This study aimed to investigate the effects of cationic liposomes containing different cationic lipids (DC-Chol and DOTAP) and different pegylation ratios on siRNA transfection in human U251 glioma cells. The data showed that the transfection efficiency of DOTAP was much higher than that of DC-Chol and PEG at 2 mol% enhanced cellular uptake of siRNA. Cationic liposome-siRNA complexes with particle size around 100 nm were prepared. PEG modification could efficiently stabilize the liposome in the presence of serum, which might protect the siRNA from serum degradation and prolong the circulation time in vivo. Efficient intracellular uptake and lysosome release of siRNA in human U251 glioma cells were observed for pegylated DOTAP-based lipososomes compared with the control transfection reagent lipofectamine 2000. The results demonstrated that this cationic liposome might be a potential vehicle for the in vivo delivery of siRNA.