Liver-specific Bid silencing inhibits APAP-induced cell death in mice.

Abstract

Acetaminophen (APAP)-induced acute liver failure (ALF) is a life-threatening disease with only a few treatment options available. Though extensive research has been conducted for more than 40years, the underlying pathomechanisms are not completely understood. Here, we studied as to whether APAP-induced ALF can be prevented in mice by silencing the BH3-interacting domain death agonist (Bid) as a potential key player in APAP pathology. For silencing Bid expression in mice, siRNABid was formulated with the liver-specific siRNA delivery system DBTC and administered 48h prior to APAP exposure. Mice which were pre-treated with HEPES (vehicleHEPES) and siRNALuci served as siRNA controls. Hepatic pathology was assessed by in vivo fluorescence microscopy, molecular biology, histology and laboratory analysis 6h after APAP or PBS exposure. Application of siRNABid caused a significant decrease of mRNA and protein expression of Bid in APAP-exposed mice. Off-targets, such as cytochrome P450 2E1 and glutathione, which are known to be consumed under APAP intoxication, were comparably reduced in all APAP-exposed mice, underlining the specificity of Bid silencing. In APAP-exposed mice non-sterile inflammation with leukocyte infiltration and perfusion failure remained almost unaffected by Bid silencing. However, the Bid silencing reduced hepatocellular damage, evident by a remarkable decrease of DNA fragmented cells in APAP-exposed mice. In these mice, the expression of the pro-apoptotic protein Bax, which recently gained importance in the cell death pathway of regulated necrosis, was also significantly reduced, in line with a decrease in both, necrotic liver tissue and plasma transaminase activities. In addition, plasma levels of HMGB1, a marker of sterile inflammation, were significantly diminished. In conclusion, the liver-specific silencing of Bid expression did not protect APAP-exposed mice from microcirculatory dysfunction, but markedly protected the liver from necrotic cell death and in consequence from sterile inflammation. The study contributes to the understanding of the molecular mechanism of the APAP-induced pathogenic pathway by strengthening the importance of Bid and Bid silencing associated effects.