A short-hairpin RNA targeting osteopontin downregulates MMP-2 and MMP-9 expressions in prostate cancer PC-3 cells

Abstract 

Osteopontin (OPN), a secreted phosphoglycoprotein, is frequently associated with cell proliferation and tumor metastatic spread in a variety of cancers. It has been reported that OPN induce matrix metalloproteinase (MMP)-2 and MMP-9 activations through nuclear factor kappaB (NF-κB)-mediated signaling pathways. In this study, we investigated the roles of OPN in human prostate cancer cells and provided clues about the possible functions of IkappaB kinase (IKK) in NF-κB-mediated OPN-induced activations of MMP-2 and MMP-9. Short-hairpin RNA (shRNA) expression vectors were used to inhibit OPN expression in PC-3 cells, human prostate cancer cell line, and IKK inhibitor VII were applied to inhibit the activities of IKK-1 and IKK-2. The results showed that OPN shRNA-mediated RNA interference can downregulate OPN, MMP-2 and MMP-9 expressions, thereby resulting in suppression of the proliferation, migration and invasion of PC-3 cells in vitro and tumor growth in vivo. Moreover, the inhibition of IKK-2 can suppress MMP-2 and MMP-9 expressions, in contrast, the inhibition of IKK-1 has no effects on the OPN, MMP-2 and MMP-9 expression levels. Thus, this study demonstrated that OPN knockdown could downregulate MMP-2 and MMP-9 expressions result in inhibiting the malignant physiological behaviors of PC-3 cell and that IKK-2 may play a crucial role in OPN-induced MMP-2 and MMP-9 expressions via NF-κB-mediated signaling pathways.

Abbreviations: DMEM-F12, mixture (1:1) Dulbecco’s-modified minimum essential medium and Ham’s F-12 medium, EGFR, epidermal growth factor receptor, EGFP, enhanced green fluorescent protein, FBS, fetal bovine serum, GAPDH, glyceraldehyde-3-phosphate dehydrogenase, IKK, IkappaB alpha kinase, MMP, matrix metalloproteinase, NF-κB, nuclear factor kappaB, OPN, osteopontin, RNAi, RNA interference, shRNA, short-hairpin RNA, siRNA, small interfering RNA, uPA, urokinase plasminogen activator

Keywords: Osteopontin, Matrix metalloproteinase, Nuclear factor kappaB, Human prostate cancer, Gene therapy