WO2016204230A1 - Procédé d'administration de cellules souches, procédé pour améliorer les symptômes de chevaux de course ou d'exhibition, récipient pour injection et ensemble d'injection de cellules souches - Google Patents

Procédé d'administration de cellules souches, procédé pour améliorer les symptômes de chevaux de course ou d'exhibition, récipient pour injection et ensemble d'injection de cellules souches Download PDF

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WO2016204230A1
WO2016204230A1 PCT/JP2016/067943 JP2016067943W WO2016204230A1 WO 2016204230 A1 WO2016204230 A1 WO 2016204230A1 JP 2016067943 W JP2016067943 W JP 2016067943W WO 2016204230 A1 WO2016204230 A1 WO 2016204230A1
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stem cell
racehorse
stem
stem cells
cells
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PCT/JP2016/067943
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English (en)
Japanese (ja)
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仁幸 小林
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仁幸 小林
株式会社アニマルステムセル
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Priority claimed from JP2015144559A external-priority patent/JP2018111649A/ja
Priority claimed from JP2015144561A external-priority patent/JP2018111650A/ja
Priority claimed from JP2015144560A external-priority patent/JP2018110604A/ja
Application filed by 仁幸 小林, 株式会社アニマルステムセル filed Critical 仁幸 小林
Priority to JP2017525296A priority Critical patent/JPWO2016204230A1/ja
Publication of WO2016204230A1 publication Critical patent/WO2016204230A1/fr

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
    • A61K35/28—Bone marrow; Haematopoietic stem cells; Mesenchymal stem cells of any origin, e.g. adipose-derived stem cells
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
    • A61K35/48—Reproductive organs
    • A61K35/50—Placenta; Placental stem cells; Amniotic fluid; Amnion; Amniotic stem cells
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
    • A61K35/48—Reproductive organs
    • A61K35/51—Umbilical cord; Umbilical cord blood; Umbilical stem cells

Definitions

  • the present invention relates to a stem cell administration method for administering stem cells to a racehorse or a racehorse, a symptom improvement method for a racehorse or a racehorse, and an injection container and a stem cell injection set used therein.
  • Patent Document 1 discloses an invention for a therapeutic agent for atopic dermatitis comprising human mesenchymal stem cells as an active ingredient.
  • Patent Document 2 also includes a step of culturing stem cells isolated from equine amniotic fluid in a medium and recovering the cultured cells, exhibiting predetermined characteristics and improved homogeneity compared to stem cells before culture.
  • An invention of a method for producing aquatic amniotic fluid-derived multipotent stem cells is disclosed. It is described in the Example of the specification of the said literature that the test which confirms the differentiation ability of the said amniotic fluid origin multipotent stem cell to a chondrocyte, an osteocyte, and an adipocyte was implemented.
  • racehorses suffer from injuries and illnesses (hereinafter also referred to as injuries) peculiar to racehorses.
  • flexor tendinitis is a disease specific to racehorses with a high incidence.
  • Flexor tendinitis means inflammation that has occurred in the flexor tendon following the muscle that flexes the limb (flexor muscle) of the tendon tissue connecting the bone and the muscle.
  • racehorses that have been confirmed to have flexor tendinitis symptoms are treated such that the affected area is cooled and rested. Effective treatment and prevention methods for flexor tendonitis other than these countermeasures have not been established yet.
  • Broken bones are another serious injury for racehorses.
  • the tendonitis and fractures described above are examples of injuries that may occur in racehorses, and are not limited to these. Prevent the occurrence of injuries in racehorses, and fully treat injuries that occur. There is a strong demand for. In contrast, administration of stem cells to racehorses is expected to satisfy at least a part of the above requirements, but no specific research has been conducted so far.
  • Patent Document 2 discloses an invention of a method for producing amniotic fluid-derived multipotent stem cells of equine animals, which is useful for treating bone, tendon, and muscle damage and deletion diseases such as racehorses.
  • the present invention provides a method for administering a stem cell to a racehorse or a racehorse, and a method for improving a symptom of a racehorse by administering a stem cell to a racehorse or a racehorse.
  • the stem cell administration method of the present invention is an administration step in which a stem cell-containing solution containing a stem cell collected from an animal or a subculture stem cell obtained by subculturing the stem cell is injected into a racehorse or racehorse with an injection needle. It is characterized by including.
  • the symptom improvement method of the racehorse or racehorse of the present invention is characterized by using the stem cell administration method of the present invention and administering a stem cell-containing solution containing stem cells to a racehorse or racehorse showing an abnormal state. .
  • a stem cell can be administered to a racehorse or a racehorse, and the effect of administration of the stem cell can be significantly provided to the racehorse or racehorse.
  • symptoms such as various injuries of racehorses or racehorses are improved, or the occurrence of injuries or the like is prevented and serious injuries or the like occur. It is possible to prevent this.
  • a racehorse to which stem cells are administered by the stem cell administration method of the present invention means a horse for a racehorse and is registered as a racehorse. This includes not only horses that have been trained, but also those that are still being trained and can be registered as racehorses.
  • racehorse refers to a horse that can participate in a competition to conduct a competition.
  • the racehorse includes a racehorse having a pedigree and registered with pedigree information and a racehorse registered without pedigree information.
  • pedigree the age of racehorses that can participate in the Olympics, national corps, and international games is defined as 10 years old or older, and the present invention is applied to racehorses to improve various symptoms and maintain good health. It is very effective to do.
  • the term “competition” includes horse riding competition, equestrian competition, cattle chase, rodeo, polo, holter, and the like, and a plurality of racing horses participate to compete for superiority and inferiority from various viewpoints.
  • a stem cell includes either a stem cell itself collected from an animal or a passage stem cell obtained by subculturing a stem cell collected from an animal or a mixture thereof.
  • the stem cells may be collected from an animal or subcultured, and then immediately prepared as a stem cell-containing solution, or may be prepared as a stem cell-containing solution after being cryopreserved and thawed.
  • the stem cell administration method in the present embodiment includes an administration step of injecting a stem cell-containing solution containing a stem cell collected from an animal or a passage stem cell obtained by subculturing the stem cell into a racehorse with an injection needle. .
  • stem cells can be administered to a racehorse to be administered by a simple technique of injection using an injection needle, rather than a technique of incising and embedding in the body.
  • the injection widely includes procedures generally known as injection, such as intraarterial injection, intravenous injection, subcutaneous injection, and intramuscular injection. According to the above stem cell administration method, the burden on the racehorse due to the procedure of stem cell administration can be avoided.
  • the racehorse administered with the stem cell administration can run in a race at a relatively short interval of about 1 week or 2 weeks.
  • racehorses are large animals, unlike pet animals brought to the animal hospital due to illness, and mice used in hospitals or laboratories, so it is not easy to move.
  • Frequent reciprocation between a facility for preparing stem cells (hereinafter also referred to as a preparation facility) and a horse for administration of stem cells may cause mental fatigue to the racehorse. Therefore, there is a demand for maintaining the health of racehorses, preventing injuries, treating them, etc. at the daily residence, such as a horse or a hospital near the horse (hereinafter also referred to as a horse).
  • the stem cell administration method in the present embodiment includes an administration step of injecting a stem cell-containing solution into a racehorse with an injection needle. Details of the administration process will be described below.
  • the administration process does not particularly limit the person who performs the administration, the administration location, and the like.
  • the administration process does not require a procedure such as incision, and since the stem cells are administered by a simple method called injection, the administration process can be performed in any place that is not a sanitary facility such as a horse. .
  • animals from which stem cells are collected include all mammals.
  • animals from which stem cells are collected may be referred to as “donor animals” as appropriate.
  • the method of administering own stem cells has been common, but in recent years, so-called solid-to-solid administration is possible in which stem cells collected from one animal are administered to another animal. Therefore, rather than administering the racehorse's own stem cells to the racehorse, mammals in general are used as donor animals.
  • preferable examples of animals include horses.
  • Stem cell-containing liquid prepared on the premise that it is administered to a to-be-administered racehorse contains stem cells collected from an equine animal, so that the biocompatibility or affinity to the to-do racehorse is good.
  • the equine family animals include horses, donkeys, zebras, and donkeys.
  • the equine animal is a racehorse.
  • a racehorse that is a donor animal may be referred to as a “donor racehorse” as appropriate.
  • the donor racehorse means a racehorse from which stem cells are collected, and includes not only an active racehorse but also a horse for a racehorse before debut and a horse that has retired from a racehorse. From the viewpoint of maintaining the pedigree of the racehorse to be administered, it is important that the stem cells used have a clear pedigree of the donor racehorse, and whether the donor racehorse is an active racehorse or not. This is because it is not a problem.
  • the donor racehorse that is a racehorse from which stem cells are collected and the administered racehorse to which the stem cells are administered are of the same breed or sibling.
  • racehorses tend to have the property of being inherited by pedigree, and the properties of the previous generation (ie, parents) are assumed to be inherited by over 30%. Yes. Lineage is an important factor in racehorses. Accordingly, providing stem cells collected from the same race or sibling donor racehorse to the administered racehorse may have a desirable effect in terms of maintaining the race of the administered racehorse. Conventionally, there has been no technical idea of maintaining pedigree by providing stem cells collected from donor racehorses that are the same pedigree or siblings to the administered racehorse, and for the first time in the present invention its effectiveness is not reported. A possible provision method is shown with the suggestion.
  • FIG. 1 showing the pedigree of the racehorse X to be administered is shown.
  • FIG. 1 is a diagram showing the pedigree from the first ancestor to the fourth ancestor of the racehorse X to be administered, and is a four-generation pedigree table.
  • the same pedigree means that the donor racehorse is an ancestor of the administered racehorse X, or the donor racehorse and the administered racehorse X are the same racehorse.
  • the donor racehorse has the same pedigree within the range of the ninth generation or less of the administered racehorse X, and more preferably the same pedigree within the range of the fifth generation or less.
  • the donor racehorse is the horse name AB in FIG. 1
  • the donor racehorse is the father of the father of the administered racehorse X, the father of the third generation, and the administered racehorse X
  • brothers related to racehorses include all brothers whose parents are the same, and half brothers whose father or mother is the same.
  • Brothers include sisters, siblings, siblings, and twins.
  • a horse name means the official name of the said racehorse registered when participating in a horse race as a racehorse. Race horses are obliged to register their horse names. In Japan, for example, names that have passed the horse name examination by a specified organization (Japan Studbook International as of April 2015) are registered as official horse names. Is done. In order to make it possible to confirm whether the donor racehorse and the racehorse are of the same breed, the stem cell-containing solution used in the administration process is the name of the donor racehorse that became the donor of the stem cells contained therein. It is preferable that can be confirmed.
  • the stem cell-containing solution is contained in a container in which the horse name information of the donor racehorse is recorded, or the stem cell-containing solution is contained. It is preferable that an accessory with horse name information is attached to the container.
  • the horse name information is information for the person using the stem cell-containing solution to know the horse name of the donor racehorse, for example, the horse name itself or a number that can be collated with a separately created horse name database. And so on.
  • the stem cells used in this embodiment are stem cells collected from animals or passage stem cells obtained by subculturing the stem cells.
  • a stem cell is a cell having self-renewal ability and multipotency (ability to differentiate into various cells).
  • the stem cells used in this embodiment are not particularly limited as to which tissue of the animal was collected.
  • the stem cells include adipose-derived stem cells (adipose stem cells), umbilical cord, umbilical cord blood or placenta (hereinafter also referred to as umbilical cord etc.)-Derived stem cells (umbilical cord stem cells), bone marrow-derived stem cells (bone marrow stem cells), and the like. Yes, but not limited to this.
  • stem cells mesenchymal stem cells, hematopoietic stem cells, neural stem cells, embryonic stem cells, and the like are known because of their differentiation potential. Any one of these stem cells or a mixture of two or more thereof may be used as the stem cells used in the present embodiment.
  • the stem cells may include mesenchymal stem cells.
  • mesenchymal stem cells can be involved in the reconstruction of many tissues in the body such as bones, blood vessels, muscles or tendons, and various effects are expected when administered to racehorses.
  • administration of mesenchymal stem cells is expected to improve the symptoms of racehorse injury and disease, maintain good physical condition, and strengthen tendons and internal organs.
  • a mesenchymal stem cell exists in a tissue having a mesenchymal tissue.
  • a bone marrow mesenchymal stem cell that is a mesenchymal stem cell collected from bone marrow, or a mesenchymal stem cell that is a mesenchymal stem cell collected from fat.
  • umbilical cord mesenchymal stem cells which are mesenchymal stem cells collected from umbilical cords and the like.
  • the stem cell-containing liquid in the present embodiment preferably contains mesenchymal stem cells, and the stem cells substantially contained in the stem cell-containing liquid are more preferably mesenchymal stem cells.
  • the stem cells include mesenchymal stem cells collected from a donor racehorse.
  • the stem cells in the present embodiment include fat-derived mesenchymal stem cells collected from fat collected from a racehorse (donor racehorse) or umbilical cord-derived mesenchymal stem cells collected from an umbilical cord or the like. Is preferred. Racehorses generally have a very low fat percentage of a few percent and it is not easy to collect fat compared to other animals.
  • stem cells containing mesenchymal stem cells fractionated from the fat of donor racehorses show a good growth rate in subsequent subcultures, and when administered to racehorses It was confirmed that the rate of colonization of the affected area was also high. Therefore, it is desirable from the standpoints of securing stem cells and the effect of administration to select adipose-derived mesenchymal stem cells collected from a donor racehorse as stem cells for administration to a racehorse to be administered.
  • umbilical cord stem cells collected from the umbilical cord or the like can be mentioned.
  • bone marrow stem cells or adipose stem cells are collected, appropriate medical practice is required, which may cause physical damage to donor animals.
  • the umbilical cord and the like can be obtained without requiring any special surgical action in giving birth to a donor animal. Therefore, there is no risk of causing physical damage to the donor animal in order to collect stem cells.
  • the donor's racehorse generally has a very low fat percentage of about several percent, it takes several weeks to heal the incision after incising the buttocks of the donor racehorse and collecting fat.
  • the umbilical cord connects the mother horse and the foal
  • the stem cells collected from the umbilical cord blood or the umbilical cord are defined as stem cells on the mother horse side. Therefore, in the present invention, the donor racehorse of stem cells collected from the umbilical cord or the like is the mother horse.
  • the method for collecting stem cells from animals is not particularly limited, and includes a method for collecting only stem cells from any tissue in animals, or a method for collecting stem cells together with any other biological component other than stem cells, and a known collection method Can be adopted as appropriate.
  • the biological component means a biological material of the animal such as a cell, a hormone or a blood component or a material composed of the biological material, and any other biological component other than the stem cell means a biological component excluding the stem cell.
  • the biological substance means a biological macromolecule (polysaccharide, nucleic acid, protein, etc.) constituting the living body, and various sugars, lipids, vitamins, hormones, etc. constituting the biological polymer.
  • the stem cell-containing solution may contain substantially only stem cells as an animal-derived component, or may contain stem cells and any other biological component.
  • the medium used for the stem cell subculture method is not particularly limited, but a serum-free medium is preferably used. Passage stem cells obtained by subculture using a serum-free medium are preferable when an animal that is a donor of stem cells and an administered animal are different individuals (that is, when administration between individuals is performed).
  • the subculture may be performed by seeding stem cells collected from animals, or may be seeded with stem cells after being cryopreserved and thawed.
  • the medium preferably contains human serum albumin, insulin, linoleic acid and transferrin, and does not contain any of ammonium metavanadate, phenol red and mercaptoethanol.
  • the medium does not contain any of ammonium metavanadate, phenol red, and mercaptoethanol, the occurrence of side effects due to administration of the stem cell-containing solution is suppressed.
  • the administration timing in relation to the time of the race to participate.
  • FIG. 2A is a flowchart showing a first example of the providing method. First, an outline of the providing method will be described.
  • the present providing method includes a cryopreservation step S1, a thawing step S2, a viable cell number determination step S3, a stem cell-containing agent preparation step S4, and a transport step S5.
  • decompression process S2, viable cell number determination process S3, stem cell containing agent preparation process S4, and transport process S5 are implemented in order of description.
  • the cryopreservation step S1 is a step of freezing stem cells collected from animals.
  • the thawing step S2 is a step of thawing the stem cells frozen in the cryopreservation step S1.
  • the viable cell number determination step S3 is a recommended recommended time from the predetermined time after the thawing step of the stem cells after the thawing step S2 until the stem cells are administered to the racehorse (administered racehorse X). This is a step of determining the absolute number of living cells using the survival rate at the time of progress and the desired number of living cells when administering stem cells to a racehorse (administered racehorse X).
  • “after the thawing step” includes a predetermined time range after the thawing step is performed, and immediately after the thawing step is performed, for example, the thawing step is performed, and then subculture is performed to perform the subculture.
  • the stem cell-containing agent preparation step S4 is a step of using the stem cells after the thawing step S2 to prepare a stem cell-containing agent 12 (see FIG. 4) containing stem cells that satisfy the absolute number of living cells.
  • the transport step S5 is a step of transporting the stem cell-containing agent 12 to an implementation area where administration is performed.
  • the viable cell number determination step S3 is performed after the cryopreservation step S1, and then the transport step S5 is performed, so that a desired number of viable stem cells can be reliably raced at the site of administration.
  • Administration to Horse X is possible. Accordingly, it is possible to administer the stem cells to the administered racehorse X at a desired location (for example, a horse) without moving the administered racehorse X from the horse etc. to the preparation facility.
  • This providing method is intended to prevent the racehorse X to be administered from reciprocating between the horse and the preparation facility, particularly when the race start interval is short or when multiple stem cell administrations are desired in a short period of time. In some situations, administration of stem cells is highly desirable and feasible.
  • the cryopreservation step S1 in this embodiment is a step of freezing stem cells collected from animals.
  • the form of the stem cell cryopreserved in the cryopreservation step S1 may be a state of a stem cell alone, but a collected material containing the collected stem cells, a stem cell-containing fraction obtained by centrifuging the collected material, etc. Any mixture, such as a subculture obtained by subculturing the collected stem cells, a stem cell-containing culture fraction obtained by centrifuging the subculture, etc. .
  • the stem cells in any state described above are cryopreserved in a state of being mixed in a cryopreservation solution containing an antifreeze solution, for example.
  • the cryopreservation solution can contain, for example, 10 to 20% by mass of dimethyl sulfoxide (DMSO) and 5 to 10% by mass of glycerol, but is not limited thereto.
  • DMSO dimethyl sulfoxide
  • the freezing temperature and freezing method in the cryopreservation step S1 are not particularly limited as long as the stem cells can be stored well for a desired period. “Saving well” means maintaining a high survival rate (for example, 70% or more and 95% or less) during or after storage.
  • a freezer having a set temperature of about ⁇ 20 ° C. or a deep freezer having a set temperature of about ⁇ 80 ° C. can be used.
  • a deep freezer with a set temperature of around ⁇ 80 ° C., and liquid nitrogen storage (for example, ⁇ 196 ° C.) is more preferable.
  • stem cells to be cryopreserved when liquid nitrogen storage (for example, ⁇ 196 ° C.) is performed, the stem cells are first stored in a deep freezer at ⁇ 80 ° C. for about 24 hours, and then frozen. It is preferable to transfer to a nitrogen tank.
  • a water bath for example, around 37 ° C. maintained at an appropriate temperature is used so that many stem cells are not killed during freezing.
  • the stem cells are preferably stem cells collected from an animal that has received stem cell administration in advance. Although the detailed mechanism is not clear, stem cells minimized from animals that have been administered stem cells are of good quality and have a high regenerative medical effect.
  • the stem cells may be stored in a state where the name of the donor racehorse that has become the donor of the cryopreserved stem cells can be confirmed.
  • the relationship of the pedigree of a donor racehorse and a to-be-administered racehorse X can be grasped
  • This providing method may include a pedigree verification step S10 as shown in FIG.
  • the pedigree collation step S10 collates the pedigree or sibling relationship between a donor racehorse that has become a cryopreserved stem cell donor and a racehorse (administered racehorse X) to be administered with a stem cell-containing agent, This is a step of determining a stem cell of a donor racehorse that is the same pedigree or sibling as a stem cell to be provided to a thawing step S2 described later.
  • the pedigree verification step S10 is preferably performed before the thawing step S2.
  • the cryopreservation step S1 can confirm the horse name information of the father racehorse and the mother of the donor racehorse that became the donor of the cryopreserved stem cells.
  • the stem cells may be stored in a state. For example, a lot number unique to the stem cell is attached to the cryopreserved stem cell container, and the name of the horse, father, and the name of the mother are the names of the donor racehorse that has become the donor of the stem cell. .
  • the pedigree verification step S10 can be appropriately performed by a means capable of verifying that the donor racehorse is an ancestor of the administered racehorse X or a sibling.
  • the pedigree verification step S10 can be executed by confirming that the donor racehorse is included in the ancestors shown in the pedigree table of the administered racehorse X shown in FIG.
  • the method for confirming the horse name of the donor racehorse in the pedigree table is not particularly limited, and the pedigree table may be confirmed visually or an information processing system may be used.
  • the information processing system for example, a database in which the pedigree relationship of the administered racehorse X is created, and the horse name input step for inputting the horse name of the donor racehorse, the horse name input in the horse name input step, You may use the horse name collation processing system which has the collation step which collates whether the same horse name exists in the said database, and the display step which displays the collation result by the collation step.
  • the umbilical cord connects the mare and the foal.
  • the stem cells collected from the umbilical cord and the like are defined as stem cells on the mother horse side. Therefore, in the present invention, the donor racehorse of stem cells collected from the umbilical cord or the like is the mother horse.
  • the thawing step S2 is a step of thawing the stem cells frozen in the cryopreservation step S1.
  • the thawing method is not particularly limited. For example, in the case of stem cells frozen and stored at about ⁇ 80 ° C. in a deep freezer, the stem cells are removed from the deep freezer and immersed in a water bath whose water temperature is adjusted to about 37 ° C. The kept state can be kept and thawed for several minutes. In the thawing step S2, consideration is given to thawing under conditions that do not damage the frozen stem cells as much as possible.
  • the thawing is performed under the same conditions as the preliminary experiment. Good.
  • the stem cells themselves thawed in the thawing step S2 are provided to the live cell number determining step S3.
  • the stem cell itself as used herein means a stem cell that has not been subcultured after being thawed in the thawing step S2.
  • the live cell number determination step S3 when the stem cell-containing agent provided by the present providing method is administered to the racehorse X to be administered, a desired number (the desired number of viable cells) of living stem cells are included in the stem cell-containing agent. It is a step of determining the absolute number of living cells for ensuring that they are contained. That is, the viable cell number determination step S3 is recommended from the predetermined time after the thawing step of the stem cells after the thawing step S2 until the stem cells are administered to the racehorse (administered racehorse X).
  • calculating process S302 is implemented.
  • the recommended time which is the recommended time from the predetermined time to the time when the stem cell-containing agent provided by the present providing method is administered to the racehorse X to be administered.
  • the number and a specific calculation step S302 for determining the number will be described.
  • the survival rate of stem cells after 48 hours from the predetermined time is about 70% with respect to 100% of the number of living cells (number of viable cells after the thawing process) at the predetermined time, and administration to the racehorse X to be administered is desired.
  • the desired number of living stem cells is 10 million
  • the absolute number Z of living cells is about 14.29 million according to the following calculation formula (1).
  • the calculation step S302 can obtain the absolute number of living cells by dividing the desired number of living cells by the survival rate and multiplying by 100.
  • Absolute number of living cells Z (10,000) 1000 (10,000) ⁇ 70 (%) ⁇ 100 (1)
  • the survival rate at the time when the recommended time elapses can be grasped by previously measuring the survival rate after an appropriate time has elapsed from a predetermined time of the stem cells to be used.
  • Stem cells donated to the steps after the live cell number determination step S3 should be handled in a temperature environment suitable for the survival of the stem cells as much as possible (eg, 0 ° C. or more and 10 ° C. or less, preferably about 4 ° C. ⁇ 3 ° C.).
  • the number of living cells of the stem cells managed in the temperature environment at an appropriate time is measured, and the appropriate number of living cells at the predetermined time (the number of living cells after the thawing step) is determined. From the ratio of the number of viable cells over time, the survival rate over time can be calculated. And among the appropriate time mentioned above, the time when sufficient survival rate was confirmed can be made into recommended time.
  • a general stem cell collected from a horse has a survival rate of a stem cell stored in a refrigerated environment from a predetermined time after the thawing step S2. It is known that about 80%, about 70% after 48 hours, and about 60% after 72 hours.
  • the refrigerated environment means a temperature environment exceeding 0 ° C. and not more than 10 ° C.
  • the “predetermined time” refers to a time when the number of viable cells after the thawing step is confirmed by an arbitrary method in the viable cell number determining step S3.
  • the predetermined time is the start of calculating the recommended time until administration of the stem cell-containing agent.
  • the predetermined time does not necessarily have to be obtained until the number of viable cells after the thawing step is exactly matched with the time when the number of viable cells is confirmed by an arbitrary method in units of seconds or minutes.
  • the viable cell number after the thawing step means the viable cell number of the stem cell provided to the viable cell number determining step S3 through the thawing step S2.
  • the viable cell number after the thawing step is immediately supplied to the viable cell number determining step S3 without subculture after the thawing step S2. It is the number of living stem cells.
  • the present providing method includes a cryopreservation step S1 and a thawing step S2 as a previous step of the live cell number determination step S3.
  • the number of living stem cells (number of living cells after the thawing step) provided to the number of living cells determination step S3 after the thawing step S2 is confirmed, and the time when the number is confirmed is called a predetermined time.
  • the method for confirming the number of viable cells after the thawing step is not particularly limited as long as it is a method capable of grasping the viable cell number of the stem cells provided in the viable cell number determining step S3.
  • the number of living cells after the thawing step S301 is further included in the number of living cells determination step S3.
  • the live cell number determining step S3 measures the number of living cells of the stem cells thawed by the thawing step S2, and determines the number of viable cells after the thawing step. It further includes an afterlife cell number measurement step S301.
  • the viable cell count measurement step S301 after the thawing step, the accuracy of the viable cell count contained in the stem cell-containing agent prepared in the stem cell-containing agent preparation step S4 described later is increased.
  • the confirmation method in the viable cell number determining step S3 is not limited to the viable cell number measuring step S301 after the thawing step.
  • FIG. 2B shows a second example of this embodiment that does not include the viable cell count measurement step S301 after the thawing step.
  • This example does not include the viable cell count measurement step S301 after the thawing step and has a cryopreserved viable cell count confirmation test S303.
  • the number of cryopreserved live cells is confirmed in advance by cryopreserving under the same temperature conditions as in the cryopreservation step S1 and confirming the viability of the number of viable stem cells thawed under the same thawing conditions as in the thawing step S2. Confirmation test S303 is performed.
  • the predetermined time in the second example is X minutes after thawing of the stem cells in the thawing step S2.
  • the number of living stem cells in the thawing step S2 may be grasped by a method other than the actual measurement of the number of living stem cells after the thawing step S2.
  • this invention includes this provision method including both cryopreservation viable cell number confirmation test S303 and the post-thaw process viable cell count measurement process S301 as a further modification. That is, the cryopreserved viable cell count confirmation test S303 is performed in advance, and the approximate number of viable cells after the cryopreservation step S1 and the thawing step S2 with respect to the viable cell count before the cryopreservation step S1 is grasped. In addition, after the thawing step S2, the number of viable cells at a predetermined time may be correctly measured by performing the post-thaw step viable cell number measuring step S301.
  • the stem cell-containing agent preparation step S4 is a step of preparing a stem cell-containing agent containing stem cells that satisfy the absolute number of living cells determined in the living cell number determining step S3 using the stem cells after the thawing step described above.
  • the present providing method has a cryopreservation step S1, a thawing step S2, and a transport step S5 described later, but has a desired number of living cells or more.
  • Stem cells can be administered to the racehorse X to be administered.
  • the stem cell satisfying the absolute number of living cells in the stem cell-containing agent preparation step S4 means a stem cell containing living cells that are more than the absolute number of living cells. That is, in the stem cell-containing agent preparation step S4, the stem cell content in the stem cell-containing agent is adjusted so that live stem cells greater than the absolute number of living cells are contained.
  • the number of viable cells after the thawing step confirmed in the viable cell number determining step S3 is Y 10,000 / ml
  • the absolute number Z of viable cells calculated by the above formula (1) is about
  • the stem cells contained in the stem cell-containing agent is (1429 ⁇ Y) ml or more.
  • the stem cell-containing agent means an agent that contains stem cells and can be administered to the racehorse X to be administered.
  • the stem cell-containing agent can be composed only of a stem cell and a medium containing the stem cell, but may contain the stem cell and the biological component other than the stem cell.
  • arbitrary additives other than a stem cell and the said medium may contain in the stem cell containing agent.
  • the stem cell-containing agent is not particularly limited.
  • the stem cell-containing agent is a liquid stem cell-containing agent in which stem cells are contained in an appropriate liquid such as Ringer's solution, a semi-solid stem cell-containing agent obtained by gelatinizing or gelling containing stem cells, or the above-mentioned semi-solid stem cells containing stem cells. It may be a solid stem cell-containing agent whose volume change is smaller than that of the containing agent. Since the liquid stem cell-containing agent can administer the stem cells by injection to the racehorse X to be administered, the administration work is easy, and after completion of the transportation process described later, the stem cells are promptly applied to the racehorse X to be administered. Can be administered.
  • the transporting step S5 is a step of transporting the stem cell-containing agent prepared in the stem cell-containing agent preparing step S4 to an implementation area where administration is performed.
  • Transportation in the transportation step S5 means movement using public facilities that can be used by the public, such as roads, railways, ships, and aviation.
  • Examples of available transportation means include, but are not limited to, mail and courier services, and various methods for transferring a stem cell-containing agent from a facility where stem cells are cryopreserved using the above-mentioned public facility. Widely encompasses the means.
  • “Implementation area” means a predetermined area including a predetermined place where administration is performed or a facility close to the place.
  • the predetermined area is an area that can be moved to a predetermined place where administration is performed without using the transportation facility described above. More specifically, for example, the implementation area is an area including a horse where administration is performed or a facility adjacent to the horse, and the stem cell-containing agent is delivered to the horse or the facility, for example, in the transport step S5. It is done.
  • the transporting step S5 is performed in a state in which a temperature environment exceeding 0 ° C. and not exceeding 10 ° C. is maintained. Thereby, it is possible to provide a high-quality stem cell-containing agent by suppressing the death and activity reduction of the stem cells in the transport step S5.
  • a courier service that can be managed at low temperature, which is a paid service of a private company, may be used.
  • the time spent for the transport step S5 is less than the recommended recommended time from the predetermined time described above until the stem cell-containing agent is administered to the racehorse X to be administered. More specifically, for example, the transportation process is preferably completed so that the stem cell-containing agent can be administered to the racehorse (administered racehorse X) in less than 72 hours from the predetermined time, for example, within 48 hours. More preferably, it is more preferably within 30 hours. According to the study of the present inventor, even when the stem cell-containing agent is maintained in a refrigerated storage state (stored at a temperature exceeding 0 ° C. and not exceeding 10 ° C.), if the storage time becomes too long, the survival rate is remarkable. It has been found that the dead stem cells may be altered and may have an undesirable effect on the administered racehorse X. In order to avoid this, it is preferable to set the time from the predetermined time to the completion of the transportation process within the above-described range.
  • the second container 40 different from the injection container 10 may be transported together with the injection container 10 containing the stem cell-containing agent 12 (see FIG. 4).
  • the second container 40 contains a live cell number confirmation sample 120 having the same composition as the stem cell-containing agent 12.
  • the number of living cells of the stem cell-containing agent 12 satisfies the number of desired living cells. .
  • the certainty and reliability of administration of the stem cell-containing agent 12 can be improved. For example, if the stem cell-containing agent 12 has fallen below the desired number of viable cells due to a transportation trouble in the transportation step S5, such as exceeding the planned transportation time and inadequate temperature control during transportation, etc. Administration of the stem cell-containing agent 12 can be avoided.
  • the measurement of the number of living cells of the confirmation sample 120 is performed after the transport step S5 and before the stem cell-containing agent 12 is administered to the racehorse X to be administered.
  • the above is not intended to prohibit the administration of the stem cell-containing agent 12 that has fallen below the desired number of living cells to the racehorse X to be administered.
  • the stem cell-containing agent 12 having a number lower than the desired number of living cells is administered, and administration performed thereafter It can be adjusted to increase the number of desired living cells at the time as appropriate.
  • confirmation sample 120 and the administration stem cell-containing agent 12 are accommodated in different containers, contamination of the stem cell-containing agent 12 at the time of measuring the number of viable cells before administration can be avoided. Therefore, even in an environment without a clean room such as a horse, the number of viable cells can be measured before administration with confidence.
  • stem cell injection set (stem cell providing set) 100 that is preferably used in the present embodiment will be described later.
  • the measurement of the number of viable cells performed using the confirmation sample 120 before the administration of the stem cell-containing agent 12 may be performed by an arbitrary method after the thawing step and the number of viable cells.
  • an arbitrary method for example, generally, there is a method in which a small amount of stem cells to be counted are collected, accurately diluted with a staining solution such as trypan blue, and the number of living cells and the number of dead cells are measured with a hemocytometer.
  • the stem cells diluted with trypan blue can be differentiated into dead cells (stem cells stained blue) and live cells (stained cells not stained) by color difference. The number of live cells read may be counted.
  • the present providing method the first mode in which the stem cell itself thawed in the thawing step S2 is subjected to the viable cell number determining step S3 is shown.
  • the method for providing the stem cell-containing agent is not limited to this, and the stem cells thawed in the thawing step S2 are subcultured, and the number of living cells is determined as the subcultured stem cells obtained by performing the subculture.
  • the 2nd aspect which uses for a process is included.
  • FIG. 3 is a flowchart showing an example of a method for providing a stem cell-containing agent according to the third example of the present invention.
  • the present providing method in the third example performs a subculture process S30 in which subculture is performed using the thawed stem cells after the thawing process S2 and before the viable cell number determination process S3. It differs from the first embodiment in that a subcultured stem cell, which is a stem cell obtained by subculture, is provided to the live cell number determination step S3. Regarding the third example, for the same steps as in the first embodiment, the description of the first embodiment is appropriately referred to.
  • the predetermined time is after the thawing process S2 and when the number of viable cells of the subcultured stem cells after the subculture process S30 is performed is confirmed. It is.
  • the second aspect is effective, for example, when the storage period in the cryopreservation step S1 is sufficiently long. That is, stem cells that have been in a frozen state for a long period of time can be refreshed by subculture after thawing and sufficiently regain their original activity.
  • the second aspect is also effective when there are few stem cells stocked in the cryopreservation step S1. This is because the number of stem cells can be increased to a desired number in the subculture, and the subcultured stem cells whose number has been increased in the subculture can be provided to the cell number determination step S3.
  • the first aspect described above is excellent in that the stem cells stored in the cryopreservation step S1 can be promptly provided when it is necessary to administer the stem cells to the racehorse X to be administered.
  • the post-thaw step viable cell number measurement step S301 is carried out and provided to the live cell number determination step S3.
  • the number of viable cells may be measured after the thawing step of the subcultured stem cells. This is because a stem cell-containing agent containing stem cells satisfying the desired viable cell count is prepared in the stem cell-containing agent preparation step S4 by correctly grasping by measuring the number of viable cells after subculture.
  • the viable cell number of the subcultured stem cells provided to the viable cell number determination step S3 corresponds to the viable cell number after the thawing step.
  • the viable cell number measurement step S301 after the thawing step can be performed by measuring the viable cell number of the subcultured stem cells provided in the viable cell number determination step S3.
  • the number of viable cells after the thawing step can be grasped, and the accuracy of the number of viable cells contained in the stem cell-containing agent prepared in the subsequent stem cell-containing agent preparation step S4 can be increased.
  • stem cells in which the number of living stem cells to be administered is adjusted to a desired number by transportation to a different administration site from the preparation facility.
  • the stem cell-containing solution in the present embodiment contains the stem cells described above and is an agent that can be injected into the racehorse to be administered through an injection needle.
  • the stem cell-containing solution can be prepared, for example, by mixing an infusion preparation such as Ringer's solution or physiological saline with a stem cell or a cell solution containing a stem cell and adjusting the viscosity to an appropriate level.
  • the cell solution may be a liquid material containing stem cells. For example, a part of a culture solution obtained by subculturing stem cells collected from a donor animal, a frozen storage solution after thawing containing stem cells However, it is not limited to these.
  • the stem cell-containing solution is preferably prepared so as to have a low viscosity so that it can be injected from the syringe into the animal to be administered.
  • the stem cell-containing liquid in the present embodiment may substantially contain only stem cells as a biological material, and may contain any other biological component in addition to the stem cells.
  • the other biological components include cytokines.
  • the stem cell-containing liquid in the present embodiment may contain cytokines together with stem cells. Cytokines are physiologically active proteins produced from cells and are generally known to be low molecular weight molecules having a molecular weight of 80,000 or less, more often 30,000 or less.
  • the physiological activity means a property that a chemical substance acts on a specific physiological regulatory function in a living body.
  • the cytokine By administering a cytokine together with a stem cell to an administered animal, the cytokine can be suitably involved in the expression of the self-replicating ability or pluripotency of the stem cell in the body of the administered animal.
  • the cytokine may be produced from stem cells contained in the stem cell-containing solution or may be produced from different cells.
  • cytokines examples include, but are not limited to, chemokines, cell growth factors, interleukins, or interferons, or derivatives thereof.
  • the chemokine is a low molecular weight protein of about 8 kDa to about 14 kDa, which is roughly classified into CC chemokine, CXC chemokine, C chemokine or CX3C chemokine.
  • Chemokines act through chemokine receptors.
  • Preferable examples of the cytokine contained in the stem cell-containing solution include CXCL12 or a derivative thereof.
  • CXCL12 or a derivative thereof can favorably promote the expression of stem cell self-renewal or pluripotency in the body of an administered animal.
  • the stem cell-containing solution containing CXCL12 or a derivative thereof preferably contains CXCR4 or a derivative thereof which is a receptor for CXCL12. This is because CXCL12 administered together with the stem cells acts effectively.
  • CXCL12 administered together with the stem cells acts effectively.
  • stem cells administered to an animal to be administered have a homing effect that reaches an affected area that is inflamed directly or sequentially through organs such as lungs, spleen, and liver.
  • the homing effect can be improved by administering CXCL12 and its receptor CXCR4 together with stem cells.
  • the injection needle used in the present embodiment is a member that can be stabbed into the body from the body surface of the racehorse to be administered and a stem cell-containing solution can be administered to the racehorse to be administered through a flow passage formed inside the injection needle.
  • the injection needle is a small-diameter cylindrical body having a tapered tip and a transverse cross section formed in an annular shape, and a flow path through which a stem cell-containing liquid can flow is extended in the longitudinal direction.
  • the injection needle is open at both ends in the longitudinal direction and has a flow path through which the stem cell-containing liquid can flow from one opening to the other opening.
  • the injection needle may be either a type that stabs the racehorse to be administered at the time of administration, or an indwelling needle that is stabbed in advance at any location of the racehorse to be administered.
  • the injection needle may be a member that can be connected to an injection cylinder (so-called syringe) or a member that can be connected to an infusion container via an infusion tube.
  • the infusion container includes an infusion bag and an infusion bottle. That is, the administration process of the present embodiment can be performed using a syringe having a syringe barrel and a syringe needle, or can be performed using an infusion device having an infusion container, an infusion tube, and an indwelling needle. .
  • the injection needle has an inner diameter of 0.6 mm or more.
  • an injection needle having an inner diameter in such a range it is possible to favorably avoid that stem cells passing through the injection needle are damaged by the injection pressure.
  • the injection amount of stem cells per unit time can be increased by using an injection needle having an inner diameter of 0.6 mm or more, the time required for injection can be shortened even if the injection rate is slowed down. The burden on the administration racehorse can be reduced.
  • the thickness of the injection needle is generally indicated by “gauge”. This indicates how much the outer diameter of the injection needle is 1 inch, for example, 23 gauge is 1/23 inch.
  • the outer diameter of the injection needle in the present embodiment is not particularly limited. In general, from the viewpoint that the inner diameter is determined according to the outer diameter, and that the inner diameter is preferably moderately thick as described above, the injection needle in the present embodiment is preferably 23 gauge or more. More preferably, it is more than gauge, more preferably more than 19 gauge.
  • the injection needle used in the present embodiment is not limited to a specific injection mode, and can be appropriately selected from injection needles used in a known injection technique.
  • the stem cell administration method of the present embodiment can be carried out by appropriately selecting from means known as injection, such as intravenous injection, intraarterial injection, subcutaneous injection, intramuscular injection, using an injection needle.
  • the means for injecting the stem cell-containing solution into the racehorse (administered racehorse) with an injection needle is preferably intravenous injection. Since direct injection into the affected area (eg, tendon) may damage the affected area, it is necessary to rely on the skill of the operator performing the injection. On the other hand, in the case of intravenous injection, the stem cell-containing liquid is once sent to the heart together with venous blood, and then depends on the skill of the operator by causing the stem cells to reach the inflamed site by the homing effect of the stem cells and the like. Without administration, the administration step can be desirably performed.
  • the stem cell administration method of this embodiment can give the outstanding effect which exceeded the effect of being only minimally invasive with respect to a racehorse by administering the stem cell containing liquid by intravenous injection with respect to a racehorse. That is, when a stem cell-containing solution is administered intravenously to a racehorse, stem cells contained in the stem cell-containing solution pass through organs such as the lungs, spleen, and liver and are consumed in part, and the rest is inflamed. Can reach the affected area.
  • the spleen is generally known as a lymph organ, but also has a function of storing a large amount of blood to be released into the circulating bloodstream during exercise or the like.
  • racehorses tend to have a larger spleen weight relative to their body weight than other animals (for example, cows) because blood that is consumed in large quantities during a horse racing game is stored in the spleen. Since the blood stored in the spleen of the racehorse is released and consumed in large quantities during the game, the spleen of the racehorse after the game is significantly fatigued, and a large amount of blood is promptly prepared for the next game. It is required to store blood.
  • the racehorse after the game tends to cause inflammation of tendons and muscles. That is, the racehorse undergoes a physical condition change not seen in other animals, in which spleen fatigue and tendon inflammation occur at the same timing.
  • the stem cells contained therein can be suitably applied to any of the inflamed parts such as the spleen and tendon.
  • FIG. 4 is an explanatory view showing an example of a stem cell injection set (stem cell providing set) 100 including the injection container 10.
  • the injection container 10 has a container body 16 that can contain the stem cell-containing liquid 12 and can be connected to an injection needle (not shown).
  • the injection container 10 may be referred to as a first container.
  • the injection container 10 is accommodated in a sealed container 32.
  • the container body 16 contains a stem cell-containing liquid 12 containing stem cells, and includes a sealing portion (cap 19) that prevents the stem cell-containing liquid 12 from leaking out of the injection container 10.
  • Stem cells contained in the stem cell-containing solution 12 in this embodiment include those collected from animals and subcultured stem cells obtained by subculturing stem cells collected from animals.
  • the container body 16 of the injection container 10 contains the stem cell-containing liquid 12 in advance, and the injection container 10 is provided with a sealing portion (cap 19). Therefore, according to the injection container 10 of the present embodiment, the stem cell-containing liquid 12 can be transported or transported in a state where it can be quickly injected. Further, since the injection container 10 is accommodated in the sealed container 32, the stem cell-containing liquid 12 can be transported or transported while maintaining a good hygiene condition. Therefore, it is possible to provide the stem cell-containing liquid 12 from the preparation facility to an arbitrary place (for example, a horse etc.) without being contaminated, and promptly administered by an easy method of injection at the provided place. The stem cell-containing solution 12 can be administered to animals.
  • the injection container 10 of the present embodiment it is possible to arbitrarily select a place where the stem cell-containing solution 12 is administered and to enable a person other than the above-mentioned operator to perform the administration operation.
  • the versatility of stem cell administration technology can be enhanced.
  • the injection container 10 is a container that has a container body 16 that can contain the stem cell-containing liquid 12 and can be connected to an injection needle. By connecting to a syringe needle (not shown), the stem cell-containing solution 12 contained in the container body 16 can be injected into the administration animal.
  • the injection container 10 is not limited to a syringe barrel, and widely includes containers that can be connected to an injection needle such as an infusion bag or an infusion bottle.
  • the stem cell-containing liquid 12 is accommodated in the injection container 10 as described above, it is easy and quick without requiring a high level of medical practice such as incision or the like of placing the stem cells in the body of an administered animal. Can be administered.
  • the stem cell-containing liquid 12 since the stem cell-containing liquid 12 is stored in the injection container 10 in advance, the stem cell-containing liquid 12 can be administered hygienically and rapidly at the site of administration.
  • the injection container 10 in this embodiment is a syringe barrel as shown in FIG.
  • the injection container 10 is an injection cylinder including a container body 16 that can contain the stem cell-containing liquid 12 and a piston 18.
  • the injection container 10, which is an injection cylinder is provided with a stopper 15 for fixing the position of the piston head 180 relative to the container body 16. Thereby, the pressure of the piston 18 is not applied to the stem cell-containing liquid 12 accommodated in the container body 16.
  • the stopper 15 can be removed from the injection container 10, and the stopper 15 can be easily removed before administering the stem cell-containing solution 12.
  • the means for removing the stopper 15 is not particularly limited.
  • the injection container 10 has a needle coupling part 17 that can be directly or indirectly connected to an injection needle (not shown) and that allows the stem cell-containing liquid 12 to flow from the container body 16 to the injection needle.
  • Connecting the needle coupling part 17 to the injection needle means that the stem cell-containing liquid 12 accommodated in the container body 16 flows through the needle coupling part 17 and is in a state where it can be injected into the animal from the injection needle.
  • the needle coupling portion 17 in the present embodiment is a cylindrical portion that is provided at one end of the container body 16 and protrudes outward.
  • the outer diameter of the needle coupling portion 17 that is a cylindrical portion is smaller than the outer diameter of the container body 16 that is a syringe barrel.
  • a flow path through which the stem cell-containing liquid 12 can flow is provided inside the needle coupling portion 17 and can be directly or indirectly connected to a flow path through which an injection solution provided inside the injection needle (not shown) flows.
  • the needle coupling part 17 and the injection needle are connected directly or indirectly.
  • the needle coupling portion 17 that is in a state before being connected to the injection needle is sealed by the sealing portion (cap 19). That the needle coupling part 17 is sealed means that the needle coupling part 17 is directly or indirectly sealed to the extent that the stem cell-containing liquid 12 accommodated in the container body 16 does not leak through the needle coupling part 17.
  • the injection container 10 can be transported or transported in a state before containing the stem cell-containing solution 12 and before the injection needle is connected, and the handleability is easy. Therefore, the injection container 10 containing the stem cell-containing liquid 12 can be transported or transported to an arbitrary place, and the stem cell-containing liquid 12 can be quickly administered to an administered animal at the arbitrary place.
  • the sealing portion is a cap 19 that covers the needle coupling portion 17.
  • the cap 19 prevents the stem cell-containing liquid 12 from leaking out from the needle coupling part 17 by maintaining the state of being fitted to the needle coupling part 17, and the stem cell-containing liquid 12 is contaminated through the needle coupling part 17.
  • a sealing member that covers the needle coupling portion 17 may be used as a modification of the sealing portion that is not shown.
  • the sealing portion may be an injection needle cap that covers the injection needle and prevents leakage of the stem cell-containing liquid 12 from the injection needle.
  • the injection container 10 of this embodiment is accommodated in a sealed container 32.
  • the term “sealed” means a state in which a gap is not visually confirmed, and it is preferable that the inside of the sealed container 32 be sealed tightly to the extent that the inside of the sealed container 32 is not contaminated with bacteria. That is, the sealed container 32 is preferably a container capable of maintaining the sterilized state of the contents.
  • the shape of the sealed container 32 is not particularly limited, and may be a bag shape that can accommodate the injection container 10 or an arbitrary container such as a predetermined shape.
  • FIG. 4 illustrates a mode in which one injection container 10 is accommodated in the sealed container 32. As a modified example, two or more injection containers 10 may be accommodated in the sealed container 32, and items other than the injection container 10 may be accommodated.
  • the stem cell-containing liquid 12 is accommodated in the container body 16 of the injection container 10.
  • the stem cell-containing liquid 12 is a liquid agent containing stem cells collected from animals.
  • the stem cell-containing solution 12 is preferably prepared so as to have a low viscosity so that the stem cell-containing solution 12 can be injected into an administered animal from a syringe.
  • Stem cells contained in the stem cell-containing solution 12 may be cryopreserved and thawed cells.
  • the stem cell-containing solution 12 can be prepared after thawing the cryopreserved stem cells as appropriate.
  • the thawing method at this time is not particularly limited, and can be appropriately performed in consideration of thawing under conditions that do not damage the frozen stem cells as much as possible.
  • the stem cells are taken out from the deep freezer and kept in a water bath prepared at a water temperature of about 40 ° C for several minutes. And can be thawed.
  • the injection container 10 includes horse name information 24 that can confirm the horse name of the racehorse (donor racehorse) as an incidental element. It is preferable.
  • the injection container 10 having the above-mentioned incidental element can recognize the origin of the stem cells contained in the stem cell-containing liquid 12 by an individual racehorse (donor racehorse) and improve the reliability of the stem cell-containing liquid 12. Can do. This makes it easy to confirm the pedigree relationship between the donor racehorse and the administered racehorse. More specifically, it can be easily determined whether the donor racehorse and the administered racehorse have the same breed.
  • the accessory element is not particularly limited as long as it is attached to the injection container 10 and can be transported or transported.
  • the horse name information 24 is written on the label 14 attached to the outer surface of the container body 16 of the injection container 10. Therefore, in the present embodiment, the accessory element is the label 14 on which the horse name information 24 is written and is affixed to the injection container 10.
  • the horse name information 24 written on the label 14 may be a horse name, a reference number for confirming the horse name, a barcode, or the like.
  • the different accessory element may be an accessory such as a manual 20 attached to the injection container 10 in which the horse name information 24 is recorded.
  • FIG. 4 shows a stem cell injection set 100 that encloses the injection container 10 and the manual 20.
  • the accessory element is accommodated together with the injection container 10 inside the sealed container 32 that accommodates the injection container 10 or is disposed outside the sealed container 32, for example.
  • the stem cell injection set 100 of this embodiment includes the number of living cells having the same composition as the above-described injection container (first container) 10 and the stem cell-containing liquid 12 contained in the injection container (first container) 10. And a second container 40 in which the confirmation sample 120 is accommodated.
  • the stem cell-containing liquid 12 can be transported or transported in a state where it can be rapidly injected, and is stored in the second container 40 when the stem cell-containing liquid 12 is administered at an arbitrary place.
  • the number of living cells in the confirmation sample 120 By measuring the number of living cells in the confirmation sample 120, the number of living cells in the stem cell-containing liquid 12 can be grasped.
  • the same composition of the stem cell-containing liquid 12 and the confirmation sample 120 means that the concentration of the stem cells, the infusion, etc. contained in these liquids are substantially the same.
  • the confirmation sample 120 can be obtained by storing a part of the liquid used as the stem cell-containing liquid 12 in the second container 40.
  • the confirmation sample 120 and the stem cell-containing liquid 12 are substantially the same liquid.
  • a stem cell is a cell having self-renewal ability and pluripotency.
  • the stem cell-containing liquid 12 is expected to express the self-replication ability and pluripotency of stem cells in the body of the administered animal by administering the stem cell-containing liquid 12, at least a part of the stem cells contained in the stem cell-containing liquid 12 is Preferably, it is a living cell.
  • the number of living stem cells contained in the stem cell-containing liquid 12 is, for example, 8 million or more, and the ratio of the number of living cells to the total number of stem cells is preferably 60% or more.
  • the number of living cells means the number of living stem cells among the stem cells contained in the stem cell-containing solution 12.
  • the method for confirming the living stem cells is not particularly limited, and examples thereof include a method of specifically staining living cells with a staining solution such as trypan blue. When stem cells are stained with trypan blue, dead cells are stained blue. Thus, living cells can be confirmed (measured) by confirming the difference in color.
  • a known cell number measuring instrument such as a hemocytometer or a microplate reader can be generally used, but is not limited thereto.
  • the number of living stem cells (the number of living cells) in the stem cell-containing liquid 12 to be administered is 8 million or more, a significant effect of stem cell administration is expected.
  • the number of living cells is preferably 10 million or more, more preferably 20 million or more, and further preferably 50 million or more. It is preferably 80 million or more.
  • the desired number of living cells can be administered within a predetermined period.
  • the ratio of the number of living cells to the total number of stem cells in the stem cell-containing solution 12 is preferably 60% or more.
  • the dead stem cells may be altered in the body of the administered animal and may have an undesirable effect on the administered racehorse. I understood. Therefore, it is better that the number of dead stem cells contained in the stem cell-containing liquid 12 is small. From this viewpoint, the ratio of the number of living cells is preferably 60% or more, and more preferably 70% or more.
  • the ratio of the number of living cells to the total number of stem cells in the stem cell-containing solution 12 can be determined by measuring the total number of stem cells and the number of living cells in the stem cell-containing solution 12 or a sample having the same composition. Of course, the number of dead cells in the sample may be measured, and the number of dead cells may be subtracted from the total number to confirm the number of living cells. For example, when the number of living cells in the stem cell-containing solution 12 is 10 million, and the ratio of the number of living cells to the total number of stem cells including the number of living cells is 70%, the total number of stem cells is about 14.29 million. It is. With such a number and ratio of living cells, a significant effect on stem cell administration is sufficiently expected, and an undesirable influence exerted on an administered animal by dead cells contained in the stem cell-containing solution 12 is sufficiently obtained. It is possible to suppress it to small.
  • stem cell injection set 100 Details of the stem cell injection set 100 will be described below. This embodiment demonstrates using the example provided with the stem cell containing liquid 12 containing the stem cell extract
  • the second container 40 is relatively small with respect to the injection container 10.
  • the shape and structure of the second container 40 are not particularly limited as long as the confirmation sample 120 can be accommodated.
  • the second container 40 includes a container body 42 and a lid portion 44 that can be repeatedly opened and closed with respect to the container body 42.
  • a confirmation sample 120 is accommodated in the second container 40.
  • the stem cell-containing liquid 12 and the confirmation sample 120 are the same stem cell-containing liquid obtained in the step of preparing the stem cell-containing liquid, and the stem cell concentrations are substantially equal. There are fewer.
  • the second container 40 is accommodated in the sealed container 34.
  • the sealed container 34 is a sealed container similar to the sealed container 32.
  • the injection container 10 and the second container 40 are accommodated in different sealed containers (sealed container 32 and sealed container 34), but they may be accommodated together in one sealed container.
  • Stem cell injection set 100 may further optionally have other articles.
  • the stem cell injection set 100 of this embodiment includes a third container 50 and a manual 20 in which an analgesic 52 is accommodated in addition to the injection container 10 and the second container 40.
  • the third container 50 contains an analgesic 52 for administration to the animal to be administered before the stem cell-containing solution 12 is administered by injection.
  • the analgesic 52 is an agent to be administered to an animal to be administered before the stem cell-containing solution 12 is administered by injection.
  • the third container 50 of the present embodiment has the same configuration as the injection container 10 except that the contained liquid is the pain relieving agent 52 and the label 14 is not attached to the outer surface of the container body 16. It may be.
  • an analgesic agent 52 is first administered to an administered animal by coupling the needle coupling portion 17 of the third container 50 to an indwelling needle (not shown) previously placed on the administered animal.
  • the third container 50 is removed from the indwelling needle, and the stem joint-containing liquid 12 can be administered by connecting the needle coupling portion 17 of the injection container 10 to the indwelling needle.
  • the administration method of the stem cell containing liquid 12 is not limited to this.
  • the third container 50 is accommodated in the sealed container 36 as in the case of the injection container 10.
  • the manual 20 includes information related to administration of the stem cell-containing solution 12.
  • the manual 20 of this embodiment includes horse name information 24, administration time information 22, horse name 26, administration method 28, and the like.
  • Horse name information 24 is the same information as the horse name information 24 attached to the label 14. By confirming the horse name information 24, it can be easily known that the stem cell-containing liquid 12 has been collected from a specific racehorse.
  • the administration time information 22 is information of a recommended time until the stem cell-containing liquid 12 accommodated in the stem cell injection set 100 is administered to the administered animal (administered racehorse X).
  • the person who provides the stem cell injection set 100 grasps the number of living stem cells in the stem cell-containing solution 12 at the time of shipment and the survival rate of the stem cells at the time of transportation, etc., and administers in order to reliably administer the desired number of living cells.
  • the recommended time may be transmitted as the administration time information 22. Specifically, for example, a description such as “please administer the stem cell-containing solution 12 before XX month ⁇ day ⁇ ” as the administration time information 22 can be provided in the manual 20.
  • the period from the preparation to the administration of the stem cell-containing solution 12 is maintained in an environment suitable for the survival of the stem cells as much as possible. That is, in the preparation of the stem cell injection set 100, the time from the confirmation of the number of living stem cells in the stem cell-containing solution 12 to the administration is preferably less than 72 hours, more preferably within 48 hours, More preferably, it is within 30 hours. It has been found that if the time to administration described above exceeds 72 hours, the survival rate of the stem cells is remarkably lowered, and the dead stem cells may be altered and may have an undesirable effect on the administered animal. It was. Therefore, it is preferable that the administration time information 22 is set in consideration of these time ranges.
  • the environment suitable for the survival of the stem cell is a temperature environment in which the stem cell exceeds 0 ° C. and is 10 ° C. or less.
  • the horse name 26 indicates the horse name of the donor racehorse from which the stem cell-containing liquid 12 was collected.
  • the horse name 26 “ABCD” is the second generation ancestor. It can be easily confirmed. However, even if the horse name 26 is omitted in the manual 20, it is easy to determine whether the racehorse X to be administered and the donor racehorse are of the same breed according to the horse name confirmed from the horse name information 24. Can be confirmed. Of course, the horse name 26 may be handled as the horse name information 24.
  • the administration method 28 includes information on a method of administering the stem cell-containing solution 12 and operations related thereto. Since the stem cell injection set 100 of the present embodiment includes the second container 40 and the third container 50 that contains the analgesic agent 52, for example, as the administration method 28, “1. Use the confirmation sample 120 contained in the second container 40”. The number of living stem cells is measured to confirm that a predetermined number of living cells are contained 2. Next, the injection container, which is the third container 50, is connected to the indwelling needle, and the racehorse X to be administered 3. The injection container 10 is connected to the indwelling needle and the stem cell-containing solution 12 is administered after the analgesic is administered.
  • the stem cell injection set 100 may further include any other article.
  • the stem cell injection set 100 cleans the remaining stem cells adhering to the inner surface of the container main body 16 after administering the injection needle (not shown) or the stem cell-containing solution 12 by injection, and then the racehorse together with the remaining stem cells. It may further comprise a washing solution or the like to be administered.
  • cleaning liquid what is necessary is just a solution which can be inject
  • the stem cell injection set 100 of this embodiment accommodates the injection container 10, the manual 20, the second container 40, and the third container 50 that constitute the stem cell injection set 100 in one container 30.
  • the container 30 is preferably a container capable of managing the temperature of the contained item within a predetermined range, and is preferably, for example, a cold insulation container having heat insulation.
  • the stem cell injection set 100 when the stem cell injection set 100 is provided to an arbitrary place by transportation, it is preferable that consideration is given to an environment suitable for the survival of the stem cells contained in the stem cell-containing solution 12. Specifically, it is preferable that the transport is performed in a state where a temperature environment of more than 0 ° C. and not more than 10 ° C. is maintained. As a result, it is possible to provide a high-quality stem cell-containing liquid 12 by suppressing the death and activity reduction of the stem cells being transported. For example, a courier service that can be managed at low temperature, which is a paid service of a private company, may be used.
  • transportation means movement using public facilities that can be used by the public, such as roads, railways, ships, and aviation. Examples of available transportation means include, but are not limited to, mail and courier, and various types of moving stem cell-containing liquids from a facility in which stem cells are cryopreserved using the above-described facilities. Widely encompasses the means.
  • the stem cell injection set 100 When the stem cell injection set 100 is over-scheduled due to troubles during transportation of the stem cell injection set 100 and the temperature control during transportation is incomplete, etc., the stem cell-containing solution 12 is killed more than expected. Can also happen. On the other hand, since the stem cell injection set 100 includes the confirmation sample 120, the number of viable stem cells was measured using the confirmation sample 120 before administration of the stem cell-containing liquid 12, and the stem cell-containing liquid 12 was alive. Whether or not the stem cells are sufficiently contained can be grasped. Therefore, the reliability of the stem cell-containing liquid 12 is high.
  • the stem cell-containing solution 12 can be provided to any place in a sterile state, and can be promptly applied to the administered animal by general-purpose means of injection at the provided place.
  • the stem cell-containing solution 12 can be administered.
  • the stem cell injection set 100 includes a second container 40 in which a sample 120 for confirming the number of living cells having the same composition as the stem cell-containing liquid 12 stored in the injection container 10 is stored together with the injection container 10. Therefore, the number of living cells of the stem cells in the stem cell-containing liquid 12 can be grasped by measuring the number of living cells of the sample 120 for confirming the number of living cells at an arbitrary timing.
  • the stem cell-containing liquid 12 of the present embodiment containing stem cells collected from a racehorse avoids mixing of non-race horse stem cells into the racehorse with respect to racehorses where the pedigree is valued. Sustainability of maintenance can be avoided.
  • the stem cell-containing liquid 12 of the present embodiment containing stem cells collected from a racehorse is mixed with a racehorse that does not have a pedigree in a racehorse that does not have a pedigree, while the stem cell-containing solution 12 of this embodiment maintains the pedigree To avoid suspicion in maintaining the racehorse lineage.
  • the method for providing a stem cell-containing solution for use in the stem cell administration method of the present invention and the stem cell injection set 100 for transporting the stem cell-containing solution and injecting it into a racehorse have been described above.
  • the description returns to the administration step of injecting the stem cell-containing solution into the racehorse with the injection needle.
  • the timing for performing the administration step is not particularly limited, for example, the administration step is preferably performed within 72 hours from the time when the stem cell-containing solution is prepared, and more preferably within 50 hours.
  • a temperature environment suitable for the cells for example, more than 0 ° C. and 10 ° C. or less. According to the study of the present inventors, even when the prepared stem cell-containing liquid is handled in a temperature environment substantially exceeding 0 ° C. and not higher than 10 ° C., the stem cells contained in the stem cell-containing liquid I knew it was going to die.
  • the survival rate is approximately 80% after 24 hours, 70% after 48 hours, and after 72 hours. 50% to 60%. Therefore, if it is less than 72 hours, a stem cell-containing solution having a survival rate exceeding 50% can be administered to a racehorse to be administered, and a large amount of dead stem cells can be prevented from being administered to a racehorse to be administered. Can do.
  • administration within 50 hours from when the stem cell-containing solution is prepared is preferable because significantly more than half of the living stem cells can be administered with respect to the total number of stem cells contained in the stem cell-containing solution.
  • the stem cell administration method in the present embodiment may include a living cell number confirmation step of confirming the number of living stem cells contained in the stem cell-containing solution in addition to the administration step described above.
  • a living cell number confirmation step By performing the live cell number confirmation step, the number of live stem cells administered to the racehorse to be administered can be grasped, and the reliability of the stem cell administration method can be improved.
  • the timing of performing the viable cell number confirmation step is not particularly limited, but it is preferably performed after the above-described transport step and before the administration step.
  • confirming the number of living stem cells contained in the stem cell-containing liquid by carrying out the viable cell number confirmation step, it is possible to confirm that the stem cell-containing liquid is normal and the reliability of the administration process improves.
  • confirming the number of living cells means grasping the number of living stem cells contained in the stem cell-containing solution.
  • the stem cell-containing solution For the confirmation of the number of living cells, using the stem cell-containing solution, measure the number of living cells contained in this, and measure the number of living cells using another sample having the same composition as the stem cell-containing solution, From the measurement result, the aspect which estimates the number of the living stem cells contained in the stem cell containing liquid is included.
  • the number of living cells included in the confirmation sample 120 having the same composition as the stem cell-containing liquid 12 and contained in the second container 40 is measured, and used for injection. This can be done by estimating the number of viable cells contained in the stem cell-containing liquid 12 filled in the container 10.
  • the whole or part of the confirmation sample 120 is collected and stained with a staining solution such as trypan blue. It is good to count by.
  • the stem cell administration method in this embodiment may further have a washing
  • a washing solution that can be injected into the racehorse is filled in the injection container during or after the administration step, and the stem cells attached to the inner wall of the injection vessel are washed with the washing solution.
  • stem cells have the property of adhering to the container wall surface. According to the administration method of this embodiment having a washing step, the stem cells attached to the injection container are efficiently administered to the racehorse to be administered. And the loss of the number of administration of stem cells can be reduced.
  • a racehorse symptom improvement method (hereinafter, also referred to as this symptom improvement method) of the present invention will be described as a second embodiment of the present invention.
  • the racehorse symptom improvement method of the present embodiment is characterized in that a stem cell-containing solution containing stem cells is administered to a racehorse showing an abnormal state using the stem cell administration method of the present invention described above.
  • the stem cell administration method in the present embodiment the description of the first embodiment described above can be referred to as appropriate.
  • the improvement of the symptom means that the abnormal state is brought into the normal state or brought close to the normal state.
  • the symptom can be improved by administering stem cells to a racehorse showing a symptom different from the normal state (that is, a racehorse in an abnormal state). More specifically, according to this symptom improvement method, for example, it is possible to improve symptoms such as racehorse injuries or prevent the occurrence of injuries.
  • the stem cell-containing fluid (or the stem cells contained therein) administered to the racehorse by injection is not normal, such as the inflamed part of the racehorse. It is presumed that it reaches a local part and exhibits a significant action for returning the local part to a normal state.
  • the abnormal state refers to a state having symptoms different from the normal state. Specifically, in one racehorse, a state without illness, injury, or fatigue after a race or training is defined as a normal state, and a defective state that has not been confirmed in the normal state is referred to as an abnormal state. That is, the abnormal state includes a state in which a fatigue state after a race or training, a disease, or the like is observed.
  • the fatigue state which is an abnormal state, is confirmed by, for example, an increase in the lactic acid level compared to the normal state, a local heat feeling observed on the body surface, or an observation of a gait different from the normal state. can do.
  • a racehorse in a fatigued state there is a risk of inviting injuries and the like, and by sufficiently resting, there is a possibility that the race interval becomes long and training is insufficient.
  • this symptom improvement method on a racehorse whose fatigue state has been confirmed, it is possible to recover fatigue earlier than usual and to improve the abnormal state.
  • injuries include motor system diseases such as tendonitis, fractures, gait denormalization (including so-called kosumi or sukumi symptoms), canal periosteitis (so-called soe symptoms) ), Lumbar paralysis (including so-called low back symptoms), hoof disease (Fumamone intercostal), ophthalmic diseases, reproductive function diseases, nervous system diseases, skin diseases, poor fur, cardiopulmonary diseases, cardiovascular system Examples include, but are not limited to, diseases such as urological diseases, fire extinguisher diseases, fractures, respiratory diseases such as wheezing (Whistling; the throat ringing), endocrine immune diseases, and nasal bleeding.
  • motor system diseases such as tendonitis, fractures, gait denormalization (including so-called kosumi or sukumi symptoms), canal periosteitis (so-called soe symptoms) ), Lumbar paralysis (including so-called low back symptoms), hoof disease (Fumamone intercostal), ophthalmic diseases, reproductive function diseases
  • gait denormalization can occur in both a racehorse in a fatigued state and a racehorse in an injured state, etc., without specifying the cause of the gait denormalization,
  • the gait can be normalized.
  • the reason for this is not clear, but by administering the stem cell-containing solution by injection such as intravenous injection, the stem cell-containing solution (or the stem cells contained therein) is put on the bloodstream and circulates through various organs. It is presumed that it is possible to reach the local area where it occurs.
  • the symptom indicating an abnormal state is any one or more of the following first to seventh symptoms. That is, the first symptom is a flexor tendinitis symptom.
  • the second symptom is a fracture.
  • the third symptom is ductal periosteitis.
  • the fourth symptom is gait denormalization.
  • the fifth symptom is a lumbar symptom.
  • the sixth symptom is an increase in lactic acid level or local hot feeling observed on the body surface.
  • the seventh symptom is a respiratory disease such as wheezing.
  • the symptom means a change from a normal (healthy) state that is recognized when the racehorse has an injury or the like or is in a state where the injury or the like is likely to occur.
  • a normal (healthy) state that is recognized when the racehorse has an injury or the like or is in a state where the injury or the like is likely to occur.
  • the first symptom is a symptom of flexor tendonitis.
  • Flexor tendinitis is inflammation that has occurred in the flexor tendon following the muscle that flexes the limb (flexor muscle) of the tendon tissue connecting the bone and the muscle. Flexionitis often occurs in the shallow flexor tendon on the posterior aspect of the forelimb (third metacarpal bone) of the racehorse, but it occurs in the deep flexor tendon and in the shallow or deep flexor tendon of the hind leg Inflammation is also included in the tendonitis referred to in the present invention.
  • Symptoms of flexor tendinitis can be confirmed by the presence or absence of clinical symptoms such as swelling around the tendon by external observation of the racehorse, heat, and pain during palpation.
  • Ultrasonic diagnosis is performed by observing an echo image of the cross-section of the tendon by applying a probe of an ultrasonic diagnostic apparatus to the flexor tendon of the racehorse.
  • a normal (healthy) flexor tendon is observed whitish in a black and white echo image.
  • flexor tendonitis develops, the inflamed part is observed in black in the flexor tendon, so that the symptom can be easily confirmed.
  • the degree of damage of flexor tendinitis can be calculated by the ratio of the area of the inflamed part (that is, the part shown in black) to the cross-sectional area of the flexor tendon shown in the echo image.
  • the second symptom is a fracture.
  • the fracture widely includes a state in which the continuity of the bone structure is broken, and includes, for example, a shear fracture, a compression fracture, a torsion fracture, a flexion fracture, and an exfoliation fracture. Also includes fissure fractures found in shin splints. Fractures can be diagnosed with clinical symptoms, but it is common to observe and examine them with echo images or X-rays. When a fracture occurs, the affected area is generally inflamed.
  • the fracture symptom can be improved significantly faster than a general fracture healing rate.
  • the third symptom is tubular periosteitis.
  • Tube periosteitis is also called soe.
  • Tubular periosteitis refers to inflammation that occurs in front of the canal bone (third metacarpal bone).
  • periosteitis is caused by overtraining a young racehorse whose bone is not completely fossified. Since there is an inflammatory symptom, the administered stem cell-containing liquid or the stem cells contained therein can act to reach the local area where the inflammatory symptom has occurred and to improve the symptom.
  • the fourth symptom is gait denormalization.
  • Gait denormalization is believed to have a number of causes, including so-called kosumi or sukumi symptoms.
  • Kozumi means a change in gait (unnormalized) of a racehorse caused by muscle pain, muscle fatigue, or tendon or ligament pain.
  • Sukumi is generally referred to as paralytic myofiuria, and indicates an abnormal condition in which the gait is disturbed suddenly immediately after a break after intense exercise.
  • the fifth symptom described above is a symptom of lumbar paralysis, and is also commonly referred to as a hip hula.
  • the above-mentioned lumbar paralysis is a disease characterized mainly by exercise paralysis of the lumbar region. There are no signs, suddenly depressed, and neuropathy such as reflexes declines, followed by movement disorders. Except for acute symptoms that heal within a few days, in the case of chronic disease or recurrence, serious injuries such as hindlimb motor dysfunction are often caused.
  • a stem cell-containing solution can be administered to a racehorse that has developed lumbar paralysis by a simple procedure called injection, thereby improving the symptoms of lumbar paralysis.
  • the sixth symptom described above is an increase in lactic acid level or local hot feeling observed on the body surface.
  • the observation includes quantitative observation and qualitative observation.
  • the sixth symptom is a symptom observed in both a racehorse in which an injury or the like has already occurred and a racehorse in which an injury or the like may occur.
  • racehorses that have already suffered injuries such as tendonitis or bone fractures tend to get hot near the inflamed local area, so it is easy to observe abnormalities by observing the body surface of the racehorse. Can be understood.
  • the lactic acid level is brought close to a normal value at an early stage, or the local feeling of heat is eliminated. Serious injury can be prevented, and injuries and the like can be prevented.
  • the seventh symptom described above is a respiratory disease such as wheezing.
  • Respiratory disorders broadly include a variety of respiratory-related diseases, many of which are normal breathing status, breathing sounds, or movements of the nostrils, lateral chest, or abdomen of a racehorse or racehorse It can be observed as abnormalities.
  • wheezing broadly includes cases in general that produce abnormal sounds such as “zeze”, “hugh”, “gorolog” from the throat, and includes symptoms derived from inflammation such as bronchitis and symptoms derived from neurological diseases. If a racehorse or a racehorse that is required to run at full power or conduct a prescribed competition suffers from a respiratory disease, there is a possibility that the race ability or the competition ability may be significantly impaired.
  • the seventh symptom can be improved after drug administration or surgery is avoided or moderately suppressed. Is possible.
  • the number of viable stem cells administered by the stem cell administration method performed in this symptom improvement method is not particularly limited.
  • the total is more preferably 50 million or more, and particularly preferably the total is 80 million or more.
  • the symptom can be significantly improved by administering the above-described range of living stem cells to a racehorse in an abnormal state including the first symptom to the sixth symptom described above.
  • the viable cell number measuring step described in the first embodiment it is preferable to carry out the viable cell number measuring step described in the first embodiment.
  • viable cells in the above-mentioned range can be obtained by previously containing living stem cells sufficiently exceeding the number of live cells to be administered. It is possible to carry out an administration process that satisfies the total number of numbers.
  • racehorses and racehorses need to undergo a strict doping test before participating in horse races or various competitions, and use of limited drugs, etc., excluding restricted substances, is unavoidable for treatment and the like.
  • the present invention for improving the symptoms of racehorses or racehorses using stem cells that are not defined as controlled substances improves the symptoms of racehorses or racehorses without worrying about the timing of administration, or It is very effective for maintaining good physical condition. Even if stem cells are defined as regulatory substances in the future, the present invention can sufficiently improve the symptoms of racehorses or horses by measuring the timing of administration of stem cells.
  • stem cells were collected from donor racehorses, and the stem cells were subcultured to obtain passage stem cells.
  • the passaged stem cells were thawed after being cryopreserved, and then administered to a racehorse by performing a live cell number determination step, a stem cell-containing agent preparation step, and a transport step.
  • the administration to the racehorse is a recommended time from the predetermined time when the number of viable cells was confirmed after the thawing process to 48 hours, and the desired number of living cells, which is the desired number of living stem cells in one administration, is about 1 ⁇ 10 7 pieces.
  • the survival rate of the stem cells when the stem cells were stored in an environment of 4 ° C. was confirmed in advance, the survival rate, which is the ratio of the number of living cells after storage to the number of living cells before storage, was about 70%. .
  • the hip of the horse was incised, and about 5 g of fat cells were collected to obtain a sample for collecting stem cells.
  • the sample is cut to about 1 mm square and then treated with a collagenase solution at 37 ° C. for 1 hour, and then the treated sample is centrifuged for about 10 minutes with a centrifugal force of 400 ⁇ g to obtain 100 ⁇ m
  • a stromal vascular fraction containing adipose-derived mesenchymal stem cells was obtained.
  • a flask containing a medium prepared by adding an antibiotic (gentasin) to a commercially available serum-free medium for animal cells was prepared.
  • the above-obtained fraction was seeded in the flask, and liquid stationary culture was performed under conditions of 37 ° C./5% CO 2 . Every two days, the entire supernatant of the medium was collected and replaced with a fresh medium, and the cells adhered in a range of about 80% to 90% with respect to 100% of the area of the bottom of the flask with a phase contrast microscope.
  • the culture was terminated, and a release agent containing protease was added to the flask to collect cells attached to the inner wall surface (especially the inner bottom surface) of the flask.
  • the viable cell count of the collected cells was measured using a hemocytometer, and subculture was repeated in the same manner as described above until an appropriate cell count was obtained.
  • the cells were mixed with a serum-free cell cryopreservation solution containing dimethyl sulfoxide to obtain a sample for freezing.
  • the cryopreservation container was taken out of the deep freezer, and the sample for freezing was thawed in a water bath adjusted to 37 ° C. to obtain a thawed sample containing stem cells.
  • Stem cells (thawed samples) obtained by thawing as described above were washed with Ringer's solution. Trypan blue was added to a part of the washed stem cells to stain the dead stem cells, and the number of viable cells was measured. As a result, 1 ml of the thawed sample had about 1.5 ⁇ 10 7 or more living cells. It was confirmed that stem cells were included. The time when the number of viable cells was measured as described above was defined as a predetermined time.
  • the recommended recommended time from the above predetermined time to administration to the racehorse to be administered is 48 hours
  • the desired number of living cells is 1 ⁇ 10 7
  • the survival rate is 70%.
  • the above-described stem cell culture, cryopreservation step, thawing step, and live cell number determination step were performed in advance, and a cryopreserved live cell number confirmation test was performed. It was confirmed that about 5% to 20% of stem cells were dead.
  • the number of living cells measured after the thawing step with respect to the number of stem cells donated to the cryopreservation step in this example was almost the same as the number of living cells grasped in this cryopreservation viable cell number confirmation test.
  • the injection stem cell-containing liquid (stem cell-containing liquid) prepared as described above has the same configuration as the injection container 10 shown in FIG. 4 and is labeled with the label containing the horse name information of the donor racehorse. And the needle coupling part was sealed with a cap. Further, in order to fix the relative position of the piston with respect to the container body, a stopper was attached to the piston to produce an injection container. The injection container containing the stem cell-containing solution was housed in a sealed container.
  • a sample for confirming the number of living cells which is the same liquid as the stem cell-containing liquid stored in the injection container, is stored in a second container different from the injection container, and the second container is stored in a sealed container.
  • a manual was prepared in which the horse name information of the donor racehorse and the recommended time information (administration time information) from the predetermined time until administration to the administered racehorse were described.
  • the injection container, the second container, and the manual prepared as described above were packed in one cold bag to obtain a stem cell injection set.
  • Stem cells obtained by thawing as described above were washed with Ringer's solution.
  • Ringer's solution When trypan blue was added to a part of the washed stem cells to stain dead stem cells and the number of viable cells was measured, 1 ml of the thawed sample contained about 1.4 ⁇ 10 7 stem cells. Confirmed that.
  • 1.5 ml of Ringer's solution was added to and mixed with 1 ml of a thawed sample containing about 1.4 ⁇ 10 7 stem cells, and this was used as a stem cell-containing solution.
  • the stem cell-containing solution prepared as described above is quickly stored in a syringe without a syringe needle connected, and the syringe is sealed so that the stored stem cell-containing solution does not leak to obtain a container for injection. It was.
  • the stem cell injection set containing the stem cell-containing solution obtained as described above was shipped to the administration facility by Yamato Transport's cool flight (temperature control conditions: 0 ° C. to 10 ° C.). About 24 hours after shipment, the stem cell injection set arrived at the administration facility.
  • the administration step was set to about 48 hours from when the stem cell-containing solution was prepared, and the stem cell injection set containing the stem cell-containing solution was stored in a temperature environment of about 4 ° C. until administration.
  • the injection container and the second container included in the stem cell injection set were stored refrigerated until administration.
  • the number of viable cells was measured using a confirmation sample, and it was confirmed that there were living stem cells having a desired viable cell number of 1 ⁇ 10 7 or more.
  • the stem cell-containing solution was administered to the racehorse to be administered using an injection container. The administration was performed by first placing an indwelling needle on the neck of the racehorse to be administered, injecting the indwelling needle connected to a container for injection, and injecting the stem cell-containing solution into the racehorse to be administered.
  • racehorse 1 The racehorse (horse name: Neovitarosa, 3 years old, male) for which the symptoms of flexor tendinitis were confirmed by a veterinarian was designated as racehorse 1 to be administered. Specifically, as shown in FIG. 5, in the administered racehorse 1, in the echo image, a damaged portion A ⁇ b> 2 that is projected as a black image (low echo image) exists in the central side portion of the transverse cross section A ⁇ b> 1 of the entire tendon. It was confirmed. In addition, it was confirmed from the external observation near the place where flexor tendinitis occurred that there was a feeling of heat. Thus, the administered racehorse 1 was diagnosed as having flexor tendinitis symptoms.
  • the area of the cross section A1 is 1.70 cm 2
  • the area of the damaged part A2 is 0.87 cm 2
  • the ratio of the area of the damaged part A2 to the area of the cross section A1 was confirmed to be 51%.
  • An indwelling needle for intravenous injection was placed in advance at the neck of the administered racehorse 1 diagnosed as flexor tendinitis.
  • An injection needle having a size of 18 gauge (inner diameter 0.94 ⁇ m) was used. Then, the syringe barrel transported and stored as described above was connected to the injection needle, and the contained stem cell-containing solution was administered to the racehorse to be administered by intravenous injection. In addition, it was previously grasped that the number of living stem cells contained in the stem cell-containing solution at the time of administration was about 10 million. By the same administration method, it was administered three times at intervals of several days. The interval between doses is shown in Table 1.
  • an echo image of a portion of the administered racehorse 1 having a symptom of flexor tendinitis was taken.
  • the echo image at that time is shown in FIG.
  • the damaged part A2 confirmed before the administration step of the stem cells was hardly observed in the echograph after administration.
  • the thermal sensation observed before administration disappeared.
  • the symptoms of flexor tendinitis in the racehorse 1 to be administered were improved.
  • the area of the low echo image indicating the damaged portion is reduced in about one month from the onset, and the low echo image is echoed in about three or four months.
  • Example 1 since the damaged part A2 almost disappeared in the echograph 21 days after the administration of the stem cells three times from the day when the onset was confirmed, the administration of the stem cells in the improvement of the symptoms of flexor tendinitis It was shown that the effect was exerted significantly.
  • Example 2 was carried out by using a racehorse (horse name: Neovanquish, 5 years old, male) in which the symptoms of mild flexor tendinitis were confirmed by a veterinarian as the racehorse 2 to be administered.
  • the administered racehorse 2 was diagnosed as having a flexion tendonitis damage rate of about 5% by echo images.
  • the administration step was carried out in the same manner as in Example 1 using the same stem cell-containing solution as used in Example 1 except that the number of administrations and the administration interval shown in Table 1 were changed. .
  • a total of about 90 million live cells of about 90 million stem cells were administered to racehorse 2 to be administered in total, about 90 million in each time, and flexor tendinitis was cured.
  • about 8 months after the start of treatment On July 15th, he raced at the Nagoya Racecourse and won. Furthermore, about a month later, on August 26, 2015, he entered the race held at the Nagoya Racecourse and gave his second win after returning. And on December 10, 2015, he entered the race held at the Nagoya Racecourse and gave his third victory after returning.
  • flexor tendinitis is cured within a very short period of time, and since it is an extremely minimally invasive procedure, the postoperative recovery is fast, so that the activity as an active racehorse can be achieved. It is possible to continue. And the degree of healing of flexor tendonitis was confirmed not only to be able to run, but to cure enough to run and win a horse racing race.
  • Example 3 Example of racehorse (horse name: Neovarkas, 2 years old, male) having a bone aneurysm with heat and pain in the canal bone, and a fracture line was observed in the vicinity of the aneurysm in X-ray photograph as administration racehorse 3 3 was performed.
  • An X-ray photograph showing the state of the fracture in the vicinity of the bone aneurysm of the administered racehorse 3 is shown in FIG. According to the diagnosis of the veterinarian, in FIG. 7, a bone aneurysm (moyamoya shadow portion) is projected in the middle portion of the canal bone, and a fracture line was confirmed in the vicinity of the bone aneurysm.
  • the same stem cell-containing liquid as that used in Example 1 was used, and the dose per administration was doubled (20 million), which was the same as Example 1.
  • the administration step was carried out in a manner.
  • X-rays of the canal including the location where the fracture of the administered racehorse 3 was confirmed were taken 11 days after the administration process was performed on the administered racehorse 3. This X-ray photograph is shown in FIG. As a result, it was observed by the veterinarian diagnosis that the fracture line that had existed before the administration had disappeared, and the improvement of fracture symptoms was confirmed. Moreover, when the affected part of the racehorse 3 to be administered was observed on the next day after the administration of the stem cells, it was observed that heat and pain were improved although the bone aneurysm remained.
  • Example 3 it was confirmed that the symptoms of fever, pain, and fractures in the canal of the cancellous bone were improved in a short period of time by the administration of a single stem cell, and the effect of the administration of the stem cell significantly affected the improvement of these symptoms. It was done. 7 and 8 are X-ray photographs obtained by reversing black and white.
  • Example 4 was conducted by using a racehorse (horse name: Neovaschelan, 3 years old, female) in which wheezing was confirmed as a racehorse 4 to be administered. Two days before the first administration step (June 3) (June 1), endoscopic examination of the pharynx was performed on the racehorse 4 to be administered. When a video taken by endoscopy was confirmed, the movement of the pair of left and right arytenoids did not synchronize in front view in the upper airway, and complete abduction of the arytenoid cartilage on one side was not maintained. In other words, an abnormal state was observed.
  • a racehorse horse name: Neovaschelan, 3 years old, female
  • endoscopic examination of the pharynx was performed on the racehorse 4 to be administered. When a video taken by endoscopy was confirmed, the movement of the pair of left and right arytenoids did not synchronize in front view in the upper air
  • the wheezing sound of the racehorse 4 to be administered was due to malfunction of the arytenoid cartilage (laryngeal hemiplegia), and the wheezing sound originated from a neurological disease. Therefore, for the racehorse 4 to be administered, the same stem cell-containing liquid as that used in Example 1 was used, and the dose per administration was increased 5 times (50 million), and the number of administration and the administration interval are shown in Table 1. The administration step was carried out in the same manner as in Example 1 except that the contents were changed to those shown. In addition, the administered racehorse 4 received normal training (training) without rest, exercise restriction, and other treatments for about one month including two administration process periods.
  • training training
  • Endoscopy of the pharynx was performed again about 3 weeks (July 1) about 3 weeks after the second administration step (June 9) for the racehorse 4 to be administered.
  • the movement of the arytenoid cartilage whose malfunction was confirmed before administration was improved and the wheezing sound of the racehorse 4 to be administered was reduced.
  • the administered racehorse 4 was diagnosed as having improved wheezing. From the results of Example 4, it was confirmed that the present invention can exert an effect on wheezing. According to the present invention, it has been confirmed that symptoms can be improved without undergoing surgical operation, particularly for wheezing caused by neurological diseases.
  • the nerves that move the organs of the pharynx start from the brain and go to the pharynx via the aorta that continues to the heart, so the length is said to be 2.5 m or more. Yes.
  • Example 4 in which improvement of symptoms for such a terminal nerve disease was confirmed suggests that the present invention works well for improvement of other neurological diseases beyond wheezing.
  • the stem cell-containing solution was administered by intravenous injection to 13 other racehorses to be administered in addition to the above 4 horses.
  • 13 cases included hoof bone fracture, cervical stenotic myelopathy, chronic (renal failure, sprain, colic, and significant weight loss due to poor physical condition. Hair loss, reproductive conception and maternal improvement, severe Chronic laminitis, ant sinus (hollow wall), severe phlegmon, difficulty walking, placentitis, miscarriage due to placentitis ( Abortion by Placentitis).
  • (A1) including an administration step of injecting a stem cell collected from an animal or a stem cell-containing solution containing a passage stem cell obtained by subculturing the stem cell into a racehorse or a racehorse with an injection needle.
  • Stem cell administration method (A2) The stem cell administration method according to (a1), wherein in the administration step, the means for injecting the stem cell-containing solution into the racehorse or the racehorse with the injection needle is intravenous injection. (A3) The stem cell administration method according to (a1) or (a2), wherein the injection needle has an inner diameter of 0.6 mm or more.
  • the stem cell administration method according to any one of (a1) to (a3), including a live cell number confirmation step of confirming the viable cell number of the stem cells contained in the stem cell-containing solution.
  • the stem cell-containing solution is contained in an injection container connected to the injection needle, In the middle of the administration step or after the completion of the administration step, the injection container is filled with a washing solution that can be injected into the racehorse, and the stem cells attached to the inner wall of the injection vessel are washed with the washing solution and the race.
  • the stem cell administration method according to any one of (a1) to (a4) above, which comprises a washing step of injecting the horse.
  • a donor race in which a donor racehorse that is a racehorse from which the stem cells are collected and an administered racehorse to which the stem cells are administered are of the same breed or sibling, or a racehorse from which the stem cells are collected The stem cell administration method according to (a9) above, wherein the horse and the administered horse to which the stem cells are administered are of the same breed or sibling.
  • A11 The stem cell administration method according to (a9) or (a10) above, wherein the stem cells are mesenchymal stem cells collected from fat collected from the racehorse or the racehorse.
  • a racehorse characterized by administering a stem cell-containing solution containing stem cells to a racehorse exhibiting an abnormal state using the stem cell administration method according to any one of (a1) to (a11) above. Or how to improve the symptoms of horses.
  • the first symptom is that of tendonitis
  • the second symptom is a fracture
  • the third symptom is duct periosteitis
  • the fourth symptom is gait denormalization
  • the fifth symptom is lumbar symptom
  • the sixth symptom is an increase in lactic acid level or local hot feeling observed on the body surface.
  • the seventh symptom is respiratory disease.
  • the administration step in the stem cell administration method is carried out once or divided into a plurality of days, so that the total number of viable cells is 10 million or more, and is administered to the racehorse or the racehorse (a12). ) Or the symptom improvement method according to (a13).
  • (B1) A method for providing a stem cell-containing agent to be administered to a racehorse or a racehorse, A cryopreservation step for freezing stem cells collected from animals; A thawing step of thawing the stem cells frozen in the cryopreservation step; The survival rate at the recommended recommended time elapse from the predetermined time when the number of viable cells after the thawing step after the thawing step is confirmed until the stem cell is administered to the racehorse or the racehorse, and A desired number of viable cells when the stem cells are administered to a racehorse or the racehorse, and a viable cell number determination step for determining an absolute number of viable cells using, Using the stem cells after the thawing step, a stem cell-containing agent preparation step of preparing a stem cell-containing agent containing the stem cells satisfying the absolute number of living cells, and A transporting step of transporting the stem cell-containing agent to an area
  • (B3) The method for providing a stem cell-containing agent according to (b1) or (b2) above, wherein the animal is an equine animal.
  • (B4) The method for providing a stem cell-containing agent according to (b3) above, wherein the equine animal is a donor racehorse or a donor racehorse.
  • (B5) The cryopreservation step stores the stem cells in a state where the name of the donor racehorse or the donor racehorse that has become a donor of the stem cells to be cryopreserved can be confirmed.
  • a method for providing a stem cell-containing agent is
  • the above-mentioned (b4) or (b5) includes a pedigree verification step that determines the stem cells of the donor racehorse or the donor racehorse that are the same pedigree or siblings as stem cells to be provided to the thawing step before the thawing step.
  • the viable cell number determination step further includes a post-thaw step viable cell number measurement step of measuring the viable cell number of the stem cell thawed by the thaw step and determining the viable cell number at the predetermined time.
  • (B9) The method according to any one of (b1) to (b8), wherein the transport process is completed so that the stem cell-containing agent can be administered to the racehorse or the racehorse within 72 hours from the predetermined time.
  • the stem cells are stem cells collected from the animal that has received stem cell administration in advance.
  • the method for providing the stem cell-containing agent comprises: The first aspect in which the stem cells themselves thawed in the thawing step are subjected to the live cell number determination step, or the stem cells obtained by subculturing the stem cells thawed in the thawing step and performing the subculture.
  • the stem cell-containing agent is provided with information of time from a predetermined time after the thawing step in the method of providing the stem cell-containing agent until the stem cell is administered to the racehorse or the racehorse.
  • the said 2nd container is a stem cell provision set as described in said (b12) in which the sample for living cell number confirmation which is the same composition as the said stem cell containing agent was accommodated.
  • the first container is an injection container that is directly or indirectly connected to an injection needle used for administering the stem cell-containing agent to the racehorse or the racehorse.
  • Stem cell donation set The animal from which the stem cell-containing agent is collected is a donor racehorse or a donor racehorse, The stem cell donating set according to any one of (b12) to (b14), wherein horse name information of the donor racehorse or the donor racehorse is included.
  • the stem cell providing set according to any one of (b12) to (b15) above, which comprises an analgesic administered before injection of the stem cell-containing agent.
  • An injection container having a container body capable of containing a stem cell-containing solution and connectable to an injection needle, Contained in a sealed container, A stem cell-containing solution containing stem cells is contained in the container body, An injection container comprising a sealing portion for preventing the stem cell-containing liquid from leaking out of the injection container.
  • C2 having a needle coupling portion that can be directly or indirectly connected to the injection needle and that allows the stem cell-containing liquid to flow from the container body to the injection needle;
  • the injection container according to (c1) or (c2), wherein the stem cell is a stem cell of an equine animal.
  • the injection container according to (c3), wherein the equine animal is a racehorse or a racehorse.
  • the injection container according to (c4) comprising horse name information capable of confirming a horse name of the racehorse or the racehorse as an incidental element of the injection container.
  • the number of viable cells of the stem cells contained in the stem cell-containing solution is 8 million or more, and the ratio of the number of viable cells to the total number of stem cells is 60% or more (c1) to (c5) The container for injection as described in any one of).
  • a stem cell injection set comprising: (C10) Stem cell-containing liquid for injection containing stem cells collected from racehorses or horses. (C11) The stem cell-containing solution for injection according to (c10), wherein the stem cell is an umbilical cord, umbilical cord blood or placenta-derived umbilical cord stem cell of the racehorse or the racehorse.

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Abstract

La présente invention concerne un procédé d'administration de cellule souches qui comprend une étape d'administration pour injecter une solution contenant des cellules souches contenant des cellules souches collectées à partir d'un animal (animal donneur) ou des cellules souches sont-cultivées obtenues par sous-culture des cellules souches dans un cheval de course (cheval de course donneur) ou un cheval d'exhibition (cheval d'exhibition donneur) au moyen d'une aiguille d'injection ; et un procédé pour améliorer les symptômes d'un cheval de course ou d'un cheval d'exhibition, dans lequel une solution contenant des cellules souches contenant des cellules souches est administrée à un cheval de course (cheval de course donneur) ou un cheval d'exhibition (cheval d'exhibition donneur) présentant un état anormal au moyen du procédé d'administration de cellules souches ci-dessus.
PCT/JP2016/067943 2015-06-17 2016-06-16 Procédé d'administration de cellules souches, procédé pour améliorer les symptômes de chevaux de course ou d'exhibition, récipient pour injection et ensemble d'injection de cellules souches WO2016204230A1 (fr)

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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP6152205B1 (ja) * 2016-08-09 2017-06-21 仁幸 小林 化粧品、医薬用組成物、およびそれらの製造方法
WO2018123628A1 (fr) * 2016-12-28 2018-07-05 ロート製薬株式会社 Composition pharmaceutique de cellules, kit pour traitement de maladie, et solution pour suspension de cellules

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2004018504A (ja) * 2002-06-20 2004-01-22 Jms Co Ltd 細胞保存容器
JP2009511212A (ja) * 2005-10-13 2009-03-19 アルゴス・セラピューティクス・インコーポレーテッド 低温で貯蔵された液体薬剤の送達に適する装置、システム、および関連する方法
JP2010504083A (ja) * 2006-09-20 2010-02-12 セールス・エンジニアリング・アクチェンゲゼルシャフト 血液、特に末梢血から成体幹細胞を増殖させるための方法及び医療分野におけるその利用

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2004018504A (ja) * 2002-06-20 2004-01-22 Jms Co Ltd 細胞保存容器
JP2009511212A (ja) * 2005-10-13 2009-03-19 アルゴス・セラピューティクス・インコーポレーテッド 低温で貯蔵された液体薬剤の送達に適する装置、システム、および関連する方法
JP2010504083A (ja) * 2006-09-20 2010-02-12 セールス・エンジニアリング・アクチェンゲゼルシャフト 血液、特に末梢血から成体幹細胞を増殖させるための方法及び医療分野におけるその利用

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
FUMIKI KATO ET AL., THOROUGHBRED KYOSOBA NO SENKUKKEN'EN NI TAISURU SHIBO SOSHIKI YURAI KANSAIBO CHIRYO SEISEKI, vol. 56, 10 August 2012 (2012-08-10), pages 368 *
FUMIO SATO ET AL.: "Uma Kotsuzui Yurai Kan'yokei Kansaibo no Bunri Baiyo Oyobi Ishoku ni Oyobosu Yuso Joken no Eikyo", JAPANESE SOCIETY OF VETERIANY SCIENCE GAKUJUTSU SHUKAI KOEN YOSHISHU, vol. 143, 2007, pages 214 *

Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP6152205B1 (ja) * 2016-08-09 2017-06-21 仁幸 小林 化粧品、医薬用組成物、およびそれらの製造方法
WO2018030428A1 (fr) * 2016-08-09 2018-02-15 仁幸 小林 Produit cosmétique, composition pharmaceutique, et procédé de production dudit produit cosmétique et de ladite composition pharmaceutique
JP2018024595A (ja) * 2016-08-09 2018-02-15 仁幸 小林 化粧品、医薬用組成物、およびそれらの製造方法
WO2018123628A1 (fr) * 2016-12-28 2018-07-05 ロート製薬株式会社 Composition pharmaceutique de cellules, kit pour traitement de maladie, et solution pour suspension de cellules
JPWO2018123628A1 (ja) * 2016-12-28 2019-10-31 ロート製薬株式会社 細胞医薬組成物、疾患治療用キット及び細胞懸濁用溶液

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