Magnetized cells and method for inducing magnetized cells

Abstract

To provide a technique capable of efficiently inducing magnetized cells by applying a magnetic field.SOLUTION: Provided are magnetized cells comprising iron oxide in which the content of iron from the iron oxide is 35 pg / cell or more.SELECTED DRAWING: Figure 1

Description

Technical Field

[0001] The present invention relates to magnetized cells that can be induced by applying a magnetic field and a method for inducing the same.

Background Art

[0002] In recent years, technology has been developed to apply a magnetic field to cells magnetized by combining cells with magnetic particles, and to accumulate the magnetized cells at a specific site in a patient's body to perform regeneration of damage to the site. Such technology is also called magnetic targeting. For example, Patent Documents 1 and 2 disclose magnetic field induction devices that can be used for magnetic targeting.

Prior Art Documents

Patent Documents

[0003]

Patent Document 1

Patent Document 2

Summary of the Invention

Problems to be Solved by the Invention

[0004] However, it is unclear under what conditions magnetized cells can be efficiently induced and retained at a desired position by applying a magnetic field.

[0005] One aspect of the present invention aims to provide a technique capable of efficiently inducing and retaining magnetized cells by applying a magnetic field.

Means for Solving the Problems

[0006] In order to solve the above problems, a magnetized cell according to one aspect of the present invention contains iron oxide, and the content of iron derived from the iron oxide is 35 pg / cell or more. According to the above configuration, by applying a magnetic field, the magnetized cells can be induced and retained at a desired position.

[0007] In a magnetized cell according to one aspect of the present invention, the content may be 125 pg / cell or less. According to the above configuration, magnetized cells containing such an amount of iron oxide can be easily produced.

[0008] In order to solve the above problems, a method for inducing magnetized cells according to one aspect of the present invention includes a step of applying a magnetic field having a magnetic flux density of 0.1 T or more to magnetized cells containing iron oxide and having an iron content derived from the iron oxide of 35 pg / cell or more, thereby inducing and retaining the magnetized cells at a desired position.

[0009] In a method for inducing magnetized cells according to one aspect of the present invention, in the above step, the magnetic field is generated using a solenoid coil, the magnetized cells are injected near the affected part of an animal, and the affected part may be arranged near the center of the solenoid coil.

Advantages of the Invention

[0010] According to one aspect of the present invention, it is possible to provide a technique for efficiently inducing magnetized cells by applying a magnetic field.

Brief Description of the Drawings

[0011]

Figure 1

Figure 2

Figure 3

Figure 4

Mode for Carrying Out the Invention

[0012] An embodiment of the present invention will be described with reference to FIG. 1. The magnetized cells according to this embodiment contain iron oxide, and the content of iron derived from the iron oxide is 35 pg / cell or more. Such magnetized cells have the property that their positions are induced by the action of a magnetic field because they contain iron oxide. In this specification, animal cells containing iron oxide as described above are referred to as "magnetized cells".

[0013] (Magnetized cells) The magnetized cells have an iron content derived from iron oxide of 35 pg / cell or more. Conventionally, there has been no detailed knowledge about whether any magnetized cells can be induced to a desired position by applying a magnetic field. The present inventors have found that, without generating an overly strong magnetic field, for example, as a condition that enables induction by a magnetic field with a magnetic flux density of 0.1 T, it is sufficient that the magnetized cells contain iron oxide containing 35 pg or more of iron per cell.

[0014] The iron content derived from iron oxide in the magnetized cells may be 35 pg / cell or more, more preferably 40 pg / cell or more, and even more preferably 45 pg / cell or more.

[0015] Also, the upper limit of the iron content derived from iron oxide in the magnetized cells is not particularly limited as long as the function of the magnetized cells is not impaired and no harmful health events occur in the animal into which the magnetized cells are introduced. From this point of view, the iron content derived from iron oxide in the magnetized cells may be, for example, 1 ng / cell or less, 500 pg / cell or less, 250 pg / cell or less, 200 pg / cell or less, 150 pg / cell or less, or 125 pg / cell or less.

[0016] The iron oxide contained in the magnetized cells preferably exhibits superparamagnetism. If the iron oxide exhibits superparamagnetism, the greater the amount of iron oxide contained in the magnetized cells, the more efficiently the induction force due to the application of a magnetic field acts on the iron oxide. Therefore, the induction and retention of the magnetized cells due to the application of a magnetic field become easier. ·Retention becomes easier.

[0017] The iron oxide contained in the magnetized cells is preferably coated with a water-soluble polysaccharide. Examples of the water-soluble polysaccharide for coating the iron oxide include dextran, dextrin, cellulose, hyaluronic acid, gelatin, mannan, pullulan, and chondroitin sulfate. Among them, carboxydextran is preferable. The water-soluble polysaccharide for coating the iron oxide may be one of these, or a mixture of two or more of them. Such a water-soluble polysaccharide can coat the iron oxide inexpensively and easily, and can effectively prevent the adverse effects of the iron oxide on cells.

[0018] A specific example of the iron oxide coated with such a water-soluble polysaccharide can be Ferucarbotran. Ferucarbotran is iron oxide particles in which maghemite (γ-Fe2O3) is coated with carboxydextran. Ferucarbotran is clinically used as a contrast agent for MRI (Magnetic Resonance Imaging), and is preferable as the iron oxide contained in the magnetized cells in terms of the established safety for the human body. Also, Ferucarbotran is preferable as the iron oxide contained in the magnetized cells in terms of exhibiting superparamagnetism.

[0019] When the magnetized cells are injected into the body of an animal by injection or the like, it is preferable that they are suspended in an infusion. Such an infusion is preferably an isotonic electrolyte infusion, and may be, for example, physiological saline, Ringer's solution, or glucose solution. The infusion may be one of these, or a mixture of two or more of them.

[0020] (Examples of the use of magnetized cells) The animal cells that serve as hosts for magnetized cells are not particularly limited as long as they are cells derived from animals. Here, as an example of using magnetized cells, the case where the host animal cells are bone marrow-derived mesenchymal stem cells (hereinafter referred to as "bone marrow MSCs") will be described.

[0021] Cartilage damage caused by the peeling of cartilage contained in human joints together with the surface layer of the bone located in the vicinity thereof is extremely difficult to regenerate naturally. In order to regenerate such cartilage damage, it is effective to accumulate cells having the function of regenerating cartilage or bone at the affected area. As such cells, for example, bone marrow MSCs are known.

[0022] FIG. 1 shows an example of a method for regenerating knee cartilage damage. Conventionally, it has been known that bone marrow MSCs exhibit a regenerative effect on knee cartilage by being accumulated at the damaged site of knee cartilage. However, simply injecting bone marrow MSCs near the damaged site (affected area) of knee cartilage hardly causes cartilage regeneration. This is presumably because the injected bone marrow MSCs do not accumulate at the damaged site of knee cartilage but are dispersed in the body.

[0023] On the other hand, magnetized cells having bone marrow MSCs as hosts have the property of being able to be induced to a desired position by the application of a magnetic field. Therefore, as shown in FIG. 1, when magnetized cells having bone marrow MSCs as hosts are injected near the damaged site of knee cartilage and a magnetic field is applied so that the magnetized cells are induced to the damaged site, the magnetized cells efficiently accumulate at the damaged site. Therefore, according to such magnetized cells, knee cartilage damage can be regenerated very efficiently.

[0024] In addition, the magnetized cells are induced to the damaged site within several seconds to at most several minutes after the start of the application of the magnetic field. Clinically, it is required that bone marrow MSCs and the like accumulate at the affected area within about 10 minutes. However, for the magnetized cells according to this embodiment, the magnetized cells can be induced to the affected area more quickly and accumulated. It is possible.

[0025] Note that the type of animal cell serving as the host for magnetized cells is not limited to bone marrow MSCs. For example, it may be mesenchymal stem cells derived from sources other than bone marrow, or stem cells other than mesenchymal stem cells. The type of stem cell may be appropriately selected according to the damaged site to be regenerated. Also, the use of magnetized cells is not limited to the regeneration of damage in the body of an animal. For example, for the purpose of using magnetized cells as markers, the magnetized cells may be accumulated near a specific organ in the body of an animal. Therefore, the animal cell serving as the host for magnetized cells may be a cell of a type other than a stem cell.

[0026] The animal cell may be a human cell, a mammalian cell other than human, or an animal cell other than these.

[0027] (Method for producing magnetized cells) The method for producing magnetized cells is not particularly limited. For example, it may be a method of adding iron oxide to the medium in which the animal cell serving as the host for magnetized cells is cultured and culturing for a predetermined time. Regarding the addition amount of iron oxide and the culturing time, appropriate addition amounts and culturing times may be appropriately selected according to the type of animal cell serving as the host for magnetized cells, the type of iron oxide, the type of medium, the number of cells during culturing, etc. Magnetized cells are obtained when such iron oxide is taken into the cell by a mechanism such as endocytosis in the animal cell.

[0028] (Method for inducing magnetized cells) The method for inducing magnetized cells according to this embodiment includes a step of applying a magnetic field with a magnetic flux density of 0.1 T or more to magnetized cells containing iron oxide and having an iron content derived from the iron oxide of 35 pg / cell or more to induce and retain the magnetized cells at a desired position.

[0029] For the induction and retention of magnetized cells, it is sufficient to apply a magnetic field with a magnetic flux density of 0.1 T or more. If the magnetic field has a magnetic flux density of 0.1 T or more, it can be easily generated using various magnetic field sources. Examples of magnetic field sources include solenoid coils, superconducting coils, superconducting magnets, and permanent magnets.

[0030] It is preferable to use a solenoid coil as the magnetic field source. In this case, since the generated magnetic field only needs to be 0.1 T or more, there is no need to apply an overly large current to the solenoid coil. Also, it is easy to make the direction of the magnetic field orthogonal to the affected part with a solenoid coil. Moreover, the solenoid coil may be designed such that the affected part can be inserted into the hollow part. With such a design, it is easy to adjust the relative position of the solenoid coil with respect to the affected part, so the induction direction of the magnetized cells can be easily adjusted. For example, since the center of the solenoid coil has the strongest magnetic field, by adjusting the position of the solenoid coil so that the affected part is arranged near the center, the induction and retention of magnetized cells in the affected part become easy.

[0031] In other words, in the method for inducing magnetized cells according to the present embodiment, in the above-described steps, the magnetic field is generated using a solenoid coil, the magnetized cells are injected near the affected part of an animal, and the affected part is preferably arranged near the center of the solenoid coil. As the magnetic field source provided with a solenoid coil, for example, the magnetic field induction device disclosed in Patent Document 1 or Patent Document 2 may be used.

[0032] The magnetic field applied to the magnetized cells only needs to have a magnetic flux density of 0.1 T or more, more preferably 0.15 T or more, and even more preferably 0.2 T or more. The stronger the applied magnetic field, the easier the induction and retention of the magnetized cells. However, when using a solenoid coil or the like as the magnetic field source, a large current needs to be applied to generate a strong magnetic field. From the viewpoint of avoiding unnecessary power consumption, the magnetic field applied to the magnetized cells may have a magnetic flux density of 1 T or less, preferably 0.5 T or less, more preferably 0.3 T or less, and even more preferably 0.2 T or less.

[0033] (Magnetized cell induction kit) The magnetized cell induction kit according to this embodiment contains iron oxide and includes magnetized cells in which the iron content derived from the iron oxide is 35 pg / cell or more. Regarding the configuration of the magnetized cells, the above description can be incorporated by reference.

[0034] Further, the magnetized cell induction kit according to this embodiment may include animal cells in a state not containing iron oxide and iron oxide, instead of the above-described magnetized cells.

[0035] Such animal cells are not particularly limited as long as they are cells derived from animals. Examples of the animal cells may include, for example, bone marrow MSCs, mesenchymal stem cells derived from sources other than bone marrow, and stem cells other than mesenchymal stem cells. The type of stem cell may be appropriately selected depending on the damaged site to be regenerated. Also, the animal cells may be cells of a type other than stem cells. Also, the animal cells may be human cells, mammalian cells other than humans, or other animal cells.

[0036] Iron oxide can be incorporated into animal cells so that the iron content derived from the iron oxide in the magnetized cells is 35 pg / cell or more. It is preferable that the iron oxide exhibits superparamagnetism or ferromagnetism. Also, it is preferable that the iron oxide is coated with a water-soluble polysaccharide such as dextran. A preferable example of such iron oxide is ferucarbotran.

[0037] The magnetized cell induction kit according to this embodiment may further include a magnetic field source capable of generating a magnetic field with a magnetic flux density of 0.1 T or more. Examples of such a magnetic field source include, for example, a solenoid coil, a superconducting coil, a superconducting magnet, and a permanent magnet. As the magnetic field source, a solenoid coil is preferable. As the magnetic field source equipped with a solenoid coil, for example, it may be a magnetic field induction device disclosed in Patent Document 1 or Patent Document 2.

[0038] In addition, the induction kit for magnetized cells according to this embodiment may further include reagents such as animal cell culture media, instruments such as syringes, equipment such as a DC stabilized power supply for supplying current to the solenoid coil, and other operation manuals.

Example

[0039] 〔1. Induction of magnetized cells by magnetic field〕 An experiment was conducted to induce magnetized cells according to an embodiment of the present invention by applying a magnetic field with a magnetic flux density of about 0.1 T.

[0040] (1-1. Experimental conditions) The magnetic field applied to the magnetized cells was generated using a solenoid coil. The solenoid coil had an inner diameter of 200 mm, an outer diameter of 300 mm, a coil diameter of 2 mm, 100 axial stages, 20 radial stages, a total number of turns of 2000 turns, and an axial length of 200 mm. By passing a current of 12 A through the solenoid coil, a magnetic field with a magnetic flux density of about 0.1 T was generated at the coil center at the end of the solenoid coil.

[0041] As magnetized cells, bone marrow MSCs incorporating ferucarbotran were prepared. The incorporation of ferucarbotran was performed by adding ferucarbotran to the medium in which bone marrow MSCs were cultured and culturing. The iron content (Fe amount) per cell after culturing is shown in Table 1 below.

[0042] The iron content per unit volume is the average value of the results measured by an ICP (Inductively Coupled Plasma) emission analyzer after dividing each sample into three equal parts. The iron content per cell was determined by calculating the iron content of the entire sample (total cells) from the iron content per unit volume and then dividing by the number of cells in the sample.

[0043]

Table 1

[0044] (1-2. Experimental results) Referring to FIGS. 2 and 3, the experimental results will be described. For the magnetized cells of Examples 1 to 4, the state before applying a magnetic field is shown in FIG. 2, and the state reaching a steady state after applying a magnetic field of about 0.1 T is shown in FIG. 3. Note that this experiment was conducted with each sample of the magnetized cells suspended in physiological saline.

[0045] As shown in FIGS. 2 and 3, before applying the magnetic field, the magnetized cells were suspended substantially uniformly in physiological saline. On the other hand, when a magnetic field of about 0.1 T was applied by the solenoid coil, the magnetized cells were induced in the axial direction of the solenoid coil. From the above results, it was shown that the magnetized cells according to one embodiment of the present invention can be induced by a magnetic field of 0.1 T or more.

[0046] [2. Induction speed of magnetized cells by magnetic field] Next, an experiment was conducted to measure the speed (induction speed) at which the magnetized cells according to one example of the present invention or the magnetized cells according to the comparative example move by the induction of a magnetic field when a magnetic field having a magnetic flux density of about 0.1 T or about 0.2 T was applied to these cells.

[0047] (2-1. Experimental conditions) The magnetized cells of Example 5 and Comparative Examples 1 and 2 were prepared by adding ferucarbotran to the medium in which bone marrow MSCs were cultured and culturing them for 12 hours each. Table 2 below shows the concentration of ferucarbotran added to the medium and the iron content in the obtained cells.

[0048] [Table 2]

[0049] The magnetized cells of Example 5 and Comparative Examples 1 and 2 were each dissolved in physiological saline and allowed to stand in a water channel having a width of 2 mm. Thereafter, a magnetic field having a magnetic flux density of about 0.1 T or about 0.2 T was applied to these cells, and the induction speed of each cell was measured.

[0050] The magnetic field applied to each cell was generated using a solenoid coil. The solenoid coil had an inner diameter of 240 mm, an outer diameter of 404 mm, a width of 119 mm, 29 axial stages, 25 radial stages, and a total number of turns of 725 turns. By passing a current of 33 A through the solenoid coil, a magnetic field with a magnetic flux density of approximately 0.1 T is generated at the coil center at the end of the solenoid coil. Also, by passing a current of 66 A through the solenoid coil, a magnetic field with a magnetic flux density of approximately 0.2 T is generated at the coil center at the end of the solenoid coil.

[0051] Further, the solenoid coil was installed such that the water channel extends along a straight line including the central axis of the solenoid coil. Note that the solenoid coil is designed such that a stable magnetic field of approximately 0.1 T or approximately 0.2 T is applied in the range where each cell is induced in the water channel.

[0052] (2 - 2. Experimental Results) Referring to FIG. 4, the experimental results will be described. FIG. 4 shows the results of measuring the induction speed at which these cells are induced in the axial direction of the solenoid coil when a magnetic field with a magnetic flux density of approximately 0.1 T or approximately 0.2 T is applied to each cell of Example 5 or Comparative Examples 1 and 2. Note that the sedimentation speed of the cells in physiological saline without applying a magnetic field was 0.1 mm / s to 0.2 mm / s. Therefore, if the "cell induction speed" exceeded 0.2 mm / s, it was determined that there was an induction effect due to the application of the magnetic field.

[0053] As shown in FIG. 4, for the magnetized cells of Example 5, an induction effect due to the application of the magnetic field was observed if the magnetic flux density was approximately 0.1 T or more. On the other hand, for the magnetized cells of Comparative Examples 1 and 2, an induction effect was observed when a magnetic field with a magnetic flux density of approximately 0.2 T was applied, but no induction effect was observed when a magnetic field with a magnetic flux density of approximately 0.1 T was applied. From the results of the group to which a magnetic field with a magnetic flux density of approximately 0.1 T was applied, it was suggested that if the magnetized cells contained 30 pg / cell or more of iron, they would show an induction speed exceeding 0.2 mm / s. Therefore, it was shown that magnetized cells containing 35 pg / cell or more of iron can be stably induced by a magnetic field with a magnetic flux density of 0.1 T or more.

[0054] [Supplementary Notes] The present invention is not limited to each of the above-described embodiments or examples, and various modifications are possible within the scope shown in the claims. Embodiments obtained by appropriately combining the technical means disclosed in different embodiments or examples are also included in the technical scope of the present invention.

Industrial Applicability

[0055] The present invention can be used for regenerative medicine of animals and the like.

Claims

**Claim 1** Magnetized cells containing iron oxide, wherein the content of iron derived from the iron oxide is 35 pg / cell or more. Magnetized cells. **Claim 2** The magnetized cells according to claim 1, wherein the content is 125 pg / cell or less. **Claim 3** A method for inducing magnetized cells, comprising the step of applying a magnetic field with a magnetic flux density of 0.1 T or more to magnetized cells containing iron oxide, wherein the content of iron derived from the iron oxide is 35 pg / cell or more, to induce the magnetized cells to a desired position. **Claim 4** In the above step, the magnetic field is generated using a solenoid coil, the magnetized cells are injected near the affected part of an animal, and the affected part is disposed near the center of the solenoid coil. The method for inducing magnetized cells according to claim 3.

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