Icariin Inhibits Pulmonary Hypertension Induced by Monocrotaline through Enhancement of NO/cGMP Signaling Pathway in Rats

Abstract

It has been reported that icariin (ICA) increased contents of nitric oxide (NO) and cyclic guanosine monophosphate (cGMP) by improving expression of endothelial nitric oxide synthase (eNOS) and inhibition of phosphodiesterase type 5 (PDE5). In addition, dysfunction of the NO/cGMP pathway may play a crucial role in the pathogenesis of pulmonary hypertension (PH). In this study, the potential protective effects of ICA on PH induced by monocrotaline (MCT, 50 mg/kg) singly subcutaneous injection were investigated and the possible mechanisms involved in NO/cGMP pathway were explored in male Sprague Dawley rats. The results showed that ICA (20, 40, and 80 mg/kg/d) treatment by intragastric administration could significantly ameliorate PH and upregulate the expression of eNOS gene and downregulate the expression of PDE5 gene in MCT-treated rats. Both ICA (40 mg/kg/d) and L-arginine (200 mg/kg/d), a precursor of NO as positive control, notably increased the contents of NO and cGMP in lung tissue homogenate, which were inversed by treatment with (N) G-nitro-L-arginine-methyl ester (L-NAME), a NOS inhibitor, and L-NAME-treatment could also inhibit the protective effects of ICA (40 mg/kg/d) on mean pulmonary artery pressure and artery remodeling and tends to inhibit right ventricle hypertrophy index. In summary, ICA is effective in protecting against MCT-induced PH in rats through enhancement of NO/cGMP signaling pathway in rats.

Figure 1

Effect of ICA on survival rates. Dose: mg/kg/d. Mortality was observed daily and the figure shows the survival rate at each time point, setting the survival rate at the start of model established as 100%. In order to easily read, the survival rates data of L-arg treatment groups, L-arg (200 mg/kg/d) with L-NAME (20 mg/kg/d) group, and ICA (40 mg/kg/d) with L-NAME (20 mg/kg/d) group are not presented in Figure 1. Compared with control group ^# p < 0.05, compared with model group ^∗ p < 0.05, Log-Rank test.

Figure 2

Representative pulmonary arterial pressure curve. Groups: (a) control; (b) model; (c) ICA (20 mg/kg/d); (d) ICA (40 mg/kg/d); (e) ICA (80 mg/kg/d); (f) SIL (25 mg/kg/d); (g) L-arg (200 mg/kg/d); (h) ICA (40 mg/kg/d) combining with L-arg (200 mg/kg/d); (i) L-NAME combining with L-arg (200 mg/kg/d).

Figure 3

Effects of ICA treatment on mPAP (a) and RVHI (b) (mean ± SD, n = 5–12). Dose: mg/kg/d. mPAP: mean pulmonary artery pressure; RVHI: right ventricular hypertrophy index. Compared with control group ^# p < 0.01; compared with model group ^∗ p < 0.05; compared with ICA (40 mg/kg/d) group ^△ p < 0.05; compared with L-arg (200 mg/kg/d) group ^▲ p < 0.05.

Figure 4

Effect of ICA on lung vascular remodeling induced by MCT in rats (mean ± SD, n = 5–12). Dose: mg/kg/d. (a)~(i) Representative images of H&E staining lung sections from every group rats. (j) Bar graph of % pulmonary artery wall area (diameter: 50 to 100 μm). Groups: (a) control; (b) model; (c) ICA (20 mg/kg/d); (d) ICA (40 mg/kg/d); (e) ICA (80 mg/kg/d); (f) SIL (25 mg/kg/d); (g) L-arg (200 mg/kg/d); (h) ICA (40 mg/kg/d) combining with L-arg (200 mg/kg/d); (i) L-NAME combining with L-arg (200 mg/kg/d). Compared with control group ^# p < 0.05; compared with model group ^∗ p < 0.05; compared with ICA (40 mg/kg/d) group ^△ p < 0.05; compared with L-arg (200 mg/kg/d) group ^▲ p < 0.05.

Figure 5

Effect of ICA on arterial blood partial pressure of oxygen and carbon dioxide (mean ± SD, n = 5–12). Dose: mg/kg/d. PaO₂: arterial blood partial pressure of oxygen; PaCO₂: arterial blood partial pressure of carbon dioxide. Compared with control group ^# p < 0.05; compared with model group ^∗ p < 0.05.

Figure 6

ICA enhances function of NO/cGMP signaling pathway in MCT-PH model rats (mean ± SD, n = 5–12). Dose: mg/kg/d. (a) Representative Western blots from lung homogenate; (b) quantitative bar graph of eNOS expression of lung tissue; (c) quantitative bar graph of PDE5 expression of lung tissue. (d) Bar graph of eNOS and PDE5 mRNA expression; (e) bar graph of NO content of lung tissue; (f) bar graph of cGMP content of lung tissue; compared with control group ^# p < 0.05; compared with model group ^∗ p < 0.05; compared with ICA (40 mg/kg/d) group ^△ p < 0.05; compared with L-arg (200 mg/kg/d) group ^▲ p < 0.05.