WO2017175774A1 - インターフェロンλ産生促進用組成物及びその製造方法 - Google Patents
インターフェロンλ産生促進用組成物及びその製造方法 Download PDFInfo
- Publication number
- WO2017175774A1 WO2017175774A1 PCT/JP2017/014153 JP2017014153W WO2017175774A1 WO 2017175774 A1 WO2017175774 A1 WO 2017175774A1 JP 2017014153 W JP2017014153 W JP 2017014153W WO 2017175774 A1 WO2017175774 A1 WO 2017175774A1
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- interferon
- lactic acid
- production
- composition
- strain
- Prior art date
Links
Images
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
- A61K35/744—Lactic acid bacteria, e.g. enterococci, pediococci, lactococci, streptococci or leuconostocs
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/70—Carbohydrates; Sugars; Derivatives thereof
- A61K31/7088—Compounds having three or more nucleosides or nucleotides
- A61K31/713—Double-stranded nucleic acids or oligonucleotides
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
- A61P1/16—Drugs for disorders of the alimentary tract or the digestive system for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/04—Antibacterial agents
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/12—Antivirals
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/12—Antivirals
- A61P31/14—Antivirals for RNA viruses
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/12—Antivirals
- A61P31/20—Antivirals for DNA viruses
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P37/00—Drugs for immunological or allergic disorders
- A61P37/02—Immunomodulators
- A61P37/04—Immunostimulants
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/52—Cytokines; Lymphokines; Interferons
- C07K14/555—Interferons [IFN]
- C07K14/57—IFN-gamma
Definitions
- the present invention relates to a composition for promoting production of interferon ⁇ and a method for producing the same.
- Interferon ⁇ is one type of physiologically active substance (cytokine) belonging to type III interferon, and production by dendritic cells, hepatocytes, intestinal epithelial cells, lung epithelial cells, etc. has been recognized so far.
- Interferon ⁇ has an innate immune activation effect such as an antiviral effect, and has antiviral activity, immunostimulatory activity, anti-infective activity, anti-hepatitis B activity, anti-hepatitis C activity, anti-proliferative activity, anti-tumor activity, anti-tumor activity, Since it can exhibit cancer activity and the like, it is expected to be used in foods and drinks having these activities, therapeutic agents, preventive agents, improving agents, mitigating agents and the like.
- interferon ⁇ has been clinically tested in the treatment of hepatitis C, and it has been confirmed that it actually shows an antiviral effect.
- interferon ⁇ has been confirmed to exhibit an antiviral action against rotavirus, RS virus, influenza virus, and the like, and plays an important role in immunization of tissues such as the liver, intestinal tract, and respiratory organs.
- interferon ⁇ can compensate for the decline in immunity that occurs with aging. Since receptors for interferon ⁇ are localized in the cells described above, interferon ⁇ suppresses side effects on the intake, unlike interferon ⁇ , where the receptor is expressed in all cells. A selective physiologically active action.
- interferon ⁇ not only by directly administering interferon ⁇ , but also by increasing the expression level of interferon ⁇ in vivo, that is, by administering a substance that promotes production of interferon ⁇ , the physiological activity expected by interferon ⁇ Can be caused in vivo.
- dendritic cells can be broadly classified into plasmacytoid dendritic cells (pDC) and myeloid dendritic cells (mDC), and mDC is positive for mDC1 and BDCA3, which are negative for CD19. It can be divided into a certain BDCA3DC.
- dendritic cell subsets are known to differ in the type and degree of cell surface molecular markers expressed, the types of cytokines produced, and the like (see, for example, Non-Patent Documents 1 and 2).
- Patent Document 1 describes Lactococcus lactis JCM20101 strain and JCM5805 strain as those that may activate pDC to promote the production of interferon ⁇ . ing.
- Tetragenococcus lactic acid bacteria are known as lactic acid bacteria that can promote the production of interferon ⁇ and interferon ⁇ as cytokines different from interferon ⁇ (see, for example, Patent Documents 2 and 3).
- the production amount of interferon ⁇ in the JCM20101 strain and the JCM5805 strain is one order of magnitude less than that of other interferons. Therefore, the present inventors decided to investigate again the interferon ⁇ production promoting action of JCM20101 strain and JCM5805 strain. At that time, among the dendritic cells, BDCA3DC, which produced a relatively large amount of interferon ⁇ , was used.
- Patent Document 1 Surprisingly, the JCM20101 strain and JCM5805 strain described in Patent Document 1 hardly exhibited an interferon ⁇ production promoting action against BDCA3DC. Further, Patent Documents 2 and 3 and Non-Patent Documents 1 and 2 do not describe a substance that exhibits an interferon ⁇ production promoting action on BDCA3DC.
- the problem to be solved by the present invention is to provide a composition for promoting production of interferon ⁇ , which contains an active ingredient exhibiting an interferon ⁇ production promoting action on BDCA3DC, and a method for producing the same.
- lactic acid bacteria that can be cultured under stress conditions, bacterial cell components, or cultures promote interferon ⁇ production against BDCA3DC. It was found to show an action.
- the JCM20101 strain and the JCM5805 strain described in Patent Document 1 contain double-stranded RNA, but do not exhibit an interferon ⁇ production promoting action against BDCA3DC, whereas Tetragenococcus
- the present inventors have found that the halophyllus (Tetragenococcus halophilus) KK221 strain exhibits a particularly excellent interferon ⁇ production promoting action against BDCA3DC.
- the present inventors have a composition for promoting the production of interferon ⁇ , which contains, as an active ingredient, lactic acid bacteria that can be cultured under stress conditions such as the KK221 strain.
- a composition for promoting the production of interferon ⁇ which contains, as an active ingredient, lactic acid bacteria that can be cultured under stress conditions such as the KK221 strain.
- the present invention has been completed based on these findings and successful examples.
- the present invention relates to the following.
- a composition for promoting the production of interferon ⁇ which contains, as an active ingredient, a lactic acid bacterium that can be cultured under stress conditions, a microbial cell component, or a culture.
- 3. The composition for promoting production of interferon ⁇ according to 1 or 2, wherein the interferon ⁇ is interferon ⁇ 3. 4).
- composition for promoting interferon ⁇ production according to any one of 1 to 3, wherein the cell, the cell component, and the culture are a cell, a cell component, and a culture containing a nucleic acid, respectively. 5. Lactobacillus is possible cultivated under the stress conditions, second lactic acid bacteria and bacterial cell 1 ⁇ 10 10 cfu per total nucleic acid containing the cells 1 ⁇ 10 10 cfu per 15,000 / ml or more double-stranded RNA 5.
- composition for promoting interferon ⁇ production according to any one of 1 to 5, wherein the composition for promoting production of interferon ⁇ is an enteric composition for promoting production of interferon ⁇ . 7).
- a method for producing a composition for promoting production of interferon ⁇ comprising culturing a lactic acid bacterium capable of being cultured under stress conditions to obtain a microbial cell, a microbial component or a culture of the lactic acid bacterium.
- lactic acid bacteria cell Use of a lactic acid bacteria cell, a cell component, or a culture that can be cultured under stress conditions for the production of a composition for promoting production of interferon ⁇ . 11. Use of lactic acid bacteria cells, cell components or cultures that can be cultured under stress conditions to promote production of interferon ⁇ .
- composition of the present invention and the composition obtained by the production method of the present invention have an interferon ⁇ production promoting action, particularly an interferon lambda production promoting action on BDCA3DC.
- An activation effect, an anti-infective effect, a resistant anti-hepatitis B effect, an anti-hepatitis C effect, an anti-proliferative activity effect, an anti-tumor effect, an anti-cancer effect and the like can be expected.
- the active ingredient used in the composition of the present invention has a track record of using food and drink additives. Therefore, the composition of the present invention is highly safe, and is an antiviral agent, an immunostimulator, an anti-infective agent, an anti-hepatitis B agent, an anti-hepatitis C agent, an intestinal tract immunostimulator, an airway immunity stimulator. It is useful as an agent, an antitumor agent, an anticancer agent and the like, and can be expected to be provided in an oral or parenteral form.
- FIG. 1 is a diagram showing the results of an interferon ⁇ 3 production promotion test of BDCA3DC by Tetragenococcus halophilus KK221 strain described in Examples.
- FIG. 2 is a diagram showing the results of the interferon ⁇ 3 production promotion test of BDCA3DC, pDC and mDC1 by Tetragenococcus halophilus KK221 strain described in Examples.
- FIG. 3 is a diagram showing the results of an interferon ⁇ 3 production promotion test of BDCA3DC by Tetragenococcus halophilus KK221 strain and cells treated with RNase A described in the Examples.
- FIG. 1 is a diagram showing the results of an interferon ⁇ 3 production promotion test of BDCA3DC by Tetragenococcus halophilus KK221 strain described in Examples.
- FIG. 2 is a diagram showing the results of the interferon ⁇ 3 production promotion test of BDCA3DC, pDC and mDC1 by Tetragenoc
- FIG. 4 is a diagram showing the results of an interferon ⁇ 3 production promotion test for BDCA3DC by the presence or absence of chloroquine by Tetragenococcus halophilus KK221 strain and Poly IC described in Examples.
- FIG. 5 is a diagram showing the results of a BDCA3DC interferon ⁇ 3 production promotion test according to the presence or absence of the addition of a TRIF inhibitor by the Tetragenococcus halophilus KK221 strain described in the Examples.
- FIG. 6 is a diagram showing the results of the interferon ⁇ 3 production promotion test of BDCA3DC by Tetragenococcus halophilus KK221 strain, Lactococcus lactis JCM20101 strain and JCM5805 strain described in Examples.
- FIG. 7 is a diagram showing the results of an interferon ⁇ production promotion test for pDC by Tetragenococcus halophilus KK221 strain, Lactococcus lactis JCM20101 strain and JCM5805 strain described in Examples.
- FIG. 8A and FIG. 8B are diagrams showing the results of each BDCA3DC cytokine production promotion test by Tetragenococcus halophilus KK221 strain and Poly IC described in Examples.
- FIG. 9 is a diagram showing the results of measuring the amount of double-stranded RNA per viable cell count by Tetragenococcus halophilus KK221 strain, Lactococcus lactis JCM20101 strain and JCM5805 strain described in Examples. is there.
- FIG. 8A and FIG. 8B are diagrams showing the results of each BDCA3DC cytokine production promotion test by Tetragenococcus halophilus KK221 strain and Poly IC described in Examples.
- FIG. 9 is a diagram
- FIG. 10 is a diagram showing the results of measuring the total amount of nucleic acids per viable count by Tetragenococcus halophilus KK221 strain, Lactococcus lactis JCM20101 strain and JCM5805 strain described in Examples.
- FIG. 11 shows the result of calculating the ratio of double-stranded RNA in the total nucleic acid per viable count by Tetragenococcus halophilus KK221 strain, Lactococcus lactis JCM20101 strain and JCM5805 strain described in Examples.
- FIG. 12 is a diagram showing the results of measuring the amount of double-stranded RNA per dry cell by Tetragenococcus halophilus KK221 strain, Lactococcus lactis JCM20101 strain and JCM5805 strain described in the Examples. is there.
- FIG. 13 is a diagram showing the results of measuring the total amount of nucleic acid per dry cell by Tetragenococcus halophilus KK221 strain, Lactococcus lactis JCM20101 strain and JCM5805 strain described in Examples.
- FIG. 13 is a diagram showing the results of measuring the total amount of nucleic acid per dry cell by Tetragenococcus halophilus KK221 strain, Lactococcus lactis JCM20101 strain and JCM5805 strain described in Examples.
- FIG. 14 shows the result of calculating the ratio of double-stranded RNA in the total nucleic acid per dry cell by Tetragenococcus halophilus KK221 strain, Lactococcus lactis JCM20101 strain and JCM5805 strain described in Examples.
- FIG. FIG. 15 is a diagram showing the results of measuring the amount of double-stranded RNA per viable count by various lactic acid bacteria described in Examples.
- FIG. 16A and FIG. 16B are solutions corresponding to 1 ⁇ 10 10 cfu of a culture sterilization treatment solution obtained by culturing Lactobacillus sake K1 strain described in Examples at 20 ° C., 34 ° C. or 38 ° C.
- FIG. 17A and FIG. 17B are solutions corresponding to 1 ⁇ 10 10 cfu of a culture sterilization treatment liquid obtained by culturing Lactobacillus sake K41 strain at 23 ° C., 29 ° C. or 36 ° C. described in the Examples. It is a figure which shows the result of having measured the amount of double stranded RNA per viable count about quantity.
- the composition for promoting the production of interferon ⁇ according to the present invention (hereinafter simply referred to as the composition of the present invention) has an action of promoting the production of interferon ⁇ and can be cultured as an active ingredient under stress conditions. It contains at least bacterial cells, bacterial cell components or cultures.
- interferon ⁇ production promoting action means, for example, at least one of promoting the expression level of interferon ⁇ gene, increasing the translation amount of interferon ⁇ protein, and activating interferon ⁇ -producing cells. Any one action is said.
- the interferon ⁇ production promoting action possessed by the composition of the present invention is not particularly limited as long as it is an interferon lambda production promoting action induced by providing the composition of the present invention.
- interferon lambda production promoting action on dendritic cells Preferably, it is an interferon ⁇ production promoting action on peripheral blood dendritic cells, and more preferably an interferon lambda production promoting action on BDCA3DC.
- the interferon ⁇ promoted by the composition of the present invention is not particularly limited, and may be any of interferon ⁇ 1, interferon ⁇ 2, and interferon ⁇ 3, preferably interferon ⁇ 3, more preferably interferon ⁇ 3 produced in BDCA3DC. is there.
- composition of the present invention has an antiviral effect, an immunostimulatory effect, an anti-infective effect, an anti-hepatitis B effect, an anti-hepatitis C effect, an anti-proliferative activity effect, an anti-tumor effect by having an interferon ⁇ production promoting action.
- the aspect of an activator, an antitumor agent, and an anticancer agent can be taken.
- the anti-disease effect exhibited by the composition of the present invention is, for example, in the case of an anti-viral effect, that a state that is assumed to be suffering from a current or future viral disease or viral disease in a dietary person. Refers to suppression, delay or improving the condition.
- the immunostimulatory effect exhibited by the composition of the present invention refers to, for example, activating the current or future immune system in a dietary person to maintain or promote various diseases and abnormalities so as to be in a normal state. .
- Lactic acid bacteria mean microorganisms that produce lactic acid as is generally known.
- Examples of lactic acid bacteria include microorganisms belonging to the genus Tetragenococcus, Pediococcus, Lactobacillus, Streptococcus, Leuconostoc, and the like. Genococcus, Pediococcus and Lactobacillus are particularly preferred.
- lactic acid bacteria include Tetragenococcus halophilus KK221 strain, Tetragenococcus halophyllus NBRC12172 strain, Pediococcus pentosaceus OS strain (NITE P-354), Pediococcus pentosaceus NRIC1915 strain, Pedio Coccus pentosaceus NRIC0099 strain, Pediococcus pentosaceus NRIC0122 strain, Lactobacillus plantarum NRIC1930 strain, Lactobacillus plantarum NRIC1067 strain, Lactobacillus delbrucky subspices bulgaricus NRIC1688 strain, Lactobacillus delbruchus Key Subspecies Lactis NRIC 1683, Lactobacillus brevis NRIC 1713, Cactobacillus pentosasus NRIC0391 strain, Lactobacillus pentosasus NRIC0396 strain, Lactobacillus pentosasus NRIC1836 strain,
- the lactic acid bacterium that can be cultured under stress conditions is preferably a lactic acid bacterium having a high double-stranded RNA content among the lactic acid bacteria described above, and more than 15,000 ng / ml per 1 ⁇ 10 10 cfu of bacterial cells. More preferably, it is at least one of lactic acid bacteria containing double-stranded RNA and lactic acid bacteria in which the ratio of double-stranded RNA in the total nucleic acid is 3.5% or more per 1 ⁇ 10 10 cfu of bacterial cells.
- the ratio of double-stranded RNA in the total nucleic acid per 1 ⁇ 10 10 cfu of lactic acid bacterium containing 15,000 ng / ml or more of double-stranded RNA per 1 ⁇ 10 10 cfu of the microbial cell Is particularly preferably 3.5% or more.
- the ratio of double-stranded RNA in the total nucleic acid per cell 1 ⁇ 10 10 cfu is 3.5% or more.
- the upper limit of the content of double-stranded RNA per 1 ⁇ 10 10 cfu of lactic acid bacteria is not particularly limited, it is usually 50,000 ng / ml or less.
- the upper limit of the ratio of double-stranded RNA in the total nucleic acid per 1 ⁇ 10 10 cfu of lactic acid bacteria is not particularly limited, but is usually 20% or less.
- Lactic acid bacteria that can be cultured under stress conditions include Lactobacillus plantarum NRIC1067 strain, Lactobacillus delbrucky subspices Lactis NRIC1683 strain, from the viewpoint of growth and homofermentation and no gas generation.
- Lactobacillus pentosas NRIC 1836 strain and Streptococcus thermophilus NRIC 0256 strain are preferred, Pediococcus pentosuseus NRIC 0099 strain, Lactobacillus paracasei subspecies paracasei NRIC 1936 strain, Lactobacillus brevis NRIC 1713 strain are more preferred More preferred are KK221 strain and Pediococcus pentosaceus OS strain (NITE P-354).
- the method for obtaining lactic acid bacteria is not particularly limited, and examples thereof include a method for utilizing commercially available or deposited lactic acid bacteria, a method for separating and utilizing lactic acid bacteria-containing materials such as soy sauce moromi, pickles, and commercially available lactic acid bacteria beverages. .
- the lactic acid bacterium contained as an active ingredient in the composition of the present invention is not particularly limited as long as it can be cultured under stress conditions among the lactic acid bacteria described above.
- the stress condition is a condition deviating from the conditions for normal cultivation of lactic acid bacteria such as high salt concentration, high or low temperature, oligotrophic, eutrophic, low pH, high pH, etc., preferably doubling time of lactic acid bacteria is suitable
- the condition is such that it is longer than the time required for culturing under various conditions.
- the culture under stress conditions with a high salt concentration means, for example, a salt concentration of preferably 0.5 to 30% (w / v), more preferably 5 to 15% (w / v). Culturing using the containing medium is mentioned. In a medium having a salinity of less than 0.5% (w / v) or more than 30% (w / v), the growth of lactic acid bacteria may be extremely slow.
- the culture under a stress condition at a high temperature or a low temperature includes, for example, culturing at an optimum temperature ⁇ 5 to 20 ° C. suitable for the growth of lactic acid bacteria, preferably at an optimum temperature ⁇ 10 ° C.
- the optimal temperature of the lactic acid bacterium to be used is 30 ° C.
- the culture is performed at 35 to 45 ° C. or 15 to 25 ° C.
- the optimal temperature of the lactic acid bacterium to be used is 35 ° C., 40 to Examples include culturing at 50 ° C. or 20 to 30 ° C.
- Lactic acid bacteria can produce more bacterial components such as double-stranded RNA by culturing under stress conditions than by culturing under no stress conditions.
- Lactic acid bacteria that can be cultured under stress conditions can be used alone or in combination of two or more.
- the lactic acid bacteria contained as an active ingredient in the composition of the present invention may be either lactic acid bacteria cultured under stress conditions or lactic acid bacteria cultured under normal conditions.
- one or more lactic acid bacteria cultured under stress conditions and one or more lactic acid bacteria cultured under normal conditions may be combined and contained in the composition of the present invention.
- the active ingredient in the composition of the present invention may be a lactic acid bacterium that can be cultured under stress conditions, that is, a microbial cell component of the lactic acid bacterium and a culture of the lactic acid bacterium in addition to the lactic acid bacterium.
- a microbial cell component of the lactic acid bacterium and a culture of the lactic acid bacterium in addition to the lactic acid bacterium.
- the lactic acid bacterium cells include cells obtained by removing a culture medium of a lactic acid bacterium culture using a generally known solid-liquid separation means such as centrifugation.
- the bacterial cell component of lactic acid bacteria includes, for example, purified or non-purified components that are present in or secreted outside the lactic acid bacteria.
- single-stranded RNA, double-stranded RNA, and single Examples include double-stranded DNA and double-stranded DNA.
- Examples of the culture of lactic acid bacteria include a culture solution obtained by culturing lactic acid bacteria.
- the active ingredient in the composition of the present invention is not particularly limited as long as it is a lactic acid bacterial cell, a bacterial cell component, and a culture that can be cultured under stress conditions. Ingredients and cultures are preferred.
- RNA is preferable, and double-stranded RNA is more preferable.
- Lactic acid bacteria that can be cultured under stress conditions, bacterial cell components, and cultures can be used alone or in combination of two or more thereof.
- RNA preferably double-stranded RNA, may be isolated from the lactic acid bacteria, bacterial cell components and culture.
- the present invention also relates to a composition for promoting production of interferon ⁇ , which contains, as an active ingredient, a lactic acid bacterium that can be cultured under stress conditions, a microbial cell component, and a double-stranded RNA isolated from the culture.
- the present invention also provides a lactic acid bacterium cell, a microbial cell component, and a culture that can be cultured under stress conditions, and a single-stranded RNA is isolated from the lactic acid bacterium cell, microbial cell component, and culture. And a method for producing a composition for promoting production of interferon ⁇ .
- the method for isolating RNA from lactic acid bacterial cells, bacterial cell components and culture is not particularly limited.
- a method for isolating double-stranded RNA from lactic acid bacterial cells the method described in Patent Document 3 is used. Is mentioned.
- lactic acid bacteria belonging to the genus Tetragenococcus are inoculated into an MRS medium (BD) containing 5 to 10% (w / v) sodium chloride, or Pediococcus, Lactobacillus, Lactococcus Lactobacillus belonging to the genus, Streptococcus genus and Leuconostoc genus is inoculated in a normal MRS medium and cultured at 20 to 35 ° C. for 24 to 72 hours, whereby lactic acid bacteria containing double-stranded RNA, Examples include obtaining body components and cultures.
- the lactic acid bacteria containing the double-stranded RNA, the bacterial cell components, and the culture are subjected to a heat sterilization treatment at 90 to 100 ° C. for 5 to 20 minutes to obtain a heat sterilization treatment solution.
- the heat sterilization treatment liquid is subjected to solid-liquid separation means such as centrifugation to recover the solid content.
- the solid is then washed and suspended in a buffer to obtain a suspension.
- lysozyme is added to the suspension and heated at 35 to 40 ° C. to obtain a lysozyme-treated solution.
- the crude nucleic acid extract is a nucleic acid mixture containing DNA, single-stranded RNA, double-stranded RNA, and the like.
- the purified nucleic acid extract is subjected to cellulose column chromatography, whereby purified double-stranded RNA can be obtained.
- Cellulose column chromatography and subsequent advanced purification means can be carried out according to the description in Patent Document 3.
- Lactic acid bacteria, bacterial cell components, and cultures are natural products that have been ingested by humans for a long time and their origins and are highly safe. Therefore, the composition of the present invention has practicality. It is high and can be applied to various uses in an oral or parenteral form as it is or by processing.
- composition of the present invention can be used by mixing with other components, for example, depending on the application.
- the composition of the present invention contains various substances as long as the object of the present invention can be achieved, in addition to the lactic acid bacteria that can be cultured under stress conditions, bacterial cell components, or cultures. it can.
- composition of the present invention examples include saccharide sweeteners, stabilizers, emulsifiers, starch, processed starch products, starch degradation products, salt, flavoring agents, coloring agents, acidulants, flavor ingredients, nutrients, fruit juices, eggs. It may further contain additives used in normal food processing such as animal and vegetable foods such as, excipients, fillers, binders, thickeners, perfume oils.
- additives used in normal food processing such as animal and vegetable foods such as, excipients, fillers, binders, thickeners, perfume oils.
- the amount of additive used is not particularly limited as long as it does not hinder the solution of the problems of the present invention, and can be set as appropriate.
- composition of the present invention is not particularly limited as long as it is a commonly used form, for example, solid, liquid, gel, suspension, cream, sheet, stick, powder, granule, granule, Various forms such as a tablet shape, a rod shape, a plate shape, a block shape, a paste shape, a capsule shape, and a caplet shape can be adopted.
- the composition of the present invention is, for example, an oral composition
- lactic acid bacteria, bacterial cell components and culture, and double-stranded RNA isolated therefrom are transferred to the small intestine via the esophagus and stomach.
- an enteric composition is not particularly limited as long as it is a composition that does not dissolve in gastric acid but dissolves in the small intestine, and examples thereof include compositions in the form of acid-resistant microcapsules and liposomes.
- the content of the microbial cells, cell components, or culture of lactic acid bacteria that can be cultured under the stress conditions contained in the composition of the present invention is not particularly limited as long as the interferon ⁇ production promoting action is recognized.
- the oral composition is, for example, preferably 0.0001% by mass or more, more preferably 0.001% by mass or more, and the upper limit is not particularly limited, but is usually 50% by mass or less, based on the whole composition. is there.
- the parenteral composition is, for example, preferably 0.00001% by mass or more, more preferably 0.0001% by mass or more, and the upper limit is not particularly limited, but is usually 10% by mass or less, based on the entire composition. It is.
- the amount of intake of the composition of the present invention is not particularly limited, and may be set as appropriate according to the symptoms and physique of the intake person.
- the composition containing lactic acid bacteria that can be cultured under stress conditions as an active ingredient The intake is usually 1 to 1000 mg / body weight 60 kg / day.
- composition of the present invention can be used alone or in addition to foods and drinks, pharmaceuticals and the like. That is, another aspect of the present invention is a food or drink containing the composition for promoting production of interferon ⁇ of the present invention, such as a functional food or drink, a specific health food or drink, a nutritional function food or drink, or a health function food or drink. , Special-purpose foods, nutritional supplements, health supplements, supplements, beauty foods, cosmetics, pharmaceuticals, quasi-drugs, animal feeds, and other raw materials for producing these products.
- the production method of the present invention at least includes culturing a lactic acid bacterium that can be cultured under stress conditions to obtain a microbial cell, a microbial cell component, or a culture of the lactic acid bacterium.
- a component showing an interferon ⁇ production promoting action as an active ingredient is obtained, the culture conditions of lactic acid bacteria and the method of collecting the fungus body, fungus body component or culture as the active ingredient are not particularly limited.
- a lactic acid bacterium is inoculated in an MRS medium having a high salt concentration or a normal salt concentration, and the lactic acid bacterium is cultured at an optimum temperature ⁇ 5 to 20 ° C. for 12 to 72 hours.
- the medium is removed from the obtained culture by a generally known solid-liquid separation means such as an ultrafiltration membrane or a centrifugal concentrator to recover the cells.
- a method including obtaining bacterial cells of lactic acid bacteria by washing the obtained bacterial cells with water or saline.
- the microbial cells obtained in this way are the compositions of the present invention. It can be used as an active ingredient.
- the method for the drying treatment is not particularly limited, and examples thereof include natural drying, air drying, spray drying, freeze drying, and the like.
- Example 1 Interferon ⁇ production promotion test of Tetragenococcus lactic acid bacteria
- lactic acid bacteria belonging to the genus Tetragenococcus an interferon ⁇ production promotion test was performed.
- KK221 strain Tetragenococcus halophyllus Th221, hereinafter referred to as KK221 strain
- the applicant deposited the KK221 strain under the following conditions. (1) Depositary Institution: National Institute of Technology and Evaluation Patent Biological Depositary Center (2) Contact: 1-chome, Higashi 1-chome, Tsukuba City, Ibaraki 305-8565 (Currently, Room No.
- a stock solution of lactic acid bacteria was prepared by inoculating MRS medium containing 10% (w / v) sodium chloride at 1 ⁇ 10 7 cells / ml.
- This lactic acid bacterium stock solution was allowed to stand for 48 to 72 hours at 30 ° C., followed by boiling sterilization treatment at 95 ° C. for 10 minutes to prepare a culture sterilization treatment solution.
- Bacterial cells obtained by centrifuging the culture sterilization treatment solution were washed with physiological saline, and then suspended in physiological saline at 1 ⁇ 10 9 cells / ml to prepare a lactic acid bacteria suspension.
- Interferon ⁇ production promotion test The interferon ⁇ production promotion activity of the prepared lactic acid bacteria suspension was evaluated as follows using dendritic cells collected and prepared from the peripheral blood of healthy non-virally infected individuals.
- the number of cells of each isolated DC was measured, and a cell stock solution was prepared using IMDM medium (GIBCO) containing 25 mM D-glucose, 25 mM HEPES and 1 ⁇ M sodium pyruvate so as to be 5 ⁇ 10 4 cells / ml. .
- IMDM medium GIC-120, GIBCO
- interferon ⁇ 3 was obtained by the chemiluminescent enzyme immunoassay method according to the document of Sugiama et al. [Sugiyama M et al. Hepatol Res. 42 (11): 1089-1099, 2012].
- FIG. 1 shows the measurement result of interferon ⁇ 3 when BDCA3DC is used as DC. As shown in FIG. 1, it was confirmed that KK221 strain promoted interferon ⁇ 3 production by BDCA3DC in a concentration-dependent manner 24 hours after the start of co-culture.
- FIG. 2 shows the measurement results of interferon ⁇ 3 when lactic acid bacteria count / DC count is set to 100 and BDCA3CD, pDC and mDC1 are used as DC. As shown in FIG. 2, it was confirmed that KK221 strain promoted interferon ⁇ 3 production of BDCA3DC (BDCA3 positive dendritic cells) and pDC (BDCA4 positive dendritic cells) 24 hours after the start of co-culture.
- BDCA3DC BDCA3 positive dendritic cells
- pDC BDCA4 positive dendritic cells
- KK221 strain has an interferon ⁇ 3 production promoting action in human dendritic cells.
- Example 2 Interferon lambda production promotion test of RNA isolated from Tetragenococcus lactic acid bacteria
- RNase A treatment KK221 strain was suspended in 10 mM Tris-HCl (pH 8.0) to 5 ⁇ 10 9 cells / ml, and bovine pancreas-derived RNase A (Sigma) was further added to 10 ⁇ g / ml. Thus, an RNase A-containing bacterial suspension was prepared.
- the RNase A-containing bacterial suspension was subjected to an enzyme treatment by incubating at 37 ° C. for 2 hours to obtain an enzyme-treated solution.
- enzyme treatment both single-stranded RNA and double-stranded RNA are degraded in the absence of NaCl.
- the bacterial cells collected by centrifuging the obtained enzyme-treated solution were washed twice with 10 mM Tris-HCl (pH 8.0), and then suspended in physiological saline to 1 ⁇ 10 9 cells / ml. It became cloudy and an RNA-degrading lactic acid bacteria suspension was prepared.
- BDCA3DC interferon ⁇ production was promoted by using either the RNA-decomposing lactic acid bacterium suspension or the untreated lactic acid bacterium suspension.
- the interferon ⁇ production promoting activity decreased in the RNA-decomposing lactic acid bacteria suspension. From this result, it was found that RNA in lactic acid bacteria that can be cultured under stress conditions activates dendritic cells and promotes production of interferon ⁇ .
- BDCA3DC is characterized by very high expression of Toll-like receptor (TLR) 3.
- This receptor is known to recognize double-stranded RNA, and the higher the content of double-stranded RNA in the fungus body or the proportion of double-stranded RNA in the total nucleic acid, the stronger the promotion of interferon ⁇ production. Can be considered.
- Example 3 Interferon ⁇ production promotion test using TLR3 pathway inhibitor
- TLR Toll-like receptor
- Compound Poly IC (product name “Poly IC”; Invivogen) was used as the TLR3 ligand.
- the untreated lactic acid bacteria suspension prepared in Example 1 was used.
- the interferon ⁇ concentration in the supernatant 24 hours after the start of co-culture was measured.
- FIG. 4 shows the results of measuring interferon ⁇ production promoting activity depending on whether or not chloroquine was added, depending on the addition of lactic acid bacteria suspension and Poly IC. As shown in FIG. 4, it was confirmed that interferon ⁇ production promoting activity was reduced by adding 10 ⁇ M chloroquine when either lactic acid bacteria suspension or PolyIC was added.
- FIG. 5 shows the measurement results of interferon ⁇ production promoting activity by adding lactic acid bacteria suspension and further adding 0 ⁇ g / ml, 1 ⁇ g / ml and 10 ⁇ g / ml of TRIF inhibitor. It was confirmed that addition of the TRIF inhibitor decreased interferon ⁇ production promoting activity depending on the concentration of the TRIF inhibitor.
- the interferon ⁇ production promoting activity in BDCA3-positive dendritic cells by lactic acid bacteria that can be cultured under stress conditions may be related to a signal involving TRIF, that is, a signal from TLR3.
- a signal involving TRIF that is, a signal from TLR3.
- Example 4 Interferon production promotion test using various lactic acid bacteria
- JCM20101 strain and JCM5805 strain Patent Document 1 belonging to Lactococcus lactis known to induce interferon ⁇ production were used to conduct an interferon ⁇ production promotion test.
- JCM20101 strain and JCM5805 strain were inoculated to MRS medium containing no salt so as to be 1 ⁇ 10 7 cells / ml to prepare lactic acid bacteria stock solutions.
- the lactic acid bacteria stock solution was allowed to stand at 30 ° C. for 24 hours, and then subjected to boiling sterilization treatment at 95 ° C. for 10 minutes to prepare a culture sterilization treatment solution.
- Bacterial cells obtained by centrifuging the culture sterilization treatment solution were washed with physiological saline, and then suspended in physiological saline at 1 ⁇ 10 9 cells / ml to prepare a lactic acid bacteria suspension.
- Patent Document 1 described that JCM20101 strain and JCM5805 strain promote the production of interferon ⁇ .
- JCM20101 strain and JCM5805 strain promote the production of interferon ⁇ .
- BDCA3DC in lactic acid bacteria suspensions using these strains, BDCA3DC Thus, interferon ⁇ production inducing action was not observed.
- the interferon ⁇ production promoting activity was measured by the above.
- Interferon ⁇ was measured using the collected culture supernatant using CBA (cytometric beads assay; BD) of interferon ⁇ . The results are shown in FIG.
- JCM20101 strain and JCM5805 strain were shown to have interferon ⁇ production promoting activity against pDC.
- the interferon ⁇ production promoting action by the JCM20101 strain and JCM5805 strain described in Patent Document 1 is an action on pDC. Is suggested.
- Example 5 Cytokine production promotion test by KK221 strain
- TNF ⁇ , IL-10, IL-1 ⁇ , IL-6, and IL-12p70 were measured using CBA (cytometric beads assay; BD), respectively. The results are shown in FIGS. 8A and 8B.
- Poly IC promotes non-specific production of TNF ⁇ , interferon ⁇ , IL-6 and the like in addition to interferon ⁇ (IL28B), whereas KK221 strain has interferon. It was confirmed that production was specifically promoted for ⁇ .
- TNF ⁇ produced and induced by Poly IC is a typical inflammatory cytokine, and an inflammatory reaction occurs in vivo by TNF ⁇ .
- the KK221 strain can specifically promote production of interferon ⁇ without causing an inflammatory reaction, it was confirmed that the strain KK221 is very useful for inducing a physiological action expected by interferon ⁇ .
- Example 6 Evaluation of amount and ratio of double-stranded RNA per number of living lactic acid bacteria
- the amount of double-stranded RNA hereinafter also abbreviated as dsRNA
- the total nucleic acid amount and the ratio of double-stranded RNA in the total nucleic acid (number of dsRNA / total nucleic acid amount x 100) per KK221, JCM20101, and JCM5805 strains was measured as follows.
- a stock solution of lactic acid bacteria was prepared by inoculating MRS medium containing 10% (w / v) sodium chloride at a concentration of 1 ⁇ 10 7 cells / ml. This lactic acid bacterium stock solution was allowed to stand at 30 ° C. for 48 hours to obtain a KK221 strain culture solution.
- a stock solution of lactic acid bacteria was prepared by inoculating the MRS medium containing no sodium chloride to inoculate JCM20101 strain and JCM5805 strain at 1 ⁇ 10 7 cells / ml, respectively.
- This lactic acid bacteria stock solution was statically cultured at 30 ° C. for 24 hours to obtain a JCM20101 strain culture solution and a JCM5805 strain culture solution.
- a part of the culture solution of KK221 strain was collected, serially diluted, and cultured using an MRS agar medium containing 5% (w / v) sodium chloride to count the number of colonies. Further, a part of the culture solution of JCM201021 strain and the culture solution of JCM5805 strain was collected, serially diluted, and cultured using an MRS agar medium not containing sodium chloride to count the number of colonies.
- Each culture sterilization treatment liquid was prepared by performing boiling sterilization treatment for 15 minutes at 95 degreeC about the remaining part of KK221 stock culture liquid, JCM20101 stock culture liquid, and JCM5805 stock culture liquid. Based on the counted number of colonies, 1 ⁇ 10 10 cfu equivalent of each culture sterilization solution was collected.
- the collected liquids were centrifuged at 8500 ⁇ g for 15 minutes to collect solids.
- the solid was then washed with STE buffer and suspended in STE buffer to obtain a suspension.
- a lysozyme (Sigma-Aldrich) solution (final concentration 5 mg / mL) suspended in STE buffer was added to the suspension and heated at 37 ° C. for 30 minutes to obtain a lysozyme-treated solution.
- 10% SDS Waako Pure Chemical Industries
- Proteinase K Takara Bio Inc.
- the proteinase-treated solution was treated with phenol / chloroform / isoamyl alcohol (Wako Pure Chemical Industries), and centrifuged at 8500 ⁇ g for 15 minutes to obtain a crude nucleic acid extract as a supernatant.
- the total amount of nucleic acid contained in the crude nucleic acid extract was measured using an ultra-trace spectrophotometer (product name “Nanodrop”, Thermo Fisher Scientific). Double-stranded RNA was quantified from the crude nucleic acid extract by ELISA.
- 9 to 11 show the amount of double-stranded RNA, the total amount of nucleic acid, and the ratio of double-stranded RNA in the total nucleic acid obtained by measurement.
- the test was performed by comparison between two groups between the KK221 strain and the JCM20101 strain or the JCM5805 strain (unmatched t-test). The significance level was 5% (“*” in the figure) or 1% (“**”).
- the double-stranded RNA amount of the KK221 strain was larger than the double-stranded RNA amount of the JCM20101 strain and the JCM5805 strain.
- the proportion of double-stranded RNA in the total nucleic acid of the KK221 strain was statistically significantly higher than that of the JCM20101 and JCM5805 strains.
- double-stranded RNA in lactic acid bacteria that can be cultured under stress conditions activates dendritic cells to promote the production of interferon ⁇ , and the amount of double-stranded RNA increases or the total nucleic acid It is considered that the interferon ⁇ production promoting action is improved as the ratio of the double-stranded RNA in the medium is higher.
- the amount of double-stranded RNA per viable cell count is greater than 12184-13819 ng / mL, or the amount of double-stranded RNA in the total nucleic acid per viable cell count It was found that lactic acid bacteria having a ratio greater than 2.6454 to 3.1530% are lactic acid bacteria having an interferon ⁇ production promoting action on BDCA3DC.
- Example 7 Evaluation of amount and ratio of double-stranded RNA per dry cell mass of various lactic acid bacteria
- KK221 strain, JCM20101 strain and JCM5805 strain were measured for the amount of dsRNA per dry cell amount, the total nucleic acid amount and the ratio of double-stranded RNA in the total nucleic acid as follows.
- a stock solution of lactic acid bacteria was prepared by inoculating MRS medium containing 10% (w / v) sodium chloride at a concentration of 1 ⁇ 10 7 cells / ml. This lactic acid bacterium stock solution was allowed to stand at 30 ° C. for 48 hours to obtain a KK221 strain culture solution.
- a stock solution of lactic acid bacteria was prepared by inoculating the MRS medium containing no sodium chloride to inoculate JCM20101 strain and JCM5805 strain at 1 ⁇ 10 7 cells / ml, respectively.
- This lactic acid bacteria stock solution was statically cultured at 30 ° C. for 24 hours to obtain a JCM20101 strain culture solution and a JCM5805 strain culture solution.
- Each culture sterilization treatment solution was prepared by subjecting the KK221 strain culture solution, JCM20101 strain culture solution, and JCM5805 strain culture solution to boiling sterilization treatment at 95 ° C. for 15 minutes.
- the cells obtained by centrifuging each culture sterilization treatment solution were washed with physiological saline, and then 3 cells obtained by centrifuging again were freeze-dried.
- RNA extraction Using the collected freeze-dried powder 5 mg, RNA extraction, total nucleic acid amount measurement, double-stranded RNA extraction, and double-stranded RNA amount measurement were performed in the same manner as in Example 6. 12 to 14 show the amount of double-stranded RNA in 5 mg of each lyophilized powder, the total amount of nucleic acid, and the ratio of double-stranded RNA in the total nucleic acid, respectively.
- the proportion of double-stranded RNA in the total nucleic acid of KK221 strain was statistically significant relative to the proportion of JCM20101 strain and JCM5805 strain. From this result, double-stranded RNA in lactic acid bacteria that can be cultured under stress conditions activates dendritic cells to promote the production of interferon ⁇ , and the percentage of double-stranded RNA in the total nucleic acid is high It is considered that the interferon ⁇ production promoting action is improved.
- lactic acid bacteria in which the ratio of double-stranded RNA in the total nucleic acid per dry cell mass is more than 3.1673-3.5898% promotes interferon ⁇ production with respect to BDCA3DC. It was found to be a lactic acid bacterium having an action.
- Example 8 Evaluation of the amount of double-stranded RNA per viable count of various lactic acid bacteria.
- the amount of double-stranded RNA per viable count of various lactic acid bacteria was measured in the same manner as in Example 6 except that the amount of 5 ⁇ 10 9 cfu equivalent of the culture sterilization treatment solution was used.
- the lactic acid bacteria used were Tetragenococcus halophilus KK221, K1121 and K1142; Lactobacillus sakei K1, K41 and K1185; Lactobacillus salivarius L , K1182 strain and K1183 strain; and Lactococcus lactis K436 strain, K550 strain and K1118 strain.
- the K1185 strain and the K1118 strain are deposited at the National Institute of Technology and Evaluation (NITE-IPOD), and the accession numbers are NBRC15893 and NBRC100933, respectively.
- the K1182 and K1183 strains were obtained from ATCC (American Type Culture Collection) and have ATCC numbers of 13419 and 25975, respectively.
- K1121, K1142, K1, K41, K598, K436 and K550 strains are lactic acid bacteria isolated from fermented foods by Kikkoman Corporation.
- FIG. 15 shows the measurement result of the double stranded RNA measurement value of each lactic acid bacterium when the positive control double stranded RNA measurement value is 100.
- the amount of double-stranded RNA in three strains of Tetragenococcus halophilus and three strains of Lactobacillus sakei was large, whereas three strains of Lactobacillus salivaius and 2 of Lactococcus lactis The strain had less double stranded RNA.
- double-stranded RNA in lactic acid bacteria that can be cultured under stress conditions activates dendritic cells via TLR3 to promote production of interferon ⁇ , and the amount of double-stranded RNA is large. It is considered that the interferon ⁇ production promoting action is improved. Therefore, these results suggested that three strains of Tetragenococcus halophilus and three strains of Lactobacillus sakei may have an interferon ⁇ production promoting effect on BDCA3DC.
- the amount of double-stranded RNA per viable cell count was determined using the 1 ⁇ 10 10 cfu equivalent of the culture sterilization solution obtained by culturing at each temperature. The measurement results are shown in FIGS. 16A and 16B and FIGS. 17A and 17B, respectively.
- the double-stranded RNA amount of the two strains of Lactobacillus sakei is lower than the optimum temperature (38 ° C. or 36 ° C.) or lower (20 ° C.). Alternatively, it was confirmed that the amount of double-stranded RNA increased when cultured at 23 ° C.).
- composition of the present invention and the composition obtained by the production method of the present invention contain an active ingredient that can be applied in both oral and parenteral modes, and have antiviral activity through the action of promoting interferon ⁇ production. It is useful for consumers who expect activity, immunostimulatory activity, anti-infective activity, anti-hepatitis B activity, anti-hepatitis C activity, anti-proliferative activity, anti-tumor activity, and anti-cancer activity. It can be used as food and drink, pharmaceuticals, quasi-drugs, cosmetics and supplements that contribute to the health and welfare of the elderly.
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Medicinal Chemistry (AREA)
- Veterinary Medicine (AREA)
- Pharmacology & Pharmacy (AREA)
- Public Health (AREA)
- Animal Behavior & Ethology (AREA)
- Organic Chemistry (AREA)
- General Chemical & Material Sciences (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Molecular Biology (AREA)
- Communicable Diseases (AREA)
- Oncology (AREA)
- Virology (AREA)
- Mycology (AREA)
- Microbiology (AREA)
- Epidemiology (AREA)
- Engineering & Computer Science (AREA)
- Biochemistry (AREA)
- Immunology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Gastroenterology & Hepatology (AREA)
- Zoology (AREA)
- Toxicology (AREA)
- Biophysics (AREA)
- Genetics & Genomics (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Biotechnology (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
- Coloring Foods And Improving Nutritive Qualities (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
Description
1.ストレス条件下で培養が可能である乳酸菌の菌体、菌体成分又は培養物を有効成分として含有する、インターフェロンλ産生促進用組成物。
2.前記インターフェロンλ産生促進用組成物は、BDCA3DCに対するインターフェロンλ産生促進用組成物である、前記1に記載のインターフェロンλ産生促進用組成物。
3.前記インターフェロンλは、インターフェロンλ3である、前記1又は2に記載のインターフェロンλ産生促進用組成物。
4.前記菌体、前記菌体成分及び前記培養物は、それぞれ核酸を含有する菌体、菌体成分及び培養物である、前記1~3のいずれか1に記載のインターフェロンλ産生促進用組成物。
5.前記ストレス条件下で培養が可能である乳酸菌は、菌体1×1010cfuあたり15,000ng/ml以上の二本鎖RNAを含有する乳酸菌及び菌体1×1010cfuあたり総核酸中の二本鎖RNAの割合が3.5%以上である乳酸菌の少なくとも一方である、前記4に記載のインターフェロンλ産生促進用組成物。
6.前記インターフェロンλ産生促進用組成物が、インターフェロンλ産生促進用腸溶組成物である、前記1~5のいずれか1に記載のインターフェロンλ産生促進用組成物。
7.前記1~6のいずれか1に記載のインターフェロンλ産生促進用組成物を含有する飲食品。
8.ストレス条件下で培養が可能である乳酸菌を培養して、該乳酸菌の菌体、菌体成分又は培養物を得ることを含む、インターフェロンλ産生促進用組成物の製造方法。
9.ストレス条件下で培養が可能である乳酸菌の菌体、菌体成分又は培養物を使用してインターフェロンλの産生を促進する方法。
10.インターフェロンλ産生促進用組成物の製造のためのストレス条件下で培養が可能である乳酸菌の菌体、菌体成分又は培養物の使用。
11.インターフェロンλの産生を促進するためのストレス条件下で培養が可能である乳酸菌の菌体、菌体成分又は培養物の使用。
テトラジェノコッカス属に属する乳酸菌を使用し、インターフェロンλ産生促進試験を実施した。
乳酸菌としてテトラジェノコッカス・ハロフィラスKK221株(Tetragenococcus halophilus Th221、以下、KK221株とよぶ。)を使用した。本出願人は、KK221株を下記の条件で寄託した。
(1)寄託機関名:独立行政法人製品評価技術基盤機構 特許生物寄託センター
(2)連絡先:〒305-8566 茨城県つくば市東1丁目1番地1 中央第6
(現:千葉県木更津市かずさ鎌足2-5-8 120号室)
電話番号0438-20-5580
(3)受託番号:FERM BP-10987
(4)識別のための表示:Tetragenococcus halophilus Th221
(5)原寄託日:2007年6月25日
(6)ブダペスト条約に基づく寄託への移管日:2008年7月16日
調製した乳酸菌懸濁液のインターフェロンλの産生促進活性を、非ウイルス感染健常者の末梢血より採取及び調製した樹状細胞を用いて以下のとおりに評価した。
非ウイルス感染健常者の末梢血200mlから低密度勾配遠心法により単核球として、磁気細胞分離システム(Militeny社)、セルソーター(BD社)を用いて樹状細胞(Dendritic cell;DC)を単離した。なお、樹状細胞は、BDCA4陽性であるplasmacytoid DC(pDC);BDCA1が陽性であり、かつ、CD19が陰性であるmyeloid cell type1(mDC1);及び、BDCA3が陽性であるmyeloid cell type2(BDCA3DC)の3種の表現型サブセットで単離した。
回収した培養上清を用いて、インターフェロンλ3を化学発光酵素免疫測定法により、スギヤマらの文献[Sugiyama M et al.Hepatol Res.42(11):1089-1099、2012]に記載の方法に準じて測定した。
1.RNaseA処理
KK221株を10mM Tris-HCl(pH8.0)を用いて5×109個/mlとなるように懸濁し、10μg/mlとなるように牛膵臓由来RNaseA(Sigma社)をさらに添加して、RNaseA含有菌懸濁液を調製した。
RNA分解乳酸菌懸濁液又は例1で調製した未処理乳酸菌混濁液と、例1で調製したBDCA3DCとを一定の割合で混合し(BDCA3DC数:乳酸菌数=1:100)、これらの共培養を行った。共培養後の上清を培養開始24時間後に回収し、例1と同様にして化学発光酵素免疫測定法により、上清中のインターフェロンλ濃度を測定した。結果を図3に示す。
乳酸菌の構成成分を認識すると想定されるBDCA3DCにおけるToll様受容体(TLR)に着目し、エンドソームへの取り込みを阻害し、細胞質内に存在するTLR3への会合を阻害するクロロキン、もしくは2本鎖RNAを認識するTLR3を介するシグナル伝達のアダプター分子であるTRIFの阻害剤(製品名「TRIF/TICAM1 Peptide」;NOVUS社)を用いて、インターフェロンλ産生促進活性を評価した。
KK221株に加えて、インターフェロンλ産生を誘導することが知られるラクトコッカス・ラクティス(Lactococcus lactis)に属するJCM20101株及びJCM5805株(特許文献1)を使用し、インターフェロンλ産生促進試験を実施した。
例1に準じて、KK221株を用いて調製した乳酸菌懸濁液又はPoly ICとBDCA3DCとを、乳酸菌数/DC数=100になるように調製して共培養することにより、各種サイトカイン産生促進活性の測定を実施した。
KK221株、JCM20101株及びJCM5805株の生菌数あたりの二本鎖RNA(以下dsRNAとも略す)量、総核酸量及び総核酸中の二本鎖RNAの割合(dsRNA量/総核酸量×100)を以下のとおりに測定した。
KK221株、JCM20101株及びJCM5805株の乾燥菌体量あたりのdsRNA量、総核酸量及び総核酸中の二本鎖RNAの割合を以下のとおりに測定した。
培養殺菌処理液の5×109cfu相当液量を用いた以外は、例6と同様にして、各種乳酸菌の生菌数あたりの二本鎖RNA量を測定した。
Claims (11)
- ストレス条件下で培養が可能である乳酸菌の菌体、菌体成分又は培養物を有効成分として含有する、インターフェロンλ産生促進用組成物。
- 前記インターフェロンλ産生促進用組成物は、BDCA3DCに対するインターフェロンλ産生促進用組成物である、請求項1に記載のインターフェロンλ産生促進用組成物。
- 前記インターフェロンλは、インターフェロンλ3である、請求項1又は2に記載のインターフェロンλ産生促進用組成物。
- 前記菌体、前記菌体成分及び前記培養物は、それぞれ核酸を含有する菌体、菌体成分及び培養物である、請求項1~3のいずれか1項に記載のインターフェロンλ産生促進用組成物。
- 前記ストレス条件下で培養が可能である乳酸菌は、菌体1×1010cfuあたり15,000ng/ml以上の二本鎖RNAを含有する乳酸菌及び菌体1×1010cfuあたり総核酸中の二本鎖RNAの割合が3.5%以上である乳酸菌の少なくとも一方である、請求項4に記載のインターフェロンλ産生促進用組成物。
- 前記インターフェロンλ産生促進用組成物が、インターフェロンλ産生促進用腸溶組成物である、請求項1~5のいずれか1項に記載のインターフェロンλ産生促進用組成物。
- 請求項1~6のいずれか1項に記載のインターフェロンλ産生促進用組成物を含有する飲食品。
- ストレス条件下で培養が可能である乳酸菌を培養して、該乳酸菌の菌体、菌体成分又は培養物を得ることを含む、インターフェロンλ産生促進用組成物の製造方法。
- ストレス条件下で培養が可能である乳酸菌の菌体、菌体成分又は培養物を使用してインターフェロンλの産生を促進する方法。
- インターフェロンλ産生促進用組成物の製造のためのストレス条件下で培養が可能である乳酸菌の菌体、菌体成分又は培養物の使用。
- インターフェロンλの産生を促進するためのストレス条件下で培養が可能である乳酸菌の菌体、菌体成分又は培養物の使用。
Priority Applications (5)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN201780021700.0A CN108883141A (zh) | 2016-04-04 | 2017-04-04 | 用于促进干扰素λ产生的组合物及其制造方法 |
JP2018510622A JP6704990B2 (ja) | 2016-04-04 | 2017-04-04 | インターフェロンλ産生促進用組成物及びその製造方法 |
CA3019975A CA3019975A1 (en) | 2016-04-04 | 2017-04-04 | Composition for promoting interferon .lambda. production and production method therefor |
US16/090,940 US20190099456A1 (en) | 2016-04-04 | 2017-04-04 | Composition for promoting interferon lambda production and production method therefor |
EP17779152.2A EP3424517A4 (en) | 2016-04-04 | 2017-04-04 | COMPOSITION FOR PROMOTING THE PRODUCTION OF INTERFERON LAMBDA AND METHOD FOR THE PRODUCTION THEREOF |
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP2016-075323 | 2016-04-04 | ||
JP2016075323 | 2016-04-04 |
Publications (1)
Publication Number | Publication Date |
---|---|
WO2017175774A1 true WO2017175774A1 (ja) | 2017-10-12 |
Family
ID=60001056
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
PCT/JP2017/014153 WO2017175774A1 (ja) | 2016-04-04 | 2017-04-04 | インターフェロンλ産生促進用組成物及びその製造方法 |
Country Status (6)
Country | Link |
---|---|
US (1) | US20190099456A1 (ja) |
EP (1) | EP3424517A4 (ja) |
JP (1) | JP6704990B2 (ja) |
CN (1) | CN108883141A (ja) |
CA (1) | CA3019975A1 (ja) |
WO (1) | WO2017175774A1 (ja) |
Cited By (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JP2019167327A (ja) * | 2018-03-26 | 2019-10-03 | 森永乳業株式会社 | ビフィドバクテリウム属細菌を有効成分とする、iii型インターフェロン産生促進用組成物 |
Citations (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2009005124A1 (ja) * | 2007-07-04 | 2009-01-08 | Kikkoman Corporation | 乳酸菌由来の2本鎖rna |
WO2012091081A1 (ja) * | 2010-12-28 | 2012-07-05 | キリンホールディングス株式会社 | 乳酸菌を含むインターフェロン産生誘導剤 |
JP2013514062A (ja) * | 2009-12-18 | 2013-04-25 | バヴァリアン・ノルディック・アクティーゼルスカブ | 従来型樹状細胞によるifn‐ラムダの産生及びその使用 |
Family Cites Families (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JP2006028047A (ja) * | 2004-07-14 | 2006-02-02 | Kikkoman Corp | インターロイキン12産生促進剤およびその製造法 |
JP2008214253A (ja) * | 2007-03-02 | 2008-09-18 | Snow Brand Milk Prod Co Ltd | 内臓脂肪減少剤 |
CN102028224B (zh) * | 2007-06-21 | 2012-12-26 | 东宇生物科技股份有限公司 | 抗过敏的乳酸菌 |
JP5531321B2 (ja) * | 2009-03-31 | 2014-06-25 | キッコーマン株式会社 | Il−10産生促進作用を有する免疫賦活組成物 |
JP2016069285A (ja) * | 2014-09-26 | 2016-05-09 | 株式会社アミノアップ化学 | 免疫促進組成物 |
-
2017
- 2017-04-04 CN CN201780021700.0A patent/CN108883141A/zh active Pending
- 2017-04-04 CA CA3019975A patent/CA3019975A1/en not_active Abandoned
- 2017-04-04 EP EP17779152.2A patent/EP3424517A4/en not_active Withdrawn
- 2017-04-04 JP JP2018510622A patent/JP6704990B2/ja active Active
- 2017-04-04 US US16/090,940 patent/US20190099456A1/en not_active Abandoned
- 2017-04-04 WO PCT/JP2017/014153 patent/WO2017175774A1/ja active Application Filing
Patent Citations (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2009005124A1 (ja) * | 2007-07-04 | 2009-01-08 | Kikkoman Corporation | 乳酸菌由来の2本鎖rna |
JP2013514062A (ja) * | 2009-12-18 | 2013-04-25 | バヴァリアン・ノルディック・アクティーゼルスカブ | 従来型樹状細胞によるifn‐ラムダの産生及びその使用 |
WO2012091081A1 (ja) * | 2010-12-28 | 2012-07-05 | キリンホールディングス株式会社 | 乳酸菌を含むインターフェロン産生誘導剤 |
Non-Patent Citations (6)
Title |
---|
JIANG, M. ET AL.: "Innate Immune Responses in Human Monocyte-Derived Dendritic Cells Are Highly Dependent on the Size and the 5 ' Phosphorylation of RNA Molecules", JOURNAL OF IMMUNOLOGY, vol. 187, no. 4, 2011, pages 1713 - 1721, XP055430974 * |
LAUTERBACH, H. ET AL.: "Mouse CD 8alpha+ DCs and human BDCA3+ DCs are major producers of IFN-lambda in response to poly IC.", JOURNAL OF EXPERIMENTAL MEDICINE, vol. 207, no. 12, 2010, pages 2703 - 2717, XP008132858 * |
SACHIYO YOSHIO ET AL.: "C-gata Mansei Kan'en Kanja ni Okeru IFNA Kosansei Jujo Saibo Subset (BDCA3+DC) no Ig i", ACTA HEPATOLOGICA JAPONICA, vol. 53, no. 1, 2012, pages A340, OWS-282, XP009514729 * |
See also references of EP3424517A4 * |
TADAOMI KAWASHIMA ET AL.: "Lactic acid bacteria induces IFN-beta secretion by dendritic cells through toll-like receptors in endosomes", PROCEEDINGS OF THE JAPANESE SOCIETY FOR IMMUNOLOG Y, vol. 39, 2009, pages 181, XP9515882 * |
YOSHIO, S. ET AL.: "Human Blood Dendritic Cell Antigen 3 (BDCA3)+ Dendritic Cells Are a Potent Producer of Interferon-lambda in Response to Hepatitis C Virus.", HEPATOLOGY, vol. 57, no. 5, 2013, pages 1705 - 1715, XP055430972 * |
Cited By (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JP2019167327A (ja) * | 2018-03-26 | 2019-10-03 | 森永乳業株式会社 | ビフィドバクテリウム属細菌を有効成分とする、iii型インターフェロン産生促進用組成物 |
Also Published As
Publication number | Publication date |
---|---|
EP3424517A1 (en) | 2019-01-09 |
EP3424517A4 (en) | 2019-11-13 |
US20190099456A1 (en) | 2019-04-04 |
CN108883141A (zh) | 2018-11-23 |
CA3019975A1 (en) | 2017-10-12 |
JP6704990B2 (ja) | 2020-06-03 |
JPWO2017175774A1 (ja) | 2019-02-21 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
ES2660813T3 (es) | Nueva bacteria de ácido láctico, fármaco, producto alimenticio o bebida, y pienso que contienen la nueva bacteria de ácido láctico | |
KR102146429B1 (ko) | 비피도박테리움 아니말리스 아종 아니말리스 균주 | |
JP5001830B2 (ja) | 免疫賦活用組成物 | |
KR102240196B1 (ko) | 신규한 락토바실러스 파라카세이 균주 | |
ES2407150T3 (es) | Bacteria ácido láctica novedosa con actividad anti-alérgica, agente anti-alérgico, alimento y composición farmacéutica que comprenden, cada uno de ellos, la bacteria ácido láctica, y proceso para la producción del agente anti-alérgico | |
JP7179343B2 (ja) | 新規な乳酸菌株およびそれを含む免疫賦活剤 | |
KR102543494B1 (ko) | 신규 프로바이오틱스 및 이의 용도 | |
JP5337535B2 (ja) | Nk活性増強剤 | |
TW201840328A (zh) | 肽聚醣辨識蛋白質產生促進用組成物 | |
KR101501210B1 (ko) | 항-염증 활성이 우수한 신규 균주 | |
JP5082048B2 (ja) | 免疫賦活作用及び/又はアレルギー抑制作用を有し、且つ胃液耐性を有する新規乳酸菌 | |
JP6821643B2 (ja) | 免疫疾患予防剤 | |
JP6557605B2 (ja) | 乳酸菌を含む腸管バリア機能亢進剤 | |
WO2015046407A1 (ja) | 免疫疾患予防剤 | |
JP4499979B2 (ja) | 病原菌感染抑制用組成物 | |
WO2017175774A1 (ja) | インターフェロンλ産生促進用組成物及びその製造方法 | |
BR122022016606B1 (pt) | Lactobacillus paracasei mortos pelo aquecimento, medicamento, alimento ou bebida, ração, agente promotor da produção de il-12 e uso | |
Kingkaew et al. | Characterization of lactic acid bacteria from fermented fish (pla-paeng-daeng) and their cholesterol-lowering and immunomodulatory effects | |
JP2010057395A (ja) | 腸管免疫調節作用を有する乳酸菌 | |
JP2009112232A (ja) | 免疫賦活作用及び/又はアレルギー抑制作用を有し、且つ胃液耐性を有する新規乳酸菌 | |
JP6480090B1 (ja) | 二本鎖rna高含有乳酸菌の製造方法及び該乳酸菌 | |
KR100930251B1 (ko) | 혈중 콜레스테롤 저하능을 갖는 비피도박테리움 롱검 균주 | |
JP2009254236A (ja) | 新規乳酸菌並びに新規乳酸菌を利用した飲食品及び免疫賦活剤 | |
TW202335676A (zh) | 腸管免疫賦活劑及IgA產生促進劑 | |
TW202335675A (zh) | 腸管免疫賦活劑、IgA産生促進劑及基因表現促進劑 |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
ENP | Entry into the national phase |
Ref document number: 2018510622 Country of ref document: JP Kind code of ref document: A |
|
ENP | Entry into the national phase |
Ref document number: 3019975 Country of ref document: CA |
|
WWE | Wipo information: entry into national phase |
Ref document number: 2017779152 Country of ref document: EP |
|
NENP | Non-entry into the national phase |
Ref country code: DE |
|
ENP | Entry into the national phase |
Ref document number: 2017779152 Country of ref document: EP Effective date: 20181004 |
|
121 | Ep: the epo has been informed by wipo that ep was designated in this application |
Ref document number: 17779152 Country of ref document: EP Kind code of ref document: A1 |