RNA interference targets on connective tissue growth factor and tissue inhibitor of metalloproteinase-1 in vitro

Objective To screen RNA interference(RNAi)targets on connective tissue growth factor(CTGF) and tissue inhibitor of metalloproteinase-1(TIMP-1)in vitro.Methods Three candidates RNAi targets for rat's CTGF and TIMP-1 gene were designed and the double-stranded RNA(dsRNA)were synthesized.Then,single or different combined dsRNAs were transfected into rat hepatic stellate cells(HSCs)activated by TGF-β1.The procol-α1,CTGF and TIMP-1 mRNA were detected by fluorescent quantitative PCR,and hepatic fibrosis indexes in HSC supernatant were detected by radioimmunoassay.Results The CTGF mRNA and procol-α1 mRNA were inhibited at 68.09% and 65.03%,respectively in CTGF-3 group when single RNAi targeted on CTGF gene was used,and the TIMP-1 mRNA and procol-α1 mRNA were inhibited at 68.55% and 62.84%,respectively in TIMP-1-3 group when single RNAi targeted on TIMP-1 gene was used;When double RNAi targeted on CTGF gene and TIMP-1 gene were used,the CTGF mRNA and TIMP-1 mRNA were inhibited at 76.60% and 79.03%,respectively in CTGF-3/TIMP-1-3 group,while CTGF-2/TIMP-1-3 group had the most effective effect on inhibiting procol-α1 mRNA with inhibition rate of 85.79%.Conclusion The most effective RNAi targets on CTGF and TIMP-1 were selected successfully and the double RNAi targeting on CTGF and TIMP-1 were stronger than any single RNAi.