WO2024110937A1 - Compositions based on a mixture of bioactive molecules and exosomes for use in the treatment of conditions requiring tissue repair and regeneration - Google Patents
Compositions based on a mixture of bioactive molecules and exosomes for use in the treatment of conditions requiring tissue repair and regeneration Download PDFInfo
- Publication number
- WO2024110937A1 WO2024110937A1 PCT/IB2023/061927 IB2023061927W WO2024110937A1 WO 2024110937 A1 WO2024110937 A1 WO 2024110937A1 IB 2023061927 W IB2023061927 W IB 2023061927W WO 2024110937 A1 WO2024110937 A1 WO 2024110937A1
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- treatment
- composition
- colostrum
- exosomes
- mixture
- Prior art date
Links
- 239000000203 mixture Substances 0.000 title claims abstract description 159
- 210000001808 exosome Anatomy 0.000 title claims abstract description 60
- 230000000975 bioactive effect Effects 0.000 title claims abstract description 44
- 238000011282 treatment Methods 0.000 title claims abstract description 35
- 230000017423 tissue regeneration Effects 0.000 title claims abstract description 15
- 238000011069 regeneration method Methods 0.000 title claims abstract description 13
- 241001465754 Metazoa Species 0.000 claims abstract description 8
- 238000002347 injection Methods 0.000 claims abstract description 7
- 239000007924 injection Substances 0.000 claims abstract description 7
- 230000000699 topical effect Effects 0.000 claims abstract description 5
- 210000003022 colostrum Anatomy 0.000 claims description 47
- 235000021277 colostrum Nutrition 0.000 claims description 47
- 238000000034 method Methods 0.000 claims description 38
- 239000003102 growth factor Substances 0.000 claims description 33
- 238000009472 formulation Methods 0.000 claims description 26
- 210000001519 tissue Anatomy 0.000 claims description 26
- 241000282414 Homo sapiens Species 0.000 claims description 25
- 108060003951 Immunoglobulin Proteins 0.000 claims description 24
- 102000018358 immunoglobulin Human genes 0.000 claims description 24
- 102000004127 Cytokines Human genes 0.000 claims description 22
- 108090000695 Cytokines Proteins 0.000 claims description 22
- 229940072221 immunoglobulins Drugs 0.000 claims description 21
- 230000001172 regenerating effect Effects 0.000 claims description 21
- 230000004936 stimulating effect Effects 0.000 claims description 18
- 239000003181 biological factor Substances 0.000 claims description 15
- 241000283690 Bos taurus Species 0.000 claims description 13
- 239000003814 drug Substances 0.000 claims description 12
- 210000000130 stem cell Anatomy 0.000 claims description 12
- 239000012528 membrane Substances 0.000 claims description 11
- 238000001356 surgical procedure Methods 0.000 claims description 11
- 208000027418 Wounds and injury Diseases 0.000 claims description 10
- 230000000844 anti-bacterial effect Effects 0.000 claims description 10
- 230000000840 anti-viral effect Effects 0.000 claims description 10
- 102000000989 Complement System Proteins Human genes 0.000 claims description 9
- 230000035876 healing Effects 0.000 claims description 9
- 108010037362 Extracellular Matrix Proteins Proteins 0.000 claims description 8
- 102000010834 Extracellular Matrix Proteins Human genes 0.000 claims description 8
- 229910052588 hydroxylapatite Inorganic materials 0.000 claims description 8
- XYJRXVWERLGGKC-UHFFFAOYSA-D pentacalcium;hydroxide;triphosphate Chemical compound [OH-].[Ca+2].[Ca+2].[Ca+2].[Ca+2].[Ca+2].[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O XYJRXVWERLGGKC-UHFFFAOYSA-D 0.000 claims description 8
- 239000003634 thrombocyte concentrate Substances 0.000 claims description 8
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 8
- 108010069112 Complement System Proteins Proteins 0.000 claims description 7
- 230000006378 damage Effects 0.000 claims description 7
- 230000007547 defect Effects 0.000 claims description 7
- 102000004190 Enzymes Human genes 0.000 claims description 6
- 108090000790 Enzymes Proteins 0.000 claims description 6
- 241000282412 Homo Species 0.000 claims description 6
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims description 6
- 206010052428 Wound Diseases 0.000 claims description 6
- 210000000481 breast Anatomy 0.000 claims description 6
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 claims description 6
- 239000012153 distilled water Substances 0.000 claims description 6
- 208000014674 injury Diseases 0.000 claims description 6
- 210000004185 liver Anatomy 0.000 claims description 6
- 208000015122 neurodegenerative disease Diseases 0.000 claims description 6
- 210000000056 organ Anatomy 0.000 claims description 6
- 239000011780 sodium chloride Substances 0.000 claims description 6
- KIUKXJAPPMFGSW-DNGZLQJQSA-N (2S,3S,4S,5R,6R)-6-[(2S,3R,4R,5S,6R)-3-Acetamido-2-[(2S,3S,4R,5R,6R)-6-[(2R,3R,4R,5S,6R)-3-acetamido-2,5-dihydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-2-carboxy-4,5-dihydroxyoxan-3-yl]oxy-5-hydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-3,4,5-trihydroxyoxane-2-carboxylic acid Chemical compound CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](O[C@H]3[C@@H]([C@@H](O)[C@H](O)[C@H](O3)C(O)=O)O)[C@H](O)[C@@H](CO)O2)NC(C)=O)[C@@H](C(O)=O)O1 KIUKXJAPPMFGSW-DNGZLQJQSA-N 0.000 claims description 5
- 238000000605 extraction Methods 0.000 claims description 5
- 229920002674 hyaluronan Polymers 0.000 claims description 5
- 229960003160 hyaluronic acid Drugs 0.000 claims description 5
- 230000003239 periodontal effect Effects 0.000 claims description 5
- 206010016654 Fibrosis Diseases 0.000 claims description 4
- 239000001506 calcium phosphate Substances 0.000 claims description 4
- 150000001875 compounds Chemical class 0.000 claims description 4
- 235000013365 dairy product Nutrition 0.000 claims description 4
- 201000010099 disease Diseases 0.000 claims description 4
- 230000002708 enhancing effect Effects 0.000 claims description 4
- 230000004761 fibrosis Effects 0.000 claims description 4
- 239000000499 gel Substances 0.000 claims description 4
- 210000004072 lung Anatomy 0.000 claims description 4
- 239000002674 ointment Substances 0.000 claims description 4
- 230000003716 rejuvenation Effects 0.000 claims description 4
- 229940078499 tricalcium phosphate Drugs 0.000 claims description 4
- 229910000391 tricalcium phosphate Inorganic materials 0.000 claims description 4
- 201000004384 Alopecia Diseases 0.000 claims description 3
- 208000023275 Autoimmune disease Diseases 0.000 claims description 3
- 108020004414 DNA Proteins 0.000 claims description 3
- 108010052285 Membrane Proteins Proteins 0.000 claims description 3
- 108010089610 Nuclear Proteins Proteins 0.000 claims description 3
- 102000007999 Nuclear Proteins Human genes 0.000 claims description 3
- 231100000360 alopecia Toxicity 0.000 claims description 3
- 150000001413 amino acids Chemical class 0.000 claims description 3
- 210000001124 body fluid Anatomy 0.000 claims description 3
- 239000010839 body fluid Substances 0.000 claims description 3
- 210000004556 brain Anatomy 0.000 claims description 3
- 210000003169 central nervous system Anatomy 0.000 claims description 3
- -1 clots Substances 0.000 claims description 3
- 238000002316 cosmetic surgery Methods 0.000 claims description 3
- 230000001086 cytosolic effect Effects 0.000 claims description 3
- 210000003238 esophagus Anatomy 0.000 claims description 3
- 210000002216 heart Anatomy 0.000 claims description 3
- 230000001969 hypertrophic effect Effects 0.000 claims description 3
- 238000011065 in-situ storage Methods 0.000 claims description 3
- 210000003734 kidney Anatomy 0.000 claims description 3
- 210000000867 larynx Anatomy 0.000 claims description 3
- 150000002632 lipids Chemical class 0.000 claims description 3
- 108020004999 messenger RNA Proteins 0.000 claims description 3
- 108091070501 miRNA Proteins 0.000 claims description 3
- 239000002679 microRNA Substances 0.000 claims description 3
- 230000000399 orthopedic effect Effects 0.000 claims description 3
- 201000008482 osteoarthritis Diseases 0.000 claims description 3
- 210000003800 pharynx Anatomy 0.000 claims description 3
- 239000004033 plastic Substances 0.000 claims description 3
- 239000000843 powder Substances 0.000 claims description 3
- 238000002278 reconstructive surgery Methods 0.000 claims description 3
- 230000008439 repair process Effects 0.000 claims description 3
- 208000002874 Acne Vulgaris Diseases 0.000 claims description 2
- 201000008283 Atrophic Rhinitis Diseases 0.000 claims description 2
- 206010003694 Atrophy Diseases 0.000 claims description 2
- 206010008570 Chloasma Diseases 0.000 claims description 2
- 208000032544 Cicatrix Diseases 0.000 claims description 2
- 208000035473 Communicable disease Diseases 0.000 claims description 2
- 208000028006 Corneal injury Diseases 0.000 claims description 2
- 208000003556 Dry Eye Syndromes Diseases 0.000 claims description 2
- 206010013774 Dry eye Diseases 0.000 claims description 2
- 208000010228 Erectile Dysfunction Diseases 0.000 claims description 2
- 208000002260 Keloid Diseases 0.000 claims description 2
- 208000003351 Melanosis Diseases 0.000 claims description 2
- 208000029549 Muscle injury Diseases 0.000 claims description 2
- 208000010886 Peripheral nerve injury Diseases 0.000 claims description 2
- 206010039088 Rhinitis atrophic Diseases 0.000 claims description 2
- 206010040943 Skin Ulcer Diseases 0.000 claims description 2
- 208000028990 Skin injury Diseases 0.000 claims description 2
- 206010040925 Skin striae Diseases 0.000 claims description 2
- 208000031439 Striae Distensae Diseases 0.000 claims description 2
- 208000002847 Surgical Wound Diseases 0.000 claims description 2
- 208000021945 Tendon injury Diseases 0.000 claims description 2
- 208000030886 Traumatic Brain injury Diseases 0.000 claims description 2
- 206010045210 Tympanic Membrane Perforation Diseases 0.000 claims description 2
- 206010047642 Vitiligo Diseases 0.000 claims description 2
- 206010047791 Vulvovaginal dryness Diseases 0.000 claims description 2
- 206010000496 acne Diseases 0.000 claims description 2
- 230000037444 atrophy Effects 0.000 claims description 2
- 201000007717 corneal ulcer Diseases 0.000 claims description 2
- 239000006071 cream Substances 0.000 claims description 2
- 239000006260 foam Substances 0.000 claims description 2
- 210000005260 human cell Anatomy 0.000 claims description 2
- 208000000069 hyperpigmentation Diseases 0.000 claims description 2
- 230000003810 hyperpigmentation Effects 0.000 claims description 2
- 201000001881 impotence Diseases 0.000 claims description 2
- 208000028867 ischemia Diseases 0.000 claims description 2
- 210000001117 keloid Anatomy 0.000 claims description 2
- 210000003041 ligament Anatomy 0.000 claims description 2
- 239000006193 liquid solution Substances 0.000 claims description 2
- 239000006210 lotion Substances 0.000 claims description 2
- 210000004086 maxillary sinus Anatomy 0.000 claims description 2
- 230000000626 neurodegenerative effect Effects 0.000 claims description 2
- 108020004707 nucleic acids Proteins 0.000 claims description 2
- 150000007523 nucleic acids Chemical class 0.000 claims description 2
- 102000039446 nucleic acids Human genes 0.000 claims description 2
- 239000006072 paste Substances 0.000 claims description 2
- 230000002085 persistent effect Effects 0.000 claims description 2
- 238000001959 radiotherapy Methods 0.000 claims description 2
- 231100000241 scar Toxicity 0.000 claims description 2
- 230000037387 scars Effects 0.000 claims description 2
- 231100000019 skin ulcer Toxicity 0.000 claims description 2
- 208000020431 spinal cord injury Diseases 0.000 claims description 2
- 239000007921 spray Substances 0.000 claims description 2
- 208000011580 syndromic disease Diseases 0.000 claims description 2
- 210000002435 tendon Anatomy 0.000 claims description 2
- 230000009529 traumatic brain injury Effects 0.000 claims description 2
- 230000000472 traumatic effect Effects 0.000 claims description 2
- 230000008736 traumatic injury Effects 0.000 claims description 2
- 231100000397 ulcer Toxicity 0.000 claims description 2
- 201000006669 vulvar dystrophy Diseases 0.000 claims description 2
- 239000004615 ingredient Substances 0.000 claims 2
- 210000004102 animal cell Anatomy 0.000 claims 1
- 210000004027 cell Anatomy 0.000 description 51
- 238000012360 testing method Methods 0.000 description 45
- 238000000338 in vitro Methods 0.000 description 25
- 230000029663 wound healing Effects 0.000 description 22
- 239000000306 component Substances 0.000 description 21
- 230000008569 process Effects 0.000 description 19
- 239000000047 product Substances 0.000 description 19
- 238000000134 MTT assay Methods 0.000 description 16
- 231100000002 MTT assay Toxicity 0.000 description 16
- 230000000694 effects Effects 0.000 description 13
- 108090000623 proteins and genes Proteins 0.000 description 13
- 238000003556 assay Methods 0.000 description 12
- 210000004369 blood Anatomy 0.000 description 12
- 239000008280 blood Substances 0.000 description 12
- 235000018102 proteins Nutrition 0.000 description 12
- 102000004169 proteins and genes Human genes 0.000 description 12
- 210000002919 epithelial cell Anatomy 0.000 description 11
- 210000003780 hair follicle Anatomy 0.000 description 11
- 210000004623 platelet-rich plasma Anatomy 0.000 description 11
- 238000002360 preparation method Methods 0.000 description 11
- 230000002062 proliferating effect Effects 0.000 description 11
- 230000035755 proliferation Effects 0.000 description 11
- 238000004113 cell culture Methods 0.000 description 10
- 210000000265 leukocyte Anatomy 0.000 description 9
- 230000006870 function Effects 0.000 description 8
- 241000124008 Mammalia Species 0.000 description 7
- 230000009471 action Effects 0.000 description 7
- 230000033115 angiogenesis Effects 0.000 description 7
- 230000004663 cell proliferation Effects 0.000 description 7
- 210000002889 endothelial cell Anatomy 0.000 description 7
- 101150026046 iga gene Proteins 0.000 description 7
- 238000004519 manufacturing process Methods 0.000 description 7
- 230000008929 regeneration Effects 0.000 description 7
- 238000011160 research Methods 0.000 description 7
- 230000004044 response Effects 0.000 description 7
- 210000003491 skin Anatomy 0.000 description 7
- 238000005199 ultracentrifugation Methods 0.000 description 7
- 102000009123 Fibrin Human genes 0.000 description 6
- 108010073385 Fibrin Proteins 0.000 description 6
- BWGVNKXGVNDBDI-UHFFFAOYSA-N Fibrin monomer Chemical compound CNC(=O)CNC(=O)CN BWGVNKXGVNDBDI-UHFFFAOYSA-N 0.000 description 6
- 102000018233 Fibroblast Growth Factor Human genes 0.000 description 6
- 108050007372 Fibroblast Growth Factor Proteins 0.000 description 6
- 230000015572 biosynthetic process Effects 0.000 description 6
- 229950003499 fibrin Drugs 0.000 description 6
- 210000002950 fibroblast Anatomy 0.000 description 6
- 239000010410 layer Substances 0.000 description 6
- 210000002381 plasma Anatomy 0.000 description 6
- 238000000746 purification Methods 0.000 description 6
- 210000002966 serum Anatomy 0.000 description 6
- 239000000243 solution Substances 0.000 description 6
- 210000000988 bone and bone Anatomy 0.000 description 5
- 230000003833 cell viability Effects 0.000 description 5
- 230000001413 cellular effect Effects 0.000 description 5
- 238000005119 centrifugation Methods 0.000 description 5
- 238000011161 development Methods 0.000 description 5
- 230000018109 developmental process Effects 0.000 description 5
- 229940088598 enzyme Drugs 0.000 description 5
- 210000003743 erythrocyte Anatomy 0.000 description 5
- 210000002744 extracellular matrix Anatomy 0.000 description 5
- GVJHHUAWPYXKBD-UHFFFAOYSA-N (±)-α-Tocopherol Chemical compound OC1=C(C)C(C)=C2OC(CCCC(C)CCCC(C)CCCC(C)C)(C)CCC2=C1C GVJHHUAWPYXKBD-UHFFFAOYSA-N 0.000 description 4
- 241000894006 Bacteria Species 0.000 description 4
- 102000008186 Collagen Human genes 0.000 description 4
- 108010035532 Collagen Proteins 0.000 description 4
- 206010061218 Inflammation Diseases 0.000 description 4
- 108010038512 Platelet-Derived Growth Factor Proteins 0.000 description 4
- 102000010780 Platelet-Derived Growth Factor Human genes 0.000 description 4
- 108060008682 Tumor Necrosis Factor Proteins 0.000 description 4
- 230000003698 anagen phase Effects 0.000 description 4
- 239000003146 anticoagulant agent Substances 0.000 description 4
- 229940127219 anticoagulant drug Drugs 0.000 description 4
- 235000013734 beta-carotene Nutrition 0.000 description 4
- 239000011648 beta-carotene Substances 0.000 description 4
- 239000013060 biological fluid Substances 0.000 description 4
- 230000003778 catagen phase Effects 0.000 description 4
- 210000001612 chondrocyte Anatomy 0.000 description 4
- 229920001436 collagen Polymers 0.000 description 4
- 230000004069 differentiation Effects 0.000 description 4
- 238000004108 freeze drying Methods 0.000 description 4
- 230000012010 growth Effects 0.000 description 4
- 210000003494 hepatocyte Anatomy 0.000 description 4
- 230000004054 inflammatory process Effects 0.000 description 4
- 210000002510 keratinocyte Anatomy 0.000 description 4
- 239000011159 matrix material Substances 0.000 description 4
- 210000002752 melanocyte Anatomy 0.000 description 4
- 230000005012 migration Effects 0.000 description 4
- 238000013508 migration Methods 0.000 description 4
- 239000012071 phase Substances 0.000 description 4
- 210000002826 placenta Anatomy 0.000 description 4
- BOLDJAUMGUJJKM-LSDHHAIUSA-N renifolin D Natural products CC(=C)[C@@H]1Cc2c(O)c(O)ccc2[C@H]1CC(=O)c3ccc(O)cc3O BOLDJAUMGUJJKM-LSDHHAIUSA-N 0.000 description 4
- 238000000926 separation method Methods 0.000 description 4
- 239000002356 single layer Substances 0.000 description 4
- 241000894007 species Species 0.000 description 4
- 239000000126 substance Substances 0.000 description 4
- 229930003231 vitamin Natural products 0.000 description 4
- 235000013343 vitamin Nutrition 0.000 description 4
- 239000011782 vitamin Substances 0.000 description 4
- 229940088594 vitamin Drugs 0.000 description 4
- OENHQHLEOONYIE-JLTXGRSLSA-N β-Carotene Chemical compound CC=1CCCC(C)(C)C=1\C=C\C(\C)=C\C=C\C(\C)=C\C=C\C=C(/C)\C=C\C=C(/C)\C=C\C1=C(C)CCCC1(C)C OENHQHLEOONYIE-JLTXGRSLSA-N 0.000 description 4
- 102000003886 Glycoproteins Human genes 0.000 description 3
- 108090000288 Glycoproteins Proteins 0.000 description 3
- 108010017213 Granulocyte-Macrophage Colony-Stimulating Factor Proteins 0.000 description 3
- 102100039620 Granulocyte-macrophage colony-stimulating factor Human genes 0.000 description 3
- 101000599951 Homo sapiens Insulin-like growth factor I Proteins 0.000 description 3
- 101001076292 Homo sapiens Insulin-like growth factor II Proteins 0.000 description 3
- 102100037852 Insulin-like growth factor I Human genes 0.000 description 3
- 102100025947 Insulin-like growth factor II Human genes 0.000 description 3
- 108010063045 Lactoferrin Proteins 0.000 description 3
- 102000010445 Lactoferrin Human genes 0.000 description 3
- 201000004681 Psoriasis Diseases 0.000 description 3
- 102100040247 Tumor necrosis factor Human genes 0.000 description 3
- 241000700605 Viruses Species 0.000 description 3
- 238000001042 affinity chromatography Methods 0.000 description 3
- 230000006907 apoptotic process Effects 0.000 description 3
- 210000003719 b-lymphocyte Anatomy 0.000 description 3
- 230000008827 biological function Effects 0.000 description 3
- 210000000845 cartilage Anatomy 0.000 description 3
- 230000024245 cell differentiation Effects 0.000 description 3
- 230000003915 cell function Effects 0.000 description 3
- 210000002808 connective tissue Anatomy 0.000 description 3
- 239000013078 crystal Substances 0.000 description 3
- 230000007123 defense Effects 0.000 description 3
- 210000001163 endosome Anatomy 0.000 description 3
- 239000002158 endotoxin Substances 0.000 description 3
- 229940126864 fibroblast growth factor Drugs 0.000 description 3
- 238000001914 filtration Methods 0.000 description 3
- 239000012467 final product Substances 0.000 description 3
- 235000020256 human milk Nutrition 0.000 description 3
- 210000004251 human milk Anatomy 0.000 description 3
- 210000002865 immune cell Anatomy 0.000 description 3
- 238000001727 in vivo Methods 0.000 description 3
- 230000002757 inflammatory effect Effects 0.000 description 3
- 239000003112 inhibitor Substances 0.000 description 3
- 230000003993 interaction Effects 0.000 description 3
- 230000003834 intracellular effect Effects 0.000 description 3
- CSSYQJWUGATIHM-IKGCZBKSSA-N l-phenylalanyl-l-lysyl-l-cysteinyl-l-arginyl-l-arginyl-l-tryptophyl-l-glutaminyl-l-tryptophyl-l-arginyl-l-methionyl-l-lysyl-l-lysyl-l-leucylglycyl-l-alanyl-l-prolyl-l-seryl-l-isoleucyl-l-threonyl-l-cysteinyl-l-valyl-l-arginyl-l-arginyl-l-alanyl-l-phenylal Chemical compound C([C@H](N)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CS)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCSC)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(=O)NCC(=O)N[C@@H](C)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CO)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CS)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](C)C(=O)N[C@@H](CC=1C=CC=CC=1)C(O)=O)C1=CC=CC=C1 CSSYQJWUGATIHM-IKGCZBKSSA-N 0.000 description 3
- 235000021242 lactoferrin Nutrition 0.000 description 3
- 229940078795 lactoferrin Drugs 0.000 description 3
- 239000007788 liquid Substances 0.000 description 3
- 210000002540 macrophage Anatomy 0.000 description 3
- 210000002901 mesenchymal stem cell Anatomy 0.000 description 3
- 210000002487 multivesicular body Anatomy 0.000 description 3
- VMGAPWLDMVPYIA-HIDZBRGKSA-N n'-amino-n-iminomethanimidamide Chemical compound N\N=C\N=N VMGAPWLDMVPYIA-HIDZBRGKSA-N 0.000 description 3
- 210000000963 osteoblast Anatomy 0.000 description 3
- 108090000765 processed proteins & peptides Proteins 0.000 description 3
- 102000005962 receptors Human genes 0.000 description 3
- 108020003175 receptors Proteins 0.000 description 3
- 230000001105 regulatory effect Effects 0.000 description 3
- 210000003296 saliva Anatomy 0.000 description 3
- 210000004116 schwann cell Anatomy 0.000 description 3
- 210000000329 smooth muscle myocyte Anatomy 0.000 description 3
- 238000003786 synthesis reaction Methods 0.000 description 3
- 230000014616 translation Effects 0.000 description 3
- 238000000108 ultra-filtration Methods 0.000 description 3
- FPIPGXGPPPQFEQ-UHFFFAOYSA-N 13-cis retinol Natural products OCC=C(C)C=CC=C(C)C=CC1=C(C)CCCC1(C)C FPIPGXGPPPQFEQ-UHFFFAOYSA-N 0.000 description 2
- 102100029459 Apelin Human genes 0.000 description 2
- 108010078791 Carrier Proteins Proteins 0.000 description 2
- 102000011632 Caseins Human genes 0.000 description 2
- 108010076119 Caseins Proteins 0.000 description 2
- 108010077840 Complement C3a Proteins 0.000 description 2
- 108010077773 Complement C4a Proteins 0.000 description 2
- 108010086672 Endosomal Sorting Complexes Required for Transport Proteins 0.000 description 2
- 102000006770 Endosomal Sorting Complexes Required for Transport Human genes 0.000 description 2
- 241000283086 Equidae Species 0.000 description 2
- 108090000385 Fibroblast growth factor 7 Proteins 0.000 description 2
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 2
- 102000004269 Granulocyte Colony-Stimulating Factor Human genes 0.000 description 2
- 108010017080 Granulocyte Colony-Stimulating Factor Proteins 0.000 description 2
- 108010051696 Growth Hormone Proteins 0.000 description 2
- 101000771523 Homo sapiens Apelin Proteins 0.000 description 2
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 description 2
- 206010020751 Hypersensitivity Diseases 0.000 description 2
- 108090000723 Insulin-Like Growth Factor I Proteins 0.000 description 2
- 102000003812 Interleukin-15 Human genes 0.000 description 2
- 108090000172 Interleukin-15 Proteins 0.000 description 2
- ROHFNLRQFUQHCH-YFKPBYRVSA-N L-leucine Chemical compound CC(C)C[C@H](N)C(O)=O ROHFNLRQFUQHCH-YFKPBYRVSA-N 0.000 description 2
- KDXKERNSBIXSRK-YFKPBYRVSA-N L-lysine Chemical compound NCCCC[C@H](N)C(O)=O KDXKERNSBIXSRK-YFKPBYRVSA-N 0.000 description 2
- 108010023244 Lactoperoxidase Proteins 0.000 description 2
- 102000045576 Lactoperoxidases Human genes 0.000 description 2
- XUMBMVFBXHLACL-UHFFFAOYSA-N Melanin Chemical compound O=C1C(=O)C(C2=CNC3=C(C(C(=O)C4=C32)=O)C)=C2C4=CNC2=C1C XUMBMVFBXHLACL-UHFFFAOYSA-N 0.000 description 2
- 101150114886 NECTIN1 gene Proteins 0.000 description 2
- 102100023064 Nectin-1 Human genes 0.000 description 2
- 239000004677 Nylon Substances 0.000 description 2
- 102000013275 Somatomedins Human genes 0.000 description 2
- 102100038803 Somatotropin Human genes 0.000 description 2
- 108010009583 Transforming Growth Factors Proteins 0.000 description 2
- 102000009618 Transforming Growth Factors Human genes 0.000 description 2
- 102000005789 Vascular Endothelial Growth Factors Human genes 0.000 description 2
- 108010019530 Vascular Endothelial Growth Factors Proteins 0.000 description 2
- FPIPGXGPPPQFEQ-BOOMUCAASA-N Vitamin A Natural products OC/C=C(/C)\C=C\C=C(\C)/C=C/C1=C(C)CCCC1(C)C FPIPGXGPPPQFEQ-BOOMUCAASA-N 0.000 description 2
- 229930003779 Vitamin B12 Natural products 0.000 description 2
- 229930003427 Vitamin E Natural products 0.000 description 2
- 210000001552 airway epithelial cell Anatomy 0.000 description 2
- OENHQHLEOONYIE-UKMVMLAPSA-N all-trans beta-carotene Natural products CC=1CCCC(C)(C)C=1/C=C/C(/C)=C/C=C/C(/C)=C/C=C/C=C(C)C=CC=C(C)C=CC1=C(C)CCCC1(C)C OENHQHLEOONYIE-UKMVMLAPSA-N 0.000 description 2
- FPIPGXGPPPQFEQ-OVSJKPMPSA-N all-trans-retinol Chemical compound OC\C=C(/C)\C=C\C=C(/C)\C=C\C1=C(C)CCCC1(C)C FPIPGXGPPPQFEQ-OVSJKPMPSA-N 0.000 description 2
- 229940024606 amino acid Drugs 0.000 description 2
- 235000001014 amino acid Nutrition 0.000 description 2
- 208000006673 asthma Diseases 0.000 description 2
- 230000003305 autocrine Effects 0.000 description 2
- 230000006399 behavior Effects 0.000 description 2
- 230000008901 benefit Effects 0.000 description 2
- TUPZEYHYWIEDIH-WAIFQNFQSA-N beta-carotene Natural products CC(=C/C=C/C=C(C)/C=C/C=C(C)/C=C/C1=C(C)CCCC1(C)C)C=CC=C(/C)C=CC2=CCCCC2(C)C TUPZEYHYWIEDIH-WAIFQNFQSA-N 0.000 description 2
- 229960002747 betacarotene Drugs 0.000 description 2
- 230000008436 biogenesis Effects 0.000 description 2
- 230000033228 biological regulation Effects 0.000 description 2
- 239000012503 blood component Substances 0.000 description 2
- 210000004204 blood vessel Anatomy 0.000 description 2
- 210000001185 bone marrow Anatomy 0.000 description 2
- BECPQYXYKAMYBN-UHFFFAOYSA-N casein, tech. Chemical compound NCCCCC(C(O)=O)N=C(O)C(CC(O)=O)N=C(O)C(CCC(O)=N)N=C(O)C(CC(C)C)N=C(O)C(CCC(O)=O)N=C(O)C(CC(O)=O)N=C(O)C(CCC(O)=O)N=C(O)C(C(C)O)N=C(O)C(CCC(O)=N)N=C(O)C(CCC(O)=N)N=C(O)C(CCC(O)=N)N=C(O)C(CCC(O)=O)N=C(O)C(CCC(O)=O)N=C(O)C(COP(O)(O)=O)N=C(O)C(CCC(O)=N)N=C(O)C(N)CC1=CC=CC=C1 BECPQYXYKAMYBN-UHFFFAOYSA-N 0.000 description 2
- 235000021240 caseins Nutrition 0.000 description 2
- 230000015556 catabolic process Effects 0.000 description 2
- 210000000170 cell membrane Anatomy 0.000 description 2
- 230000012292 cell migration Effects 0.000 description 2
- 239000013553 cell monolayer Substances 0.000 description 2
- 239000003795 chemical substances by application Substances 0.000 description 2
- AGVAZMGAQJOSFJ-WZHZPDAFSA-M cobalt(2+);[(2r,3s,4r,5s)-5-(5,6-dimethylbenzimidazol-1-yl)-4-hydroxy-2-(hydroxymethyl)oxolan-3-yl] [(2r)-1-[3-[(1r,2r,3r,4z,7s,9z,12s,13s,14z,17s,18s,19r)-2,13,18-tris(2-amino-2-oxoethyl)-7,12,17-tris(3-amino-3-oxopropyl)-3,5,8,8,13,15,18,19-octamethyl-2 Chemical compound [Co+2].N#[C-].[N-]([C@@H]1[C@H](CC(N)=O)[C@@]2(C)CCC(=O)NC[C@@H](C)OP(O)(=O)O[C@H]3[C@H]([C@H](O[C@@H]3CO)N3C4=CC(C)=C(C)C=C4N=C3)O)\C2=C(C)/C([C@H](C\2(C)C)CCC(N)=O)=N/C/2=C\C([C@H]([C@@]/2(CC(N)=O)C)CCC(N)=O)=N\C\2=C(C)/C2=N[C@]1(C)[C@@](C)(CC(N)=O)[C@@H]2CCC(N)=O AGVAZMGAQJOSFJ-WZHZPDAFSA-M 0.000 description 2
- 238000004737 colorimetric analysis Methods 0.000 description 2
- 230000000295 complement effect Effects 0.000 description 2
- 239000000470 constituent Substances 0.000 description 2
- 239000002537 cosmetic Substances 0.000 description 2
- 238000009295 crossflow filtration Methods 0.000 description 2
- 238000006731 degradation reaction Methods 0.000 description 2
- 206010012601 diabetes mellitus Diseases 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 238000000502 dialysis Methods 0.000 description 2
- 238000009792 diffusion process Methods 0.000 description 2
- 208000035475 disorder Diseases 0.000 description 2
- 230000002500 effect on skin Effects 0.000 description 2
- 239000012636 effector Substances 0.000 description 2
- 238000011013 endotoxin removal Methods 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 210000002615 epidermis Anatomy 0.000 description 2
- 210000000981 epithelium Anatomy 0.000 description 2
- 210000001723 extracellular space Anatomy 0.000 description 2
- 239000000945 filler Substances 0.000 description 2
- 239000012530 fluid Substances 0.000 description 2
- 238000007710 freezing Methods 0.000 description 2
- 230000008014 freezing Effects 0.000 description 2
- WIGCFUFOHFEKBI-UHFFFAOYSA-N gamma-tocopherol Natural products CC(C)CCCC(C)CCCC(C)CCCC1CCC2C(C)C(O)C(C)C(C)C2O1 WIGCFUFOHFEKBI-UHFFFAOYSA-N 0.000 description 2
- 239000000122 growth hormone Substances 0.000 description 2
- 239000001963 growth medium Substances 0.000 description 2
- 230000009931 harmful effect Effects 0.000 description 2
- 230000028993 immune response Effects 0.000 description 2
- 210000000987 immune system Anatomy 0.000 description 2
- 230000001939 inductive effect Effects 0.000 description 2
- 230000028709 inflammatory response Effects 0.000 description 2
- 230000002401 inhibitory effect Effects 0.000 description 2
- 230000005764 inhibitory process Effects 0.000 description 2
- 230000000977 initiatory effect Effects 0.000 description 2
- 229940068935 insulin-like growth factor 2 Drugs 0.000 description 2
- 230000002608 insulinlike Effects 0.000 description 2
- 229940057428 lactoperoxidase Drugs 0.000 description 2
- 238000011068 loading method Methods 0.000 description 2
- 210000004698 lymphocyte Anatomy 0.000 description 2
- 230000035800 maturation Effects 0.000 description 2
- 238000005259 measurement Methods 0.000 description 2
- 230000004060 metabolic process Effects 0.000 description 2
- 238000001471 micro-filtration Methods 0.000 description 2
- 210000001616 monocyte Anatomy 0.000 description 2
- 210000003205 muscle Anatomy 0.000 description 2
- 229920001778 nylon Polymers 0.000 description 2
- 230000014207 opsonization Effects 0.000 description 2
- 230000000278 osteoconductive effect Effects 0.000 description 2
- 230000001575 pathological effect Effects 0.000 description 2
- 239000000546 pharmaceutical excipient Substances 0.000 description 2
- 230000035479 physiological effects, processes and functions Effects 0.000 description 2
- 239000011148 porous material Substances 0.000 description 2
- 230000003389 potentiating effect Effects 0.000 description 2
- 238000001556 precipitation Methods 0.000 description 2
- 230000001737 promoting effect Effects 0.000 description 2
- 238000001243 protein synthesis Methods 0.000 description 2
- 230000005855 radiation Effects 0.000 description 2
- 230000009467 reduction Effects 0.000 description 2
- 239000013074 reference sample Substances 0.000 description 2
- 230000009719 regenerative response Effects 0.000 description 2
- 230000025600 response to UV Effects 0.000 description 2
- 210000002363 skeletal muscle cell Anatomy 0.000 description 2
- 210000004927 skin cell Anatomy 0.000 description 2
- 208000017520 skin disease Diseases 0.000 description 2
- 150000003384 small molecules Chemical class 0.000 description 2
- 210000004872 soft tissue Anatomy 0.000 description 2
- 230000001954 sterilising effect Effects 0.000 description 2
- 238000002560 therapeutic procedure Methods 0.000 description 2
- 231100000419 toxicity Toxicity 0.000 description 2
- 230000001988 toxicity Effects 0.000 description 2
- 230000001228 trophic effect Effects 0.000 description 2
- VBEQCZHXXJYVRD-GACYYNSASA-N uroanthelone Chemical compound C([C@@H](C(=O)N[C@H](C(=O)N[C@@H](CS)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CS)C(=O)N[C@H](C(=O)N[C@@H]([C@@H](C)CC)C(=O)NCC(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(=O)N[C@@H](CO)C(=O)NCC(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CS)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O)C(C)C)[C@@H](C)O)NC(=O)[C@H](CO)NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CO)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@@H](NC(=O)[C@H](CC=1NC=NC=1)NC(=O)[C@H](CCSC)NC(=O)[C@H](CS)NC(=O)[C@@H](NC(=O)CNC(=O)CNC(=O)[C@H](CC(N)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CS)NC(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)CNC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)[C@H](CO)NC(=O)[C@H](CO)NC(=O)[C@H]1N(CCC1)C(=O)[C@H](CS)NC(=O)CNC(=O)[C@H]1N(CCC1)C(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)[C@H](CO)NC(=O)[C@@H](N)CC(N)=O)C(C)C)[C@@H](C)CC)C1=CC=C(O)C=C1 VBEQCZHXXJYVRD-GACYYNSASA-N 0.000 description 2
- 230000002792 vascular Effects 0.000 description 2
- 235000019155 vitamin A Nutrition 0.000 description 2
- 239000011719 vitamin A Substances 0.000 description 2
- 235000019163 vitamin B12 Nutrition 0.000 description 2
- 239000011715 vitamin B12 Substances 0.000 description 2
- 235000019165 vitamin E Nutrition 0.000 description 2
- 229940046009 vitamin E Drugs 0.000 description 2
- 239000011709 vitamin E Substances 0.000 description 2
- 229940045997 vitamin a Drugs 0.000 description 2
- 230000037314 wound repair Effects 0.000 description 2
- AZKSAVLVSZKNRD-UHFFFAOYSA-M 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide Chemical compound [Br-].S1C(C)=C(C)N=C1[N+]1=NC(C=2C=CC=CC=2)=NN1C1=CC=CC=C1 AZKSAVLVSZKNRD-UHFFFAOYSA-M 0.000 description 1
- 229930091051 Arenine Natural products 0.000 description 1
- 101001123995 Bacillus subtilis (strain 168) FMN reductase [NAD(P)H] Proteins 0.000 description 1
- 102100021943 C-C motif chemokine 2 Human genes 0.000 description 1
- 101710155857 C-C motif chemokine 2 Proteins 0.000 description 1
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 description 1
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 description 1
- 241000282832 Camelidae Species 0.000 description 1
- 102000019034 Chemokines Human genes 0.000 description 1
- 108010012236 Chemokines Proteins 0.000 description 1
- 208000006545 Chronic Obstructive Pulmonary Disease Diseases 0.000 description 1
- 102000007644 Colony-Stimulating Factors Human genes 0.000 description 1
- 108010071942 Colony-Stimulating Factors Proteins 0.000 description 1
- 208000011231 Crohn disease Diseases 0.000 description 1
- 201000003883 Cystic fibrosis Diseases 0.000 description 1
- 201000004624 Dermatitis Diseases 0.000 description 1
- 238000002965 ELISA Methods 0.000 description 1
- 102400001368 Epidermal growth factor Human genes 0.000 description 1
- 101800003838 Epidermal growth factor Proteins 0.000 description 1
- 241000283073 Equus caballus Species 0.000 description 1
- 102000003951 Erythropoietin Human genes 0.000 description 1
- 108090000394 Erythropoietin Proteins 0.000 description 1
- 206010063560 Excessive granulation tissue Diseases 0.000 description 1
- 102000003972 Fibroblast growth factor 7 Human genes 0.000 description 1
- 108010067306 Fibronectins Proteins 0.000 description 1
- 102000016359 Fibronectins Human genes 0.000 description 1
- 241000192125 Firmicutes Species 0.000 description 1
- 239000004471 Glycine Substances 0.000 description 1
- 229920002683 Glycosaminoglycan Polymers 0.000 description 1
- 102000005744 Glycoside Hydrolases Human genes 0.000 description 1
- 108010031186 Glycoside Hydrolases Proteins 0.000 description 1
- 241000590002 Helicobacter pylori Species 0.000 description 1
- 206010020772 Hypertension Diseases 0.000 description 1
- 102000008394 Immunoglobulin Fragments Human genes 0.000 description 1
- 102000003746 Insulin Receptor Human genes 0.000 description 1
- 108010001127 Insulin Receptor Proteins 0.000 description 1
- 108010002352 Interleukin-1 Proteins 0.000 description 1
- 102000000589 Interleukin-1 Human genes 0.000 description 1
- 108010002350 Interleukin-2 Proteins 0.000 description 1
- 108010002586 Interleukin-7 Proteins 0.000 description 1
- 102000000704 Interleukin-7 Human genes 0.000 description 1
- 108010002335 Interleukin-9 Proteins 0.000 description 1
- 206010022998 Irritability Diseases 0.000 description 1
- 206010023330 Keloid scar Diseases 0.000 description 1
- 235000019766 L-Lysine Nutrition 0.000 description 1
- ONIBWKKTOPOVIA-BYPYZUCNSA-N L-Proline Chemical compound OC(=O)[C@@H]1CCCN1 ONIBWKKTOPOVIA-BYPYZUCNSA-N 0.000 description 1
- 235000019454 L-leucine Nutrition 0.000 description 1
- 239000004395 L-leucine Substances 0.000 description 1
- 229930182821 L-proline Natural products 0.000 description 1
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 description 1
- 239000004472 Lysine Substances 0.000 description 1
- 102000005741 Metalloproteases Human genes 0.000 description 1
- 108010006035 Metalloproteases Proteins 0.000 description 1
- 102000029749 Microtubule Human genes 0.000 description 1
- 108091022875 Microtubule Proteins 0.000 description 1
- 102000016943 Muramidase Human genes 0.000 description 1
- 108010014251 Muramidase Proteins 0.000 description 1
- 241000699666 Mus <mouse, genus> Species 0.000 description 1
- 241000699670 Mus sp. Species 0.000 description 1
- 108010062010 N-Acetylmuramoyl-L-alanine Amidase Proteins 0.000 description 1
- OVRNDRQMDRJTHS-UHFFFAOYSA-N N-acelyl-D-glucosamine Natural products CC(=O)NC1C(O)OC(CO)C(O)C1O OVRNDRQMDRJTHS-UHFFFAOYSA-N 0.000 description 1
- MNLRQHMNZILYPY-MDMHTWEWSA-N N-acetyl-alpha-D-muramic acid Chemical compound OC(=O)[C@@H](C)O[C@H]1[C@H](O)[C@@H](CO)O[C@H](O)[C@@H]1NC(C)=O MNLRQHMNZILYPY-MDMHTWEWSA-N 0.000 description 1
- OVRNDRQMDRJTHS-FMDGEEDCSA-N N-acetyl-beta-D-glucosamine Chemical compound CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O OVRNDRQMDRJTHS-FMDGEEDCSA-N 0.000 description 1
- MBLBDJOUHNCFQT-LXGUWJNJSA-N N-acetylglucosamine Natural products CC(=O)N[C@@H](C=O)[C@@H](O)[C@H](O)[C@H](O)CO MBLBDJOUHNCFQT-LXGUWJNJSA-N 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 206010029113 Neovascularisation Diseases 0.000 description 1
- 102000014413 Neuregulin Human genes 0.000 description 1
- 108050003475 Neuregulin Proteins 0.000 description 1
- 241000283973 Oryctolagus cuniculus Species 0.000 description 1
- 241001494479 Pecora Species 0.000 description 1
- 208000012641 Pigmentation disease Diseases 0.000 description 1
- 108010009736 Protein Hydrolysates Proteins 0.000 description 1
- 241000588769 Proteus <enterobacteria> Species 0.000 description 1
- 241000700159 Rattus Species 0.000 description 1
- 206010057190 Respiratory tract infections Diseases 0.000 description 1
- 206010039710 Scleroderma Diseases 0.000 description 1
- 108010003723 Single-Domain Antibodies Proteins 0.000 description 1
- 208000000453 Skin Neoplasms Diseases 0.000 description 1
- 206010040829 Skin discolouration Diseases 0.000 description 1
- 229920002385 Sodium hyaluronate Polymers 0.000 description 1
- 208000007107 Stomach Ulcer Diseases 0.000 description 1
- 210000001744 T-lymphocyte Anatomy 0.000 description 1
- 108700031126 Tetraspanins Proteins 0.000 description 1
- 102000043977 Tetraspanins Human genes 0.000 description 1
- 108090000190 Thrombin Proteins 0.000 description 1
- 102000004338 Transferrin Human genes 0.000 description 1
- 108090000901 Transferrin Proteins 0.000 description 1
- 208000036142 Viral infection Diseases 0.000 description 1
- 239000005862 Whey Substances 0.000 description 1
- 102000007544 Whey Proteins Human genes 0.000 description 1
- 108010046377 Whey Proteins Proteins 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 230000003213 activating effect Effects 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 239000004480 active ingredient Substances 0.000 description 1
- 230000006978 adaptation Effects 0.000 description 1
- 239000000654 additive Substances 0.000 description 1
- 230000032683 aging Effects 0.000 description 1
- 210000004381 amniotic fluid Anatomy 0.000 description 1
- 230000001195 anabolic effect Effects 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 230000019552 anatomical structure morphogenesis Effects 0.000 description 1
- 238000010171 animal model Methods 0.000 description 1
- 230000001093 anti-cancer Effects 0.000 description 1
- 230000003110 anti-inflammatory effect Effects 0.000 description 1
- 239000000427 antigen Substances 0.000 description 1
- 230000007503 antigenic stimulation Effects 0.000 description 1
- 108091007433 antigens Proteins 0.000 description 1
- 102000036639 antigens Human genes 0.000 description 1
- 239000004599 antimicrobial Substances 0.000 description 1
- 210000000709 aorta Anatomy 0.000 description 1
- 210000002403 aortic endothelial cell Anatomy 0.000 description 1
- 230000001640 apoptogenic effect Effects 0.000 description 1
- 208000010668 atopic eczema Diseases 0.000 description 1
- 230000001363 autoimmune Effects 0.000 description 1
- 244000052616 bacterial pathogen Species 0.000 description 1
- 230000001420 bacteriolytic effect Effects 0.000 description 1
- 230000004888 barrier function Effects 0.000 description 1
- 210000003912 basophilic leucocyte Anatomy 0.000 description 1
- 239000011324 bead Substances 0.000 description 1
- ZYXYTGQFPZEUFX-UHFFFAOYSA-N benzpyrimoxan Chemical compound O1C(OCCC1)C=1C(=NC=NC=1)OCC1=CC=C(C=C1)C(F)(F)F ZYXYTGQFPZEUFX-UHFFFAOYSA-N 0.000 description 1
- 210000000941 bile Anatomy 0.000 description 1
- 239000003462 bioceramic Substances 0.000 description 1
- 230000008499 blood brain barrier function Effects 0.000 description 1
- 210000000601 blood cell Anatomy 0.000 description 1
- 210000001218 blood-brain barrier Anatomy 0.000 description 1
- 230000004097 bone metabolism Effects 0.000 description 1
- 230000010478 bone regeneration Effects 0.000 description 1
- 210000000621 bronchi Anatomy 0.000 description 1
- 239000011575 calcium Substances 0.000 description 1
- 229910052791 calcium Inorganic materials 0.000 description 1
- 239000001110 calcium chloride Substances 0.000 description 1
- 229910001628 calcium chloride Inorganic materials 0.000 description 1
- 201000011510 cancer Diseases 0.000 description 1
- 230000019259 carbohydrate homeostasis Effects 0.000 description 1
- 239000005018 casein Substances 0.000 description 1
- 229940021722 caseins Drugs 0.000 description 1
- 230000030833 cell death Effects 0.000 description 1
- 230000032823 cell division Effects 0.000 description 1
- 230000010261 cell growth Effects 0.000 description 1
- 210000002421 cell wall Anatomy 0.000 description 1
- 230000033077 cellular process Effects 0.000 description 1
- 230000036755 cellular response Effects 0.000 description 1
- 239000004568 cement Substances 0.000 description 1
- 210000001175 cerebrospinal fluid Anatomy 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 230000035605 chemotaxis Effects 0.000 description 1
- 230000000973 chemotherapeutic effect Effects 0.000 description 1
- 238000004587 chromatography analysis Methods 0.000 description 1
- 230000035602 clotting Effects 0.000 description 1
- 230000000052 comparative effect Effects 0.000 description 1
- 230000004154 complement system Effects 0.000 description 1
- 210000004087 cornea Anatomy 0.000 description 1
- 210000004351 coronary vessel Anatomy 0.000 description 1
- 230000002596 correlated effect Effects 0.000 description 1
- 230000037029 cross reaction Effects 0.000 description 1
- 208000035250 cutaneous malignant susceptibility to 1 melanoma Diseases 0.000 description 1
- 230000009089 cytolysis Effects 0.000 description 1
- 210000004292 cytoskeleton Anatomy 0.000 description 1
- 238000007405 data analysis Methods 0.000 description 1
- 239000008367 deionised water Substances 0.000 description 1
- 229910021641 deionized water Inorganic materials 0.000 description 1
- 238000004925 denaturation Methods 0.000 description 1
- 230000036425 denaturation Effects 0.000 description 1
- 210000003298 dental enamel Anatomy 0.000 description 1
- 239000004053 dental implant Substances 0.000 description 1
- 230000008021 deposition Effects 0.000 description 1
- CGMRCMMOCQYHAD-UHFFFAOYSA-J dicalcium hydroxide phosphate Chemical compound [OH-].[Ca++].[Ca++].[O-]P([O-])([O-])=O CGMRCMMOCQYHAD-UHFFFAOYSA-J 0.000 description 1
- 102000038379 digestive enzymes Human genes 0.000 description 1
- 108091007734 digestive enzymes Proteins 0.000 description 1
- 230000023011 digestive tract development Effects 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 239000003640 drug residue Substances 0.000 description 1
- 238000007877 drug screening Methods 0.000 description 1
- 238000001962 electrophoresis Methods 0.000 description 1
- 210000002308 embryonic cell Anatomy 0.000 description 1
- 230000002124 endocrine Effects 0.000 description 1
- 230000003511 endothelial effect Effects 0.000 description 1
- 229940116977 epidermal growth factor Drugs 0.000 description 1
- 210000005175 epidermal keratinocyte Anatomy 0.000 description 1
- 230000010393 epithelial cell migration Effects 0.000 description 1
- 229940105423 erythropoietin Drugs 0.000 description 1
- 150000002148 esters Chemical class 0.000 description 1
- 230000007717 exclusion Effects 0.000 description 1
- 230000028023 exocytosis Effects 0.000 description 1
- 230000003721 exogen phase Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 210000003722 extracellular fluid Anatomy 0.000 description 1
- 230000019305 fibroblast migration Effects 0.000 description 1
- 238000011049 filling Methods 0.000 description 1
- 239000012634 fragment Substances 0.000 description 1
- 230000002538 fungal effect Effects 0.000 description 1
- 239000007789 gas Substances 0.000 description 1
- 230000002496 gastric effect Effects 0.000 description 1
- 201000005917 gastric ulcer Diseases 0.000 description 1
- 210000004907 gland Anatomy 0.000 description 1
- 229960002449 glycine Drugs 0.000 description 1
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 1
- 230000002710 gonadal effect Effects 0.000 description 1
- 210000001126 granulation tissue Anatomy 0.000 description 1
- 230000003394 haemopoietic effect Effects 0.000 description 1
- 210000004209 hair Anatomy 0.000 description 1
- 230000009583 hair follicle growth Effects 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 229940037467 helicobacter pylori Drugs 0.000 description 1
- 210000003958 hematopoietic stem cell Anatomy 0.000 description 1
- 231100000304 hepatotoxicity Toxicity 0.000 description 1
- 244000038280 herbivores Species 0.000 description 1
- 210000003630 histaminocyte Anatomy 0.000 description 1
- 229940088597 hormone Drugs 0.000 description 1
- 239000005556 hormone Substances 0.000 description 1
- 230000004727 humoral immunity Effects 0.000 description 1
- 230000000774 hypoallergenic effect Effects 0.000 description 1
- 238000003384 imaging method Methods 0.000 description 1
- 230000002519 immonomodulatory effect Effects 0.000 description 1
- 230000001900 immune effect Effects 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 239000012535 impurity Substances 0.000 description 1
- 239000012678 infectious agent Substances 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 230000008595 infiltration Effects 0.000 description 1
- 238000001764 infiltration Methods 0.000 description 1
- 230000015788 innate immune response Effects 0.000 description 1
- 102000006495 integrins Human genes 0.000 description 1
- 108010044426 integrins Proteins 0.000 description 1
- 230000016507 interphase Effects 0.000 description 1
- 210000002490 intestinal epithelial cell Anatomy 0.000 description 1
- 210000000936 intestine Anatomy 0.000 description 1
- 230000010262 intracellular communication Effects 0.000 description 1
- 230000010438 iron metabolism Effects 0.000 description 1
- 238000002955 isolation Methods 0.000 description 1
- 210000003127 knee Anatomy 0.000 description 1
- 239000008101 lactose Substances 0.000 description 1
- 229960003136 leucine Drugs 0.000 description 1
- 239000003446 ligand Substances 0.000 description 1
- 239000007791 liquid phase Substances 0.000 description 1
- 230000003908 liver function Effects 0.000 description 1
- 230000007056 liver toxicity Effects 0.000 description 1
- 210000002751 lymph Anatomy 0.000 description 1
- 229960003646 lysine Drugs 0.000 description 1
- 229960000274 lysozyme Drugs 0.000 description 1
- 235000010335 lysozyme Nutrition 0.000 description 1
- 239000004325 lysozyme Substances 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 201000001441 melanoma Diseases 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- 230000037353 metabolic pathway Effects 0.000 description 1
- 244000000010 microbial pathogen Species 0.000 description 1
- 239000004005 microsphere Substances 0.000 description 1
- 210000004688 microtubule Anatomy 0.000 description 1
- 235000013336 milk Nutrition 0.000 description 1
- 210000004080 milk Anatomy 0.000 description 1
- 239000008267 milk Substances 0.000 description 1
- 230000003278 mimic effect Effects 0.000 description 1
- 230000011278 mitosis Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000009456 molecular mechanism Effects 0.000 description 1
- 210000003097 mucus Anatomy 0.000 description 1
- 201000006417 multiple sclerosis Diseases 0.000 description 1
- 210000000066 myeloid cell Anatomy 0.000 description 1
- 229950006780 n-acetylglucosamine Drugs 0.000 description 1
- 239000013642 negative control Substances 0.000 description 1
- 210000005036 nerve Anatomy 0.000 description 1
- 230000008271 nervous system development Effects 0.000 description 1
- 210000004498 neuroglial cell Anatomy 0.000 description 1
- 210000002569 neuron Anatomy 0.000 description 1
- 201000001119 neuropathy Diseases 0.000 description 1
- 230000007823 neuropathy Effects 0.000 description 1
- 230000037311 normal skin Effects 0.000 description 1
- 210000001331 nose Anatomy 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 238000011275 oncology therapy Methods 0.000 description 1
- 238000000399 optical microscopy Methods 0.000 description 1
- 238000010883 osseointegration Methods 0.000 description 1
- 230000004072 osteoblast differentiation Effects 0.000 description 1
- 230000003647 oxidation Effects 0.000 description 1
- 238000007254 oxidation reaction Methods 0.000 description 1
- 230000003076 paracrine Effects 0.000 description 1
- 230000036961 partial effect Effects 0.000 description 1
- 244000052769 pathogen Species 0.000 description 1
- 230000007110 pathogen host interaction Effects 0.000 description 1
- 230000001717 pathogenic effect Effects 0.000 description 1
- 210000002379 periodontal ligament Anatomy 0.000 description 1
- 210000001428 peripheral nervous system Anatomy 0.000 description 1
- 102000013415 peroxidase activity proteins Human genes 0.000 description 1
- 108040007629 peroxidase activity proteins Proteins 0.000 description 1
- 239000008363 phosphate buffer Substances 0.000 description 1
- 239000011574 phosphorus Substances 0.000 description 1
- 229910052698 phosphorus Inorganic materials 0.000 description 1
- 239000000049 pigment Substances 0.000 description 1
- 210000004180 plasmocyte Anatomy 0.000 description 1
- 230000010118 platelet activation Effects 0.000 description 1
- 229920000642 polymer Polymers 0.000 description 1
- 238000006116 polymerization reaction Methods 0.000 description 1
- 239000013641 positive control Substances 0.000 description 1
- OXCMYAYHXIHQOA-UHFFFAOYSA-N potassium;[2-butyl-5-chloro-3-[[4-[2-(1,2,4-triaza-3-azanidacyclopenta-1,4-dien-5-yl)phenyl]phenyl]methyl]imidazol-4-yl]methanol Chemical compound [K+].CCCCC1=NC(Cl)=C(CO)N1CC1=CC=C(C=2C(=CC=CC=2)C2=N[N-]N=N2)C=C1 OXCMYAYHXIHQOA-UHFFFAOYSA-N 0.000 description 1
- 239000002243 precursor Substances 0.000 description 1
- 238000004321 preservation Methods 0.000 description 1
- 239000003755 preservative agent Substances 0.000 description 1
- 230000000750 progressive effect Effects 0.000 description 1
- 230000009696 proliferative response Effects 0.000 description 1
- 229960002429 proline Drugs 0.000 description 1
- 230000001681 protective effect Effects 0.000 description 1
- 230000002797 proteolythic effect Effects 0.000 description 1
- 210000001147 pulmonary artery Anatomy 0.000 description 1
- 230000002685 pulmonary effect Effects 0.000 description 1
- 239000002510 pyrogen Substances 0.000 description 1
- 201000002793 renal fibrosis Diseases 0.000 description 1
- 230000003362 replicative effect Effects 0.000 description 1
- 210000005000 reproductive tract Anatomy 0.000 description 1
- 230000000717 retained effect Effects 0.000 description 1
- 206010039073 rheumatoid arthritis Diseases 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 239000000523 sample Substances 0.000 description 1
- 238000006748 scratching Methods 0.000 description 1
- 230000002393 scratching effect Effects 0.000 description 1
- 230000003248 secreting effect Effects 0.000 description 1
- 230000028327 secretion Effects 0.000 description 1
- 238000004062 sedimentation Methods 0.000 description 1
- 230000019491 signal transduction Effects 0.000 description 1
- 210000002027 skeletal muscle Anatomy 0.000 description 1
- 210000001626 skin fibroblast Anatomy 0.000 description 1
- 231100000430 skin reaction Toxicity 0.000 description 1
- 230000000391 smoking effect Effects 0.000 description 1
- 229940010747 sodium hyaluronate Drugs 0.000 description 1
- 159000000000 sodium salts Chemical class 0.000 description 1
- YWIVKILSMZOHHF-QJZPQSOGSA-N sodium;(2s,3s,4s,5r,6r)-6-[(2s,3r,4r,5s,6r)-3-acetamido-2-[(2s,3s,4r,5r,6r)-6-[(2r,3r,4r,5s,6r)-3-acetamido-2,5-dihydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-2-carboxy-4,5-dihydroxyoxan-3-yl]oxy-5-hydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-3,4,5-trihydroxyoxane-2- Chemical compound [Na+].CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](O[C@H]3[C@@H]([C@@H](O)[C@H](O)[C@H](O3)C(O)=O)O)[C@H](O)[C@@H](CO)O2)NC(C)=O)[C@@H](C(O)=O)O1 YWIVKILSMZOHHF-QJZPQSOGSA-N 0.000 description 1
- 230000007928 solubilization Effects 0.000 description 1
- 238000005063 solubilization Methods 0.000 description 1
- 230000006641 stabilisation Effects 0.000 description 1
- 238000011105 stabilization Methods 0.000 description 1
- 238000004659 sterilization and disinfection Methods 0.000 description 1
- 210000002784 stomach Anatomy 0.000 description 1
- 210000002948 striated muscle cell Anatomy 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 239000002344 surface layer Substances 0.000 description 1
- 230000031068 symbiosis, encompassing mutualism through parasitism Effects 0.000 description 1
- 230000002195 synergetic effect Effects 0.000 description 1
- 210000001138 tear Anatomy 0.000 description 1
- 230000003797 telogen phase Effects 0.000 description 1
- 125000003831 tetrazolyl group Chemical group 0.000 description 1
- 230000001225 therapeutic effect Effects 0.000 description 1
- 229960004072 thrombin Drugs 0.000 description 1
- 239000012049 topical pharmaceutical composition Substances 0.000 description 1
- 231100000027 toxicology Toxicity 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- 239000012581 transferrin Substances 0.000 description 1
- 230000009466 transformation Effects 0.000 description 1
- 238000000844 transformation Methods 0.000 description 1
- 230000032258 transport Effects 0.000 description 1
- 235000019731 tricalcium phosphate Nutrition 0.000 description 1
- 102000003390 tumor necrosis factor Human genes 0.000 description 1
- 210000004026 tunica intima Anatomy 0.000 description 1
- 210000003606 umbilical vein Anatomy 0.000 description 1
- 241000712461 unidentified influenza virus Species 0.000 description 1
- 210000002700 urine Anatomy 0.000 description 1
- 230000009385 viral infection Effects 0.000 description 1
- 230000003612 virological effect Effects 0.000 description 1
Classifications
-
- 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
- A61K35/20—Milk; Whey; Colostrum
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/70—Carbohydrates; Sugars; Derivatives thereof
- A61K31/715—Polysaccharides, i.e. having more than five saccharide radicals attached to each other by glycosidic linkages; Derivatives thereof, e.g. ethers, esters
- A61K31/726—Glycosaminoglycans, i.e. mucopolysaccharides
- A61K31/728—Hyaluronic acid
-
- 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
- A61K35/14—Blood; Artificial blood
- A61K35/19—Platelets; Megacaryocytes
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
- A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L26/00—Chemical aspects of, or use of materials for, wound dressings or bandages in liquid, gel or powder form
- A61L26/0057—Ingredients of undetermined constitution or reaction products thereof
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/36—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix
- A61L27/3604—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix characterised by the human or animal origin of the biological material, e.g. hair, fascia, fish scales, silk, shellac, pericardium, pleura, renal tissue, amniotic membrane, parenchymal tissue, fetal tissue, muscle tissue, fat tissue, enamel
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P15/00—Drugs for genital or sexual disorders; Contraceptives
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P17/00—Drugs for dermatological disorders
-
- 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
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2400/00—Materials characterised by their function or physical properties
- A61L2400/06—Flowable or injectable implant compositions
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2430/00—Materials or treatment for tissue regeneration
- A61L2430/10—Materials or treatment for tissue regeneration for reconstruction of tendons or ligaments
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2430/00—Materials or treatment for tissue regeneration
- A61L2430/32—Materials or treatment for tissue regeneration for nerve reconstruction
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2430/00—Materials or treatment for tissue regeneration
- A61L2430/34—Materials or treatment for tissue regeneration for soft tissue reconstruction
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2430/00—Materials or treatment for tissue regeneration
- A61L2430/38—Materials or treatment for tissue regeneration for reconstruction of the spine, vertebrae or intervertebral discs
Definitions
- compositions based on a mixture of bioactive molecules and exosomes for use in the treatment of conditions requiring tissue repair and regeneration are included in the compositions based on a mixture of bioactive molecules and exosomes for use in the treatment of conditions requiring tissue repair and regeneration.
- the present invention relates to topical or injection compositions based on a mixture of bioactive molecules (e.g., isolated from biological tissues and fluids of mammals: placenta, colostrum, and prepartum serum) and exosomes for use in the treatment of conditions requiring tissue repair and regeneration, in humans and animals.
- bioactive molecules e.g., isolated from biological tissues and fluids of mammals: placenta, colostrum, and prepartum serum
- exosomes for use in the treatment of conditions requiring tissue repair and regeneration, in humans and animals.
- bioactive molecules mainly containing growth factors, cytokines, stem cell stimulating factors, complement proteins, antibacterial and antiviral factors, immunoglobulins can be isolated from some tissues and biological fluids of mammals such as the placenta and prepartum serum, but in particular from colostrum.
- Growth factors and cytokines are chemical messengers which mediate intracellular communication to regulate a variety of cellular functions. They can also exert their action through endocrine mechanisms, but, due to the short half-life and slow diffusion into intercellular spaces thereof, they typically act locally as autocrine or paracrine signals.
- cytokines and “growth factors” are used to indicate these molecules in a superimposable and indistinct manner, however, cytokines behave like growth factors for many cell types, for others as regulators of cell division and apoptosis or cell death and also play an important role in the innate and specific immune response and in the inflammatory phase of the tissue repair process.
- Growth factors and cytokines are thus capable of stimulating a variety of cellular processes including proliferation, migration, differentiation, and morphogenesis during the development or healing of injured tissues.
- growth factors Three main groups of growth factors are distinguished by structural homologies: insulin-like growth factors, fibroblast growth factors and transforming growth factors.
- the growth factors belonging to the first group are represented by IGF1 and IGF2 (Insulin-like Growth Factor 1 and 2), mainly synthesized by the liver and with an insulin-like structure.
- IGF1 and IGF2 carry out an insulin-like action, regulating carbohydrate homeostasis, and exert an anabolic action on bone metabolism similar to that of growth hormone (GH). Moreover, they regulate cell proliferation and differentiation, especially at the cartilage and muscle levels.
- Endothelial cells produce and respond to Fibroblast Growth Factors (FGFs): these are involved in the wound healing process by stimulating angiogenesis, collagen synthesis, epithelial cell migration and extracellular matrix synthesis.
- FGFs Fibroblast Growth Factors
- KGF Keratinocyte Growth Factor
- KGF also belongs to the same family of growth factors, playing an important role in the re-epithelialization process, stimulating the proliferation and differentiation of keratinocytes.
- TGF-a and TGF-p are instead multifunctional cytokines that play a fundamental role in tissue regeneration.
- TGF-p produced by most granulation tissue cells, including activated macrophages, stimulates fibroblast migration and proliferation, increases collagen and fibronectin synthesis, and counteracts extracellular matrix (ECM) degradation by inhibiting metalloproteinases. It acts in an autocrine manner, regulating the production thereof and stimulating monocytes to produce other growth factors, such as fibroblast growth factor (FGF), Platelet-Derived Growth Factor (PDGF) and Tumor Necrosis Factor a (TNF-a). TGF-p is also an anti-inflammatory cytokine which is used to limit and resolve inflammatory responses, inhibiting lymphocyte proliferation and the activity of other leukocytes.
- FGF fibroblast growth factor
- PDGF Platelet-Derived Growth Factor
- TGF-a Tumor Necrosis Factor a
- TGF-a is a peptide which is structurally correlated to Epidermal Growth Factor (EGF) and with a similar action.
- EGF acts on epithelial cells, stimulating cell mitosis and therefore the re-epithelialization, but also on fibroblasts, smooth muscle cells and endothelial cells, promoting angiogenesis and the process of forming new blood vessels starting from pre-existing vessels, a fundamental process in wound healing.
- the newly formed vessels must be stabilized by the intake of smooth muscle cells and by the deposition of connective tissue.
- PDGF produced by platelets, and TGF-p participate in this stabilization process: the former recalls smooth muscle cells and acts by activating the expression of TGF-p, which suppresses endothelial proliferation and migration and increases ECM protein production.
- VEGF Vascular Endothelial Growth Factors
- TNF-a belongs to the category of inflammatory cytokines mainly produced by cells of the monocyte-macrophage system. It plays a role in the wound repair process as a stimulator of angiogenesis.
- cytokines linked to innate immunity such as TNF-a, IL-1 , MCP-1 and IL-15
- those linked to the regulation of acquired immunity such as IL-2, IL- 4, IFN-y, IL-9 and IL-17
- IL-2, IL- 4, IFN-y, IL-9 and IL-17 have been highlighted as important components of the mixture of bioactive molecules.
- the cytokine IL-17 present in high concentrations in the mixture and produced in mammals by a specific class of cells referred to as Th17, allows the release of chemokines and growth factors from mesenchymal cells; moreover, the modulatory action thereof has also been demonstrated in many diseases such as asthma, Crohn's disease, multiple sclerosis, psoriasis and rheumatoid arthritis.
- the cytokine IL-17 can thus be considered one of the most important mediators of inflammatory, autoimmune, fungal and viral diseases.
- High levels of IL-15 which induces the proliferation of T, B and natural killer (NK) cells, stimulating the maturation of the immune system and also exerting an important anticancer activity, were also measured in the mixture.
- SCF Stem Cell Factor
- G-CSF Granulocyte Colony Stimulating Factor
- GM-CSF Granulocyte Macrophage Colony Stimulating Factor
- Complement proteins are components of the homonymous system which supports the body in immune and inflammatory responses.
- the system consists of over 60 proteins, but only about thirty of these circulate in the blood. In fact, these molecules are present as inactive precursors in interstitial fluids and serum and are activated when a foreign agent triggers the body's endogenous response, which thus produces antibodies to neutralize the threat.
- Lactoferrin and Transferrin are among the main proteins involved in iron metabolism. Lactoferrin, in particular, has several other functions, including acting as a direct antimicrobial agent against a wide range of bacteria and viruses. Moreover, Lactoferrin has been shown to stimulate the growth of several cell lines in vitro, including fibroblasts and intestinal epithelial cells, suggesting that the presence thereof, for example in colostrum, can be important in regulating intestinal development in infants.
- Lysozyme is an enzyme of the glycosidase group provided with a bacteriolytic action and contained in nasal mucus, saliva, tears, tissues, blood serum and breast milk. It hydrolyzes the glycosidic bond between N-acetylmuramic acid and N-acetyl-glucosamine, major constituents of the cell wall mucopeptides of many gram-positive bacteria, causing the lysis thereof. It is synthesized by white blood cells and macrophages and, with the proteolytic activity thereof, participates in the defense of the body.
- Lactoperoxidase is a peroxidase enzyme secreted by the mammary, salivary, and other mucosal glands, including the lungs, bronchi, and nose, which acts as a natural first line of defense against bacteria and viruses.
- Lactoperoxidase catalyzes the oxidation of various inorganic and organic substrates by hydrogen peroxide.
- the oxidized products, derived from the action of this enzyme, have potent and non-specific bactericidal and antiviral activities, including the destruction of the influenza virus.
- Immunoglobulins are glycoprotein molecules with antibody activity, produced by B lymphocytes in response to antigenic stimulation: the final cell figure in the series of transformations which the B lymphocyte undergoes is the plasma cell capable of secreting mature antibodies, which represent the effector molecules of humoral immunity.
- immunoglobulins 5 classes are recognized: IgG, IgA, IgM, IgD and IgE, distinguishable based on the structure of the molecule and identifiable in clinical practice based on the different molecular weight by means of electrophoresis.
- Immunoglobulins belonging to the same class have identical physicochemical features (allowing, for example, the passage of the antibody molecule through the placenta or breast milk) and the same effector functions, such as the ability to bind to immune cells (opsonization) or to activate the complement.
- IgGs which form 85% of Immunoglobulins present in the blood, activate complement and promote opsonization.
- IgMs which form up 5-10% of Immunoglobulins present in the blood, are the first to form in response to an unknown antigen.
- IgAs which form 10-20% of the Immunoglobulins present in the blood, represent the highest portion of Immunoglobulins present in exocrine secretions (milk, saliva), forming the body's first defense barrier against the entry of pathogenic germs.
- IgDs form less than 1 % of Immunoglobulins and the biological function thereof is not fully known.
- IgEs physiologically present in minimal amounts in the blood, are responsible for allergic reactions, bind to basophilic leukocytes and mast cells and cause the degranulation thereof.
- the mixture of bioactive molecules also contains APLN peptides (not properly recognized as growth factors but with functions similar to proteins belonging to this class), non-peptide trophic factors and vitamins, in particular vitamin A, vitamin E, vitamin B12, but also traces of other vitamins, such as D and provitamin A (beta carotene). Tryptic inhibitors are also present exclusively in the colostrum -derived mixture. These compounds allow bioactive molecules, when administered orally, not to be broken down by gastric digestive enzymes and to reach the intestine intact, where they exert the activity thereof or are absorbed. Some recent researches have further shown how these inhibitors can prevent the adhesion of the bacterium responsible for gastric ulcer (Helicobacter pylori) to the stomach walls.
- APLN peptides not properly recognized as growth factors but with functions similar to proteins belonging to this class
- non-peptide trophic factors in particular vitamin A, vitamin E, vitamin B12, but also traces of other vitamins, such as D and provitamin A (bet
- FIG. 1 growth factors and stem cell stimulating factors
- Figure 2 cytokines
- Figure 3 immunoglobulins, antibacterial and antiviral factors, complement proteins
- the inventors of the present patent application have surprisingly found that by combining a mixture of colostrum-isolated bioactive molecules with exosomes, its activity is unexpectedly enhanced in all fields of application.
- the present invention relates to a composition comprising a mixture of colostrum-isolated bioactive molecules and exosomes.
- the exosomes are obtained from colostrum.
- Figures 1 , 2 and 3 list the main factors present in the mixture of bioactive molecules isolated from mammalian tissues and biological fluids and in particular from colostrum (growth factors and stem cell stimulating factors - cytokines - immunoglobulins, antibacterial and antiviral factors, complement proteins).
- Figure 4 shows the flow diagram for the exosome purification procedure.
- FIG. 5 shows the protocol for the production of PRP.
- Figure 6 shows the protocol for the production of PRF.
- FIG. 7 shows the protocol for the production of PRGF.
- Figures 8 to 20 show the results obtained with the MTT Assay using different cell lines.
- Figure 21 shows a depiction of the Wound Healing Assay.
- Figure 22 shows the results obtained with the Wound Healing Assay at 24, 48 and 72 hours.
- Figure 23 shows the results obtained with the "Test on a biological model of human hair follicle".
- the composition described comprises a mixture of bioactive molecules and exosomes.
- the mixture of bioactive molecules is isolated from colostrum.
- Such a mixture of bioactive molecules comprises: growth factors, cytokines, stem cell stimulating factors, C3a/C4a complement proteins, antibacterial and antiviral factors, immunoglobulins and possibly also APLN peptides (not properly recognized as growth factors but with functions similar to proteins belonging to this class), non-peptide trophic factors and vitamins, in particular vitamin A, vitamin E, vitamin B12, but also traces of other vitamins, such as D and provitamin A (beta carotene).
- Tryptic inhibitors are also present in the colostrum-derived mixture. Mixture of colostrum-isolated bioactive molecules
- the colostrum used for the purposes of the present invention is obtained from mammals, including humans, and preferably is obtained from cattle. In fact, colostrum from herbivores has been found to contain several bioactive molecules to a greater extent.
- Bovine colostrum is the richest source of biological factors and therefore the most advantageous.
- dairy breeds have been shown to produce colostrum with the highest concentration of growth factors, cytokines, stem cell stimulating factors, complement proteins, antibacterial and antiviral factors, immunoglobulins. Taking the immunoglobulin concentration as an indicator of colostrum quality, the breed which produces the colostrum richest in bioactive factors is Jersey with 9.0% immunoglobulins, followed by Ayrshire with 8.1%, Brown Swiss with 6.6%, Guernsey with 6.3%, and Friesian (Holstein) with 5.6%.
- the colostrum used for the purposes of the present invention is preferably collected before the newborn has had the opportunity to nurse and empty the udder of the first fraction produced, regardless of the time elapsed from the moment of birth. In fact, the greatest concentration of active ingredients is present precisely in the first fraction of colostrum produced by the udder. In a preferred aspect, the cows are preferably at the second or third birth.
- the method for preparing the colostrum mixture rich in biological factors does not include steps or operations which could damage such factors, reducing the activity thereof.
- the colostrum isolated mixture rich in biological factors is obtained according to the process described by P. Sacerdote et aL, comprising the steps of:
- step 1 a 100 kg amount of colostrum is placed in a reactor and diluted 1 :10 with deionized water added with 0.9% NaCI.
- step 2) of skimming the whole mass is centrifuged at 12,400 g at a temperature of 20-25°C to remove the fat part.
- step 3 the previously obtained liquid phase is subjected to ultrafiltration through a membrane with 300 KDa cut-off, always maintaining the temperature of 20-25°C, to remove the large proteins and pathogenic microorganisms.
- the caseins are removed from the large proteins.
- step 4 of dialysis the colostrum isolate mixture rich in biological factors thus obtained is then dialyzed with a 5 KDa membrane to remove any preservatives or drug residues present in the starting colostrum and especially the lactose in solution.
- the product obtained is subjected to a series of sterilizing cross-flow filtration steps with 0.2 pm membranes and then frozen.
- the product obtained is characterized by a mixture of biological factors, also comprising the immunoglobulins (IgG, IgA and IgM) provided with a natural specificity against many bacteria and viruses.
- immunoglobulins IgG, IgA and IgM
- the mixture is subjected to a lyophilization step.
- a sterile, preservative-free, hypoallergenic powder is thus obtained with very high solubility and with the highest possible concentration of active factors.
- some key colostrum growth factors are used as markers, which are quantified by exploiting individual known ELISA tests.
- the process for preparing the mixture of colostrum-isolated active factors thus comprises a further purification step.
- a step is carried out on the concentrated product before freezing and lyophilization, and is characterized by an IgG and IgA depletion step, which, in a preferred aspect of the invention, is carried out by means of affinity chromatography.
- the colostrum purification step further comprises a step of IgM depletion, for example carried out by tangential filtration or cartridge filters.
- affinity chromatography is used by means of:
- CaptureSelectTM IgG-Fc (ms) Affinity Matrix (Thermo Fisher Scientific) or equivalent product. This system has been specifically designed to purify IgGs from different species (human, mouse, rat, rabbit, cow, horse, and sheep) by high affinity binding to the Fc region of IgG.
- CaptureSelectTM Bovine IgA Affinity Matrix (Thermo Fisher Scientific) or equivalent product. This system has been specifically designed for the purification of bovine IgA from whey/colostrum. In CaptureSelectTM Bovine IgA Affinity Matrix, affinity ligands are created using proprietary technology based on single domain antibody fragments derived from camelids.
- the depletion of IgMs and immunoglobulin aggregates occurs by means of a tangential flow filtration step which uses membranes with a molecular cut-off of about 750 kDa - 0.02 pm, or by using cartridge filters with pores of the same size.
- the pentamers of the M immunoglobulins which have an average molecular weight of 800-900 kDa, can be removed by tangential filtration, or cartridge filters, using membranes of appropriate size, while the other bioactive factors, with a lower molecular weight, can cross them without being retained.
- a further purification step of the colostrum isolate mixture is necessary to deprive it of the possible presence of bacterial endotoxins (exogenous pyrogens).
- the removal of the endotoxins occurs by means of specific kits which use an affinity chromatography, such as the Bio-Rad® Proteus Endotoxin Removal kits or equivalent products.
- affinity chromatography such as the Bio-Rad® Proteus Endotoxin Removal kits or equivalent products.
- cartridge filters with directly incorporated positively charged nylon membrane can be used which, by strongly interacting with the negatively charged endotoxins, can adsorb consistent concentrations of the latter present in solution. The best results in terms of endotoxin removal were obtained using BEA Technologies POSINYL nylon 6.6 membrane cartridge filters or equivalent product.
- the process was developed with a view to industrial application. All the process steps are transferable with a view to a large-scale purification process.
- the mixture of bioactive molecules according to the present invention is much more effective in terms of proliferative and regenerative properties, with respect to the constituent factors tested alone or in small artificially-created groups.
- the growth factors and other bioactive molecules naturally present in the mixture do not act individually, but fully carry out the action thereof only if they are in an appropriate combination, determined by nature during evolution, where all the components work in unison like the different instruments of an orchestra.
- the interaction between growth factors, cytokines, stem cell stimulating factors, complement proteins, antibacterial and antiviral factors, and exclusively for oral administration, immunoglobulins determines the initiation of a cascade of intracellular biochemical signals, followed by activation or inhibition of genes which control different cellular functions, and in particular cell proliferation and differentiation.
- bioactive molecules described are genetically highly conserved in the different species and thus for humans it is possible to use, for reparative and regenerative purposes, the biological factors isolated from other species of mammals such as cattle, horses, camels, etc.
- the disclosed composition further comprises exosomes.
- Exosomes represent the most extensively studied group among the three major subgroups of extracellular vesicles or EVs (exosomes, microvesicles, and apoptotic vesicles or ApoEVs). Among these, exosomes are the smallest vesicles, with a variable diameter of 50-150 nm and a cup-shaped morphology when viewed under an electron microscope.
- the biogenesis process of the exosome includes several steps: 1 ) the inward folding of a section of the plasma membrane forms the early endosomes; 2) the first endosomes mature into late endosomes and then into multivesicular bodies (MVBs) characterized by intraluminal vesicles (ILVs); 3) once they reach maturity, these MVBs are pulled by means of a network of microtubules and cytoskeletons towards the cell membrane where, by exocytosis, they will fuse therewith, releasing the ILVs into the extracellular environment which will become exosomes.
- This process of biogenesis and exosome formation is finely directed by the Endosomal Sorting Complexes Required for Transport (ESCRT) and accessory proteins.
- Exosomes possess a specific molecular load which includes membrane proteins, cytosolic and nuclear proteins, extracellular matrix proteins, lipids, nucleic acids (mRNA, miRNA and DNA), growth factors, cytokines and enzymes, underlining the involvement thereof in the regulation, by means of different molecular mechanisms, of various biological functions.
- exosomes The absorption of exosomes depends on the interactions between the proteins expressed on the surface thereof and the recipient cells.
- adhesion molecules on the surface of exosomes such as tetraspanins, glycoproteins, and integrins, determine which cells can “accept” exosomes.
- Exosomes are secreted by most cell types including immune cells (B cells, T cells), neuronal cells, epithelial cells, endothelial cells, embryonic cells, and mesenchymal stem cells (MSCs).
- B cells immune cells
- T cells neuronal cells
- epithelial cells epithelial cells
- endothelial cells endothelial cells
- embryonic cells embryonic cells
- mesenchymal stem cells MSCsenchymal stem cells
- Exosomes have been detected in almost all body fluids, under both physiological and pathological conditions, including: urine, blood, serum, breast milk, amniotic fluid, cerebrospinal fluid, saliva, bile, and lymph.
- Exosomes are also contained in colostrum, albeit in smaller quantities.
- exosomes can be considered “transporters”, therefore it was thought to exploit this ability thereof to enter other cells to transfer useful "loads” including chemotherapeutic and immunomodulatory agents, with the ability to direct the release thereof towards a given target.
- useful "loads” including chemotherapeutic and immunomodulatory agents, with the ability to direct the release thereof towards a given target.
- exosomes allow crossing the blood-brain barrier and bringing the molecules of interest to the desired CNS sites.
- inserting these useful "loads” inside the vesicles requires the manipulation of the vesicles themselves (exogenous loading in which the small molecules are incorporated from the outside into the isolated EVs) or of the cells from which they come (endogenous loading in which the cell receives the necessary means to incorporate the small molecules in the EVs during the formation process).
- exosomes do not induce toxicity when injected repeatedly into mice and are well tolerated, thus the therapeutic use thereof is very promising.
- UC ultracentrifugation
- the UC method for isolating exosomes is mainly divided into two steps: first, a series of continuous centrifugations at medium-low speed to remove dead cells, cellular debris and large extracellular vesicles. The UC is then carried out at 100,000xg to separate the exosomes, followed by a series of washes in phosphate buffer (PBS), at the same UC speed, to remove impurities such as contaminatingproteins.
- PBS phosphate buffer
- the inventors of the present patent application have surprisingly found that a highly concentrated exosome preparation is capable of enhancing the activity of the mixture of biological molecules of the present invention.
- the exosomes are isolated and concentrated from bovine colostrum.
- a preferable condition is that the exosomes are isolated and concentrated from colostrum, rather than from other sources, and that the latter are added to the mixture in a functional amount.
- a highly concentrated exosome preparation is a preparation containing an amount of exosomes >1x10 9 /mL, preferably about 20-50x10 9 /mL.
- the two components can be present, independently of each other, in the following amounts (in distilled water or saline):
- the two components can be present, independently of each other, in the following amounts:
- the two components can be present, independently of each other, in the following amounts:
- the present patent application describes formulations comprising a composition according to the invention.
- such formulations are represented by: liquid solutions, lotions, foams, sprays, creams, salves, ointments, pastes, gels, membranes, clots, powders or other forms suitable for the medical use.
- Such formulations can one or more additives, excipients and the other components necessary to obtain such formulations according to what is known in the art.
- the formulations described can optionally comprise one or more further effective components.
- This can be present in various forms (esters, salts, hydrolysate, crosslinked, liposomal, etc.), among which sodium hyaluronate, the sodium salt thereof, in amounts between 5 and 50 mg/mL of distilled water or saline, preferably 25 mg/mL, is preferred.
- Hyaluronic acid can further confer the ability to restore tissue tone and elasticity, as well as enhance and accelerate the re-epithelialization process.
- These can be present in a mixture, in amounts between 10 and 50 mg/mL of distilled water or saline, preferably 20 mg/mL.
- Such amino acids can further confer the ability to stimulate collagen regeneration.
- Hydroxyapatite is the main inorganic component of bone and makes up 60-70% of the calcified skeleton and 98% of tooth enamel. It is biocompatible, binds quickly to adjacent hard and soft tissues and has a strong osteoconductive capacity. The clinical indications of hydroxyapatite concern the reconstruction of bone tissue and the lining of endo-osseous dental implants to promote osseointegration.
- Hydroxyapatite is also used as a component of skin fillers.
- the amounts used vary in relation to the surfaces to be reconstructed or covered.
- 1 mL of gel is used containing from 20 to 50% of hydroxyapatite microspheres in 5 mL of distilled water or saline, preferably 40% of microspheres.
- P-tricalcium phosphate is a polycrystalline bioceramic, with osteoconductive properties, which in contact with water releases hydroxyapatite crystals. It is used for the resolution of deep intraosseous periodontal defects (periodontal regeneration), for filling post-extraction bone cavities, for bone regeneration, etc.
- p- tricalcium phosphate has a progressive resorption, unlike what occurs with hydroxyapatite, with a release of calcium and phosphorus ions which, for example in the case of intraosseous periodontal defects, contribute to the neo-apposition of bone, cement, and periodontal ligament. It is used in the amount of 1 g/mL of distilled water or saline.
- compositions of the invention can further comprise Platelet Concentrates (PCs).
- PCs Platelet Concentrates
- the platelet concentrates are autologous.
- APCs Autologous Platelet Concentrates
- the final product has a platelet concentration above the baseline level, therefore it has a higher number of bioactive molecules derived from the platelets themselves.
- the logic of the clinical use of such preparations is based on the concept of exploiting the thus enriched content thereof to stimulate different biological functions, such as chemotaxis, angiogenesis, proliferation and differentiation, so as to promote the healing of hard and soft tissues.
- Platelet-Rich Plasma really began with Marx in 1998 when he published a comparative clinical study in which the regenerative potential of PRP was demonstrated in a series of patients undergoing mandibular reconstruction. PRP has then been associated with the concept of platelet growth factors and the potential contribution thereof in inducing tissue healing.
- the patient’s blood is collected in test tubes containing anticoagulants and processed by means of two centrifugation steps.
- Figure 5 diagrammatically shows the specific protocol.
- the PRP thus obtained can be applied to the site to be treated with a syringe or activated by thrombin and/or calcium chloride to trigger platelet activation and stimulate fibrin polymerization.
- the first low- intensity centrifugation allows the separation of the blood into three distinct layers: red blood cells at the bottom, a cell plasma (Platelet-Poor Plasma or PPP) at the top and a whitish layer referred to as the buffy coat, located therebetween, containing the highest concentration of platelets and leukocytes.
- PPP Pure-PRP
- the PPP and the buffy coat surface layer are transferred to another test tube and centrifuged at high intensity (hard spin), then most of the PPP and leukocytes are discarded and the P-PRP can be collected.
- L-PRP leukocyte rich PRP
- the PPP, the entire buffy coat layer and some residual red blood cells are collected and transferred to another test tube to be centrifuged at high intensity (hard spin), then most of the PPP is discarded thus obtaining an L-PRP containing the buffy coat with most of the platelets and leukocytes, some residual red blood cells and PPP.
- PRP leukocyte rich PRP
- a protocol was developed for producing a blood component referred to as Platelet-Rich Fibrin (PRF).
- PRF Platelet-Rich Fibrin
- blood is collected in test tubes without anticoagulant and centrifuged at a moderate speed.
- Three layers are thus formed inside the test tube: red blood cells and acellular plasma are located, respectively, in the lower and upper part thereof, while the fibrin clot, positioned therebetween, forms the PRF ( Figure 6). Since the PRF clot naturally forms inside the test tube, it has a robust fibrin matrix in which most of the platelets and leukocytes are trapped.
- Injectable PRF (i-PRF) has also been recently developed, which can be obtained with a further, even more delicate, centrifugation step. It has a liquid form, is very rich in white blood cells and can be used for infiltration into tissues and joints.
- PRGF Plasma Poor in Growth Factors
- Figure 7 Plasma Poor in Growth Factors
- PRGF can be applied as a liquid fraction in the target site or it can be pre-activated by adding 0.2 mL of 10% CaCIs to induce clot formation.
- the form thereof can vary based on the different applications (liquid form, gel, membranes or fibrin clots, etc.), as well as preparation protocols, there is multiple clinical evidence on the effectiveness of platelet concentrates in different fields of medicine.
- compositions and formulations detailed above are described for the medical use.
- these can be advantageously used for the treatment of those conditions requiring tissue repair and regeneration.
- the application is in humans and animals.
- compositions and formulations described find use in oral and maxillofacial regenerative surgical procedures; in orthopedics and sports medicine; in dermatology and aesthetic medicine; in gynecology; in andrology; in plastic and reconstructive surgery; in ophthalmology; in otolaryngology; in neuroscience; in the repair of organ injuries (brain, heart, lungs, liver, kidneys, larynx, pharynx, esophagus, etc.).
- composition can possibly be modified by adding the following components (independently of each other):
- composition can possibly be modified by adding the following components (independently of each other):
- compositions based on a mixture of bioactive molecules and a functional amount of exosomes, were evaluated on a limited but highly significant number of cell cultures.
- compositions comprising the exosomes isolated and concentrated from colostrum, with respect to the other sources, emerges.
- the entire test panel was then performed in vitro with the composition based on a mixture of bioactive molecules from colostrum + highly concentrated exosomes from colostrum.
- the MTT assay is used to measure the cellular metabolic activity, as an indicator of cell viability and proliferation.
- This colorimetric test is based on the reduction of a yellow tetrazolium salt (3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide or MTT) to purple formazan crystals by metabolically active cells.
- Vital cells contain NAD(P) H-dependent oxidoreductase enzymes which reduce MTT to formazan.
- the insoluble formazan crystals are dissolved using a solubilization solution and the resulting colored solution is quantified by measuring absorbance at 500-600 nanometers using a multi-well spectrophotometer. The darker the solution, the greater the number of viable and metabolically active cells.
- This colorimetric test can be used for multiple applications, such as quantifying cell growth and viability and measuring cell proliferation in response to growth factors, cytokines, and nutrients.
- the following cell lines were used, necessary to demonstrate the proliferative and regenerative activity of the preparations of the present invention on the different tissues and organs.
- keratinocytes are the most common type of skin cell, as they form the structural component of the epidermis.
- Keratinocytes are used for scientific applications including studying the behavior of growth factors, wound repair, skin cancers, response to UV radiation, psoriasis, eczema, viral infections, cell differentiation, and cosmetics research.
- fibroblasts play an important role in maintaining the structural integrity of connective tissue and protein synthesis of the extracellular matrix such as collagens, glycosaminoglycans and glycoproteins, molecules also involved in wound healing.
- Fibroblasts are mediators in inflammation, cancer, angiogenesis, and pathological tissue fibrosis and are frequently used in studies related to wound healing, tissue engineering, and regenerative applications. Therefore, the cell line of primary skin fibroblasts finds application in several fields of research: in the response to pathogens, skin ageing, wound healing and skin diseases, including scleroderma.
- HAEC Normal, Human
- Endothelial cells form the tunica intima, the thin layer of cells lining the inner surface of blood vessels. This surface acts as a selective filter to regulate the passage of gases, fluids, immune cells and various molecules.
- Primary endothelial cell cultures can be isolated from the umbilical vein, aorta, pulmonary and coronary arteries and dermal vascular tissue and represent useful tools for the study of angiogenesis, cancer therapy, wound healing, burn therapy, tissue engineering and regeneration.
- HCEpC Primary Corneal Epithelial Cells; Normal, Human (HCEpC): primary corneal epithelial cell cultures are used as a model for studying eye re- epithelialization following photorefractive keratectomy (PRK) or Laser-ASsisted In situ Keratomileusis (LASIK). They are also used to study the regeneration of damaged corneas.
- PRK photorefractive keratectomy
- LASIK Laser-ASsisted In situ Keratomileusis
- HVEC Primary Vaginal Epithelial Cells; Normal, Human (HVEC): primary vaginal epithelial cell cultures can be used to study the cellular physiology of the reproductive tract, the cellular response to infectious agents, and in other areas of research, including the regenerative activity of the damaged or atrophic vaginal epithelium.
- HSkMC Normal, Human
- Primary Osteoblasts Normal, Human (NHOst): primary osteoblast cultures provide a homogeneous and rapidly proliferating model for the in vitro study of normal osteoblast differentiation, the physiology thereof, and the effects of hormones, growth factors, and other cytokines on osteoblast function and differentiation in vivo.
- HSC Normal, Human
- Schwann cells represent the main type of glial cells. The proliferation in vitro thereof can be stimulated by various growth factors including PDGF, FGF, neuregulin and others.
- Primary Schwann cell cultures provide a relatively simple, well-defined, and accessible model for studying neuropathies, nerve regeneration, and several diseases of nervous system development.
- HEMa Human, Adult
- Primary epidermal melanocyte cultures are used for the in vitro study of wound healing and as a model for toxicity and irritability studies, for studies of skin response to UV radiation, psoriasis and other skin diseases and also in cosmetic research (e.g., skin lightening compounds and skin protective compounds).
- HSAEC Normal, Human
- the wound healing assay also known as the "scratch test," is a well- established two-dimensional (2D) technique which can be used to study wound healing in vitro. This method was one of the first to be developed for the study of cell migration and measures the speed with which cells, in a cell monolayer, migrate to fill a free space.
- the technique reproduces the wound by creating an empty space in a confluent cellular monolayer and consists of four main steps which are described below ( Figure 21 shows an overview thereof).
- the first step of the test is the culture of a confluent cell monolayer.
- This monolayer represents the in vivo condition of the tissue prior to injury, as an intact epithelium.
- Epithelial or endothelial cells are normally used to produce the monolayer.
- Human corneal epithelial cells HCEpC were used to evaluate the effectiveness of the composition of the present invention in regenerative medicine.
- Optical microscopy using phase contrast imaging can be used to observe cells migrating into the wound area.
- a series of time-lapse images can be acquired as the cells migrate into the cell free space. These time points are then collected at 24, 48 and 72 hours after the start of the experiment. The migration images can then be used to collect measurements or be evaluated visually.
- the method used for the purposes of the present patent calculates the change in wound area over time as a percentage of wound closure.
- the hair follicles used in this test show a growth phase (anagen) of 15 days before moving on to the regression phase (catagen).
- the biological model used for this test is represented by hair follicles microsectioned from fragments of human skin.
- the purpose of the test is to verify the growth of the hair follicle in the presence or not of the substances to be tested: the hair follicles were thus cultured in the William culture medium without the addition of other compositions (negative control), in the same culture medium with the addition of the composition of the present patent, i.e., the composition based on a mixture of bioactive molecules from colostrum + highly concentrated exosomes from colostrum (positive control), or with the addition of the other two reference products (mixture alone or exosomes alone). Hair follicle growth was followed for 18 days, with measurements of the length thereof being taken at regular intervals.
- the in vitro tests performed to evaluate the proliferative and regenerative effectiveness of the composition of the present invention in the multiple medical uses are summarized below.
- the in vitro tests were performed using the MTT Assay with NHAC-kn cells (treating osteoarthritis, treating degenerative disease of the intervertebral discs); with NHOst, HAEC cells (managing fractures and nonunions); with HDFa cells (treating tendon injuries, enhancing the post-surgery healing process of tendons and ligaments) and with HSkMC cells (treating muscle injuries).
- NHAC-kn cells treating osteoarthritis, treating degenerative disease of the intervertebral discs
- NHOst, HAEC cells managing fractures and nonunions
- HDFa cells treating tendon injuries, enhancing the post-surgery healing process of tendons and ligaments
- HSkMC cells treating muscle injuries
- the in vitro tests were performed using the MTT Assay with HEKa, HDFa cells (skin rejuvenation, treating acne, treating stretch marks, treating melasma, treating periorbital hyperpigmentation); with HEMa cells (treating vitiligo), as well as with the "Test on a biological model of human hair follicle" for treating alopecia.
- the in vitro tests were performed using the Wound Healing Assay and the MTT Assay with HVEC, HDFa cells (treating skin injuries, healing surgical wounds, treating vaginal atrophy and vaginal dryness, treating vulvar dystrophy, aesthetic gynecology in vaginal rejuvenation).
- the in vitro tests were performed using the MTT Assay with HAEC cells (treating erectile dysfunction).
- the in vitro tests were performed using the Wound Healing Assay and the MTT Assay with HEKa, HDFa, HAEC cells (treating burns, treating wounds and skin ulcers, treating keloids and hypertrophic scars post-excision); and with HDFa, HSkMC cells (breast reconstruction).
- the in vitro tests were performed using the Wound Healing Assay and the MTT Assay with HCEpC cells (treating persistent corneal epithelial defects, treating corneal abrasions and ulcers, corneal surgery, treating dry eye syndrome, treating ocular surface syndrome after Laser-ASsisted In situ Keratomileusis - LASIK or after photorefractive keratectomy - PRK).
- the in vitro tests were performed using the Wound Healing Assay and the MTT Assay with HEKa, HDFa, HAEC cells (treating atrophic rhinitis, treating tympanic membrane perforation).
- the in vitro tests were performed using the MTT Assay with HSwC cells (treating traumatic brain injuries, treating traumatic spinal cord injuries, treating neurodegenerative and autoimmune diseases of the CNS, treating peripheral nerve injuries).
- the results obtained with the composition of the present patent application were far superior to the results obtained with all the other products tested or with the reference sample (control), both in terms of proliferative and regenerative effectiveness (MTT Assay, Figures 8 to 20; Wound Healing Assay, Figure 22; Test on a biological model of human hair follicle, Figure 23).
- the bioactive molecules of the mixture can act, promoting the reparative process; this occurs by stimulating neoangiogenesis, even in poorly vascularized tissues, and the regeneration of damaged tissues which are replaced with cells of the same type, thus avoiding the replacement thereof with connective tissue (fibrosis).
- the application of the composition of the present invention thus prevents the onset of hypertrophic or keloid scars, which represent one of the most common wound healing disorders.
- exosomes of the present invention remain in a living and vital state even up to 6-8 months and even in the absence of lyophilization, thus demonstrating a high and unexpected stability.
- one or more excipients can be modified according to the pharmaceutical form selected.
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- General Health & Medical Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- Medicinal Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Pharmacology & Pharmacy (AREA)
- Epidemiology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Biomedical Technology (AREA)
- Zoology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Chemical & Material Sciences (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Organic Chemistry (AREA)
- Dermatology (AREA)
- Hematology (AREA)
- Molecular Biology (AREA)
- Biotechnology (AREA)
- Virology (AREA)
- Immunology (AREA)
- Developmental Biology & Embryology (AREA)
- Cell Biology (AREA)
- Reproductive Health (AREA)
- Physical Education & Sports Medicine (AREA)
- Endocrinology (AREA)
- Urology & Nephrology (AREA)
- Botany (AREA)
- Oral & Maxillofacial Surgery (AREA)
- Transplantation (AREA)
- Materials Engineering (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
Abstract
The present invention relates to topical or injection compositions for use in the treatment of conditions requiring tissue repair and regeneration in humans and animals, comprising a mixture of bioactive molecules and exosomes.
Description
“Compositions based on a mixture of bioactive molecules and exosomes for use in the treatment of conditions requiring tissue repair and regeneration”
DESCRIPTION
The present invention relates to topical or injection compositions based on a mixture of bioactive molecules (e.g., isolated from biological tissues and fluids of mammals: placenta, colostrum, and prepartum serum) and exosomes for use in the treatment of conditions requiring tissue repair and regeneration, in humans and animals.
It is known that a mixture of bioactive molecules mainly containing growth factors, cytokines, stem cell stimulating factors, complement proteins, antibacterial and antiviral factors, immunoglobulins can be isolated from some tissues and biological fluids of mammals such as the placenta and prepartum serum, but in particular from colostrum.
Growth factors and cytokines
Growth factors and cytokines are chemical messengers which mediate intracellular communication to regulate a variety of cellular functions. They can also exert their action through endocrine mechanisms, but, due to the short half-life and slow diffusion into intercellular spaces thereof, they typically act locally as autocrine or paracrine signals.
These messengers bind to receptors present on the cell surface thus initiating, at the membrane level and in the intracellular environment, a cascade of events involved in the signal transduction process.
Often, the terms "cytokines" and "growth factors" are used to indicate these molecules in a superimposable and indistinct manner, however, cytokines behave like growth factors for many cell types, for others as regulators of cell division and apoptosis or cell death and also play an important role in the innate and specific immune response and in the inflammatory phase of the tissue repair process.
Growth factors and cytokines are thus capable of stimulating a variety of
cellular processes including proliferation, migration, differentiation, and morphogenesis during the development or healing of injured tissues.
Three main groups of growth factors are distinguished by structural homologies: insulin-like growth factors, fibroblast growth factors and transforming growth factors.
The growth factors belonging to the first group, also referred to as somatomedins, are represented by IGF1 and IGF2 (Insulin-like Growth Factor 1 and 2), mainly synthesized by the liver and with an insulin-like structure. IGF1 and IGF2 carry out an insulin-like action, regulating carbohydrate homeostasis, and exert an anabolic action on bone metabolism similar to that of growth hormone (GH). Moreover, they regulate cell proliferation and differentiation, especially at the cartilage and muscle levels.
Endothelial cells produce and respond to Fibroblast Growth Factors (FGFs): these are involved in the wound healing process by stimulating angiogenesis, collagen synthesis, epithelial cell migration and extracellular matrix synthesis. Keratinocyte Growth Factor, known as KGF, also belongs to the same family of growth factors, playing an important role in the re-epithelialization process, stimulating the proliferation and differentiation of keratinocytes.
Transforming Growth Factors TGF-a and TGF-p are instead multifunctional cytokines that play a fundamental role in tissue regeneration.
TGF-p produced by most granulation tissue cells, including activated macrophages, stimulates fibroblast migration and proliferation, increases collagen and fibronectin synthesis, and counteracts extracellular matrix (ECM) degradation by inhibiting metalloproteinases. It acts in an autocrine manner, regulating the production thereof and stimulating monocytes to produce other growth factors, such as fibroblast growth factor (FGF), Platelet-Derived Growth Factor (PDGF) and Tumor Necrosis Factor a (TNF-a). TGF-p is also an anti-inflammatory cytokine which is used to limit and resolve inflammatory responses, inhibiting lymphocyte
proliferation and the activity of other leukocytes.
TGF-a is a peptide which is structurally correlated to Epidermal Growth Factor (EGF) and with a similar action. EGF acts on epithelial cells, stimulating cell mitosis and therefore the re-epithelialization, but also on fibroblasts, smooth muscle cells and endothelial cells, promoting angiogenesis and the process of forming new blood vessels starting from pre-existing vessels, a fundamental process in wound healing. The newly formed vessels must be stabilized by the intake of smooth muscle cells and by the deposition of connective tissue. PDGF, produced by platelets, and TGF-p participate in this stabilization process: the former recalls smooth muscle cells and acts by activating the expression of TGF-p, which suppresses endothelial proliferation and migration and increases ECM protein production.
Vascular Endothelial Growth Factors (VEGF) also make an important contribution in the angiogenesis process, stimulating the migration and proliferation of endothelial cells and contributing to the formation of the vascular lumen. TNF-a, mentioned above, belongs to the category of inflammatory cytokines mainly produced by cells of the monocyte-macrophage system. It plays a role in the wound repair process as a stimulator of angiogenesis.
Both the cytokines linked to innate immunity, such as TNF-a, IL-1 , MCP-1 and IL-15, and those linked to the regulation of acquired immunity, such as IL-2, IL- 4, IFN-y, IL-9 and IL-17, have been highlighted as important components of the mixture of bioactive molecules.
The cytokine IL-17, present in high concentrations in the mixture and produced in mammals by a specific class of cells referred to as Th17, allows the release of chemokines and growth factors from mesenchymal cells; moreover, the modulatory action thereof has also been demonstrated in many diseases such as asthma, Crohn's disease, multiple sclerosis, psoriasis and rheumatoid arthritis. The cytokine IL-17 can thus be considered one of the most important mediators of
inflammatory, autoimmune, fungal and viral diseases. High levels of IL-15, which induces the proliferation of T, B and natural killer (NK) cells, stimulating the maturation of the immune system and also exerting an important anticancer activity, were also measured in the mixture.
Stem cell stimulating factors
Stem Cell Factor (SCF) is involved in the development of hematopoietic, gonadal, and pigmented cell lines. It has a very wide range of activities with direct effects on the development of myeloid and lymphoid cells and potent synergistic effects with other growth factors such as GM-CSF, IL-7 and erythropoietin.
Granulocyte Colony Stimulating Factor (G-CSF) and Granulocyte Macrophage Colony Stimulating Factor (GM-CSF) are key substances for the proliferation, maturation and expansion of bone marrow progenitor cells. These factors are capable of binding specific receptor proteins located on the surfaces of hematopoietic stem cells, present in bone marrow. The bond between receptor and colony-stimulating factor activates intracellular metabolic pathways that cause the cells to proliferate and differentiate into a specific type of blood cell.
The role thereof as stimulating factors in stem cells is thus now well defined.
C3a/C4a complement proteins
Complement proteins are components of the homonymous system which supports the body in immune and inflammatory responses.
They are mostly synthesized (90%) by the liver, with the rest instead synthesized by fibroblasts, monocytes and macrophages.
The system consists of over 60 proteins, but only about thirty of these circulate in the blood. In fact, these molecules are present as inactive precursors in interstitial fluids and serum and are activated when a foreign agent triggers the body's endogenous response, which thus produces antibodies to neutralize the threat.
There are nine main proteins of the complement system, indicated from C1
to C9. These, together with all the other proteins in the system, interact in a cascade pattern, forming complexes which work to support the immune system in different situations such as infections, inflammation or apoptosis.
Fractions C3/a and C4/a are the most represented in the mixture of bioactive molecules isolated from mammalian tissues and biological fluids.
Antibacterial and antiviral factors
Lactoferrin and Transferrin are among the main proteins involved in iron metabolism. Lactoferrin, in particular, has several other functions, including acting as a direct antimicrobial agent against a wide range of bacteria and viruses. Moreover, Lactoferrin has been shown to stimulate the growth of several cell lines in vitro, including fibroblasts and intestinal epithelial cells, suggesting that the presence thereof, for example in colostrum, can be important in regulating intestinal development in infants.
Lysozyme is an enzyme of the glycosidase group provided with a bacteriolytic action and contained in nasal mucus, saliva, tears, tissues, blood serum and breast milk. It hydrolyzes the glycosidic bond between N-acetylmuramic acid and N-acetyl-glucosamine, major constituents of the cell wall mucopeptides of many gram-positive bacteria, causing the lysis thereof. It is synthesized by white blood cells and macrophages and, with the proteolytic activity thereof, participates in the defense of the body.
Lactoperoxidase is a peroxidase enzyme secreted by the mammary, salivary, and other mucosal glands, including the lungs, bronchi, and nose, which acts as a natural first line of defense against bacteria and viruses.
Lactoperoxidase catalyzes the oxidation of various inorganic and organic substrates by hydrogen peroxide. The oxidized products, derived from the action of this enzyme, have potent and non-specific bactericidal and antiviral activities, including the destruction of the influenza virus.
Immunoglobulins
Immunoglobulins are glycoprotein molecules with antibody activity, produced by B lymphocytes in response to antigenic stimulation: the final cell figure in the series of transformations which the B lymphocyte undergoes is the plasma cell capable of secreting mature antibodies, which represent the effector molecules of humoral immunity.
5 classes of immunoglobulins are recognized: IgG, IgA, IgM, IgD and IgE, distinguishable based on the structure of the molecule and identifiable in clinical practice based on the different molecular weight by means of electrophoresis.
Immunoglobulins belonging to the same class have identical physicochemical features (allowing, for example, the passage of the antibody molecule through the placenta or breast milk) and the same effector functions, such as the ability to bind to immune cells (opsonization) or to activate the complement.
IgGs, which form 85% of Immunoglobulins present in the blood, activate complement and promote opsonization. IgMs, which form up 5-10% of Immunoglobulins present in the blood, are the first to form in response to an unknown antigen. IgAs, which form 10-20% of the Immunoglobulins present in the blood, represent the highest portion of Immunoglobulins present in exocrine secretions (milk, saliva), forming the body's first defense barrier against the entry of pathogenic germs. IgDs form less than 1 % of Immunoglobulins and the biological function thereof is not fully known. IgEs, physiologically present in minimal amounts in the blood, are responsible for allergic reactions, bind to basophilic leukocytes and mast cells and cause the degranulation thereof.
Other components
The mixture of bioactive molecules also contains APLN peptides (not properly recognized as growth factors but with functions similar to proteins belonging to this class), non-peptide trophic factors and vitamins, in particular vitamin A, vitamin E, vitamin B12, but also traces of other vitamins, such as D and provitamin A (beta carotene).
Tryptic inhibitors are also present exclusively in the colostrum -derived mixture. These compounds allow bioactive molecules, when administered orally, not to be broken down by gastric digestive enzymes and to reach the intestine intact, where they exert the activity thereof or are absorbed. Some recent researches have further shown how these inhibitors can prevent the adhesion of the bacterium responsible for gastric ulcer (Helicobacter pylori) to the stomach walls.
The main bioactive molecules described above are shown in Figure 1 (growth factors and stem cell stimulating factors), Figure 2 (cytokines) and Figure 3 (immunoglobulins, antibacterial and antiviral factors, complement proteins) .
Summary of the invention
The inventors of the present patent application have surprisingly found that by combining a mixture of colostrum-isolated bioactive molecules with exosomes, its activity is unexpectedly enhanced in all fields of application.
Object of the invention
In a first object, the present invention relates to a composition comprising a mixture of colostrum-isolated bioactive molecules and exosomes.
According to a particular aspect, the exosomes are obtained from colostrum.
In a second object of the invention, formulations comprising the composition of the invention are described.
In a third object, it is described the medical use of the disclosed composition.
The multiple medical uses of the disclosed composition represent further aspects of the present invention.
Brief description of the drawings
Figures 1 , 2 and 3 list the main factors present in the mixture of bioactive molecules isolated from mammalian tissues and biological fluids and in particular from colostrum (growth factors and stem cell stimulating factors - cytokines -
immunoglobulins, antibacterial and antiviral factors, complement proteins).
Figure 4 shows the flow diagram for the exosome purification procedure.
Figure 5 shows the protocol for the production of PRP.
Figure 6 shows the protocol for the production of PRF.
Figure 7 shows the protocol for the production of PRGF.
Figures 8 to 20 show the results obtained with the MTT Assay using different cell lines.
Figure 21 shows a depiction of the Wound Healing Assay.
Figure 22 shows the results obtained with the Wound Healing Assay at 24, 48 and 72 hours.
Figure 23 shows the results obtained with the "Test on a biological model of human hair follicle".
Detailed description of the invention
According to a first object of the invention, the composition described comprises a mixture of bioactive molecules and exosomes.
For the purposes of the present invention, the mixture of bioactive molecules is isolated from colostrum.
Alternative sources to colostrum are represented by other tissues and biological fluids of mammals.
In particular, it is the placenta and the prepartum serum.
Such a mixture of bioactive molecules comprises: growth factors, cytokines, stem cell stimulating factors, C3a/C4a complement proteins, antibacterial and antiviral factors, immunoglobulins and possibly also APLN peptides (not properly recognized as growth factors but with functions similar to proteins belonging to this class), non-peptide trophic factors and vitamins, in particular vitamin A, vitamin E, vitamin B12, but also traces of other vitamins, such as D and provitamin A (beta carotene).
Tryptic inhibitors are also present in the colostrum-derived mixture.
Mixture of colostrum-isolated bioactive molecules
It is possible to obtain a mixture of biological factors from the colostrum with an appropriate method, comprising: growth factors, cytokines, stem cell stimulating factors, complement proteins C3a/C4a, antibacterial and antiviral factors, immunoglobulins.
The colostrum used for the purposes of the present invention is obtained from mammals, including humans, and preferably is obtained from cattle. In fact, colostrum from herbivores has been found to contain several bioactive molecules to a greater extent.
Bovine colostrum is the richest source of biological factors and therefore the most advantageous. In particular, dairy breeds have been shown to produce colostrum with the highest concentration of growth factors, cytokines, stem cell stimulating factors, complement proteins, antibacterial and antiviral factors, immunoglobulins. Taking the immunoglobulin concentration as an indicator of colostrum quality, the breed which produces the colostrum richest in bioactive factors is Jersey with 9.0% immunoglobulins, followed by Ayrshire with 8.1%, Brown Swiss with 6.6%, Guernsey with 6.3%, and Friesian (Holstein) with 5.6%.
These biological factors are highly conserved in phylogenesis and therefore have a considerable cross-reaction activity between the various species, therefore it is possible to use factors isolated from other mammals such as cattle, horses, camelids, etc. on humans.
The colostrum used for the purposes of the present invention is preferably collected before the newborn has had the opportunity to nurse and empty the udder of the first fraction produced, regardless of the time elapsed from the moment of birth. In fact, the greatest concentration of active ingredients is present precisely in the first fraction of colostrum produced by the udder. In a preferred aspect, the cows are preferably at the second or third birth.
Advantageously, the method for preparing the colostrum mixture rich in
biological factors does not include steps or operations which could damage such factors, reducing the activity thereof.
For example, no heat sterilization steps are carried out which can lead to the loss of large amounts of biological factors in the final product following the degradation caused by high temperatures. Or, the separation of casein, necessary as it can cause allergic reactions, cannot include a precipitation step at low pH values which can lead to the denaturation of many proteins, including various biological factors present in the colostrum.
Therefore, for the preparation of the mixture of colostrum-isolated bioactive molecules according to the present invention, extraction processes which allow obtaining a mixture of factors in a biologically active form will be used.
Processes which can be employed for the preparation of the mixture rich in biological factors are described for example by P. Sacerdote et al. ("Biological components in a standardized derivative of bovine colostrum" - Journal of Dairy Science, Volume 96, Issue 3, 1 March 2013, pages 1745 - 1754) and in international patent application WO2017134559. Other extraction processes may be used, as long as they meet the above requirements.
In a preferred aspect of the invention, the colostrum isolated mixture rich in biological factors is obtained according to the process described by P. Sacerdote et aL, comprising the steps of:
1 ) dilution;
2 ) skimming;
3 ) ultrafiltration;
4 ) dialysis;
5 ) microfiltration.
In another preferred aspect of the invention, in step 1 ), a 100 kg amount of colostrum is placed in a reactor and diluted 1 :10 with deionized water added with 0.9% NaCI.
In a further preferred aspect of the invention, in step 2) of skimming, the whole mass is centrifuged at 12,400 g at a temperature of 20-25°C to remove the fat part.
According to a particularly preferred aspect of the invention, in step 3), the previously obtained liquid phase is subjected to ultrafiltration through a membrane with 300 KDa cut-off, always maintaining the temperature of 20-25°C, to remove the large proteins and pathogenic microorganisms. For example, the caseins are removed from the large proteins.
In another particularly preferred aspect of the invention, in step 4) of dialysis, the colostrum isolate mixture rich in biological factors thus obtained is then dialyzed with a 5 KDa membrane to remove any preservatives or drug residues present in the starting colostrum and especially the lactose in solution.
In a further particularly preferred aspect of the invention, in the microfiltration step 5), the product obtained is subjected to a series of sterilizing cross-flow filtration steps with 0.2 pm membranes and then frozen.
Therefore, the product obtained is characterized by a mixture of biological factors, also comprising the immunoglobulins (IgG, IgA and IgM) provided with a natural specificity against many bacteria and viruses.
As the last step of the preparatory process, the mixture is subjected to a lyophilization step.
A sterile, preservative-free, hypoallergenic powder is thus obtained with very high solubility and with the highest possible concentration of active factors.
In a preferred aspect, to evaluate the quality of the processes implemented to obtain a standardized final product, some key colostrum growth factors are used as markers, which are quantified by exploiting individual known ELISA tests.
If it is desired to obtain a mixture of active factors isolated from colostrum which can be used in injectable form in those countries where health legislation prohibits the injection administration of heterologous immunoglobulins, it is possible
to add further steps to the method described above which allow obtaining a product deprived of the immunoglobulins. The product thus obtained has a high proliferative and regenerative activity because, with the same weight, the content of bioactive molecules is much higher.
According to another preferred aspect of the present invention, the process for preparing the mixture of colostrum-isolated active factors thus comprises a further purification step. In particular, such a step is carried out on the concentrated product before freezing and lyophilization, and is characterized by an IgG and IgA depletion step, which, in a preferred aspect of the invention, is carried out by means of affinity chromatography.
The colostrum purification step further comprises a step of IgM depletion, for example carried out by tangential filtration or cartridge filters.
METHOD: in a preferred aspect in the method of the present invention, affinity chromatography is used by means of:
1. CaptureSelect™ IgG-Fc (ms) Affinity Matrix (Thermo Fisher Scientific) or equivalent product. This system has been specifically designed to purify IgGs from different species (human, mouse, rat, rabbit, cow, horse, and sheep) by high affinity binding to the Fc region of IgG.
2. CaptureSelect™ Bovine IgA Affinity Matrix (Thermo Fisher Scientific) or equivalent product. This system has been specifically designed for the purification of bovine IgA from whey/colostrum. In CaptureSelect™ Bovine IgA Affinity Matrix, affinity ligands are created using proprietary technology based on single domain antibody fragments derived from camelids.
In a further preferred aspect of the present invention, the depletion of IgMs and immunoglobulin aggregates occurs by means of a tangential flow filtration step which uses membranes with a molecular cut-off of about 750 kDa - 0.02 pm, or by using cartridge filters with pores of the same size. More in detail, the pentamers of the M immunoglobulins, which have an average molecular weight of 800-900 kDa,
can be removed by tangential filtration, or cartridge filters, using membranes of appropriate size, while the other bioactive factors, with a lower molecular weight, can cross them without being retained. Different filtration methods were tested as well as a variety of filters and the best results in terms of IgM and aggregate depletion were obtained using cartridge filters with pores of 750 kDa - 0.02 pm, type ENTEGRIS Savana® PS cartridge filter 0.02 micron or equivalent product.
In a particularly preferred aspect, again in relation to the injective use of the product, a further purification step of the colostrum isolate mixture is necessary to deprive it of the possible presence of bacterial endotoxins (exogenous pyrogens).
In another particularly preferred aspect of the present invention, the removal of the endotoxins occurs by means of specific kits which use an affinity chromatography, such as the Bio-Rad® Proteus Endotoxin Removal kits or equivalent products. Alternatively, cartridge filters with directly incorporated positively charged nylon membrane can be used which, by strongly interacting with the negatively charged endotoxins, can adsorb consistent concentrations of the latter present in solution. The best results in terms of endotoxin removal were obtained using BEA Technologies POSINYL nylon 6.6 membrane cartridge filters or equivalent product.
In particular, all these further steps, necessary to purify the mixture for injective use, are carried out on the concentrated product before freezing and lyophilization.
The process was developed with a view to industrial application. All the process steps are transferable with a view to a large-scale purification process.
The mixture of bioactive molecules according to the present invention is much more effective in terms of proliferative and regenerative properties, with respect to the constituent factors tested alone or in small artificially-created groups. In fact, the growth factors and other bioactive molecules naturally present in the mixture do not act individually, but fully carry out the action thereof only if they are
in an appropriate combination, determined by nature during evolution, where all the components work in unison like the different instruments of an orchestra.
In fact, the interaction between growth factors, cytokines, stem cell stimulating factors, complement proteins, antibacterial and antiviral factors, and exclusively for oral administration, immunoglobulins, determines the initiation of a cascade of intracellular biochemical signals, followed by activation or inhibition of genes which control different cellular functions, and in particular cell proliferation and differentiation.
The close correlation between these bioactive molecules in inducing and modulating an effective regenerative response by injured tissues emerges from the above. When lacking entirely or in part, the action of some of these factors will have a null, partial or even harmful response.
Moreover, all the bioactive molecules described are genetically highly conserved in the different species and thus for humans it is possible to use, for reparative and regenerative purposes, the biological factors isolated from other species of mammals such as cattle, horses, camels, etc.
Exosomes
For the purposes of the present invention, the disclosed composition further comprises exosomes.
Exosomes represent the most extensively studied group among the three major subgroups of extracellular vesicles or EVs (exosomes, microvesicles, and apoptotic vesicles or ApoEVs). Among these, exosomes are the smallest vesicles, with a variable diameter of 50-150 nm and a cup-shaped morphology when viewed under an electron microscope.
The biogenesis process of the exosome includes several steps: 1 ) the inward folding of a section of the plasma membrane forms the early endosomes; 2) the first endosomes mature into late endosomes and then into multivesicular bodies (MVBs) characterized by intraluminal vesicles (ILVs); 3) once they reach maturity, these MVBs
are pulled by means of a network of microtubules and cytoskeletons towards the cell membrane where, by exocytosis, they will fuse therewith, releasing the ILVs into the extracellular environment which will become exosomes. This process of biogenesis and exosome formation is finely directed by the Endosomal Sorting Complexes Required for Transport (ESCRT) and accessory proteins.
Exosomes possess a specific molecular load which includes membrane proteins, cytosolic and nuclear proteins, extracellular matrix proteins, lipids, nucleic acids (mRNA, miRNA and DNA), growth factors, cytokines and enzymes, underlining the involvement thereof in the regulation, by means of different molecular mechanisms, of various biological functions.
The absorption of exosomes depends on the interactions between the proteins expressed on the surface thereof and the recipient cells. Several studies suggest that adhesion molecules on the surface of exosomes, such as tetraspanins, glycoproteins, and integrins, determine which cells can “accept” exosomes.
Exosomes are secreted by most cell types including immune cells (B cells, T cells), neuronal cells, epithelial cells, endothelial cells, embryonic cells, and mesenchymal stem cells (MSCs).
Exosomes have been detected in almost all body fluids, under both physiological and pathological conditions, including: urine, blood, serum, breast milk, amniotic fluid, cerebrospinal fluid, saliva, bile, and lymph.
Exosomes are also contained in colostrum, albeit in smaller quantities.
Due to the characteristics features thereof, exosomes can be considered "transporters", therefore it was thought to exploit this ability thereof to enter other cells to transfer useful "loads" including chemotherapeutic and immunomodulatory agents, with the ability to direct the release thereof towards a given target. For example, exosomes allow crossing the blood-brain barrier and bringing the molecules of interest to the desired CNS sites. However, inserting these useful "loads" inside the vesicles requires the manipulation of the vesicles themselves (exogenous loading in which the small
molecules are incorporated from the outside into the isolated EVs) or of the cells from which they come (endogenous loading in which the cell receives the necessary means to incorporate the small molecules in the EVs during the formation process).
Moreover, exosomes do not induce toxicity when injected repeatedly into mice and are well tolerated, thus the therapeutic use thereof is very promising.
There are several main methods for isolating exosomes: (1 ) ultracentrifugation, (2) size exclusion (e.g., ultrafiltration or chromatography), (3) immunological separation (e.g., antibody-bead capture), and (4) polymer-based precipitation.
The advantages in using each method vary based on the different origin of the exosomes, the degree of purity to be obtained (protein-exosome ratio, with the aim of minimizing the non-exosomal proteins which can be present in biofluids), yield or quality (preservation of the integrity and function of the exosomes).
For the purposes of the present patent, ultracentrifugation (UC) was used, recognized as the gold standard for the extraction and separation of exosomes and currently the most widely used isolation technique. UC allows separating and collecting the various components present in the original solution based on their differences in size and density, thus it is a suitable method for separating components with significant differences in the sedimentation coefficient from large sample volumes.
The UC method for isolating exosomes is mainly divided into two steps: first, a series of continuous centrifugations at medium-low speed to remove dead cells, cellular debris and large extracellular vesicles. The UC is then carried out at 100,000xg to separate the exosomes, followed by a series of washes in phosphate buffer (PBS), at the same UC speed, to remove impurities such as contaminatingproteins. This method is summarized in the flow diagram in Figure 4.
The inventors of the present patent application have surprisingly found that a highly concentrated exosome preparation is capable of enhancing the activity of the
mixture of biological molecules of the present invention.
Moreover, even more surprisingly, the inventors of the present patent application have found that a highly concentrated exosome preparation from colostrum, rather than from other sources, is capable of further enhancing the activity of the mixture of biological molecules of the present invention.
In a preferred aspect of the present invention, the exosomes are isolated and concentrated from bovine colostrum.
The inventors of the present patent application have thus surprisingly found that in order for the activity of the mixture of bioactive molecules to be enhanced in all fields of application, a preferable condition is that the exosomes are isolated and concentrated from colostrum, rather than from other sources, and that the latter are added to the mixture in a functional amount.
For the purposes of the present invention, a highly concentrated exosome preparation is a preparation containing an amount of exosomes >1x109/mL, preferably about 20-50x109/mL.
In the preparations of the invention, the two components can be present, independently of each other, in the following amounts (in distilled water or saline):
In a preferred aspect of the invention, the two components can be present, independently of each other, in the following amounts:
For example, the two components can be present, independently of each other, in the following amounts:
In accordance with a second object, the present patent application describes
formulations comprising a composition according to the invention.
In particular, such formulations are represented by: liquid solutions, lotions, foams, sprays, creams, salves, ointments, pastes, gels, membranes, clots, powders or other forms suitable for the medical use.
Such formulations can one or more additives, excipients and the other components necessary to obtain such formulations according to what is known in the art.
According to a preferred aspect of the present invention, the formulations described can optionally comprise one or more further effective components.
For example, one or more of the following can be included:
Hyaluronic acid
This can be present in various forms (esters, salts, hydrolysate, crosslinked, liposomal, etc.), among which sodium hyaluronate, the sodium salt thereof, in amounts between 5 and 50 mg/mL of distilled water or saline, preferably 25 mg/mL, is preferred.
Hyaluronic acid can further confer the ability to restore tissue tone and elasticity, as well as enhance and accelerate the re-epithelialization process.
Amino acids
In particular, glycine, L-proline, L-leucine and L-lysine are considered.
These can be present in a mixture, in amounts between 10 and 50 mg/mL of distilled water or saline, preferably 20 mg/mL.
Such amino acids can further confer the ability to stimulate collagen regeneration.
Hydroxyapatite
Hydroxyapatite (HA) is the main inorganic component of bone and makes up 60-70% of the calcified skeleton and 98% of tooth enamel. It is biocompatible, binds quickly to adjacent hard and soft tissues and has a strong osteoconductive capacity.
The clinical indications of hydroxyapatite concern the reconstruction of bone tissue and the lining of endo-osseous dental implants to promote osseointegration.
Hydroxyapatite is also used as a component of skin fillers.
In the case of bone reconstruction or dental implantology, the amounts used vary in relation to the surfaces to be reconstructed or covered.
In the case of fillers, 1 mL of gel is used containing from 20 to 50% of hydroxyapatite microspheres in 5 mL of distilled water or saline, preferably 40% of microspheres.
/3 -tricalcium phosphate
P-tricalcium phosphate is a polycrystalline bioceramic, with osteoconductive properties, which in contact with water releases hydroxyapatite crystals. It is used for the resolution of deep intraosseous periodontal defects (periodontal regeneration), for filling post-extraction bone cavities, for bone regeneration, etc. p- tricalcium phosphate has a progressive resorption, unlike what occurs with hydroxyapatite, with a release of calcium and phosphorus ions which, for example in the case of intraosseous periodontal defects, contribute to the neo-apposition of bone, cement, and periodontal ligament. It is used in the amount of 1 g/mL of distilled water or saline.
Platelet concentrates
According to a further aspect of the present invention, the compositions of the invention can further comprise Platelet Concentrates (PCs).
In a preferred aspect of the invention, the platelet concentrates are autologous.
Autologous Platelet Concentrates (APCs) are blood components obtained by centrifuging the patient’s blood in order to collect the most active components: platelets, fibrin and, in some cases, even leukocytes. The final product has a platelet concentration above the baseline level, therefore it has a higher number of bioactive molecules derived from the platelets themselves. The logic of the clinical
use of such preparations is based on the concept of exploiting the thus enriched content thereof to stimulate different biological functions, such as chemotaxis, angiogenesis, proliferation and differentiation, so as to promote the healing of hard and soft tissues.
The use of the term Platelet-Rich Plasma (PRP) really began with Marx in 1998 when he published a comparative clinical study in which the regenerative potential of PRP was demonstrated in a series of patients undergoing mandibular reconstruction. PRP has then been associated with the concept of platelet growth factors and the potential contribution thereof in inducing tissue healing.
According to the protocol for the production of PRP, the patient’s blood is collected in test tubes containing anticoagulants and processed by means of two centrifugation steps. Figure 5 diagrammatically shows the specific protocol. The PRP thus obtained can be applied to the site to be treated with a syringe or activated by thrombin and/or calcium chloride to trigger platelet activation and stimulate fibrin polymerization.
After the collection of blood in test tubes with anticoagulant, the first low- intensity centrifugation (soft spin) allows the separation of the blood into three distinct layers: red blood cells at the bottom, a cell plasma (Platelet-Poor Plasma or PPP) at the top and a whitish layer referred to as the buffy coat, located therebetween, containing the highest concentration of platelets and leukocytes. For the production of Pure-PRP (P-PRP), the PPP and the buffy coat surface layer are transferred to another test tube and centrifuged at high intensity (hard spin), then most of the PPP and leukocytes are discarded and the P-PRP can be collected.
To obtain the leukocyte rich PRP (L-PRP), the PPP, the entire buffy coat layer and some residual red blood cells are collected and transferred to another test tube to be centrifuged at high intensity (hard spin), then most of the PPP is discarded thus obtaining an L-PRP containing the buffy coat with most of the platelets and leukocytes, some residual red blood cells and PPP.
There are currently more than 20 different commercial systems for preparing PRP which can lead to products with different features, in particular regarding the composition and the cellular concentration rate with respect to the baseline. On average, a 5-8x concentration is obtained, although a ratio of up to 1 1 x has been reported with PRP.
In 2001 , a protocol was developed for producing a blood component referred to as Platelet-Rich Fibrin (PRF). In this case blood is collected in test tubes without anticoagulant and centrifuged at a moderate speed. Three layers are thus formed inside the test tube: red blood cells and acellular plasma are located, respectively, in the lower and upper part thereof, while the fibrin clot, positioned therebetween, forms the PRF (Figure 6). Since the PRF clot naturally forms inside the test tube, it has a robust fibrin matrix in which most of the platelets and leukocytes are trapped.
Injectable PRF (i-PRF) has also been recently developed, which can be obtained with a further, even more delicate, centrifugation step. It has a liquid form, is very rich in white blood cells and can be used for infiltration into tissues and joints.
In parallel with the introduction of PRP and PRF, another platelet concentrate protocol referred to as Plasma Rich in Growth Factors (PRGF) was suggested in 1999.
In short, blood is collected in test tubes with anticoagulant. After a low- intensity centrifugation, the red blood cells and the buffy coat layer are deposited on the bottom of the test tube and the plasma component on top of these. The latter is then manually separated into two fractions. The lower portion of about 2 mL, above the buffy coat, is PRGF, while the upper portion is Plasma Poor in Growth Factors (PPGFs) (Figure 7). PRGF can be applied as a liquid fraction in the target site or it can be pre-activated by adding 0.2 mL of 10% CaCIs to induce clot formation.
Although the form thereof can vary based on the different applications (liquid form, gel, membranes or fibrin clots, etc.), as well as preparation protocols, there is multiple clinical evidence on the effectiveness of platelet concentrates in different fields of medicine.
In accordance with a third object of the present invention, the compositions and formulations detailed above are described for the medical use.
In particular, these can be advantageously used for the treatment of those conditions requiring tissue repair and regeneration.
For the purposes of the present invention, the application is in humans and animals.
In a preferred aspect, the compositions and formulations described find use in oral and maxillofacial regenerative surgical procedures; in orthopedics and sports medicine; in dermatology and aesthetic medicine; in gynecology; in andrology; in plastic and reconstructive surgery; in ophthalmology; in otolaryngology; in neuroscience; in the repair of organ injuries (brain, heart, lungs, liver, kidneys, larynx, pharynx, esophagus, etc.).
The same uses described above can be applied to the veterinary field.
The invention is described here in greater detail in the following experimental part.
EXAMPLE 1
The composition can possibly be modified by adding the following
components (independently of each other):
EXAMPLE 2
The composition can possibly be modified by adding the following components (independently of each other):
EXAMPLE 3
Proliferative and regenerative effectiveness tests
In compliance with the principle of the 3Rs (Replacement, Reduction, Refinement) according to which the researcher should initially try, with the greatest
possible effort, to replace or substitute the animal model thereof with an alternative model, all the proliferative and regenerative effectiveness tests of the compositio ns of the invention have been performed in vitro on cell cultures or on a biological model of human hair follicle, considering that the techniques replacing animal experimentation in regenerative medicine are highly effective and predictive.
In a first step all the compositions, based on a mixture of bioactive molecules and a functional amount of exosomes, were evaluated on a limited but highly significant number of cell cultures. The proliferative and regenerative responses obtained using the mixture of bioactive molecules from colostrum plus highly concentrated exosomes obtained from various sources (body fluids, tissues and cells, including adult mesenchymal stem cells) were then compared.
From the tests carried out, a superior proliferative and regenerative activity of the compositions comprising the exosomes isolated and concentrated from colostrum, with respect to the other sources, emerges. The entire test panel was then performed in vitro with the composition based on a mixture of bioactive molecules from colostrum + highly concentrated exosomes from colostrum.
The in vitro tests performed, to evaluate the effectiveness of the composition of the present invention in regenerative medicine, were as follows:
1. MTT Assay for Cell Viability and Proliferation.
2. Wound Healing Assay.
3. Test on a biological model of human hair follicle.
1 . MTT Assay for Cell Viability and Proliferations
The MTT assay is used to measure the cellular metabolic activity, as an indicator of cell viability and proliferation. This colorimetric test is based on the reduction of a yellow tetrazolium salt (3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide or MTT) to purple formazan crystals by metabolically active cells. Vital cells contain NAD(P) H-dependent oxidoreductase enzymes which reduce MTT to formazan. The insoluble formazan crystals are dissolved
using a solubilization solution and the resulting colored solution is quantified by measuring absorbance at 500-600 nanometers using a multi-well spectrophotometer. The darker the solution, the greater the number of viable and metabolically active cells.
This colorimetric test can be used for multiple applications, such as quantifying cell growth and viability and measuring cell proliferation in response to growth factors, cytokines, and nutrients.
For the execution of the MTT test, the following cell lines were used, necessary to demonstrate the proliferative and regenerative activity of the preparations of the present invention on the different tissues and organs.
Primary Epidermal Keratinocytes; Normal, Human, Adult (HEKa): keratinocytes are the most common type of skin cell, as they form the structural component of the epidermis.
Keratinocytes are used for scientific applications including studying the behavior of growth factors, wound repair, skin cancers, response to UV radiation, psoriasis, eczema, viral infections, cell differentiation, and cosmetics research.
Primary keratinocyte cultures have been extensively tested and validated to form skin equivalents which mimic the architectural features and behavior of normal skin cells.
Primary Dermal Fibroblast; Normal, Human, Adult (HDFa): fibroblasts play an important role in maintaining the structural integrity of connective tissue and protein synthesis of the extracellular matrix such as collagens, glycosaminoglycans and glycoproteins, molecules also involved in wound healing.
Fibroblasts are mediators in inflammation, cancer, angiogenesis, and pathological tissue fibrosis and are frequently used in studies related to wound healing, tissue engineering, and regenerative applications.
Therefore, the cell line of primary skin fibroblasts finds application in several fields of research: in the response to pathogens, skin ageing, wound healing and skin diseases, including scleroderma.
Primary Aortic Endothelial Cells; Normal, Human (HAEC): Endothelial cells form the tunica intima, the thin layer of cells lining the inner surface of blood vessels. This surface acts as a selective filter to regulate the passage of gases, fluids, immune cells and various molecules. Primary endothelial cell cultures can be isolated from the umbilical vein, aorta, pulmonary and coronary arteries and dermal vascular tissue and represent useful tools for the study of angiogenesis, cancer therapy, wound healing, burn therapy, tissue engineering and regeneration.
Primary Corneal Epithelial Cells; Normal, Human (HCEpC): primary corneal epithelial cell cultures are used as a model for studying eye re- epithelialization following photorefractive keratectomy (PRK) or Laser-ASsisted In situ Keratomileusis (LASIK). They are also used to study the regeneration of damaged corneas.
Primary Vaginal Epithelial Cells; Normal, Human (HVEC): primary vaginal epithelial cell cultures can be used to study the cellular physiology of the reproductive tract, the cellular response to infectious agents, and in other areas of research, including the regenerative activity of the damaged or atrophic vaginal epithelium.
Primary Skeletal Muscle Cells; Normal, Human (HSkMC): primary skeletal muscle cells are isolated from skeletal muscle and, cultured in vitro, provide an ideal model for studying the biology and metabolism of striated muscle cells, diabetes, insulin receptors, muscle tissue repair, and myotube development.
Primary Articular Chondrocytes; Normal, Human (NHAC-kn): primary articular chondrocytes are isolated from tissue taken from the knee. Joint cartilage is characterized by high wear resistance and poor regenerative qualities.
Primary articular chondrocyte cultures provide an ideal model for studying osteoarthritis, cartilage failure, inhibition of protein synthesis, and chondrocyte apoptosis.
Primary Osteoblasts; Normal, Human (NHOst): primary osteoblast cultures provide a homogeneous and rapidly proliferating model for the in vitro study of normal osteoblast differentiation, the physiology thereof, and the effects of hormones, growth factors, and other cytokines on osteoblast function and differentiation in vivo.
Primary Schwann Cells; Normal, Human (HSwC): in the peripheral nervous system, Schwann cells represent the main type of glial cells. The proliferation in vitro thereof can be stimulated by various growth factors including PDGF, FGF, neuregulin and others. Primary Schwann cell cultures provide a relatively simple, well-defined, and accessible model for studying neuropathies, nerve regeneration, and several diseases of nervous system development.
Primary Epidermal Melanocytes; Normal, Human, Adult (HEMa): primary melanocytes are specialized epithelial cells found primarily in the epidermis, where they produce the pigment melanin which gives skin the color thereof while protecting it from ultraviolet (UV) damage. Melanocytes are thus useful in research into pigmentation disorders and for the development of effective therapies for such disorders, in particular malignant melanoma.
Primary epidermal melanocyte cultures are used for the in vitro study of wound healing and as a model for toxicity and irritability studies, for studies of skin response to UV radiation, psoriasis and other skin diseases and also in cosmetic research (e.g., skin lightening compounds and skin protective compounds).
Primary Hepatocytes; Normal, Human (HH): primary hepatocytes are often referred to as the “gold standard” for in vitro liver tests. Most of the specific cellular functions of hepatocytes can be maintained in culture, which makes them the best model for replicating liver function in vitro, e.g., metabolism, liver toxicity,
cell biology, and host-pathogen interactions. Primary hepatocyte cultures are thus a cost-effective and economical alternative to in vivo testing.
Primary Small Airway Epithelial Cells; Normal, Human (HSAEC): primary small airway epithelial cell cultures provide an ideal model for studying respiratory infections, asthma, chronic obstructive pulmonary disease, cystic fibrosis, inflammation response, harmful effects of smoking.
Primary Renal Epithelial Cells; Normal, Human (HREC): primary renal epithelial cell cultures provide an ideal model for research related to hypertension, diabetes, oncology, renal fibrosis, inflammation, autoimmune diseases, drug screening, and toxicology.
The results obtained with the MTT Assay for Cell Viability and Proliferations, using the cell cultures described above, are shown in Figure 8 to Figure 20.
2. Wound Healing Assay
The wound healing assay, also known as the "scratch test,", is a well- established two-dimensional (2D) technique which can be used to study wound healing in vitro. This method was one of the first to be developed for the study of cell migration and measures the speed with which cells, in a cell monolayer, migrate to fill a free space.
The technique reproduces the wound by creating an empty space in a confluent cellular monolayer and consists of four main steps which are described below (Figure 21 shows an overview thereof).
1. Cell culture preparation
The first step of the test is the culture of a confluent cell monolayer. This monolayer represents the in vivo condition of the tissue prior to injury, as an intact epithelium. Epithelial or endothelial cells are normally used to produce the monolayer. Human corneal epithelial cells (HCEpC) were used to evaluate the effectiveness of the composition of the present invention in regenerative medicine.
2. Scratch-making
After the cells have become confluent, the next step is to create a cell-free space in the monolayer, usually by means of mechanical scratching.
3. Data acquisition
Optical microscopy using phase contrast imaging can be used to observe cells migrating into the wound area.
Once the microscope is set up, a series of time-lapse images (snapshot method) can be acquired as the cells migrate into the cell free space. These time points are then collected at 24, 48 and 72 hours after the start of the experiment. The migration images can then be used to collect measurements or be evaluated visually.
4. Data analysis - "scratch" closing
Once the gap closure images have been acquired, different analysis methods can be used to quantify the cell migration rate. The method used for the purposes of the present patent calculates the change in wound area over time as a percentage of wound closure.
The results of the Wound Healing Assay, used to evaluate the effectiveness of the composition of the present invention in wound healing, are shown in Figure 22.
3. Test on a biological model of human hair follicle
Hair grows in cycles. Starting from the growth phase (anagen), the hair follicle and the shaft thereof go through the regression phase (catagen), the resting phase (telogen) and finally the exogenous phase (shedding phase). The hair follicles used in this test show a growth phase (anagen) of 15 days before moving on to the regression phase (catagen).
The biological model used for this test is represented by hair follicles microsectioned from fragments of human skin. The purpose of the test is to verify the growth of the hair follicle in the presence or not of the substances to be tested: the hair follicles were thus cultured in the William culture medium without the addition
of other compositions (negative control), in the same culture medium with the addition of the composition of the present patent, i.e., the composition based on a mixture of bioactive molecules from colostrum + highly concentrated exosomes from colostrum (positive control), or with the addition of the other two reference products (mixture alone or exosomes alone). Hair follicle growth was followed for 18 days, with measurements of the length thereof being taken at regular intervals.
The results of this test, used to evaluate the effectiveness of the composition of the present invention in the treatment of alopecia, are shown in Figure 23.
Proliferative and regenerative effectiveness
The in vitro tests performed to evaluate the proliferative and regenerative effectiveness of the composition of the present invention in the multiple medical uses are summarized below. a) To evaluate the effectiveness of the composition of the present invention in oral and maxillofacial regenerative surgical procedures, the in vitro tests were performed using the Wound Healing Assay and the MTT Assay with HEKa, HDFa, HAEC cells (healing of post-extractive alveoli, treating periodontal defects, endodontics and endodontic surgery) and with NHOst cells (dental implantology, elevation of the maxillary sinus). b) To evaluate the effectiveness of the composition of the present invention in orthopedics and sports medicine, the in vitro tests were performed using the MTT Assay with NHAC-kn cells (treating osteoarthritis, treating degenerative disease of the intervertebral discs); with NHOst, HAEC cells (managing fractures and nonunions); with HDFa cells (treating tendon injuries, enhancing the post-surgery healing process of tendons and ligaments) and with HSkMC cells (treating muscle injuries). c) To evaluate the effectiveness of the composition of the present invention in dermatology and aesthetic medicine, the in vitro tests were performed using the MTT Assay with HEKa, HDFa cells (skin rejuvenation, treating acne, treating
stretch marks, treating melasma, treating periorbital hyperpigmentation); with HEMa cells (treating vitiligo), as well as with the "Test on a biological model of human hair follicle" for treating alopecia. d) To evaluate the effectiveness of the composition of the present invention in gynecology, the in vitro tests were performed using the Wound Healing Assay and the MTT Assay with HVEC, HDFa cells (treating skin injuries, healing surgical wounds, treating vaginal atrophy and vaginal dryness, treating vulvar dystrophy, aesthetic gynecology in vaginal rejuvenation). e) To evaluate the effectiveness of the composition of the present invention in andrology, the in vitro tests were performed using the MTT Assay with HAEC cells (treating erectile dysfunction). f) To evaluate the effectiveness of the composition of the present invention in plastic and reconstructive surgery, the in vitro tests were performed using the Wound Healing Assay and the MTT Assay with HEKa, HDFa, HAEC cells (treating burns, treating wounds and skin ulcers, treating keloids and hypertrophic scars post-excision); and with HDFa, HSkMC cells (breast reconstruction). g) To evaluate the effectiveness of the composition of the present invention in ophthalmology, the in vitro tests were performed using the Wound Healing Assay and the MTT Assay with HCEpC cells (treating persistent corneal epithelial defects, treating corneal abrasions and ulcers, corneal surgery, treating dry eye syndrome, treating ocular surface syndrome after Laser-ASsisted In situ Keratomileusis - LASIK or after photorefractive keratectomy - PRK). h) To evaluate the effectiveness of the composition of the present invention in otolaryngology, the in vitro tests were performed using the Wound Healing Assay and the MTT Assay with HEKa, HDFa, HAEC cells (treating atrophic rhinitis, treating tympanic membrane perforation). i) To evaluate the effectiveness of the composition of the present invention in neuroscience, the in vitro tests were performed using the MTT Assay with HSwC
cells (treating traumatic brain injuries, treating traumatic spinal cord injuries, treating neurodegenerative and autoimmune diseases of the CNS, treating peripheral nerve injuries).
I) To evaluate the effectiveness of the composition of the present invention in the repair of organ injuries (brain, heart, lungs, liver, kidneys, larynx, pharynx, esophagus, etc.) resulting from ischemia, infectious diseases, degenerative diseases, fibrosis, radiotherapy treatments, traumatic injuries, burns, etc., the in vitro tests were performed using the MTT Assay with HH, HSAEC, HREC cells, in addition to what has already been reported for the individual tissues forming the different organs.
The in vitro tests described above to evaluate the proliferative and regenerative effectiveness of certain substances on human cells/tissues/organs are comparable to those used for animals, thus all tests performed to evaluate the effectiveness of the composition of the present invention in human medicine and surgery are also to be considered valid in veterinary medicine and surgery.
In all the in vitro tests performed, the results obtained with the composition of the present patent application (i.e., the composition based on the mixture of bioactive molecules + highly concentrated exosomes) were far superior to the results obtained with all the other products tested or with the reference sample (control), both in terms of proliferative and regenerative effectiveness (MTT Assay, Figures 8 to 20; Wound Healing Assay, Figure 22; Test on a biological model of human hair follicle, Figure 23).
Student's t (p < 0.05 significant values; p < 0.01 highly significant values) was used to compare the results obtained with the in vitro tests. The statistical calculations show that the difference in the post-treatment averages between the composition of the present patent, the other products tested and the reference sample (control) is significant and, in almost all cases, highly significant.
The above description further demonstrates the greater effectiveness of the
composition based on a mixture of bioactive molecules plus highly concentrated exosomes, of the present invention, with respect to the other tested products (mixture of bioactive molecules alone, highly concentrated exosomes alone). Such a greater effectiveness is clearly higher than what could be expected from the mere arithmetic sum of the results obtained by the individual products.
In particular, it has been surprisingly found that by combining the exosomes with the bioactive molecules of the mixture, the correct growth and regeneration of tissues is modulated and essential nourishment is provided to cells in the growth and proliferation phase.
From the above description of the present invention, the advantages given by the suggested formulations will be immediately apparent.
In particular, it has been observed that the effectiveness of these formulations is closely linked to the synergy of the two components (mixture of bioactive molecules and highly concentrated exosomes) and the functional interdependence thereof; in fact, the bioactive molecules of the mixture act synergistically with the exosomes and the content thereof (membrane proteins, cytosolic and nuclear proteins, extracellular matrix proteins, lipids, mRNA, miRNA and DNA nucleic acids, growth factors, cytokines and enzymes) exponentially increasing the activity thereof.
Moreover, it appears that this interaction results in an increase in the halflife of the bioactive molecules of the mixture, which would otherwise be very short, and/or an increase in the diffusion rate thereof in the intercellular spaces. Both of these factors would contribute to resolving the defect of transport of the bioactive molecules of the mixture through the healthy tissues to those areas where the action thereof would be required.
By virtue of this synergy, the bioactive molecules of the mixture can act, promoting the reparative process; this occurs by stimulating neoangiogenesis, even
in poorly vascularized tissues, and the regeneration of damaged tissues which are replaced with cells of the same type, thus avoiding the replacement thereof with connective tissue (fibrosis). The application of the composition of the present invention thus prevents the onset of hypertrophic or keloid scars, which represent one of the most common wound healing disorders.
It should be noted that for the exosomes of the present invention, the only clearly demonstrated function, i.e., the function of transporters, is excluded, since for the execution of the tests described in the experimental part no manipulation of the exosomes was carried out in order to incorporate the bioactive molecules therein.
It was also observed that the exosomes of the present invention remain in a living and vital state even up to 6-8 months and even in the absence of lyophilization, thus demonstrating a high and unexpected stability.
In order to meet contingent and specific needs, those skilled in the art may make adaptations and modifications to the invention described above, and may replace components with other similar ones, without however departing from the scope of the claims set forth below.
For example, one or more excipients can be modified according to the pharmaceutical form selected.
Claims
CLAIMS A composition comprising a mixture of bioactive molecules and exosomes, wherein said mixture is preferably isolated from colostrum. A composition according to the preceding claim, comprising said mixture of bioactive molecules in an amount of about 10-600 mg/mL, preferably about 25-400 mg/mL, still more preferably 50-200 mg/mL, in distilled water or saline. A composition according to claim 1 or 2, comprising exosomes in an amount of at least 1x109/mL. A composition according to the preceding claim, wherein said exosomes are included in an amount of about 5-200x109/mL, preferably about 10- 100x109/mL and more preferably about 20-50x109/mL. A composition according to any one of the preceding claims, wherein said colostrum-isolated, biological factor-rich mixture is isolated from human or animal colostrum, preferably animal colostrum, more preferably from dairy cows, even more preferably of the Jersey breed. A composition according to any one of the preceding claims, wherein said colostrum-isolated, biological factor-rich mixture is isolated from colostrum collected before the newborn has had the opportunity to nurse and empty the udder of the first fraction produced, regardless of the time elapsed since the moment of the birth, preferably at the second or third birth. A composition according to any one of the preceding claims, wherein said colostrum-isolated, biological factor-rich mixture comprises growth factors, cytokines, stem cell stimulating factors, complement proteins, antibacterial and antiviral factors, immunoglobulins. A composition according to any one of the preceding claims, wherein said colostrum-isolated, biological factor-rich mixture is free of IgG, IgA, IgM immunoglobulins at least for 90% of the total, preferably for at least 95% of
the total, even more preferably for at least 99% of the total and much more preferably it is totally free. A composition according to any one of the preceding claims, wherein the exosomes are isolated and concentrated from human or animal cells, tissues or body fluids. A composition according to the preceding claim, wherein said exosomes are isolated from colostrum, even more preferably from dairy cow colostrum, much more preferably from colostrum collected before the newborn has had the opportunity to nurse and empty the udder of the first fraction produced, regardless of the time elapsed since the moment of the birth, preferably at the second or third birth. A composition according to any one of the preceding claims, wherein the exosomes contain membrane proteins, cytosolic and nuclear proteins, extracellular matrix proteins, lipids, nucleic acids (mRNA, miRNA and DNA), growth factors, cytokines and enzymes. A composition according to any one of the preceding claims, further comprising one or more of the following compounds: hyaluronic acid, platelet concentrates, amino acids, hydroxyapatite, p-tricalcium phosphate. A composition according to any one of the preceding claims, which is a topical or injection composition. A topical composition according to any one of the preceding claims, having the following formulation:
and further comprising, or as an alternative to hyaluronic acid, one or more of the following ingredients:
An injection composition according to any one of claims 1 to 13, having the following formulation:
and further comprising, or as an alternative to hyaluronic acid, one or more of the following ingredients:
A topical or injection formulation comprising the composition according to any one of the preceding claims, in a form selected from the group
comprising: liquid solutions, lotions, foams, sprays, creams, salves, ointments, pastes, gels, membranes, clots, powders or other pharmaceutically suitable forms. A composition or formulation according to any one of the preceding claims for the medical use, in humans or animals. A composition or formulation according to any one of the preceding claims for the medical use in the treatment of diseases requiring tissue repair and regeneration. A composition or formulation according to claim 17 or 18, for the medical use in oral and maxillofacial regenerative surgical procedures, comprising procedures selected from the group comprising: healing of post-extraction alveoli, treatment of periodontal defects, endodontics and endodontic surgery, dental implantology, maxillary sinus lift. A composition or formulation according to claim 17 or 18, for the medical use in orthopedics and sports medicine: in the treatment of osteoarthritis, in the treatment of degenerative disease of the intervertebral discs, in the treatment of tendon injuries, enhancing the post-surgery healing process of tendons and ligaments, in the treatment of muscle injuries, managing fractures and nonunions. A composition or formulation according to claim 17 or 18, for the medical use in dermatology and aesthetic medicine: in skin rejuvenation, in the treatment of acne, in the treatment of stretch marks, in the treatment of melasma, in the treatment of periorbital hyperpigmentation, in the treatment of vitiligo, in the treatment of alopecia. A composition or formulation according to claim 17 or 18, for the medical use in gynecology: in in the treatment of skin injuries and healing surgical wounds, in the treatment of vaginal atrophy and vaginal dryness, in the treatment of vulvar dystrophy, aesthetic gynecology in vaginal rejuvenation.
A composition or formulation according to claim 17 or 18, for the medical use in andrology: in the treatment of erectile dysfunction. A composition or formulation according to claim 17 or 18, for the medical use in plastic and reconstructive surgery: in the treatment of burns, in the treatment of wounds, in the treatment of skin ulcers, in the treatment of keloids and hypertrophic scars post-excision, breast reconstruction. A composition or formulation according to claim 17 or 18, for the medical use in ophthalmology: in treating persistent corneal epithelial defects, treating corneal abrasions and ulcers, corneal surgery, treating dry eye syndrome, treating the ocular surface syndrome after Laser-Assisted In situ Keratomileusis (LASIK) or after Photorefractive Keratectomy (PRK). A composition or formulation according to claim 17 or 18, for the medical use in otolaryngology: in the treatment of atrophic rhinitis, in the treatment of tympanic membrane perforation. A composition or formulation according to claim 17 or 18, for the medical use in neuroscience: in the treatment of traumatic brain injuries, in the treatment of traumatic spinal cord injuries, in the treatment of neurodegenerative and autoimmune diseases of the CNS, in the treatment of peripheral nerve injuries. A composition or formulation according to claim 17 or 18, for the medical use in the repair of organ injuries (brain, heart, lungs, liver, kidneys, larynx, pharynx, esophagus, etc.) resulting from: ischemia, infectious diseases, degenerative diseases, fibrosis, radiotherapy treatments, traumatic injuries, burns, etc.
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
IT202200024306 | 2022-11-25 | ||
IT102022000024306 | 2022-11-25 |
Publications (1)
Publication Number | Publication Date |
---|---|
WO2024110937A1 true WO2024110937A1 (en) | 2024-05-30 |
Family
ID=85172648
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
PCT/IB2023/061927 WO2024110937A1 (en) | 2022-11-25 | 2023-11-27 | Compositions based on a mixture of bioactive molecules and exosomes for use in the treatment of conditions requiring tissue repair and regeneration |
Country Status (1)
Country | Link |
---|---|
WO (1) | WO2024110937A1 (en) |
Citations (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US10166259B1 (en) * | 2016-03-15 | 2019-01-01 | 3P Biotechnologies, Inc. | Isolation of exosomes from colostrum powder and exosomal drug formulations using the same |
WO2021107706A1 (en) * | 2019-11-28 | 2021-06-03 | 한국과학기술연구원 | Novel use of milk exosomes |
KR102262765B1 (en) * | 2019-10-15 | 2021-06-08 | 전주대학교 산학협력단 | Composition for reinforcing bone density or treating osteoporosis comprising a ranch raw milk derived exosome |
US20210228634A1 (en) * | 2018-06-06 | 2021-07-29 | Board Of Regents Of The University Of Nebraska | Extracellular vesicles and methods of using |
WO2022079684A1 (en) * | 2020-10-15 | 2022-04-21 | Biomedical Research S.R.L. | Composition based on autologous platelet concentrates and a colostrum isolate mixture of biological factors for use in the treatment of conditions requiring tissue repair and regeneration |
-
2023
- 2023-11-27 WO PCT/IB2023/061927 patent/WO2024110937A1/en unknown
Patent Citations (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US10166259B1 (en) * | 2016-03-15 | 2019-01-01 | 3P Biotechnologies, Inc. | Isolation of exosomes from colostrum powder and exosomal drug formulations using the same |
US20210228634A1 (en) * | 2018-06-06 | 2021-07-29 | Board Of Regents Of The University Of Nebraska | Extracellular vesicles and methods of using |
KR102262765B1 (en) * | 2019-10-15 | 2021-06-08 | 전주대학교 산학협력단 | Composition for reinforcing bone density or treating osteoporosis comprising a ranch raw milk derived exosome |
WO2021107706A1 (en) * | 2019-11-28 | 2021-06-03 | 한국과학기술연구원 | Novel use of milk exosomes |
WO2022079684A1 (en) * | 2020-10-15 | 2022-04-21 | Biomedical Research S.R.L. | Composition based on autologous platelet concentrates and a colostrum isolate mixture of biological factors for use in the treatment of conditions requiring tissue repair and regeneration |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
JP6826135B2 (en) | Composition for treating skin wounds containing exosomes derived from thrombin-treated stem cells | |
EP3541398B1 (en) | A method for preparing a growth factors containing platelet releasate, and its uses | |
KR20180127538A (en) | Process, tube and device for the preparation of wound healant composition | |
US9402881B2 (en) | Tissue regeneration and wound treatment methods with platelet derived compositions | |
US20090081181A1 (en) | Isolation of growth and differentiating factors from colostrum | |
da Fonseca et al. | Human platelet lysate–A potent (and overlooked) orthobiologic | |
EP2480239A1 (en) | Composition comprising a haematic component and its use for the treatment of degenerative joint disease | |
RU2645082C2 (en) | COMBINATION OF GROWTH FACTORS, CYTOKINES, ANTIBACTERIAL/ANTIVIRAL FACTORS, STEM CELL FACTORS, C3a/C4a COMPLEMENT PROTEINS AND CHEMOTAXIC FACTORS | |
EP3881858A1 (en) | Heat-treated platelet-derived growth factor extract for use in a method of preventing or treating a tissue defect | |
WO2017082441A1 (en) | Veterinary medicinal composition for treatment of wound of animal comprising activated platelet-rich plasma as active ingredient | |
US20230414663A1 (en) | Composition based on autologous platelet concentrates and a colostrum isolate mixture of biological factors for use in the treatment of conditions requiring tissue repair and regeneration | |
WO2024110937A1 (en) | Compositions based on a mixture of bioactive molecules and exosomes for use in the treatment of conditions requiring tissue repair and regeneration | |
KR20150061806A (en) | Veterinary composition for wound healing of animal containing activated platelet rich plasma | |
CA3076046A1 (en) | Method for producing enhanced anti-inflammatory/anti-catabolic agents from autologous physiological fluid with shortened incubation time | |
Alsousou et al. | Therapeutic platelet-rich plasma in wound healing | |
Shome et al. | Recent advances in platelet-rich plasma and its derivatives: therapeutic agents for tissue engineering and regenerative medicine | |
RU2795884C2 (en) | Method for obtaining platelet releasate containing growth factors | |
Lunjanasatienchai et al. | Effect of human exfoliated deciduous teeth stem cell-derived conditioned medium on human periodontal ligament stem cell proliferation and osteogenic differentiation | |
Bharti et al. | Therapeutic applications of canine platelets and their derivatives: a narrative review | |
Brundo et al. | How to enhance the effects of Platelet Rich Plasma on tissue regeneration: AMPLEX PLUS technology, a new strategy. | |
Uniejewska | Possibilities of using platelet-rich plasma in medicine, cosmetology and veterinary medicine. Review article. | |
Goczyńska et al. | Innovative applications of platelet derivatives in light of information presented during the 2022 virtual congress of the International Society of Blood Transfusion (ISBT); selected issues | |
PT1365795E (en) | Use of the g-csf as additional treatment in the reconstruction of connective tissue | |
WO2005097170A1 (en) | Angiogenesis promoter and angiogenic therapy | |
KR20240096758A (en) | method for preparing a growth factors containing platelet releasate |