Application of Glucosamine in Preparation of Radiotherapy Sensitizer
A technology of glucosamine and drugs, applied in the preparation of radiotherapy sensitization drugs, the new application field of glucosamine, which can solve the problems such as no reports of glucosamine radiotherapy sensitization
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Embodiment 1
[0029] (1) Cell culture: A549 cells and LLC cells were cultured in DMEM medium containing 10% fetal bovine serum; BEAS-2B cells were cultured in RMPI medium containing 10% fetal bovine serum. All cells were placed at 37°C, 5% CO 2 Cultured in an incubator, subcultured every 2-3 days, and the cells in the logarithmic growth phase were used for experiments.
[0030] (2) CCK-8 colorimetric method: cells in logarithmic growth phase (5*10 4 / mL) were inoculated into 96-well plates, and 6 parallel wells were set up for each concentration. Different concentrations of glucosamine were added to continue culturing for 24 and 48 hours. Add 10 μL CCK-8 reagent to the cell culture medium and continue culturing for 1-4 hours. The absorbance at 450 nm was measured using a microplate reader. Finally, the formula of cell survival rate=dosage group value / control group value×100% was used to calculate the cell survival rate.
[0031] The result is as figure 1 As shown, the results show tha...
Embodiment 2
[0033] (1) Cell culture is the same as in Example 1;
[0034] (2) Cell clone formation method: take A549 cells in the logarithmic growth phase, and inoculate different numbers of cells into six-well plates according to different irradiation dose requirements (0, 2, 4, 8Gy doses of 200, 400, 800 and 1600 cells were inoculated respectively ). After 24 hours, the glucosamine combined with radiation group was treated with 5 mmol / L glucosamine for 1 hour, and the control group was treated with the same amount of PBS as a negative control. The two groups of cells were irradiated with different doses (0-8Gy) of γ-rays at the same time, and continued to culture for 24 hours after irradiation. Both groups were replaced with ordinary medium, and continued to stop the culture until obvious cell clones visible to the naked eye appeared in the culture dish. . Discard the medium, wash twice with PBS, fix with anhydrous methanol for 30 minutes, stain with Gimsa stain for 30 minutes, rinse ...
Embodiment 3
[0036] (1) Cell culture is the same as in Example 1;
[0037] (2) Cell apoptosis detection by flow cytometry: A549 cells and BEAS-2B cells (1*10 5 / mL) into six-well plates, add 5mmol / L glucosamine to treat A549 cells and BEAS-2B cells, then give the cells 8Gy of 60Coγ-ray irradiation, and then continue to culture for 24h, use EDTA-free trypsin to digest the cells After being centrifuged at 1500 rpm for 5 minutes, the cells were washed three times with PBS, and cell apoptosis was detected by flow cytometry after double staining with Annexin V-FITC and PI. The result is as image 3 Shown, the glucosamine (5mmol / L) treatment of non-toxic side effect concentration can obviously increase the radiotherapy sensitivity of A549 cell, but has no radiosensitization effect to BEAS-2B cell: After 5mmol / L glucosamine treats A549 cell, Radiation-induced apoptosis increased by 13%; after 5mmol / L glucosamine treated BEAS-2B cells, radiation-induced apoptosis decreased by 2%. These research...
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