Nuclear Factor-Kappa B Inhibition Can Enhance Apoptosis of Differentiated Thyroid Cancer Cells Induced by 131I

Objective: To evaluate changes of nuclear factor-kappa B (NF-kappa B) during radioiodine 131(I-131) therapy and whether NF-kappa B inhibition could enhance I-131-induced apoptosis in differentiated thyroid cancer (DTC) cells in a synergistic manner. Methods: Three human DTC cell lines were used. NF-kappa B inhibition was achieved by using a NF-kappa B inhibitor (Bay 11-7082) or by p65 siRNA transfection. Methyl-thiazolyl-tetrazolium assay was performed for cell viability assessment. DNA-binding assay, luciferase reporter assay, and Western blot were adopted to determine function and expression changes of NF-kappa B. Then NF-kappa B regulated anti-apoptotic factors XIAP, cIAP1, and Bcl-xL were measured. Apoptosis was analyzed by Western blot for caspase 3 and PARP, and by flow cytometry as well. An iodide uptake assay was performed to determine whether NF-kappa B inhibition could influence radioactive iodide uptake. Results: The methyl-thiazolyl-tetrazolium assay showed significant decrease of viable cells by combination therapy than by mono-therapies. The DNA-binding assay and luciferase reporter assay showed enhanced NF-kappa B function and reporter gene activities due to I-131, yet significant suppression was achieved by NF-kappa B inhibition. Western blot proved I-131 could increase nuclear NF-kappa B concentration, while NF-kappa B inhibition reduced NF-kappa B concentration. Western blot also demonstrated significant up-regulation of XIAP, cIAP1, and Bcl-xL after I-131 therapy. And inhibition of NF-kappa B could significantly down-regulate these factors. Finally, synergism induced by combined therapy was displayed by significant enhancements of cleaved caspase 3 and PARP from Western blot, and of Annexin V positively staining from flow cytometry. The iodine uptake assay did not show significant changes when NF-kappa B was inhibited. Conclusion: We demonstrated that I-131 could induce NF-kappa B activation, which would attenuate I-131 efficacy in DTC cells. NF-kappa B inhibition by Bay 11-7082 or by p65 siRNA transfection was effective in suppressing NF-kappa B regulated anti-apoptotic changes and in combined regimen apoptosis was achieved synergistically.