Effect of the combined use of oncolytic adenovirus carrying short hairpin RNA targeting special AT rich sequence binding protein-1 with L-oxaliplatin on the proliferation and apoptosis of colorectal cancer cells

Abstract

Objective To observe the effects of the combined use of oncolytic adenovirus carrying short hairpin RNA (shRNA) targeting special AT rich sequence binding protein-1 (SATB1) (ZD55-SATB1) with L-oxaliplatin (L-OHP) on the proliferation and apoptosis of colorectal cancer SW620 cells. Methods SW620 cells were divided into 5 groups and treated with phosphate buffer (PBS), L-OHP (16 μg/ml), ZD55-enhanced green fluorescent protein (EGFP, 10 multiplicity of infection, MOI), ZD55-SATB1 (10 MOI), ZD55-SATB1 (10 MOI) plus L-OHP (16 μg/ml) respectively. The viability of SW620 cells treated with L-OHP (0, 4, 8, 32 μg/ml) alone and ZD55-SATB1 (0, 0.1, 1.0, 10.0, 50.0 MOI) alone for 96 h was assessed by cell counting kit-8 (CCK-8) assay. And the viability of SW620 cells of 5 groups with 48 h treatment was assessed by CCK-8 assay. Cell apoptosis was determined by Annexin V propidium iodide (PI)/fluoresceine isothiocyanate (FITC) and Hoechst 33258 assay. The expression of E1A, SATB1 and apoptosis associated protein cysteinyl aspartate-specific protease (Caspase)-3, Caspase-8, B cell lymphoma/leukemia-2 (bcl-2) and bcl-2 associated X protein (bax) in SW620 cells was detected by Western blotting. Results CCK-8 assay showed that ZD55-SATB1 (10 MOI) plus L-OHP (16 μg/ml) (35.22±2.42)%, compared with corresponding dose ZD55-SATB1 (10 MOI) (52.18±2.85)% (P=0.028), ZD55-EGFP (10 MOI) (60.19±0.82)% (P=0.036) and L-OHP (16 μg/ml) (63.29±2.01)% (P=0.014), could significantly inhibit the viability of SW620 cells. Hoechst33258 staining showed the combination of ZD55-SATB1 (10 MOI) with L-OHP (16 μg/ml) (59.49±2.53)% produced obvious apoptosis of the SW620 cells as compared with ZD55-SATB1 (10 MOI) (44.24±8.81)% (P=0.019), ZD55-EGFP (10 MOI) (23.59±6.28)% (P=0.026) and L-OHP (16 μg/ml) (8.61±2.83)% (P=0.015), and the difference was statistically significant. Annexin V assay showed that ZD55-SATB1 (10 MOI) plus L-OHP (16 μg/ml) (55.84±2.14)%, as compared with corresponding dose ZD55-SATB1 (10 MOI) (36.17±1.51)% (P=0.018), ZD55-EGFP (10 MOI) (25.43±3.25)% (P=0.012) and L-OHP (16 μg/ml) (23.86±1.92)% (P=0.017), could significantly accelerate the apoptosis of SW620 cells. Western blotting showed that the adenovirus ZD55-SATB1 and ZD55-EGFP were efficient replication in SW620 cells and L-OHP had no effect on adenovirus replication. In addition, SATB1 expression in the ZD55-SATB1 plus L-OHP group was lower than PBS group, L-OHP group, ZD55-SATB1 group and ZD55-EGFP group. Western blotting analysis also confirmed that ZD55-SATB1 (10 MOI) plus L-OHP (16 μg/ml) could increase the expression level of proapoptotic protein Caspase-3, Caspase-8, bax, concomitant with down-regulating the expression level of antiapoptotic proteins bcl-2 (P=0.000). Conclusion The combined use of ZD55-SATB1 with L-OHP could significantly inhibit the proliferation and induce proapoptotic effect in SW620 cells. And the combination of the treatment of oncolytic adenovirus and L-OHP is better than L-OHP or adenovirus alone. Key words: Special AT rich sequence binding protein-1; Colorectal cancer; Oxaliplatin; Proliferation