-kappaB activation was reduced by cotransfection with catalase and by the presence of high concentrations of pyruvate in the medium, suggesting the involvement of H2O2. The p38 mitogen-activated protein kinase inhibitor SB-203580 also suppressed the GMF-activated NF-kappaB, suggesting the involvement of the p38 signal transduction cascade. On the other hand, the phorbol ester phorbol 12-myristate 13-acetate activated NF-kappaB whether or not GMF was overexpressed. Along with NF-kappaB activation was an enhanced expression of superoxide dismutase (SOD), which was suppressed if NF-kappaB nuclear translocation was blocked by its specific decoy DNA, implicating NF-kappaB as an upstream mediator of this antioxidant enzyme. The p38 inhibitor SB-203580 also blocked the GMF-activated SOD. As NF-kappaB and SOD are both pro-survival signals, the results suggest a cytoprotective role for endogenous GMF in glial cells.

• Authors: R Lim, A Zaheer, M A Yorek, C J Darby, L W Oberley
• Journal: Journal of neurochemistry (J Neurochem) Vol. 74 Issue 2 Pg. 596-602 (Feb 2000) ISSN: 0022-3042 [Print] England
• PMID: 10646510 (Publication Type: Journal Article, Research Support, U.S. Gov't, Non-P.H.S., Research Support, U.S. Gov't, P.H.S.)

Chemical References

• Glia Maturation Factor
• NF-kappa B
• RNA, Messenger
• Pyruvic Acid
• Catalase
• Superoxide Dismutase

Topics

• Animals
• Catalase (genetics, physiology)
• Glia Maturation Factor (genetics, physiology)
• Glioma (metabolism)
• Humans
• NF-kappa B (drug effects, physiology)
• Pyruvic Acid (pharmacology)
• RNA, Messenger (metabolism)
• Rats
• Superoxide Dismutase (genetics)
• Transfection
• Tumor Cells, Cultured