Application of Sulfated Mannoglucuronide Hexasaccharide in Prevention and Treatment of Atherosclerosis
A technology for atherosclerosis and uronic acid hexasaccharide, applied in the prevention and treatment of atherosclerosis, in the field of sulfated mannose glucuronic acid hexasaccharide, which can solve the problems such as the application of sulfated mannose glucuronic acid oligosaccharide , to achieve the effect of inhibiting absorption effect and small toxic side effects
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Embodiment 1
[0056]Example 1 Preparation of sulfated mannose glucuronic acid hexaose
[0057]1. Preparation of mannose glucuronic acid oligosaccharides
[0058]The dried kelp was extracted with 30 times the mass of distilled water at 100°C for 3 hours. The extract was filtered and concentrated. After concentration, ethanol was added to the final concentration of 75% to precipitate. After standing for 12 hours, the precipitate was collected, and then dried in vacuum to obtain kelp polysaccharide. . The kelp polysaccharide sample was dissolved in a sulfuric acid solution with a mass concentration of 4% (the material-to-liquid ratio was 60mg / mL) and heated to reflux for 5 hours, neutralized with barium hydroxide to pH=6-7, centrifuged, and the supernatant was concentrated to the original One-fifth of the volume, the concentrate is chromatographed on activated carbon column, first equilibrated with distilled water, and then eluted with a gradient of 50%-90% ethanol, and the 50%-90% ethanol eluate is conce...
Embodiment 2
[0065]Example 2 Regulation of Sulfated Mannoglucuronic Acid Hexose on Inflammatory Factors
[0066]Inoculate THP-1 cells in a 6-well plate (4×105Cells / well, RPMI-1640 complete culture medium containing 10% FBS), add phorbol ester (PMA) at a final concentration of 100ng / ml, and continue to culture for 48h to induce monocytes to differentiate into macrophages. The experimental group was added 0.25mg / ml, 0.5mg / ml, 1mg / ml sulfated mannoglucuronic acid hexaose. Incubate for 48 hours, collect the culture solution, and remove the precipitate at 4000 rpm for 5 min. ELISA kit detects the release levels of TNF-α and IL-1β.
[0067]The result isfigure 1 As shown, sulfated mannose glucuronide hexaose at a concentration of 0.5 mg / ml can significantly reduce the inflammatory factor TNF-α (figure 1 A) and IL-1β (figure 1 The expression level of B) is increased to 1 mg / ml, and the effect of reducing the expression level of inflammatory factors is more obvious than that of the 0.5 mg / ml group, showing a c...
Embodiment 3
[0068]Example 3 Inhibitory regulation of sulfated mannoglucuronic acid hexaose on NF-κB signaling pathway
[0069]Inoculate THP-1 cells in a 60mm culture dish (2×106Cells / well, RPMI-1640 complete culture medium containing 10% FBS), add phorbol ester (PMA) at a final concentration of 100ng / ml, and continue to culture for 48h to induce monocytes to differentiate into macrophages. The experimental group was added 0.5mg / ml and 1mg / ml sulfated mannose glucuronic acid hexaose. Incubate for 48 hours, remove the medium, wash with PBS, add 100 μL of RIPA lysis buffer containing protease and phosphatase inhibitors, scrape the cells with a cell scraper, and transfer the cell lysate to a 1.5 mL EP tube to lyse the cells on ice for 30 minutes; 4 Centrifuge at 13000 rpm for 15 min at ℃, and transfer the supernatant to a new 1.5 mL EP tube. Use the BCA protein concentration determination kit for concentration determination, and store in -80℃ refrigerator. Western blot was used to detect the molecular...
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