CN116731929A - Lactobacillus mucilaginosus ZS40 and application thereof - Google Patents

Lactobacillus mucilaginosus ZS40 and application thereof Download PDF

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CN116731929A
CN116731929A CN202310817035.3A CN202310817035A CN116731929A CN 116731929 A CN116731929 A CN 116731929A CN 202310817035 A CN202310817035 A CN 202310817035A CN 116731929 A CN116731929 A CN 116731929A
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张妤涵
王梦薇
赵欣
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Chongqing University of Education
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Abstract

The invention provides lactobacillus mucilaginosus ZS40 and application thereof, and belongs to the technical field of microorganisms. The lactobacillus mucilaginosus ZS40 is named as: lactobacillus fermentum Lactobacillus fermentum ZS, accession number: china general microbiological culture Collection center, preservation address: the national institute of microbiology, college of sciences, national institute of sciences, no. 3, national center for sciences, no.1, north chen west way, north, region of korea, beijing city, deposit number: CGMCC No.18226, date of preservation: 2019, 07, 15. The fermented lactobacillus mucilaginosus ZS40 can effectively improve the weight reduction and hematochezia condition of a C57 mouse induced by AOM-DSS, inhibit the NF- κB classical signal path, reduce the level of inflammatory factors and vascular cell adhesion factors in serum, reduce the inflammatory infiltration of colon tissues of the mouse, and have the effect of preventing and/or treating colon cancer.

Description

Lactobacillus mucilaginosus ZS40 and application thereof
Technical Field
The invention relates to the technical field of microorganisms, in particular to lactobacillus mucilaginosus ZS40 and application thereof.
Background
Colon cancer (Colon cancer) is a type of malignant disease that threatens human health and life, and has high morbidity and mortality. Colon cancer is second only to lung and prostate cancer in men with incidence of security, and third in women with incidence of lung and breast cancer. In China, colon cancer is a new case of up to 40 ten thousand per year, and the global annual new case is about 1000 ten thousand. Colon cancer may not have any symptoms in early stages, and may be manifested as abdominal distension, dyspepsia in middle and late stages, followed by changes in bowel movement habits, abdominal pain, or hematochezia. After tumor ulcer, hemophilation or toxin absorption, symptoms such as anemia, low fever, power, emaciation or edema of lower limbs are often caused. Because early colon cancer symptoms are not obvious, detection means for early colon cancer are not perfect, so that colon cancer is found, namely, a late stage result is obtained, and at present, part of patients can be treated in a surgical mode for treating colon cancer, the surgical risk is high, and meanwhile, the postoperative effect is not ideal.
The human colon micro-ecological system is complex and is an important metabolic organ. The number of viable bacteria per gram of the content in the intestinal tract is far more than 10 11 And even up to 10 14 And the total microbial biomass of the human body is 78 percent. In general, the intestinal flora may act to protect the colon, but when the flora is reduced by more than 50%, the ability to protect the colon from carcinogens is lost. Therefore, the development of new strains for preventing and treating colon cancer foods or medicines has important significance and great market value.
Disclosure of Invention
In view of the above, the invention aims to provide a lactobacillus fermentum ZS40 and application thereof, and the lactobacillus fermentum ZS40 provided by the invention can inhibit the NF- κB classical signal path, relieve colon tissue inflammation, and has the effect of preventing and/or treating colon cancer.
In order to achieve the above object, the present invention provides the following technical solutions:
a strain of lactobacillus mucilaginosus ZS40, said lactobacillus mucilaginosus ZS40 being designated as: lactobacillus fermentum Lactobacillus fermentum ZS, accession number: china general microbiological culture Collection center, preservation address: the national institute of microbiology, college of sciences, national institute of sciences, no. 3, national center for sciences, no.1, north chen west way, north, region of korea, beijing city, deposit number: CGMCC No.18226, date of preservation: 2019, 07, 15.
Preferably, the 16S rDNA gene sequence of the lactobacillus mucilaginosus ZS40 is shown as SEQ ID NO. 1.
Preferably, the lactobacillus mucilaginosus ZS40 is derived from traditional fermented yoghurt in Zhaosu county, xinjiang.
The invention also provides an application of the lactobacillus mucilaginosus ZS40 in preparing a food or a medicine for preventing colon cancer.
The invention also provides application of the lactobacillus mucilaginosus ZS40 in preparing a medicament for treating colon cancer.
The invention also provides an application of the lactobacillus mucilaginosus ZS40 in preparing a medicament for inhibiting the NF- κB classical signal path.
The invention also provides an application of the lactobacillus mucilaginosus ZS40 in preparing a medicament for relieving colon tissue inflammation.
Preferably, the medicament comprises one or two of a live strain of lactobacillus fermentum ZS40 and a metabolite of lactobacillus fermentum ZS40.
Preferably, the content of lactobacillus mucilaginosus ZS40 in the medicine is 10 9 CFU～10 11 CFU。
The beneficial technical effects are as follows: the invention provides a lactobacillus mucilaginosus ZS40 and application thereof, wherein the lactobacillus mucilaginosus ZS40 is named as: lactobacillus fermentum Lactobacillus fermentum ZS, accession number: china general microbiological culture Collection center, preservation address: the national institute of microbiology, college of sciences, national institute of sciences, no. 3, national center for sciences, no.1, north chen west way, north, region of korea, beijing city, deposit number: CGMCC No.18226, date of preservation: 2019, 07, 15. The fermented lactobacillus mucilaginosus ZS40 can effectively improve the weight reduction and hematochezia condition of a C57 mouse induced by AOM-DSS, inhibit the NF- κB classical signal path, reduce the level of inflammatory factors and vascular cell adhesion factors in serum, reduce the inflammatory infiltration of colon tissues of the mouse, and have the effect of preventing and/or treating colon cancer.
Drawings
FIG. 1 shows body weight changes in CRC mice from different treatment groups;
FIG. 2A is a colon histopathological section of CRC mice from different treatment groups; fig. 2B is a colon severity pathology score for CRC mice of different treatment groups, with a score of 0: the villi and the epithelium are complete; 1 marker: slight swelling and separation of submucosa or lamina propria; 2 markers: a moderate swelling separation of submucosa or lamina propria, plasma cell infiltration; 3 flags: severe swelling of submucosa or lamina propria, infiltration of plasma cells, local villus atrophy and shedding; 4, the following steps: the intestinal villi disappears, and the pathological score of the necrosis of the intestinal wall is more than or equal to 2 minutes, which is regarded as intestinal injury;
fig. 3 shows colon pathology (immunohistochemical staining), magnification 20×, a for CRC mice from different treatment groups: CD34; b: CD117;
FIG. 4 shows the expression of mRNA indicators in colon tissue of mice;
FIG. 5 shows the expression of colon tissue proteins from CRC mice from different treatment groups.
Detailed Description
The invention provides a lactobacillus mucilaginosus ZS40, wherein the lactobacillus mucilaginosus ZS40 is named as follows: lactobacillus fermentum Lactobacillus fermentum ZS, accession number: china general microbiological culture Collection center, preservation address: the national institute of microbiology, college of sciences, national institute of sciences, no. 3, national center for sciences, no.1, north chen west way, north, region of korea, beijing city, deposit number: CGMCC No.18226, date of preservation: 2019, 07, 15.
In the invention, the 16S rDNA gene sequence of the lactobacillus mucilaginosus ZS40 is shown as SEQ ID NO. 1. The specific nucleotide sequence of SEQ ID NO.1 is as follows:
SEQ ID NO.1：
GGCTGGCTCCTAAAGGTTACCCCACCGACTTTGGGTGTTACAAACTCTCATGGTGTGACGGGCGGTGTGTACAAGGCCCGGGAACGTATTCACCGCGGCATGCTGATCCGCGATTACTAGCGATTCCGACTTCGTGCAGGCGAGTTGCAGCCTGCAGTCCGAACTGAGAACGGTTTTAAGAGATTTGCTTGCCCTCGCGAGTTCGCGACTCGTTGTACCGTCCATTGTAGCACGTGTGTAGCCCAGGTCATAAGGGGCATGATGATCTGACGTCGTCCCCACCTTCCTCCGGTTTGTCACCGGCAGTCTCACTAGAGTGCCCAACTTAATGCTGGCAACTAGTAACAAGGGTTGCGCTCGTTGCGGGACTTAACCCAACATCTCACGACACGAGCTGACGACGACCATGCACCACCTGTCATTGCGTTCCCGAAGGAAACGCCCTATCTCTAGGGTTGGCGCAAGATGTCAAGACCTGGTAAGGTTCTTCGCGTAGCTTCGAATTAAACCACATGCTCCACCGCTTGTGCGGGCCCCCGTCAATTCCTTTGAGTTTCAACCTTGCGGTCGTACTCCCCAGGCGGAGTGCTTAATGCGTTAGCTCCGGCACTGAAGGGCGGAAACC CTCCAACACCTAGCACTCATCGTTTACGGCATGGACTACCAGGGTATCTAATCCTGTTCGCTACCCATGCTTTCGAGTCTCAGCGTCAGTTGCAGACCAGGTAGCCGCCTTCGCCACTGGTGTTCTTCCATATATCTACGCATTCCACCGCTACACATGGAGTTCCACTACCCTCTTCTGCACTCAAGTTATCCAGTTTCCGATGCACTTCTCCGGTTAAGCCGAAGGCTTTCACATCAGACTTAGAAAACCGCCTGCACTCTCTTTACGCCCAATAAATCCGGATAACGCTTGCCACCTACGTATTACCGCGGCTGCTGGCACGTAGTTAGCCGTGACTTTCTGGTTAAATACCGTCAACGTATGAACAGTTACTCTCATACGTGTTCTTCTTTAACAACAGAGCTTTACGAGCCGAAACCCTTCTTCACTCACGCGGTGTTGCTCCATCAGGCTTGCGCCCATTGTGGAAGATTCCCTACTGCTGCCTCCCGTAGGAGTATGGGCCGTGTCTCAGTCCCATTGTGGCCGATCAGTCTCTCAACTCGGCTATGCATCATCGCCTTGGTAGGCCGTTACCCCACCAACAAGCTAATGCACCGCAGGTCCATCCAGAAGTGATAGCGAGAAGCCATCTTTTAAGCGTTGTTCATGCGAACAACGCTGTTATGCGGTATTAGCATCTGTTTCCAAATGTTGTCCCCCGCTTCTGGGCAGGTTACCTACGTGTTACTCACCCGTCCGCCACTCGTTGGCGACCAAAATCAATCAGGTGCAAGCACCATCAATCAA。
in the invention, the lactobacillus mucilaginosus ZS40 is derived from traditional fermented yoghurt in Zhaosu county in Xinjiang; the experimental result of the lactobacillus tolerance artificial gastric juice shows that the capability of fermenting lactobacillus mucilaginosus ZS40 to resist artificial gastric juice reaches 79.3 percent.
In the invention, the lactobacillus tolerating artificial gastric juice experiment is as follows: a solution of 0.35% pepsin and 0.20% NaCl was prepared. The pH of the artificial gastric juice was adjusted to pH=3.0 with 1mol/L HCl and sterilized by filtration using a disposable sterile syringe and sterile filter head (0.22 μm filter). Taking 1mL of activated bacterial liquid, centrifuging at 3000rpm for 10min, and washing the bottom bacterial body for 2 times by using sterile physiological saline. Adding sterile physiological saline, resuspending thalli, mixing the evenly mixed bacterial liquid and the filtered and sterilized artificial gastric juice according to the volume ratio of 1:9, and culturing for 3 hours at 37 ℃ and 100 rpm. Plate counting method calculates viable counts of 0h and 3h, and calculates survival rate of experimental strain in gastric juice (ph=3.0).
The invention also provides an application of the lactobacillus mucilaginosus ZS40 in preparing a food or a medicine for preventing colon cancer.
In the present invention, the type of the food is not particularly limited, and a type acceptable in food is adopted by lactobacillus mucilaginosus ZS40. The preparation method of the food is not particularly limited, and the preparation method of the corresponding dosage form is adopted. In the present invention, the food preferably comprises one or two of a live strain of lactobacillus mucilaginosus ZS40 and a metabolite of lactobacillus mucilaginosus ZS40, and more preferably comprises a live strain of lactobacillus mucilaginosus ZS 40; the content of lactobacillus mucilaginosus ZS40 in the food is preferably 10 9 CFU～10 11 CFU, more preferably 10 11 CFU。
In the present invention, the dosage form of the drug is not particularly limited, and a pharmaceutically acceptable dosage form of lactobacillus mucilaginosus ZS40 is adopted. The preparation method of the medicine is not particularly limited, and the preparation method of the corresponding dosage form is adopted.
The invention also provides application of the lactobacillus mucilaginosus ZS40 in preparing a medicament for treating colon cancer.
In the present invention, the dosage form, the preparation method and the content of the active ingredient of the medicine are the same as those described above, and are not described herein. The fermented lactobacillus mucilaginosus ZS40 can effectively improve the weight reduction and hematochezia condition of a C57 mouse induced by AOM-DSS, inhibit the NF- κB classical signal path, reduce the level of inflammatory factors and vascular cell adhesion factors in serum, reduce the inflammatory infiltration of colon tissues of the mouse, and have the effect of preventing and/or treating colon cancer.
The invention also provides an application of the lactobacillus mucilaginosus ZS40 in preparing a medicament for inhibiting the NF- κB classical signal path.
In the present invention, the dosage form, the preparation method and the content of the active ingredient of the medicine are the same as those described above, and are not described herein. NF- κB signaling pathways regulate key processes in the development and progression of various types of cancer. NF- κb is probably one of the most common regulators of cancer due to its widely involved target genes and tissues. In colon cancer, activation of the NF-. Kappa.B classical pathway eventually leads to increased expression of NF-. Kappa.B target genes such as IL-8, bcl-2, cox-2. The high expression of pro-inflammatory factors in turn will further stimulate the classical NF- κb pathway, affecting cell proliferation or turnover. Thus, NF- κb may lead to the occurrence of colonitis-associated colon cancer by maintaining a sustained inflammatory process of the intestinal mucosa. The expression of IκB kinase protein activated by lactobacillus mucilaginosus ZS40 blocks the important pathway of NF- κB classical pathway and reduces the level of target proteins IL-1 beta, TNF-alpha, cox-2, etc.
The invention also provides an application of the lactobacillus mucilaginosus ZS40 in preparing a medicament for relieving colon tissue inflammation.
In the present invention, the dosage form, the preparation method and the content of the active ingredient of the medicine are the same as those described above, and are not described herein. The fermented lactobacillus mucilaginosus ZS40 can reduce the expression level of cytokines TNF-alpha, MIP-1 beta, MIP-2 and VCAM-1, reduce the expression level of CD34 and CD117 and relieve colon tissue inflammation.
For a better understanding of the present invention, the following examples are further illustrated, but are not limited to the following examples. Materials, reagents and the like used in the examples and test examples of the present invention can be obtained commercially unless otherwise specified; the methods used in the examples and test examples of the present invention are conventional methods unless otherwise specified.
EXAMPLE 1 isolation and characterization of Lactobacillus mucilaginosus ZS40
1. Separation and purification of total lactic acid bacteria
A sample of yogurt was obtained from traditional fermentation in Zhaosu county, xinjiang.
1.1 rejuvenation of lactic acid bacteria
Mixing the samples uniformly, taking 10mL to 100mL of 10% sterilized skim milk of the fermented yogurt sample in a sterile environment, standing and culturing at 37 ℃ for 24-72 h, fermenting to a semi-solid state, and stopping fermentation.
1.2 separation culture by plate streaking
Mixing and rejuvenating fermented milk, taking 1mL of the fermented milk into a sterile test tube, adding 9mL of 0.9wt% sterile physiological saline, soaking and mixing uniformly, carrying out gradient dilution by adopting a 10-time dilution method, absorbing diluent with proper dilution, and separating single bacterial colony by adopting a five-zone streak method.
1.3 Strain purification
Under the aseptic condition, according to morphological characteristics, typical colonies with different morphologies on an MRS-agar plate are picked, streaked, separated and purified on an MRS-agar culture dish, and cultured for 24-72 hours at 37 ℃. And (3) determining whether the strain is purified or not through colony morphology and microscopic observation, and preserving the purified strain as an experimental strain.
1.4 isolation of lactic acid bacteria
Under aseptic conditions, single colonies on MRS-agar plates are picked, gram staining is carried out, and whether the shapes of the bacteria are single or not is observed through an oil lens.
1.5 16S rDNA molecular biological characterization
(1) Extracting the genome of the strain: separating by streaking on MRS-agar plate, culturing for 24-72 h, picking single colony, activating in MRS-broth culture medium for three times, and culturing at 37deg.C for 24h according to 1% inoculum size. The bacterial liquid was centrifuged (8000 rpm,10min,4 ℃) and the medium was washed off with PBS to collect the bacterial cells, and the total DNA of the strain was extracted according to the bacterial genome extraction kit instructions.
(2) Amplification of DNA: the extracted total DNA is used as a template and amplified by a PCR method. Amplified PCR fragments were visualized by gel electrophoresis.
(3) And (3) strain identification: the extracted total DNA was sent to Shanghai Biotechnology for total DNA sequence analysis.
Finally separating to obtain the lactobacillus mucilaginosus ZS40.
Example 2 Effect of Lactobacillus mucilaginosus ZS40 on NF- κB signaling pathway in AOM/DSS-induced colon cancer mouse model
2 experimental materials and methods
2.1 Strain activation and cultivation
Lactobacillus mucilaginosus ZS40 and Bulgaria strains are respectively inoculated into MRS liquid culture medium in an inoculum size of 2 percent, and are cultured for 24 hours in a shaking table at a constant temperature of 37 ℃ and at a constant speed of 100 rpm. The bacterial liquid was centrifuged at 12000rpm for 10min, the supernatant was discarded, and the bacterial liquid was resuspended in 0.9% physiological saline, and the purity of the bacterial liquid was examined by gram staining. Diluting lactobacillus mucilaginosus ZS40 bacterial liquid to 10 by adopting a gradient dilution method 11 CFU and 10 9 CFU, lactobacillus bulgaricus diluted to 10 9 CFU is ready for use.
2.2 laboratory animals
Male C57BL/6J mice purchased from Chongqing medical university (4 weeks old, weight 16 g-18 g) were fed at room temperature for 12h/12h of light/dark period and standard rodent feed (Jiangsu province collaborative medical bioengineering Co., ltd.) and purified water. After 7d of acclimation, animals were randomly assigned to 6 groups, 20 in each of the Normal (NC), model (CRC), ZS40 high dose (ZS 40-H), ZS40 low dose (ZS 40-L), bulgaria fluid (BLA), and drug control (SD).
The NC group and the CRC group are treated by the gastric lavage of 0.2mL physiological saline every day; ZS40-H group, ZS40-L group and BLA group were each treated with 0.2mL of the same compound at a concentration of 10 11 CFU、10 9 CFU and 10 11 The bacterial liquid of the CFU is subjected to gastric lavage treatment to provide the number of viable bacteria ingested daily; SD group was treated by gavage with 0.2mL daily of 2.5% sulfasalazine solution. After 7d, azomethane (AOM) reagent was intraperitoneally injected at a dose of 10mg/kg, and fed for 7d, during which purified water was supplied. After 7d, the purified water was replaced with 2.5wt% Dextran Sodium Sulfate (DSS) solution, and after 7d, the purified water was replaced with purified water 14d againThe 3 cycles were repeated according to the drinking conditions.
At the end of the experimental period (week 12), all animals were sacrificed. Taking whole blood, centrifuging to obtain serum, rapidly cutting liver and colon, photographing, sampling, immediately freezing in liquid nitrogen, and storing in a refrigerator at-80 ℃ for later use.
2.3 histological observations of animals
A histological examination of colon tissue samples was performed for each animal. These tissue samples were fixed in tissue fixative (Chongqing Seville Biotechnology Co., ltd., chongqing, china), fixed at 4℃and paraffin sections were prepared by feeding them to Chongqing Seville BioCo., ltd. Within 24 hours to obtain hematoxylin and eosin (H & E) stained and CD34, CD117 immunohistochemical stained sections, and section observations were made using a positive microscope (Osbalin).
2.4 serum ELISA
For serum preparation, blood collected from the eyeball was allowed to stand at 4℃for 1 hour and centrifuged at 3000rpm for 15min. The levels of interleukin 1 beta (IL-1 beta), interleukin 8 (IL-8), tumor necrosis factor-alpha (TNF-alpha), macrophage inflammatory protein 1 beta (MIP-1 beta), macrophage inflammatory protein 2 (MIP-2), vascular endothelial cell adhesion molecule 1 (VCAM-1) were measured using a commercial ELISA kit (Shanghai ELISA kit, inc., china).
2.5 real-time immunofluorescence PCR assay
mRNA expression of IL-1β, TNF- α, P65, TRAF1/2, IκBα, TRAF-6, IKKα, IKKKβ, bcl-XL, bcl-2, and Cox-2 was determined by RT-qPCR. Total RNA was extracted and cDNA reverse transcribed according to the requirements of RNA extraction kit instructions (Soy Biotechnology Co., beijing, china) and cDNA reverse transcription kit (ASP, shanghai, china), and quantified by measuring absorbance at 260nm using a UV-2401PC micro-spectrophotometer. RT-qPCR was then performed in a StepOne Plus PCR (Sieimer fly), normalized with actin (actin), and final RT-qPCR product expression usingMethod calculation. The primer sequences are shown in Table 1.
TABLE 1 primer sequences
2.6Western-Blot analysis
Liver and colon samples were homogenized with pre-chilled RIPA buffer (zemoeid) (PMSF, protease inhibitor, phosphatase inhibitor have been mixed) and then centrifuged at 12,000xg for 20min at 4 ℃. Protein concentration was determined with BCA protein assay kit (Yeasen, shanghai, china). For western blot analysis, 50 μg of protein extract was separated by 10% NuPAGE (zemoer), then transferred onto PVDF membrane. Blocking with 5% skim milk at 28℃for 1h, incubating PVDF membrane with anti-IL-1β, anti-TNF- α, anti-IκBα, anti-P65, anti-COX-2, anti-Actin (all antibodies were purchased from Invitrogen), washing with 1 XTBE 5 times, then incubating with HRP-second anti-ibody (purchased from Invitrogen) for 1h at room temperature, washing PVDF membrane with 1 XTBE 5 times. Antibody binding was observed by enhanced chemiluminescent ECL (solebao, beijing). Protein expression was quantified using ImageJ software (NIH).
2.7 data analysis
Data are expressed as mean ± Standard Deviation (SD). Analysis was performed using Graph Pad 7.0 and SPSS 21.0 statistical software packages. Differences between the mean values of the individual groups were evaluated using one-way anova and the Duncan multi-range test, with p values <0.05 considered statistically significant.
3. Results
3.1 body weight and bowel movement of CRC mice from different treatment groups
The body weight changes of the CRC mice from the different treatment groups are shown in fig. 1 and table 2. As can be seen from fig. 1 and table 2, the final body weight of mice in the bacterial fluid-treated group was significantly higher than that in the CRC group (p < 0.05). The final body weights of the ZS40-H, ZS40-L, BLA and SD groups were approximated to NC (p < 0.05). All AOM-DSS treatments significantly reduced the final body weight of mice (p < 0.05) compared to NC groups. In addition, the final body weights of mice in ZS40-H, ZS40-L, BLA, and SD groups were also significantly higher than those in CRC group (24.39.+ -. 0.7 g) (p < 0.05), 28.38.+ -. 0.8g, 25.33.+ -. 0.7g, 25.61.+ -. 0.7g, and 27.62.+ -. 1.0g, respectively.
TABLE 2 body weight changes (unit: g) of CRC mice from different treatment groups
The hematochezia of the experimental mice was observed and recorded during the experiment, and the results are shown in table 3. As can be seen from Table 3, from week 4 after AOM drug injection, the blood pattern was increased in the CRC group mice, and after each administration of DSS aqueous solution, the blood pattern was aggravated. The bacterial liquid intervention treatment combination and the drug treatment group can delay the occurrence of the phenomenon, wherein the phenomenon is most obvious in ZS40-H and SD groups. This suggests that ZS40 was able to reduce AOM-DSS-induced weight loss and hematochezia in C57 mice.
TABLE 3 hematochezia conditions (Unit: only) of CRC mice of different treatment groups
3.2 serum indicators of CRC mice from different treatment groups
Serum indices of CRC mice from different treatment groups are shown in table 4. As can be seen from Table 4, the levels of IL-1β, IL-8, TNF- α, MIP-1β, MIP-2 and VCAM-1 were significantly increased in the serum of mice in the CRC group compared to NC mice. Indicating that colon cancer can cause these cytokines and inflammation
ZS40-H intervention reduced serum levels of IL-1β, IL-8, TNF- α, MIP-1β, MIP-2 and VCAM-1 in CRC mice as compared to CRC. In addition, mice in ZS40-L, BLA intervention and SD control groups effectively reduced IL-1β, IL-8, TNF- α, MIP-1β, MIP-2 and VCAM-1 levels compared to CRC groups, SD showed better effects in reducing inflammatory factor levels (p < 0.05). This suggests that ZS40 was able to reduce inflammatory and vascular cell adhesion factor levels in serum of AOM-DSS induced C57 mice.
TABLE 4 serum indices of CRC mice from different treatment groups
Note that: the letter difference on the same column indicates p <0.05, and the difference is significant.
3.3 State of colon tissue of CRC mice of different treatment groups
The colon histopathological sections of the CRC mice from the different treatment groups are shown in FIG. 2A. As can be seen from fig. 2A, inflammatory factors accumulate in large amounts in colon cells of CRC group, and ZS40-H group significantly reduced the accumulation of inflammatory factors in colon tissue of CRC mice. In addition, the ZS40-L group, the BLA group and the SD group also significantly reduced inflammatory infiltration of the colon tissue of mice compared to the CRC group. Colon severity pathology scores for the different treatment groups of CRC mice are shown in figure 2B. As can be seen from fig. 2B, the pathological score of the CRC group was significantly higher than that of the NC group, and the ZS40-H group and the SD group significantly reduced the pathological score of the colon severity than that of the CRC group.
The CD34, CD117 expression for the CRC mice of the different treatment groups is shown in figure 3. As can be seen from fig. 3, the expression level of CD34 and CD117 in the colon tissue samples of the mice in the CRC group was significantly increased compared to the NC group, and the expression level of CD34 and CD117 in the ZS40-H group, the ZS40-L group, the BLA group and the SD group was decreased, wherein the decrease in the ZS40-H group and the SD group was more remarkable.
3.4 Effect of different treatment groups on colon tissue mRNA expression of CRC mice
The colon tissue mRNA expression of the CRC mice from the different treatment groups is shown in FIG. 4. As can be seen from FIG. 4, ZS40-H and ZS40-L groups have the strongest ability to reduce NF-. Kappa.B, IL-1β, TNF-. Alpha., TRAF-6, NEMO, IKKKα/β, IκBα, TRAF2, p100, bcl-xL, A20, cox-2 expression in CRC mice. ZS40-H group and ZS40-L group reduce the expression level of I kappa B kinase protein and target protein IL-8, bcl-2 and Cox-2, and block NF-kappa B classical pathway.
3.5 Effect of different treatment groups on CRC mouse colon tissue protein expression
The expression of colon tissue proteins from CRC mice from different treatment groups is shown in FIG. 5. As can be seen from FIG. 5, the expression levels of p65, IL-1. Beta., TNF-. Alpha.and COX-2 were elevated in the colon tissue of the mice in the CRC group as compared to the NC group, while the expression levels of these protein factors were reduced in the ZS40-H group, the ZS40-L group, the BLA group and the SD group, with the reduction effect being most remarkable in the ZS40-H group and the SD group; in addition, protein expression of IκBα was shown to be down-regulated in the CRC group, with varying degrees of up-regulated expression tendencies in the remaining control groups, with ZS40-H and SD groups being most pronounced.
In summary, lactobacillus mucilaginosus ZS40 can inhibit the classical NF- κB signaling pathway, reduce the occurrence of colon cancer tumor, and inhibit the NF- κB signaling pathway at both high and low doses, but the inhibition of proinflammatory factor by ZS40 at high dose and the regulation of key proteins in signaling pathway are more obvious, which may be related to the regulation of intestinal microbiota of the organism after lactobacillus ZS40 is ingested into the body.
The foregoing is merely a preferred embodiment of the present invention and it should be noted that modifications and adaptations to those skilled in the art may be made without departing from the principles of the present invention, which are intended to be comprehended within the scope of the present invention.

Claims (9)

1. A strain of lactobacillus mucilaginosus ZS40, characterized in that the lactobacillus mucilaginosus ZS40 is named: lactobacillus fermentum Lactobacillus fermentum ZS, accession number: china general microbiological culture Collection center, preservation address: the national institute of microbiology, college of sciences, national institute of sciences, no. 3, national center for sciences, no.1, north chen west way, north, region of korea, beijing city, deposit number: CGMCC No.18226, date of preservation: 2019, 07, 15.
2. The lactobacillus mucilaginosus ZS40 according to claim 1, characterized in that the 16S rDNA gene sequence of the lactobacillus mucilaginosus ZS40 is shown in SEQ ID No. 1.
3. Lactobacillus mucilaginosus ZS40 according to claim 1, characterized in that the lactobacillus mucilaginosus ZS40 is derived from traditionally fermented yoghurt in singapore county.
4. Use of lactobacillus mucilaginosus ZS40 according to any of claims 1 to 3 for preparing a food or a medicament for preventing colon cancer.
5. Use of lactobacillus mucilaginosus ZS40 according to any of claims 1 to 3 for the preparation of a medicament for the treatment of colon cancer.
6. Use of lactobacillus mucilaginosus ZS40 according to any of claims 1 to 3 for the preparation of a medicament for inhibiting NF- κb classical signaling pathway.
7. Use of lactobacillus mucilaginosus ZS40 according to any of claims 1 to 3 for the preparation of a medicament for alleviating inflammation of colon tissue.
8. The use according to any one of claims 4 to 7, wherein the medicament comprises one or both of a live strain of lactobacillus fermentum ZS40, a metabolite of lactobacillus fermentum ZS40.
9. The use according to any one of claims 4 to 7, wherein the content of lactobacillus mucilaginosus ZS40 in the medicament is 10 9 CFU～10 11 CFU。
CN202310817035.3A 2023-07-04 2023-07-04 Lactobacillus mucilaginosus ZS40 and application thereof Pending CN116731929A (en)

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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN117535175A (en) * 2023-10-12 2024-02-09 善恩康生物科技（苏州）有限公司 Composite probiotics and application thereof in preparation of products for preventing or assisting in treating colorectal cancer
CN117736942A (en) * 2024-02-20 2024-03-22 山东中科嘉亿生物工程有限公司 Fermented lactobacillus mucilaginosus JYLF-315 for improving skin aging and metagen preparation and application thereof

Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN117535175A (en) * 2023-10-12 2024-02-09 善恩康生物科技（苏州）有限公司 Composite probiotics and application thereof in preparation of products for preventing or assisting in treating colorectal cancer
CN117535175B (en) * 2023-10-12 2024-05-31 善恩康生物科技（苏州）有限公司 Composite probiotics and application thereof in preparation of products for preventing or assisting in treating colorectal cancer
CN117736942A (en) * 2024-02-20 2024-03-22 山东中科嘉亿生物工程有限公司 Fermented lactobacillus mucilaginosus JYLF-315 for improving skin aging and metagen preparation and application thereof
CN117736942B (en) * 2024-02-20 2024-05-31 山东中科嘉亿生物工程有限公司 Fermented lactobacillus mucilaginosus JYLF-315 for improving skin aging, and metagen preparation and application thereof

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