by Zhaowei Meng, Shanshan Lou, Jian Tan, Ke Xu, Qiang Jia, Wei Zheng
To evaluate changes of nuclear factor-kappa B (NF-?B) during radioiodine 131 (131I) therapy and whether NF-?B inhibition could enhance 131I-induced apoptosis in differentiated thyroid cancer (DTC) cells in a synergistic manner. Methods
Three human DTC cell lines were used. NF-?B inhibition was achieved by using a NF-?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-?B. Then NF-?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-?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-?B function and reporter gene activities due to 131I, yet significant suppression was achieved by NF-?B inhibition. Western blot proved 131I could increase nuclear NF-?B concentration, while NF-?B inhibition reduced NF-?B concentration. Western blot also demonstrated significant up-regulation of XIAP, cIAP1, and Bcl-xL after 131I therapy. And inhibition of NF-?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-?B was inhibited. Conclusion
We demonstrated that 131I could induce NF-?B activation, which would attenuate 131I efficacy in DTC cells. NF-?B inhibition by Bay 11-7082 or by p65 siRNA transfection was effective in suppressing NF-?B regulated anti-apoptotic changes and in combined regimen apoptosis was achieved synergistically.