JP2007054374A - Manufacturing method for bone supplementation body - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To reduce burden which is applied to a patient by enabling a bone supplementation body for quickly restoring a lesion when being transplanted in the body of the patient to be easily and quickly manufactured. <P>SOLUTION: This manufacturing method for the bone supplementation body comprising following steps is provided. In the step S1, a peripheral blood is extracted. In the step S2, the macrophage is collected from the extracted peripheral blood. In the step S4, the collected macrophage is seeded in a bone supplementation material. <P>COPYRIGHT: (C)2007,JPO&INPIT

Description

  The present invention relates to a method for producing a bone filling material.

  In recent years, it has become possible to regenerate a living tissue and repair a bone defect by repairing a bone defect caused by excision of a tumor, trauma, or the like with a bone filling material. For bone tissue regeneration, bone prosthetic materials that become cells, growth factors, and scaffolds (carriers) are indispensable. Hydroxyapatite (HAP) and tricalcium phosphate (TCP) are known as bone prosthetic materials, but from the idea that no foreign matter remains in the body, for example, it is made of a calcium phosphate porous material such as β-TCP. Scaffolding material is used. When β-TCP is compensated for in a bone defect portion, so-called remodeling is performed in which osteoclasts absorb β-TCP and osteoblasts form new bone. That is, the bone prosthetic material supplemented in the bone defect portion is replaced with autologous bone over time.

On the other hand, it has been proposed to use cultured bone produced by in vitro culturing bone marrow mesenchymal stem cells collected from patients with bone filling materials in order to increase the repair speed of bone defects after surgery. Yes. Since bone bone defects are filled with cultured bone containing many bone marrow mesenchymal stem cells that have been proliferated using the bone substitute material as a scaffold by culturing, it is replaced with autologous bone compared to the method of filling only the bone substitute material. It is possible to significantly reduce the number of days until the time is taken (see, for example, Non-Patent Document 1).
Uemura et al., “Tissue engineering in bone using biodegradable β-TCP porous material -Osferion, a new material that increases strength in vivo”, Medical Asahi, Asahi Shimbun, October 1, 2001, No. 30 Volume 10, No. 10, p. 46-49

  However, the cost of cultured bone is relatively high because of the long time required for culture. In addition, the collection of bone marrow fluid from the iliac and the like is relatively invasive and places a burden on the patient. In addition, the patient has to wait for a long period from the operation of collecting bone marrow fluid to the operation of transplanting cultured bone, and there is a disadvantage that it cannot be applied to an emergency patient. In addition, there is also a problem that it is difficult to produce systematic cultured bone because there are many cell losses due to subculture work during culture and there are individual differences in the stage of differentiation.

  The present invention has been made in view of the above-described circumstances, and can easily and quickly manufacture a bone filling body that quickly repairs an affected part by being transplanted into a patient's body, thereby reducing the burden on the patient. An object of the present invention is to provide a method for producing a bone filling material that can be used.

In order to achieve the above object, the present invention provides the following means.
The present invention provides a method for producing a bone filling material by collecting peripheral blood, collecting macrophages from the collected peripheral blood, and seeding the collected macrophages on a bone filling material.

  According to the present invention, since a bone filling material is produced using macrophages collected from peripheral blood, a cell source can be performed in a minimally invasive manner by collecting peripheral blood, and the burden on the patient can be greatly reduced. . Peripheral blood can be collected in large quantities, and since there are many macrophages in the peripheral blood, many macrophages can be obtained without culturing.

  Since macrophages can differentiate into osteoblasts and osteoclasts, osteoclasts differentiated from macrophages can be used as bone substitutes by filling the bone defect with bone substitutes that have been seeded on bone substitutes. It is absorbed, and osteoblasts form a bone matrix and bone tissue is repaired. Therefore, according to the method for manufacturing a bone substitute according to the present invention, the bone substitute can be quickly manufactured with minimal invasiveness, and the burden on the patient can be greatly reduced.

In the above invention, it is preferable to activate the collected macrophages.
In this way, macrophages can be differentiated into osteoblasts and osteoclasts at an earlier stage, and more rapid repair of bone defects can be achieved.

  Moreover, in the said invention, activation of a macrophage is good also as being performed by addition of a cytokine. Alternatively, macrophage activation may be performed by introducing proteins and peptides.

  According to the method for manufacturing a bone filling device according to the present invention, a bone filling material for quickly repairing an affected part by being transplanted into the body of a patient can be easily and quickly manufactured, and the burden on the patient can be reduced. There is an effect.

A method for manufacturing a bone prosthesis according to an embodiment of the present invention will be described below with reference to FIG.
As shown in FIG. 1, the method for manufacturing a bone substitute according to the present embodiment includes step S1 for collecting peripheral blood of a patient, step S2 for collecting macrophages from the collected peripheral blood, and activating the collected macrophages. And step S4 for seeding the activated macrophages on the bone grafting material.

In step S2 of collecting macrophages from peripheral blood, for example, the macrophages adhering to the inner wall of the centrifuge tube are collected by centrifuging the peripheral blood contained in the centrifuge tube using the adhesion of the macrophages. Collection of macrophages from the inner wall of the centrifuge tube can be performed, for example, by adding trypsin. By performing a temperature-responsive treatment on the inner wall of the centrifuge tube, the macrophages may be detached by the temperature change and collected. Thereby, macrophages in peripheral blood can be easily collected.
Peripheral blood cell sources include mononuclear cells, human monocyte-derived dendritic cells, T cells, lymphocytes, osteogenic cells, and PPP components in peripheral blood.

The step S3 for activating macrophages is, for example, bFGF, BMP-2, TGF-b, PDGF, EGF, IGF, IL-1, IL-11, IL-17, M-CSF, RANKL, EFMF, VFGF, HGF. Alternatively, it can be performed by adding a cytokine such as IHG.
Alternatively, macrophages are activated by introduction of proteins and peptides, administration of antigenic peptides, administration of calcium phosphate, administration of peptides and cytokines, administration of genes such as CD40 stimulation, liposome administration, polysaccharide modification, etc. May be.
In addition, substances having an affinity such as monocyte activating factor, leukocyte activating factor of mononuclear cells, chemokinesin, and chemotaxin may be added.

  As bone replacement material, block or granule solid or liquid 8-phosphate calcium, β-tricalcium phosphate porous body, hydroxyapatite, polylactic acid, polyglycolic acid, etc. are arbitrarily adopted according to the form of the affected area to be supplemented can do.

  Since the seeded macrophages are activated in the bone substitute produced by the production method according to the present embodiment, the bone substitute is easily differentiated into osteoblasts and osteoclasts, and is compensated for in a bone defect portion or the like. As a result, the differentiated osteoclasts absorb the bone filling material, and the osteoblasts can form a bone matrix to quickly repair the bone defect.

  As described above, according to the method for manufacturing a bone filling material according to the present embodiment, in comparison with the case where bone marrow fluid is collected by perforating the iliac bone or the like, peripheral blood is collected only by inserting a needle of a syringe into a blood vessel. Therefore, there is an advantage that the burden on the patient can be greatly reduced because the cell source can be collected by the method, and is extremely minimally invasive, does not require surgery.

  Unlike bone marrow fluid, which is limited in volume, macrophages are collected from peripheral blood that is present in large amounts in the body. Therefore, unlike mesenchymal stem cells in bone marrow fluid, a relatively large amount of macrophages can be obtained without going through a culture process. be able to. Therefore, there is an advantage that a culture process is not required, and equipment, chemicals, and time required for culture can be saved, and a bone substitute can be manufactured at low cost. Furthermore, since the culturing step is not required, there is an advantage that transplantation surgery can be performed even for patients who need treatment early.

  In this embodiment, macrophages activated by cytokine administration or the like are seeded on the bone grafting material, but instead of this, the collected macrophages are seeded on the bone grafting material as it is to obtain the bone grafting material. You may decide to manufacture. The macrophages in the bone graft produced in this way are inferior in their ability to differentiate into osteoblasts and osteoclasts compared to the activated case, but the bone tissue adjacent to the bone defect site of the transplant destination By obtaining a cytokine from the above, it is possible to gradually differentiate into osteoblasts and osteoclasts and repair the bone defect.

It is a flowchart which shows the manufacturing method of the bone filling body which concerns on one Embodiment of this invention.

Explanation of symbols

S1 peripheral blood collection step S2 macrophage collection step S3 macrophage activation step S4 macrophage seeding step

Claims (4)

Collecting peripheral blood,
Collecting macrophages from the collected peripheral blood,
A method for producing a bone filling material, wherein the collected macrophages are seeded on a bone filling material.   The method for producing a bone substitute according to claim 1, wherein the collected macrophages are activated.   The method for producing a bone substitute according to claim 2, wherein the macrophage is activated by addition of cytokine.   The method for producing a bone substitute according to claim 2, wherein the macrophage is activated by introducing a protein and a peptide.

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