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Natural-tissue-derived decellularized and decalcified bone material and preparation method thereof

A decellularized and demineralized bone technology, applied in medical science, prosthesis, etc., can solve the problems of lack of uniformity of bone repair effect, loss of natural biological ability of repairing bone, and reduction of ECM biological activity, etc. Microenvironment, efficient and convenient uniformity, the effect of improving biological activity

Inactive Publication Date: 2016-03-30
GUANGXI MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the bone decellularization protocols of different companies or laboratories are very different due to issues such as the location of the material, the type of bone tissue, and the size of the bone tissue.
However, different intensities of decellularization methods will have great differences in the retention of extracellular matrix, which makes the bone repair effect of clinical patients lack uniformity.
At the same time, in the prior art, strong acid decalcification (concentrated hydrochloric acid, formic acid, acetic acid, etc.) is often used to obtain decalcified bone tissue or decellularized decalcified bone material. This treatment process will greatly reduce the biological activity of ECM, making Bone material has largely lost its natural biological ability to repair bone

Method used

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  • Natural-tissue-derived decellularized and decalcified bone material and preparation method thereof
  • Natural-tissue-derived decellularized and decalcified bone material and preparation method thereof
  • Natural-tissue-derived decellularized and decalcified bone material and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Example 1 Preparation and Research of Completely Decalcified and Decellularized Cancellous Bone Material

[0042] (1) Take the porcine ilium cancellous bone, transfer it to an environment at 4°C, and rinse it with sterile saline for 3 times, 20 minutes each time, to remove blood, residual muscle tissue, hair, ligament and other tissues; the size of the sample depends on the clinical practice Depending on the needs of the operation, usually the size is 2cm*2cm*2cm

[0043] (2) In 1000ml of normal saline buffer solution containing protease inhibitors (concentration is 1%, protease inhibitor content is 10KIU / ml), shake at 150rpm at a constant temperature of 45°C for 24 hours, and rinse with normal saline for 5 hours;

[0044] (3) In 1000ml of chloroform and methanol solution with equal volume ratio, degrease with constant temperature 45°C shaker at 150rpm for 12 hours, and rinse with normal saline for 5 hours;

[0045] (4) In 1000ml of TritonX-containing PBS buffer soluti...

Embodiment 2

[0052] Example 2 Preparation and Research of Completely Decalcified and Decellularized Compact Bone Material

[0053] The compact bone of the pig's limbs is taken, and the size of the sample depends on the actual clinical operation needs, and the size is usually 1*1*0.5cm; the rest is carried out with reference to the method of Example 1 to obtain completely decalcified and decellularized compact bone materials.

[0054] The complete decalcified and decellularized compact bone material obtained in this case was evaluated histologically, quantitatively detected for antigenic components, and detected for osteogenic repair ability. The results are as follows: figure 2 , 4 , 6, 8. figure 2 HE staining showed that the extracellular matrix of completely decalcified and decellularized cancellous bone materials remained intact, the nuclear components were completely removed, and no cells and their fragments remained; Figure 4 DAPI staining further indicated that the nuclear compo...

Embodiment 3

[0055] Example 3 Preparation and Research of Completely Decalcified, Decellularized Cancellous Bone and Compact Bone Materials

[0056] Get porcine ilium cancellous bone and limb cortical bone, step (2) in 1000ml containing protease inhibitor physiological saline buffer solution (concentration is 5%, protease inhibitor content is 10KIU / ml), constant temperature 45 ℃ shaker Shake at 150rpm for 8 hours; for the rest, refer to the method of Example 1 to obtain completely decalcified, decellularized cancellous bone and compact bone materials.

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Abstract

The invention discloses a preparation method of a natural-tissue-derived decellularized and decalcified bone material. According to the method, any bone tissue of a mammal is treated with a protease inhibitor-containing normal saline buffer, an organic solvent, Tirton X-containing PBS (phosphate buffer saline), SDS (sodium dodecyl sulfonate)-containing PBS, pancreatin-containing PBS, deoxyribonuclease-containing PBS, an EDTA (ethylene diamine tetraacetic acid) isotonic solution and ultrasonic waves, and the decellularized and decalcified bone material is obtained. Cancellous bones and cortical bones can be decellularized completely and simultaneously, the conditions are mild, damage to an ECM (extracellular matrix) is avoided, rapidness and stability are realized, and the obtained decellularized and decalcified bone material has the advantages of good biocompatibility, high plasticity, high biomechanical strength and the like and can be used for clinically repairing bone regeneration and repair disorder such as bone defects, nonunion and the like caused by various causes.

Description

technical field [0001] The invention belongs to the technical field of bone tissue repair and regeneration, and in particular relates to a decellularized combined decalcified bone material derived from natural tissue and a preparation method thereof. Background technique [0002] At present, osteonecrosis and bone defect caused by trauma, tumor, infection and other factors are very common in clinical practice. For this reason, artificial biomaterials have been gradually applied to the repair of bone tissue in patients. However, due to the deficiencies in biocompatibility, degradability, osteoconductivity, and osteoinduction, autologous bone grafting is still considered the most ideal bone repair material. However, in actual clinical operations, autologous bone transplantation cannot be well satisfied with the repair and treatment of patients with osteonecrosis or bone defect due to the number of materials taken and secondary damage. [0003] The extracellular matrix (ECM) d...

Claims

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Application Information

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IPC IPC(8): A61L27/36A61L27/50
Inventor 赵劲民宗少晖苏伟秦安林贤丰韦庆军
Owner GUANGXI MEDICAL UNIVERSITY
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