Antarctic Krill Oil from Euphausia superba Ameliorates Carrageenan-Induced Thrombosis in a Mouse Model

Figures

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  Figure 1. Effects of FJH-KO on carrageenan-induced thrombosis in mice. (A) Schematic diagram of carrageenan-induced thrombosis model with FJH-KO and aspirin treatment. (B) Thrombus length, (C) thrombosis rate (thrombosis length/whole tail length), and (D) bleeding time (s) in the tail of carrageenan-induced thrombosis mice. * p < 0.05 vs. the vehicle control group, # p < 0.05 vs. the carrageenan model group (n = 8).

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  Figure 2. Effect of orally administered FJH-KO on the plasma levels of (A) thromboxane B2 (TXB2), (B) 6-keto prostaglandin F1α (6-keto PGF1α), (C) P-selectin, (D) endothelin-1 (ET-1), (E) β-thromboglobulin (β-TG), (F) platelet factor 4 (PF4), (G) serotonin, and (H) plasminogen in carrageenan-induced thrombosis model mice. * p < 0.05 vs. the vehicle control group, # p < 0.05 vs. the carrageenan model group (n = 8).

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  Figure 3. Effect of orally administered FJH-KO on the plasma levels of (A) tumor necrosis factor alpha (TNF-α), (B) interleukin (IL)-1β, and (C) IL-6 in carrageenan-induced thrombosis model mice. * p < 0.05 vs. the vehicle control group, # p < 0.05 vs. the carrageenan model group (n = 8).

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  Figure 4. Effect of FJH-KO on the expression of blood clot-related mRNAs in carrageenan-induced mouse blood vessels. (A) Endothelial nitric oxide synthase (eNOS), (B) intercellular adhesion molecule 1 (ICAM-1), (C) vascular cell adhesion molecule 1 (VCAM-1), (D) P-selectin, (E) E-selectin, (F) monocyte chemoattractant protein 1 (MCP-1), (G) TNF-α, (H) IL-1β, and (I) IL-6. * p < 0.05 vs. the vehicle control group, # p < 0.05 vs. the carrageenan model group (n = 8).

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  Figure 5. Effects of FJH-KO on carrageenan-induced thrombosis in mouse tissues. (A) Hematoxylin and eosin (H&E) staining of the tail 2, 4, and 6 cm from the tail tip (Scale bar: 200 μm). Vessels and thrombi are represented by blue and red dotted lines, respectively. (B) H&E staining of lung tissue (Scale bar: 200 μm). Venous thrombosis is represented by black arrows. (C) H&E staining of liver tissue (Scale bar: 200 μm). Hepatic vein thrombosis is represented by blue arrows.

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  Figure 6. Effect of FJH-KO on eNOS phosphorylation and nitric oxide (NO) production in EA.hy926 cells. (A) Cells were treated with 10–300 μg/mL FJH-KO for 24 h. Cell viability and cytotoxicity were determined using MTT and LDH assays, respectively. (B) Cells were treated with 10–100 μg/mL FJH-KO for 3 h. NO generation was detected using DAF-2DA (Scale bar: 200 μm). (C,D) Cells were treated with 100 μg/mL FJH-KO for 1–6 h or 10–100 μg/mL FJH-KO for 3 h and assessed via Western blotting. (E) Cells were pretreated with 100 μM L-NAME for 1 h and were then treated with 100 μg/mL FJH-KO for 3 h. NO generation was detected using DAF-2DA (Scale bar: 200 μm).

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  Figure 7. Effect of FJH-KO on NO production and THP-1 monocyte adhesion in HUVEC cells. (A) Cells were treated with 10–100 μg/mL FJH-KO for 3 h. NO generation was detected using DAF-2DA (Scale bar: 200 μm). (B) Cells were treated with 10–100 μg/mL FJH-KO for 3 h and assessed via Western blotting. (C) Cells were pretreated with 100 μM L-NAME for 1 h and were then treated with 100 μg/mL FJH-KO for 3 h. NO generation was detected using DAF-2DA (Scale bar: 200 μm). (D) Cells were treated with 10–100 μg/mL of FJH-KO for 3 h, followed by incubation with 10 ng/mL TNF-α for 12 h. Endothelial cells were co-cultured with THP-1 cells for 1 h, and the adherence of endothelial cells to monocytes was assessed via fluorescence microscopy (Scale bar: 400 μm). All experiments were performed in triplicate. Data are represented as the mean ± standard deviation. * p < 0.01 vs. control cells, # p < 0.01 vs. FJH-KO-treated cells.

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  Figure 8. Effects of FJH-KO on TNF-α-induced THP-1 monocyte adhesion in EA.hy926 cells. (A) Cells were treated with 10–100 μg/mL FJH-KO for 3 h, followed by incubation with 10 ng/mL TNF-α for 12 h. Endothelial cells were co-cultured with THP-1 cells for 1 h, and the adherence of endothelial cells to monocytes was assessed via fluorescence microscopy (Scale bar: 400 μm). (B,C) Cells were pretreated with 100 μg/mL FJH-KO for 3 h, followed by 100 ng/mL TNF-α for 12 h. mRNA levels of (B) ICAM-1 and (C) VCAM-1 were measured using RT-PCR. (D,E) Cells were pretreated with 100 μM L-NAME for 1 h and then treated with 100 μg/mL FJH-KO for 3 h, followed by incubation with 10 ng/mL TNF-α for 12 h. mRNA levels of (D) ICAM-1 and (E) VCAM-1 were measured using RT-PCR. (F) Cells were pretreated with 10–100 μg/mL FJH-KO for 3 h and were then treated with 10 ng/mL TNF-α for 1 h. (G) Cells were pretreated with 100 μM L-NAME for 1 h and were then treated with 100 μg/mL FJH-KO for 3 h, followed by incubation with 10 ng/mL TNF-α for 1 h. NF-κB phosphorylation was assessed using Western blotting. (H) Cells were pretreated with 100 μM L-NAME for 1 h and were then treated with 100 μg/mL FJH-KO for 3 h, followed by…

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  Figure 9. Schematic diagram of the protective effect of FJH-KO on thrombosis by regulating endothelial dysfunction and inflammation.