Figure 2: AAV-based miR-125b-5p overexpression inhibits ALF in vivo.

(a) Schematic representation of the experimental design in APAP-induced ALF model. (b) Quantitative reverse transcriptase–PCR (qRT–PCR) revealed overexpression of miR-125b-5p in the liver of mice injected with AAV-Ttr-miR-125b-5p compared with liver of mice injected with control AAV. *P<0.05, two-tailed Student’s t-test. (c) Kaplan–Meier survival analyses of mice with or without hepatic miR-125b-5p overexpression (n=10 per group) after triggering ALF. P<0.05, log-rank test. (d) Serum ALT and AST analyses showed less liver damage at 6 h after APAP injection in miR-125b-5p-overexpressing mice. *P<0.05, two-tailed Student’s t-test. (e) Haematoxylin and eosin (HE) staining showed less injury at 6 h after induction of ALF. Scale bars, 100 μm. (f) Serum GDH, a mechanistic biomarker (g) liver GSH/GSSG ratio and (h) serum mtDNA, another mechanistic biomarker, revealed less hepatocyte damage in miR-125b-5p-overexpressing mice after APAP injection. *P<0.05 and **P<0.01, two-tailed Student’s t-test. (i) Schematic representation of the experimental design in FAS-induced ALF model. (j) qRT–PCR revealed overexpression of miR-125b-5p in liver of mice injected with AAV-Ttr-miR-125b-5p compared with liver of mice injected with control AAV. **P<0.01, two-tailed Student’s t-test. (k) Kaplan–Meier survival analyses of mice with or without hepatic miR-125b-5p overexpression (n=10 per group) after triggering ALF. P<0.0001, log-rank test. (l) Serum ALT and AST analyses showed less liver damage at 6 h after induction of FAS-induced ALF in miR-125b-5p-overexpressing mice. *P<0.05, two-tailed Student’s t-test. (m) HE staining indicated less apoptosis at 6 h after induction of ALF. Scale bars, 100 μm. (n) Caspase-3/7 activity assay showed lower caspase-3/7 activity in mice with miR-125b-5p overexpression, compared with control mice. **P<0.01, two-tailed Student’s t-test. (o) TUNEL assay revealed less cell death in miR-125b-5p-overexpressing mice at 6 h after induction of ALF. Scale bars, 100 μm. (p) Quantification of TUNEL staining shown in o. **P<0.01, two-tailed Student’s t-test. Data are presented as mean±s.e.m. from at least three independent experiments.