MiR-25 exerts cardioprotective effect in a rat model of myocardial ischemia-reperfusion injury by targeting high-mobility group box 1

Fig. 1.. MiR-25 and high-mobility group box 1 (HMGB1) changed inversely in a rat model of myocardial ischemia-reperfusion injury (IRI). The miR-25 expression (A) and the mRNA and protein levels of HMGB1 (B, C) in myocardial tissues obtained from Sprague-Dawley rats subjected to ischemia for 45 min followed by reperfusion for 2, 4, and 6 h and sham-operated rats using quantitative real-time PCR (qRT-PCR) and western blot. **p < 0.01 vs the sham group. n = 6 for each group.

Fig. 2.. MiR-25 agomir administration reduced serum myocardial enzyme activities and inflammatory cytokines production during myocardial ischemia-reperfusion injury (IRI). Enzyme-linked immunosorbent assay (ELISA) detection of serum levels of creatine kinase muscle-brain fraction (CK-MB) (A), lactic acid dehydrogenase (LDH) (B), tumor necrosis factor-α (TNF-α) (C), and interleukin (IL)-1β (D) in Sprague-Dawley rats injected of agomir negative control (agomir-NC) or miR-25-Agomir (10 mg/kg) at 48 h before ischemia-reperfusion (I/R; reperfusion for 2 h) induction. **p < 0.01 vs the sham group; ##p < 0.01 vs the Agomir-NC group. n = 6 for each group.

Fig. 3.. MiR-25 agomir administration alleviated myocardial ischemia-reperfusion injury (IRI)–induced pathological damage and apoptosis. Representative images of Evans blue/triphenyltetrazolium chloride (TTC) staining (A) and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining (B) in myocardial tissues and the calculation of infarct size and apoptotic cells. A, The myocardial infarct size was evaluated by Evans blue/TTC double staining. The percentage of infarct areas was significantly reduced after the administration of miR-25 agomir. Scale bar: 5 mm. B, TUNEL staining was performed to evaluate myocardial apoptosis. Scale bar: 20 μm. IRI induction markedly increased the population of apoptotic cells, whereas miR-25 agomir administration resulted in a prominent reduction of apoptosis in myocardium tissues. C, The protein levels of high-mobility group box 1 (HMGB1), caspase-3, and B-cell lymphoma-2 (Bcl-2) in myocardial tissues were determined by western blot. *p < 0.05, **p < 0.01 vs the sham group; #p < 0.05, ##p < 0.01 vs the agomir negative control (Agomir-NC) group. n = 6 for each group.

Fig. 4.. Lentivirus-mediated shRNA interference targeting high-mobility group box 1 (LV-sh HMGB1) could exert protective effect on myocardial ischemia-reperfusion injury (IRI). Enzyme-linked immunosorbent assay (ELISA) detection of serum levels of creatine kinase muscle-brain fraction (CK-MB) (A), lactic acid dehydrogenase (LDH) (B), tumor necrosis factor-α (TNF-α) (C), and interleukin (IL)-1β (D) in Sprague-Dawley rats injected of LV-sh NC or LV-sh HMGB1 at 48 h before ischemia-reperfusion (I/R; reperfusion for 2 h) induction. E, Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining was performed to evaluate myocardial apoptosis. Scale bar: 25 μm. **p < 0.01 vs the sham group; ##p < 0.01 vs the LV-sh NC group. n = 6 for each group.