Figure 1.
The molecular structure of quercetin.
Table 1.
Primers sequences and real-time PCR amplification parameters.
Figure 2.
Effects of quercetin on systolic blood pressure measured by tail-cuff plethysmography in the four groups.
The characteristics of the systolic blood pressures from week 1 to week 12 in the four groups are shown in Figure 2. Each value represents the mean ± sem (n = 8 in each group). *P<0.05, **P<0.01, vs. the spontaneously hypertensive rats (SHR) group; #P<0.05, vs. the SHR+QH group.
Figure 3.
Effects of quercetin on left ventricular weight index (LVW/BW) in the different treatment group.
After 12 weeks treatment, the animals were anesthetized and the left ventricular weight (LVW) and body weight (BW) ratio was determined. Each value represents the mean ± sem (n = 8 in each group). *P<0.05, vs. the Wistar–Kyoto rats (WKY) group; **P<0.05, vs. the spontaneously hypertensive rats (SHR) group; #P<0.05, vs. the SHR+QH group.
Figure 4.
Histopathological analysis reveals that quercetin suppresses myocardial hypertrophy in spontaneously hypertensive rats.
Tissue sections were stained with hematoxylin and eosin (H&E; a1–a4; ×400) and Masson's trichrome (b1–b4, ×400). (c) Mean cardiomyocyte diameter was determined by measuring 100 cells from each section. (d) The ratio of the area of myocardial fibrosis (excluding perivascular collagen) to the total myocardial area (percentage) was quantified from images of Masson's trichrome-stained tissue. The scale bar denotes 100 µm. Both mean cardiomyocyte diameter and collagen volume were quantified using Image-Pro Plus software. At least 8–10 areas were evaluated in each section and each value denotes the mean ± sem (n = 8 in each group). CVF, collagen volume fraction. *P<0.05, vs. the Wistar–Kyoto rats (WKY) group; **P<0.05, vs. the spontaneously hypertensive rats (SHR) group; #P<0.05, vs. the SHR+QH group.
Figure 5.
Transmission electron microscopic observations of myocardial tissue of rats in the different treatment group.
Ultrastructure of cardiac sections (a1–a4; ×8000). (a1) Observations of myocardial tissue of the WKY vehicle-treated group, which showed normal myocardial tissue. (a2) Myocardial tissue in the SHR group. Swollen mitochondria, deposition of a large number of dense granules in the mitochondria and cellular necrosis was observed. (a3, a4) Myocardial tissue of the quercetin-treated SHR groups. The cardiomyocyte mitochondria were less swollen, the number of dense granules was lower and the structure was relatively better organized compared with the SHR group. Arrows indicate obviously swollen and loosely arranged mitochondria.
Figure 6.
Quercetin administration improves cardiac function in spontaneously hypertensive rats.
Representative M-mode echocardiogram showing improved wall motion after treatment (a–d). After 12 weeks of treatment with quercetin, the left ventricular end-diastolic internal diameter (LVIDd) improved and the left ventricular end-diastolic posterior wall thickness (LVPWd), and end-diastolic interventricular septal thickness (IVSd) differences were attenuated. Data are the mean ± sem (n = 8 for each group). *P<0.05, vs. the Wistar–Kyoto rats (WKY) group ; #P<0.05, vs. the spontaneously hypertensive rats (SHR) group.
Table 2.
Echocardiographic data from the four treatment groups at week 1 and week12.
Figure 7.
Quercetin administration activates protein expression of PPAR-γ and inhibits protein expression of AP-1 in cardiomyocyte.
Figure (a2) shows few cells with nuclei positive for PPAR-γ in spontaneously hypertensive rat cardiomyocytes. However, SHRs (a3–a4) treated with quercetin showed significantly more positive nuclear staining that followed a dose-dependent manner. Compared with Wistar–Kyoto (WKY) rats (b1, c1), SHRs (b2, c2) treated with vehicle showed a large number of positively stained cardiomyocyte nuclei. In contrast, SHRs treated with quercetin (b3–b4, c3–c4) had significantly and dose-dependently fewer positively stained cardiomyocyte nuclei. The scale bar denotes 100 µm (×400). (b–d) At least 8–10 areas were evaluated in each section and were quantified using Image-Pro Plus software. Arrows indicate positive nuclear staining. Results are shown as the mean ± sem. *P<0.05, vs. the Wistar–Kyoto rats (WKY) group; **P<0.05, vs. the spontaneously hypertensive rats (SHR) group; #P<0.05, vs. the SHR+QH group.
Figure 8.
Quercetin enhances PPAR-γ mRNA expression and suppresses AP-1 mRNA and BNP and ANP mRNA levels after treatment.
After 12 weeks of treatment with quercetin, the myocardial tissues mRNA level were analysis by real-time polymerase chain reaction. Values denote the expression level relative to the Wistar–Kyoto (WKY) group (n = 8 per group). All samples were analyzed in triplicate and are expressed as the mean ± sem. *P<0.05, vs. the Wistar–Kyoto rats (WKY) group; **P<0.05, vs. the spontaneously hypertensive rats (SHR) group; #P<0.05, vs. the SHR+QH group.
Figure 9.
Western blotting reveals that quercetin enhances PPAR-γ protein expression and suppresses AP-1 protein expression.
After 12 weeks of treatment with quercetin, the myocardial tissues protein expression were analysis by Western blotting. All samples were analyzed in triplicate and expressed as the mean ± sem. *P<0.05, vs. the Wistar–Kyoto rats (WKY) group; **P<0.05, vs. the spontaneously hypertensive rats (SHR) group; #P<0.05, vs. the SHR+QH group.
Figure 10.
Inhibitory effects of quercetin on Ang II–stimulated [3H]leucine (1 µCi/ml) incorporation into H9C2 cells.
H9C2 cells were pretreated with different concentrations of quercetin for 30[3H]leucine(1 µCi/ml) for 48 h. All values were expressed as a percentage of incorporation in the comparison to Ang II treated group. Data are mean ± sem and each experiment performed in triplicate independently. ** P<0.01, vs Ang II-treated group.
Figure 11.
AngII induced hypertrophic growth of H9C2 cells was inhibited by quercetin.
After treated with quercetin (100 µg/ml) and AngII (200 µM) alone or in combination for 48 h (a), H9C2 cells hypertrophy was analyzed by measuring cell surface area using Image Pro Plus software 7.0. (b). Ang II (200 µM) caused an increase in H9C2 cells size and was attenuated by quercetin (100 µg/mL). The values denote the relative area ± sem (n = 3). **P<0.01, vs. the Ang II-treated group.
Figure 12.
Cells treated with quercetin enhances PPAR-γ nuclear immunofluorescence and suppresses AP-1 nuclear immunofluorescence activity.
After H9C2 cells were treated with quercetin (100 µg/ml) and AngII (200 µM) or in combination for 24 hours, (a) Coverslips were mounted on a slide and processed for fluorescence (×400), then analyzed using Image Pro Plus 7.0. Green: PPAR-γ; Red: c-fos and c-jun; Blue: nuclei (DAPI). Results are representative images of cells from three independent experiments. (b–d) The protein levels of PPAR-γ, c-fos and c-jun were quantified by densitometry, and the data denote the relative intensity compared with control cells. The values denote the relative area ± sem (n = 3). **P<0.01, vs. the Ang II-treated group.
Figure 13.
Cells transfected with PPAR-γ siRNA showed a reversal of the quercetin inhibition on AP-1 protein expression.
H9C2 cells were transiently transfected with siRNA targeted to PPAR-γ and this siRNA led to decrease in protein levels of PPAR-γ compared with NCsiRNA and untreated controls (P<0.01; a, c). After H9C2 cells were transfected with PPAR-γ NCsiRNA or PPAR-γ siRNA (200 nM) for 24 h, cells were incubated with quercetin and Ang II alone or in combination for 24 h. Cells were incubated for a further 48 h prior to isolating protein samples for c-fos and c-jun western blotting analysis (b, d). All samples were analyzed in triplicate, and data are expressed as the mean ± sem. **P<0.01, vs. Ang II-treated group.
Figure 14.
Transfection with PPAR-γ siRNA reversed the quercetin-mediated inhibition of ANP and BNP mRNA expression.
After H9C2 cells were transfected with each siRNA (200 nM) for 24 h, cells were treated with quercetin and AngII alone or in combination for 24 h. Cells were incubated for a further 24 h before RNA isolation for analysis of ANP and BNP mRNA by real-time PCR (a–b). Data are expressed as the mean ± sem. Values are the means from three independent experiments. **P<0.01, vs. Ang II-treated group.