Bisacurone ameliorates myocardial ischemia/reperfusion injury in rats: regulation of inflammatory and apoptosis pathways via CHOP/GRP78 proteins

Fig. 1. Effect of bisacurone on ischemia-reperfusion injury (IRI)-induced alterations in electrocardiographic parameters. Representative images of electrocardiographic recordings from the sham (A), IRI control (B), IRI + diltiazem (10 mg/kg) (C), IRI + bisacurone (50 µg/kg) (D), IRI + bisacurone (100 µg/kg) (E), and bisacurone (100 µg/kg) (F) treated rats

Fig. 2. Effect of bisacurone on ischemia-reperfusion injury (IRI)-induced alterations in cardiac ANP, BNP, cTn-I, and HO-1 mRNA expression determined by relative quantification of reverse transcriptase-polymerase chain reaction analysis. (A) Quantitative representation of mRNA expression of ANP (B), BNP (C), cTn-I (D), and HO-1 (E) in rats. Data are expressed as mean [95% CI] (n = 6) and were analyzed by one-way ANOVA followed by Tukey’s multiple range tests. *p < 0.05 as compared to the IRI-control group, #p < 0.05 as compared to the sham, $p < 0.05 as compared to one another. Lane 1: Ladder 1000 bp; Lane 2: mRNA expression of sham group; Lane 3: mRNA expression of IRI control group; Lane 4: mRNA expression of IRI + Dil (10); Lane 5: mRNA expression of IRI + Bis (25); Lane 6: mRNA expression of IRI + Bis (50); Lane 7: mRNA expression of IRI + Bis (100) and Lane 8: mRNA expression of per se i.e., Bis (100) treated groups. IRI: Ischemia-reperfusion Injury control rats; Dil (10): diltiazem (10 mg/kg, p.o.) treated rats; Bis (25): bisacurone (25 µg/kg, p.o.); Bis (50): bisacurone (50 µg/kg, p.o.) and Bis (100): bisacurone (100 µg/kg, p.o.) treated rats. ANP: Atrial natriuretic peptide, BNP:…

Fig. 3. Effect of bisacurone on ischemia-reperfusion injury (IRI)-induced alterations in cardiac TNF-α, IL-1β, and TGF-β mRNA expressions determined by relative quantification of reverse transcriptase-polymerase chain reaction analysis (A), quantitative representation of protein expression of TNF-α (B), IL-1β (C), and TGF-β (D) in rats. Data are expressed as mean [95% CI] (n = 6). *p < 0.05 as compared to the IRI-control group, #p < 0.05 as compared to the sham, $p < 0.05 as compared to one another. Lane 1: Ladder 1000 bp; Lane 2: mRNA expression of sham group; Lane 3: mRNA expression of IRI control group; Lane 4: mRNA expression of IRI + Dil (10); Lane 5: mRNA expression of IRI + Bis (25); Lane 6: mRNA expression of IRI + Bis (50); Lane 7: mRNA expression of IRI + Bis (100) and Lane 8: mRNA expression of per se i.e., Bis (100) treated groups. IRI: Ischemia-reperfusion Injury control rats; Dil (10): diltiazem (10 mg/kg, p.o.) treated rats; Bis (25): bisacurone (25 µg/kg, p.o.); Bis (50): bisacurone (50 µg/kg, p.o.) and Bis (100): bisacurone (100 µg/kg, p.o.) treated rats. ILs: Interleukins; TNF-α: Tumor necrosis factor-alpha, TGF-β, Transforming growth factor beta

Fig. 4. Effect of bisacurone on ischemia-reperfusion injury (IRI)-induced alterations in cardiac Bax, Bcl-2, and Caspase-3 mRNA expression determined by relative quantification of reverse transcriptase-polymerase chain reaction analysis (A), and quantitative representation of protein expression of Bax (B), Bcl-2 (C), and Caspase-3 (D) in rats. Effect of bisacurone on ischemia-reperfusion injury (IRI)-induced alterations in the percentage of apoptotic cells in the cardiac tissue (E). A representative image of FACS analysis was obtained using Annexin V/FITCPI staining (F–J). Four distinct cell distribution patterns were observed: normal viable cells (lower left quadrant), apoptotic cells (lower right quadrant), late apoptotic or necrotic cells (upper right quadrant), and necrotic cells (upper left quadrant). Representative images of FACS analysis from sham (F), IRI control (G), IRI + diltiazem (10 mg/kg) (H), IRI + bisacurone (50 µg/kg) (I), and IRI + bisacurone (100 µg/kg) (J) treated rats. Data are expressed as mean [95% CI] (n = 6). *p < 0.05 as compared to the IRI-control group, #p < 0.05 as compared to the sham, $p < 0.05 as compared to one another. Lane 1: Ladder 1000 bp; Lane 2:…

Fig. 5. Effect of bisacurone on ischemia-reperfusion injury (IRI)-induced alterations in cardiac GRP78 (A) and CHOP (B) protein expression in rats. Data are expressed as mean [95% CI] (n = 6). *p < 0.05 as compared to the IRI-control group, #p < 0.05 as compared to the sham, $p < 0.05 as compared to one another. Lane 1: protein expression in the sham group; Lane 2: protein expression of IRI control group; Lane 3: protein expression of IRI + Dil (10); Lane 4: protein expression of IRI + Bis (25); Lane 5: protein expression of IRI + Bis (50); Lane 6: protein expression of IRI + Bis (100) and Lane 7: protein expression of per se i.e., Bis (100) treated groups. IRI: Ischemia-reperfusion Injury control rats; Dil (10): diltiazem (10 mg/kg, p.o.) treated rats; Bis (25): bisacurone (25 µg/kg, p.o.); Bis (50): bisacurone (50 µg/kg, p.o.) and Bis (100): bisacurone (100 µg/kg, p.o.) treated rats. GRP: Glucose-regulated protein, CHOP: C/-EBP homologous protein and GAPDH: Glyceraldehyde 3-phosphate dehydrogenase

Fig. 6. Effect of bisacurone on ischemia-reperfusion injury (IRI)-induced alterations in cardiac histopathology in rats. Photomicrographs of heart sections from sham (A), IRI control (B), IRI + diltiazem (10 mg/kg) (C), IRI + bisacurone (50 µg/kg) (D), IRI + bisacurone (100 µg/kg) (E), and bisacurone (100 µg/kg) (F)-treated rats. Images at 40X. Quantitative representation of the histological scores (G). Data are expressed as mean [95% CI] (n = 3), and one-way ANOVA followed by the Kruskal-Wallis test was applied for post hoc analysis. *p < 0.05 as compared to the IRI-control group, #p < 0.05 as compared to the sham, $p < 0.05 as compared to one another. Myocardial degeneration (black arrow) and interstitial inflammation (red arrow)

Fig. 7. Effect of bisacurone on ischemia-reperfusion injury (IRI)-induced alterations in cardiac ultrastructure in rats. Photomicrographs of sections of cardiac tissue from sham (A), IRI control (B), IRI + diltiazem (10 mg/kg) (C), and IRI + bisacurone (100 µg/kg) (D) treated rats at 11,580 X magnification