Aim To investigate the potential applica-tion of a non-viral gene carrier Tat-LK15 for delivering siRNA targeting nNOS in vitro, which provides evi-dence of Tat-LK15 mediating siRNA targeting nNOS in vivo for treatment of neuropathic pain. Methods 1. Tat-LK15 was mixed with siRNA, then the mixture was analyzed the best ratio by Gel retardation. The trans-fection efficiency of FAM-siRNA mediated by Tat-LK15 on RGC-5 cells was examined by Flow Cytome-try. The apoptosis ratio of RGC-5 was identified by Flow Cytometry 24 h after treated with the different do-ses of Tat-LK15 (1, 2. 5, 5, 10 and 20 μg). 2. The model of RGC-5 cell overexpression of nNOS protein was prepared. 3. RGC-5 cells were randomly divided into 5 groups:control group,model group, Tat-S group ( Tat-LK15 mediate nNOS/siRNA transfection model cell) , Lipo-S group ( LipofectamineTM RNAiMAX me-diate nNOS/siRNA transfection model cell) and Tat-N group ( Tat-LK15 mediate NCsiRNA transfection model cell) . Real-time Quantitative polymerase chain reac-tion( Q-PCR) and Western blot were used to evaluate nNOS expression level assay. Results It indicated that the Tat-LK15/siRNA complex completely formed at the weight ratio of 2∶ 1 (μg/μg) , and the transfec-tion efficiency was (84. 4 ± 3. 9)%. It caused cotytox-icity when Tat-LK15 dose was 20 μg ( 6. 1 μmol · L-1 ) , and the apoptosis rate more than control group [(10. 3 ± 1. 1)% vs (7. 4 ± 0. 9)%, P0．05)。结论Tat-LK15能高效转染siRNA,细胞毒性低,离体条件下可有效运载siRNA沉默nNOS的基因表达。
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