CN117959350A - Osteoarthritis treatment composition based on composite stem cell SVF - Google Patents
Osteoarthritis treatment composition based on composite stem cell SVF Download PDFInfo
- Publication number
- CN117959350A CN117959350A CN202410371140.3A CN202410371140A CN117959350A CN 117959350 A CN117959350 A CN 117959350A CN 202410371140 A CN202410371140 A CN 202410371140A CN 117959350 A CN117959350 A CN 117959350A
- Authority
- CN
- China
- Prior art keywords
- svf
- stem cell
- treatment composition
- bone marrow
- mass
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
- 201000008482 osteoarthritis Diseases 0.000 title claims abstract description 30
- 210000000130 stem cell Anatomy 0.000 title claims abstract description 29
- 239000000203 mixture Substances 0.000 title claims abstract description 25
- 239000002131 composite material Substances 0.000 title claims abstract description 21
- 210000001185 bone marrow Anatomy 0.000 claims abstract description 33
- 210000002901 mesenchymal stem cell Anatomy 0.000 claims abstract description 32
- 239000003814 drug Substances 0.000 claims abstract description 26
- 210000004027 cell Anatomy 0.000 claims description 26
- 238000004821 distillation Methods 0.000 claims description 25
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 18
- 238000002360 preparation method Methods 0.000 claims description 14
- 235000001287 Guettarda speciosa Nutrition 0.000 claims description 13
- 244000020518 Carthamus tinctorius Species 0.000 claims description 12
- 235000003255 Carthamus tinctorius Nutrition 0.000 claims description 12
- 238000001914 filtration Methods 0.000 claims description 12
- 210000000577 adipose tissue Anatomy 0.000 claims description 10
- 241000830536 Tripterygium wilfordii Species 0.000 claims description 8
- 235000015398 thunder god vine Nutrition 0.000 claims description 8
- 244000131316 Panax pseudoginseng Species 0.000 claims description 7
- 235000003181 Panax pseudoginseng Nutrition 0.000 claims description 7
- CURLTUGMZLYLDI-UHFFFAOYSA-N Carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 claims description 6
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims description 6
- 238000009835 boiling Methods 0.000 claims description 6
- 238000005119 centrifugation Methods 0.000 claims description 6
- 239000003480 eluent Substances 0.000 claims description 6
- 239000007924 injection Substances 0.000 claims description 6
- 238000002347 injection Methods 0.000 claims description 6
- 239000007788 liquid Substances 0.000 claims description 6
- 210000005087 mononuclear cell Anatomy 0.000 claims description 6
- 239000002674 ointment Substances 0.000 claims description 6
- 239000002504 physiological saline solution Substances 0.000 claims description 6
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 6
- 210000004698 lymphocyte Anatomy 0.000 claims description 4
- 238000002156 mixing Methods 0.000 claims description 4
- 230000001105 regulatory effect Effects 0.000 claims description 4
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims description 3
- 102000029816 Collagenase Human genes 0.000 claims description 3
- 108060005980 Collagenase Proteins 0.000 claims description 3
- RTAQQCXQSZGOHL-UHFFFAOYSA-N Titanium Chemical compound [Ti] RTAQQCXQSZGOHL-UHFFFAOYSA-N 0.000 claims description 3
- 229910052799 carbon Inorganic materials 0.000 claims description 3
- 229910002092 carbon dioxide Inorganic materials 0.000 claims description 3
- 239000001569 carbon dioxide Substances 0.000 claims description 3
- 229960002424 collagenase Drugs 0.000 claims description 3
- 239000003636 conditioned culture medium Substances 0.000 claims description 3
- 238000001816 cooling Methods 0.000 claims description 3
- 238000012258 culturing Methods 0.000 claims description 3
- 230000029087 digestion Effects 0.000 claims description 3
- 238000007865 diluting Methods 0.000 claims description 3
- 238000000605 extraction Methods 0.000 claims description 3
- 239000000706 filtrate Substances 0.000 claims description 3
- 210000003692 ilium Anatomy 0.000 claims description 3
- 230000001376 precipitating effect Effects 0.000 claims description 3
- 238000010992 reflux Methods 0.000 claims description 3
- 239000011347 resin Substances 0.000 claims description 3
- 229920005989 resin Polymers 0.000 claims description 3
- 239000011780 sodium chloride Substances 0.000 claims description 3
- 230000001954 sterilising effect Effects 0.000 claims description 3
- 238000005406 washing Methods 0.000 claims description 3
- 239000008215 water for injection Substances 0.000 claims description 3
- 239000000463 material Substances 0.000 claims description 2
- 244000061520 Angelica archangelica Species 0.000 claims 2
- 230000000694 effects Effects 0.000 abstract description 8
- 238000011084 recovery Methods 0.000 abstract description 6
- 230000002757 inflammatory effect Effects 0.000 abstract description 5
- 210000001188 articular cartilage Anatomy 0.000 abstract description 4
- 210000000988 bone and bone Anatomy 0.000 abstract description 4
- 235000016709 nutrition Nutrition 0.000 abstract description 4
- 230000035764 nutrition Effects 0.000 abstract description 4
- 238000013329 compounding Methods 0.000 abstract description 2
- 230000009818 osteogenic differentiation Effects 0.000 abstract description 2
- 238000002660 stem cell treatment Methods 0.000 abstract description 2
- 230000000052 comparative effect Effects 0.000 description 14
- 241000125175 Angelica Species 0.000 description 11
- 210000000845 cartilage Anatomy 0.000 description 10
- 230000028993 immune response Effects 0.000 description 7
- 238000000034 method Methods 0.000 description 6
- 235000003143 Panax notoginseng Nutrition 0.000 description 5
- 241000180649 Panax notoginseng Species 0.000 description 5
- 238000012360 testing method Methods 0.000 description 5
- 230000002792 vascular Effects 0.000 description 4
- 206010061218 Inflammation Diseases 0.000 description 3
- 102000003777 Interleukin-1 beta Human genes 0.000 description 3
- 108090000193 Interleukin-1 beta Proteins 0.000 description 3
- 239000004480 active ingredient Substances 0.000 description 3
- 206010003246 arthritis Diseases 0.000 description 3
- 238000001514 detection method Methods 0.000 description 3
- 230000004054 inflammatory process Effects 0.000 description 3
- 230000008439 repair process Effects 0.000 description 3
- 238000011476 stem cell transplantation Methods 0.000 description 3
- 230000003110 anti-inflammatory effect Effects 0.000 description 2
- 201000010099 disease Diseases 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 150000004676 glycans Chemical class 0.000 description 2
- 239000012535 impurity Substances 0.000 description 2
- 210000000629 knee joint Anatomy 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 229920001282 polysaccharide Polymers 0.000 description 2
- 239000005017 polysaccharide Substances 0.000 description 2
- 238000012545 processing Methods 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 230000004044 response Effects 0.000 description 2
- 230000001225 therapeutic effect Effects 0.000 description 2
- 102000015427 Angiotensins Human genes 0.000 description 1
- 108010064733 Angiotensins Proteins 0.000 description 1
- 238000002965 ELISA Methods 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 229930189092 Notoginsenoside Natural products 0.000 description 1
- 210000001744 T-lymphocyte Anatomy 0.000 description 1
- DFBIRQPKNDILPW-CIVMWXNOSA-N Triptolide Chemical compound O=C1OCC([C@@H]2C3)=C1CC[C@]2(C)[C@]12O[C@H]1[C@@H]1O[C@]1(C(C)C)[C@@H](O)[C@]21[C@H]3O1 DFBIRQPKNDILPW-CIVMWXNOSA-N 0.000 description 1
- 230000006909 anti-apoptosis Effects 0.000 description 1
- 230000002300 anti-fibrosis Effects 0.000 description 1
- 230000003078 antioxidant effect Effects 0.000 description 1
- 210000003719 b-lymphocyte Anatomy 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000017531 blood circulation Effects 0.000 description 1
- 210000004204 blood vessel Anatomy 0.000 description 1
- 210000003321 cartilage cell Anatomy 0.000 description 1
- 210000001612 chondrocyte Anatomy 0.000 description 1
- 238000002648 combination therapy Methods 0.000 description 1
- 230000002596 correlated effect Effects 0.000 description 1
- 230000007850 degeneration Effects 0.000 description 1
- 230000000593 degrading effect Effects 0.000 description 1
- 229940088598 enzyme Drugs 0.000 description 1
- 239000012530 fluid Substances 0.000 description 1
- 230000008105 immune reaction Effects 0.000 description 1
- 230000004957 immunoregulator effect Effects 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 230000028709 inflammatory response Effects 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 239000002609 medium Substances 0.000 description 1
- 230000003349 osteoarthritic effect Effects 0.000 description 1
- 238000011056 performance test Methods 0.000 description 1
- 239000008055 phosphate buffer solution Substances 0.000 description 1
- 230000035755 proliferation Effects 0.000 description 1
- 230000000717 retained effect Effects 0.000 description 1
- 229910052702 rhenium Inorganic materials 0.000 description 1
- WUAPFZMCVAUBPE-UHFFFAOYSA-N rhenium atom Chemical compound [Re] WUAPFZMCVAUBPE-UHFFFAOYSA-N 0.000 description 1
- KINGXFAMZNIVNL-SXQDSXCISA-N safflor yellow A Natural products OC[C@@H]1O[C@H]2[C@H](OC3=C2C(=O)C(=C(O)C=Cc4ccc(O)cc4)C(=O)[C@]3(O)[C@@H]5O[C@H](CO)[C@@H](O)[C@H](O)[C@H]5O)[C@@H](O)[C@H]1O KINGXFAMZNIVNL-SXQDSXCISA-N 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- 208000024891 symptom Diseases 0.000 description 1
- 238000002560 therapeutic procedure Methods 0.000 description 1
- 238000002054 transplantation Methods 0.000 description 1
- YKUJZZHGTWVWHA-UHFFFAOYSA-N triptolide Natural products COC12CC3OC3(C(C)C)C(O)C14OC4CC5C6=C(CCC25C)C(=O)OC6 YKUJZZHGTWVWHA-UHFFFAOYSA-N 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/23—Apiaceae or Umbelliferae (Carrot family), e.g. dill, chervil, coriander or cumin
- A61K36/232—Angelica
-
- 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
- A61K35/35—Fat tissue; Adipocytes; Stromal cells; Connective tissues
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/25—Araliaceae (Ginseng family), e.g. ivy, aralia, schefflera or tetrapanax
- A61K36/258—Panax (ginseng)
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/28—Asteraceae or Compositae (Aster or Sunflower family), e.g. chamomile, feverfew, yarrow or echinacea
- A61K36/286—Carthamus (distaff thistle)
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/37—Celastraceae (Staff-tree or Bittersweet family), e.g. tripterygium or spindletree
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P19/00—Drugs for skeletal disorders
- A61P19/02—Drugs for skeletal disorders for joint disorders, e.g. arthritis, arthrosis
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P19/00—Drugs for skeletal disorders
- A61P19/04—Drugs for skeletal disorders for non-specific disorders of the connective tissue
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/30—Extraction of the material
- A61K2236/33—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones
- A61K2236/331—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones using water, e.g. cold water, infusion, tea, steam distillation or decoction
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/30—Extraction of the material
- A61K2236/39—Complex extraction schemes, e.g. fractionation or repeated extraction steps
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/50—Methods involving additional extraction steps
- A61K2236/53—Liquid-solid separation, e.g. centrifugation, sedimentation or crystallization
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/50—Methods involving additional extraction steps
- A61K2236/55—Liquid-liquid separation; Phase separation
Landscapes
- Health & Medical Sciences (AREA)
- Natural Medicines & Medicinal Plants (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Veterinary Medicine (AREA)
- Animal Behavior & Ethology (AREA)
- Medicinal Chemistry (AREA)
- Public Health (AREA)
- General Health & Medical Sciences (AREA)
- Pharmacology & Pharmacy (AREA)
- Chemical & Material Sciences (AREA)
- Biotechnology (AREA)
- Epidemiology (AREA)
- Botany (AREA)
- Mycology (AREA)
- Microbiology (AREA)
- Medical Informatics (AREA)
- Immunology (AREA)
- Cell Biology (AREA)
- Alternative & Traditional Medicine (AREA)
- Biomedical Technology (AREA)
- Organic Chemistry (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- General Chemical & Material Sciences (AREA)
- Developmental Biology & Embryology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Physical Education & Sports Medicine (AREA)
- Zoology (AREA)
- Virology (AREA)
- Rheumatology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Hematology (AREA)
- Orthopedic Medicine & Surgery (AREA)
- Pain & Pain Management (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
Abstract
The invention relates to the technical field of stem cell treatment, and in particular provides an osteoarthritis treatment composition based on composite stem cell SVF, which comprises the following components: 40-50% of SVF, 16-20% of bone marrow mesenchymal stem cells and the balance of traditional Chinese medicine extract. According to the invention, through compounding SVF and bone marrow mesenchymal stem cells in proportion, the osteogenic differentiation capacity of the bone marrow mesenchymal stem cells and the nutrition effect of SVF are considered, and the unique traditional Chinese medicine extract is matched, so that the recovery of bone articular cartilage is promoted, and simultaneously, inflammatory factors are effectively inhibited, so that the immune rejection is weakened.
Description
Technical Field
The invention relates to the technical field of stem cell treatment, in particular to an osteoarthritis treatment composition based on composite stem cell SVF.
Background
Osteoarthritis is a common disease and is mainly characterized by symptoms such as articular cartilage degeneration and pain. At present, stem cell transplantation treatment is an advanced treatment method, and after stem cells are injected into osteoarthritis joints, the stem cells can play a role in regulating immune and inflammatory reactions, reduce pain, secrete anti-apoptosis factors and anti-fibrosis factors to inhibit disease progress, promote endogenous stem cell cartilage repair, and can be differentiated into cartilage cells to help repair cartilage. The bone forming effect of the bone marrow mesenchymal stem cells is better, liang Jianji and the like describe the treatment condition of the bone marrow mesenchymal stem cells on arthritis in the 'joint cavity injection autologous bone marrow mesenchymal stem cells treatment light and medium bone arthritis'; furthermore, zhang Pingping et al, in bone marrow mesenchymal stem cells for the treatment of osteoarthritis: it is possible and future's trend to use mesenchymal stem cells for arthritis; meanwhile, as the knee joint cavity does not have rich blood vessels to provide nutrition, the survival time of the high-purity stem cells injected into the knee joint cavity is short, and the effective treatment time is greatly reduced. The newly separated adipose-derived Stromal Vascular Fraction (SVF) containing adipose-derived stem cells has higher nutrition and therapeutic effects because of the inclusion of the stromal vascular fraction, and the research of the cell fraction and the application of the vascular stroma fraction (SVF) in adipose tissue, such as primordial Bobo, is directed to the clinical therapeutic application of SVF.
However, the current stem cell transplantation therapy for osteoarthritis still has a limitation that there is little research on the treatment of multiple stem cells, such as the combination therapy of bone marrow mesenchymal stem cells and SVF, even though the response of the two to inflammatory environment, safety and immune compatibility at the time of transplantation may be different, the combined osteoarthritic treatment scheme of the combined stem cells SVF still is one of the leading fields to be explored.
Accordingly, the present invention aims to develop a composition for treating osteoarthritis by stem cell transplantation that is more effective and has low immune response.
Disclosure of Invention
In order to solve the above problems, the present invention aims to provide an osteoarthritis treatment composition based on composite stem cell SVF.
The invention provides an osteoarthritis treatment composition based on composite stem cell SVF, which consists of the following components:
40-50% of SVF, 16-20% of bone marrow mesenchymal stem cells and the balance of traditional Chinese medicine extract;
the preparation method of the bone marrow mesenchymal stem cells comprises the following steps:
Collecting autologous bone marrow in ilium through a bone marrow puncture needle, centrifuging 10 min at 1500r/min for 15ml each time, sucking upper adipose tissue, suspending and precipitating with equal mass PBS, injecting equal mass human lymphocyte separating liquid, centrifuging 20 min at 1500r/min, taking middle mononuclear cells after centrifugation, washing 2 times with PBS, sucking mononuclear cells, and re-suspending in a human mesenchymal stem cell conditioned medium with 30 times of mass; inoculating the cells with the cell density of 2X 10 5/mL into a culture flask, culturing the cells in a culture box with 5% carbon dioxide at 37 ℃ for 48 hours, adding physiological saline to wash the cells, and adjusting the cell density to 1X 10 5/mL to obtain the bone marrow mesenchymal stem cells;
The preparation method of the SVF comprises the following steps:
Collecting autologous adipose tissue by operation, adding into an SVF cell preparation device, adding 0.1% type I collagenase with equal adipose tissue mass into the SVF cell preparation device, shaking table digestion at 37deg.C for 50min, centrifugation at 180 r/min for 10 min, filtering with 400 μm filter screen, adjusting cell density to 1× 5/mL with physiological saline, and collecting to obtain SVF;
The preparation method of the traditional Chinese medicine extract comprises the following steps:
taking 10-14% of dried angelica, 14-18% of safflower, 24-30% of pseudo-ginseng and the balance of tripterygium wilfordii, crushing the crushed materials with the fineness of 50 meshes, adding 15 times of pure water, carrying out reflux extraction for 4 times, each time for 45min, filtering, and carrying out first reduced pressure distillation for 90min at the temperature of 60-70 ℃ under the pressure of 0.3 MPa; adding 85% ethanol with 10 times of the mass of the residues, uniformly mixing, standing for 5 hours, filtering with 100 meshes, collecting filtrate, carrying out second reduced pressure distillation at 60-70 ℃ under 0.3MPa to recover ethanol until no more outflow occurs, and collecting residues to obtain a traditional Chinese medicine ointment;
regulating the pH value of the traditional Chinese medicine ointment to 5.0, adsorbing by using a macroporous resin column, collecting the tail liquid, eluting by using 60% ethanol, wherein the using amount of the eluent is 2BV, the flow rate is 2BV/h, collecting the eluent, carrying out third reduced pressure distillation for 70min at the temperature of 55-60 ℃ under the pressure of 0.4MPa, adding NaCl into the collected residues according to the concentration of 0.013g/ml, boiling for 40min, adding active carbon for injection according to the concentration of 0.005g/ml, boiling for 45min, filtering by using a titanium rod filter when cooling to 30 ℃, diluting to 5 times of mass by adding pure water for injection, and sterilizing for 30min at the temperature of 115 ℃ to obtain the traditional Chinese medicine extract.
Further, the mass fraction of the SVF is 47%.
Further, the mass fraction of the bone marrow mesenchymal stem cells is 18%.
Further, the mass fraction of the angelica is 12%.
Further, the mass fraction of the safflower is 16%.
Further, the mass fraction of the pseudo-ginseng is 28%.
Further, the temperature of the first reduced pressure distillation was 67 ℃.
Further, the temperature of the second reduced pressure distillation was 65 ℃.
Further, the temperature of the third reduced pressure distillation was 58 ℃.
The invention has the following beneficial effects:
according to the invention, through compounding SVF and bone marrow mesenchymal stem cells in proportion, the osteogenic differentiation capacity of the bone marrow mesenchymal stem cells and the nutrition effect of SVF are considered, and the unique traditional Chinese medicine extract is matched, so that the recovery of bone articular cartilage is promoted, and simultaneously, inflammatory factors are effectively inhibited, so that the immune rejection is weakened.
The components of angelica polysaccharide, safflower yellow and the like in the angelica and the safflower can promote blood circulation, reduce the generation of angiotensin, reduce inflammatory reaction and relieve pain; notoginseng radix contains notoginsenoside and polysaccharide, has antiinflammatory and antioxidant effects, and can inhibit activity of cartilage degrading enzyme, and promote proliferation and repair of chondrocyte; tripterygium wilfordii contains active ingredients such as triptolide, and the ingredients have anti-inflammatory and immunoregulatory effects, and can inhibit the activities of T cells and B cells and reduce immune responses such as inflammation. The traditional Chinese medicine is processed into the traditional Chinese medicine extract special for the invention under the special preparation method, the medicine properties of the traditional Chinese medicine extract and the traditional Chinese medicine extract are matched with each other, under the injection type treatment background, the recovery of cartilage tissues is accelerated, the response of SVF and bone marrow mesenchymal stem cells to the immune environment is coordinated, and the inflammatory response is reduced.
The three times of reduced pressure distillation is carried out at the temperature which is matched with the respective distillation pressure to remove impurities, and the processing of the traditional Chinese medicine active ingredients is carried out at the temperature, so that the processing effect is not good when the temperature is too low, the impurities are difficult to volatilize, and the active ingredients are damaged when the temperature is too high.
Detailed Description
In order to more clearly illustrate the overall concept of the present invention, the following describes the overall scheme of the present invention in detail by way of examples; in the following description, numerous specific details are set forth in order to provide a more thorough understanding of the present invention; it will be apparent, however, to one skilled in the art that the invention may be practiced without one or more of these details; in other instances, well-known features have not been described in detail in order to avoid obscuring the invention.
SVF of the present invention refers to adipose-derived stromal vascular fraction; the PBS of the invention is a phosphate buffer solution with ph of 7.4 purchased from Xiamen seamark technologies Co., ltd; the present inventors lymphocyte isolates were purchased from rhenium (Shanghai) Biochemical technologies Co., ltd, cat# LBX110; the SVF cell preparation apparatus of the present invention was purchased from Shanghai Polymu medical instruments Co., ltd., model BCV-100.
Unless otherwise specified, the starting components in the examples below are commercially available, and the laboratory apparatus used is a laboratory conventional laboratory apparatus, and the performance test methods are known in the art.
The preferred embodiment is as follows:
Example 1:
the mesenchymal stem cells are prepared and obtained by the following method:
Collecting autologous bone marrow in ilium through a bone marrow puncture needle, centrifuging 10 min at 1500r/min for 15ml each time, sucking upper adipose tissue, suspending and precipitating with equal mass PBS, injecting equal mass human lymphocyte separating liquid, centrifuging 20 min at 1500r/min, taking middle mononuclear cells after centrifugation, washing 2 times with PBS, sucking mononuclear cells, and re-suspending in a human mesenchymal stem cell conditioned medium with 30 times of mass; inoculating the cells with the cell density of 2X 10 5/mL into a culture flask, culturing the cells in a culture box with 5% carbon dioxide at 37 ℃ for 48 hours, adding physiological saline to wash the cells, and adjusting the cell density to 1X 10 5/mL to obtain the bone marrow mesenchymal stem cells;
The SVF is prepared by the following method:
Collecting autologous adipose tissue by operation, adding into an SVF cell preparation device, adding 0.1% type I collagenase with equal adipose tissue mass into the SVF cell preparation device, shaking table digestion at 37deg.C for 50min, centrifugation at 180 r/min for 10 min, filtering with 400 μm filter screen, adjusting cell density to 1× 5/mL with physiological saline, and collecting to obtain SVF;
The traditional Chinese medicine extract is prepared by the following method:
Taking 12% of dried angelica, 16% of safflower, 28% of pseudo-ginseng and the balance of tripterygium wilfordii, crushing the mixture with the fineness of 50 meshes, adding 15 times of pure water, carrying out reflux extraction for 4 times, 45 minutes each time, filtering, and carrying out first reduced pressure distillation for 90 minutes at 67 ℃ under the pressure of 0.3 MPa; adding 85% ethanol 10 times of the residue, mixing, standing for 5 hr, filtering with 100 mesh sieve, collecting filtrate, distilling under reduced pressure at 65deg.C and 0.3MPa for the second time to recover ethanol until no more flow out, and collecting residue to obtain Chinese medicinal unguent;
regulating the pH value of the traditional Chinese medicine ointment to 5.0, adsorbing by using a macroporous resin column, collecting the tail liquid, eluting by using 60% ethanol, wherein the using amount of the eluent is 2BV, the flow rate is 2BV/h, collecting the eluent, carrying out third reduced pressure distillation for 70min at 58 ℃ under the pressure of 0.4MPa, adding NaCl into the collected residues according to the ratio of 0.013g/ml, boiling for 40min, adding active carbon for injection according to the ratio of 0.005g/ml, boiling for 45min, cooling to 30 ℃, filtering by using a titanium rod filter, diluting to 5 times of mass by adding pure water for injection, and sterilizing for 30min at 115 ℃ to obtain the traditional Chinese medicine extract.
The osteoarthritis treatment composition based on the fat stem cell SVF is prepared by mixing 47% of SVF, 18% of bone marrow mesenchymal stem cells and the rest of traditional Chinese medicine extract.
Examples 2 to 17:
example 2 differs from example 1 only in that the mass fraction of SVF is 40%;
Example 3 differs from example 1 only in that the mass fraction of SVF is 50%;
example 4 differs from example 1 only in that the mass fraction of bone marrow mesenchymal stem cells is 16%;
example 5 differs from example 1 only in that the mass fraction of bone marrow mesenchymal stem cells is 20%;
Example 6 differs from example 1 only in that the mass fraction of angelica is 10%;
example 7 differs from example 1 only in that the mass fraction of angelica is 14%;
example 8 differs from example 1 only in that the mass fraction of safflower is 14%;
example 9 differs from example 1 only in that the mass fraction of safflower is 18%;
example 10 differs from example 1 only in that the mass fraction of pseudo-ginseng is 24%;
Example 11 differs from example 1 only in that the mass fraction of pseudo-ginseng is 30%;
example 12 differs from example 1 only in that the temperature of the first reduced pressure distillation is 60 ℃;
example 13 differs from example 1 only in that the temperature of the first reduced pressure distillation is 70 ℃;
Example 14 differs from example 1 only in that the temperature of the second reduced pressure distillation is 60 ℃;
Example 15 differs from example 1 only in that the temperature of the second reduced pressure distillation is 70 ℃;
Example 16 differs from example 1 only in that the temperature of the third reduced pressure distillation is 55 ℃;
Example 17 differs from example 1 only in that the temperature of the third reduced pressure distillation is 60 ℃;
Comparative examples 1 to 12:
Comparative example 1 differs from example 1 only in that the mass ratio of the traditional Chinese medicine extract to the equivalent mass is 47:18 and bone marrow mesenchymal stem cells;
comparative example 2 differs from example 1 only in that the traditional Chinese medicine extract was replaced with SVF of equal quality;
Comparative example 3 differs from example 1 only in that the traditional Chinese medicine extract is replaced with bone marrow mesenchymal stem cells of equal mass;
comparative example 4 differs from example 1 only in that SVF was exchanged for equal mass of bone marrow mesenchymal stem cells;
Comparative example 5 differs from example 1 only in that the bone marrow mesenchymal stem cells were replaced with equal mass of SVF;
Comparative example 6 differs from example 1 only in that 18% by mass of dried angelica, 16% by mass of safflower, 28% by mass of notoginseng and the balance of tripterygium wilfordii were taken.
Comparative example 7 differs from example 1 only in that 12% of dried angelica, 22% of safflower, 28% of notoginseng and the balance of tripterygium wilfordii were taken;
comparative example 8 differs from example 1 only in that 12% of dried angelica, 16% of safflower, 40% of notoginseng and the balance of tripterygium wilfordii were taken;
Comparative example 9 differs from example 1 only in that 6% by mass of dried angelica, 10% by mass of safflower, 18% by mass of notoginseng and the balance of tripterygium wilfordii were taken;
comparative example 10 differs from example 1 only in that the temperature of the first reduced pressure distillation was 80 ℃;
comparative example 11 differs from example 1 only in that the temperature of the second reduced pressure distillation was 80 ℃;
comparative example 12 differs from example 1 only in that the temperature of the third reduced pressure distillation was 70 ℃.
Taking the osteoarthritis treatment composition based on the composite stem cell SVF obtained in each example, 4mL of the composition was slowly injected into the joint of each human suffering from osteoarthritis by using a syringe in subjects suffering from the same degree of osteoarthritis, the same degree being based on that the cartilage loss ratio of the affected part of the subjects is 20% observed by MRI, the subjects are 90 persons, the subjects are divided into 3 groups of experimental groups of 87 persons and 3 blank groups of 3 persons, 29 persons in each experimental group are respectively injected with the osteoarthritis treatment composition based on the composite stem cell SVF obtained in each 29 persons in examples 1-17 and 29 examples in comparative examples 1-12, the injections are repeated once again in the second and third months, the blank group of 3 persons is not treated, and the following detection is carried out at the 35 th week after the treatment.
Immune response test:
the amount of the IL-1 beta inflammatory factor is used for representing the immune response, and the two factors are positively correlated; the joint fluid is taken, the IL-1 beta inflammatory factor expression is detected by adopting an ELISA method, the content detection is shown in table 1, the detection result unit is pg/ml, and the whole digits are reserved.
Cartilage recovery test:
The joint of the patient was subjected to MRI observation to detect the joint recovery state, i.e., the cartilage loss area. The area of articular cartilage loss after 35 weeks of treatment is shown in table 1, in units of whole digits remaining.
The above test results were taken as an average of 3 persons treated with the composite stem cell SVF-based osteoarthritis treatment composition and an average of 3 persons in the blank group obtained in each example and the whole number was retained.
Table 1: immune response test and cartilage recovery test results for each example:
From the data in table 1, it is clear that the osteoarthritis treatment composition of the present invention, especially of example 1, based on the composite stem cell SVF, has lower IL-1 β content and lower cartilage loss area compared to the other examples; that is, the embodiment of the present invention, and in particular, the composite stem cell SVF-based osteoarthritis treatment composition of embodiment 1 of the present invention has more effective cartilage restoring effect and low immune response effect, and can treat osteoarthritis more effectively and with low immune response.
The foregoing is merely exemplary of the present invention and is not intended to limit the present invention; various modifications and variations of the present invention will be apparent to those skilled in the art; any modification, equivalent replacement, improvement, etc. which come within the spirit and principles of the invention are to be included in the scope of the claims of the present invention.
Claims (9)
1. An osteoarthritis treatment composition based on a composite stem cell SVF, characterized in that the composite stem cell SVF-based osteoarthritis treatment composition consists of:
40-50% of SVF, 16-20% of bone marrow mesenchymal stem cells and the balance of traditional Chinese medicine extract;
the preparation method of the bone marrow mesenchymal stem cells comprises the following steps:
Collecting autologous bone marrow in ilium through a bone marrow puncture needle, centrifuging 10 min at 1500r/min for 15ml each time, sucking upper adipose tissue, suspending and precipitating with equal mass PBS, injecting equal mass human lymphocyte separating liquid, centrifuging 20 min at 1500r/min, taking middle mononuclear cells after centrifugation, washing 2 times with PBS, sucking mononuclear cells, and re-suspending in a human mesenchymal stem cell conditioned medium with 30 times of mass; inoculating the cells with the cell density of 2X 10 5/mL into a culture flask, culturing the cells in a culture box with 5% carbon dioxide at 37 ℃ for 48 hours, adding physiological saline to wash the cells, and adjusting the cell density to 1X 10 5/mL to obtain the bone marrow mesenchymal stem cells;
The preparation method of the SVF comprises the following steps:
Collecting autologous adipose tissue by operation, adding into an SVF cell preparation device, adding 0.1% type I collagenase with equal adipose tissue mass into the SVF cell preparation device, shaking table digestion at 37deg.C for 50min, centrifugation at 180 r/min for 10 min, filtering with 400 μm filter screen, adjusting cell density to 1× 5/mL with physiological saline, and collecting to obtain SVF;
The preparation method of the traditional Chinese medicine extract comprises the following steps:
taking 10-14% of dried angelica, 14-18% of safflower, 24-30% of pseudo-ginseng and the balance of tripterygium wilfordii, crushing the crushed materials with the fineness of 50 meshes, adding 15 times of pure water, carrying out reflux extraction for 4 times, each time for 45min, filtering, and carrying out first reduced pressure distillation for 90min at the temperature of 60-70 ℃ under the pressure of 0.3 MPa; adding 85% ethanol with 10 times of the mass of the residues, uniformly mixing, standing for 5 hours, filtering with 100 meshes, collecting filtrate, carrying out second reduced pressure distillation at 60-70 ℃ under 0.3MPa to recover ethanol until no more outflow occurs, and collecting residues to obtain a traditional Chinese medicine ointment;
regulating the pH value of the traditional Chinese medicine ointment to 5.0, adsorbing by using a macroporous resin column, collecting the tail liquid, eluting by using 60% ethanol, wherein the using amount of the eluent is 2BV, the flow rate is 2BV/h, collecting the eluent, carrying out third reduced pressure distillation for 70min at the temperature of 55-60 ℃ under the pressure of 0.4MPa, adding NaCl into the collected residues according to the concentration of 0.013g/ml, boiling for 40min, adding active carbon for injection according to the concentration of 0.005g/ml, boiling for 45min, filtering by using a titanium rod filter when cooling to 30 ℃, diluting to 5 times of mass by adding pure water for injection, and sterilizing for 30min at the temperature of 115 ℃ to obtain the traditional Chinese medicine extract.
2. The composite stem cell SVF-based osteoarthritis treatment composition of claim 1, wherein the mass fraction of SVF is 47%.
3. The composite stem cell SVF-based osteoarthritis treatment composition of claim 1, wherein the mass fraction of bone marrow mesenchymal stem cells is 18%.
4. The composite stem cell SVF-based osteoarthritis treatment composition of claim 1, wherein the angelica is 12% in mass fraction.
5. The composite stem cell SVF-based osteoarthritis treatment composition of claim 1, wherein the mass fraction of safflower is 16%.
6. The composite stem cell SVF-based osteoarthritis treatment composition of claim 1, wherein the mass fraction of pseudo-ginseng is 28%.
7. The composite stem cell SVF-based osteoarthritis treatment composition of claim 1, wherein the temperature of the first reduced pressure distillation is 67 ℃.
8. The composite stem cell SVF-based osteoarthritis treatment composition of claim 1, wherein the temperature of the second reduced pressure distillation is 65 ℃.
9. The composite stem cell SVF-based osteoarthritis treatment composition of claim 1, wherein the temperature of the third reduced pressure distillation is 58 ℃.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202410371140.3A CN117959350B (en) | 2024-03-29 | 2024-03-29 | Osteoarthritis treatment composition based on composite stem cell SVF |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202410371140.3A CN117959350B (en) | 2024-03-29 | 2024-03-29 | Osteoarthritis treatment composition based on composite stem cell SVF |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN117959350A true CN117959350A (en) | 2024-05-03 |
| CN117959350B CN117959350B (en) | 2024-07-05 |
Family
ID=90858331
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202410371140.3A Active CN117959350B (en) | 2024-03-29 | 2024-03-29 | Osteoarthritis treatment composition based on composite stem cell SVF |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN117959350B (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN118542930A (en) * | 2024-07-30 | 2024-08-27 | 广东赛尔生物科技有限公司 | Stem cell and gelatin sponge solution composition for knee joint injection |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106011056A (en) * | 2016-01-20 | 2016-10-12 | 北京希普生国际生物医学研究院 | Preparation method of mesenchymal stem cells for skin acne treatment, preparation method and product thereof |
| CN108653329A (en) * | 2018-05-28 | 2018-10-16 | 广东唯泰生物科技有限公司 | It is a kind of to be used to treat drug of osteoarthritis and preparation method thereof |
| CN110564835A (en) * | 2018-06-06 | 2019-12-13 | 屈正 | research method of effect of celastrus orbiculatus extract containing drug serum on HBV and MIF |
| US20220125848A1 (en) * | 2019-02-07 | 2022-04-28 | Direct Biologics Llc | Method for treating osteoarthritis with a combination of mesenchymal stem cell exosomes, synovial mesencymal stem cells, and scaffolds |
| CN116115661A (en) * | 2022-12-20 | 2023-05-16 | 湖南华启细胞生物科技有限公司 | Adipose-derived mesenchymal stem cell preparation for treating joint degenerative disease and preparation method thereof |
-
2024
- 2024-03-29 CN CN202410371140.3A patent/CN117959350B/en active Active
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106011056A (en) * | 2016-01-20 | 2016-10-12 | 北京希普生国际生物医学研究院 | Preparation method of mesenchymal stem cells for skin acne treatment, preparation method and product thereof |
| CN108653329A (en) * | 2018-05-28 | 2018-10-16 | 广东唯泰生物科技有限公司 | It is a kind of to be used to treat drug of osteoarthritis and preparation method thereof |
| CN110564835A (en) * | 2018-06-06 | 2019-12-13 | 屈正 | research method of effect of celastrus orbiculatus extract containing drug serum on HBV and MIF |
| US20220125848A1 (en) * | 2019-02-07 | 2022-04-28 | Direct Biologics Llc | Method for treating osteoarthritis with a combination of mesenchymal stem cell exosomes, synovial mesencymal stem cells, and scaffolds |
| CN116115661A (en) * | 2022-12-20 | 2023-05-16 | 湖南华启细胞生物科技有限公司 | Adipose-derived mesenchymal stem cell preparation for treating joint degenerative disease and preparation method thereof |
Non-Patent Citations (1)
| Title |
|---|
| 王纬玲;: "祛痹胶囊的药理作用研究", 延安大学学报(医学科学版), no. 03, 15 July 2009 (2009-07-15) * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN118542930A (en) * | 2024-07-30 | 2024-08-27 | 广东赛尔生物科技有限公司 | Stem cell and gelatin sponge solution composition for knee joint injection |
Also Published As
| Publication number | Publication date |
|---|---|
| CN117959350B (en) | 2024-07-05 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN117959350B (en) | Osteoarthritis treatment composition based on composite stem cell SVF | |
| CN117815335B (en) | Osteoarthritis treatment composition based on adipose-derived stem cells SVF and preparation method thereof | |
| JP5048673B2 (en) | A kind of extract that prevents or treats thrombotic diseases | |
| JP6471259B2 (en) | Cell preparation for hair regeneration | |
| CN108478600B (en) | Application of mesenchymal stem cell exosome in preparation of medicine for preventing and treating osteoporosis | |
| JP5572777B2 (en) | Cell preparation containing adipocytes | |
| CN104284601B (en) | Secrete for promoting insulin-like growth factor and the white radix-polygoni multiflori extract of bone growth and Radix Dipsaci extract and manufacture method thereof | |
| CN100446779C (en) | Method for preparing acanthopanax senticousus extracting liquid/extraction | |
| CN114984047B (en) | Application of plasma exosome in preparation of medicine for treating osteoporosis | |
| EP3785721A1 (en) | Stem cell filtrate formulation and method for preparing same | |
| US9956317B2 (en) | Clinical applications of formulations containing adipose-derived stem cells | |
| CN110833187A (en) | Enzyme stock solution and preparation method thereof | |
| CN110179888A (en) | A kind of arcotic oral solution and preparation method thereof for gastric operation | |
| CN110559327B (en) | Liver-protecting product and preparation method thereof | |
| CN111558019B (en) | Traditional Chinese medicine composition with effects of benefiting qi, nourishing blood, regulating menstruation and relieving pain and preparation method and application thereof | |
| CN110897971B (en) | Hair care cleaning composition containing selaginella tamariscina and preparation method thereof | |
| CN113274421A (en) | Anti-inflammatory exosome preparation for knee joint treatment and preparation method and application thereof | |
| US20200155608A1 (en) | Therapeutic Treatments Via Intravenous Infusion Of Mesenchymal Stem Cells | |
| CN109288880A (en) | The extracting method of Bulbilus boussingaultiae pseudobaselloidis colloid | |
| CN113116970A (en) | Stem cell skin repair liquid containing gardenia jasminoides extract and preparation method thereof | |
| CN104257755A (en) | Composition with effect of treating diabetes as well as preparation method and application of composition | |
| CN113662970B (en) | Application of umbilical cord stem cell preparation in preparation of medicines for treating osteoarthritis | |
| SONNANSTINE | BIOLOGICS IN ORTHOPEDICS | |
| CN1583164A (en) | Preparing procedure of transfer factor for intravenous injection | |
| Ruiz‐Lopez et al. | Complications in Regenerative Medicine |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |