Transfection of nuclear factor-κB decoy oligodeoxynucleotide inhibits inflammatory factors expression in renal tubular epithelial cells

Objective To evaluate the effect of NF-KB decoy oligodeoxynucleotides (ODN) strategy mediated by protamine liposome on NRK-52E renal tubular epithelial cells in vitro. Methods Lipsome (1 : 1 DOTAP/cholesterol) and ODNs complexes were prepared at + /-charge ratio of 4 and then transfected into TNF-αstimulated, complete cultured (contained 20% fetal bovine serum) NRK-52E cells. The transfection efficiency was observed by fluorescence microscope and evaluated by flow cytometry. NF-κB DNA binding abilities were analyzed by electrophoretic mobility shift assay (EMSA) . Genes expression of endothelin-1 (ET-1), iNOS, intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecular-1 (VCAM-1) and monocyte chemoattractant protein-1 (MCP-1) were assessed by reverse transcription polymerase chain reaction (RT-PCR). Results Mediated by protamine liposome produced high transfection efficiency compared to naked transfection (transcription efficiency was 93. 20% and 39. 0%, respectively) . NF-κB DNA binding abilities activated by TNF-α were abolished by NF-κB decoy ODN, instead of scrambled ODN. TNF-α-induced gene expression of ET-1, iNOS, ICAM-1, VCAM-1 and MCP-1 was significantly inhibited by NF-κB decoy ODN (all P 0. 01) . Conclusion Protamine liposome mediated ODN transfection method is highly efficient in vitro and not affected by serum. Transfection of NF-KB decoy ODN results in attenuation of NF-κB activities, leading to the inhibition of inflammatory genes expression in TNF-α activated renal tubular epithelial cells.