TGF-beta1 gene silencing for treating liver fibrosis.
Small interfering RNA (siRNA) and short hairpin RNA (shRNA) targeting different regions of transforming growth factor beta1 (TGF-beta1) mRNA were designed and the silencing effect was determined after transfection into immortalized rat liver stellate cells (HSC-T6). There was not only significant decrease in TGF-beta1, tissue inhibitor of metalloproteinase 1 (TIMP-1), alpha-smooth muscle actin (alpha-SMA) and type I collagen after transfection with TGF-beta1 siRNAs, but also synergism in gene silencing when siRNAs targeting two different start sites were used as a pool for transfection. The two siRNA sequences which efficiently inhibited TGF-beta1 gene expression were converted to shRNAs via cloning into the pSilencer1.0. There was significant decrease in TGF-beta1 and TIMP-1 when HSC-T6 cells were transfected with pshRNA targeting the same regions of TGF-beta1 mRNA as siRNAs. Furthermore, TGF-beta1 gene silencing in HSC-T6 cells significantly decreased the levels of inflammatory cytokines, such as tumor necrosis factor-alpha (TNF-alpha) and interleukin-1 beta (IL-1beta). In conclusion, both siRNA and shRNA showed sequence-specific and dose dependent TGF-beta1 gene silencing and have the potential to treat liver fibrosis.
Cheng
Kun
K
Department of Pharmaceutical Sciences, University of Tennessee Health Science Center, Memphis, Tennessee 38103, USA.
Yang
Ningning
N
Mahato
Ram I
RI
D000199:Actins / Q000235:genetics / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D024042:Collagen Type I / Q000235:genetics / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D020868:Gene Silencing / Q000502:physiology*; D015316:Genetic Therapy / Q000379:methods*; D055166:Hepatic Stellate Cells; D053583:Interleukin-1beta / Q000378:metabolism; D008103:Liver Cirrhosis / Q000235:genetics / Q000628:therapy*; D034741:RNA, Small Interfering / Q000235:genetics / Q000502:physiology*; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D053773:Transforming Growth Factor beta1 / Q000235:genetics* / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
D000199:Actins; D024042:Collagen Type I; D053583:Interleukin-1beta; D034741:RNA, Small Interfering; D019715:Tissue Inhibitor of Metalloproteinase-1; D053773:Transforming Growth Factor beta1; D014409:Tumor Necrosis Factor-alpha; C498862:Acta2 protein, rat