Figure 3: MiR-125b-5p mimic delivery suppresses ALF in vivo.

(a) Schematic representation of the experimental design in APAP-induced ALF model. MiR-125b-5p mimic was injected 1 h after the induction of ALF. (b) Quantitative reverse transcriptase–PCR (qRT–PCR) revealed significantly higher levels of miR-125b-5p in the liver of mice injected with miR-125b-5p mimic compared with respective controls. *P<0.05, two-tailed Student’s t-test. (c) Kaplan–Meier survival analyses of mice with or without hepatic miR-125b-5p mimic (n=6 per group) after induction of ALF. P<0.05, log-rank test. (d) Serum ALT and AST analyses showed less liver damage after APAP injection in miR-125b-5p mimic-injected mice. *P<0.05, two-tailed Student’s t-test. (e) Haematoxylin and eosin (HE) staining showed less injury 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 mimic-injected mice after APAP injection. **P<0.01, two-tailed Student’s t-test. (i) Schematic representation of the experimental design in FAS-induced ALF model. Similar to APAP model, mice were injected with miR-125b-5p mimic 1 h after the injection of lethal dose of FAS. (j) qRT–PCR revealed overexpression of miR-125b-5p in the liver of mice injected with miR-125b-5p mimic compared with respective controls. **P<0.01, two-tailed Student’s t-test. (k) Kaplan–Meier survival analyses of mice with or without hepatic miR-125b-5p mimic (n=6 per group) after triggering ALF. It is important to mention that mice that survived in miR-125b-5p mimic-injected group remained alive at later time points as well (monitored for 1 month). P<0.05, log-rank test. (l) Serum ALT and AST analyses showed less liver damage after induction of FAS-induced ALF in miR-125b-5p mimic-injected mice. **P<0.01, two-tailed Student’s t-test. (m) HE staining indicated less apoptosis after induction of ALF in miR-125b-5p mimic-injected mice. Scale bars, 100 μm. (n) Caspase-3/7 activity assay revealed lower caspase-3/7 activity in mice with miR-125b-5p injected mice, compared with control mice. **P<0.01, two-tailed Student’s t-test. (o) TUNEL assay showed less cell death in miR-125b-5p-injected mice 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.