Hepatic stellate cells (HSCs) are the primary cell type responsible for liver fibrogenesis. Transforming growth factor beta 1 (TGF-β1) and platelet-derived growth factor (PDGF) are key profibrotic cytokines that regulate HSC activation and proliferation with functional convergence. Dual RNA interference against their receptors may achieve therapeutic effects. A novel RNAi strategy based on HSC-specific GFAP promoter-driven and lentiviral-expressed artificial microRNAs (amiRNAs) was devised that consists of an microRNA-30a backbone and effective shRNAs against mouse Pdgfrβ and Tgfbr2. Then， its antifibrotic efficacy was tested in primary and cultured HSCs and in mice affected with carbon tetrachloride-induce... More
Hepatic stellate cells (HSCs) are the primary cell type responsible for liver fibrogenesis. Transforming growth factor beta 1 (TGF-β1) and platelet-derived growth factor (PDGF) are key profibrotic cytokines that regulate HSC activation and proliferation with functional convergence. Dual RNA interference against their receptors may achieve therapeutic effects. A novel RNAi strategy based on HSC-specific GFAP promoter-driven and lentiviral-expressed artificial microRNAs (amiRNAs) was devised that consists of an microRNA-30a backbone and effective shRNAs against mouse Pdgfrβ and Tgfbr2. Then， its antifibrotic efficacy was tested in primary and cultured HSCs and in mice affected with carbon tetrachloride-induced hepatic fibrosis. The study shows that amiRNA-mediated Pdgfrβ and Tgfbr2 co-silencing inhibits HSC activation and proliferation. After recombinant lentiviral particles were delivered into the liver via tail-vein injection， therapeutic amiRNAs were preferentially expressed in HSCs and efficiently co-knocked down in situ Tgfbr2 and Pdgfrβ expression， which correlates with downregulated expression of target or effector genes of their signaling， which include Pai-1， P70S6K， and D-cyclins. amiRNA-based HSC-specific co-silencing of Tgfbr2 and Pdgfrβ significantly suppressed hepatic expression of fibrotic markers α-Sma and Col1a1， extracellular matrix regulators Mmps and Timp1， and phenotypically ameliorated liver fibrosis， as indicated by reductions in serum alanine aminotransferase activity， collagen deposition， and α-Sma-positive staining. The findings provide proof of concept for the use of amiRNA-mediated co-silencing of two profibrogenic pathways in liver fibrosis treatment and highlight the therapeutic potential of concatenated amiRNAs for gene therapy.
