A kind of thermosensitive gel material combination and its preparation method and application
A temperature-sensitive gel and gel technology, applied in the field of regenerative medicine, can solve the problems of increased cost and high requirements of 3D printing technology, and achieve the effect of promoting regeneration
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[0042] The present invention also provides a preparation method of the temperature-sensitive gel material combination, comprising the following steps:
[0043] (1) mixing chitosan with acetic acid solution to obtain chitosan solution;
[0044] (2) β-glycerophosphate disodium and hyaluronic acid are mixed and dissolved in water to obtain a hyaluronic acid solution;
[0045] (3) mixing chitosan solution and hyaluronic acid solution to obtain chitosan hyaluronic acid solution;
[0046] (4) mixing SDF-1 with a 40-60% volume chitosan hyaluronic acid solution to obtain a chitosan hyaluronic acid solution loaded with SDF-1;
[0047] (5) Mix 40-60% volume chitosan hyaluronic acid solution with 1-ethyl-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide, then add Apt19S aptamer to obtain Apt19S-loaded chitosan hyaluronic acid solution.
[0048] In the present invention, chitosan is mixed with acetic acid solution to obtain chitosan solution.
[0049] In the present inventio...
Embodiment 1
[0092] 48 mg of chitosan (CS) was dissolved in 1.6 mL of 0.1 M acetic acid solution, placed on a magnetic stirrer and stirred continuously for 2 h (1000 rpm), and sterilized by autoclaving at 121° C. for 10 min to obtain the chitosan solution. 240 mg of β-GP and 4 mg of hyaluronic acid were dissolved in 0.4 mL of deionized water, and sterilized by filtration with a 0.22 μm filter to obtain a dissolved hyaluronic acid solution. Chitosan solution and hyaluronic acid solution were ice-bathed (0°C) for 15min, then hyaluronic acid was added dropwise to the chitosan solution and stirred with a magnetic stirrer for 10min (1000rpm) to obtain a chitosan hyaluronic acid solution . Divide the chitosan hyaluronic acid solution into two equal parts, 1 mL each.
[0093] Take 1 mL of chitosan hyaluronic acid solution, dropwise add 1 μl of SDF-1 solution with a concentration of 100 μg / mL dissolved in PBS, and stir magnetically for 10 min (1000 rpm) to obtain a loaded SDF with a SDF-1 concent...
Embodiment 2
[0098] 100 μL of the third-generation rBMSCs culture medium (containing 1×10 3 cells) were seeded into the upper chamber of the Transwell chamber, and 600 μL of α-MEM medium containing 200 ng / mL SDF-1 and 600 μL of α-MEM medium containing amino-modified Apt19S with a concentration of 2 nmol / mL were added to the lower chamber of Transwell. at 37°C, CO 2 After culturing in an incubator with a concentration of 5% for 24 hours, the unmigrated cells in the upper chamber were wiped off with a cotton swab. The cells that migrated to the lower chamber were fixed with 4% formaldehyde for 15 minutes, rinsed twice with PBS, air-dried, and 0.5% crystal violet for 30 minutes. Rinse 3 times with PBS. Then, 5 fields of view were randomly selected from each chamber under the microscope, and the BMSC cells that migrated to the lower chamber were counted. The results showed that the addition of SDF-1 and amino-modified Apt19S aptamer could significantly promote the migration of BMSC cells com...
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