A Decoy Oligodeoxynucleotide Against Sp1 Can Suppress Cell Proliferation And Type Ⅰ Collagen Gene Expression In Cultured NIH3T3

0bjective To investigate the effect of decoy oligodeoxynucleotide (decoy ODN) against transcription factor Sp1 on cell proliferation and expression of typeⅠcollagen gene in mouse embryo fibroblasts(NIH3T3).Methods Dumb-bell decoy ODN targeting Sp1 was designed and synthesized.Sp1 decoy ODN was transfected into NIH3T3 cells by cationic JetPEI.The transfection efficiency was detected by flow cytometry.The distribution of decoy ODN in the cells was investigated by laser scanning confocal fluorescence microscope.The specific binding of decoy ODN with Sp1 was checked by electrophoretic mobility shift assay (EMSA).The cell proliferation was examined by WST-8.The effect of decoy ODN on the type Ⅰcollagen gene expression was analyzed by semi-quantity PCR and real-time PCR.Results Sp1 decoy ODN could be effectively transfected into NIH3T3 cells mediated by cationic JetPEI, transfection efficiency detected by flow cytometry was up to 99.24%. Spatio-temporal dsitributin of decoy ODN in the cells was presented by laser scanning confocal fluorescence microscope, almost all ODN was transported into cell nucleus after 12h. The specific binding of decoy ODN with Sp1 was verified by electrophoretic mobility shift assay (EMSA). 100nM of Sp1 decoy ODN could effectively inhibit cell proliferation of NIH3T3, especially on the second day and third day after transfection. Meanwhile, 100nM of Sp1 decoy ODN could effectively srepress expression of pro-α1(Ⅰ),pro-α2(Ⅰ)collagen gene mRNA analyzed by semi-quantity PCR and real-time PCR.Conclusion Dumb-bell Sp1 decoy ODN could inhibit the cell proliferation and the expression of typeⅠcollagen gene by antagonizing Sp1.