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Bone repair material containing rH-BMP2, and preparation method and application thereof

A technology of bone repair and rh-bmp2, which is applied in the field of regenerative medicine, can solve problems such as difficulty in achieving balance, fast degradation, poor mechanical properties, etc., and achieve the effect of improving degradation/release time and reducing allergies

Active Publication Date: 2019-04-30
ZHEJIANG RISING BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Natural polymer materials such as gelatin, chitosan, hyaluronic acid, alginic acid, and cellulose have become good carriers of BMP-2 due to their good biocompatibility and degradation characteristics, but they are used as bone repair materials containing rH-BMP2 Sometimes there are problems of poor mechanical properties and rapid degradation, even if it is combined with other materials, it is difficult to achieve a good balance

Method used

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  • Bone repair material containing rH-BMP2, and preparation method and application thereof
  • Bone repair material containing rH-BMP2, and preparation method and application thereof
  • Bone repair material containing rH-BMP2, and preparation method and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1B

[0038] The preparation of embodiment 1BMP-2

[0039] In order to facilitate the experiment process and quality control, the BMP-2 used in the experiment was prepared by the applicant:

[0040] According to the rH-BMP2 amino acid and gene sequence in Genbank, the rH-BMP2 gene sequence suitable for expression in Escherichia coli was constructed, and its nucleotide sequence (including partial restriction site) was SEQ ID NO.1 in the sequence table; The gene fragment was digested and inserted into pET plasmid vector, and then transformed into E. coli host after ligation; positive clones were screened, and the expression of rH-BMP2 was induced; fermentation, breaking up the bacteria, collection of inclusion bodies, inclusion body lysis and protein renaturation; anion and cation exchange layer The rH-BMP2 was obtained by analysis and purification, and electrophoresis showed a single band of about 12kDa (consistent with the reported rH-BMP2), and the amino acid sequence of the produc...

Embodiment 2

[0042] The preparation of embodiment 2 modified gelatin

[0043] The preparation method of chitosan modified gelatin is as follows: by mass ratio, gelatin: chitosan is 10:2; Heating and dissolving in 70 degree centigrade water bath uses distilled water gelatin to be mixed with the solution of 5% (w / v); Use 2% Chitosan was formulated into a 2% (w / v) solution with acetic acid (pH about 5); the chitosan solution was added to the gelatin solution and stirred evenly, and stirred for 1 hour at 30 degrees Celsius to fully cross-link.

[0044] The preparation method of linoleic acid-modified gelatin is as follows: by mass ratio, gelatin: linoleic acid is 10:1; Heating and dissolving in 70 degree centigrade water bath uses distilled water to prepare the solution of 5% (w / v) of gelatin; Use 2% Acetic acid (pH about 5) to prepare linoleic acid into a 0.5% (w / v) solution; mix the two solutions at a volume ratio of 1:1, and add 0.5% (w / v) linoleic acid to it , stirred at 30 degrees Celsiu...

Embodiment 3

[0047] A bone repair material containing rH-BMP2, the bone repair material containing rH-BMP2 consists of the following components by weight percentage:

[0048]

[0049]Calcium chloride and half of the modified gelatin are mixed and dissolved in water, and rH-BMP2 is added to freeze-dry. The freeze-dried powder is mixed with hydroxyapatite and the remaining amount of modified gelatin, dissolved in a small amount of water, filled and freeze-dried. After drying, it is irradiated and sterilized to obtain the bone repair material containing rH-BMP2. Store in refrigerator at 4°C when not in use.

[0050] The CCK-8 cell proliferation toxicity test conducted according to the kit instructions shows that the cytotoxicity of the material is CTSO-I grade, which preliminarily confirms its safety.

[0051] A control bone repair material containing rH-BMP2 was also prepared, in which chitosan was used instead of modified gelatin, and other processes and components remained unchanged. ...

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Abstract

The invention belongs to the fields of biomedical materials containing active proteins and regenerative medicine. Specifically, the invention provides a bone repair material containing rH-BMP2, wherein the bone repair material includes modified gelatin and rH-BMP2. The material can be used for repairing bone damage. The first object comprises that provided is the bone repair material containing rH-BMP2, wherein the bone repair material is composed of the following components by the mass percentage: 1%-5% of rH-BMP2, 60%-86% of modified gelatin, 5%-20% of hydroxyapatite, and 8%-15% of calcium chloride. The bone repair material containing rH-BMP2 improves the mechanical properties and achieves great progress in degradation / drug release time when being used as a sustained-release carrier, andsignificantly reduces the allergy. In in-vitro release experiments and animal experiments, the effect of chitosan is better than that of chitosan and other conventional similar materials. Cytotoxicity experiments also preliminarily prove the safety of the bone repair material.

Description

technical field [0001] The invention belongs to the field of biomedical materials containing active protein and the field of regenerative medicine. Specifically, the application provides a bone repair material containing rH-BMP2, which includes modified gelatin and rH-BMP2. The material can be used for repairing bone damage. Background technique [0002] Bone morphogenetic protein-2 (bone morphogenetic protein-2, BMP-2) is a key factor for promoting bone formation discovered by Marshall R. Urist in 1965 when conducting osteogenesis experiments. BMP-2 belongs to the TGF-α family, which is secreted by osteoblasts and acts on osteoblasts. BMP-2 is the main signaling molecule for cell differentiation into mineral deposition osteoblasts and plays a role in inducing osteoblast differentiation. . It is expressed during limb growth, endochondral ossification, and fracture, and plays an important role in bone growth and development, regeneration and repair. Since BMP-2 has a signi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61L27/02A61L27/12A61L27/22A61L27/58A61L27/54C08H1/00
CPCA61L27/025A61L27/12A61L27/222A61L27/227A61L27/54A61L27/58A61L2300/252A61L2300/412A61L2300/604A61L2430/02C08H1/00C08L89/00
Inventor 曹建新
Owner ZHEJIANG RISING BIOTECH CO LTD
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