WO2022134658A1 - 一株能够预防、缓解银屑病的短双歧杆菌及其应用 - Google Patents

一株能够预防、缓解银屑病的短双歧杆菌及其应用 Download PDF

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WO2022134658A1
WO2022134658A1 PCT/CN2021/117116 CN2021117116W WO2022134658A1 WO 2022134658 A1 WO2022134658 A1 WO 2022134658A1 CN 2021117116 W CN2021117116 W CN 2021117116W WO 2022134658 A1 WO2022134658 A1 WO 2022134658A1
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ccfm1078
bifidobacterium breve
psoriasis
breve
mice
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French (fr)
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陈卫
陆文伟
邓雅丹
翟齐啸
杨波
崔树茂
毛丙永
王鸿超
赵建新
张灏
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江南大学
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    • 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
    • A61K35/745—Bifidobacteria
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
    • A23C9/00—Milk preparations; Milk powder or milk powder preparations
    • A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L19/00—Products from fruits or vegetables; Preparation or treatment thereof
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L2/00—Non-alcoholic beverages; Dry compositions or concentrates therefor; Their preparation
    • A23L2/385—Concentrates of non-alcoholic beverages
    • A23L2/39—Dry compositions
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • A23L33/135—Bacteria or derivatives thereof, e.g. probiotics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A61P17/06—Antipsoriatics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • 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
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23V2400/00—Lactic or propionic acid bacteria
    • A23V2400/51—Bifidobacterium
    • A23V2400/519—Breve

Definitions

  • the invention relates to a Bifidobacterium breve capable of preventing and relieving psoriasis and its application, belonging to the technical field of microorganisms and medical technology.
  • T helper cell 17 is a newly discovered subgroup of T cells that can secrete interleukin-17 (IL-17), which has a role in autoimmune diseases and the body's defense response. Significance. After naive CD4 + T cells receive antigen stimulation, they can differentiate into different subtypes of T cells under different conditions and perform different functions. The differentiation direction is regulated by various factors such as the nature of the antigen, hormones and cytokines in the local environment, among which the type of cytokines and the balance between cytokines play an important role in regulating the differentiation of Th cells.
  • IL-23 is a member of the IL-12 family and is a heterodimer, including the IL-23-specific p19 subunit and the p40 subunit shared with IL-12.
  • the corresponding IL-23 receptor is also a heterodimer, composed of two subunits, IL-12R ⁇ 1 and IL-23R.
  • IL-23R is mainly expressed in T cells, natural killer T cells, monocytes and dendritic cells.
  • IL-23 plays an important role in the differentiation of Th17 cells and can directly induce naive T cells to produce IL-22 and IL-17.
  • the IL ⁇ 23/Th17 axis is involved in the occurrence of various diseases, including psoriasis, arthritis, and inflammatory bowel disease.
  • Psoriasis is a chronic, genetically and environmentally induced, immune-mediated inflammatory disease that is non-contagious, prone to relapse, and may progressively worsen with age or vary in severity , and some cases even life-long unhealed.
  • the global prevalence of psoriasis released by the World Health Organization in 2016 was 0.09%-11.43%. In China, the prevalence of psoriasis is less than 0.5%.
  • the prevalence of psoriasis appears to be higher in countries with higher latitudes, which may be related to different UV intensities caused by different latitudes. Psoriasis is more common in adults than in children. Although onset can occur at any age, the highest incidence is between the ages of 18-39 or 50-69.
  • psoriasis The pathogenesis of psoriasis is not fully understood. Increased release of pro-inflammatory cytokines from immune-related cells and chronic activation of the innate and adaptive immune systems have been suggested as mechanisms leading to long-term damage to multiple tissues and organs. A variety of psoriasis-related abnormalities have been reported, including antigen presentation, activation of NF- ⁇ B signaling, differentiation of helper T cell populations (especially TH17 cells, the main source of IL-17), and IL-17 Response enhancement, these abnormalities promote the host's immune response and immune cell infiltration. Liu Aimin, Qiao Ju, Guo Qinghao, etc. all reported that the expression levels of IL-23, IL-22, and IL-17 in the serum of patients with psoriasis were increased.
  • Psoriasis currently does not have a complete cure for medical methods and drugs, which is still a worldwide medical problem. Treatment for psoriasis depends on several factors. In the early stage, local treatment was mostly used. The lesion area determines the formula and dosage of local treatment. Corticosteroids are the main method for local treatment of psoriasis. When traditional treatment regimens are ineffective, biologics can be considered. In recent years, a variety of high-efficiency biological agents can be used to treat moderate and severe psoriasis, including tumor necrosis factor (TNF) inhibitors, T cell regulators, IL-12/23p40 inhibitors and IL-17A inhibitors. Recent studies have emphasized the importance of IL-23 and IL-17, so the research focus is biased towards biologics targeting IL-23 and IL-17. But biologics are expensive.
  • TNF tumor necrosis factor
  • Probiotics are a class of active microorganisms that are beneficial to the host by colonizing the human body and changing the composition of the flora in a certain part of the host.
  • the metabolites of probiotics are important mediators connecting the host and intestinal flora, and have biological effects. For example, butyrate can reduce the level of IL-17 by promoting the differentiation of Treg cells, thereby affecting the balance of Th17/Treg. Relevant foreign studies have also confirmed this possibility. Chen et al.
  • probiotics that can inhibit the release of IL-23/Th17 axis-related inflammatory factors, so as to solve the current situation of psoriasis that is difficult to treat, easy to repeat, and has large side effects.
  • the present invention provides Bifidobacterium breve CCFM1078 or products containing Bifidobacterium breve CCFM1078 in inhibiting the release of IL-23/Th17 axis-related inflammatory factors, preventing and/or alleviating psoriasis. use.
  • the Bifidobacterium breve CCFM1078 has been deposited in the Guangdong Provincial Microbial Culture Collection Center on May 6, 2020, the preservation number is GDMCC No. 61011, and the preservation address is 5th Floor, Building 59, No. 100, Xianlie Middle Road, Guangzhou City.
  • the alleviation of psoriasis comprises the use of (a) or (b) at least one of:
  • the inhibiting the release of IL-23/Th17 axis-related inflammatory factors is specifically: reducing the level of IL-23, IL-22 and IL-17 in the skin of a psoriasis-like individual.
  • the product comprises food, medicine or health food.
  • the viable count of Bifidobacterium breve CCFM1078 in the product is not less than 1 ⁇ 10 6 CFU/mL or 1 ⁇ 10 6 CFU/g.
  • the medicine contains Bifidobacterium breve CCFM1078, a pharmaceutical carrier and/or a pharmaceutical excipient.
  • the drug carrier comprises microcapsules, microspheres, nanoparticles and liposomes.
  • the pharmaceutical excipients include excipients and additives.
  • the pharmaceutical excipients comprise anti-adhesives, penetration enhancers, buffers, plasticizers, surfactants, antifoaming agents, thickeners, inclusion agents, absorbents, humectants, solvents , propellant, solubilizer, cosolvent, emulsifier, colorant, pH adjuster, binder, disintegrant, filler, lubricant, wetting agent, integration agent, osmotic pressure regulator, stabilizer, auxiliary Flow agents, flavoring agents, preservatives, foaming agents, suspending agents, coating materials, fragrances, diluents, flocculants and deflocculants, filter aids and release retarders.
  • the additive comprises microcrystalline cellulose, hydroxypropyl methylcellulose, and refined lecithin.
  • the dosage form of the medicine comprises granules, capsules, tablets, pills or oral liquids.
  • the food is a food containing Bifidobacterium breve CCFM1078 or a fermented metabolite thereof.
  • the food is a dairy product, soy product or fruit and vegetable product produced by using Bifidobacterium breve CCFM1078 or a starter containing Bifidobacterium breve CCFM1078.
  • the food product is a solid beverage containing Bifidobacterium breve CCFM1078.
  • the preparation method of the starter is as follows: Bifidobacterium breve CCFM1078 is inoculated into the medium according to the inoculum amount of 2-4% of the total mass of the medium, and cultured at 37° C. for 30 hours to obtain a culture medium. Centrifuge the culture solution to collect the bacteria; wash the bacteria three times with a phosphate buffer with a pH of 7.2 and then resuspend it with a freeze-drying protective agent to obtain a resuspension; freeze the resuspension by vacuum freezing Dried to obtain a starter of Bifidobacterium breve CCFM1078.
  • the mass ratio of the freeze-drying protective agent and the bacterial cells is 2:1.
  • the medium is prepared by dissolving 10% skim milk, 0.5% glucose, 1.5% tryptone and 0.3% yeast extract in water based on the total mass of the medium.
  • the pH of the medium is 6.8.
  • the present invention provides a strain of Bifidobacterium breve CCFM1078 that inhibits the release of IL-23/Th17 axis-related inflammatory factors, thereby preventing and/or alleviating psoriasis.
  • CCFM1078 When it is used in psoriasis-like mice, the following effects can be achieved Effect:
  • Bifidobacterium breve CCFM1078 has great application prospects in the preparation of products for preventing and/or alleviating psoriasis (such as food, medicine or health food, etc.).
  • Bifidobacterium breve CCFM1078 has been deposited in the Guangdong Provincial Microbial Culture Collection Center on May 6, 2020, the preservation number is GDMCC No.61011, and the preservation address is Building 59, Yard, No. 100, Middle Xianlie Road, Guangzhou 5th Floor.
  • Figure 1 Lesion skin conditions of different groups of experimental mice.
  • Figure 2 Pathological sections of the skin of different groups of experimental mice.
  • Figure 3 IL-23 content in the skin of different groups of experimental mice.
  • Figure 4 IL-22 content in the skin of different groups of experimental mice.
  • Figure 5 IL-17 content in the skin of different groups of experimental mice.
  • the detection method of the number of viable bacteria adopt the national standard "GB 4789.35-2016 National Food Safety Standard for Food Microbiology Detection of Lactic Acid Bacteria".
  • Acidity detection method using the national standard GB 431334-2010.
  • Bifidobacterium breve B. breve 1 is another strain isolated from different stool samples using the same method.
  • Bifidobacterium breve CCFM1078 was inserted into mMRS solid medium and cultivated for 48 hours at 37°C, and its colonies were observed and observed under a microscope. Campylobacter species are slightly irregular, rounded-ended, usually singly, in pairs, and in small clusters.
  • Bifidobacterium breve CCFM1078 was inserted into MRS liquid medium and cultured at 37°C for 30 hours, then transferred to fresh MRS liquid medium, cultured under the same conditions for 30 hours, centrifuged at 6000 ⁇ g for 15 minutes, and washed with 0.9% saline. After the cells were centrifuged again at 6000 ⁇ g for 10 min, the cells were collected, resuspended with 30% (m/v) sucrose solution, and frozen at 80°C for use.
  • Bifidobacterium breve B.breve 1 was prepared by the same method as above.
  • Example 2 Relief effect of Bifidobacterium breve on skin lesions of mice with psoriasis
  • the experimental period was 3 weeks in total, and the model was established in the third week.
  • the back of the mice was depilated the day before the modeling, with an area of about 2.5cm ⁇ 2.5cm.
  • imiquimod cream 10 mg on the ear and 62.5 mg on the back, was applied to the ear and back hair removal areas of the mice in the model group and the experimental group every day, while the normal group only applied the same amount of Vaseline.
  • the methotrexate group was gavaged with 1 mg/kg/day of methotrexate solution (the methotrexate solution is a solution prepared by dissolving methotrexate in sterile physiological saline).
  • CCFM1078 group was given 0.2 mL of CCFM1078 bacterial suspension (prepared according to the method of Example 1) with a viable count of 1 ⁇ 10 9 CFU/mL every day, and B.breve 1 group was given 0.2 mL of viable count every day.
  • B.breve 1 bacterial suspension prepared according to the method of Example 1) of 1 ⁇ 10 9 CFU/mL, the normal group and the model group were only given the same amount of sterile normal saline as a control, and all groups were free to drink water and feeding. The back of the mice was photographed and recorded every day during the modeling period, and the mice were sacrificed on the first day of the fourth week. The results are shown in Figure 1.
  • the skin of the back depilation area of the mice in the normal group is smooth, without scales and erythema; while the skin of the back depilation area of the mice in the model group is wrinkled, covered with obvious scales, and accompanied by erythema; while the CCFM1078 group is compared with the model
  • the skin of the dorsal depilation area was smooth, basically free of scales and erythema, which was basically similar to the skin condition of the methotrexate group; while the skin of the dorsal depilation area of the mice in the B.breve 1 group was the same as the model group, with a wrinkled feeling and obvious covering Scaly.
  • Example 3 Inhibitory effect of Bifidobacterium breve on keratin thickening in mice with psoriasis
  • the 6-8-week-old SPF grade BALB/c female mice were divided into 4 groups, namely the normal group, the model group and the experimental group.
  • the B.breve 1 group of Fidobacteria B.breve 1, 6 animals in each group were raised in the Experimental Animal Center of Jiangnan University, fed with common feed, constant temperature 21-26°C, humidity 40-70%, noise less than or equal to 60dB, animal illumination 15-20LX (all animal experimental procedures were reviewed and approved by the Animal Welfare and Ethics Management Committee of Jiangnan University).
  • the experimental period was 3 weeks in total, and the model was established in the third week.
  • the back of the mice was depilated the day before the modeling, with an area of about 2.5cm ⁇ 2.5cm.
  • imiquimod cream 10 mg on the ear and 62.5 mg on the back, was applied to the ear and back hair removal areas of the mice in the model group and the experimental group every day, while the normal group only applied the same amount of Vaseline.
  • CCFM1078 group was given 0.2 mL of CCFM1078 bacterial suspension (prepared according to the method of Example 1) with a viable count of 1 ⁇ 10 9 CFU/mL every day, and B.breve 1 group was given daily by gavage 0.2mL of B.breve 1 bacterial suspension (prepared according to the method of Example 1) with a viable bacterial count of 1 ⁇ 10 9 CFU/mL, the normal group and the model group were only given the same amount of sterile normal saline as a control, all All groups were allowed to drink water and food ad libitum. The mice were sacrificed on the first day of the 4th week, and part of the skin of the dorsal depilation area of the mice was taken to make pathological sections for histopathological analysis. The results are shown in Figure 2.
  • the skin epidermis layer of the mice in the blank group is composed of only one or two layers of cells, and no inflammatory reaction is seen in each layer of the epidermis; while the mice in the model group have significantly thickened keratin and severe inflammation; Compared with the model group, the phenomenon of keratin thickening was not obvious, and the inflammation was mild, which was basically similar to the skin condition of the methotrexate group; while the skin keratin thickening of the mice in the B.breve 1 group was obvious, and the inflammation was serious.
  • Example 4 Effect of Bifidobacterium breve on IL-23 in the skin of psoriatic mice
  • the 6-8-week-old SPF grade BALB/c female mice were divided into 4 groups, namely the normal group, the model group and the experimental group.
  • the B.breve 1 group of Fidobacteria B.breve 1, 6 animals in each group were raised in the Experimental Animal Center of Jiangnan University, fed with common feed, constant temperature 21-26°C, humidity 40-70%, noise less than or equal to 60dB, animal illumination 15-20LX (all animal experimental procedures were reviewed and approved by the Animal Welfare and Ethics Management Committee of Jiangnan University).
  • the experimental period was 3 weeks in total, and the model was established in the third week.
  • the back of the mice was depilated the day before the modeling, with an area of about 2.5cm ⁇ 2.5cm.
  • imiquimod cream 10 mg on the ear and 62.5 mg on the back, was applied to the ear and back hair removal areas of the mice in the model group and the experimental group every day, while the normal group only applied the same amount of Vaseline.
  • the CCFM1078 group was given 0.2 mL of CCFM1078 bacterial suspension with a viable count of 1 ⁇ 10 9 CFU/mL every day
  • the B.breve 1 group was given 0.2 mL of B with a viable count of 1 ⁇ 10 9 CFU/mL every day.
  • the normal group and the model group were only given the same amount of sterile normal saline as a control, and all groups were given free access to water and food.
  • mice were sacrificed on the first day of the 4th week, and part of the skin of the dorsal depilation area of the mice was taken and stored at -80°C, and the content of IL-23 in the skin was detected by ELISA kit. The results are shown in Figure 3.
  • Bifidobacterium breve CCFM1078 can significantly down-regulate the typical up-regulated pro-inflammatory factors in psoriatic mice to normal levels, especially the level of IL-23, which is significantly better than another B. breve strain B.breve 1 .
  • Example 5 Effect of Bifidobacterium breve on IL-22 in the skin of psoriatic mice
  • the 6-8-week-old SPF grade BALB/c female mice were divided into 4 groups, namely the normal group, the model group and the experimental group.
  • the B.breve 1 group of Fidobacteria B.breve 1, 6 animals in each group were raised in the Experimental Animal Center of Jiangnan University, fed with common feed, constant temperature 21-26°C, humidity 40-70%, noise less than or equal to 60dB, animal illumination 15-20LX (all animal experimental procedures were reviewed and approved by the Animal Welfare and Ethics Management Committee of Jiangnan University).
  • the experimental period was 3 weeks in total, and the model was established in the third week.
  • the back of the mice was depilated the day before the modeling, with an area of about 2.5cm ⁇ 2.5cm.
  • imiquimod cream 10 mg on the ear and 62.5 mg on the back, was applied to the ear and back hair removal areas of the mice in the model group and the experimental group every day, while the normal group only applied the same amount of Vaseline.
  • the CCFM1078 group was given 0.2 mL of CCFM1078 bacterial suspension with a viable count of 1 ⁇ 10 9 CFU/mL every day
  • the B.breve 1 group was given 0.2 mL of B with a viable count of 1 ⁇ 10 9 CFU/mL every day.
  • .breve 1 bacterial suspension, the normal group and the model group were only given the same amount of sterile normal saline as a control, and all groups were given free access to water and food.
  • the mice were sacrificed on the first day of the 4th week, and part of the skin of the dorsal depilation area of the mice was taken and stored at -80°C, and the content of IL-22 in the skin was detected by ELISA kit.
  • Bifidobacterium breve CCFM1078 can significantly down-regulate the typical up-regulated pro-inflammatory factors in psoriatic mice to normal levels, especially the level of IL-22, which is significantly better than another strain of B. breve B.breve 1 .
  • Example 6 Effect of Bifidobacterium breve on IL-17 in the skin of psoriatic mice
  • the 6-8-week-old SPF grade BALB/c female mice were divided into 4 groups, namely the normal group, the model group and the experimental group.
  • the B.breve 1 group of Fidobacteria B.breve 1, 6 animals in each group were raised in the Experimental Animal Center of Jiangnan University, fed with common feed, constant temperature 21-26°C, humidity 40-70%, noise less than or equal to 60dB, animal illumination 15-20LX (all animal experimental procedures were reviewed and approved by the Animal Welfare and Ethics Management Committee of Jiangnan University).
  • the experimental period was 3 weeks in total, and the model was established in the third week.
  • the back of the mice was depilated the day before the modeling, with an area of about 2.5cm ⁇ 2.5cm.
  • imiquimod cream 10 mg on the ear and 62.5 mg on the back, was applied to the ear and back hair removal areas of the mice in the model group and the experimental group every day, while the normal group only applied the same amount of Vaseline.
  • CCFM1078 group was gavaged with 0.2 mL of CCFM1078 bacterial suspension every day
  • B.breve 1 group was gavaged with 0.2 mL of B.breve 1 bacterial suspension every day
  • the normal group and model group were only gavaged with the same amount of sterile normal saline.
  • all groups were given free access to water and food.
  • the mice were sacrificed on the first day of the 4th week, and part of the skin of the dorsal depilation area of the mice was taken and stored at -80°C. The content of IL-17 in the skin was detected by an ELISA kit. The results are shown in Figure 5 .
  • Bifidobacterium breve CCFM1078 can significantly down-regulate the typical up-regulated pro-inflammatory factors in psoriasis mice to normal levels, especially the level of IL-17, which is significantly better than another B. breve strain B.breve 1 .
  • Example 7 Preparation of bacterial powder containing Bifidobacterium breve CCFM1078
  • the seed liquid of Bifidobacterium breve CCFM1078 stored in the bacteria preservation tube was inoculated into mMRS medium according to the inoculum amount of 3% of the total mass of the medium, and cultured at 37 °C for 30 h to obtain a culture medium; the culture medium was centrifuged to collect bacteria.
  • Example 8 Preparation of yogurt containing Bifidobacterium breve CCFM1078
  • Example 9 Comparison of Bifidobacterium breve CCFM1078 and Bifidobacterium infantis 35624
  • Bifidobacterium breve CCFM1078 reduced the levels of Th17/IL-23 axis-related inflammatory factors in the skin lesions of psoriasis-like mice, specifically IL-23, IL-22, and IL-17, and Bifidobacterium infantis ( Bifidobacterium infantis) 35624 targets different sites.
  • Bifidobacterium breve CCFM1078 has a significant inhibitory effect on the thickening of the stratum corneum in the skin lesions of psoriasis-like mice, and effectively improves the erythema and scaling symptoms of the skin lesions. Therefore, the relieving effect of Bifidobacterium breve CCFM1078 on psoriasis is different from that of Bifidobacterium infantis (Bifidobacterium infantis) 35624.

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Abstract

一株能够预防、缓解银屑病的短双歧杆菌及其应用，属于微生物技术领域以及医药技术领域。提供了短双歧杆菌(Bifidobacterium breve)CCFM1078在抑制IL‐23/Th17轴相关炎症因子释放，及缓解银屑病方面的新用途。短双歧杆菌CCFM1078，能够缓解银屑病样小鼠病变皮肤、抑制皮肤表皮层增厚，使皮肤中IL-23水平、IL-22水平和IL-17水平分别降低20.3％、22.0％和18.3％；在制备预防和/或治疗银屑病的产品(如食品、药品或保健食品等)中，具有巨大的应用前景。

Description

一株能够预防、缓解银屑病的短双歧杆菌及其应用 技术领域
本发明涉及一株能够预防、缓解银屑病的短双歧杆菌及其应用，属于微生物技术领域以及医药技术领域。
背景技术
辅助性T细胞17(T helper cell 17,Th17)是一种新发现的能够分泌白介素-17(interleukin-17，IL-17)的T细胞亚群，在自身免疫性疾病和机体防御反应中具有重要的意义。初始CD4 +T细胞接受抗原刺激后，在不同的条件下可分化成不同亚型的T细胞，执行不同的功能。其分化方向受抗原的性质、局部环境中的激素及细胞因子等多种因素的调控，其中细胞因子的种类和细胞因子之间的平衡对Th细胞的分化具有重要的调节作用。IL-23属于IL-12家族成员，为异二聚体，包括IL-23特异性的p19亚单位和与IL-12共享的p40亚单位。IL-23受体相应的也为异二聚体，由IL-12Rβ1和IL-23R两个亚单位组成。IL-23R主要表达在T细胞、自然杀伤T细胞、单核细胞和树突细胞。IL-23在Th17细胞的分化中具有重要作用，可直接诱导原始T细胞产生IL-22和IL-17。IL‐23/Th17轴参与了多种疾病的发生，包括银屑病、关节炎、炎性肠病等。
银屑病是一种慢性的、遗传和环境共同诱发、免疫介导的炎症性疾病，无传染性，易复发，可能会随着年龄的增长而病情逐渐恶化，或者严重程度上时好时坏，有的病例甚至终身不愈。世界卫生组织2016年发布的银屑病全球患病率为0.09％-11.43％。在中国，银屑病患病率小于0.5％。高纬度国家的银屑病患病率似乎更高，这可能与纬度不同引起的紫外线强度不同有关。相比儿童，银屑病更易在成年人中发病。虽然可在任意年龄阶段发病，但在18-39岁或50-69岁之间的发病率最高。银屑病发病机制尚未完全明确。免疫相关细胞释放的促炎性细胞因子增多以及先天性和适应性免疫系统的慢性激活被认为是导致多个组织和器官长期受损的机制。目前多种银屑病相关的异常已被报道，包括抗原提呈、NF-κB信号通路的激活、辅助性T细胞群(尤其是IL-17的主要来源的TH17细胞)的分化和IL-17反应的增强，这些异常促进了宿主的免疫应答和免疫细胞的浸润。刘爱民、乔菊、郭庆浩等均报道了在银屑病患者血清内IL-23、IL-22、IL-17表达水平升高。
银屑病目前没有彻底治愈的医疗方法和药物，这仍然是一个世界性医疗难题。银屑病的治疗方法取决于几个因素。早期多采用局部治疗，病变的区域决定了局部治疗的配方和剂量，皮质类固醇是目前局部治疗银屑病的主要方法。当传统治疗方案疗效不佳时，可考虑应用生 物制剂。近年来,多种高效生物制剂可用来治疗中、重度银屑病,主要包括肿瘤坏死因子(TNF)抑制剂、T细胞调节剂、IL-12/23p40抑制剂及IL-17A抑制剂。近年来的研究强调了IL-23和IL-17的重要性,因此研究重点偏向了针对IL-23和IL-17的生物制剂。但生物制剂价格高昂。
益生菌是通过定殖在人体内，改变宿主某一部位菌群组成的一类对宿主有益的活性微生物。益生菌的代谢产物短链脂肪酸是联系宿主和肠道菌群的重要中介物质，具有生物学效应。如丁酸，可通过促进Treg细胞分化，进而影响Th17/Treg平衡，从而降低IL-17水平。国外相关研究也证实了该种可能，Chen等人用戊糖乳杆菌(Lactobacillus pentosus)GMNL-77干预银屑病样小鼠，结果显示，该戊糖乳杆菌降低小鼠皮肤内IL-23、IL-22、IL-17水平,且小鼠病情得到缓解。David等人发现26个银屑病患者连续口服婴儿双歧杆菌(Bifidobacterium infantis)35624 6周后，患者病情得到缓解。
因此，或许可通过开发能够抑制IL‐23/Th17轴相关炎症因子释放的益生菌，从而解决银屑病难治疗、易反复、副作用大的现状。
发明内容
本发明提供了短双歧杆菌(Bifidobacterium breve)CCFM1078或含短双歧杆菌(Bifidobacterium breve)CCFM1078的产品在抑制IL‐23/Th17轴相关炎症因子释放，预防和/或缓解银屑病方面的新用途。所述短双歧杆菌CCFM1078已于2020年5月6日保藏于广东省微生物菌种保藏中心，保藏编号为GDMCC No.61011，保藏地址为广州市先烈中路100号大院59号楼5楼。
在一种实施方式中，所述缓解银屑病包括(a)或(b)至少一种用途：
(a)有效改善皮肤褶皱、鳞屑和/或红斑症状，
(b)抑制皮肤角质增厚。
在一种实施方式中，所述抑制IL‐23/Th17轴相关炎症因子释放具体为：降低银屑病样个体皮肤中IL-23水平、IL-22水平和IL-17水平。
在一种实施方式中，所述产品包含食品、药品或保健食品。
在一种实施方式中，所述产品中短双歧杆菌CCFM1078的活菌数不低于1×10 6CFU/mL或1×10 6CFU/g。
在一种实施方式中，所述药品含有短双歧杆菌CCFM1078、药物载体和/或药用辅料。
在一种实施方式中，所述药物载体包含微囊、微球、纳米粒以及脂质体。
在一种实施方式中，所述药用辅料包含赋形剂以及附加剂。
在一种实施方式中，所述药用辅料包含抗黏合剂、渗透促进剂、缓冲剂、增塑剂、表面活性剂、消泡剂、增稠剂、包合剂、吸收剂、保湿剂、溶剂、抛射剂、增溶剂、助溶剂、乳化剂、着色剂、pH值调节剂、黏合剂、崩解剂、填充剂、润滑剂、润湿剂、整合剂、渗透压调节剂、稳定剂、助流剂、矫味剂、防腐剂、发泡剂、助悬剂、包衣材料、芳香剂、稀释剂、絮凝剂与反絮凝剂、助滤剂以及释放阻滞剂。
在一种实施方式中，所述附加剂包含微晶纤维素、羟丙基甲基纤维素以及精制卵磷脂。
在一种实施方式中，所述药品的剂型包含颗粒剂、胶囊剂、片剂、丸剂或口服液。
在一种实施方式中，所述食品为含有短双歧杆菌CCFM1078或其发酵代谢物的食品。
在一种实施方式中，所述食品是使用短双歧杆菌CCFM1078或含短双歧杆菌CCFM1078的发酵剂生产得到的乳制品、豆制品或果蔬制品。
在一种实施方式中，所述食品是含有短双歧杆菌CCFM1078的固体饮料。
在一种实施方式中，所述发酵剂的制备方法为：将短双歧杆菌CCFM1078按照占培养基总质量2～4％的接种量接种到培养基中，于37℃下培养30h，得到培养液；将培养液离心，收集菌体；将菌体用pH为7.2的磷酸盐缓冲液清洗3次后用冻干保护剂重悬，得到重悬液；将重悬液采用真空冷冻法进行冻干，得到短双歧杆菌CCFM1078的发酵剂。
在一种实施方式中，所述冻干保护剂和菌体的质量比为2:1。
在一种实施方式中，所述冻干保护剂含有脱脂奶粉、麦芽糊精和L-谷氨酸钠；其中脱脂奶粉:麦芽糊精:L-谷氨酸钠＝(8～10):(8～10):1。
在一种实施方式中，所述培养基是由占培养基总质量10％的脱脂乳、0.5％的葡萄糖、1.5％的胰蛋白胨以及0.3％的酵母浸膏溶解于水中制得的。
在一种实施方式中，所述培养基的pH为6.8。
有益效果：本发明提供一株抑制IL‐23/Th17轴相关炎症因子释放、进而预防和/或缓解银屑病的短双歧杆菌CCFM1078，将其用于银屑病样小鼠，可达到如下效果：
(1)银屑病样小鼠病变皮肤情况好转；
(2)银屑病样小鼠皮肤表皮层增厚被抑制；
(3)银屑病样小鼠皮肤中IL-23水平降低20.3％；
(4)银屑病样小鼠皮肤中IL-22水平降低22.0％；
(5)银屑病样小鼠皮肤中IL-17水平降低18.3％；
因此，短双歧杆菌CCFM1078在制备预防和/或缓解银屑病的产品(如食品、药品或保健食品等)中，具有巨大的应用前景。
生物材料保藏
短双歧杆菌(Bifidobacterium breve)CCFM1078，已于2020年5月6日保藏于广东省微生物菌种保藏中心，保藏编号为GDMCC No.61011，保藏地址为广州市先烈中路100号大院59号楼5楼。
附图说明
图1：不同组别实验小鼠病变皮肤情况。
图2：不同组别实验小鼠皮肤病理切片。
图3：不同组别实验小鼠皮肤中IL-23含量。
图4：不同组别实验小鼠皮肤中IL-22含量。
图5：不同组别实验小鼠皮肤中IL-17含量。
具体实施方式
下述实施例中涉及的培养基如下：
mMRS培养基配方(1L)：蛋白胨10g，牛肉膏10g，酵母粉5g，葡萄糖20g，K 2HPO 42g，柠檬酸二铵2g，乙酸钠2g，吐温80 1mL，MgSO 4·7H 2O 0.5g，半胱氨酸盐酸盐0.5g，MnSO 4·4H 2O 0.25g，pH为7.2～7.4。
mMRS固体培养基配方(1L)：蛋白胨10g，牛肉膏10g，酵母粉5g，葡萄糖20g，K 2HPO 4 2g，柠檬酸二铵2g，乙酸钠2g，吐温80 1mL，MgSO 4·7H 2O 0.5g，半胱氨酸盐酸盐0.5g，MnSO 4·4H 2O 0.25g，琼脂20g，pH为7.2～7.4。
下述实施例中涉及的检测方法如下：
活菌数的检测方法：采用国标《GB 4789.35-2016食品安全国家标准食品微生物学检测乳酸菌检测》。
酸度检测方法：采用国标GB 431334-2010。
短双歧杆菌B.breve 1为采用相同方法分离从不同的粪便样品中分离获得的另一菌株。
实施例1：短双歧杆菌的培养
将短双歧杆菌CCFM1078接入mMRS固体培养基中于37℃培养48h后，观察其菌落并在显微镜下对其菌体进行观察，发现其菌落乳白色、不规则型、圆形凸起、光滑，其菌体形状呈轻微不规则、圆形末端的弯曲杆菌，通常单个、成对和小簇存在。
将短双歧杆菌CCFM1078接入MRS液体培养基中于37℃培养30h后，转入新鲜的MRS液体培养基中，同样条件培养30h，6000×g离心菌体15min，用0.9％生理盐水洗涤菌体后于6000×g再次离心10min，收集菌体，用30％(m/v)蔗糖溶液重悬，冻存于80℃待用。
短双歧杆菌B.breve 1采用上述相同方法制备菌液。
实施例2：短双歧杆菌对银屑病小鼠病变皮肤的缓解作用
将6-8周龄的SPF级BALB/c雌性小鼠分成5组，分别为正常组、模型组、阳性参照组(以下简写为阳参组)和实验组。阳参组使用药物甲氨蝶呤，实验组包括灌胃短双歧杆菌CCFM1078的CCFM1078组和灌胃短双歧杆菌B.breve 1的B.breve 1组，每组6只，饲养在江南大学实验动物中心，普通饲料喂养，恒定温度21-26℃，湿度40-70％，噪声小于等于60dB，动物照度15-20LX(所有动物实验程序皆由江南大学动物福利与伦理管理委员会进行审查并批准)。
实验周期共计3周，第3周进行造模，造模前一天对小鼠背部进行脱毛处理，面积约为2.5cm×2.5cm。造模期间每天对模型组和实验组小鼠耳部和背部脱毛区涂抹咪喹莫特乳膏，耳部10mg，背部62.5mg，正常组仅涂抹等量的凡士林。实验期间，甲氨蝶呤组灌胃1mg/kg/天的甲氨蝶呤溶液(甲氨蝶呤溶液是将甲氨蝶呤溶解在无菌生理盐水中配制的溶液)。造模期间，CCFM1078组每天灌胃0.2mL活菌数为1×10 9CFU/mL的CCFM1078菌悬液(按照实施例1的方法制备)，B.breve 1组每天灌胃0.2mL活菌数为1×10 9CFU/mL的B.breve 1菌悬液(按照实施例1的方法制备)，正常组和模型组仅灌胃等量的无菌生理盐水作为对照，所有分组均为自由饮水和摄食。造模期间每天对小鼠背部情况拍照记录，于第4周第1天处死小鼠。结果见图1。
由图1可知，正常组小鼠背部脱毛区皮肤光滑，无鳞屑无红斑；而模型组小鼠背部脱毛区皮肤具有褶皱感，并覆有明显的鳞屑，伴有红斑；而CCFM1078组较之模型组，背部脱毛区皮肤光滑，且基本无鳞屑不红斑，基本与甲氨蝶呤组皮肤状况相似；而B.breve 1组小鼠背部脱毛区皮肤如模型组一样，具有褶皱感并覆有明显鳞屑。
以上实验结果表明，相比于短双歧杆菌B.breve 1，短双歧杆菌(Bifidobacterium breve)CCFM1078更能够缓解银屑病小鼠病变皮肤的症状。
实施例3：短双歧杆菌对银屑病小鼠角质增厚的抑制作用
将6-8周龄的SPF级BALB/c雌性小鼠分成4组，分别为正常组、模型组和实验组，其中，实验组包括灌胃短双歧杆菌CCFM1078的CCFM1078组和灌胃短双歧杆菌B.breve 1的B.breve 1组，每组6只，饲养在江南大学实验动物中心，普通饲料喂养，恒定温度21-26℃，湿度40-70％，噪声小于等于60dB，动物照度15-20LX(所有动物实验程序皆由江南大学动物福利与伦理管理委员会进行审查并批准)。
实验周期共计3周，第3周进行造模，造模前一天对小鼠背部进行脱毛处理，面积约为2.5cm×2.5cm。造模期间每天对模型组和实验组小鼠耳部和背部脱毛区涂抹咪喹莫特乳膏，耳部10mg，背部62.5mg，正常组仅涂抹等量的凡士林。第1-3周，实验期间，CCFM1078组每天灌胃0.2mL活菌数为1×10 9CFU/mL的CCFM1078菌悬液(按照实施例1的方法制备)，B.breve 1组每天灌胃0.2mL活菌数为1×10 9CFU/mL的B.breve 1菌悬液(按照实施例1的方法制备)，正常组和模型组仅灌胃等量的无菌生理盐水作为对照，所有分组均为自由饮水和摄食。于第4周第1天处死小鼠，取小鼠背部脱毛区部分皮肤制作病理学切片，进而进行组织病理学分析。结果见图2。
由图2可知，空白组小鼠皮肤表皮层仅由一层或两层细胞组成，表皮各层未见炎症反应；而模型组小鼠角质明显增厚，并伴有严重的炎症；而CCFM1078组较之模型组，角质增厚现象不明显，且炎症现象较轻，基本与甲氨蝶呤组皮肤状况相似；而B.breve 1组小鼠皮肤角质增厚明显，且炎症现象严重。
以上实验结果表明，相比于短双歧杆菌B.breve 1，短双歧杆菌CCFM1078更能够抑制银屑病小鼠病变皮肤的角质增厚现象。
实施例4：短双歧杆菌对银屑病小鼠皮肤中IL-23的影响
将6-8周龄的SPF级BALB/c雌性小鼠分成4组，分别为正常组、模型组和实验组，其中，实验组包括灌胃短双歧杆菌CCFM1078的CCFM1078组和灌胃短双歧杆菌B.breve 1的B.breve 1组，每组6只，饲养在江南大学实验动物中心，普通饲料喂养，恒定温度21-26℃，湿度40-70％，噪声小于等于60dB，动物照度15-20LX(所有动物实验程序皆由江南大学动物福利与伦理管理委员会进行审查并批准)。
实验周期共计3周，第3周进行造模，造模前一天对小鼠背部进行脱毛处理，面积约为2.5cm×2.5cm。造模期间每天对模型组和实验组小鼠耳部和背部脱毛区涂抹咪喹莫特乳膏，耳部10mg，背部62.5mg，正常组仅涂抹等量的凡士林。实验期间，CCFM1078组每天灌胃0.2mL活菌数为1×10 9CFU/mL的CCFM1078菌悬液，B.breve 1组每天灌胃0.2mL活菌数为1×10 9CFU/mL的B.breve 1菌悬液，正常组和模型组仅灌胃等量的无菌生理盐水作为对照，所有分组均为自由饮水和摄食。于第4周第1天处死小鼠，取小鼠背部脱毛区部分皮肤于-80℃保存，通过ELISA试剂盒检测皮肤中IL-23的含量，结果见图3。
由图3可知，小鼠灌胃短双歧杆菌CCFM1078后，皮肤中IL-23的含量降低了20.3％，较模型组显著降低(p＜0.01)，说明本发明中的菌株，尽管抑制效果不如药物甲氨蝶呤(较 之模型组降低43.8％，甚至IL-23水平低于空白组)，但的确能够抑制炎症反应；而小鼠灌胃短双歧杆菌B.breve 1后，皮肤中IL-23的含量仅降低了4.4％，与模型组无显著差异。
以上实验结果表明，短双歧杆菌CCFM1078能显著下调银屑病小鼠中典型上调促炎因子至正常水平，尤其是降低IL-23水平，显著优于另外一株短双歧杆菌B.breve 1。
实施例5：短双歧杆菌对银屑病小鼠皮肤中IL-22的影响
将6-8周龄的SPF级BALB/c雌性小鼠分成4组，分别为正常组、模型组和实验组，其中，实验组包括灌胃短双歧杆菌CCFM1078的CCFM1078组和灌胃短双歧杆菌B.breve 1的B.breve 1组，每组6只，饲养在江南大学实验动物中心，普通饲料喂养，恒定温度21-26℃，湿度40-70％，噪声小于等于60dB，动物照度15-20LX(所有动物实验程序皆由江南大学动物福利与伦理管理委员会进行审查并批准)。
实验周期共计3周，第3周进行造模，造模前一天对小鼠背部进行脱毛处理，面积约为2.5cm×2.5cm。造模期间每天对模型组和实验组小鼠耳部和背部脱毛区涂抹咪喹莫特乳膏，耳部10mg，背部62.5mg，正常组仅涂抹等量的凡士林。实验期间，CCFM1078组每天灌胃0.2mL活菌数为1×10 9CFU/mL的CCFM1078菌悬液，B.breve 1组每天灌胃0.2mL活菌数为1×10 9CFU/mL的B.breve 1菌悬液，正常组和模型组仅灌胃等量的无菌生理盐水作为对照，所有分组均为自由饮水和摄食。于第4周第1天处死小鼠，取小鼠背部脱毛区部分皮肤于-80℃保存，通过ELISA试剂盒检测皮肤中IL-22的含量，结果见图4。
由图4可知，小鼠灌胃短双歧杆菌CCFM1078后，皮肤中IL-22的含量降低了22.0％，较模型组显著降低(p＜0.05)，说明本发明中的菌株，尽管抑制效果不如药物甲氨蝶呤(较之模型组降低41.8％，甚至IL-22水平低于空白组)，但的确能够抑制炎症反应；而小鼠灌胃短双歧杆菌B.breve 1后，皮肤中IL-22的含量仅降低了0.5％，与模型组无显著差异。
以上实验结果表明，短双歧杆菌CCFM1078能显著下调银屑病小鼠中典型上调促炎因子至正常水平，尤其是降低IL-22水平，显著优于另外一株短双歧杆菌B.breve 1。
实施例6：短双歧杆菌对银屑病小鼠皮肤中IL-17的影响
将6-8周龄的SPF级BALB/c雌性小鼠分成4组，分别为正常组、模型组和实验组，其中，实验组包括灌胃短双歧杆菌CCFM1078的CCFM1078组和灌胃短双歧杆菌B.breve 1的B.breve 1组，每组6只，饲养在江南大学实验动物中心，普通饲料喂养，恒定温度21-26℃，湿度40-70％，噪声小于等于60dB，动物照度15-20LX(所有动物实验程序皆由江南大学动物福利与伦理管理委员会进行审查并批准)。
实验周期共计3周，第3周进行造模，造模前一天对小鼠背部进行脱毛处理，面积约为2.5cm×2.5cm。造模期间每天对模型组和实验组小鼠耳部和背部脱毛区涂抹咪喹莫特乳膏，耳部10mg，背部62.5mg，正常组仅涂抹等量的凡士林。实验期间，CCFM1078组每天灌胃0.2mL的CCFM1078菌悬液，B.breve 1组每天灌胃0.2mL的B.breve 1菌悬液，正常组和模型组仅灌胃等量的无菌生理盐水作为对照，所有分组均为自由饮水和摄食。于第4周第1天处死小鼠，取小鼠背部脱毛区部分皮肤于-80℃保存，通过ELISA试剂盒检测皮肤中IL-17的含量，结果见图5。
由图5可知，小鼠灌胃短双歧杆菌CCFM1078后，皮肤中IL-17的含量降低了18.3％，较模型组显著降低(p＜0.05)，说明本发明中的菌株，尽管抑制效果不如药物甲氨蝶呤(较之模型组降低24.6％)，但的确能够抑制炎症反应；而小鼠灌胃短双歧杆菌B.breve 1后，皮肤中IL-17的含量甚至升高了。
以上实验结果表明，短双歧杆菌CCFM1078能显著下调银屑病小鼠中典型上调促炎因子至正常水平，尤其是降低IL-17水平，显著优于另外一株短双歧杆菌B.breve 1。
实施例7：含有短双歧杆菌CCFM1078菌粉的制备
将保藏于保菌管的短双歧杆菌CCFM1078的种子液按照占培养基总质量3％的接种量接种到mMRS培养基中，于37℃下培养30h，得到培养液；将培养液离心，收集菌体；将菌体用pH为7.2的磷酸盐缓冲液清洗3次后用海藻糖浓度为100g/L的海藻糖冻干保护剂重悬，并控制冻干保护剂和菌体的质量比为2:1，得到重悬液；将重悬液-80℃预冷1.5h后立即转移至冷冻干燥机干燥24h，得到短双歧杆菌CCFM1078菌粉。
实施例8：含有短双歧杆菌CCFM1078酸奶的制备
将奶粉、菊粉、甜菊糖、水按重量比20：5：5：75进行混料，均质，制成发酵原材料；以121℃超高温灭菌300s杀菌，然后冷却到42℃，接种保加利亚乳杆菌和嗜热链球菌的混合菌粉，在42℃下发酵12h后，使保加利亚乳杆菌和嗜热链球菌的菌体浓度控制为10 5CFU/g和10 7CFU/g，之后进行调配；将发酵产物冷却至37℃，加入短双歧杆菌CCFM1078冻干菌粉，短双歧杆菌CCFM1078冻干菌粉的投料量为10 9CFU短双歧杆菌CCFM1078/ml酸奶，搅拌，罐装，在4℃下保存2天自然完成后熟，制成益生菌酸奶。
实施例9：短双歧杆菌CCFM1078与婴儿双歧杆菌35624的比较
David等人发现26个银屑病患者连续口服婴儿双歧杆菌(Bifidobacterium infantis)356246周后，患者病情得到缓解。具体表现为银屑病患者血清内C-反应蛋白、IL-6、TNF-α水平降低，而未对患者的皮损状况进行描述。而短双歧杆菌CCFM1078则是降低了银屑病样小鼠 皮损部位Th17/IL-23轴相关炎症因子水平，具体为IL-23、IL-22以及IL-17，与婴儿双歧杆菌(Bifidobacterium infantis)35624针对的位点不同。且短双歧杆菌CCFM1078对银屑病样小鼠皮损部位的角质层增厚现象具有明显抑制作用，并且有效改善了皮损的红斑、鳞屑症状。因此短双歧杆菌CCFM1078对银屑病的缓解作用与婴儿双歧杆菌(Bifidobacterium infantis)35624具有差异性。
虽然本发明已以较佳实施例公开如上，但其并非用以限定本发明，任何熟悉此技术的人，在不脱离本发明的精神和范围内，都可做各种的改动与修饰，因此本发明的保护范围应该以权利要求书所界定的为准。

Claims (13)

  1. 短双歧杆菌(Bifidobacterium breve)CCFM1078或含短双歧杆菌CCFM1078的产品在抑制IL‐23/Th17轴相关炎症因子释放，预防和/或缓解银屑病方面的应用，其特征在于，所述短双歧杆菌CCFM1078已于2020年5月6日保藏于广东省微生物菌种保藏中心，保藏编号为GDMCC No.61011。
  2. 根据权利要求1所述的应用，其特征在于，所述缓解银屑病包括(a)或(b)至少一种用途：
    (a)有效改善皮肤褶皱、鳞屑和/或红斑症状，
    (b)抑制皮肤角质增厚。
  3. 所述抑制IL‐23/Th17轴相关炎症因子释放具体为：降低银屑病样个体皮肤中IL-23水平、IL-22水平和IL-17水平。
  4. 根据权利要求1所述的应用，其特征在于，所述产品包含食品、药品或保健食品。
  5. 根据权利要求1～4任一所述的应用，其特征在于，所述产品中短双歧杆菌CCFM1078的活菌数不低于1×10 6CFU/mL或1×10 6CFU/g。
  6. 根据权利要求5所述的应用，其特征在于，所述药品含有短双歧杆菌CCFM1078、药物载体和/或药用辅料。
  7. 根据权利要求6所述的应用，其特征在于，所述药物载体包含微囊、微球、纳米粒以及脂质体；所述药用辅料包含赋形剂以及附加剂。
  8. 根据权利要求7所述的应用，其特征在于，所述附加剂包含微晶纤维素、羟丙基甲基纤维素以及精制卵磷脂。
  9. 根据权利要求6～8任一所述的应用，其特征在于，所述药品的剂型包含颗粒剂、胶囊剂、片剂、丸剂或口服液。
  10. 根据权利要求4所述的应用，其特征在于，所述食品含有短双歧杆菌CCFM1078或其发酵代谢物。
  11. 根据权利要求4所述的应用，其特征在于，所述食品是使用短双歧杆菌CCFM1078或含有短双歧杆菌CCFM1078的发酵剂生产得到的乳制品、豆制品或果蔬制品。
  12. 根据权利要求10所述的应用，其特征在于，所述食品是含有短双歧杆菌CCFM1078的固体饮料。
  13. 根据权利要求1所述的应用，其特征在于，所述含短双歧杆菌CCFM1078的产品为发酵剂，所述发酵剂按如下方法制备：将短双歧杆菌CCFM1078接种到培养基中，于35～37℃下培养20～30h，得到培养液；将培养液离心，收集菌体；将菌体清洗后用冻干保护剂重悬，得到重悬液；再将重悬液冻干，得到短双歧杆菌CCFM1078的发酵剂。
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