Application of hydrazine curcumin in the preparation of anti-squamous cell carcinoma drugs
A technology of hydrazine curcumin and squamous cell carcinoma, which is applied in the field of medicine, can solve the problem that curcumin cannot be widely used in clinical practice, and achieve high bioavailability
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Embodiment 1
[0020] MTT method to detect the lethality of hydrazine curcumin on cells:
[0021] Take the cells in the logarithmic growth phase, press 4×10 3 Each well was inoculated in a 96-well plate, and sterile PBS was added to the edge wells. After culturing for 24 hours, hydrazine curcumin was added at a concentration of 5, 10, 15, 20, 25, 30, 35, 40, and 50 μmol / L, respectively. A negative control group (0.1% dimethyl sulfoxide DMSO) and a blank control group were set up, and 3 replicate wells were set up in each group. 0.1% DMSO could inhibit the proliferation of human skin squamous cell carcinoma A431 cells. After culturing for 24, 48, and 72 hours, centrifuge at 1000r / min, discard the culture medium, add 180μl of serum-free RPMI1640 medium and 20μl of MTT solution (5mg / ml) to each well, continue to cultivate for 4h, centrifuge at 1000r / min, and discard the supernatant , add 100 μl DMSO to each well, shake for 10 minutes. Measure the optical density A value at a wavelength of 490...
Embodiment 2
[0029] Transwell chamber invasion assay was used to detect the effect of hydrazine curcumin on cell invasion ability. The logarithmic growth phase cells were divided into the following four groups: negative control group (0.1% DMSO); hydrazinocurcumin 5 μmol / L treatment group; hydrazinocurcumin 10 μmol / L treatment group; hydrazinocurcumin 15 μmol / L treatment group .
[0030] The results are attached figure 1 Shown, the control group and 5,10,15 μ mol / L hydrazine curcumin treatment group penetrating cell numbers are respectively (462.33±36.71) / visual field, (310.33±37.66) / visual field, (189.10±17.06) / visual field and ( 126.33±8.62) / field of view. Compared with the control group, the number of transmembrane cells gradually decreased with the increase of the concentration of hydrazinocurcumin, and the experimental effect was the most obvious in the 15 μmol / L group, and the differences among the experimental groups were statistically significant (F=85.67, P﹤0.05).
Embodiment 3
[0032] Adhesion test was used to detect the effect of hydrazine curcumin on cell adhesion ability. According to the four groups classified above, the cells were pretreated for 24 hours. Prepare a fibronectin solution with a final concentration of 10 μg / ml, coat 96-well cells with 50 μl per well on a culture plate, seal and seal for 12 hours at 4°C; wash 5 times with PBS, add serum-free RPMI1640 medium containing 1% BSA, 50 μl / well, block at 37°C for 2 hours; digest the cells with 0.25% trypsin, and adjust the cell concentration to 2×10 4 Each well was inoculated in a 96-well plate, and three replicate wells were set for each group, and incubated for 1.5 h; washed 5 times with PBS, added 2 mg / ml MTT, 50 μl / well, and incubated for 3 h; discarded MTT, added 150 μl of DMSO per well, Shake for 15 minutes, measure the A value at the 490nm wavelength of the microplate reader, and calculate the amount of adhered cells according to the following formula:
[0033] Amount of adherent c...
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