JP2002209573A - Supporter and transplanter for cell and/or biological tissue, and method for supporting - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a means for putting the surface of a transplanter having cells/biological tissues placed thereon on the surface of a recipient organ or a subcutaneous tissue to transplant the cells and/or the biological tissues to the surface of the recipient organ or subcutaneous tissue without forming a lump, by which the sure, safe and simple transplantation of the cells and/or the tissues can be realized. SOLUTION: The supporter 1 comprises a biologically harmless material having many fine voids on the surface. When a culture liquid containing the cells/biological tissues are applied and adhered to the surface of the supporter, the culture liquid is naturally dropped, or a negative pressure is applied from the back side of the supporter. Thus, the culture liquid is absorbed or passed in the supporter through the voids existing on the surface. Consequently, the cells/biological tissues are adhered to the surface of the supporter to make the transplanter.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a transplant for transplanting a part of tissue (hereinafter referred to as a living tissue) collected from a cell / living body to the surface of an organ or the surface of a subcutaneous tissue, and a method for supporting them.

[0002]

2. Description of the Related Art Conventional cell / living tissue transplantation involves transplanting cells / living tissue from a donor to the recipient's organ (liver, kidney, blood vessel, muscle, brain, etc.). Often inside. When cells / living tissues are to be transplanted onto the surface of a recipient organ or tissue, the cells / living tissue is directly placed on the organ surface or tissue surface.

[0003]

However, in conventional cell / living tissue transplantation, when cells / living tissue is transplanted into an organ, normally functioning organs (liver, kidney, spleen, blood vessel, muscle, brain, etc.) ) Is invasive. For example, transplantation of islets of Langerhans (hereinafter referred to as pancreatic islets), which is a pancreatic tissue transplant currently being performed in clinical practice, is mainly performed in the portal vein. It is invasive in that it punctures the portal vein, which is a blood vessel.
Portal vein obstruction (emboli) by injecting islets into the portal vein
There is also a risk of complications such as this. In addition, transplantation techniques require specialized skills. When cells / living tissues are transplanted into an organ, the cells / living tissues tend to be injected as a lump. For example, in a parathyroid tissue transplant, tissue is injected into the muscle of the recipient and the tissue forms a mass. Blood flow and nutrients from the recipient are difficult to reach near the center of the mass, which is disadvantageous for engraftment of cells / living tissues.

[0004] On the other hand, cells /
When a living tissue is transplanted, engraftment is difficult because the cell / living tissue easily shifts from the surface of the target site, and it is difficult to accurately transplant the cell / living tissue to the target site. Therefore, the present invention has been made in view of such problems,
Safe and secure without complications caused by transplantation operation,
It is another object of the present invention to provide a means for realizing simple cell / tissue transplantation.

[0005]

The above object is achieved by a carrier or implant according to the present invention. That is, the carrier of cells and / or living tissue is used to transplant cells / living tissue onto the surface of an organ or the surface of a subcutaneous tissue. When a culture solution containing cells / living tissue is applied to the surface of the carrier, the culture solution is absorbed into the carrier or passes through the carrier due to the voids present on the surface of the carrier, while the cells and / or The carrier is characterized in that the carrier has a surface shape such that all the living tissues do not pass through the voids and are not buried inside the carrier. The minimum pore size of the pores present on the surface of the carrier is smaller than the individual cells and / or living tissue to be placed, or at least a part of the cells / living tissue that entered the cavity emerges from the surface of the carrier. Shape.

[0006] The carrier contains a culture solution and / or a substance (eg, a cell growth factor) which affects cells / living tissues in the material of the carrier and / or in the space inside the carrier. Since the culture solution and / or the substance affecting the cells / living tissue is gradually released from the carrier, the cells / living tissue placed on the surface of the carrier can be cultured or expanded before or after transplantation. .

[0007] These carriers and / or transplants of cells and / or living tissues are related to a relatively safe and simple new cell / living tissue transplantation technique. The carrier of the cells and / or living tissue according to the present invention is prepared by placing the surface of the carrier on which the cell / living tissue (hepatocytes, pancreatic islets, parathyroid tissue, thymus tissue, brain tissue, skin tissue, etc.) is attached, as By placing it facing the organ surface or subcutaneous tissue surface,
Organs (liver, spleen, muscle,
Brain or omentum) or subcutaneous tissue. The carrier of the cell / living tissue may be devised so as to be fixed to the surface of the organ or the surface of the subcutaneous tissue by a clip, an adhesive, a suture, or the like. Thus, the cell / living tissue can be safely and reliably transplanted without dislocation of the cell / living tissue from the target site on the surface of the organ or subcutaneous tissue of the recipient. Moreover, the transplantation procedure is simple, not invasive, and does not cause any complications. Transplanted cells / living tissues are less likely to form clumps than conventional transplantation methods, so that blood flow and nutrients can easily reach the recipient's organ, which is advantageous for engraftment.

In the transplant according to the present invention, since the substance contained in the carrier that affects cells / living tissue, for example, cell growth factor, is slowly released from the bearing, the cell / living tissue placed on the surface of the transplant is Growth is promoted. Thereafter, if the cell / living tissue is placed with the surface of the transplanted surface facing the surface of the recipient organ or the surface of the subcutaneous tissue, the cell growth factor is gradually released from the carrier. Angiogenesis from the surface of the subcutaneous tissue is promoted, and the cells / living tissue on the surface of the implant are nourished early. This is advantageous for the cells / living tissue to engraft in the recipient organ or subcutaneous tissue. When the culture medium is contained in the carrier, the culture medium is gradually released from the carrier, so that the cells / living tissue placed on the surface of the implant are cultured.

[0009]

DESCRIPTION OF THE PREFERRED EMBODIMENTS Hereinafter, the present invention will be described based on embodiments. FIG. 1 is a perspective view of a carrier 1 according to one embodiment.
Shown in The carrier 1 is made of a harmless material, but the carrier 1 itself is made of a bioabsorbable material that is absorbed in the recipient's body while leaving cells and / or living tissue attached to the surface. In this case, materials such as hydrophilic chitin, collagen, gelatin, starch, fibrin, serum albumin, alginic acid, hyaluronic acid, cellulose oxide, and bioabsorbable ceramics can be used. The carrier 1 does not necessarily need to be bioabsorbable, and may be a material that is not absorbed in the body. In this case, as the non-bioabsorbable material, cellulose, polyvinyl alcohol, polyacrylic acid, ceramics and the like can be used. The above are hydrophilic and suitable materials, but not limited thereto, and may be hydrophobic depending on the production method. In the case of a hydrophobic material, as a bioabsorbable material, polylactic acid, polyglycolic acid, lactic acid-glycolic acid copolymer, polycyanoacrylate, polyanhydride, polyorthoester, chitin, chitosan, polycarbonate, etc. Materials such as polyvinyl chloride, polyolefins, and polypropylene can be used as non-bioabsorbable materials.

As an example of the carrier 1, a non-woven fabric is used as shown in FIG. The carrier 1 having such a form has a large number of minute voids 2 on the surface. When a culture solution containing cells / living tissue is applied to the surface of the carrier, the cavity 2 absorbs the culture solution into the carrier by gravity or by applying a negative pressure (negative pressure) from the back surface of the carrier. Or through a carrier. On the other hand, the void 2 is an individual cell /
The shape is such that not all of the living tissue enters the inside of the carrier, but some or all of the living tissue can protrude from the surface of the carrier. Specifically, as an example, FIG. 2 shows a partially enlarged view of the surface of the carrier, and the minimum diameter 3 of the pore diameter on the surface of the carrier of each of the voids 2 is part or all of the individual cells to be placed.
/ It may be smaller than part or all of the living tissue 10. Incidentally, if all of the minimum diameters 3 of the individual pore diameters are larger than all the cells / living tissues 10 to be attached, it is not preferable because all the cells / living tissues 10 are buried inside the carrier. It is also preferred that a culture solution containing cells / living tissue is widely spread on the surface of the carrier. It is not preferable to apply the coating in one place because it easily forms a mass of cells / living tissue.

The carrier 1 shown in FIG. 1 has a square shape and the dimensions a × b × t = 20 mm × 20 mm × 1 mm. However, this shape and size are merely examples, An optimal shape and size may be used. In addition, a carrier in which a part or all of the minimum diameter 3 of the pore diameter on the surface of the carrier of each void 2 is smaller than a part or all of the individual cell / living tissue 10 to be attached may be used. In order to attach the cells / living tissue to the surface of the support 1 as described above, a culture solution containing the cells / living tissue is applied to the surface of the support 1. At this time, the culture solution is absorbed into the carrier 1 through the gap 2 or passed through the carrier by gravity or by applying a negative pressure (negative pressure) from the back surface of the carrier. Then, since the minimum diameter 3 of the pore size on the surface of the carrier of the void 2 is smaller than a part or all of the cell / living tissue 10, as shown in the partial enlarged sectional view of the carrier in FIG. A part or all of 10 is placed on the surface of the support 1.
At this time, the cells / living tissue 10 may be placed in a state of being layered. In this state, as shown in FIG. 4, the cell / living tissue 10 is placed by placing the surface of the carrier 1 on which the cell / living tissue 10 is placed facing the organ surface or subcutaneous tissue surface of the recipient. It is implanted on the surface of an organ or subcutaneous tissue 20.

In the embodiment shown in FIGS. 1 and 2, the shape of the space 2 on the surface of the carrier is considered to be a spindle shape, but other than the spindle shape, it is substantially circular, almost gourd-shaped, substantially polygonal, or substantially star-shaped. May be.

FIG. 5 is a partially enlarged cross-sectional view of a modified example of the shape of the gap. That is, the carrier 1 may have the shape of the gap 2 as shown in the partially enlarged sectional view of FIG. Some or all of the minimum diameter 3 of the pore diameter on the carrier surface of the void 2 is a cell / cell even if the pore diameter is larger than a part or all of the biological tissue 10.
It suffices that a part of the cell / living tissue protrudes from the surface without the whole living tissue entering the gap 2.

FIG. 6 is a partially enlarged cross-sectional view of an example of a shape change in which the surface of the carrier shown in FIG. In FIGS. 1, 3 and 5, the surface of the carrier forms a plane, and there are gaps in the plane, but the surface of the carrier may have a shape as shown in FIG. The voids 2 are present on the uneven surface. When placing the carrier / body with the living tissue facing the recipient's organ surface or subcutaneous tissue surface, the cell /
It is sufficient that the carrier has a shape of the surface of the carrier such that the living tissue can come into contact with the surface of the organ or the subcutaneous tissue of the recipient. Since the non-woven fabric can be freely deformed, it can be easily used according to the shape of the organ or the subcutaneous tissue. However, it is convenient to prepare a non-deformed material in advance according to the shape of the organ or the subcutaneous tissue. A reinforcing material such as a mesh may be included inside or on the back surface of the support to maintain the shape.

The carrier has not only a surface but also a space 2 inside the carrier for absorbing or passing a liquid such as a culture solution. Specifically, for example, there is a nonwoven fabric shown in FIG. 7, 8 and 9 are partially enlarged views of a carrier according to another embodiment. FIG. 7 is a partially enlarged view of a carrier having a porous void 2, FIG. 8 is a polygonal column void 2, and FIG. The porous void 2 shown in FIG. 7 may be a void such as pumice or a void such as a sponge. The cross section of the groove in FIG. 9 does not need to be linear, that is, may be circular or curved.

A transplant can be prepared by spontaneously dropping a culture solution containing cells / living tissue onto the surface of the carrier. In this case, the space on the surface of the carrier and the internal space are preferably connected to each other (communication space) or independent spaces, and the material of the support is preferably hydrophilic. Also, by applying a culture solution containing cells / living tissue to the surface of the carrier and applying a negative pressure (negative pressure) from the back surface of the carrier to aspirate the culture solution, the cells / living tissue is transferred to the carrier It can be attached to a surface to produce an implant. In this case, the pores on the surface and inside of the carrier are preferably communicating pores, and the material of the carrier may be hydrophobic as well as hydrophilic.

The carrier may be composed of a material containing a culture solution and / or a substance (cell growth factor, etc.) that affects cells / living tissue, or a culture solution and / or a substance affecting cells / living tissue. May be contained in the voids, and these are gradually released. FIGS. 1, 7, 8 and 9 show examples of the voids on the surface and inside of the carrier.

It is not necessary to attach cells / living tissues to the carrier 1 and immediately carry out transplantation. After culturing the cells / living tissue placed on the surface of the carrier 1 for several days in a culture solution,
Alternatively, transplantation can be performed by thawing after cryopreservation.

Next, an example of an animal experiment on cell / living tissue transplantation according to the present invention will be described. <Examples> (Animals and Methods Used) ACI rats (body weight 180-
220 g). Recipients were syngeneic ACI rats, genetically approximately equivalent to the donor and diabetic by intravenous injection of streptozotocin 60 mg / kg (body weight 220
２５０250 g).

First, the pancreas was excised from the donor, and the islets were separated and purified by collagenase digestion and dextran discontinuous gradient method. The minimum diameter of the pore diameter on the carrier surface of the void is 150 μm at the maximum and the minimum diameter of the pore diameter on the carrier surface of the void is 0.1 μm, and the average of the minimum diameter of the pore diameter on the carrier surface is 75 μm. Non-woven carrier (20 mm x 20 mm)
× 1 mm large) were prepared. 75 μm or more and 300 μm
By spontaneously dropping a culture solution containing a total of 2,000 pancreatic islets of a size smaller than the entire surface of the carrier, the islets were placed on the surface, and a transplant containing the culture solution inside the carrier was produced. The islet was transplanted to the liver surface by placing the transplanted islet mounting surface in contact with the liver surface of the recipient. After transplantation, non-fasting blood glucose levels and body weight of the recipients (8 animals) were measured, and histological examination of the transplanted islets was performed. (Results) The non-fasting blood glucose level (mean ± standard error) of 7 out of 8 recipients (87.5%) was 501 ± pre-transplantation value.
From the high value of 26 mg / dl, it gradually decreased, and all became low values (normal values) of 200 mg / dl or less within 4 weeks, and the low value was maintained (262 ± 53 on day 10 after transplantation, 208 ± 20 days after transplantation). 4
8, 30 days 148 ± 8, 50 days 131 ± 9, 100
Day 139 ± 5, Day 150 135 ± 6). This indicates that the transplanted islets function normally and secrete enough insulin. The weight of these seven recipients was reduced before transplantation but increased after transplantation. This indicates that the transplant improved the diabetic status of the recipient. Histological examination 150 days after transplantation confirmed engraftment of pancreatic islets in the liver of the recipient. No complications due to the transplantation operation were observed. As described above, in the new method for transplanting islets into the liver surface using the transplant, the islets were cultured in a culture solution gradually released from the carrier at the early stage of transplantation, and survived and showed good functions. It was confirmed that this was a safe and simple cell / tissue transplantation technique without complications due to the transplantation operation.

<Comparative Example> In the same animal as in the example,
A carrier having a gap larger than the gap of the carrier used in the example was prepared. In other words, the minimum diameter of the pore diameter on the surface of the support of the void is 1000 μm at the maximum and 300 μm at the minimum, and the average of the minimum diameter of the pore diameter on the surface of the support of the void is 650 μm.
m, a non-woven support made of hydrophilic chitin (20 mm ×
An experiment was performed using two (20 mm × 1 mm size). 75
A transplant containing pancreatic islets was prepared by spontaneously dropping a culture solution containing a total of 2000 pancreatic islets having a size of not less than μm and less than 300 μm over the entire surface of the carrier. The pancreatic islets were transplanted to the recipient by placing the transplanted islet mounting surface in contact with the liver surface of the recipient. After transplantation, the recipient's non-fasting blood glucose levels remained high without showing low levels. This indicates that the transplanted islets do not function properly and do not secrete enough insulin.
The recipient's body weight was reduced before transplantation but decreased without increasing after transplantation. This indicates that the transplant has not improved the recipient's diabetic status. From the above, if the minimum diameter of the pore diameter of all the voids present on the surface of the carrier is larger than all the transplanted islets, all the transplanted islets are buried inside the carrier without being placed on the surface of the carrier. The transplanted islets cannot be placed on the liver surface,
It was confirmed that the transplant was not successful.

[0022]

As can be seen from the above experimental results, according to the present invention, safe, reliable, and simple cell and / or tissue transplantation can be realized without complications due to the transplantation operation. Although the above effects were confirmed in rats, similar effects can be expected for humans. In addition, application to transplantation of regenerated cells and / or regenerated tissues can be expected. With safe, reliable and easy transplantation in the clinic, cells,
Not only does this provide mental stability and reduced financial burden for those who require tissue transplantation, but it also makes it possible for many general surgeons to be performed in multiple centers.

[Brief description of the drawings]

FIG. 1 is a perspective view of a cell and / or tissue carrier according to one embodiment.

FIG. 2 is a partially enlarged plan view schematically showing a space on a surface and cells and / or tissues in the carrier of FIG. 1;

FIG. 3 is a partially enlarged cross-sectional view when a culture solution containing cells and / or tissues is applied to a carrier.

FIG. 4 is a partial schematic diagram showing a state in which the surface of a carrier on which cells and / or tissues are placed is placed facing an organ surface or a tissue surface of a recipient.

FIG. 5 is a partially enlarged cross-sectional view showing an example of changing the shape of the gap of the carrier.

FIG. 6 is a partially enlarged cross-sectional view of a modified example in which the surface of the carrier shown in FIG. 1 has irregularities when a culture solution containing cells and / or tissues is applied to the carrier.

FIG. 7 is a partially enlarged view showing an example of a change in the shape of voids on the surface and inside of the carrier in FIG. 1;

FIG. 8 is a partially enlarged view showing another example of the shape change of the voids on the surface and inside of the carrier in FIG. 1;

FIG. 9 is a partially enlarged view showing still another example of the shape change of the voids on the surface and inside of the carrier in FIG. 1;

[Explanation of symbols]

DESCRIPTION OF SYMBOLS 1 Carrier 2 Gap 3 Minimum diameter of pore diameter on the carrier surface of cavity 2 10 Cells and / or tissue 20 Organ or subcutaneous tissue

Claims (5)

[Claims] 1. A carrier for mounting cells and / or living tissues on its surface, wherein the surface has a large number of voids on its surface for absorbing or passing through a culture solution containing cells and / or living tissues to be mounted. Cell and / or biological tissue carrier made of the following materials 2. A transplant in which cells and / or living tissues are attached to the surface of the carrier according to claim 1. 3. A carrier for culturing and growing cells and / or living tissue, wherein the carrier according to claim 1 is made to contain a culture solution and / or a substance which affects cells or living tissue. 4. The transplant according to claim 2, wherein the culture medium and / or a substance that affects cells or living tissue is present in the carrier portion of the transplant. 5. A cell comprising a step of applying a culture solution containing cells and / or living tissue to the surface of a carrier, and then absorbing or passing the culture solution through a gap to place the cells and / or living tissue on the surface of the carrier. And / or a method for supporting biological tissue