Targeted inhibition of platelet-derived growth factor receptor-β subunit in hepatic stellate cells ameliorates hepatic fibrosis in rats

Abstract

The activation of hepatic stellate cells (HSCs) is the key event of the pathogenesis of hepatic fibrosis. Platelet-derived growth factor (PDGF) is the most potent mitogen for HSCs, and PDGF receptor-beta subunit (PDGFR-beta) is required for the proliferation of HSCs induced by PDGF. In this study, a high gene-silencing-efficacy PDGFR-beta small interference RNA (siRNA) was synthesized that could suppress the PDGFR-beta expression and inhibit the activation and proliferation but could not induce the apoptosis of HSCs in vitro. To avoid the side effect of nonspecific interference of PDGFR-beta, we constructed an HSCs-specific short hairpin RNA (shRNA) expression plasmid in which PDGFR-beta shRNA was driven by a glial fibrillary acidic protein (GFAP) promoter. The double-staining immunofluorescence examination indicated that GFAP promoter could target the transgene expression into HSCs in carbon tetrachloride induced acute injured rat's liver and bile duct ligation (BDL)-induced chronic injured rat's liver. Furthermore, HSCs-specific PDGFR-beta shRNA could relieve liver injury and hepatic fibrosis in the rat's model induced by BDL. This study demonstrates that PDGFR-beta siRNA may be presented as an antifibrogenic agent. The application of HSCs-specific RNA interference induced by the GFAP promoter might supply a new powerful tool for cell-specific gene therapy of hepatic fibrogenesis.

A novel population of subepithelial platelet-derived growth factor receptor α-positive cells in the mouse and human colon

“…Adult cells in multiple organs expressing PDGF ligands and receptors often have important roles in pathophysiology. For example, PDGF receptor (PDGFR)-␣ and -␤ are found in fibroblasts in lung and mesangial cells in kidney, and PDGFR␤ is found in hepatic stellate cells, which are involved in lung fibrosis, glomerulosclerosis, and liver cirrhosis (5,11,43).…”

GFAP Promoter-Driven RNA Interference on TGF-β1 to Treat Liver Fibrosis

“…GFAP is an intermediate filament protein identified as a biomarker for both quiescent and activated HSCs (22). Furthermore, GFAP promoter has been utilized for HSC-specific gene expression (13,14) and is effective under the control of pol II, which can only recognize miRNA but not shRNA. Therefore, we selected miR-30 to be incorporated into the constructed plasmids.…”

“…It is the first time to construct TGF-β1 primiRNA mimic plasmids driven by glial fibrillary acidic protein (GFAP)-promoter, a known biomarker for HSCs of fibrotic livers (12,13). Since GFAP promoter relies on RNA polymerase II (pol II) to achieve RNAi (14), this promoterdriven shRNA is unlikely to have inefficient nuclear export. Therefore, we inserted shRNA sequences targeting TGF-β1 into pri-miRNA backbone (Fig.…”

Vaccination with Platelet-Derived Growth Factor B Kinoids Inhibits CCl4-Induced Hepatic Fibrosis in Mice

“…In vitro studies have demonstrated that PDGF-B is the most potent mitogenic factor for HSCs (Pinzani, 2002). Consequently, blockage of PDGF-B signaling inhibits experimental hepatic fibrosis (Borkham-Kamphorst et al, 2004a,b;Gonzalo et al, 2007;Chen et al, 2008).…”

“…Several approaches have been reported to block the PDGF-B signaling including reducing the synthesis of active PDGF-B (Borkham-Kamphorst et al, 2004b) or PDGFR␤ (Chen et al, 2008) by gene silencing, neutralizing PDGF-B with specific antibodies (Abs) (Ogawa et al, 2010), decoying PDGF-B with soluble PDGF-B receptors (Borkham-Kamphorst et al, 2004a), and suppressing the postreceptor signal transduction pathways (Gonzalo et al, 2007). Although the efficacies of these measures have been validated in experimental hepatic fibrosis or in cultured HSCs, they are not seemingly possible to be employed in clinical practice.…”