CN116712463B - Application of bacillus coagulans MZY531 in preparation of anti-liver cancer drugs - Google Patents
Application of bacillus coagulans MZY531 in preparation of anti-liver cancer drugs Download PDFInfo
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- CN116712463B CN116712463B CN202310981306.9A CN202310981306A CN116712463B CN 116712463 B CN116712463 B CN 116712463B CN 202310981306 A CN202310981306 A CN 202310981306A CN 116712463 B CN116712463 B CN 116712463B
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- bacillus coagulans
- liver cancer
- mzy531
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Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
- A61K35/66—Microorganisms or materials therefrom
- A61K35/74—Bacteria
- A61K35/741—Probiotics
- A61K35/742—Spore-forming bacteria, e.g. Bacillus coagulans, Bacillus subtilis, clostridium or Lactobacillus sporogenes
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- 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
-
- 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
- C12R2001/07—Bacillus
-
- 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
Abstract
The application of bacillus coagulans MZY531 in preparing anti-liver cancer drugs relates to the research and development field of anti-liver cancer drugs. The bacillus coagulans MZY531 is preserved in China center for type culture Collection, with the preservation number of 2021, 6 and 2: cctccc NO: m2021662. Experiments prove that the bacillus coagulans MZY531 has the following functions: inducing apoptosis of H22 liver cancer cells of mice; inhibiting proliferation of H22 liver cancer cells of mice; reducing the activity of H22 liver cancer cells of mice; down-regulating the expression of PI3K/AKT/mTOR signal path related proteins in H22 liver cancer cells of mice; up-regulating the expression of Bax protein and Caspase-3 protein in H22 liver cancer cell of mouse; inhibit the expression of Bcl-2 protein in H22 liver cancer cells of mice. The invention lays an important foundation for the research of the bacillus coagulans MZY531 as an anti-liver cancer drug.
Description
Technical Field
The invention relates to the technical field of research and development of anti-liver cancer drugs, in particular to bacillus coagulansBacillus coagulans) Application of MZY531 in preparing anti-liver cancer drugs.
Background
Bacillus coagulans @Bacillus coagulans) Is a kind of gram positive bacteria forming spores and producing lactic acid. Bacillus coagulans has attracted considerable attention from researchers as an important probiotic with which people often come into contact in daily life. In addition to lactic acid production, the bacteria are capable of producing spores at the proper growth temperature and pH, while having high temperature resistance properties, and can grow at the pH of the stomach and intestinal tract. Bacillus coagulans promotes host health by having the effects of resisting oxidation, maintaining normal flora of the digestive tract, preventing enteritis, regulating immune response, and the like.
At present, chemical medicines are mostly used for treating various malignant tumors clinically. Although the chemical anticancer drugs have high drug effect, the long-term use of the chemical anticancer drugs can cause side effects such as anemia, immunity decline and the like, so that the research on the bacillus coagulans which is safe and beneficial to human health further digs the specific mechanism of inducing the apoptosis of liver cancer cells, and has important significance for developing the probiotics with the anti-liver cancer effect or the preparation thereof and clinically supplementing meal to assist in treating liver cancer. At present, researches on toxic effects and anti-tumor effects of bacillus coagulans on H22 liver cancer cells are not reported yet.
Disclosure of Invention
The invention aims to provide bacillus coagulansBacillus coagulans) Application of MZY531 in preparing anti-liver cancer drugs.
The technical scheme adopted by the invention for solving the technical problems is as follows:
bacillus coagulans of the inventionBacillus coagulans) Application of MZY531 in preparing anti-liver cancer drugs.
As a preferred embodiment, the bacillus coagulansBacillus coagulans) MZY531 was deposited with the chinese collection for typical cultures at 2021, month 6 and 2, accession number: cctccc NO: m2021662.
As a preferred embodiment, the bacillus coagulansBacillus coagulans) The functions of MZY531 include:
(1) Inducing apoptosis of H22 liver cancer cells of mice;
(2) Inhibiting proliferation of H22 liver cancer cells of mice;
(3) Reducing the activity of H22 liver cancer cells of mice;
(4) Down-regulating the expression of PI3K/AKT/mTOR signal path related proteins in H22 liver cancer cells of mice;
(5) Up-regulating the expression of Bax protein and Caspase-3 protein in H22 liver cancer cell of mouse;
(6) Inhibit the expression of Bcl-2 protein in H22 liver cancer cells of mice.
As a more preferred embodiment, the PI3K/AKT/mTOR signaling pathway related proteins in the mouse H22 hepatoma cell include: PI3K proteins, p-PI3K proteins, AKT proteins, p-AKT proteins, mTOR proteins, and p-mTOR proteins.
The beneficial effects of the invention are as follows:
the invention adopts bacillus coagulansBacillus coagulans）MZY531 treat H22 liver cancer cell of mouse, and determine activity of H22 liver cancer cell of mouse by CCK-8 method, and treat Bacillus coagulansBacillus coagulans) The MZY531 treated H22 liver cancer cell of the mouse is subjected to TUNEL staining, flow cytometry and Western blot method to detect apoptosis of the H22 liver cancer cell of the mouse, and the bacillus coagulans is discussedBacillus coagulans) mZY531 has therapeutic effects on inhibiting proliferation of mouse H22 liver cancer cells and inducing apoptosis of mouse H22 liver cancer cells. The above research results show that bacillus coagulansBacillus coagulans) MZY531 exerts its therapeutic effect by decreasing the viability of mouse H22 hepatoma cells, inhibiting the proliferation of mouse H22 hepatoma cells, and increasing apoptosis of mouse H22 hepatoma cells.
The invention also determines the bacillus coagulans by detecting the expression of PI3K/AKT/mTOR signal path related proteins PI3K, p-PI3K, AKT and p-AKT, mTOR, p-mTOR in the mouse H22 liver cancer cellsBacillus coagulans) The signal path activated by MZY531 in H22 hepatoma cell of mouse shows that bacillus coagulans @Bacillus coagulans) MZY531 can regulate apoptosis of mouse H22 hepatoma cells by down-regulating the levels of PI3K/AKT/mTOR signaling pathway related proteins (PI 3K, p-PI3K, AKT, p-AKT, mTOR, and p-mTOR) in mouse H22 hepatoma cells. In addition, the bacillus coagulans is determined by detecting the change of the total protein and the activated protein of the apoptosis signal path in the H22 hepatoma cell of the mouseBacillus coagulans) The possible mechanism of MZY531 to induce apoptosis of H22 liver cancer cell in mouse shows that bacillus coagulans @Bacillus coagulans) MZY531 up-regulates the expression of Bax protein and Caspase-3 protein, and obviously inhibits the expression of Bcl-2 protein, and by increasing bacillus coagulans @Bacillus coagulans) The dose of MZY531 can effectively induce apoptosis of H22 liver cancer cells of mice, and has obvious dose dependency.
The invention relates to bacillus coagulansBacillus coagulans) The MZY531 lays an important foundation for the research of anti-liver cancer drugs.
Drawings
FIG. 1 shows Bacillus coagulans of example 2Bacillus coagulans) Shadow of MZY531 on mouse H22 liver cancer cell viabilityAnd (5) sounding.
FIG. 2 shows apoptosis of H22 hepatoma cells of each group of mice under 20X field of view as measured by TUNEL method in example 2.
FIG. 3 shows the results of TUNEL staining quantitative analysis of each group in example 2.
FIG. 4 shows Bacillus coagulans of example 2Bacillus coagulans) Effects of MZY531 on the apoptosis cycle of mouse H22 hepatoma cells. In FIG. 3, Q1-Q4 represent necrotic cells, late apoptotic cells, viable cells, and early apoptotic cells, respectively.
FIG. 5 shows Bacillus coagulans of example 2Bacillus coagulans) Electropherograms of MZY531 on PI3K/AKT/mTOR signaling pathway related protein expression in mouse H22 hepatoma cells.
FIG. 6 shows Bacillus coagulans of example 2Bacillus coagulans) Effects of MZY531 on PI3K/AKT/mTOR signaling pathway related proteins p-PI3K/PI3K in mouse H22 hepatoma cells.
FIG. 7 shows Bacillus coagulans of example 2Bacillus coagulans) Effects of MZY531 on the PI3K/AKT/mTOR signaling pathway-associated protein p-AKT/AKT in mouse H22 hepatoma cells.
FIG. 8 shows Bacillus coagulans of example 2Bacillus coagulans) Effects of MZY531 on PI3K/AKT/mTOR signaling pathway related proteins p-mTOR/mTOR in mouse H22 hepatoma cells.
FIG. 9 shows Bacillus coagulans of example 2Bacillus coagulans) Electropherograms of MZY531 for Bax protein, bcl-2 protein and Caspase-3 protein expression in mouse H22 hepatoma cells.
FIG. 10 shows Bacillus coagulans of example 2Bacillus coagulans) Effects of MZY531 on Bax protein in mouse H22 hepatoma cells.
FIG. 11 shows Bacillus coagulans of example 2Bacillus coagulans) Effects of MZY531 on Bcl-2 protein in mouse H22 hepatoma cells.
FIG. 12 shows Bacillus coagulans of example 2Bacillus coagulans) Effects of MZY531 on Caspase-3 protein in mouse H22 hepatoma cells.
Detailed Description
The technical solutions of the embodiments of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention, and it is apparent that the described embodiments are only some embodiments of the present invention, 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.
The experimental methods in the following examples are conventional methods unless otherwise specified. The test materials used in the examples described below, unless otherwise specified, were purchased from conventional biochemical reagent stores. The quantitative tests in the following examples were all set up in triplicate and the results averaged.
Solid LB medium: the solvent is water, 10g/L of tryptone, 5g/L of yeast extract powder, 10g/L of sodium chloride and 15g/L of agar; ph=7.0 unless otherwise specified.
Liquid LB medium; the difference from the solid LB medium is only that no agar is added to the liquid LB medium.
Liquid GPY medium: 20g of glucose, 10g of tryptone, 10g of yeast extract powder, 5g of sodium chloride, 0.7g of calcium chloride and 0.3g of manganese chloride, and distilled water with the volume of 1000 mL, placing the materials in an autoclave at 121 ℃, sterilizing for 20min, taking out at 40 ℃, and placing the materials in a refrigerator at 4 ℃ for standby.
Tryptone and yeast extract: all purchased from Beijing Obocin Biotechnology Inc.
Sodium chloride, calcium chloride, manganese chloride and glucose: all purchased from the company of the light complex technology development limited of Tianjin.
Methanol: purchased from Tianjin Xintong fine chemical industry Co.
RPMI-1640 medium, phosphate Buffer (PBS), trypsin, mouse H22 liver cancer cells: all purchased from Jiangsu Kaiki Biotechnology Co., ltd.
Fetal Bovine Serum (FBS): purchased from the company of Hangzhou biotechnology, inc. on the Zhejiang day.
BCA kit and tissue lysate (RIPA): all purchased from beijing solebao technologies limited.
Protease inhibitor (PMSF): purchased from Jiangsu kang as a century biotechnology company.
Kit for rapidly preparing PAGE gel: purchased from Shanghai elegance Biotechnology Co.
Beta-actin, PI3K, p-PI3K, AKT, p-AKT, mTOR and p-mTOR: all purchased from beijing boaosen biotechnology limited.
Caspase-3 and Bax and Bcl-2: all from GeneTex, inc.
1. Isolation of Bacillus coagulans strains
Sample material selection: traditional fermented food pickled Chinese cabbage.
10 months in 2019, a fresh pickled Chinese cabbage sample is collected from the city of capital in Sichuan province, placed in a transport medium, quickly placed in an ice box and sent to a laboratory for strain separation. Placing a pickled Chinese cabbage sample in a water bath at 80 ℃ for 10min, grinding, carrying out gradient dilution, uniformly coating on a solid LB (LB) culture medium plate, culturing for 48h at 50 ℃, picking single bacterial colony, inoculating on the solid LB culture medium plate, continuously culturing, and repeatedly carrying out streak culture and purification on the solid LB culture medium plate to obtain a plurality of pure bacterial colonies. The pure cultured strain is inoculated into a liquid LB culture medium for culture, and then 60% glycerol is added for preservation in a refrigerator at-80 ℃. Of these, 1 strain was designated MZY531.
2. Identification of strains
Physiological and biochemical identification results of strain MZY 531: gram stain positive, catalase test negative, benzidine test negative, indole test negative, and acetyl methyl methanol test positive; starch is not hydrolyzed, gelatin is not liquefied, hydrogen sulfide is not generated, and acid and gas are not generated by fermenting glucose; a motionless bacillus; can grow at 25 ℃ and 65 ℃ and has the optimal growth temperature of 45-50 ℃; the pH is suitably 5.0-7.0; tolerance to 6.5% nacl; the strain MZY531 grows in a liquid LB culture medium in a uniform turbidity manner, and the strain is white in sedimentation after long-term storage.
16S rDNA sequence homology analysis: extracting genome DNA, adopting a primer pair consisting of 27F (SEQ ID NO: 2) and 1492R (SEQ ID NO: 3) to carry out PCR amplification, and then sequencing the amplified product, wherein the 16s rDNA sequence is shown as SEQ ID NO:1 of a sequence table.
The obtained 16S rDNA sequence is subjected to homology comparison and analysis by BLAST program and sequence information of known strains in GenBank library, and the strains to be detected are identified; genus classification was performed with a 16S rDNA sequence homology of greater than 99%.
According to the above identification result, strain MZY531 was identified as Bacillus coagulansBacillus coagulans。
3. Preservation of strains
The invention relates to a bacillus coagulans strainBacillus coagulans) MZY531 was preserved in the China center for type culture collection, abbreviated as CCTCC, at the address: in the Jiuqiu No. 299 university of Wuhan in Wuchang district of Wuhan, hubei province (Wuhan university collection), the preservation number is: cctccc NO: m2021662.
EXAMPLE 2 Bacillus coagulansBacillus coagulans) mZY531 inhibits proliferation of H22 liver cancer cell and induces apoptosis of H22 liver cancer cell.
1. Strain culture
Freezing and preserving bacillus coagulansBacillus coagulans) Inoculating MZY531 into liquid GPY culture medium, incubating at 50deg.C and 180rpm for 20 hr, centrifuging at 3000 rpm and 4deg.C for 10min, collecting bacterial precipitate, re-suspending the bacteria in sterile physiological saline, and adjusting bacterial concentration to 1.0X10 9 CFU/ml, stored at 4℃for further use.
2. Cell culture
Incubating H22 liver cancer cells of mice under the conditions of 95% relative humidity, 5% carbon dioxide and 37 ℃, wherein the culture medium is RPMI-1640 culture medium, and the RPMI-1640 culture medium contains 10% fetal bovine serum and 1% (penicillin and streptomycin mixture); changing the culture medium every 1-2 days; after the cell monolayer covered 80% of the surface of the cell culture plate, the supernatant broth was collected and the bottom was then washed with Phosphate Buffered Saline (PBS); the cells at the bottom are digested by trypsin, and the collected liquid is used for subculture or experiment; in all experiments, cells in the logarithmic phase were used.
3. Cell viability test
To verify bacillus coagulansBacillus coagulans) The influence of MZY531 on proliferation and activity of H22 hepatoma cells of mice was tested by CCK-8 method. Will be 1X 10 4 Inoculating H22 liver cancer cells of mice into 96-well plates, and respectively adding bacillus coagulans according to MOI=0, 1, 10, 50 and 100 CFU/cellBacillus coagulans) MZY531 was intervened, fluorouracil 5-FU (100. Mu.g/mL) was used as a positive control, and after 24 hours, 10% by volume of CCK-8 solution was added, incubated for 3 hours, and absorbance was measured at 450 nm.
Referring to FIG. 1, the results show that Bacillus coagulans @Bacillus coagulans) MZY531 significantly inhibited the growth of mouse H22 hepatoma cells and was dose dependent. Along with bacillus coagulansBacillus coagulans) The activity of the H22 hepatoma cells of the mice gradually decreases due to the increase of the concentration of MZY531. After mice H22 hepatoma cells were treated at MOI=100 CFU/cell concentration for 24H, the cell viability reached 52.74.+ -. 1.36%. Calculated bacillus coagulansBacillus coagulans) Half-inhibitory concentration value of MZY531 was moi=10 7 CFU/cell。
4. TUNEL staining analysis experiment
Study of Bacillus coagulans Using TUNEL staining analysisBacillus coagulans) Effects of MZY531 on mouse H22 hepatoma cells.
Mouse H22 liver cancer cells were grown at 2X 10 per well 5 The number of individual cells is inoculated into a 6-well plate in which a climbing plate is placed in advance, and bacillus coagulans is usedBacillus coagulans) MZY531 (moi=0, 50, 100 CFU/cell) and fluorouracil 5-FU (100 μg/mL) were separately intervened for 24h, and after three times of Phosphate Buffered Saline (PBS) washing the slide, cells were fixed with 4% paraformaldehyde for 30min; after fixation, the cells were washed 3 times with Phosphate Buffer (PBS), 50. Mu.L-100. Mu.L of 0.3% Triton X-100 membrane-breaking working solution was added, incubated at room temperature for 20min, and washed 3 times with Phosphate Buffer (PBS); dripping Buffer in TUNEL kit to cover cells after the climbing sheet is dried, and incubating for 10min at normal temperature; covering cells with mixed solution of TDT enzyme, dUTP and Buffer (the volume ratio of TDT enzyme, dUTP and Buffer is 1:5:50) in TUNEL kit, and incubating in a constant temperature incubator at 37deg.C for 2h; phosphate Buffer (PBS)) Washing for 3 times, each time for 5min, removing Phosphate Buffer (PBS) and dropwise adding DAPI (4', 6-diamidino-2-phenylindole) dye liquor for counterstaining, and incubating for 10min at room temperature in dark place; images were observed and collected under a fluorescence microscope. Referring to FIG. 2 (Sporange in FIG. 2 is CY3 fluorescein label), DAPI ultraviolet excitation wavelength is 330nm-380nm, emission wavelength is 420nm, and blue light is emitted; CY3 excitation wavelength is 510nm-560nm, emission wavelength is 590nm, and red light is emitted; the cell nucleus of the DAPI-dyed living cells is blue under the excitation of ultraviolet, the CY3 fluorescein marks the apoptotic cells and the cell nucleus is red; imageJ software counted TUNEL positive cell numbers.
Observing TUNEL staining results, nuclei of TUNEL positive cells were stained red, and the number of TUNEL positive cells reflected the extent of apoptosis. Referring to FIG. 3, as the MOI increases, the number of H22 hepatoma cells in mice gradually decreases, and the number of TUNEL positive cells increases, which indicates that Bacillus coagulans @Bacillus coagulans) MZY531 can induce apoptosis of H22 liver cancer cells of mice and inhibit proliferation of H22 liver cancer cells of mice.
5. Flow cytometry for detecting apoptosis
The H22 liver cancer cells of the mice are treated by single-hole 2×10 5 The number of individual cells was seeded in 6-well plates; bacillus coagulans is used as raw materialBacillus coagulans) MZY531 (moi=0, 50, 100 CFU/cell) and fluorouracil 5-FU (100 μg/mL) were each interfered with 24h, and the cells were digested with EDTA-free trypsin and centrifuged (4 ℃,1000×g,2 min); count 1×10 5 The mice H22 hepatoma cells of (2) were suspended in 100. Mu.L of 1 Xcombined buffer, stained with Annexin V/PI solution, and incubated at 37℃for 15 min; fluorescence emission spectra of 488 nm laser excitation at 530 nm (FITC channel) and 617nm (FITC channel) were analyzed by flow cytometry.
Referring to FIG. 4, the results of flow cytometry show that Bacillus coagulans @Bacillus coagulans) MZY531 can induce apoptosis of H22 liver cancer cells of mice; bacillus coagulans @Bacillus coagulans) Apoptosis rates of H22 hepatoma cells of mice after 24H treatment with MZY531 (moi=50 CFU/cell and 100 CFU/cell) were 25.53% and 42.39%, respectively, significantly higher than control (5.34%), and moi=100 CFU%The cell group was closer to the positive group 5-FU (47.59%).
6. Western blot
Taking mouse H22 liver cancer cells in logarithmic growth phase, and 3×10 each hole 5 The number of individual cells is inoculated into a 6-well plate and bacillus coagulans is used forBacillus coagulans) MZY531 (moi=0, 50, 100 CFU/cell) and fluorouracil 5-FU (100 μg/mL); after 24h, 100. Mu.L of a mixture of tissue lysate (RIPA), protease inhibitor (PMSF) and Phosphatase Inhibitor (PI) was added to lyse cells, wherein the volume ratio of tissue lysate (RIPA), protease inhibitor (PMSF) and Phosphatase Inhibitor (PI) was 100:1:1, protein was extracted, and lysed in an ice bath for 60 min, followed by centrifugation (10000 rpm,4 ℃ C., 10 min) using a low-temperature centrifuge, and finally the supernatant was collected for use, and the protein concentration was measured by BCA method. According to the instruction method of the BCA kit instruction, a 96-well plate method is selected, the absorbance of each tissue sample at 570 nm is measured by using an enzyme-labeled instrument, and the protein concentration of each tissue sample is calculated according to a standard curve; SDS-PAGE was performed and transferred to PVDF membrane, western blot analysis was performed using a primary anti-dilution of the following monoclonal antibodies: PI3K, p-PI3K, AKT, p-AKT, mTOR, p-mTOR, and Bcl-2 related death promoters Bax, caspase-3, and Bcl-2, were incubated with horseradish peroxidase-conjugated anti-rabbit secondary antibodies at 37℃for 1 h, and the bands of interest were quantified using a chemiluminescent imaging analyzer (Image Quant LAS 4000), beta-actin (beta-actin) as a loading control.
To determine bacillus coagulansBacillus coagulans) The invention detects the expression of PI3K/AKT/mTOR signal path related proteins PI3K, p-PI3K, AKT and p-AKT, mTOR, p-mTOR in the H22 hepatoma cell of the mouse by activating signal path of MZY531 in the H22 hepatoma cell of the mouse. Compared with the control group, bacillus coagulansBacillus coagulans) The expression of the phosphorylated proteins p-PI3K, p-AKT and p-mTOR was significantly reduced in MZY531 treated groups (FIG. 5). The phosphorylation levels of PI3K and AKT in mouse H22 hepatoma cells were also reduced (FIGS. 6-8), indicating Bacillus coagulans @Bacillus coagulans) MZY531 can pass throughRegulate the apoptosis of the mouse H22 liver cancer cells by regulating the content of PI3K/AKT/mTOR signal path related proteins (PI 3K, p-PI3K, AKT, p-AKT and p-mTOR) in the mouse H22 liver cancer cells.
To study bacillus coagulansBacillus coagulans) The invention detects the change of total protein and activated protein of apoptosis signal path in H22 liver cancer cell of mice. Referring to FIGS. 9-12, the results show that Bacillus coagulans @Bacillus coagulans) MZY531 up-regulates the expression of Bax protein and Caspase-3 protein, and remarkably inhibits the expression of Bcl-2 protein; in addition, low dosage of bacillus coagulansBacillus coagulans) MZY531 can promote apoptosis of mouse H22 hepatoma cells, but the effect is not particularly obvious; increase bacillus coagulansBacillus coagulans) The dose of MZY531 can effectively induce apoptosis of H22 liver cancer cells of mice, and has obvious dose dependency.
The invention discloses application of bacillus coagulans MZY531 in preparation of anti-liver cancer drugs, and a person skilled in the art can properly improve process parameters by referring to the content of the drugs. It is expressly noted that all such similar substitutions and modifications will be apparent to those skilled in the art, and are deemed to be included in the present invention. While the invention has been described with reference to preferred embodiments, it will be apparent to those skilled in the relevant art that variations and modifications can be made in the invention described herein without departing from the spirit or scope of the invention.
Claims (3)
1. The application of bacillus coagulans (Bacillus coagulans) MZY531 in preparing anti-liver cancer drugs is characterized in that the bacillus coagulans (Bacillus coagulans) MZY531 is preserved in China center for type culture Collection (China, with the preservation number: cctccc NO: m2021662.
2. The use according to claim 1, characterized in that the function of the bacillus coagulans (Bacillus coagulans) MZY531 is:
(1) Inducing apoptosis of H22 liver cancer cells of mice;
(2) Inhibiting proliferation of H22 liver cancer cells of mice;
(3) Reducing the activity of H22 liver cancer cells of mice;
(4) Down-regulating the expression of PI3K/AKT/mTOR signal path related proteins in H22 liver cancer cells of mice;
(5) Up-regulating the expression of Bax protein and Caspase-3 protein in H22 liver cancer cell of mouse;
(6) Inhibit the expression of Bcl-2 protein in H22 liver cancer cells of mice.
3. The use according to claim 2, wherein the PI3K/AKT/mTOR signaling pathway related proteins in the mouse H22 hepatoma cell are: PI3K proteins, p-PI3K proteins, AKT proteins, p-AKT proteins, mTOR proteins, and p-mTOR proteins.
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| CN108796027A (en) * | 2018-07-17 | 2018-11-13 | 杭州园泰生物科技有限公司 | A method of producing carotenoid |
| CN113444668A (en) * | 2021-07-26 | 2021-09-28 | 吉林省命之元生物科技有限公司 | Bacillus coagulans with hypoglycemic effect and application thereof |
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| CN113444668A (en) * | 2021-07-26 | 2021-09-28 | 吉林省命之元生物科技有限公司 | Bacillus coagulans with hypoglycemic effect and application thereof |
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