Experimental Study Of The Sense Oligonucleotide On The Inhibition Of Collagen Genes' Promoter

Objective :To construct an experimental method for observation on the effects of exogenous sense oligonucleotide on collagen genes'promoter, and to discuss the probability that the exogenous sense oligonucleotide would reduce the transcription of collagen genes'promoter.Materials and methods : First, fibroblasts derived from keloids were cultured in vitro. Then one chimeric gene with the sequence between -2483bp to + 42 bp of collagen gene promoter was fused to Reporter Vectors—PGL3 as an recombinant, and double-stranded oligonucleotides with binding sites to Sp1,Ap1 were synthesized. At last the recombinants were transfected to human keloid fibroblasts as the control,but the sense oligonucleotides described above were simultaneously transfected with the recombinants to keloid fibroblasts, as the experiment group. Luciferase assay system was used to detect the expression of luciferase in PGL3 .Results : The PGL3 recombinants and the synthesized double-stranded oligonucleotides could be transfected to keloid fibroblasts. The PGL3 recombinants and the exogenous double-stranded oligonucleotides could bind competitively to transcription factor Ap1 and Sp1. Luciferase assay showed that the difference of luciferase expression in PGL3 in experiment groups were significantly higher than that in control, P <0.01.Conclusions : The PGL3 recombinant with gene sequence between -2483bp to + 42 bp of collagen gene promoter is suitable for the research of activation of gene promoter in keloid fibroblasts. The phenomena that the collagen gene promoter were reduced as sense double-strand oligonucleotides transfected into the cells indicates that the pathogenesis of cicatrix can be controlled with sense oligonucleotides technique.