AP-1 Inhibits Expression of MMP-2/9 and Its Effects on Rat Smooth Muscle Cells

• Published in: The Journal of surgical research 2009-11, Vol.157 (1), p.e31-e37
• Main Authors: Zhang, Hong-Wei, Ph.D, Wang, Xin, Ph.D, Zong, Zhi-Hong, Ph.D, Huo, Xin, Ph.D, Zhang, Qiang, Ph.D
• Format: Article
• Language: English
• Subjects: Animals; Aorta, Thoracic - cytology; AP-1; AP-1siRNA; Cell Differentiation - physiology; Cell Division - physiology; Cell Movement - physiology; Enzyme Activation - physiology; Enzyme-Linked Immunosorbent Assay; Extracellular Signal-Regulated MAP Kinases - metabolism; Gene Expression Regulation, Enzymologic - physiology; invasion; JNK Mitogen-Activated Protein Kinases - genetics; JNK Mitogen-Activated Protein Kinases - metabolism; Male; Matrix Metalloproteinase 2 - genetics; Matrix Metalloproteinase 2 - metabolism; Matrix Metalloproteinase 2 - secretion; Matrix Metalloproteinase 9 - genetics; Matrix Metalloproteinase 9 - metabolism; Matrix Metalloproteinase 9 - secretion; MMP-2; MMP-9; Muscle, Smooth, Vascular - cytology; Muscle, Smooth, Vascular - enzymology; Proto-Oncogene Proteins c-fos - genetics; Proto-Oncogene Proteins c-fos - metabolism; Rats; Rats, Wistar; RNA, Small Interfering; Surgery; Transcription Factor AP-1 - genetics; Transcription Factor AP-1 - metabolism; Transfection; vascular smooth muscle cells
• ISSN: 0022-4804; 1095-8673
• DOI: 10.1016/j.jss.2009.02.015

Background We designed AP-1 small interfering RNA (siRNA) to study the relationship between AP-1 expression and matrix metalloproteinase (MMP)-2 activation. Methods To suppress the expression of AP-1 gene, we used RNA interference to silence the AP-1 gene post-transcriptionally in rat smooth muscle cells. Effects of AP-1 siRNA on mRNA expression of AP-1 were examined using reverse transcriptase polymerase chain reaction (RT-PCR). Phosphorylation levels of c-Jun, c-Fos, and the activity of AP-1 were determined by Western blotting and AP-1 DNA-binding activity assay. To observe the expression of SM α-actin and downstream genes of AP-1, we simultaneously examined the activity of cell matrix metal proteinases and the migration ability of smooth muscle cells using a modified Boyden-chamber assay. Effects of AP-1 siRNA on rat vascular smooth muscle cells (VSMC) in vitro were evaluated using cell cycle analysis, MTT-tests, BrdU-ELISA and immunofluorescence. Results AP-1 siRNA decreased not only AP-1 mRNA and AP-1 binding activity, but also c-Jun and c-Fos phosphorylation levels and the expression levels of urokinase-type plasminogen (u-PA), MMPs, TGF-β1 and bFGF in VSMCs. The AP-1 siRNA also significantly inhibited PDGF/IL-1-induced MMP-2 and MMP-9 gelatinolytic activity in VSMCs. The invasion and migration of VSMC were also induced greatly. Conclusions AP-1 inhibited expression of MMP-2/9 and AP-1 siRNA was able to effectively inhibit the proliferation, migration and de-differentiation of rat smooth muscle cells.