Preparation method of FGF1 sericin protein gel with activity for promoting cell proliferation and product thereof

A technology of sericin and cell proliferation, applied in medical science, prostheses, etc., can solve the problems of weak cell adhesion, increased production cost, few products that have obtained clinical approval and finalization, and achieves strong water absorption performance and simple method Effect

Active Publication Date: 2018-09-28
SOUTHWEST UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, limited by the complex preparation process of traditional silk biomaterials, single functionality, and its own defects such as weak cell adhesion, even if it can be improved by adding functional substances later, the source and efficacy of functional substances The risk factors brought about, as well as the increase in production costs, have limited the marketization of silk biomaterials, and so far few clinical approvals and finalized products have been obtained.

Method used

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  • Preparation method of FGF1 sericin protein gel with activity for promoting cell proliferation and product thereof
  • Preparation method of FGF1 sericin protein gel with activity for promoting cell proliferation and product thereof
  • Preparation method of FGF1 sericin protein gel with activity for promoting cell proliferation and product thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Example 1, the method for extracting sericin in human FGF1 silk

[0034] The cocoon shells were ground into powder by liquid nitrogen and used for later use; at a concentration of 50 mg / ml, extraction buffer 1 (50 mM Tris-HCl, 8 M urea, pH 7.0) was used for extraction at 80 ° C for 1 h; extraction buffer 2 ( 9.3M LiBr) at 60°C for 4h; using extraction buffer 3 (0.02M Na 2 CO 3 ) in a boiling water bath for 1 h to obtain a sericin solution containing human FGF1 protein.

Embodiment 2

[0035] Example 2, the preparation method of sericin gel (SS-FGF1) containing human FGF1 protein

[0036] Centrifuge the extract of extraction buffer 1 at 80°C, 13400rpm-18,000rpm for 30min, take the supernatant and refold the supernatant using the dithiothreitol / glutathione (DTT / GSSH) redox system FGF1 protein in . Refolding process: At 4°C, the supernatant was fully dialyzed with a dialysis bag (MWCO 1000Da, Spectrum Laboratory, Inc, USA) for 12 hours, and then replaced with a dialysate (8M urea, 1mM dithiothreitol (DTT), 50mM Tris–Cl ( pH 7.0), and 250mM NaCl); using half-dilution method using refolding solution (2.0mM reduced glutathione (GSH), 0.2mM oxidized glutathione (GSSG), 1mM DTT, 50mM Tris–Cl (pH 7.0), and 250mM NaCl ) to replace the dialysate, dialysis for 12 hours each time, repeat 4 times; dialyze with ultrapure water 6 times, remove the refolding solution, 12 hours each time. After renaturation is completed, the sericin aqueous solution is centrifuged at 4° C....

Embodiment 3

[0040] Example 3, performance detection of sericin gel containing human FGF1 protein

[0041] a. Electron microscope observation (SEM)

[0042] SS-FGF1 with a length and width of 1 cm was freeze-dried, and the inner layer was sprayed with gold, and observed and photographed with a scanning electron microscope (Supra 55sapphire, Zeiss). All experiments were performed at room temperature with a voltage of 3.0 kV. The number of samples in each experiment was 5 independent samples, and each sample was taken 3 times. SEM results showed that the interior of SS-FGF1 was a porous loose structure with a void size of 137±48 μm ( image 3 ).

[0043] b. Infrared spectroscopy (FTIR)

[0044] Freeze-dry the SS-FGF1 whose length and width are respectively 1 cm, and take its inner layer as a test sample. Determination was carried out using an infrared spectrometer (Thermo fisher scientific). An infrared spectrum analyzer (Thermo fisherscientific) was used to analyze the infrared absorp...

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Abstract

The invention relates to a preparation method of FGF1 sericin protein gel with activity for promoting cell proliferation and a product thereof. The preparation method comprises the following steps that sericin protein containing human FGF1 protein is extracted by human FGF1 gene modifying silk, and a sericin protein solution containing human FGF1 protein is obtained; after renaturation, an FGF1 sericin protein aqueous solution is obtained after dialysis with water, and FGF1 sericin protein hydrogel is obtained by centrifugation; and the FGF1 sericin protein hydrogel is placed at 4 DEG C, an FGF1 sericin protein gel material is formed by cross-linking of a chemical cross-linking agent or inducement of an inducer. The prepared gel material has the activity of promoting cell proliferation andhas broad application potential in tissue engineering.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to a preparation method of FGF1 sericin gel with the activity of promoting cell proliferation, and also relates to a product prepared by the method. Background technique [0002] Silk is mainly composed of silk fibroin in the inner layer and sericin in the outer layer, the content of which is 75% and 25% respectively. For a long time, a large amount of sericin protein produced by the silk reeling industry has been regarded as "waste" and discharged, resulting in serious environmental safety hazards. With the deepening of research, researchers found that sericin has the functions of anti-oxidation, moisturizing, promoting cell proliferation and accelerating wound healing, and it is a potential raw material for biomaterials. Sericin can be prepared such as sericin ointment, sericin-gelatin film, sericin-carboxymethyl cellulose, sericin poly(ethylene) after chemical crosslinking, physical b...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61L27/26A61L27/54A61L27/52A61L27/58
Inventor 王峰夏庆友王元成田驰赵萍
Owner SOUTHWEST UNIVERSITY
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