Journal
PHARMACEUTICAL RESEARCH
Volume 28, Issue 4, Pages 752-761Publisher
SPRINGER/PLENUM PUBLISHERS
DOI: 10.1007/s11095-011-0384-y
Keywords
GFAP promoter; liver fibrosis; pri-miRNA; mimics; shRNA; TGF-beta 1
Funding
- National Institutes of Health [EB003922]
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The objective was to determine the role of promoters and miRNA backbone in shRNA-based hepatic stellate cell (HSC)-specific transforming growth factor (TGF)-beta 1 gene silencing. This is expected to avoid the side effect of non-specific TGF-beta 1 gene silencing. Two most potent shRNAs targeting 769 and 1033 start sites of rat TGF-beta 1 mRNA were cloned into pSilencer 1.0 vector for enhanced TGF-beta 1 gene silencing. We then constructed HSC-specific pri-miRNA mimic and pri-miRNA cluster mimic expression plasmids in which shRNA expression was driven by a glial fibrillary acidic protein (GFAP) promoter to achieve HSC-specific TGF-beta 1 gene silencing to avoid nonspecific inhibition of TGF-beta 1 expression in other cells and organs. These TGF-beta 1 pri-miRNA-producing plasmids showed the inhibition of proliferation and induced apoptosis of activated HSC-T6 cells. TGF-beta 1 pri-miRNA cluster mimic plasmids decreased TGF-beta 1 and collagen gene expression at both mRNA and protein levels. GFAP promoter driven TGF-beta 1 pri-miRNA producing plasmids have the potential to be used for site-specific gene therapeutics to treat liver fibrosis.
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