Role of trefoil factor3 / phosphatidylinositl 3-kinase / protein kinase B / nuclear factor κB in the proliferation and apoptosis of human papillary thyroid carcinoma cell line TPC-1 / 解剖学报

ABSTRACT

Objective To investigate the effects of trefoil factor 3 (TFF3) on the proliferation and apoptosis of human thyroid papillary carcinoma cell line (TPC-1) and its molecular mechanism. Methods The lentiviral expression vector of overexpression and knockdown of TFF3 gene were constructed, 293T cell was packaged to produce lentiviral particles, virus solution was collected and transfected into TPC-1 cells, enhanced cell TFF3-TPC-1 and enhanced control group ConTFF3-TPC-1; silencing cell shRNA-TFF3-TPC-1 and silencing control cells shCon -TPC-1. Western blotting, and Real-time PCR were used to detect the expression of TFF3 protein and mRNA of four groups. Growth curve and colony formation assay were used to detect the proliferation. Flow cytometry was used to analyze the apoptosis level of the four groups; Western blotting and immunocytochemistry were used to detect apoptosis-related protein and pathway protein phosphatidylinositol 3-kinase/ protein kinase B (PI3K/ Akt), nuclear factor-κB (NF-κB) expression. Results 1. Overexpression and inhibition of expression of TFF3 stable cell TFF3-TPC-1 and shRNA-TFF3-TPC-1 were constructed suscessfully. 2. The proliferation and cloning ability of TFF3-TPC-1 cells were significantly higher than those of ConTFF3-TPC-1 cells(P<0. 05 or P< 0. 01), the proliferation and cloning ability of shRNA-TFF3-TPC-1 cells were significantly lower than those of shCon-TPC-1 cells(P<0. 01); 3. The apoptosis rate of TFF3-TPC-1 cells was lower than that of ConTFF3-TPC-1 (0. 75%±0. 08% vs 5. 62%±0. 3%, P<0. 01),and the apoptosis rate of shRNA-TFF3-TPC-1 was higher than that of shConTPC-1 (22. 2% ± 1. 2% vs 5. 34% ± 0. 4%, P<0. 01); 4. After silencing TFF3 gene, the expressions of Bax, cytochrome C (Cyt-C), cleaved-Caspase-9, cleaved-Caspase-3 were up-regulated, and the expressions of Bcl-2, Akt, p-Akt and NF-κB-P65 were down-regulated (P<0. 05 or P<0. 01). Conclusion TFF3 may regulate the proliferation and apoptosis of TPC-1 cells by affecting the PI3K/ Akt/ NF-κB signaling pathway.