Formononetin regulates endothelial nitric oxide synthase to protect vascular endothelium in deep vein thrombosis rats

Figures

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  Figure 1.. Effects of formononetin on IVC thrombosis in DVT rats. (a) The images, lengths and weights of IVC thrombosis in each group. (b) The pathological changes of venous thrombosis in each group were analyzed by H&E (scale = 200 μm), which showed longitudinal sections. The arrow represents the thrombosis. Compared with the DVT group, ##p < 0.01. Compared with the 10 mg/kg formononetin group, &&p < 0.01.

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  Figure 2.. Effects of formononetin on inflammatory factors and coagulation factors in DVT rats. (a) The contents of IL-1β and IL-18 in serum analyzed by ELISA. (b) The contents of D-dimer, F1 + 2, TM, and TF in plasma analyzed by ELISA. Compared to the sham group, *p < 0.05, **p < 0.01. Compared to the DVT group, #p < 0.05, ##p < 0.01. Compared to the 10 mg/kg formononetin group, &p < 0.05, &&p < 0.01.

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  Figure 3.. Effects of formononetin on p-eNOS expression in the venous endothelial cells of DVT rats. The expression of eNOS was analyzed by immunofluorescence ((a), yellow) and western blot (b). Scale = 50 μm. The results were analyzed by Image J software. Compared to the sham group, *p < 0.05, **p < 0.01. Compared to the DVT group, #p < 0.05, ##p < 0.01. Compared to the 10 mg/kg formononetin group, &p < 0.05, &&p < 0.01. The activity of formononetin to eNOS was predicted by the molecular docking analysis (c).

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  Figure 4.. Formononetin improved DVT via regulating the p-eNOS in rats. (a) The images, lengths and weights of IVC thrombosis in each group. (b) The pathological changes of venous thrombosis in each group were analyzed by H&E (scale = 200 μm), which showed longitudinal sections. 40 mg/kg formononetin and 40 mg/kg NOS inhibitor L-NAME were orally treated after surgery. Compared to the sham group, **p < 0.01. Compared to the DVT group, ##p < 0.01. Compared to the formononetin group, &p < 0.05.

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  Figure 5.. Formononetin decreased NF-κB pathway via regulating the p-eNOS in the DVT rats. (a) The expression of p-eNOS was measured by Immunofluorescence ((a), yellow) and western blot (b). Scale = 20 μm. (b) Western blot was used to analyze the expression of ICAM-1, Iκκβ and p-NF-κB p65/NF-κB p65. Compared to the sham group, *p < 0.05, **p < 0.01. Compared to the DVT group, #p < 0.05, ##p < 0.01. Compared to the formononetin group, &&p < 0.01.

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  Figure 6.. Gender-related differences in DVT rats after formononetin (40 mg/kg) treatment. (a) The differences of thrombus length and weights between male and female rats. (b) The difference of levels of IL-1β and IL-18 in serum between male and female rats. (c) The difference of p-eNOS/eNOS protein expression between male and female rats was analyzed by western blot. No difference between male and female was found. Male = 8 and female = 8.