The Effects Of NF-κB On Epithelial -Mesenchymal Transition,Migration And Invasion Of Human Colon Cancer Cells

Objective To observe the role of NF-ΚB in the induction of epithelialmesenchymal transition (EMT), cell proliferation activity, migration and invasion of human colon cancer cell line HCT116, as well as its potential mechanism by controlling the expression of nuclear factorkappaB (NF-ΚB).Methods The HCT116 cells were divided into three groups:NF-ΚB activation group, NF-ΚB suppression group and control group. And treated with 20 ng/ml of tumor necrosis factor-a (TNF-a),20 μmol/1 of ammonium pyrrolidinedithiocarbamate(PDTC), and equal volume of culture medium, respectively. Four days later, cell proliferation activity was detected by MTT assay, cell morphological changes associated with EMT were observed and photographed by a phase contrast microscope, and the migration ability and invasiveness were assessed by transwell chamber assays. The protein expression of p65, P-p65, E-cadherin and N-cadherin were analyzed by Western blotting, and the mRNA expression of N-cadherin and E-cadherin were detected by quantitative real-time PCR.Results TNF-a promoted but PDTC inhibited the cell proliferation of colon can cells. The group of TNF-a caused a complete EMT phenotype, which manifested as the formation of large filopodia-like processes and spindle-cell shape, While the group of PDTC inhibited the occurrence of typical EMT phenomenon, and they arranged more closely. Meanwhile the TNF-a group and PDTC group showed significantly different cell invasion and migration. The former one promoted cell invasion (118.50±1.95) and migration (167.00±6.36) compared with the controls (97.75±3.77,140.00±4.32). And the other suppressed cell invasion (51.00±1.83) and migration (80.00±2.53) compared with the controls (97.75±3.77,140.00±4.32) (all P<0.05). As we suspected, the TNF-a group up-regulated the expression of P-p65 and N-cadherin, suppressed the expression of E-cadherin. In contrast, PDTC down-regulated the expression of P-p65 and N-cadherin, upregulated the expression of E-cadherin. Follows were the detailed results, the protein expression of P-p65, N-cadherin in TNF-a group (respectively 0.72±0.04,0.33±0.01) was higher than control group (respectively 0.42±0.01,0.29±0.01) (all P<0.05). While E-cadherin in TNF-a group (0.15±0.01) was lower than control group (0.42±0.03) (P<0.05).The mRNA expression of N-cadherin in TNF-a group (3.90 ± 0.47) was larger than control group(1.00±0.00). However, the E-cadherin(0.26±0.08) was no more than the control group (1.00 ± 0.00) (all P< 0.05).Conclusion NF-ΚB induces the occurrence of typical EMT phenomenon and promotes cell proliferation, invasion and migration in human colon cancer cell line HCT116. The activation of NF-ΚB may affect cell invasion and migration by inducing the occurrence of EMT.