JunB/AP-1 and NF-kappa B-mediated induction of nitric oxide synthase by bovine type I collagen in serum-stimulated murine macrophages.
Type I collagen comprises the majority of the total body collagens. In particular, bovine type I collagen is utilized for medical purposes and used widely in a variety of cell culture models as a fibrous component of extracellular matrix. This study was designed to explore the effects of type I collagen on the expression of inducible nitric oxide synthase (iNOS) in serum-stimulated Raw264.7 cells and to study the molecular mechanistic basis. Bovine, but not rat or murine, type I collagen increased NO production in serum-stimulated cells, which resulted from the induction of iNOS, as monitored by Northern and Western blot analyses. Bovine type I collagen in combination with serum activated JunB and JunB/AP-1 transcription complex, as evidenced by supershift and immunodepletion of the retarded AP-1 band with anti-JunB antibody. AP-1 complex was immunodepleted in part by anti-c-Jun or anti-JunD antibody. Extracellular signal-regulated kinase1/2 (ERK1/2), p38 kinase, and c-Jun N-terminal kinase (JNK) were all activated by bovine type I collagen in serum-stimulated cells. PD98059, but not SB203580 or JNK1(-) transfection, inhibited both ERK1/2 phosphorylation and JunB/AP-1 activation. Either PD98059 or MKK1(-) transfection suppressed the iNOS induction. The induction of iNOS accompanied activation of NF-kappa B with degradation of I-kappa B alpha. AP-1 and/or NF-kappa B decoy oligonucleotides and pyrrolidine dithiocarbamate suppressed the iNOS induction, which confirmed involvement of AP-1 and NF-kappa B as transcription factors. These results demonstrated that bovine type I collagen induces iNOS in serum-stimulated murine macrophages through JunB/AP-1 and NF-kappa B activation and that activation of ERK1/2 plays an essential role in JunB/AP-1 activation.
Cho
Min Kyung
MK
National Research Laboratory (MDT), College of Pharmacy and Research Institute of Pharmaceutical Sciences, Seoul National University, Seoul 151-742, South Korea.
Suh
Seung Hoon
SH
Kim
Sang Geon
SG
D001483:Base Sequence; D001769:Blood; D015152:Blotting, Northern; D024042:Collagen Type I / Q000502:physiology*; D017931:DNA Primers; D004789:Enzyme Activation; D004790:Enzyme Induction; D020935:MAP Kinase Signaling System; D008262:Macrophage Activation; D016328:NF-kappa B / Q000502:physiology*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D052247:Nitric Oxide Synthase Type II; D018808:Transcription Factor AP-1 / Q000502:physiology*
D024042:Collagen Type I; D017931:DNA Primers; D016328:NF-kappa B; D018808:Transcription Factor AP-1; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II