Figure 1.
The chemical structure of Astragaloside IV.
Figure 2.
The effect of post-ischemia treatment with Ast IV on the viability of SIR-injured cardiomyocytes.
Cardiomyocyte viability was assessed using the MTT assay. The results are expressed as the mean±SEM, n = 6. aaP<0.01 vs. Control; bP<0.05 vs. SIR; bbP<0.01 vs. SIR; cP<0.05 vs. SIR+Ast IV(12.5 µΜ); ccP<0.01 vs. SIR+Ast IV(12.5 µΜ); dP<0.05 vs. SIR+Ast IV(25 µΜ); eP<0.05 vs. SIR+Ast IV(50 µΜ); eeP<0.01 vs. SIR+Ast IV(50 µΜ). SIR, simulated ischemia reperfusion; Ast IV, Astragaloside IV.
Figure 3.
The effects of Ast IV and 2-MeOE2 post-ischemia treatment on cell viability, LDH release, and apoptotic index of SIR-injured cardiomyocytes.
(A). Cardiomyocyte viability was assessed using the MTT assay. (B). An ELISA assay was performed to detect the release of LDH in culture medium. (C). Representative flow cytometry apoptotic results are shown. Four subpopulations and their fractions are indicated: normal cells (lower left), dead cells (upper left), early apoptotic cells (lower right), and late apoptotic cells (upper right). The apoptotic index is expressed as the number of apoptotic cells/the total number of counted cells ×100%. The results are expressed as the mean±SEM, n = 6. aaP<0.01 vs. Control; bbP<0.01 vs. SIR; ccP<0.01 vs. SIR+Ast IV. SIR, simulated ischemia reperfusion; Ast IV, Astragaloside IV; 2-MeOE2, 2-methoxyestradiol; LDH, lactate dehydrogenase.
Figure 4.
The effects of Ast IV and 2-MeOE2 post-ischemia treatment on HIF-1α, iNOS, Bcl2, and Caspase3 protein expression in SIR-injured cardiomyocytes.
(A). Western blot image for HIF-1a and iNOS protein expression. (B). Western blot image for the expression of Bcl2 and Caspase3. The results are expressed as the mean±SEM, n = 6. aP<0.05 vs. Control; aaP<0.01 vs. Control; bbP<0.01 vs. SIR; ccP<0.01 vs. SIR+Ast IV. SIR, simulated ischemia reperfusion; Ast IV, Astragaloside IV; 2-MeOE2, 2-methoxyestradiol.
Figure 5.
The effects of Ast IV and 2-MeOE2 post-ischemia treatment on cardiac function of IR-injured isolated hearts.
(A). Representative line graph for the LVDP curve. (B). Representative line graph for the HR curve.(C). Representative line graph for the +dP/dt max curve. (D). Representative line graph for the CF curve. The results are expressed as the mean±SEM, n = 6. aP<0.05 vs. Control; aaP<0.01 vs. Control; bP<0.05 vs. IR; bbP<0.01 vs. IR; cP<0.05 vs. IR+Ast IV; ccP<0.01 vs. IR+Ast IV. IR, ischemia reperfusion; Ast IV, Astragaloside IV; 2-MeOE2, 2-methoxyestradiol; HR, heart rate; LVDP, left ventricular peak developing pressure; +dP/dt max, the maximum rate of pressure change in the ventricle; CF, coronary flow.
Figure 6.
The effects of Ast IV and 2-MeOE2 post-ischemia treatment on the infarct size, apoptotic index, and LDH release of IR-injured isolated hearts.
(A). Representative images of the myocardial infarct size are shown. The infarction size is expressed as the percentage of infarct relative to the mass at risk. (B). Representative images of apoptotic cardiomyocytes are shown. The apoptotic cells were detected by immunofluorescent staining with TUNEL (green)and DAPI (blue) staining was used to label the nuclei. (C). The amount of LDH was normalized against the dry weight of the heart and is expressed as IU/g. The results are expressed as the mean±SEM, n = 6. aaP<0.01 vs. Control; bbP<0.01 vs. IR; ccP<0.01 vs. IR+Ast IV. IR, ischemia reperfusion; Ast IV, Astragaloside IV; 2-MeOE2, 2-methoxyestradiol; LDH, lactate dehydrogenase.
Figure 7.
The effects of Ast IV and 2-MeOE2post-ischemia treatment on HIF-1α, iNOS, Bcl2, and Caspase3 protein expression in IR-injured isolated hearts.
(A). Representative images of HIF-1α and iNOS expression are shown. (B). Representative images of Bcl2 and Caspase3 expression are shown. The results are expressed as the mean±SEM, n = 6. aaP<0.01 vs. Control; bbP<0.01 vs. IR; ccP<0.01 vs. IR+Ast IV. IR, ischemia reperfusion; Ast IV, Astragaloside IV; 2-MeOE2, 2-methoxyestradiol.
Figure 8.
The effects of Ast IV and HIF-1α siRNA post-ischemia treatment on SIR-injured cardiomyocytes.
(A) Cardiomyocyte viability was assessed using the MTT assay. (B) Representative flow cytometry apoptotic results are shown. Four subpopulations and their fractions are indicated: normal cells (lower left), dead cells (upper left), early apoptotic cells (lower right), and late apoptotic cells (upper right). The apoptotic index is expressed as the number of apoptotic cells/the total number of counted cells ×100%. (C) Representative images of HIF-1α and iNOS protein expression are shown. The results are expressed as the mean±SEM, n = 6. aaP<0.01 vs. Control; bbP<0.01 vs. SIR+Con siRNA; ccP<0.01 vs. SIR+Con siRNA+Ast IV. ddP<0.01 vs. SIR+HIF-1α siRNA+Ast IV. SIR, simulated ischemia reperfusion; Ast IV, Astragaloside IV.