RNA Interference Selectivly Down-Regulated Connective Tissue Growth Factor by in Cardiac Fibroblast

Objective To investigate whether the expression of connective tissue growth factor(CTGF) in neonatal rat cardiac fibroblasts can be down-regulated by RNA interference.Methods The neonatal rat cardiac fibroblasts were isolated and cultured.Then the cells were transfected by plasmids CTGF-shRNA which carrying an U6 promoter and a CTGF-specific shRNA-coding template sequence,or by plasmid CTGF-con which carrying an U6 promoter and nonspecific shRNA-coding sequence.After selection with G418 administration for one month,stable transfection were obtained in the cells.The mRNA expression of CTGF,TGF-β_1,and FN were measured by reverse transcription-polymerase chain reaction(RT-PCR).Meanwhile,the protein expressions of CTGF,FN were detected by Western blot.Results Basal expression of CTGF was found in the control fibroblast cells.TGF-β_1 induced remarkable the mRNA and protein expression of CTGF and FN.The plasmid CTGF-shRNA resulted in 65%and 78%decrease in CTGF mRNA and protein expression,while the mRNA and protein expression of FN were decreased by 81%and 63%,associated with upregulation of TGF-β_1 by 32%.On the contrary,there was no significant difference in the cells transfected by the plasmid of CTGF-con.Conclusion These results indicate that RNAi can specifically knock down CTGF expression of neonatal rat cardiac fibroflast which may present a new strategy of gene therapy for myocardial fibrosis.