Abstract： Objective To investigate the inhibitory effects of NF-κB decoy oligodeoxynucleotides (ODN) on the expression of co-stimulatory molecules in Kupffer cells (KCs). Methods KCs were isolated by in situ collagenase perfusion and randomly divided into three groups: control group, LPS stimulation group (1 mg/L LPS), and NF-κB decoy ODN group, in which KCs were transfected with NF-κB decoy ODN (4 μg/1×105 KCs) before LPS stimulation. After 8 h of LPS stimulation, KCs were collected and CD80, CD40,and CD54 mRNA expression was assayed by reverse transcription-polymerase chain reaction. Results LPS stimulation significantly increased the expression of co-stimulatory molecules (CD80,CD40,and CD54) in KCs by 2.53,2.51 ,and 2.74 fold respectively over that of the control group (P<0.01). NF-κB decoy ODN could efficiently suppress KCs activation. In decoy group, the expression of CD80 and CD40 mRNA was significantly decreased by 0.52 and 0.55 fold respectively over that of the LPS group (P<0.01),while that of CD54 mRNA was not (P>0.05). Conclusion NF-κB decoy ODN can. efficiently inhibit the expression of co-stimulatory molecules in KCs,which provides reliable experimental data for the clinical application of NF-κB decoy ODN to inhibit acute rejection after liver transplantation in vivo.