Figure 6

Tissue inhibitor of metalloproteinase (TIMP)-2 suppressed the DNA binding and transcriptional activities of NF-κB but enhanced the DNA binding and transcriptional activities of Nrf2 and cAMP-response element binding protein (CREB). Nuclear extracts were prepared from BV2 cells transfected with TIMP-2 or control vector after treatment with lipopolysaccharide (LPS) (100 ng/ml) for 3 h. Electrophoretic mobility shift assay (EMSA) was performed using NF-κB (A), anti-oxidant response element (ARE) (C), or cAMP response element (CRE) (E) probe. Competition assays indicate that the DNA-protein complex is sequence specific because the amount of complex was diminished by a molar excess of cold oligonucleotides but not by mutant (mNF-κB) or nonspecific oligonucleotides (Sp-1 or PU1). ‘F’ indicates free probe. Transcriptional activity of the reporter plasmids (B) (κB)₃-luc, (D) ARE-luc and (F) CRE-luc in TIMP-2-overexpressed BV2 cells. Cells co-transfected with reporter plasmid and TIMP-2 vector were treated with LPS (100 ng/ml) for 6 h, and reporter gene assay was performed. Data are reported as the means ± S.E.M. for three separate experiments. *P <0.05, significantly different from the LPS + control vector group.