CN116333932A - Lactobacillus johnsonii LS04 and application thereof in preparation of medicines for treating or preventing allergic diseases - Google Patents
Lactobacillus johnsonii LS04 and application thereof in preparation of medicines for treating or preventing allergic diseases Download PDFInfo
- Publication number
- CN116333932A CN116333932A CN202310249339.4A CN202310249339A CN116333932A CN 116333932 A CN116333932 A CN 116333932A CN 202310249339 A CN202310249339 A CN 202310249339A CN 116333932 A CN116333932 A CN 116333932A
- Authority
- CN
- China
- Prior art keywords
- lactobacillus johnsonii
- tryptase
- treating
- allergic diseases
- medicines
- 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.)
- Pending
Links
- 241001468157 Lactobacillus johnsonii Species 0.000 title claims abstract description 50
- 208000026935 allergic disease Diseases 0.000 title claims abstract description 17
- 239000003814 drug Substances 0.000 title claims abstract description 13
- 229940079593 drug Drugs 0.000 title abstract description 7
- 238000002360 preparation method Methods 0.000 title description 8
- 210000003630 histaminocyte Anatomy 0.000 claims abstract description 16
- 210000003651 basophil Anatomy 0.000 claims abstract description 7
- 238000004321 preservation Methods 0.000 claims abstract description 6
- 238000011282 treatment Methods 0.000 claims abstract description 5
- 230000001580 bacterial effect Effects 0.000 claims description 16
- 239000000725 suspension Substances 0.000 claims description 7
- 230000002401 inhibitory effect Effects 0.000 claims description 5
- 238000004519 manufacturing process Methods 0.000 claims description 5
- 238000011321 prophylaxis Methods 0.000 claims 2
- 102000001400 Tryptase Human genes 0.000 abstract description 24
- 108060005989 Tryptase Proteins 0.000 abstract description 24
- 241000252212 Danio rerio Species 0.000 abstract description 12
- 229920002055 compound 48/80 Polymers 0.000 abstract description 8
- 238000001727 in vivo Methods 0.000 abstract description 7
- 230000002265 prevention Effects 0.000 abstract description 4
- 208000003455 anaphylaxis Diseases 0.000 abstract description 3
- 244000005700 microbiome Species 0.000 abstract description 3
- 230000004936 stimulating effect Effects 0.000 abstract description 2
- 206010002198 Anaphylactic reaction Diseases 0.000 abstract 2
- 206010020751 Hypersensitivity Diseases 0.000 description 10
- 210000004027 cell Anatomy 0.000 description 10
- 239000008055 phosphate buffer solution Substances 0.000 description 8
- 239000006041 probiotic Substances 0.000 description 8
- 235000018291 probiotics Nutrition 0.000 description 8
- 239000001963 growth medium Substances 0.000 description 7
- 239000000243 solution Substances 0.000 description 7
- 210000001035 gastrointestinal tract Anatomy 0.000 description 6
- 238000011534 incubation Methods 0.000 description 6
- 230000005764 inhibitory process Effects 0.000 description 6
- 108020004465 16S ribosomal RNA Proteins 0.000 description 4
- 238000000576 coating method Methods 0.000 description 4
- 229960000265 cromoglicic acid Drugs 0.000 description 4
- IMZMKUWMOSJXDT-UHFFFAOYSA-N cromoglycic acid Chemical compound O1C(C(O)=O)=CC(=O)C2=C1C=CC=C2OCC(O)COC1=CC=CC2=C1C(=O)C=C(C(O)=O)O2 IMZMKUWMOSJXDT-UHFFFAOYSA-N 0.000 description 4
- 239000013641 positive control Substances 0.000 description 4
- 239000002244 precipitate Substances 0.000 description 4
- 239000007787 solid Substances 0.000 description 4
- 241000894006 Bacteria Species 0.000 description 3
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 3
- 230000004913 activation Effects 0.000 description 3
- 230000007815 allergy Effects 0.000 description 3
- 230000003266 anti-allergic effect Effects 0.000 description 3
- 239000011248 coating agent Substances 0.000 description 3
- 238000010790 dilution Methods 0.000 description 3
- 239000012895 dilution Substances 0.000 description 3
- 210000003608 fece Anatomy 0.000 description 3
- 238000009630 liquid culture Methods 0.000 description 3
- 239000002904 solvent Substances 0.000 description 3
- 108060003951 Immunoglobulin Proteins 0.000 description 2
- 240000000907 Musa textilis Species 0.000 description 2
- 206010070834 Sensitisation Diseases 0.000 description 2
- 230000000172 allergic effect Effects 0.000 description 2
- 239000000427 antigen Substances 0.000 description 2
- 102000036639 antigens Human genes 0.000 description 2
- 108091007433 antigens Proteins 0.000 description 2
- 208000010668 atopic eczema Diseases 0.000 description 2
- 210000001124 body fluid Anatomy 0.000 description 2
- 239000010839 body fluid Substances 0.000 description 2
- 238000004113 cell culture Methods 0.000 description 2
- 238000006243 chemical reaction Methods 0.000 description 2
- 238000012258 culturing Methods 0.000 description 2
- 230000006378 damage Effects 0.000 description 2
- 239000003085 diluting agent Substances 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 230000002496 gastric effect Effects 0.000 description 2
- 102000018358 immunoglobulin Human genes 0.000 description 2
- 239000002609 medium Substances 0.000 description 2
- 238000000034 method Methods 0.000 description 2
- 238000009629 microbiological culture Methods 0.000 description 2
- 210000004877 mucosa Anatomy 0.000 description 2
- 210000004400 mucous membrane Anatomy 0.000 description 2
- 239000002504 physiological saline solution Substances 0.000 description 2
- 230000000529 probiotic effect Effects 0.000 description 2
- 230000001105 regulatory effect Effects 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 230000008313 sensitization Effects 0.000 description 2
- 241000894007 species Species 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- RSYYQCDERUOEFI-UHFFFAOYSA-N 2-benzamido-5-(diaminomethylideneazaniumyl)pentanoate Chemical compound NC(N)=NCCCC(C(O)=O)NC(=O)C1=CC=CC=C1 RSYYQCDERUOEFI-UHFFFAOYSA-N 0.000 description 1
- 229920001817 Agar Polymers 0.000 description 1
- 206010002216 Anaphylactoid reaction Diseases 0.000 description 1
- 108090000145 Bacillolysin Proteins 0.000 description 1
- 108020004414 DNA Proteins 0.000 description 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 1
- 241000124008 Mammalia Species 0.000 description 1
- DEOKFPFLXFNAON-UHFFFAOYSA-N N-α-Benzoyl-DL-arginine 4-nitroanilide hydrochloride Chemical compound Cl.C=1C=C([N+]([O-])=O)C=CC=1NC(=O)C(CCCN=C(N)N)NC(=O)C1=CC=CC=C1 DEOKFPFLXFNAON-UHFFFAOYSA-N 0.000 description 1
- 102000035092 Neutral proteases Human genes 0.000 description 1
- 108091005507 Neutral proteases Proteins 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 230000001154 acute effect Effects 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 208000030961 allergic reaction Diseases 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- 238000003113 dilution method Methods 0.000 description 1
- 239000012636 effector Substances 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 239000012634 fragment Substances 0.000 description 1
- 230000006870 function Effects 0.000 description 1
- 230000009610 hypersensitivity Effects 0.000 description 1
- 210000002865 immune cell Anatomy 0.000 description 1
- 230000034435 immune system development Effects 0.000 description 1
- 230000006698 induction Effects 0.000 description 1
- 230000006799 invasive growth in response to glucose limitation Effects 0.000 description 1
- 238000002955 isolation Methods 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 210000004698 lymphocyte Anatomy 0.000 description 1
- 239000003550 marker Substances 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000000877 morphologic effect Effects 0.000 description 1
- 239000012466 permeate Substances 0.000 description 1
- 230000000270 postfertilization Effects 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 230000035755 proliferation Effects 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 230000028327 secretion Effects 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 238000012163 sequencing technique Methods 0.000 description 1
- 230000000638 stimulation Effects 0.000 description 1
- 238000003860 storage Methods 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 230000000451 tissue damage Effects 0.000 description 1
- 231100000827 tissue damage Toxicity 0.000 description 1
- 230000009092 tissue dysfunction Effects 0.000 description 1
- 108700024526 zebrafish sox32 Proteins 0.000 description 1
Images
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
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
- C12N1/205—Bacterial isolates
-
- 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
- A61K35/66—Microorganisms or materials therefrom
- A61K35/74—Bacteria
- A61K35/741—Probiotics
- A61K35/744—Lactic acid bacteria, e.g. enterococci, pediococci, lactococci, streptococci or leuconostocs
- A61K35/747—Lactobacilli, e.g. L. acidophilus or L. brevis
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P37/00—Drugs for immunological or allergic disorders
- A61P37/08—Antiallergic agents
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/225—Lactobacillus
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Organic Chemistry (AREA)
- Medicinal Chemistry (AREA)
- Microbiology (AREA)
- Animal Behavior & Ethology (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- Pharmacology & Pharmacy (AREA)
- Genetics & Genomics (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Wood Science & Technology (AREA)
- Zoology (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Biotechnology (AREA)
- Mycology (AREA)
- Biochemistry (AREA)
- Epidemiology (AREA)
- Molecular Biology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Pulmonology (AREA)
- Immunology (AREA)
- General Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Virology (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
Abstract
The invention discloses lactobacillus johnsonii LS04 and application thereof in preparing medicines for treating or preventing allergic diseases, and belongs to the technical field of microorganisms. The preservation number of the lactobacillus johnsonii LS04 is CGMCC No.22094. Lactobacillus johnsonii LS04 can remarkably inhibit compound 48/80 from stimulating zebra fish to secrete Tryptase in vivo, can inhibit mast cells or basophils degranulation in vivo, and has the potential of being applied to in vivo treatment or prevention of anaphylactic reaction. The lactobacillus johnsonii LS04 disclosed by the invention has a great potential application prospect in the aspect of treating or preventing anaphylactic reaction.
Description
Technical Field
The invention relates to the technical field of microorganisms, in particular to lactobacillus johnsonii LS04 and application thereof in preparing medicines for treating or preventing allergic diseases.
Background
Allergic reactions refer to reactions of tissue damage or dysfunction that occur in an immunized body when it is again stimulated with the same antigen. Worldwide, allergic people are increasing. Researches show that the ratio of the number of harmful bacteria to the number of probiotics in the gastrointestinal tract has great correlation with the occurrence of allergic diseases, and the probiotics in the gastrointestinal tract are more, so that the probiotics can protect the mucous membrane of the gastrointestinal tract, promote the proliferation of immunoglobulin, generate natural antibodies and enable immune cells to easily permeate the mucous membrane of the gastrointestinal tract; while the harmful bacteria in the gastrointestinal tract are more, the harmful bacteria easily damage the gastrointestinal mucosa, cause lymphocyte decrease and insufficient immunoglobulin types, however, the damage of the gastrointestinal mucosa can cause the increase of antigen amount, thereby causing sensitization to generate allergic diseases; thus, the bacterial species with which humans are exposed are sufficient to influence the trend of the immune system development, and it is desirable to further control the development of allergic reactions by adjusting the number and type of probiotics in the gastrointestinal tract.
The lack of powerful scientific research evidence of the function of probiotics seriously affects the popularization of probiotics and products thereof.
Accordingly, providing lactobacillus johnsonii LS04 and its use in the manufacture of a medicament for the treatment or prevention of allergic diseases is a problem to be solved by the person skilled in the art.
Disclosure of Invention
In view of this, the present invention provides lactobacillus johnsonii LS04 and its use in the preparation of a medicament for the treatment or prevention of allergic diseases.
Mast cells are the main effector cells of the anaphylactoid reaction, the number of activated mast cells being related to the degree of allergy, the more activated mast cells, the more severe the degree of allergy. Tryptase (Tryptase) is a pre-synthesized neutral protease in mast cells, is the most abundant medium, is released outside cells through degranulation of mast cells, has high selectivity in storage and expression in mast cells, and can be used as a marker for activation and degranulation of mast cells. The Tryptase is normally secreted only in small amounts into the body fluid, and in large amounts into the body fluid when the hypersensitivity causes degranulation of mast cells. The predictive accuracy of the acute allergic positive and negative reactions of Tryptase was 92.6% and 54.3%, respectively, and the half-life of the Tryptase was 2 hours, so that the expression level of the Tryptase was frequently used as one of the detection markers of the allergic reaction.
Mast cells of zebra fish, which are structurally and functionally similar to mammals, are involved in immune and allergic reactions in the body. N-benzoyl-DL-arginine paranitroanilide hydrochloride (BAPNA) is a substrate specific to the Tryptase, and the expression level of the Tryptase in the zebra fish can be detected. The sensitization and antiallergic effects of a substance can be quantitatively analyzed by detecting the expression level of the Tryptase in the zebra fish body within 24 hours after the substance induces the zebra fish body.
In order to achieve the above purpose, the present invention adopts the following technical scheme:
lactobacillus johnsonii LS04 has a collection number of CGMCC No.22094, and is preserved in China general microbiological culture Collection center, CGMCC for short, and the collection date of the institute of microorganisms of national academy of sciences of China, no.3, beijing area, chaoyang area, is 2021, and the collection date of the Lactobacillus johnsonii is 29 days, and is named as Lactobacillus johnsonii.
Further, the lactobacillus johnsonii LS04 is applied to the preparation of medicines for treating or preventing allergic diseases.
Further, the lactobacillus johnsonii LS04 is applied to the preparation of medicines for inhibiting mast cells or basophils degranulation.
Further, the lactobacillus johnsonii LS04 is a bacterial suspension.
Compared with the prior art, the invention discloses lactobacillus johnsonii LS04 and application thereof in preparing medicines for treating or preventing allergic diseases, wherein the lactobacillus johnsonii LS04 is obtained by separating and screening the faeces of long-life old people in the city of abaca county in Guangdong, the lactobacillus johnsonii LS04 can remarkably inhibit compound 48/80 from stimulating zebra fish to secrete Tryptase in vivo, can inhibit mast cells or basophils degranulation in vivo, has the potential of being applied to treating or preventing allergic reactions in vivo, and provides theoretical reference and guiding basis for developing probiotic preparations for pretreatment or prevention of allergic reactions by using lactobacillus johnsonii LS 04.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings that are required to be used in the embodiments or the description of the prior art will be briefly described below, and it is obvious that the drawings in the following description are only embodiments of the present invention, and that other drawings can be obtained according to the provided drawings without inventive effort for a person skilled in the art.
FIG. 1 is a drawing showing colony morphology of Lactobacillus johnsonii LS04 of the present invention on MRS agar plates;
FIG. 2 is a graph showing the effect of Lactobacillus johnsonii LS04 of the present invention on the expression level of compound 48/80 stimulated zebra fish to secrete Tryptase;
FIG. 3 is a graph showing the inhibition of compound 48/80 stimulated zebra fish to secrete Tryptase by Lactobacillus johnsonii LS04 of the present invention.
Detailed Description
The following description of the embodiments of the present invention will be made clearly and completely with reference to the accompanying drawings, in which it is apparent that the embodiments described are only some embodiments of the present invention, 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.
Sodium cromoglycate and N-benzoyl-DL-arginine p-nitroanilide hydrochloride (BAPNA) were purchased from Beijing Warce Shake chemical Co., ltd, compoud48/80 was purchased from Sigma Co., USA; lactobacillus johnsonii 33200 (ATCC 33200) was purchased from bio-technology limited of beijing Bai-o-borg.
EXAMPLE 1 isolation, identification and preservation of Lactobacillus johnsonii LS04
(1) Separating:
1) The feces (about 0.1 g) of the elder with long life are dissolved in a 1.5mL centrifuge tube filled with 1mL sterile physiological saline, and are fully blown and evenly mixed by a 1mL sterile gun head for standby.
2) Into each of 6 sterile 1.5mL centrifuge tubes, 900. Mu.L of sterile physiological saline was added.
3) Diluting the sample to 10 by adopting a gradient dilution method -2 、10 -3 、10 -4 、10 -5 、10 -6 、10 -7 。
4) From the package 10 -4 100 mu L of sample diluent is sucked into a centrifuge tube of the sample diluent and respectively inoculated on an MRS solid culture medium and a BS solid culture medium, and 100 mu L of bacterial liquid is spread and dried, and the coating method is mild, fast in action and needs to be operated near the flame of an alcohol lamp. After coating, the side of the dish was marked, including information on name, sample number, medium name, incubation time, dilution gradient, incubation conditions (anaerobic/aerobic), etc.
5) Repeating the above steps to obtain 10 -5 、10 -6 、10 -7 Dilution coating of dilution gradient.
6) After the coating, the dishes were cultured at 37℃under anaerobic conditions for 48 hours, and then were subjected to observation and recording.
7) Single colony on the plate is picked up by an inoculating loop and streaked into MRS solid culture medium, anaerobic culture is carried out for 48 hours at 37 ℃, and pure colony is obtained by separation.
8) Pure bacterial colony on the flat plate is inoculated in MRS liquid culture medium, anaerobic culture is carried out for 12-16 h at 37 ℃, 20% glycerol is added, and the flat plate is placed in a refrigerator at-80 ℃ for preservation.
(2) Molecular biological identification of strains: genomic DNA was extracted from the obtained strain, and a full-length fragment of 16S rDNA was amplified by PCR technique using the universal primers 27F and 1492R of 16S rDNA, followed by sequencing to identify the species of the strain.
The primer sequences of the universal primers 27F and 1492R are as follows:
27F：5’-AGAGTTTGATCCTGGCTCAG-3’；SEQ ID NO.1；
1492R：5’-GGTTACCTTGTTACGACTT-3’；SEQ ID NO.2。
experimental results: the strain screened from the faeces of the elderly in the city of the abaca county of Guangdong province is identified by morphological observation and 16S rDNA, wherein the strain LS04 is identified as Lactobacillus johnsonii, and the 16S rDNA sequence is shown as SEQ ID NO. 3.
TTAGACGGCTGACTCCTATAAAGGTTATCCCACCGGCTTTGGGTGTTACAGACTCTCATGGTGTGACGGGCGGTGTGTACAAGGCCCGGGAACGTATTCACCGCGGCGTGCTGATCCGCGATTACTAGCGATTCCAGCTTCGTGTAGGCGAGTTGCAGCCTACAGTCCGAACTGAGAACGGCTTTAAGAGATCCGCTTGCCTTCGCAGGTTCGCTTCTCGTTGTACCGTCCATTGTAGCACGTGTGTAGCCCAGGTCATAAGGGGCATGATGACTTGACGTCATCCCCACCTTCCTCCGGTTTGTCACCGGCAGTCTCATTAGAGTGCCCAACTTAATGATGGCAACTAATGACAAGGGTTGCGCTCGTTGCGGGACTTAACCCAA CATCTCACGACACGAGCTGACGACAGCCATGCACCACCTGTCTCAGCGTCCCCGAAGGGAACACCTAATCTCTTAGGTTTGCACTGGATGTCAAGACCTGGTAAGGTTCTTCGCGTTGCTTCGAATTAAACCACATGCTCCACCGCTTGTGCGGGCCCCCGTCAATTCCTTTGAGTTTCAACCTTGCGGTCGTACTCCCCAGGCGGAGTGCTTAATGCGTTAGCTGCAGCACTGAGAGGCGGAAACCTCCCAACACTTAGCACTCATCGTTTACGGCATGGACTACCAGGGTATCTAATCCTGTTCGCTACCCATGCTTTCGAGCCTCAGCGTCAGTTGCAGACCAGAGAGCCGCCTTCGCCACTGGTGTTCTTCCATATATCTACGCATTCCACCGCTACACATGGAGTTCCACTCTCCTCTTCTGCACTCAAGTTCAACAGTTTCTGATGCAATTCTCCGGTTGAGCCGAAGGCTTTCACATCAGACTTATTGAACCGCCTGCACTCGCTTTACGCCCAATAAATCCGGACAACGCTTGCCACCTACGTATTACCGCGGCTGCTGGCACGTAGTTAGCCGTGACTTTCTAAGTAATTACCGTCAAATAAAGGCCAGTTACTACCTCTATCTTTCTTCACTACCAACAGAGCTTTACGAGCCGAAACCCTTCTTCACTCACGCGGCGTTGCTCCATCAGACTTTCGTCCATTGTGGAAGATTCCCTACTGCTGCCTCCCGTAGGAGTTTGGGCCGTGTCTCAGTCCCAATGTGGCCGATCAGTCTCTCAACTCGGCTATGCATCATTGCCTTGGTAAGCCGTTACCTTACCAACTAGCTAATGCACCGCAGGTCCATCCAAGAGTGATAGCAGAACCATCTTTCAAACTCTAGACATGCGTCTAGTGTTGTTATCCGGTATTAGCATCTGTTTCCAGGTGTTATCCCAGTCTCTTGGGCAGGTTACCCACGTGTTACTCACCCGTCCGCCGCTCGCTTGTATCTAGTTTCATTTAGTGCAAGCACTAAAATCATCTAGGCAAGCTCGCTCGACTGCAG；SEQ ID NO.3。
The strain LS04 single colony is inoculated on MRS solid culture medium, grows well under the aerobic condition of 37 ℃, and has milky white, round shape, regular edge and smooth surface (figure 1). The strain LS04 is preserved in China general microbiological culture Collection center (CGMCC), the national institute of microbiology, national academy of sciences of China, no.3, the North Chen West Lu 1, the Korean area of Beijing, with a preservation date of 2021, 03 months and 29 days, and is classified and named as Lactobacillus johnsonii, with a preservation number of CGMCC No.22094.
EXAMPLE 2 preparation of Lactobacillus johnsonii LS04 bacterial suspension (thallus)
Inoculating lactobacillus johnsonii LS04 after activation culture in MRS liquid culture medium, culturing at 37 ℃ for 24 hours, and centrifuging at 4 ℃ for 10 minutes at 6000r/min to obtain bacterial precipitate; after the bacterial cell precipitate is washed twice by PBS, the bacterial cell is resuspended by PBS, and the cell concentration is regulated to be 1 multiplied by 10 6 CFU/mL gave a bacterial suspension (cell).
EXAMPLE 3 preparation of Lactobacillus johnsonii 33200 bacterial suspension (thallus)
Inoculating lactobacillus johnsonii 33200 after activation culture in MRS liquid culture medium, culturing at 37deg.C for 24 hr, and centrifuging at 4deg.C for 10min at 6000r/min to obtain bacterial precipitate; after the bacterial cell precipitate is washed twice by PBS, the bacterial cell is resuspended by PBS, and the cell concentration is regulated to be 1 multiplied by 10 6 CFU/mL gave a bacterial suspension (cell).
Example 4 Effect of Lactobacillus johnsonii LS04 on Compound 48/80 stimulation of Tryptase secretion in zebra fish
Healthy wild-type AB-line zebra fish that developed to 5dpf (days post fertilization) were selected and placed in 96-well cell culture plates, 10 strips/well. The experiment set was a normal group, a model group, an intervention group (positive control group, lactobacillus johnsonii 33200, lactobacillus johnsonii LS 04), and a solvent zeroing group, each group was provided with 6 duplicate wells. PBS was added to the normal group, PBS was added to the model group, sodium cromolyn solution (100. Mu.g/mL) was added to the positive control group, and Lactobacillus johnsonii 33200 was added to 1X 10 6 CFU/mL of Lactobacillus johnsonii 33200, lactobacillus johnsonii LS04 was added to 1X 10 6 CFU/mL Lactobacillus johnsonii LS04, solvent zeroing group (non zebra fish added)) Adding PBS (phosphate buffer solution) into 100 mu L of each hole, placing the mixture in a biochemical incubator for incubation at 28 ℃ for 24 hours, and replacing a new solution; after 48h incubation, 150. Mu.L PBS was added to the normal group, the model group, the positive control group, lactobacillus johnsonii 33200, lactobacillus johnsonii LS04, and solvent zeroing were added to each of the compound 48/80 (8. Mu.g/mL), 150. Mu.L solution was placed in 2mL centrifuge tubes after incubation at 28℃for 2h, 150. Mu.L solution was added to each of the centrifuge tubes, and 150. Mu.L BAPNA solution (20 mg/mL) was added to each of the centrifuge tubes, after incubation at 37℃for 48h, 200. Mu.L solution was placed in 96-well cell culture plates, and absorbance (OD) was measured at 405nm using a microplate reader. The expression level and inhibition rate of the Tryptase are expressed as follows:
SPSS 19.0 software was used to statistically process the data, the experimental data were all expressed as x+ -SD data, and analyzed by T-test, as compared to the normal group: #### p<0.001, compared to model group: * P (P)<0.05，****P<0.001。
The results of the expression level and the inhibition rate of the Tryptase are shown in fig. 2 and 3, and the results show that compared with the normal group (100.00+/-10.18%), the expression level of the Tryptase in the model group (349.76 +/-40.89%) is obviously increased, which indicates that the compound 48/80 induction allergy model is successfully established.
The positive control group (sodium cromolyn) showed a level of expression of 119.53.+ -. 9.18% and a level of inhibition of 92.18.+ -. 3.67% with a significant difference (P) compared to the model group (expression level of Tryptase: 349.76.+ -. 40.89%<0.001). Therefore, the cromolyn sodium has antiallergic effect, and is consistent with clinical results, which shows that the antiallergic test is effective. Lactobacillus johnsonii 33200 group (1X 10) 6 CFU/mL) the expression level of the Tryptase in the zebra fish was 297.00 ±19.25%, while the inhibition rate of the Tryptase was 21.12±7.71%, and the expression level of the Tryptase in the model group (expression level: 349.76 + -40.89%) is significantly different than the above (P)<0.05). In addition, lactobacillus johnsonii LS04 group (1X 10) 6 CFU/mL) of the expression level of the Tryptase in the zebra fish was 168.34 ±20.23%, while the inhibition rate of the Tryptase was 72.64 ±8.10%, and the expression level of the Tryptase in the model group (expression level: 349.76 + -40.89%) is significantly different than the above (P)<0.001 Shows good probiotic efficacy in inhibiting degranulation of mast cells or basophils. Thus, the above results indicate that at the same concentration, lactobacillus johnsonii LS04 has a stronger inhibitory effect on compound 48/80 to induce degranulation of mast cells or basophils in vivo than lactobacillus johnsonii 33200, and has the potential to treat or prevent allergic diseases.
The previous description of the disclosed embodiments is provided to enable any person skilled in the art to make or use the present invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein may be applied to other embodiments without departing from the spirit or scope of the invention. Thus, the present invention is not intended to be limited to the embodiments shown herein but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.
Claims (5)
1. Lactobacillus johnsonii LS04 is characterized by having a preservation number of CGMCC No.22094.
2. Use of lactobacillus johnsonii LS04 as claimed in claim 1 in the manufacture of a medicament for the treatment or prophylaxis of allergic diseases.
3. Use of lactobacillus johnsonii LS04 according to claim 2 for the manufacture of a medicament for the treatment or prophylaxis of allergic diseases, wherein said lactobacillus johnsonii LS04 is a bacterial suspension.
4. Use of lactobacillus johnsonii LS04 as claimed in claim 1 in the manufacture of a medicament for inhibiting degranulation of mast cells or basophils.
5. Use of lactobacillus johnsonii LS04 as claimed in claim 4 for the manufacture of a medicament for inhibiting degranulation of mast cells or basophils, wherein the lactobacillus johnsonii LS04 is a bacterial suspension.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202310249339.4A CN116333932A (en) | 2023-03-15 | 2023-03-15 | Lactobacillus johnsonii LS04 and application thereof in preparation of medicines for treating or preventing allergic diseases |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202310249339.4A CN116333932A (en) | 2023-03-15 | 2023-03-15 | Lactobacillus johnsonii LS04 and application thereof in preparation of medicines for treating or preventing allergic diseases |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN116333932A true CN116333932A (en) | 2023-06-27 |
Family
ID=86880046
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202310249339.4A Pending CN116333932A (en) | 2023-03-15 | 2023-03-15 | Lactobacillus johnsonii LS04 and application thereof in preparation of medicines for treating or preventing allergic diseases |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN116333932A (en) |
-
2023
- 2023-03-15 CN CN202310249339.4A patent/CN116333932A/en active Pending
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN114350578B (en) | Lactobacillus plantarum LP1Z for producing lysozyme and efficiently antagonizing multidrug-resistant helicobacter pylori and application thereof | |
| CN115975880B (en) | Lactobacillus mucilaginosus CYQ09 and application thereof | |
| CN106011040B (en) | One plant degradation Ofloxacin anthropi and its application | |
| CN112410269A (en) | Bacillus subtilis and application thereof in degradation of zearalenone | |
| CN114891675B (en) | Stomach-derived lactobacillus plantarum LPF-01 and application thereof | |
| CN113122485A (en) | Lactobacillus plantarum HNU082 with antagonistic effect and application thereof | |
| CN113186136B (en) | Multiple drug-resistant helicobacter pylori and application thereof | |
| Khisti et al. | Isolation and identification of Saccharomyces cerevisiae from caterpillar frass and their probiotic characterization | |
| CN112322553B (en) | Clostridium difficile resistant lactococcus lactis and application thereof | |
| CN110835620B (en) | Lactobacillus plantarum and application thereof | |
| CN116333932A (en) | Lactobacillus johnsonii LS04 and application thereof in preparation of medicines for treating or preventing allergic diseases | |
| CN115772487A (en) | Lactobacillus reuteri C2 and application thereof in preparation of monoamine oxidase inhibiting medicines | |
| CN113980843B (en) | Application of lactobacillus acidophilus LA-03 in preparation of helicobacter pylori resistant medicines | |
| CN113215045B (en) | Lactobacillus gasseri LGV03 and application thereof | |
| CN115992079A (en) | Lactobacillus fermentum E1 and application thereof in preparation of uric acid reducing drugs | |
| CN116463249A (en) | Lactobacillus salivarius LF01 and application thereof in preparation of medicines for treating or preventing allergic diseases | |
| CN115025130A (en) | Application of lactobacillus reuteri E9 in preparing medicine for treating or preventing allergic diseases | |
| CN116463250A (en) | Bacillus subtilis BS02 and application thereof in preparation of medicines for treating or preventing allergic diseases | |
| CN112322539B (en) | Enterococcus faecium R-NTR-1 from ocean and screening method and application thereof | |
| CN116059259A (en) | Application of lactobacillus fermentum E1 in preparation of medicines for treating or preventing allergic diseases | |
| CN116426417A (en) | Lactobacillus curvatus LC02 and application thereof in preparing medicines for treating or preventing allergic diseases | |
| CN116376758A (en) | Lactobacillus jensenii LS06 and application thereof in preparation of medicines for treating or preventing allergic diseases | |
| CN116179438A (en) | Pediococcus pentosaceus P1 and application thereof in preparing medicines for treating or preventing allergic diseases | |
| CN113308416B (en) | Lactobacillus plantarum capable of inhibiting kidney stone formation and application thereof | |
| CN115786197B (en) | Lactobacillus paracasei and preparation method and application of bacteriocin thereof, antibacterial peptide and identification method and application thereof |
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 |