CN117503800A - Gastric mucosa epithelial cell extract and preparation method and application thereof - Google Patents
Gastric mucosa epithelial cell extract and preparation method and application thereof Download PDFInfo
- Publication number
- CN117503800A CN117503800A CN202410010945.5A CN202410010945A CN117503800A CN 117503800 A CN117503800 A CN 117503800A CN 202410010945 A CN202410010945 A CN 202410010945A CN 117503800 A CN117503800 A CN 117503800A
- Authority
- CN
- China
- Prior art keywords
- epithelial cell
- extract
- gastric
- gastric mucosal
- cell extract
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
- 239000000284 extract Substances 0.000 title claims abstract description 56
- 210000002919 epithelial cell Anatomy 0.000 title claims abstract description 48
- 210000001156 gastric mucosa Anatomy 0.000 title claims abstract description 43
- 238000002360 preparation method Methods 0.000 title claims abstract description 13
- 238000000605 extraction Methods 0.000 title description 3
- 230000002496 gastric effect Effects 0.000 claims abstract description 48
- 230000008439 repair process Effects 0.000 claims abstract description 29
- 230000001737 promoting effect Effects 0.000 claims abstract description 20
- 230000000638 stimulation Effects 0.000 claims abstract description 10
- 210000004027 cell Anatomy 0.000 claims description 41
- 239000007788 liquid Substances 0.000 claims description 18
- 238000000034 method Methods 0.000 claims description 18
- 230000006698 induction Effects 0.000 claims description 15
- 239000008267 milk Substances 0.000 claims description 14
- 210000004080 milk Anatomy 0.000 claims description 14
- 235000013336 milk Nutrition 0.000 claims description 14
- 108090000284 Pepsin A Proteins 0.000 claims description 12
- 102000057297 Pepsin A Human genes 0.000 claims description 12
- 229940111202 pepsin Drugs 0.000 claims description 12
- 108090000746 Chymosin Proteins 0.000 claims description 11
- 229940080701 chymosin Drugs 0.000 claims description 11
- 239000001963 growth medium Substances 0.000 claims description 11
- GNOLWGAJQVLBSM-UHFFFAOYSA-N n,n,5,7-tetramethyl-1,2,3,4-tetrahydronaphthalen-1-amine Chemical compound C1=C(C)C=C2C(N(C)C)CCCC2=C1C GNOLWGAJQVLBSM-UHFFFAOYSA-N 0.000 claims description 11
- 239000006228 supernatant Substances 0.000 claims description 11
- 229940002010 banana extract Drugs 0.000 claims description 10
- 238000005406 washing Methods 0.000 claims description 9
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 claims description 8
- 239000002609 medium Substances 0.000 claims description 8
- 108091003079 Bovine Serum Albumin Proteins 0.000 claims description 7
- 239000008213 purified water Substances 0.000 claims description 7
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 7
- 239000012091 fetal bovine serum Substances 0.000 claims description 6
- 238000001914 filtration Methods 0.000 claims description 6
- 102000006747 Transforming Growth Factor alpha Human genes 0.000 claims description 5
- 101800004564 Transforming growth factor alpha Proteins 0.000 claims description 5
- 229940104640 chinese yam extract Drugs 0.000 claims description 5
- 230000001954 sterilising effect Effects 0.000 claims description 5
- 235000004879 dioscorea Nutrition 0.000 claims description 4
- 239000000047 product Substances 0.000 claims description 4
- 206010021143 Hypoxia Diseases 0.000 claims description 3
- 238000005119 centrifugation Methods 0.000 claims description 3
- 238000012258 culturing Methods 0.000 claims description 3
- 239000012634 fragment Substances 0.000 claims description 3
- 229910052760 oxygen Inorganic materials 0.000 claims description 3
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims description 2
- 239000012528 membrane Substances 0.000 claims description 2
- 239000000203 mixture Substances 0.000 claims description 2
- 239000011148 porous material Substances 0.000 claims description 2
- 238000004659 sterilization and disinfection Methods 0.000 claims description 2
- 238000012546 transfer Methods 0.000 claims description 2
- 210000004400 mucous membrane Anatomy 0.000 abstract description 9
- 206010061218 Inflammation Diseases 0.000 abstract description 5
- 230000004054 inflammatory process Effects 0.000 abstract description 5
- 210000001519 tissue Anatomy 0.000 abstract description 4
- 230000008338 local blood flow Effects 0.000 abstract description 2
- 230000000694 effects Effects 0.000 description 12
- 239000003814 drug Substances 0.000 description 11
- 208000002193 Pain Diseases 0.000 description 9
- 230000000052 comparative effect Effects 0.000 description 8
- 230000028327 secretion Effects 0.000 description 8
- 208000024891 symptom Diseases 0.000 description 8
- 102100026019 Interleukin-6 Human genes 0.000 description 7
- 210000002784 stomach Anatomy 0.000 description 7
- 208000007882 Gastritis Diseases 0.000 description 6
- 206010061297 Mucosal erosion Diseases 0.000 description 6
- 210000004877 mucosa Anatomy 0.000 description 6
- 230000008901 benefit Effects 0.000 description 5
- 230000006378 damage Effects 0.000 description 5
- 230000002757 inflammatory effect Effects 0.000 description 5
- 239000000463 material Substances 0.000 description 5
- 208000025865 Ulcer Diseases 0.000 description 4
- 206010000059 abdominal discomfort Diseases 0.000 description 4
- 239000002253 acid Substances 0.000 description 4
- 229940079593 drug Drugs 0.000 description 4
- 230000002829 reductive effect Effects 0.000 description 4
- 239000000243 solution Substances 0.000 description 4
- 238000012360 testing method Methods 0.000 description 4
- 231100000397 ulcer Toxicity 0.000 description 4
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 description 3
- 206010067715 Gastrointestinal sounds abnormal Diseases 0.000 description 3
- 229910052791 calcium Inorganic materials 0.000 description 3
- 239000011575 calcium Substances 0.000 description 3
- 208000023652 chronic gastritis Diseases 0.000 description 3
- 238000003745 diagnosis Methods 0.000 description 3
- 210000004185 liver Anatomy 0.000 description 3
- 230000007774 longterm Effects 0.000 description 3
- 206010067484 Adverse reaction Diseases 0.000 description 2
- 102000029816 Collagenase Human genes 0.000 description 2
- 108060005980 Collagenase Proteins 0.000 description 2
- 206010067171 Regurgitation Diseases 0.000 description 2
- 208000027418 Wounds and injury Diseases 0.000 description 2
- 210000001015 abdomen Anatomy 0.000 description 2
- 230000003187 abdominal effect Effects 0.000 description 2
- 238000010521 absorption reaction Methods 0.000 description 2
- 230000009471 action Effects 0.000 description 2
- 230000006838 adverse reaction Effects 0.000 description 2
- 230000036528 appetite Effects 0.000 description 2
- 235000019789 appetite Nutrition 0.000 description 2
- 244000309466 calf Species 0.000 description 2
- 230000004663 cell proliferation Effects 0.000 description 2
- 239000006285 cell suspension Substances 0.000 description 2
- 229960002424 collagenase Drugs 0.000 description 2
- 230000029087 digestion Effects 0.000 description 2
- 230000000857 drug effect Effects 0.000 description 2
- 230000003628 erosive effect Effects 0.000 description 2
- 235000013305 food Nutrition 0.000 description 2
- 210000001035 gastrointestinal tract Anatomy 0.000 description 2
- 241000411851 herbal medicine Species 0.000 description 2
- 230000006872 improvement Effects 0.000 description 2
- 208000014674 injury Diseases 0.000 description 2
- 210000003734 kidney Anatomy 0.000 description 2
- 235000012054 meals Nutrition 0.000 description 2
- 230000004060 metabolic process Effects 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 230000001681 protective effect Effects 0.000 description 2
- 102000004169 proteins and genes Human genes 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 230000009467 reduction Effects 0.000 description 2
- 230000008929 regeneration Effects 0.000 description 2
- 238000011069 regeneration method Methods 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- QZAYGJVTTNCVMB-UHFFFAOYSA-N serotonin Chemical compound C1=C(O)C=C2C(CCN)=CNC2=C1 QZAYGJVTTNCVMB-UHFFFAOYSA-N 0.000 description 2
- 210000002966 serum Anatomy 0.000 description 2
- 208000018556 stomach disease Diseases 0.000 description 2
- UCSJYZPVAKXKNQ-HZYVHMACSA-N streptomycin Chemical compound CN[C@H]1[C@H](O)[C@@H](O)[C@H](CO)O[C@H]1O[C@@H]1[C@](C=O)(O)[C@H](C)O[C@H]1O[C@@H]1[C@@H](NC(N)=N)[C@H](O)[C@@H](NC(N)=N)[C@H](O)[C@H]1O UCSJYZPVAKXKNQ-HZYVHMACSA-N 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 229940126680 traditional chinese medicines Drugs 0.000 description 2
- 229940126673 western medicines Drugs 0.000 description 2
- 210000002417 xiphoid bone Anatomy 0.000 description 2
- LOGFVTREOLYCPF-KXNHARMFSA-N (2s,3r)-2-[[(2r)-1-[(2s)-2,6-diaminohexanoyl]pyrrolidine-2-carbonyl]amino]-3-hydroxybutanoic acid Chemical compound C[C@@H](O)[C@@H](C(O)=O)NC(=O)[C@H]1CCCN1C(=O)[C@@H](N)CCCCN LOGFVTREOLYCPF-KXNHARMFSA-N 0.000 description 1
- IIZPXYDJLKNOIY-JXPKJXOSSA-N 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine Chemical compound CCCCCCCCCCCCCCCC(=O)OC[C@H](COP([O-])(=O)OCC[N+](C)(C)C)OC(=O)CCC\C=C/C\C=C/C\C=C/C\C=C/CCCCC IIZPXYDJLKNOIY-JXPKJXOSSA-N 0.000 description 1
- 208000004998 Abdominal Pain Diseases 0.000 description 1
- 206010000060 Abdominal distension Diseases 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- 206010004716 Binge eating Diseases 0.000 description 1
- 208000032841 Bulimia Diseases 0.000 description 1
- 102000004127 Cytokines Human genes 0.000 description 1
- 108090000695 Cytokines Proteins 0.000 description 1
- 206010061819 Disease recurrence Diseases 0.000 description 1
- 206010013710 Drug interaction Diseases 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 206010015137 Eructation Diseases 0.000 description 1
- 206010061164 Gastric mucosal lesion Diseases 0.000 description 1
- 206010017865 Gastritis erosive Diseases 0.000 description 1
- 239000012981 Hank's balanced salt solution Substances 0.000 description 1
- 206010019375 Helicobacter infections Diseases 0.000 description 1
- 102000008070 Interferon-gamma Human genes 0.000 description 1
- 108010074328 Interferon-gamma Proteins 0.000 description 1
- 102000003777 Interleukin-1 beta Human genes 0.000 description 1
- 108090000193 Interleukin-1 beta Proteins 0.000 description 1
- 102000004889 Interleukin-6 Human genes 0.000 description 1
- 108090001005 Interleukin-6 Proteins 0.000 description 1
- 102100033420 Keratin, type I cytoskeletal 19 Human genes 0.000 description 1
- 102100023972 Keratin, type II cytoskeletal 8 Human genes 0.000 description 1
- 102000018329 Keratin-18 Human genes 0.000 description 1
- 108010066327 Keratin-18 Proteins 0.000 description 1
- 108010066302 Keratin-19 Proteins 0.000 description 1
- 108010070511 Keratin-8 Proteins 0.000 description 1
- 244000207740 Lemna minor Species 0.000 description 1
- 235000006439 Lemna minor Nutrition 0.000 description 1
- 241000234295 Musa Species 0.000 description 1
- 235000018290 Musa x paradisiaca Nutrition 0.000 description 1
- 229930182555 Penicillin Natural products 0.000 description 1
- JGSARLDLIJGVTE-MBNYWOFBSA-N Penicillin G Chemical compound N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)CC1=CC=CC=C1 JGSARLDLIJGVTE-MBNYWOFBSA-N 0.000 description 1
- 241000219000 Populus Species 0.000 description 1
- 235000001855 Portulaca oleracea Nutrition 0.000 description 1
- 208000032023 Signs and Symptoms Diseases 0.000 description 1
- 208000005718 Stomach Neoplasms Diseases 0.000 description 1
- 208000007107 Stomach Ulcer Diseases 0.000 description 1
- 102000009618 Transforming Growth Factors Human genes 0.000 description 1
- 108010009583 Transforming Growth Factors Proteins 0.000 description 1
- 102000004142 Trypsin Human genes 0.000 description 1
- 108090000631 Trypsin Proteins 0.000 description 1
- 108060008682 Tumor Necrosis Factor Proteins 0.000 description 1
- 241000700605 Viruses Species 0.000 description 1
- 206010000210 abortion Diseases 0.000 description 1
- 231100000176 abortion Toxicity 0.000 description 1
- 230000003213 activating effect Effects 0.000 description 1
- 239000004480 active ingredient Substances 0.000 description 1
- 230000001464 adherent effect Effects 0.000 description 1
- 230000003110 anti-inflammatory effect Effects 0.000 description 1
- 230000006907 apoptotic process Effects 0.000 description 1
- 230000004596 appetite loss Effects 0.000 description 1
- 230000001174 ascending effect Effects 0.000 description 1
- 230000004888 barrier function Effects 0.000 description 1
- 239000007640 basal medium Substances 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000002457 bidirectional effect Effects 0.000 description 1
- 208000014679 binge eating disease Diseases 0.000 description 1
- 230000033228 biological regulation Effects 0.000 description 1
- 230000000740 bleeding effect Effects 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 230000017531 blood circulation Effects 0.000 description 1
- 235000001465 calcium Nutrition 0.000 description 1
- 230000015556 catabolic process Effects 0.000 description 1
- 239000006143 cell culture medium Substances 0.000 description 1
- 208000015864 chronic erosive gastritis Diseases 0.000 description 1
- 238000003759 clinical diagnosis Methods 0.000 description 1
- 235000009508 confectionery Nutrition 0.000 description 1
- 238000000354 decomposition reaction Methods 0.000 description 1
- 230000007123 defense Effects 0.000 description 1
- 238000006731 degradation reaction Methods 0.000 description 1
- 230000037213 diet Effects 0.000 description 1
- 235000005911 diet Nutrition 0.000 description 1
- 208000010643 digestive system disease Diseases 0.000 description 1
- XEYBRNLFEZDVAW-ARSRFYASSA-N dinoprostone Chemical compound CCCCC[C@H](O)\C=C\[C@H]1[C@H](O)CC(=O)[C@@H]1C\C=C/CCCC(O)=O XEYBRNLFEZDVAW-ARSRFYASSA-N 0.000 description 1
- 229960002986 dinoprostone Drugs 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 229940088598 enzyme Drugs 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 210000001808 exosome Anatomy 0.000 description 1
- 239000012894 fetal calf serum Substances 0.000 description 1
- 230000006870 function Effects 0.000 description 1
- 230000027119 gastric acid secretion Effects 0.000 description 1
- 206010017758 gastric cancer Diseases 0.000 description 1
- 210000004051 gastric juice Anatomy 0.000 description 1
- 201000005917 gastric ulcer Diseases 0.000 description 1
- 230000005176 gastrointestinal motility Effects 0.000 description 1
- 210000004907 gland Anatomy 0.000 description 1
- 230000012010 growth Effects 0.000 description 1
- 230000035876 healing Effects 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 235000003642 hunger Nutrition 0.000 description 1
- 230000007954 hypoxia Effects 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 239000003112 inhibitor Substances 0.000 description 1
- 229960003130 interferon gamma Drugs 0.000 description 1
- 229940100601 interleukin-6 Drugs 0.000 description 1
- 230000003834 intracellular effect Effects 0.000 description 1
- 230000001788 irregular Effects 0.000 description 1
- 230000007794 irritation Effects 0.000 description 1
- 230000003907 kidney function Effects 0.000 description 1
- 229940067606 lecithin Drugs 0.000 description 1
- 235000010445 lecithin Nutrition 0.000 description 1
- 239000000787 lecithin Substances 0.000 description 1
- 230000003908 liver function Effects 0.000 description 1
- 235000021266 loss of appetite Nutrition 0.000 description 1
- 208000019017 loss of appetite Diseases 0.000 description 1
- 210000003712 lysosome Anatomy 0.000 description 1
- 230000001868 lysosomic effect Effects 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 235000016709 nutrition Nutrition 0.000 description 1
- 230000035764 nutrition Effects 0.000 description 1
- 230000003204 osmotic effect Effects 0.000 description 1
- 229940049954 penicillin Drugs 0.000 description 1
- 230000035790 physiological processes and functions Effects 0.000 description 1
- 239000002504 physiological saline solution Substances 0.000 description 1
- 230000000291 postprandial effect Effects 0.000 description 1
- 239000003755 preservative agent Substances 0.000 description 1
- 230000002335 preservative effect Effects 0.000 description 1
- 230000035755 proliferation Effects 0.000 description 1
- XEYBRNLFEZDVAW-UHFFFAOYSA-N prostaglandin E2 Natural products CCCCCC(O)C=CC1C(O)CC(=O)C1CC=CCCCC(O)=O XEYBRNLFEZDVAW-UHFFFAOYSA-N 0.000 description 1
- 235000018102 proteins Nutrition 0.000 description 1
- 238000003908 quality control method Methods 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 210000004894 snout Anatomy 0.000 description 1
- 230000037351 starvation Effects 0.000 description 1
- 201000011549 stomach cancer Diseases 0.000 description 1
- 229960005322 streptomycin Drugs 0.000 description 1
- 238000001356 surgical procedure Methods 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- 230000002195 synergetic effect Effects 0.000 description 1
- 230000017423 tissue regeneration Effects 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 239000012588 trypsin Substances 0.000 description 1
- 102000003390 tumor necrosis factor Human genes 0.000 description 1
- 235000013343 vitamin Nutrition 0.000 description 1
- 239000011782 vitamin Substances 0.000 description 1
- 229940088594 vitamin Drugs 0.000 description 1
- 229930003231 vitamin Natural products 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
- C12N5/0625—Epidermal cells, skin cells; Cells of the oral mucosa
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/105—Plant extracts, their artificial duplicates or their derivatives
-
- 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
- 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
- A61K35/36—Skin; Hair; Nails; Sebaceous glands; Cerumen; Epidermis; Epithelial cells; Keratinocytes; Langerhans cells; Ectodermal cells
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/88—Liliopsida (monocotyledons)
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/88—Liliopsida (monocotyledons)
- A61K36/894—Dioscoreaceae (Yam family)
- A61K36/8945—Dioscorea, e.g. yam, Chinese yam or water yam
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/43—Enzymes; Proenzymes; Derivatives thereof
- A61K38/46—Hydrolases (3)
- A61K38/48—Hydrolases (3) acting on peptide bonds (3.4)
- A61K38/488—Aspartic endopeptidases (3.4.23), e.g. pepsin, chymosin, renin, cathepsin E
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/0087—Galenical forms not covered by A61K9/02 - A61K9/7023
- A61K9/0095—Drinks; Beverages; Syrups; Compositions for reconstitution thereof, e.g. powders or tablets to be dispersed in a glass of water; Veterinary drenches
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
- A61P1/04—Drugs for disorders of the alimentary tract or the digestive system for ulcers, gastritis or reflux esophagitis, e.g. antacids, inhibitors of acid secretion, mucosal protectants
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y304/00—Hydrolases acting on peptide bonds, i.e. peptidases (3.4)
- C12Y304/23—Aspartic endopeptidases (3.4.23)
- C12Y304/23003—Gastricsin (3.4.23.3)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y304/00—Hydrolases acting on peptide bonds, i.e. peptidases (3.4)
- C12Y304/23—Aspartic endopeptidases (3.4.23)
- C12Y304/23004—Chymosin (3.4.23.4), i.e. rennin
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/10—Growth factors
- C12N2501/12—Hepatocyte growth factor [HGF]
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/10—Growth factors
- C12N2501/135—Platelet-derived growth factor [PDGF]
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/10—Growth factors
- C12N2501/148—Transforming growth factor alpha [TGF-a]
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- Natural Medicines & Medicinal Plants (AREA)
- Medicinal Chemistry (AREA)
- Pharmacology & Pharmacy (AREA)
- Organic Chemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Zoology (AREA)
- Epidemiology (AREA)
- Biotechnology (AREA)
- Biomedical Technology (AREA)
- Wood Science & Technology (AREA)
- Mycology (AREA)
- Cell Biology (AREA)
- Genetics & Genomics (AREA)
- Microbiology (AREA)
- General Engineering & Computer Science (AREA)
- Immunology (AREA)
- Biochemistry (AREA)
- Botany (AREA)
- Food Science & Technology (AREA)
- Medical Informatics (AREA)
- Nutrition Science (AREA)
- Polymers & Plastics (AREA)
- Developmental Biology & Embryology (AREA)
- Dermatology (AREA)
- Alternative & Traditional Medicine (AREA)
- Virology (AREA)
- Gastroenterology & Hepatology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
Abstract
The embodiment of the invention discloses a gastric mucosa epithelial cell extract and a preparation method and application thereof. The invention sequentially carries out common culture, induced culture, anoxic culture and low-temperature stimulation on the gastric mucosal epithelial cells, can obviously improve the levels of various repair factors secreted by the gastric mucosal epithelial cells, and the obtained gastric mucosal epithelial cell extract has strong keratogenesis and renewal capacity, simultaneously inhibits inflammatory reaction, improves local blood circulation, protects local mucous membrane and plays an important role in promoting the repair of damaged tissues of the gastric mucosal.
Description
Technical Field
The embodiment of the invention relates to the technical field of biological medicines, in particular to a gastric mucosa epithelial cell extract and a preparation method and application thereof.
Background
Along with the faster and faster pace of life, the pressure of life and work is greater and greater, sleep and diet are irregular, helicobacter pylori infection and stomach discomfort are more and more people, and common factors in life are as follows: the method is characterized in that foods such as cold and hot spicy stimulation are frequently drunk, binge eating and the like are carried out, under the frequent stimulation of bad habits, gastric mucosa is damaged to different degrees, barrier function of the gastric mucosa is weakened, unbalance can occur to microenvironment, long-time gastric mucosa is damaged, viruses and bacteria are invaded, a series of inflammatory factors such as interleukin-1 beta, interleukin-6, tumor necrosis factor, interferon-gamma and the like are released, the continuous secretion of the inflammatory factors and the continuous reduction of some protective factors such as PGE2 are carried out, the difficulty of continuous destruction and repair of tissues is increased, the normal physiological functions and repair capability of the gastric mucosa are reduced, and the secretion of anti-inflammatory factors and tissue repair factors is insufficient. These factors are necessary for cell proliferation and repair, and lack of these factors can affect repair of mucous membrane, long-term damage of gastric mucous membrane, and repeated irritation of inflammation results in poorer defense capability, which causes frequent gastric diseases, mainly gastritis, erosion, ulcer, perforation and the like of gastric mucous membrane, and gastric cancer incidence is also in an ascending trend.
Therefore, repairing damaged gastric mucosa is the root of solving the problem, promoting the repair of gastric mucosa, and there is a need to improve the repair status of gastric mucosa by providing appropriate microenvironment, suppressing inflammatory reaction, promoting cell proliferation, providing sufficient repair factors, and the like.
At present, the treatment of gastric mucosal lesions mainly comprises:
1. western medicines or traditional Chinese medicines are used for treatment, have large side effects, often have a plurality of side effects, and are easy to cause complications; the traditional Chinese medicine is poor in taste, difficult to adhere to, cannot achieve a good treatment effect, often causes frequent repeated attacks of stomach diseases, generally needs long-term administration, is harmful to health after long-term use, and is unfavorable for self-repair of stomach.
2. For patients with serious gastric mucosa injury, repeated bleeding and special ulcer parts, some patients need to be treated by surgery to cut the ulcer parts so as to prevent disease recurrence, and the patients bear great pain.
At present, the treatment of symptoms is most, the medicines for promoting the regeneration and repair of gastric mucosal tissues are few, the treatment means is single, western medicines or traditional Chinese medicines are generally used for treatment, the medicines generally cause the metabolism burden of the liver and the kidney through the metabolism of the liver and the kidney, are easy to damage, have complex western medicine components, often have a plurality of side effects and are easy to cause complications. The quality of the traditional Chinese medicine materials is uneven, the problems of low extraction efficiency of the herbal medicine, unstable content of active ingredients and the like exist, the ingredients of the various herbal medicines are complicated, the difficulty in drug effect evaluation and quality control is high, the stability and the drug effect are not guaranteed, drug interaction or adverse reaction easily occurs, and the traditional Chinese medicine is difficult to adhere due to poor taste, so that a better treatment effect cannot be achieved.
Disclosure of Invention
Therefore, the embodiment of the invention provides a gastric mucosa epithelial cell extract and a preparation method and application thereof, so as to solve the problems of poor effect, poor safety and the like of the existing gastric mucosa repair promoting product.
In order to achieve the above object, the embodiment of the present invention provides the following technical solutions:
according to a first aspect of an embodiment of the present invention, the present invention provides a method for preparing a gastric mucosal epithelial cell extract, the method comprising:
inoculating gastric mucosa epithelial cells into a 6-hole culture plate for culture, subculturing when the cells are fused to 80% -90%, and performing related identification, and adding an induction culture medium into the cells when the cells are transferred to the 5 th generation, placing the cells at 37 ℃ and 5% CO 2 Induced culture in incubator for 72 hr, then transfer to 37deg.C, 10% O 2 Culturing in an incubator for 24 hours under oxygen deficiency, repeating the steps of induction culture and anoxic culture, transferring to 7 th generation, pouring out the induction culture medium after the cells are fused to 90% -100%, washing the cells for three times with normal saline, adding purified water for low-temperature stimulation, collecting the supernatant, removing cell fragments, filtering and sterilizing to obtain gastric mucosa epithelial cell extract; wherein the induction culture medium consists of a basal culture medium, HGF, PDGF and TGF-alpha, and the final concentrations of HGF, PDGF, TGF-alpha are respectively as follows: 8-15ng/ml, 12-18ng/ml, 6-10ng/ml.
HGF and PDGF can promote the generation, survival and regeneration of tissue cells and inhibit the apoptosis of the cells; TGF-alpha has the function of activating paracellular secretion, can stimulate exosomes to secrete a large amount of transforming growth factors, thereby promoting proliferation of cells and having bidirectional regulation. It was found that the induction medium containing HGF, PDGF and TGF- α in the above final concentration ranges significantly increased the level of active factor secretion in the gastric mucosal epithelial cell extract. The invention sequentially carries out common culture, induction culture, anoxic culture and low-temperature stimulation on the gastric mucosal epithelial cells, and the obtained gastric mucosal epithelial cell extract has strong keratogenesis and renewal capacity, simultaneously inhibits inflammatory reaction, improves local blood circulation, protects local mucous membrane and plays an important role in promoting the repair of damaged tissues of the gastric mucosal.
Further, the final concentrations of HGF, PDGF, TGF- α were: 10g/ml, 15ng/ml, 8ng/ml.
Further, the basal medium is DMEM/F-12 containing 10v/v% fetal bovine serum.
Further, the culture medium used for the culture and the subculture is a medium containingDMEM/F-12 of 10v/v% fetal bovine serum; the conditions of the culture and the subculture are as follows: 37 ℃,5% CO 2 。
Further, the step of adding purified water for low-temperature stimulation comprises the following steps: based on the cell area, according to 3.5-4.5ml/cm 2 Purified water is added, and then the mixture is kept stand in a refrigerator at the temperature of 4 ℃ for 1 to 2 hours. The research shows that when the low-temperature stimulation is carried out under the conditions, the cells can release a large amount of intracellular factors under the actions of starvation and osmotic pressure, and meanwhile, the lysosome enzyme activity is reduced under the low-temperature environment of 4 ℃, so that the degradation of the factors is reduced, and the factor content in the extracting solution is further improved.
Further, the step of removing cell debris is: after centrifugation at 750 Xg for 10 minutes, the supernatant was centrifuged at 2000 Xg for 20 minutes, and the supernatant was again centrifuged at 1000 Xg for 30 minutes;
the filtering adopts a 70 mesh screen;
the sterilization adopts a filter membrane with the pore diameter of 0.22 mu m.
According to a second aspect of embodiments of the present invention, there is provided a gastric mucosal epithelial cell extract made by the method as described in any one of the above.
According to a third aspect of embodiments of the present invention, there is provided the use of a gastric mucosal epithelial cell extract as described above in the preparation of a product for promoting repair of gastric mucosa.
According to a fourth aspect of embodiments of the present invention, the present invention provides an oral liquid for promoting repair of gastric mucosa, the oral liquid comprising: gastric mucosal epithelial cell extract, milk extract, yam extract, chymosin, pepsin and banana extract as described above; based on each 1000ml of gastric mucosa epithelial cell extract, the dosages of the milk extract, the Chinese yam extract and the banana extract are respectively 60-100g, 40-70g and 30-60g, and the final concentrations of chymosin and pepsin are 3500-4500U and 1000-2000U.
The milk extract contains rich substances such as milk calcium, protein, vitamins, lecithin and the like, can provide nutrition for the stomach, enhance the immunity of the organism, and can form a protective film on the erosion surface or ulcer surface of the stomach so as to play a synergistic effect in repairing gastric mucosa. The rhizoma Dioscoreae extract contains mucin, can prevent gastric mucosa injury, and protect gastric wall under the action of pepsin, and is beneficial for preventing gastric ulcer and gastritis. The banana extract contains 5-hydroxytryptamine, which can protect human gastric mucosa, reduce gastric acid secretion, and also contains a special chemical substance, which can promote the growth of gastric mucosa cells, and can enhance the acid-resistant ability of gastric mucosa cells, thereby protecting and repairing gastric mucosa. Pepsin and chymosin mainly have the effects of promoting protein decomposition, promoting secretion of gastrointestinal glands, improving blood circulation of digestive tract, and improving absorption capacity of digestive tract to nutrient components and resistance to inflammation. According to the invention, further research shows that the gastric mucosa epithelial cell extract is combined with the raw materials, plays a role in synergism in mucosa protection and repair, can effectively repair and regenerate damaged mucosa tissues, and achieves the purpose of relieving/treating stomach discomfort.
Further, the dosages of the milk extract, the Chinese yam extract and the banana extract are respectively 80g, 60g and 50g, and the final concentrations of chymosin and pepsin are 4000U and 1500U.
The embodiment of the invention has the following advantages:
(1) The safety is high: the product is a pure biological combined stomach nourishing food preparation, has no toxic or side effect, and is safer when being used.
(2) Stable process and high acquisition performance: the preparation technology of the unique oral liquid for high expression of the mucosa repair cytokines and promoting the mucosa repair has the advantages of very stable process and effective acquisition of targeted active mucosa repair factors.
(3) The effectiveness is high: by utilizing the unique advantages of the mucous membrane cells, the active repair factors secreted by the cells have the characteristics of small molecular weight and easy absorption, act on the damaged part, repair and regenerate damaged mucous membrane tissues, and fundamentally solve the problems existing in the stomach.
(4) No drug or preservative is added.
(5) The taste is fresh and sweet, and the problem that people suffering from stomach discomfort cannot adhere to the stomach due to poor taste of the medicine is solved.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below. It will be apparent to those of ordinary skill in the art that the drawings in the following description are exemplary only and that other implementations can be obtained from the extensions of the drawings provided without inventive effort.
FIG. 1 is a graph showing the comparison of the secretion of active factors in gastric mucosal epithelial cell extracts provided by the invention;
FIG. 2 is a photograph of a gastric mucosal endoscope of a patient before and after treatment, before left-treatment, after right-treatment, according to the present invention;
fig. 3 is a photograph of a gastric mucosal endoscope of another patient provided by the present invention before and after treatment, left-before treatment, right-after treatment.
Detailed Description
Other advantages and advantages of the present invention will become apparent to those skilled in the art from the following detailed description, which, by way of illustration, is to be read in connection with certain specific embodiments, but not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
In the following, milk extract (milk calcium) was purchased from milk calcium biotechnology, model 013289; the rhizoma Dioscoreae extract is obtained from Scott organism, model number 90147-49-2; banana extracts were purchased from the snout organism; chymosin is purchased from Shandong Duckweed Polybion Co., ltd, model 9001-98-3; pepsin was purchased from the nanning Pang Bo organism.
The steps for separating gastric mucosal epithelial cells are as follows:
taking the stomach of abortion pig after 15 weeks, repeatedly washing in sterile D-Hanks solution containing penicillin and streptomycin for at least three timesThe mucosa lamina propria is peeled by the ophthalmic forceps and sheared into 1mm 3 Placing the fragments in 0.1% type I collagenase and 0.1% type II collagenase, digesting for 60-80 min at 37 ℃, filtering and collecting cell suspension by a 100-mesh filter screen, centrifuging for 10min at 600 Xg, discarding the supernatant, and centrifuging and washing for 3 times by using D-Hanks liquid to obtain the gastric mucosal epithelial cells.
Example 1
The embodiment provides a gastric mucosal epithelial cell extract, and the preparation method thereof is as follows:
(1) Primary culture: gastric mucosal epithelial cells at 1×10 5 Inoculating into 6-well culture plate, adding DMEM/F-12 containing 10v/v% foetal calf serum, placing into incubator (37deg.C, 5% CO) 2 ) The non-adherent cells were aspirated for 10-15 minutes and the culture continued.
(2) Subculture: the following identification was performed when cells were fused to 80%: positive markers, positive for the identification of keratin 8, 18 and 19 antibodies, and the cell morphology conforming to the gastric mucosal epithelial cell morphology observed under a microscope. Simultaneously starting passage, removing cell culture medium when cells are fused to 80%, adding 20ml of D-Hanks into each bottle for washing, removing washing liquid after washing, adding 2ml-3ml of 5% trypsin into each bottle for digestion for about 3 minutes, observing cell shedding, adding 10ml of DMEM/F-12 complete culture medium (containing 10v/v% fetal calf serum) for stopping digestion, collecting cell suspension, adding 33ml of D-Hanks, centrifuging for 10 minutes at 500 Xg, removing washing liquid after washing for 2 times, and removing cells according to 1X 10 4 /cm 2 Inoculated in 150cm of complete medium (10 v/v% foetal calf serum) containing 30ml of DMEM/F-12 2 Placing the culture flask into an incubator (37 ℃ C., 5% CO) 2 ) Is cultured. According to this culture procedure, cells were expanded to the 5 th generation, induced, and the cells were subjected to induction culture in an incubator (37 ℃ C., 5% CO) with the medium being changed to an induction medium (DMEM/F-12 containing 10% fetal bovine serum, 10ng/ml HGF, 15ng/ml PDGF, 8ng/ml TGF-. Alpha.) 2 ) Culturing for 72 hr, transferring the induced cells into three-steam incubator, and artificially preparing anoxic microenvironment (37deg.C, 5% CO) 2 、10%O 2 ) Culture was continued for 24 hours. The steps of induction culture and hypoxia culture were repeated to pass the cells to passage 7.
(3) Low temperature stimulation and collection of supernatant: after the cells were grown, the induction medium was poured out, and the cells were washed three times with 0.9% physiological saline, based on the cell area, at a rate of 4ml/cm 2 Adding purified water, standing in a refrigerator at 4deg.C for 1 hr, collecting cell supernatant, centrifuging at 750Xg for 10min, collecting supernatant, centrifuging at 2000 Xg for 20 min, collecting supernatant again, centrifuging at 1000g for 30 min, removing cell debris by gradient centrifugation, filtering with 70 mesh sieve, and sterilizing with 0.22 μm to obtain gastric mucosa epithelial cell extractive solution.
Comparative example 1
The comparative example provides a gastric mucosal epithelial cell extract, which is prepared by a method differing from example 1 only in that an induction medium consisting of DMEM/F-12, 5ng/ml HGF, 15ng/ml PDGF containing 10% fetal bovine serum is used.
Example 2
The embodiment provides an oral liquid for promoting gastric mucosa repair, which is prepared by the following steps:
according to the method, 80g of milk extract, 60g of Chinese yam extract, 50g of banana extract, 4000U of chymosin and 1500U of pepsin are added into each 1000ml of gastric mucosa epithelial cell extract in example 1 for preparation, and all materials are mixed and stirred uniformly.
Example 3
The embodiment provides an oral liquid for promoting gastric mucosa repair, which is prepared by the following steps:
according to the method, 80g of milk extract, 50g of Chinese yam extract, 30g of banana extract, 3500U of chymosin and 2000U of pepsin are added into each 1000ml of gastric mucosa epithelial cell extract in example 1 for preparation, and all materials are mixed and stirred uniformly.
Example 4
The embodiment provides an oral liquid for promoting gastric mucosa repair, which is prepared by the following steps:
100g of milk extract, 60g of yam extract, 50g of banana extract, 4000U of chymosin and 1000U of pepsin were added per 1000ml of the gastric mucosa epithelial cell extract of example 1 to prepare materials, and all materials were mixed.
Comparative example 2
The present comparative example provides an oral liquid for promoting repair of gastric mucosa, and the preparation method thereof differs from example 2 only in that the gastric mucosa epithelial cell extract in this example is replaced with the gastric mucosa epithelial cell extract of comparative example 1 in equal amount.
Test example 1
The secretion amount of active factors in the gastric mucosal epithelial cell extracts prepared in example 1 and comparative example 1 was measured using an Elisa kit, and the results are shown in table 1 below. The results were plotted and shown in FIG. 1.
TABLE 1 (Unit: pg/ml)
The results show that: the secretion level of the active factors in the gastric mucosa epithelial cell extract prepared by the embodiment of the invention is higher.
Test example 2
Clinical data: 60 cases of patients with erosive gastritis are selected from the outpatient diagnosis. Inclusion criteria: meets the clinical diagnosis standard of chronic erosive gastritis in Chinese chronic gastritis consensus; 2) Meets the diagnosis standard (symptoms: upper abdominal discomfort, fullness, pain, loss of appetite, eructation, acid regurgitation, etc.).
All patients were randomly divided into test 1-4 groups, 15 cases/group, and the test 1-4 groups were treated with the gastric mucosal epithelial cell extract of example 1, the oral liquid of example 2, the gastric mucosal epithelial cell extract of comparative example 1, and the oral liquid of comparative example 2, respectively. Orally taken after meal, 150-200 ml/day for 1 month. Symptoms and signs were observed before and after treatment, the extent of gastric mucosal erosion was endoscopically examined, gastric juice was examined for changes in inflammatory factors, and blood routine and liver and kidney function.
Standard of efficacy: reference to "diagnosis and treat guide for digestive diseases" of traditional Chinese medicine
And (3) healing: the clinical symptoms and gastric mucosal erosion basically disappear;
the effect is shown: the clinical symptoms are obviously improved, and the area of gastric mucosal erosion reduction is more than or equal to 50%;
the method is effective: the clinical symptoms are improved, and the area of gastric mucosal erosion is reduced by less than 50%;
invalidation: the clinical symptoms and gastric mucosal erosion conditions are not obviously improved;
total effective rate= (recovery + onset + efficacy)/total number of cases 100%.
The treatment results are shown in table 2, and each group of treatment achieves better clinical curative effect, wherein the oral liquid in the embodiment 2 has more remarkable curative effect in promoting gastric mucosa repair. No serious adverse reactions occurred in patients in treatment groups 1-4.
TABLE 2
Typical cases:
1. the patient has a certain poplar, a woman, 36 years old, a history of chronic gastritis for 2 years, intermittent subxiphoid discomfort, abdominal distension with hidden pain, obvious postprandial effect and poor appetite, and the oral gastrointestinal motility medicine is not relieved; checking: abdominal weakness, subxiphoid depression discomfort, no tenderness, rebound pain, negative Murphy's symptoms and normal borborygmus. The oral liquid of the example 2 is orally taken, and after 1 month, the discomfort under the xiphoid process is obviously relieved, and the appetite is restored to be normal; checking: abdominal weakness, the subxiphoid depression vanishes, no tenderness, rebound pain and normal borborygmus.
The changes of gastric inflammatory factors before and after treatment are shown in Table 3, and the gastric mucosal endoscopic photographs are shown in FIG. 2.
TABLE 3 Table 3
2. The patient is somehow in his or her plum, 45 years old, the history of chronic gastritis is 3 years old, the patient is intermittently provided with abdominal pain, acid regurgitation is accompanied with belch after meal, and the oral acid inhibitor is slightly relieved; checking: soft abdomen, light pain under the xiphoid process, no rebound pain, negative Murphy's sign and normal borborygmus. Oral administration of the oral liquid of example 2, after 1 month, clinical symptoms were relieved; checking: soft abdomen, no pressure pain, rebound pain and normal borygmus.
Changes in gastric inflammatory factors before and after treatment are shown in Table 4, and changes in gastric mucosal erosion degree are shown in FIG. 3.
TABLE 4 Table 4
While the invention has been described in detail in the foregoing general description and specific examples, it will be apparent to those skilled in the art that modifications and improvements can be made thereto. Accordingly, such modifications or improvements may be made without departing from the spirit of the invention and are intended to be within the scope of the invention as claimed.
Claims (10)
1. A method for preparing gastric mucosal epithelial cell extract, comprising:
inoculating gastric mucosa epithelial cells into a 6-hole culture plate for culture, subculturing when the cells are fused to 80% -90%, and performing related identification, and adding an induction culture medium into the cells when the cells are transferred to the 5 th generation, placing the cells at 37 ℃ and 5% CO 2 Induced culture in incubator for 72 hr, then transfer to 37deg.C, 10% O 2 Culturing in an incubator for 24 hours under oxygen deficiency, repeating the steps of induction culture and anoxic culture, transferring to 7 th generation, pouring out the induction culture medium after the cells are fused to 90% -100%, washing the cells for three times with normal saline, adding purified water for low-temperature stimulation, collecting the supernatant, removing cell fragments, filtering and sterilizing to obtain gastric mucosa epithelial cell extract; wherein the induction culture medium consists of a basal culture medium, HGF, PDGF and TGF-alpha, and the final concentrations of HGF, PDGF, TGF-alpha are respectively as follows: 8-15ng/ml, 12-18ng/ml, 6-10ng/ml.
2. The method for preparing gastric mucosal epithelial cell extract according to claim 1, wherein the final concentrations of HGF, PDGF, TGF- α are respectively: 10g/ml, 15ng/ml, 8ng/ml.
3. The method for preparing gastric mucosal epithelial cell extract according to claim 1, wherein said basic medium is DMEM/F-12 containing 10v/v% fetal bovine serum.
4. The method for preparing gastric mucosal epithelial cell extract according to claim 1, wherein the culture medium used for the culture and the subculture is DMEM/F-12 containing 10v/v% fetal bovine serum;
the conditions of the culture and the subculture are as follows: 37 ℃,5% CO 2 。
5. The method for preparing gastric mucosal epithelial cell extract according to claim 1, wherein the step of adding purified water for low temperature stimulation comprises: based on the cell area, according to 3.5-4.5ml/cm 2 Purified water is added, and then the mixture is kept stand in a refrigerator at the temperature of 4 ℃ for 1 to 2 hours.
6. The method for preparing gastric mucosal epithelial cell extract according to claim 1, wherein said step of removing cell debris comprises: after centrifugation at 750 Xg for 10 minutes, the supernatant was centrifuged at 2000 Xg for 20 minutes, and the supernatant was again centrifuged at 1000 Xg for 30 minutes;
the filtering adopts a 70 mesh screen;
the sterilization adopts a filter membrane with the pore diameter of 0.22 mu m.
7. A gastric mucosal epithelial cell extract, characterized in that it is made by the method of any one of claims 1-6.
8. Use of the gastric mucosal epithelial cell extract of claim 7 in the preparation of a product for promoting repair of gastric mucosa.
9. An oral liquid for promoting repair of gastric mucosa, characterized in that the oral liquid comprises:
the gastric mucosal epithelial cell extract, milk extract, yam extract, chymosin, pepsin and banana extract of claim 7; based on each 1000ml of gastric mucosa epithelial cell extract, the dosages of the milk extract, the Chinese yam extract and the banana extract are respectively 60-100g, 40-70g and 30-60g, and the final concentrations of chymosin and pepsin are 3500-4500U and 1000-2000U.
10. The oral liquid for promoting gastric mucosal repair according to claim 9, wherein the milk extract, the yam extract and the banana extract are used in an amount of 80g, 60g and 50g, respectively, and the final concentration of chymosin and pepsin is 4000U and 1500U.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202410010945.5A CN117503800B (en) | 2024-01-04 | 2024-01-04 | Gastric mucosa epithelial cell extract and preparation method and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202410010945.5A CN117503800B (en) | 2024-01-04 | 2024-01-04 | Gastric mucosa epithelial cell extract and preparation method and application thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN117503800A true CN117503800A (en) | 2024-02-06 |
| CN117503800B CN117503800B (en) | 2024-04-05 |
Family
ID=89757018
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202410010945.5A Active CN117503800B (en) | 2024-01-04 | 2024-01-04 | Gastric mucosa epithelial cell extract and preparation method and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN117503800B (en) |
Citations (31)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1075874A (en) * | 1993-01-01 | 1993-09-08 | 温州医学院 | Medicine for treating chronic atrophic gastritis |
| WO2000003002A2 (en) * | 1998-07-09 | 2000-01-20 | Universite De Sherbrooke | Pure and viable human epithelial cell preparations from digestive tissue |
| CN1331250A (en) * | 2000-04-14 | 2002-01-16 | 株式会社源 | Glucoprotein having inhibitory activity to pyroric spirillar colonization |
| CN1539961A (en) * | 2003-04-21 | 2004-10-27 | 宋于刚 | Method for separatng cell of extracorporeal gastic mucosa and relevant purification method |
| CN101103815A (en) * | 2007-08-17 | 2008-01-16 | 广东医学院 | Banana heath-care biscuit with blood pressure lowering, blood lipoid reducing and angiocardiopathy preventing, and its formula technology |
| CN101214349A (en) * | 2007-12-29 | 2008-07-09 | 高彦越 | Medicine for treating peptic ulcer |
| CN101244248A (en) * | 2008-02-02 | 2008-08-20 | 孟德芹 | Method of preparing traditional Chinese medicine for treating chronic superficial gastritis |
| CN101264105A (en) * | 2007-03-15 | 2008-09-17 | 赵新民 | Lamb stomach extract vitamin B12 preparations composing prescription |
| CN101264106A (en) * | 2007-03-15 | 2008-09-17 | 赵新民 | Preparation of lamb stomach extract and its preparation |
| CN101664406A (en) * | 2008-07-09 | 2010-03-10 | 香港中文大学 | Composition for treating and/or preventing gastric ulcer and use thereof |
| CN102188641A (en) * | 2010-03-18 | 2011-09-21 | 罗红英 | Medicament for treating stomach diseases |
| CN102335383A (en) * | 2010-07-23 | 2012-02-01 | 赵鹏 | Gastric cancer health-care product and preparation method thereof |
| TW201233393A (en) * | 2011-02-01 | 2012-08-16 | Univ Chaoyang Technology | A pharmaceutical composition for inhibition of helicobacter pylori growth and helicobacter pylori-induced inflammation in human gastric epithelial cells |
| CN103289950A (en) * | 2013-06-28 | 2013-09-11 | 浙江大学 | Methods for in vitro separation, culture, preservation and recovery of milk cow omasum epithelial cells |
| CN103404758A (en) * | 2013-05-27 | 2013-11-27 | 胡安然 | Special diet for gastric ulcer patients |
| CN103750324A (en) * | 2014-01-27 | 2014-04-30 | 泰州比尔夫生物科技有限公司 | Milk basic protein hericium erinaceus powder composite piece |
| CN105238741A (en) * | 2015-11-14 | 2016-01-13 | 河南农业大学 | Complete medium for culturing ruminal epithelium cells of dairy cows and application thereof |
| CN105907700A (en) * | 2016-04-28 | 2016-08-31 | 王晓冰 | Primary rat or mouse gastric mucosal epithelial cell isolation and culture method |
| CN106361993A (en) * | 2016-10-25 | 2017-02-01 | 陕西中医药大学 | Medicinal composition for preventing and treating gastric mucosal lesion and preparation method of medicinal composition |
| CN106967674A (en) * | 2017-05-22 | 2017-07-21 | 江西农业大学 | A kind of isolated culture method of sheep rumen epithelial cell |
| CN107636149A (en) * | 2015-04-03 | 2018-01-26 | 普罗帕格尼克斯公司 | The vitro proliferation of epithelial cell |
| CN107881197A (en) * | 2017-11-17 | 2018-04-06 | 扬州大学 | One kind immortalizes cow rumen epithelial cell line and its construction method |
| CN108948130A (en) * | 2017-05-23 | 2018-12-07 | 广州资生生物科技有限公司 | A kind of new technology without using chemicals low temperature preparation gastric mucosal cell extract |
| CN108969584A (en) * | 2018-10-24 | 2018-12-11 | 海南医学院 | A kind of composition and its preparation method and application |
| CN112342183A (en) * | 2020-10-30 | 2021-02-09 | 广东省农业科学院动物科学研究所 | Method for establishing piglet gastrointestinal tract in-vitro environment model and application |
| CN112458040A (en) * | 2020-12-23 | 2021-03-09 | 四川农业大学 | Isolation culture method of rumen epithelial cells of adult yaks |
| CN112725270A (en) * | 2021-03-29 | 2021-04-30 | 北京益华生物科技有限公司 | Human-derived bone marrow mesenchymal stem cell induction culture medium and induction method |
| CN113088482A (en) * | 2021-04-16 | 2021-07-09 | 四川农业大学 | Method for separating and culturing rumen epithelial cells of calves |
| CN114058592A (en) * | 2021-11-16 | 2022-02-18 | 四川农业大学 | Immortalized yak rumen epithelial cell line and construction method thereof |
| CN114107172A (en) * | 2021-11-12 | 2022-03-01 | 上海市农业科学院 | Method for constructing non-infectious gastric mucosa injury cell model and application |
| CN117050929A (en) * | 2023-04-17 | 2023-11-14 | 西北农林科技大学 | Method for separating and culturing rumen epithelial cells of dairy cows |
-
2024
- 2024-01-04 CN CN202410010945.5A patent/CN117503800B/en active Active
Patent Citations (31)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1075874A (en) * | 1993-01-01 | 1993-09-08 | 温州医学院 | Medicine for treating chronic atrophic gastritis |
| WO2000003002A2 (en) * | 1998-07-09 | 2000-01-20 | Universite De Sherbrooke | Pure and viable human epithelial cell preparations from digestive tissue |
| CN1331250A (en) * | 2000-04-14 | 2002-01-16 | 株式会社源 | Glucoprotein having inhibitory activity to pyroric spirillar colonization |
| CN1539961A (en) * | 2003-04-21 | 2004-10-27 | 宋于刚 | Method for separatng cell of extracorporeal gastic mucosa and relevant purification method |
| CN101264106A (en) * | 2007-03-15 | 2008-09-17 | 赵新民 | Preparation of lamb stomach extract and its preparation |
| CN101264105A (en) * | 2007-03-15 | 2008-09-17 | 赵新民 | Lamb stomach extract vitamin B12 preparations composing prescription |
| CN101103815A (en) * | 2007-08-17 | 2008-01-16 | 广东医学院 | Banana heath-care biscuit with blood pressure lowering, blood lipoid reducing and angiocardiopathy preventing, and its formula technology |
| CN101214349A (en) * | 2007-12-29 | 2008-07-09 | 高彦越 | Medicine for treating peptic ulcer |
| CN101244248A (en) * | 2008-02-02 | 2008-08-20 | 孟德芹 | Method of preparing traditional Chinese medicine for treating chronic superficial gastritis |
| CN101664406A (en) * | 2008-07-09 | 2010-03-10 | 香港中文大学 | Composition for treating and/or preventing gastric ulcer and use thereof |
| CN102188641A (en) * | 2010-03-18 | 2011-09-21 | 罗红英 | Medicament for treating stomach diseases |
| CN102335383A (en) * | 2010-07-23 | 2012-02-01 | 赵鹏 | Gastric cancer health-care product and preparation method thereof |
| TW201233393A (en) * | 2011-02-01 | 2012-08-16 | Univ Chaoyang Technology | A pharmaceutical composition for inhibition of helicobacter pylori growth and helicobacter pylori-induced inflammation in human gastric epithelial cells |
| CN103404758A (en) * | 2013-05-27 | 2013-11-27 | 胡安然 | Special diet for gastric ulcer patients |
| CN103289950A (en) * | 2013-06-28 | 2013-09-11 | 浙江大学 | Methods for in vitro separation, culture, preservation and recovery of milk cow omasum epithelial cells |
| CN103750324A (en) * | 2014-01-27 | 2014-04-30 | 泰州比尔夫生物科技有限公司 | Milk basic protein hericium erinaceus powder composite piece |
| CN107636149A (en) * | 2015-04-03 | 2018-01-26 | 普罗帕格尼克斯公司 | The vitro proliferation of epithelial cell |
| CN105238741A (en) * | 2015-11-14 | 2016-01-13 | 河南农业大学 | Complete medium for culturing ruminal epithelium cells of dairy cows and application thereof |
| CN105907700A (en) * | 2016-04-28 | 2016-08-31 | 王晓冰 | Primary rat or mouse gastric mucosal epithelial cell isolation and culture method |
| CN106361993A (en) * | 2016-10-25 | 2017-02-01 | 陕西中医药大学 | Medicinal composition for preventing and treating gastric mucosal lesion and preparation method of medicinal composition |
| CN106967674A (en) * | 2017-05-22 | 2017-07-21 | 江西农业大学 | A kind of isolated culture method of sheep rumen epithelial cell |
| CN108948130A (en) * | 2017-05-23 | 2018-12-07 | 广州资生生物科技有限公司 | A kind of new technology without using chemicals low temperature preparation gastric mucosal cell extract |
| CN107881197A (en) * | 2017-11-17 | 2018-04-06 | 扬州大学 | One kind immortalizes cow rumen epithelial cell line and its construction method |
| CN108969584A (en) * | 2018-10-24 | 2018-12-11 | 海南医学院 | A kind of composition and its preparation method and application |
| CN112342183A (en) * | 2020-10-30 | 2021-02-09 | 广东省农业科学院动物科学研究所 | Method for establishing piglet gastrointestinal tract in-vitro environment model and application |
| CN112458040A (en) * | 2020-12-23 | 2021-03-09 | 四川农业大学 | Isolation culture method of rumen epithelial cells of adult yaks |
| CN112725270A (en) * | 2021-03-29 | 2021-04-30 | 北京益华生物科技有限公司 | Human-derived bone marrow mesenchymal stem cell induction culture medium and induction method |
| CN113088482A (en) * | 2021-04-16 | 2021-07-09 | 四川农业大学 | Method for separating and culturing rumen epithelial cells of calves |
| CN114107172A (en) * | 2021-11-12 | 2022-03-01 | 上海市农业科学院 | Method for constructing non-infectious gastric mucosa injury cell model and application |
| CN114058592A (en) * | 2021-11-16 | 2022-02-18 | 四川农业大学 | Immortalized yak rumen epithelial cell line and construction method thereof |
| CN117050929A (en) * | 2023-04-17 | 2023-11-14 | 西北农林科技大学 | Method for separating and culturing rumen epithelial cells of dairy cows |
Non-Patent Citations (3)
| Title |
|---|
| STEFANO MILANI等: "Role of Growth Factors and Their Receptors in Gastric Ulcer Healing", 《MICROSCOPY RESEARCH AND TECHNIQUE》, vol. 53, 9 May 2001 (2001-05-09), pages 360 - 371 * |
| WUYI KONG等: "Hepatocyte Growth Factor Stimulates Fetal Gastric Epithelial Cell Growth in Vitro", 《JOURNAL OF SURGICAL RESEARCH》, vol. 78, 31 December 1998 (1998-12-31), pages 161 - 168 * |
| 杜改梅等: "1-1 Ghrelin 对断奶仔猪胃毅膜上皮细胞中H+-K+-ATPase和胃蛋白酶活性及胃猫膜上皮细胞增殖活力的体外作用", 《全国动物生理生化第九次学术交流会论文摘要汇编》, 31 December 2006 (2006-12-31), pages 77 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN117503800B (en) | 2024-04-05 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN108339112B (en) | Nutritional composition for promoting wound healing, bedsore repair and postoperative stress ulcer healing | |
| CN102058781B (en) | Traditional Chinese medicinal film agent for treating burns and scalds | |
| Xu | Burns regenerative medicine and therapy | |
| CN114306206A (en) | External traditional Chinese medicine composition for treating thyroid nodule and preparation method and application of external traditional Chinese medicine gel plaster | |
| CN108324921B (en) | A pharmaceutical composition with cyst removing and gastric mucosa protecting effects, and its preparation method | |
| CN113413454A (en) | Composition for promoting gastric mucosa injury repair and preparation method thereof | |
| CN117503800B (en) | Gastric mucosa epithelial cell extract and preparation method and application thereof | |
| US11806376B2 (en) | Pharmaceutical composition for treating oral ulcer and preparation method and application thereof | |
| CN109966364A (en) | A kind of Chinese medicine composition and preparation method thereof for treating skin trauma | |
| Jennings et al. | Clinical and Blood Pictures in Adult Scurvy | |
| CN108434301A (en) | External medicine composition and preparation method thereof for diabetes wound repair | |
| CN112316000A (en) | Traditional Chinese medicine composition for treating epigastric pain and patch and preparation method thereof | |
| CN109394885A (en) | A kind of Chinese materia medica preparation of anti-nonalcoholic fatty liver | |
| CN114796283B (en) | Application of five-cereal worm extract in preparation of medicine for treating digestive diseases and pharmaceutical composition | |
| CN1354002A (en) | Medicine for curing gastrointestinal diseases and its production method | |
| US20230065462A1 (en) | Pharmaceutical composition and use thereof | |
| CN105194717B (en) | Promote the sponginum and preparation method of scalpel edge healing | |
| CN113559206A (en) | Compound traditional Chinese medicine for preventing and treating atrial fibrillation | |
| CN102846778B (en) | Chinese medicinal formulation for treating burn and scald and preparation method thereof | |
| Newman | transurethral surgery in relation to bilharziosis of the bladder | |
| CN114767622A (en) | Lycium barbarum glycopeptide hydrogel for preventing or treating oral ulcer and preparation method and application thereof | |
| CN117323368A (en) | Traditional Chinese medicine for treating radiation proctitis and preparation method thereof | |
| CN108379417A (en) | A kind of Chinese medicine composition and preparation method thereof for treating the diabetes surface of a wound | |
| CN112451634A (en) | Formula for treating duodenal gastritis and erosive gastritis | |
| CN1073365A (en) | Chinese medicine dysentery spirit (Kowloon capsule) and compound method |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |