ShRNA-mediated knock-down of CXCR7 increases TRAIL-sensitivity in MCF-7 breast cancer cells.

ABSTRACT

This study aims to investigate the effects of CXCR7-shRNA on TRAIL-mediated apoptosis and suppression of invasive migration and the underlying mechanisms. (1) We constructed CXCR-7-shRNA lentiviral vectors and confirmed their silencing efficiency in MCF-7 cells by RT-PCR analysis. (2) The effects of CXCR7 and/or TRAIL on cell proliferation were examined by MTT assay. (3) Trans well invasion assay was used to examine the effects of CXCR7 silencing and/or TRAIL on MCF-7 cell invasive migration. (4) Expression of Caspase-3, and Caspase-8, and MMP-2 and MMP-9 proteins was examined by Western blot analysis. (1) Viral titers were 2.95 × 10(8) TU/ml, 3.01 × 10(8) TU/ml, 3.26 × 10(8) TU/ml, and 3.08 × 10(8) TU/ml, respectively. (2) CHXR7 shRNAs markedly decreased CXCR7 mRNA expression in MCF-7 cells, among which CXCR7-shRNA-1 showed significantly higher rate of inhibition (P < 0.05). (3) Combination of TRAIL and CXCR7-shRNA-1 resulted in marked suppression of cell proliferation in time-dependent manner (P < 0.05). (4) Cell invasion capacity was inhibited in each experimental group as compared to blank control group at 48 h post treatments (P < 0.05). Among them, combination of TRAIL and CXCR7-shRNA had the highest inhibitory effect (P < 0.05). (5) Western blot analysis indicated that TRAIL alone does not affect CXCR7 expression, but either TRAIL + CXCR7 shRNA or CXCR7 shRNA alone markedly suppressed CXCR7 protein expression. Furthermore, combination of TRAIL and CXCR-7-shRNA significantly increased Caspase-3 and Caspase-8 expression and decreased MMP-2 and MMP-9 expression (P < 0.05). Knock-down of CXCR-7 expression leads to augmented TRAIL-mediated suppression of MCF-7 cell proliferation and invasion.