JP2023053926A - plasmacytoid dendritic cell activator - Google Patents

Abstract

To provide a plasmacytoid dendritic cell activator.SOLUTION: Provided is a plasmacytoid dendritic cell activator that contains at least one selected from the group consisting of euglena, paramylum, processed paramylum, and β-1,3-glucan.SELECTED DRAWING: None

Description

The present invention relates to a plasmacytoid dendritic cell activator and the like.

Recently, with the outbreak and epidemic of SARS-CoV-2, awareness of virus infection prevention has increased. Plasmacytoid dendritic cells (pDC) are known as one of the cells responsible for the virus infection defense mechanism. pDCs are activated by virus infection, produce type I interferons (interferon-α, interferon-β, etc.), and exhibit antiviral effects. Therefore, if pDC can be artificially activated, the production of type I interferon can be promoted in the absence of viral infection and in the presence of viral infection, and preventive or therapeutic effects on viral infections can be exhibited.

Euglena is a microalga belonging to the genus Euglena (= Euglena genus) and is used as a food material. In addition, application of Euglena extract to the skin is also performed (Patent Document 1). Paramylon is a β-1,3-glucan produced by Euglena, and is reported to be useful for wound healing and allergy suppression. However, the relationship between euglena or β-1,3-glucan and pDC is still unknown.

Japanese Patent Publication No. 2008-526954

An object of the present invention is to provide a plasmacytoid dendritic cell activator.

As a result of intensive research in view of the above problems, the present inventors have found that plasmacytoid dendritic cells containing at least one selected from the group consisting of euglena, paramylon, processed paramylon, and β-1,3-glucan. We have found that the above problems can be solved by using a cell activator. As a result of further research based on this knowledge, the present invention was completed. That is, the present invention includes the following aspects.

Item 1. A plasmacytoid dendritic cell activator containing at least one selected from the group consisting of euglena, paramylon, processed paramylon, and β-1,3-glucan.

Section 2. Item 2. The plasmacytoid dendritic cell activator according to Item 1, containing at least one selected from the group consisting of euglena, paramylon, and processed products of paramylon.

Item 3. Item 3. The plasmacytoid dendritic cell activator according to Item 1 or 2, which contains Euglena, and the Euglena is Euglena gracilis.

Section 4. Item 3. The plasmacytoid dendritic cell activator according to Item 1 or 2, which contains Euglena, and the Euglena is Euglena gracilis EOD-1 strain (accession number FERM BP-11530).

Item 5. Item 5. The plasmacytoid dendritic cell activator according to any one of Items 1 to 4, for use in promoting type I interferon production.

Item 6. Item 6. The plasmacytoid dendritic cell activator according to any one of Items 1 to 5, which is a food composition, nutritional supplement, food additive, or pharmaceutical.

Item 7. Item 7. The plasmacytoid dendritic cell activator according to any one of Items 1 to 6, which is an oral composition.

Item 8. A type I interferon production promoter containing at least one selected from the group consisting of euglena, paramylon, processed paramylon, and β-1,3-glucan.

According to the present invention, a plasmacytoid dendritic cell activator can be provided.

Fig. 2 shows the measurement results of HLA-DR expression intensity in plasmacytoid dendritic cells (Test Example 1). The vertical axis indicates the expression intensity of HLA-DR (Mean Fluorescence Intensity; M.F.I.). The horizontal axis indicates the group to be measured. * indicates significant (P value <0.05, Mann-Whitney U test) between groups indicated by bars.

As used herein, the expressions "contain" and "include" include the concepts "contain", "include", "consist essentially of" and "consist only of".

In one aspect of the present invention, a plasmacytoid dendritic cell activator (this In the specification, it may be indicated as "the agent of the present invention"). This will be explained below.

1. Euglena Euglena is a microalgae belonging to the genus Euglena (= Euglena genus), and is not particularly limited as long as it is. Specific examples of Euglena include Euglena gracilis , Euglena longa , Euglena caudata , Euglena oxyuris , Euglena tripteris , Euglena proxima , Euglena viridis , Euglena sociabilis , Euglena ehrenbergii , Euglena deses , Euglena pisciformis , Euglena spirogyra , Euglena acus , Euglena geniculata , Euglena intermedia , Euglena mutabilis , Euglena sanguinea , Euglena stellata , Euglena terricola , Euglena klebsi , Euglena rubra , Euglena cyclopicola and the like. Among these, from the viewpoint that the effects of the present invention can be more reliably exhibited, Euglena gracilis is preferable, and Euglena gracilis EOD-1 strain [as of June 28, 2013, Independent Administrative Agency Product Evaluation Under the provisions of the Budapest Treaty, the Patent Organism Depositary Center, National Institute of Technology, NITE-IPOD (2-5-8 Kazusa Kamatari, Kisarazu City, Chiba Prefecture, Japan 292-0818, Room 120) under the provisions of the Budapest Treaty, with the accession number FERM BP-11530. Internationally deposited].

The form of Euglena is not particularly limited as long as it contains the cell body of Euglena or most of its components. Examples of the form of Euglena include dry powder form of Euglena, suspension of Euglena, Euglena extract, etc. Among them, dry powder form of Euglena is preferred.

The paramylon content of Euglena in a dry state is, for example, 50% or more, preferably 60% or more, and more preferably 70% or more.

Euglena may be used singly or in combination of two or more.

2. β-1,3-Glucan and paramylon β-1,3-glucan have a single sugar chain (or sugar chain structure) consisting of glucose linked only by β1,3 bonds as the main chain. There are no particular restrictions. β-1,3-Glucans include not only straight-chain ones but also branched-chain ones.

The weight average molecular weight of the β-1,3-glucan derivative is not particularly limited, but is, for example, 1×10 ⁴ to 2×10 ⁶ , preferably 5×10 ⁴ to 1×10 ⁶ , more preferably 1×10 ⁵ to There are 1×10 ⁶ . In addition, the weight average molecular weight can be measured by the GPC method.

The β-1,3-glucan may be one obtained by chemical synthesis, but natural β-1,3-glucan produced by various organisms is preferable from the viewpoint of availability. Examples of natural β-1,3-glucans include paramylon, curdlan, laminaran, callose, lentinan, schizophyllan and the like. Among these, paramylon is particularly preferred. Paramylon will be described below.

Paramylon is a β-1,3-glucan derived from Euglena, and is not particularly limited as far as it is concerned.

Euglena from which paramylon is derived is the same as described in the above "1. Euglena".

The mass average molecular weight of paramylon is not particularly limited, but is, for example, 1×10 ⁴ to 5×10 ⁶ , preferably 2×10 ⁴ to 1×10 ⁶ , more preferably 5×10 ⁴ to 1×10 ⁶ , still more preferably. is between 1×10 ⁵ and 5×10 ⁵ .

The mass average molecular weight can be measured by SEC-MALS analysis under the following conditions:
Detector: Multi-angle scattering detector (DAWN HELEOS II manufactured by Wyatt Technology) Differential refractometer detector (Optilab T-rEX manufactured by Wyatt Technology) Column used: 2 TSKgel α-M (manufactured by Tosoh)
Mobile phase: DMSO with 0.05M potassium bromide
Flow rate: 0.5 mL/min.

In Euglena cells, paramylon normally exists as paramylon particles, which are highly-accumulated triple-helix structures formed by β-1,3-glucan chains on certain regular bases.

The shape of the paramylon particles is not particularly limited, but is usually a flattened spheroid.

The particle size distribution of paramylon particles is not particularly limited, but is, for example, 0.5 to 15 μm, preferably 1 to 6 μm. Also, the average particle size of the paramylon particles is not particularly limited, but is, for example, 1 to 10 μm, preferably 2 to 4 μm.

The form of paramylon is not particularly limited as long as it contains paramylon. Examples of the form of Euglena include a dry powder form of paramylon, a suspension of paramylon, etc. Among them, a dry powder form of paramylon is preferred.

Paramylon may be used singly or in combination of two or more.

3. Method for Producing Euglena and Paramylon Euglena can be prepared in large quantities by a method including a step of culturing Euglena contained in a liquid (culturing step). The culture step can be performed, for example, according to a known method (eg, the method described in Japanese Patent No. 5883532). In the culturing step, typically, Euglenoid microalgae are cultured under aerobic conditions while stirring a liquid (culture solution) containing water, Euglena, and nutrients that Euglena can use.

Nutrients include sugars (monosaccharides such as glucose (glucose) and fructose (fructose)), minerals (such as sodium, potassium, magnesium, calcium, iron, zinc, molybdenum, copper, phosphorus, nitrogen, sulfur, or boron). etc.), B vitamins (e.g. vitamin B1 (thiamine), vitamin B2 (riboflavin), niacin, pantothenic acid, vitamin B6 (pyridoxine, pyridoxal, or pyridoxamine), vitamin B12 (cyanocobalamin), folic acid, biotin, etc.) be done. The concentration of nutrients in the culture medium is not particularly limited as long as it is a concentration that allows Euglena to survive, proliferate, and the like.

The light conditions for the culturing step are not particularly limited, and the culturing step may be performed under either light or dark conditions. When cultured in heterotrophic culture, it is cultured under dark conditions. As a light condition, a normal light intensity for growing algae can be adopted. Dark conditions include, for example, less than 10 μmol/m ² /s, preferably completely dark conditions with no light.

The culture temperature in the culture step is not particularly limited as long as it is a temperature at which Euglena can grow. As the culture temperature (temperature of the culture solution), for example, 20°C to 35°C is adopted.

The pH of the liquid in the culturing step is not particularly limited as long as it allows Euglena to proliferate. A pH of 3.0 to 5.5, for example, is adopted as the pH at which Euglena can grow.

After the culturing step, it is preferable to concentrate Euglena by liquid centrifugation, gravity separation, or the like. The obtained Euglena can be subjected to additional processing (for example, suspension in liquid, dispersion in water or oil, extract extraction, dry powderization, etc.) depending on the desired form.

Paramylon particles can be produced by separating, isolating, or purifying from Euglena according to or according to known methods (eg, the method described in Japanese Patent No. 5883532). Paramylon particles can be easily obtained, for example, by recovering intracellular components obtained by disrupting Euglena cell membranes. Moreover, if necessary, the paramylon particles may be purified. Various methods for purifying paramylon particles are known (eg, Japanese Patent No. 5883532), and can be performed according to those methods. Examples of the purification step include a surfactant treatment step and a washing step. The resulting Euglena can be subjected to additional processing (eg, suspension in liquid, dispersion in water or oil, dry powderization, etc.) depending on the desired form.

Four. Paramylon-processed product The paramylon-processed product is obtained by subjecting paramylon to a processing treatment, such as a physical treatment or a chemical treatment, and is not particularly limited as long as it is processed. Examples of paramylon-processed products include fiberized paramylon and amorphous paramylon. Amorphous paramylon can be obtained by chemical treatment according to or in accordance with a known method, for example, using the method described in JP-A-2011-184592.

Fiberized paramylon is preferable as the processed paramylon. Fiberized paramylon will be described below.

Fibrosis paramylon is Euglena-derived β-1,3-glucan, and is not particularly limited as long as it is in a fibrous form. Until now, amorphous paramylon obtained by chemically treating paramylon particles (alkali treatment, etc.) has been reported. Since it is an amorphous mass, it is not included in fibrillated paramylon.

The weight average molecular weight of fiberized paramylon is not particularly limited, but is, for example, 1×10 ⁴ to 2×10 ⁷ , preferably 1×10 ⁵ to 5×10 ⁵ .

The weight average molecular weight can be measured by SEC-MALS analysis by the following method: Detector: Multi-angle scattering detector (DAWN HELEOS II manufactured by Wyatt Technology) Differential refractometer detector (Optilab T manufactured by Wyatt Technology -rEX) Columns used: 2 TSKgel α-M (manufactured by Tosoh)
Mobile phase: DMSO with 0.05M potassium bromide
Flow rate: 0.5 mL/min.

The fiber diameter of the fiberized paramylon is not particularly limited, but is, for example, 10 to 500 nm, preferably 20 to 300 nm, more preferably 50 to 200 nm. The diameter of the fibers of fibrillated paramylon can usually be measured based on an electron microscope image of fibrillated paramylon.

The settling volume of fibrillated paramylon in water is not particularly limited, but is, for example, 30 to 300 mL/g, preferably 50 to 250 mL/g, more preferably 70 to 200 mL/g.

Settling volume in water can be measured according to or in accordance with the following method:
Measure according to the method described in "Supervised by the Japan Dietary Fiber Society, Edited by the Japan Dietary Fiber Society Editorial Committee (2008) Dietary Fiber -Basics and Applications- 3rd Edition, p.111 Daiichi Publishing, Tokyo" . Specifically, it is as follows. Weigh 125 mg of the sample slurry test sample into a 25 mL volume plastic tube, calculated as dry mass, and shake the plastic tube vigorously by hand to agitate the contents. Then transfer the contents to a 25 mL graduated cylinder and add pure water to bring the volume to 25 mL. After stirring the liquid in the graduated cylinder, leave it at 37°C for 24 hours. This causes the sample to settle, resulting in two layers, one containing mainly the precipitated sample (lower layer) and the other containing mainly water (upper layer), separated through the interface. Calculate the volume of the lower layer from the scale of the graduated cylinder, divide the obtained volume by the sample mass (dry mass), and calculate the volume settled in water (mL/g). Perform the test 3 or 4 times and calculate the average value and standard deviation.

Fibrillating paramylon has a relatively high resistance to enzymatic degradation. For example, the amount of monomer (glucose) produced by the decomposition of β-glucanase is, for example, 0.1-50 mg, preferably 1-10 mg, per 1 g of fibrillated paramylon.

This amount can be measured according to or according to the following methods:
Reaction solution [test substance 30 mg (dry weight), buffer solution (B0156 manufactured by Tokyo Chemical Industry Co., Ltd., potassium hydrogen phthalate-sodium hydroxide buffer (pH 4.0)) 5 mL, enzyme solution (endo- 1,3-β-Glucanase (enzyme content: 50 units/mL)) 0.1 mL, pure water, reaction volume 10 mL], shake horizontally at 40°C for 24 hours at 45 rpm. After shaking, immediately store frozen and lyophilize for concentration. After freeze-drying, add 0.5 mL of pure water to each sample and stir (20-fold concentration). The operation of centrifuging (10000G, 5 minutes, 4°C) and collecting the supernatant is repeated twice. The glucose concentration in the recovered supernatant is measured using a measurement kit (Glucose CII-Test Wako manufactured by Wako Pure Chemical Industries, Ltd.). Based on the measured values, calculate the amount of glucose produced (mg) per 1 g of the test substance.

Fiberized paramylon has relatively low solubility in alkaline solutions. For example, fiberized paramylon does not dissolve in 0.1-0.3M sodium hydroxide aqueous solution. Here, "does not dissolve" means, for example, that the absorbance (660 nm) of the solution after suspending the fibrillated paramylon in the aqueous solution (for example, immediately after to 1 hour later) is, for example, 0.1 or more, preferably 1.0. means greater than or equal to

Solubility can be measured according to or in accordance with the following methods:
Suspend 250 mg (dry weight) of the test substance in 10 mL of test solution (pure water, 0.1 M NaOH aqueous solution, 0.3 M NaOH aqueous solution) in a vial. The absorbance at 660 nm of the liquid in the vial is measured after shaking the vial vigorously by hand for 20 seconds and after shaking on a shaker at 80 rpm for 1 hour. The absorbance is measured using a spectrophotometer V-730 manufactured by JASCO Corporation.

The relative value of crystallinity of fiberized paramylon to granular paramylon (crystallinity of fiberized paramylon/crystallinity of granular paramylon) is, for example, 0.60 to 0.90, preferably 0.65 to 0.80.

Crystallinity can be measured according to or according to the following methods:
XRD measurement is performed on the test substance. The conditions are as follows. Equipment: PANalytical X'Pert3 Powder, tube voltage: 45kV, tube current: 40mA, measurement range: 5.005 to 50.018°, measurement interval: 0.013°, analysis software: HighScore. The degree of crystallinity is analyzed by the ratio of the intensity of the amorphous part to the intensity of the crystalline part at 2θ=5 to 80°. The analysis was performed after removing the background from each measurement sample (background setting Auto, venting factor 0, granularity 100), and the amorphous part was determined by the tangent line passing through 2θ = 14, 29°. do. The pending factor and granularity condition for determining each amorphous portion is 0/20.

Fiberized paramylon may be in a form dispersed in a solvent such as water, or may be in a dry form. Fiberized paramylon, even in dry form, can be redispersed in water.

As used herein, the term "dry form" indicates that the water content is 15% by mass or less, preferably 10% by mass or less, and more preferably 5% by mass or less.

As the fibrillated paramylon, a fibrillated product of paramylon particles obtained by physically fibrillating the paramylon particles can be preferably used. In addition, a fibrillated product of Euglena obtained by applying this fibrillation treatment to Euglena can also be used as fiberized paramylon.

The fibrillation treatment hardly breaks the hydrogen bonds of β-1,3-glucans present in the paramylon particles (for example, 10% or less, 5% or less, 2% or less of the hydrogen bonds of β-1,3-glucans , 1% or less), or unraveling part or all of the β-1,3-glucan chains present in the paramylon particles or the triple helix structure formed by them. is not particularly limited as long as the processing can be performed. Preferably, the β-1,3-glucan present in the paramylon particles is fibrillated without breaking the hydrogen bonds to form fibrils. Any known treatment capable of grinding (shearing) or pulverizing (preferably grinding (shearing)) fine particles such as paramylon particles can be employed as defibration treatment.

The fibrillation treatment can be performed using a known device such as a grinder (shearer) or a grinder. Examples of apparatuses used for fibrillation treatment include a stone grinder, a jet mill, a twin-screw kneader, a high-pressure homogenizer, a high-pressure emulsifier, a twin-screw extruder, and a bead mill. Among these, stone mills and bead mills are preferred.

The defibration treatment can be performed wet or dry. A wet fibrillation treatment is preferable because the fiberized paramylon can be more efficiently dispersed in the solution. The solvent in the wet method is not particularly limited as long as it can disperse the fiberized paramylon, and water can be preferably used.

The defibration treatment may be performed singly or in combination of two or more. In addition, it may be a partially fibrillated paramylon, and is intended in the present invention as long as it contains fibrillated paramylon.

Five. At least one selected from the group consisting of euglena , paramylon, processed paramylon, and β-1,3-glucan (hereinafter sometimes referred to as the “active ingredient of the present invention”) is a plasmacytoid tree It can be used as an active ingredient of a plasmacytoid dendritic cell activator, since it has an activating effect on dendritic cells.

Plasmacytoid dendritic cells (pDC: plasmacytoid dendritic cells) can be defined as CD123-positive and BDCA4-positive cells. Activation of plasmacytoid dendritic cells is determined using indicators such as increased expression of activation markers (HLA-DR, etc.) in plasmacytoid dendritic cells, the ratio of plasmacytoid dendritic cells among dendritic cells, etc. be able to. Plasmacytoid dendritic cells can be sorted and the expression of activation markers can be measured by staining with antibodies against surface proteins followed by FACS analysis, mass cytometry, and the like.

Plasmacytoid dendritic cells, when activated, produce increased levels of type I interferon. Type I interferons include, for example, interferon-α and interferon-β. The active ingredient of the present invention and the agent of the present invention can promote production of type I interferon in the absence of virus infection and virus infection. Therefore, the active ingredient of the present invention and the agent of the present invention can be used to promote production of type I interferon. In one aspect, the present invention relates to a type I interferon production promoter containing the active ingredient of the present invention. In the present specification, this agent may also be referred to as "the agent of the present invention" together with the plasmacytoid dendritic cell activating agent described above.

The active ingredient of the present invention and the agent of the present invention can exhibit antiviral effects based on activation of plasmacytoid dendritic cells and promotion of type I interferon production. Type I interferon has the effect of activating the viral replication inhibitory mechanism, activating the acquired immune system such as CD8 ⁺ T cells, CD4 ⁺ T cells, B cells, etc., and activating NK cells. action, action to eliminate viruses from the living body, action to promote immune response in the early and late stages of virus infection, and the like. The active ingredient of the present invention and the agent of the present invention can be used for antiviral purposes or for the prevention or amelioration of viral infections. The term "improvement" means amelioration or alleviation of symptoms or conditions, prevention or delay of worsening of symptoms or conditions, reversal, prevention or delay of progression of symptoms or conditions, and includes "treatment".

Target viruses are not particularly limited. Viruses include, for example, influenza virus (e.g., type A, type B, etc.), rubella virus, Ebola virus, coronavirus, measles virus, varicella-zoster virus, mumps virus, arbovirus, respiratory syncytial virus, SARS virus, hepatitis virus ( enveloped viruses such as hepatitis B virus, hepatitis C virus, etc.), yellow fever virus, AIDS virus, rabies virus, hantavirus, dengue virus, nipah virus, lyssa virus; adenovirus, norovirus, rotavirus , human papillomavirus, poliovirus, enterovirus, coxsackievirus, human parvovirus, encephalomyocarditis virus, poliovirus, rhinovirus, and other non-enveloped viruses (viruses without envelopes).

Coronaviruses are viruses that belong to the subfamily Orthocoronavirus. Examples of coronaviruses include alphacoronavirus, betacoronavirus, gammacoronavirus, deltacoronavirus, and the like. Betacoronavirus genus includes SARS-associated coronavirus (SARSr-CoV), broad coronavirus HKU1, MERS coronavirus, and the like. SARSr-CoV includes SARS-CoV-2, SARS-CoV-1, and the like.

Furthermore, the active ingredient of the present invention can be used for the following uses, purposes, and targets:
(a) Maintains, enhances, and supports immune function (b) Supports maintenance of immune function (c) Maintains, enhances, and supports immunity (d) Maintains, enhances, and supports immunity (e) ) acting on plasmacytoid dendritic cells (f) maintaining, enhancing, and supporting the function of plasmacytoid dendritic cells (g) activating plasmacytoid dendritic cells (h) plasmacytoid dendritic cells (i) Activate the immune control tower (j) Support the activation of the immune control tower (k) Maintain/enhance/support the function of the immune control tower (l) Immune cells (m) Supports the activation of all immune cells (n) Promotes the production of type I interferon (o) Maintains/improves the number of plasmacytoid dendritic cells (p) Plasmacytoid dendrites It can also be used to maintain/promote differentiation into cells.

The agent of the present invention is used in various fields, for example, food compositions (including health foods, health promoting agents, dietary supplements (such as supplements)), food additives, cosmetics, cosmetic additives, pharmaceuticals, reagents, feeds, etc. can be used as The agent of the present invention is preferably an oral composition.

The form of the agent of the present invention is not particularly limited, and can take a form commonly used in each application depending on the application.

The form of the agent of the present invention, when used as a food composition, includes liquid, gel or solid foods such as juices, soft drinks, teas, soups, beverages such as soy milk, salad oils, dressings, yoghurts and jellies. , puddings, sprinkles, powdered milk for infants, cake mixes, dairy products (eg, powder, liquid, gel, solid, etc.), bread, confectionery (eg, cookies, etc.), and the like.

Examples of the form of the agent of the present invention include emulsions, cosmetic liquids, face creams, hand creams, lotions, body soaps, shampoos, rinses, cosmetic gels, packs, foundations, lip balms, and facial cleansers when the application is cosmetics. agents and the like.

Forms of the agent of the present invention include ointments, liquids for external use (liniments, lotions, etc.), sprays (aerosols for external use, pump sprays, etc.), creams, and gels when the application is a medicine. , patches (plasters, tapes such as plasters (reservoir type, matrix type, etc.), cataplasms, patches, microneedles, etc.), eye drops, ophthalmic ointments, nasal drops, suppositories, semi-rectal Formulations suitable for parenteral intake such as solid formulations and enemas (especially formulations for external use); Formulations suitable for oral intake such as pills, granules, fine granules, powders, hard capsules, soft capsules, dry syrups, liquids (including drinks, suspensions, and syrups), and jellies ( oral dosage forms).

As the form of the agent of the present invention, when the application is an additive, a health promoting agent, a nutritional supplement (such as a supplement), for example, a tablet (orally disintegrating tablet, chewable tablet, effervescent tablet, troche, jelly) drops, etc.), pills, granules, fine granules, powders, hard capsules, soft capsules, dry syrups, liquids (including suspensions and syrups), jellies, and the like.

The agent of the present invention may further contain other ingredients as necessary. Other components include food compositions (including health foods, health-promoting agents, dietary supplements (supplements, etc.)), food additives, cosmetics, cosmetic additives, pharmaceuticals, reagents, ingredients that can be added to feeds, etc. Although not particularly limited as long as it ingredients, fragrances, chelating agents, and the like.

The content of the active ingredient in the agent of the present invention depends on the application, the mode of use, the state of the application target, etc., and is not limited, but for example, 0.0001 to 100% by mass, preferably 0.001 to 50% by mass. can do.

The amount of application (e.g., administration, ingestion, inoculation, etc.) of the agent of the present invention is not particularly limited as long as it is an effective amount for expressing plasmacytoid dendritic cell activating action. , generally 0.1 to 10000 mg/kg body weight per day. The above dosage is preferably applied in divided doses once or more (eg, 1 to 3 times) per day, and can be adjusted as appropriate depending on age, disease state, and symptoms.

In a preferred embodiment of the present invention, the Euglena dosage (dry weight) is preferably 100-10000 mg, more preferably 1000-7000 mg, and even more preferably 2000-5000 mg per day. The application period is preferably 1 week or longer, more preferably 4 weeks or longer, and still more preferably 8 weeks or longer, and can be applied for a longer period (10 weeks or longer, 15 weeks or longer, or 20 weeks or longer). . Since the active ingredient of the present invention is naturally derived and highly safe, the upper limit of the application period is not particularly limited, but is, for example, 3 years, 1 year, 6 months, and 4 months.

EXAMPLES The present invention will be described in detail below based on examples, but the present invention is not limited by these examples.

Production example 1
Euglena gracilis EOD-1 strain (NITE-IPOD) dry powder (manufactured by Kobelco Eco-Solutions, paramylon content of 70% or more) is made into capsules with the following formulation (capsule composition Components: pullulan and sorghum pigment) were used as test foods, and capsule tablets containing cellulose (capsule components: pullulan and sorghum pigment) were produced as control foods.

Test example 1
The outline of the test is as follows.
・Test meal intake period: 12 weeks ・Test meal intake: 15 capsules/day (5 capsules each before breakfast, lunch, and dinner)
- Subjects: 8 (Table 2).

Two types of test diets were used, a test diet and a control diet (placebo) (Manufacturing Example 1). .

15 capsules of the test meal were taken daily (3 times a day: 5 capsules each before breakfast, lunch, and dinner) for 12 weeks. The total content of ingredients in 15 capsules is as follows. Test food: Euglena gracilis EOD-1 strain containing 3.75 g (containing 2.6 g of paramylon). Control diet: Contains 2.6 g of cellulose.

At the end of the study (12 weeks after the start of intake), blood was collected from each subject using a BD Vacutainer CPT ^™ collection tube for mononuclear cell separation (Japan BD). Peripheral blood mononuclear cells (PBMC) are isolated from blood, suspended in 1 mL of CELLBANKER 1 (Nippon Zenyaku Kogyo), and stored at -80°C until the date of measurement. bottom. Frozen PBMCs were thawed, stained with FITC anti-human CD123 (Bio Legend), APC anti-human BDCA-4 (Bio Legend), and Per Cp-Cy ^TM Anti-Human HLA-DR (BD Biosciences), followed by FACS. After washing with a buffer (0.5% BSA in PBS buffer), the cells were fixed with 4% paraformaldehyde and measured using a BD FACS Via ^™ Flow Cytometer (BD Japan). Using BD FACS Via ^TM Research Software (BD Japan), CD123-positive and BDCA4-positive cells were defined as plasmacytoid dendritic cells (pDC), and HLA-DR expression intensity (Mean Fluorescence Intensity; MFI) in pDC was measured. Analyzed.

The results are shown in FIG. The expression level of HLA-DR in pDC was higher in the test diet intake group than in the control diet intake group. HLA-DR is known to be an indicator of pDC activation. From this, it was found that pDC was activated by ingestion of the test food. The above results were presumed to be due to paramylon contained in Euglena.

Test example 2
The outline of the test is as follows.
・Test meal intake period: 12 weeks ・Test meal intake: 2 capsules/day
・Subjects: 23 males and females aged 50 to 65 (12 in the test diet group, 11 in the control diet group).

Two types of test diets were used, a test diet and a control diet (placebo) (Manufacturing Example 1). .

Two capsules of the test meal were taken daily for 12 weeks. The total content of ingredients in 2 capsules is as follows. Test food: Euglena gracilis EOD-1 strain containing 444 mg (containing 350 mg of paramylon). Control diet: containing 350 mg cellulose.

Before the start of intake of the test diet and after 12 weeks of intake, 5 mL of blood was collected from each subject using a heparinized blood collection tube and used as heparinized whole blood. PBMC were isolated by centrifugation and stained using Maxpar Direct Immune Profiling Assay kit (Fluidigm). The percentage of plasmacytoid dendritic cells (pDC) among the dendritic cells was measured by mass cytometry. Measurements by mass cytometry were performed with Helios (Fluidigm). Data analysis was performed using Flow Jo software (BD). The value obtained by subtracting the average pDC ratio (%) in dendritic cells after 12 weeks of ingestion from the average ratio (%) of pDC in dendritic cells before the start of intake of the test diet was calculated as Amount of change (%).

Table 3 shows the results. In Table 3, the p-value indicates the p-value obtained by Welch's t-test.

The amount of change (%) in the control diet group was -8.80, while the amount of change (%) in the test diet group was -2.86. From this, it was found that Euglena has the effect of suppressing the decreasing tendency of the proportion of pDC in dendritic cells (that is, improving the proportion = improving the activity of the whole pDC). The above results were presumed to be due to paramylon contained in Euglena.

Claims (8)

A plasmacytoid dendritic cell activator containing at least one selected from the group consisting of euglena, paramylon, processed paramylon, and β-1,3-glucan. 2. The plasmacytoid dendritic cell activator according to claim 1, containing at least one selected from the group consisting of euglena, paramylon, and processed products of paramylon. 2. The plasmacytoid dendritic cell activator according to claim 1, which contains Euglena, and said Euglena is Euglena gracilis. 2. The plasmacytoid dendritic cell activator according to claim 1, which contains Euglena, and the Euglena is Euglena gracilis strain EOD-1 (accession number FERM BP-11530). The plasmacytoid dendritic cell activator according to any one of claims 1 to 4, for use in promoting type I interferon production. The plasmacytoid dendritic cell activator according to any one of claims 1 to 4, which is a food composition, dietary supplement, food additive, or pharmaceutical. The plasmacytoid dendritic cell activator according to any one of claims 1 to 4, which is an oral composition. A type I interferon production promoter containing at least one selected from the group consisting of euglena, paramylon, processed paramylon, and β-1,3-glucan.

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