JP2010246446A - Device for treating adipose tissue and apparatus for separating cell - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a device for treating adipose tissue and an apparatus for separating cell which smoothly discharge cell suspension while avoiding the decrease of recovery rate of fat-derived cells. <P>SOLUTION: The device for treating adipose tissue 1 includes: a container 2 having an outlet 2a; a filter 3 which, in a container 2, covers the outlet 2a, partitions the inside of the container 2, catches the adipose tissue A, and passes fat-derived cells separated from the adipose tissue A; a valve V1 which opens and closes the outlet 2a; a pump 5 connected to the outlet 2a and sucking the inside of the container 2; injection means 8 which holds a solution C dissolving collagen and injects the solution C into a space partitioned by the filter 3 and including the outlet 2a; the pressure detection part 7, provided near the outlet 2a, and detects the pressure in the container 2; and a control unit 9 which controls the valve V1 and the injection means 8 based on the pressure detected by the pressure detection part 7 while liquid is discharged from the outlet 2a. <P>COPYRIGHT: (C)2011,JPO&INPIT

Description

  The present invention relates to an adipose tissue processing apparatus and a cell separation apparatus.

  2. Description of the Related Art Conventionally, an apparatus that separates and extracts fat-derived cells including stem cells and the like from adipose tissue collected from the body is known (see, for example, Patent Document 1). When the adipose tissue is digested with the digestive enzyme solution, the adipose-derived cells are separated from the adipose tissue to produce a cell suspension, and the adipose-derived cells can be obtained by centrifuging the cell suspension. At this time, the cell suspension generated in the processing container is selectively fed into the centrifuge container while being separated from the adipose tissue by the filter.

International Publication No. 2005/012480 Pamphlet

  However, when the cell suspension is discharged from the processing container, the filter is clogged with blood cell components collected at the same time as the adipose tissue, residues of decomposed adipose tissue (hereinafter referred to as debris), and the cell suspension is in the middle. There is a problem that it will not be discharged. In particular, there is an inconvenience that collagen, which is a main component of adipose tissue, is not sufficiently decomposed by digestive enzymes or reassociates after being decomposed and is caught on the filter and covers the filter. Further, there is a problem that fat-derived cells are trapped in the collagen layer formed on the surface of the filter in this way, and the recovery rate of fat-derived cells is lowered.

  The present invention has been made in view of the above-described circumstances, and provides an adipose tissue processing apparatus and a cell separation apparatus that can smoothly discharge a cell suspension while preventing a reduction in the recovery rate of adipose-derived cells. The purpose is to do.

In order to achieve the above object, the present invention provides the following means.
The present invention includes a container having a discharge port, a filter that covers the discharge port in the container, partitions the container, captures adipose tissue, and passes fat-derived cells separated from the adipose tissue, A valve that opens and closes a discharge port; a pump that is connected to the discharge port and sucks the inside of the container; and holds a solution for dissolving collagen; the solution includes the discharge port partitioned by the filter An injection means for injecting into the space, a pressure detector provided in the vicinity of the outlet, and detecting the pressure in the container, and a pressure detected by the pressure detector while discharging the liquid from the outlet And a fat tissue processing apparatus comprising a control unit for controlling the valve and the injection means.

  According to the present invention, fat-derived cells can be separated from adipose tissue by closing the valve and mixing the adipose tissue and digestive enzyme solution in the container to digest the adipose tissue. After that, when the valve is opened and the inside of the container is sucked by the suction means, the fat-derived cells are discharged from the discharge port as a cell suspension suspended in the digestive enzyme solution while restricting the discharge of the adipose tissue by the filter. Can be recovered from the container.

  In this case, if the filter is clogged by debris generated from the adipose tissue during discharge of the cell suspension and the cell suspension does not pass through the filter, the pressure in the vicinity of the discharge port decreases. When the pressure detected by the pressure detection unit decreases, the control unit closes the valve and injects the solution by the injection unit, so that the filter is immersed in the solution. As a result, the collagen constituting the debris is dissolved by the lysis solution to eliminate clogging of the filter, and the cell suspension can be discharged smoothly by opening the valve again. In addition, fat-derived cells captured by debris are also discharged, and a reduction in the recovery rate of fat-derived cells can be prevented.

In the said invention, the said solution may be acidic.
By doing in this way, the association of collagen can be prevented and debris can be efficiently dissolved.

Moreover, in the said invention, the said container accommodates the said fat tissue and the digestive enzyme liquid which digests this fat tissue, and the said injection | pouring means heats the said solution to a temperature higher than the said digestive enzyme liquid. A heating unit may be provided, and the solution may contain a digestive enzyme having a higher concentration than the digestive enzyme solution.
By doing in this way, the collagen which was not fully decomposed | disassembled by the digestive enzyme liquid at the time of digestion of an adipose tissue can be decomposed | disassembled more reliably by a solution.

Moreover, in the said invention, it is preferable that the said solution contains at least 1 among collagenase, dispase, trypsin, pepsin, and hyaluronidase.
By doing in this way, collagen can be dissolved efficiently.

Moreover, in the said invention, you may provide the determination means which determines that the collagen derived from the said fat tissue capture | acquired by the said filter was melt | dissolved with the said solution.
In this way, it can be quickly determined that collagen has been dissolved and clogging of the filter has been eliminated.

The present invention also includes the adipose tissue processing apparatus according to any one of the above provided with a stirring mechanism for stirring the inside of the container, and the fat-derived cells discharged from the discharge port of the container of the fat tissue processing apparatus. Provided is a cell separation device comprising a concentration unit for centrifuging and concentrating a cell suspension, and a liquid supply path connected to the discharge port for supplying the cell suspension to the concentration unit.
According to the present invention, when adipose tissue is mixed with a digestive enzyme solution and digested in a container of an adipose tissue processing apparatus, and the resulting cell suspension is sent to a concentration unit through a solution feeding path and centrifuged, A cell mass of fat-derived cells can be obtained.

  In this case, even if the filter of the container is clogged with debris during the transfer of the cell suspension from the container to the concentration section, it is detected by the pressure detection section as a drop in pressure, and the debris is dissolved by the lysis solution. The Thereby, the cell suspension can be smoothly discharged from the container while the fat-derived cells trapped in the debris are also discharged to the concentrating portion to prevent a reduction in the recovery rate of the fat-derived cells.

  According to the present invention, it is possible to smoothly discharge a cell suspension while preventing a reduction in the recovery rate of fat-fat-derived cells.

1 is an overall configuration diagram of an adipose tissue processing apparatus according to an embodiment of the present invention. It is a whole block diagram of the cell separation device concerning one embodiment of the present invention.

An adipose tissue processing apparatus 1 and a cell separation apparatus 100 according to an embodiment of the present invention will be described below with reference to FIGS. 1 and 2.
As shown in FIG. 1, the adipose tissue processing apparatus 1 according to this embodiment is connected to a processing container 2 having a discharge port 2a, a filter 3 provided in the processing container 2, and a discharge port 2a. A valve V1 and a liquid feed pump 5 provided in the middle of the liquid feed tube 4, a shaker (stirring mechanism) 6 for shaking the processing container 2, and a pressure sensor 7 for detecting the pressure in the processing container 2. The injection means 8 for injecting the solution C from the discharge port 2a into the processing container 2 and the controller 9 for controlling the valve V1 and the injection means 8 are provided.

  The processing container 2 has a cylindrical shape, and a bottom surface 2b is inclined in one direction inside, and a discharge port 2a is opened at the lowest position. The processing container 2 contains adipose tissue A collected from the body and a digestive enzyme solution B that digests the adipose tissue A. As the digestive enzyme solution B, for example, a neutral solution such as collagenase, dispase, trypsin, pepsin or hyaluronidase is used.

The filter 3 has a hole having a diameter larger than that of the fat-derived cell D and smaller than that of the fat tissue A, and is disposed substantially horizontally above the discharge port 2a to partition the inside of the processing container 2 in the vertical direction. As a result, the adipose tissue A is restricted from being discharged from the discharge port 2a.
The valve V1 is provided in the vicinity of the discharge port 2a, and the liquid feed pump 5 is disposed with the valve V1 interposed between the valve V1 and the discharge port 2a.

The shaker 6 includes a base 6a on which the processing container 2 is placed, and a motor 6b that translates and rotates the base 6a around an eccentric axis in the vertical direction.
The base 6a has a cylindrical shape that is open at the top and supports the side surface and the bottom surface of the processing container 2, and is digested in the processing container 2 placed on the base 6a by a sheet heater 6c provided on the bottom surface. The enzyme solution B is kept at about 37 ° C. The motor 6b translates and rotates a shaft 6d fixed perpendicularly to the bottom surface of the base 6a along a predetermined circumferential orbit in a horizontal plane.

  When the motor 6b is driven, the processing container 2 on the base 6a is shaken in a horizontal plane, the adipose tissue A is agitated and digested in the digestive enzyme solution B, and is derived from the fat contained in the adipose tissue A. Cell D separates. After this, when shaking is stopped and the processing container 2 is allowed to stand, the digested adipose tissue A is upward, the cell suspension containing the fat-derived cells D in the digestive enzyme solution B is downward, and the specific gravity is lower. The layers are separated according to the difference. When the valve V1 is opened in this state and the liquid feeding pump 5 is operated, the cell suspension is discharged from the discharge port 2a into the liquid feeding tube 4, and the adipose tissue A is captured by the filter 3 and remains in the processing container 2. It is supposed to be.

  The pressure sensor 7 is provided on the side wall of the processing container 2 between the filter 3 and the discharge port 2a. The pressure sensor 7 detects the pressure in the processing container 2 while discharging the cell suspension from the discharge port 2 a, and outputs information on the detected pressure to the control unit 9.

The injection means 8 includes a storage portion 8a that stores a solution C for dissolving collagen, and a plug heater (heating portion) 8b that heats the storage portion 8a to a temperature higher than 37 ° C.
The accommodating portion 8a is connected to a branch tube 8c branched from the liquid feeding tube 4 between the discharge port 2a and the valve V1, and the valve V2 and the discharge port 2a from the accommodating portion 8a are located in the middle of the branch tube 8c. An infusion pump 8d that feeds in the direction is provided.

  The solution C contains a proteolytic enzyme that degrades collagen, for example, collagenase, dispase, trypsin, pepsin, hyaluronidase, etc. at a higher concentration than the digestive enzyme solution B, like the digestive enzyme solution B. . Further, the solution C is adjusted to be acidic, for example, pH 5 or lower.

  The controller 9 is connected to the pressure sensor 7, the valves V1 and 2, and the pumps 5 and 8d. At the time of discharging the cell suspension, the control unit 9 sends only the discharge time required for discharging the cell suspension calculated based on the volume of the digestive enzyme solution B in the processing container 2 and the flow rate of the liquid feeding pump 5. The liquid pump 5 is actuated. When the pressure inputted from the pressure sensor 7 becomes smaller than a predetermined threshold before the operation time of the liquid feeding pump 5 reaches the discharge time, the control unit 9 closes the valve V1 and stops the liquid feeding pump 5. Thereafter, the valve V2 is opened to operate the infusion pump 8d. At this time, the amount of the solution C fed by the injection pump 8d is set so that the solution C is filled up to the height position of the filter 3 in the processing container 2.

  Further, the controller 9 opens the valve V1 and operates the liquid feed pump 5 again until the discharge time is reached when a predetermined time has elapsed after the solution C has been fed. Yes. The predetermined time is set to a time sufficiently shorter than the time required for digesting the adipose tissue A, for example, several minutes, while the collagen contained in the debris captured by the filter 3 is sufficiently dissolved by the solution C. The

As shown in FIG. 2, the cell separation device 100 according to the present embodiment includes an adipose tissue processing apparatus 1, a concentration unit 20 that centrifuges and concentrates the fat-derived cells D, a processing container 2, a concentration unit 20, A liquid supply path 30 for connecting the cleaning liquid bag 10 and the waste liquid bag 11 is provided.
The concentration unit 20 includes a centrifuge 12 to which a centrifuge container 12a is attached. A pipe 12b is inserted into the centrifuge container 12a, and an open front end surface of the pipe 12b is disposed near the bottom surface of the centrifuge container 12a and facing the bottom surface.

The cleaning solution bag 10 contains a lactated Ringer solution (LR solution) that is a cleaning solution for the fat-derived cells D.
The liquid supply path 30 extends from the discharge port 2 a of the processing container 2, and is arranged at the other end of the pipe 12 b, the liquid supply tube 4 connected to the cleaning liquid bag 10 and the waste liquid bag 11, and an intermediate position of the liquid supply tube 4. And the discharge pump 13 are provided. The path through which the liquid is fed is selected by opening and closing the valves V1 and V3 to 5 and switching between the liquid feeding pump 5 and the discharge pump 13.

  When the discharge of the cell suspension from the processing container 2 to the centrifuge container 12a is completed, the control unit 9 rotates the centrifuge 12 and centrifuges the cell suspension into a precipitate mass and a supernatant of fat-derived cells D. To do. Subsequently, the control unit 9 opens the valves V4 and 5 and operates the discharge pump 13 to discharge the supernatant in the centrifuge container 12a to the waste liquid bag 11, and then opens the valves V3 and 4 for delivery. The liquid pump 5 is operated to supply the LR liquid to the centrifuge container 12a, and the centrifuge is performed. At this time, the LR liquid is supplied from the tip of the pipe 12b toward the fat-derived cells D aggregated at the bottom of the centrifuge container 12a, and the fat-derived cells D are stirred in the LR liquid. . The supply of LR liquid, centrifugation, and discharge of the supernatant are repeated several times, and the fat-derived cells D are washed in the centrifuge container 12a.

Operations of the adipose tissue processing apparatus 1 and the cell separation apparatus 100 configured as described above will be described below.
When the processing vessel 2 is shaken and the adipose tissue A is digested with the digestive enzyme solution B, the valves 1 and 4 are opened, the liquid feeding pump 5 is operated, and the cell suspension is discharged to the centrifuge vessel 12a.

When the filter 3 is clogged during the discharge and the cell suspension is not discharged, the pressure in the space partitioned downward by the filter 3 in the processing container 2 is reduced by the suction of the liquid feed pump 5, and the injection means 8 Solution C is injected and filled with solution C. After a while, the discharge of the cell suspension is resumed, and when the cell suspension is sufficiently discharged from the processing container 2, the liquid feed pump 5 is stopped.
Subsequently, the fat-derived cells D are washed in the concentration unit 20 to remove the digestive enzyme solution B and the lysate C, and when the cell mass precipitated at the bottom in the centrifuge container 12a is collected together with a small amount of supernatant, A concentrated solution of fat-derived cells D with high purity can be obtained.

  Thus, according to the present embodiment, even if the filter 3 is clogged by debris during the discharge of the cell suspension and is not discharged, it is quickly detected by the pressure sensor 7 and is dissolved by the solution C. Clogging is eliminated. Thereby, there exists an advantage that a cell suspension can be fully and smoothly discharged | emitted from the inside of the processing container 2. FIG. In addition, since the fat-derived cells D trapped in the collagen layer formed on the surface of the filter 3 are also released, the fat-derived cells D are prevented from being left in the processing container 2, and the fat-derived cells D There is an advantage that a reduction in recovery rate can be prevented.

  In addition, by making the solution C acidic, re-association of the degraded collagen can be prevented, and debris can be more reliably dissolved. Furthermore, by making the temperature and concentration of the solution C higher than that of the digestive enzyme solution B, collagen that has not been sufficiently decomposed during digestion of the adipose tissue A is also reliably and rapidly decomposed, effectively clogging the filter 3. Can be resolved. Moreover, the influence of the lysate C on the fat-derived cells D can be suppressed by washing and quickly removing the lysate C sent to the centrifuge container 12a together with the cell suspension.

In the above invention, the discharge of the cell suspension is resumed when a predetermined time has elapsed after injecting the lysis solution C into the processing container 2. Instead, a transmission light sensor (determination means) that measures the transmittance in the vicinity of the discharge port 2a is provided, and when the transmittance detected by the transmission light sensor decreases, the discharge of the cell suspension is resumed. It is good as well.
The transmitted light sensor includes, for example, a light emitting unit that emits detection light and a light receiving unit that receives detection light, which are arranged in the radial direction in the vicinity of the discharge port 2a. The transmittance in the vicinity of the discharge port 2a is detected from the intensity of the detection light detected by the light receiving portion through the light.

  When the debris that has clogged the filter 3 is lysed by the lysis solution C, red blood cells and adipose tissue A residues contained in the debris, fat-derived cells D, and cells that have been dammed above the filter 3 The suspension passes through the filter 3 and the transmittance in the vicinity of the discharge port 2a decreases. Thereby, it can be determined more reliably and promptly that the clogging of the filter 3 has been eliminated. Moreover, after the clogging of the filter 3 is eliminated, the fat-derived cells D are prevented from being unnecessarily brought into contact with the lysis solution C in the processing container 2, and damage to the fat-derived cells D can be reduced.

DESCRIPTION OF SYMBOLS 1 Adipose tissue processing apparatus 2 Processing container (container)
2a Discharge port 2b Bottom surface 3 Filter 4 Liquid feed tube 5 Liquid feed pump (pump)
6 Shaker (stirring mechanism)
6a base 6b motor 6c seat heater 6d shaft 7 pressure sensor (pressure detector)
8 Injection means 8a Accommodating part 8b Plug heater (heating part)
8c Branch tube 8d Infusion pump 9 Control unit 10 Washing solution bag 11 Waste solution bag 12 Centrifuge 12a Centrifugal container 12b Pipe 13 Discharge pump 20 Concentration unit 30 Fluid delivery path 100 Cell separation device A Adipose tissue B Digestive enzyme solution C Lysate D Fat-derived cells V1-V5 valve

Claims (7)

A container having a discharge port;
A filter that covers the discharge port in the container, partitions the container, captures fat tissue, and passes fat-derived cells separated from the fat tissue;
A valve for opening and closing the outlet;
A pump connected to the outlet and sucking the inside of the container;
An injection means for holding a solution for dissolving collagen and injecting the solution into a space including the discharge port partitioned by the filter;
A pressure detector provided in the vicinity of the discharge port for detecting the pressure in the container;
A fat tissue processing apparatus comprising: a control unit that controls the valve and the injection unit based on a pressure detected by the pressure detection unit while discharging the liquid from the discharge port.   The adipose tissue processing apparatus according to claim 1, wherein the solution is acidic. The container contains the adipose tissue and a digestive enzyme solution for digesting the adipose tissue;
The adipose tissue processing apparatus according to claim 1, wherein the injection unit includes a heating unit that heats the lysate to a temperature higher than that of the digestive enzyme solution. The container contains the adipose tissue and a digestive enzyme solution for digesting the adipose tissue;
The adipose tissue processing apparatus according to claim 1, wherein the lysing solution contains a digestive enzyme having a higher concentration than the digestive enzyme solution.   The adipose tissue processing apparatus according to claim 1, wherein the lysate contains at least one of collagenase, dispase, trypsin, pepsin, and hyaluronidase.   The adipose tissue processing apparatus according to claim 1, further comprising a determination unit that determines that the collagen derived from the adipose tissue captured by the filter is dissolved by the lysis solution. The adipose tissue processing apparatus according to any one of claims 1 to 6, further comprising a stirring mechanism for stirring the inside of the container;
A concentration unit for centrifuging and concentrating the cell suspension containing the fat-derived cells discharged from the discharge port of the container of the adipose tissue processing apparatus;
A cell separation apparatus comprising: a liquid supply path that is connected to the discharge port and supplies the cell suspension to the concentration unit.

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