Method for treating allergic disease, and composition and device for use in the method

Abstract

To provide a novel method for treating allergic diseases, or a composition for use in the method.SOLUTION: The present invention provides a gaseous composition for use in a method for treating an allergic disease in a subject, the composition being a gaseous composition comprising hydrogen gas as an active ingredient, the method including a step of causing the subject to inhale the composition comprising hydrogen gas as an active ingredient such that the inhaled hydrogen gas concentration falls within a predetermined range.SELECTED DRAWING: Figure 8

Description

Technical Field

[0001] The present disclosure relates to a method for treating allergic diseases, as well as a composition and an apparatus for use in the method.

Background Art

[0002] Allergic rhinitis is a type I allergic disease of the nasal mucosa characterized by paroxysmal and recurrent sneezing, watery rhinorrhea, and nasal congestion. The prevalence of allergic rhinitis has been increasing year by year, and currently, it is said that about 50% of the population in Japan suffers from allergic rhinitis, which has become a serious social problem. The number of patients with allergic diseases is increasing rapidly not only in Japan but also worldwide, and effective preventive and treatment measures are eagerly desired.

[0003] As a method for preventing or treating allergic diseases, a method using drugs such as steroid drugs and antihistamines is mainstream. However, steroid drugs have many side effects such as susceptibility to infectious diseases due to their immunosuppressive effects, while antihistamines have problems such as causing drowsiness as side effects.

[0004] Therefore, various methods for preventing or treating allergic diseases that can be used more safely have been studied. For example, Patent Document 1 discloses a composition for improving allergic rhinitis symptoms containing β-carotene contained in various vegetables including carrots as an active ingredient. On the other hand, Patent Document 2 discloses a composition for suppressing allergic rhinitis containing 15-hydroxyicosapentaenoic acid, which is a metabolite of omega-3 fatty acids ingested through diet, as an active ingredient.

[0005] However, there are still patients suffering from allergic rhinitis. Against this background, a more effective and easily applicable method for dealing with allergic rhinitis is desired.

Prior Art Documents

Patent Documents

[0006]

Patent Document 1

Patent Document 2

Patent Document 3

Patent Document 4

Patent Document 5

Summary of the Invention

Problems to be Solved by the Invention

[0007] The present invention has been made in view of the problems of the prior art as described above, and in one aspect, an object thereof is to provide a new method for treating allergic diseases or a composition for use in the method.

Means for Solving the Problems

[0008] In order to solve the above problems, the present inventors established a model animal for allergic diseases and conducted various studies. As a result, it was found that when hydrogen gas was inhaled by model mice showing symptoms of allergic rhinitis at a predetermined inhalation concentration, the symptoms of allergic rhinitis were significantly improved.

[0009] In recent years, it has been found that hydrogen gas has various health promotion effects. For example, its applications to ischemia-reperfusion injury, cerebral infarction (Patent Document 3), Parkinson's disease (Patent Document 4), and dementia (Patent Document 5) are known. On the other hand, as far as the present inventors know, there has been no solid evidence regarding its application to allergic diseases. Against this background, the above finding that the symptoms of allergic rhinitis are significantly improved when hydrogen gas is inhaled at a predetermined inhalation concentration was also surprising to the present inventors.

[0010] Here, the inhalation concentration of hydrogen gas means the concentration of hydrogen gas in the gas inhaled by the subject, and does not necessarily match the supply concentration of hydrogen gas. For example, hydrogen gas is often supplied to the subject using a nasal cannula or the like. In this case, the hydrogen gas supplied to the subject is inhaled by the subject together with the air present in the surroundings. Therefore, the hydrogen gas supplied to the subject is diluted by the inhaled air. For example, when hydrogen gas with a supply amount of 120 mL / min and a concentration of 4% is supplied to the subject, assuming an adult breathing volume of 7,000 mL / min, the inhalation concentration of hydrogen gas in the subject is lower than 4%, and is estimated to be about 0.07% (= 120 mL / min × 4% / 7,000 mL / min). That is, the hydrogen concentration in the gas actually inhaled by the subject tends to be much lower than the supply concentration of hydrogen gas.

[0011] The present inventors focused on the discrepancy between the inhalation concentration of hydrogen gas and the supply concentration of hydrogen gas, and considered that the reason why the expected therapeutic effects have not been obtained in various treatments claiming the health promotion effect of hydrogen gas lies here. Therefore, the present inventors established an experimental system capable of precisely controlling the hydrogen gas concentration in the gas inhaled by the subject, and as a result of further intensive research efforts, when hydrogen gas was inhaled by a subject suffering from allergic disease symptoms at an inhalation concentration within a predetermined range, particularly at an inhalation concentration of 2.0 ± 0.5 v / v%, the inventors obtained the finding that the symptoms of allergic diseases, particularly allergic rhinitis, were significantly improved, and completed the present invention. As far as the present inventors know, there is no precedent for a study that has examined in detail the relationship between the inhalation concentration of hydrogen gas and allergic diseases, and it can be said that it is extremely useful.

[0012] That is, in one aspect, the present invention is a method for treating allergic diseases in a subject, which solves the above problems by providing a method including a step of causing the subject to inhale a composition containing hydrogen gas as an active ingredient so that the inhalation concentration of hydrogen gas falls within a predetermined range.

[0013] Also, in another aspect, the present invention A composition for use in a method of treating allergic diseases in a subject, wherein the composition is a gaseous composition containing hydrogen gas as an active ingredient, and the method includes a step of causing the subject to inhale the composition containing hydrogen gas as an active ingredient such that the inhalation concentration of hydrogen gas falls within a predetermined range. The present invention solves the above problems by providing the composition.

Advantages of the Invention

[0014] According to the present invention, allergic diseases or their symptoms can be effectively treated.

Brief Description of the Drawings

[0015]

Figure 1

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Figure 8

Embodiments for Carrying Out the Invention

[0016] Hereinafter, the present invention will be described in more detail.

[0017] In one aspect, the present invention provides a method for treating an allergic disease in a subject, comprising the step of causing the subject to inhale a composition containing hydrogen gas as an active ingredient such that the inhalation concentration of hydrogen gas falls within a predetermined range.

[0018] As used herein, the term "allergic disease" means a disease or its symptoms caused by an immune reaction to an antigen. There is no particular limitation on the specific types of allergic diseases. However, as shown in the experimental examples described later, for example, allergic rhinitis and / or allergic conjunctivitis may be particularly suitable. Examples of antigens that cause these allergic diseases include, but are not limited to, dust, house dust, mites, pet hair and skin, pollen, mold, food, drugs, etc.

[0019] "Allergic rhinitis" is a type I allergic disease of the nasal mucosa characterized by paroxysmal and recurrent sneezing, watery rhinorrhea, and nasal congestion. As used herein, the term "allergic rhinitis" includes both perennial allergic rhinitis and seasonal allergic rhinitis (hay fever) unless otherwise specified.

[0020] On the other hand, as used herein, "allergic conjunctivitis" is an inflammatory disease of the conjunctiva involving type I allergy, and refers to a condition accompanied by any subjective and objective symptoms such as eye itching, foreign body sensation, watering, and tearing. Allergic conjunctivitis can be vernal catarrh, atopic keratoconjunctivitis, perennial allergic conjunctivitis, and seasonal allergic conjunctivitis. As used herein, the term "allergic conjunctivitis" includes any of these allergic conjunctivitis unless otherwise specified.

[0021] As used herein, "treatment" is a concept that includes the cure, improvement, suppression, alleviation, and prevention of a disease or its symptoms. That is, as used herein, when referring to the treatment of an allergic disease, it means the cure, improvement, suppression, alleviation, and prevention of the allergic disease, as well as the cure, improvement, suppression, alleviation, and prevention of at least one or more of the symptoms of the allergic disease. Therefore, the treatment of allergic rhinitis can be the cure, improvement, suppression, alleviation, and prevention of one or more symptoms selected from sneezing, watery rhinorrhea, and nasal congestion, while the treatment of allergic conjunctivitis can be the cure, improvement, suppression, alleviation, and prevention of one or more symptoms selected from eye itching, foreign body sensation, watering eyes, and tearing. In addition, as shown in the experimental examples described later, inhalation of hydrogen gas at a predetermined inhalation concentration can have a particularly remarkable effect on suppressing sneezing in allergic rhinitis. Therefore, the above method according to one aspect of the present invention can be particularly preferably used for the cure, improvement, suppression, alleviation, and prevention of sneezing in allergic rhinitis.

[0022] Moreover, according to the findings discovered by the present inventors, the method for treating an allergic disease according to one aspect of the present invention can exhibit a particularly remarkable inhibitory effect on allergic diseases when applied to a subject with an increased eosinophil count compared to a healthy subject. That is, in a certain preferred embodiment, the subject to whom the method for treating an allergic disease according to one aspect of the present invention can be applied can be a subject whose eosinophil count is increased or suspected of being increased compared to a healthy subject.

[0023] The subject to which the method for treating allergic diseases according to one aspect of the present invention can be applied may be a subject whose eosinophil count is increased or suspected of being increased compared to a healthy subject. The eosinophil count does not necessarily have to be measured, but the eosinophil count may be measured if necessary. That is, in one preferred embodiment, the method for treating allergic diseases according to one aspect of the present invention may include a step of measuring the eosinophil count of the subject and a step of determining whether the measured eosinophil count is increased compared to a healthy subject. In this case, when it is determined that the measured eosinophil count is increased compared to a healthy subject, the subject may be made to inhale a composition containing hydrogen gas as an active ingredient. The healthy subject does not necessarily have to be a different individual from the subject, and past data measured for the subject in a healthy state may be used as a comparison control. There is no particular limitation on the degree of increase in the eosinophil count when compared to a healthy subject, but it is preferably a subject in which the eosinophil count has increased by 2 times or more, more preferably 3 times or more, even more preferably 4 times or more, and even more preferably 5 times or more compared to a healthy subject.

[0024] The eosinophil count can be measured by an appropriate method by those skilled in the art, and there is no particular limitation on the specific measurement method. For example, as shown in the experimental examples described later, a specimen collected from the subject may be applied to a slide glass, the specimen applied to the slide glass may be stained, and observed with an optical microscope. There is no particular limitation on the staining method, but for example, eosinostain staining (Hansel staining) or Giemsa staining may be used. There is no particular limitation on the specimen used for measuring the eosinophil count, but for example, in the case of a subject suffering from or suspected of suffering from allergic rhinitis or allergic conjunctivitis, nasal mucus or tear fluid is preferably used respectively.

[0025] The subject to which the method for treating allergic diseases according to one aspect of the present invention can be applied may be a human or an animal other than a human. The animal other than a human may preferably be a mammal, for example, a dog, a rabbit, a cow, a horse, a camel, a giraffe, a rhinoceros, a monkey, an elephant, a tiger, a cat, a hamster, a guinea pig, a sheep, a pig, a mouse, a guinea pig, a goat, a lion, a rat, a meerkat, but is not limited thereto. Note that mammals other than humans are also known to suffer from allergic diseases.

[0026] The method for treating allergic diseases according to one aspect of the present invention is a method including a step of causing a subject to inhale a composition containing hydrogen gas as an active ingredient so that the inhalation concentration of hydrogen gas falls within a predetermined range. Hereinafter, the composition containing hydrogen gas as an active ingredient will be described.

[0027] "Hydrogen gas" means gaseous hydrogen molecules. A hydrogen molecule is a molecule in which two hydrogen atoms are bonded. As isotopes of the hydrogen atom constituting the hydrogen molecule, mainly, 1 H (light hydrogen) and 2 H (deuterium; D) are known to exist stably in nature. The hydrogen atoms constituting the hydrogen molecule used in the present invention may be 1 H or 2 H (D). That is, the hydrogen molecule used in the present invention may be H 2 , HD, D 2 , or a mixture thereof. In the present specification, unless otherwise specified, the hydrogen atom represented by "H" means 1 H (light hydrogen), and the hydrogen atom represented by "D" means 2 H (deuterium).

[0028] There is no particular limitation on the source of hydrogen gas. For example, a hydrogen cylinder may be used, or a hydrogen generator (e.g., a combination of metallic aluminum and calcium oxide, or metallic aluminum and calcium hydroxide) may be used, or a hydrogen gas generator that generates hydrogen gas by utilizing electrolysis of water or the like may be used. Using a hydrogen cylinder is the simplest, but from the viewpoint of safety and the like, it is particularly preferable to use a hydrogen gas generator. That is, if a hydrogen gas generator is used, a composition containing hydrogen gas as an active ingredient can be prepared in situ during use, so there is no need to prepare, carry in, and store a hydrogen cylinder.

[0029] The composition used in the method for treating allergic diseases according to one aspect of the present invention is a gaseous composition containing hydrogen gas as an active ingredient. The composition may be a mixture containing hydrogen gas and other gases, and as the other gases, typically, air, oxygen gas, nitrogen gas, or a mixture thereof may be used. Thus, the gaseous composition containing hydrogen gas and other gases can be prepared by mixing hydrogen gas supplied from various sources with a diluting gas, such as air, oxygen gas, nitrogen gas, or a mixture thereof, at a predetermined ratio.

[0030] The method for treating allergic diseases in one aspect of the present invention is characterized by causing a subject to inhale the above composition so that the inhalation concentration of hydrogen gas falls within a predetermined range. The above predetermined range for the inhalation concentration of hydrogen gas may be, for example, 1 to 4%, more preferably 1.5 to 3.5%, still more preferably 1.5 to 3.0%, particularly preferably 1.5 to 2.5%, that is, 2.0 ± 0.5%. As shown in the experimental examples described later, according to the findings discovered by the present inventors, when the inhalation concentration of hydrogen gas is 2.0 ± 0.5%, a particularly remarkable inhibitory effect on allergic diseases can be achieved. In this specification, the numerical range indicated using "~" means a numerical range including both the upper and lower limits thereof. Also, unless otherwise specified, the concentration of a gas expressed using "%" means "% (v / v)".

[0031] As long as the inhalation concentration of hydrogen gas in the subject is within the above-specified range, there is no particular limitation on the concentration of hydrogen gas in the composition containing hydrogen gas as an active ingredient and used in the method for treating allergic diseases according to one aspect of the present invention. However, hydrogen gas is an explosive gas. For example, it is known that there is a risk of explosion when the hydrogen concentration in the air is 4.1% or more. Therefore, especially when used outside large medical institutions, for example, in small medical institutions or ordinary households, the concentration of hydrogen gas in the composition containing hydrogen gas as an active ingredient and used in the method for treating allergic diseases according to one aspect of the present invention is preferably 4% or less. Most simply, it may also be the concentration within the above-specified range described for the inhalation concentration of hydrogen gas.

[0032] There is no particular limitation on the specific method of causing the subject to inhale a composition containing hydrogen gas as an active ingredient so that the inhalation concentration of hydrogen gas in the subject is within the above-specified range. Most simply, the subject may be present in the space where the composition is supplied so that the concentration of hydrogen gas in the composition is within the above-specified range. Such a space can be, for example, a housing capable of accommodating the subject and / or a treatment room. The housing may have any shape as long as it can accommodate the subject, and can be, for example, in the shape of a cage, a capsule, a box, etc.

[0033] On the other hand, the method of causing the subject to inhale a composition containing hydrogen gas as an active ingredient is not limited to these. For example, a nasal cannula may be used, or a mask for gas inhalation may be used. When a composition containing hydrogen gas as an active ingredient is inhaled using a nasal cannula or a mask for gas inhalation, the gas supplied to the subject, that is, the concentration of hydrogen gas in the composition containing hydrogen gas as an active ingredient does not necessarily match the inhalation concentration of hydrogen gas in the subject. In this case, the supply amount of the composition containing hydrogen gas as an active ingredient and / or the concentration of hydrogen gas in the supplied composition may be appropriately adjusted according to the breathing volume of the subject, etc., so that the inhalation concentration of hydrogen gas in the subject is within the above-specified range.

[0034] In the method for treating allergic diseases according to one aspect of the present invention, there is no particular limitation on the inhalation time of hydrogen gas. For example, it can be several minutes or more, 10 minutes or more, 20 minutes or more, 30 minutes or more, 1 hour or more, 90 minutes or more, 2 hours or more, 3 hours or more, or 6 hours or more. Although there is no particular limitation on the upper limit, it is usually preferably about 24 hours.

[0035] There is also no particular limitation on the frequency of inhaling hydrogen gas. For example, it can be once a month or more, twice a month or more, once a week or more, twice a week or more, three times a week or more, or it may be inhaled once a day or multiple times a day. Also, when the symptoms of allergic diseases are strongly felt, it may be administered once each time.

[0036] In addition, in this specification, unless otherwise specified, the term "inhalation" includes both inhalation through the mouth and inhalation through the nose.

[0037] On the other hand, the present invention provides, in another aspect, an apparatus for use in the above method for treating allergic diseases in a subject.

[0038] The apparatus according to one aspect of the present invention may include means for generating a composition containing hydrogen gas as an active ingredient. The means for generating a composition containing hydrogen gas as an active ingredient may basically be of any kind, but preferably may be a hydrogen gas generator, and more preferably may be, for example, a hydrogen gas generator that generates hydrogen gas by electrolyzing tap water, mineral water, etc. (hereinafter referred to as water, etc.). A hydrogen gas generator that generates hydrogen gas by electrolyzing water, etc. has the advantage that it can generate and supply hydrogen even in places where it is difficult to install a large hydrogen gas generator such as in ordinary households or small stores.

[0039] There is no particular limitation on the configuration of a hydrogen gas generator that generates hydrogen gas by electrolysis of water or the like. For example, a hydrogen gas generator comprising an electrolytic cell that electrolyzes water or the like to generate hydrogen gas and oxygen gas, and a discharging means that discharges the generated hydrogen gas can be used. The hydrogen gas generated in the electrolytic cell may be discharged as it is, or may be discharged as a mixed gas of hydrogen gas and oxygen gas. Further, if necessary, it may be discharged as a mixed gas mixed with a diluting gas composed of air, nitrogen gas, oxygen gas, other gases, or a mixture thereof. As a specific example of the hydrogen gas generator applicable to the present invention, for example, the hydrogen gas generator disclosed in JP-A-2017-012501 is exemplified, but it is not limited thereto.

[0040] In one suitable aspect, the above-described apparatus according to one aspect of the present invention may include means for supplying the generated above-described composition to a predetermined space. Such means may be, for example, a gas supply pipe, a tube, or the like that connects the above-described means for generating a composition containing hydrogen gas as an active ingredient and the predetermined space. When the predetermined space to which the generated above-described composition is supplied has a sufficient size, the above-described apparatus according to one aspect of the present invention may be installed in the space.

[0041] As described above, the predetermined space to which the generated above-described composition is supplied may be, for example, a housing capable of accommodating an object and / or a treatment room, and the housing may be, for example, a housing having a shape such as a cage type, a capsule type, or a box type. Such a housing may be formed independently of the above-described apparatus, or may be formed as a part of the above-described apparatus.

[0042] Also, in a certain preferred aspect, the apparatus according to one aspect of the present invention may include means for maintaining the hydrogen gas concentration in the predetermined space within a predetermined range. The concentration of hydrogen gas in the space can be controlled within a predetermined range by a person skilled in the art using an appropriate method. Although there is no particular limitation on the specific method, for example, a hydrogen gas concentration meter may be installed in the space, and based on the concentration of hydrogen gas measured by the hydrogen gas concentration meter, the supply amount and / or concentration of the hydrogen gas supplied to the space may be adjusted to maintain the hydrogen gas concentration within a predetermined range.

[0043] Hereinafter, the present invention will be described in more detail with reference to experimental examples, but the scope of the present invention is not limited to these experimental examples in any way.

[0044] Experimental Example 1: Effect of hydrogen gas on allergic rhinitis model mice A hydrogen gas inhalation test was conducted using allergic rhinitis model mice. The outline of this experiment is shown in FIG. 1.

[0045] <Experimental animals> In this experiment, female ICR mice (5 weeks old at the start of the experiment) (Japan SLC Inc., Shizuoka) were used. The mice were housed in a plastic cage lined with sawdust in an animal breeding room controlled at a room temperature of 24 ± 2°C and a humidity of 55 ± 15%. During the breeding period, the lighting time was set from 8:00 to 20:00, and the mice were allowed to freely ingest feed and drinking water. This experiment was carried out based on the "Okayama University Animal Experiment Regulations" and the "Okayama University Animal Experiment Committee Guidelines".

[0046] <Reagents> In this experiment, ovalbumin (OVA) (Sigma Aldrich, MO, USA), aluminum hydroxide gel (ALUM) (LSL Co., Ltd., Tokyo), and thiopental sodium (Tanabe Mitsubishi Pharma Corporation, Osaka) were used. OVA and ALUM were dissolved or suspended in phosphate buffer solution (PBS) (Nacalai Tesque Inc., Kyoto) for use.

[0047] <Preparation of allergic rhinitis model mice> A solution prepared by suspending 10 μg of OVA as an antigen and 100 μg of ALUM as an adjuvant in 200 μL of PBS was intraperitoneally administered to female ICR mice (5 weeks old at the start of the experiment) (Japan SLC, Inc., Shizuoka) on the 0th and 5th days of sensitization for systemic sensitization. Furthermore, from 14 days after the first sensitization (day 0 of sensitization), as local sensitization, 2 μL (total 4 μL) of an OVA solution (100 mg / mL) was instilled into both nasal cavities once a day with a micropipette.

[0048] <Inhalation of hydrogen gas> The mice were placed in an observation box (MK-ICM, 25 cm × 15 cm × 20 cm, Muromachi Kikai Co., Ltd., Osaka). Then, a hydrogen gas generator (OHG-PRO) (OPS Co., Ltd., Tokyo) was connected to the observation box, and the hydrogen gas generator was started to generate hydrogen gas. While confirming that the hydrogen concentration in the observation box reached 2% with a hydrogen gas detector (XP-3310, Shin Cosmos Electric Co., Ltd., Osaka), the mice in the observation box were allowed to inhale hydrogen gas for a predetermined time.

[0049] The mice in the hydrogen gas inhalation group were allowed to inhale hydrogen gas once a day for 2 hours for 28 days from the 0th day of sensitization. On the other hand, the mice in the control group were not subjected to hydrogen gas inhalation.

[0050] <Measurement of allergic rhinitis symptoms> As allergic rhinitis symptoms, the number of sneezing reactions and nose scratching behaviors was evaluated. The number of sneezing reactions and nose scratching behaviors was measured every week from the first sensitization, that is, on the 0th, 7th, 14th, 21st, and 28th days after the first sensitization. On the days of measuring the number of sneezing reactions and nose scratching behaviors, the above-mentioned 2-hour hydrogen gas inhalation was performed before starting the measurement of the number of sneezing reactions and nose scratching behaviors.

[0051] The measurement of the number of sneezing reactions and nose scratching behaviors was performed according to the following procedure. First, before the start of the test, the mice were placed in an observation box (MK-ICM, 25 cm × 15 cm × 20 cm, Muromachi Kikai Co., Ltd., Osaka) and acclimated to the environment for 10 minutes. After acclimating to the environment, the mice were taken out of the observation box, and an OVA solution (100 mg / mL) was instilled into both nasal cavities with a micropipette at 2 μL each (total 4 μL), and then the mice were returned to the observation box. The number of sneezing reactions and nose scratching behaviors was measured by visually observing them for 20 minutes. In this test, a hydrogen gas generator was connected to the observation box, and the test was performed while confirming that the hydrogen gas concentration in the observation box was 2%.

[0052] <Statistical analysis> All experimental data were expressed as mean ± standard error of the mean (S.E.M.). For comparison among multiple groups, Dunnett's test was used, and for comparison between two groups, a t-test was used to compare the statistical significant differences with the control group. A case with a p-value less than 5% was determined to have a significant difference.

[0053] <Results> The measurement results of allergic rhinitis symptoms in mice that inhaled hydrogen gas at an inhalation concentration of 2% for 2 hours once a day from day 0 to day 28 after the start of sensitization with OVA and ALUM are shown in Figure 2. As shown in Figure 2, the number of sneezing reactions observed in the hydrogen gas inhalation group tended to be less compared to the control group, and a significant decrease was observed on day 28 of sensitization compared to the control group. On the other hand, continuous inhalation of 2% hydrogen gas did not show an inhibitory effect on nose scratching behavior. The above results indicate that inhalation of hydrogen gas at an inhalation concentration of 2% is useful for suppressing the symptoms of allergic rhinitis, particularly for suppressing sneezing reactions.

[0054] Experimental Example 2: Effect of hydrogen gas inhalation on allergic rhinitis model mice An allergic rhinitis model mouse was used to conduct a hydrogen gas inhalation test in the same manner as in Experimental Example 1, except that the number of hydrogen gas inhalations was changed from once a day for 2 hours to 3 times (6 hours each) on days 14, 21, and 28 of sensitization. The outline of this experiment is shown in Figure 3.

[0055] <Measurement of allergic rhinitis symptoms> As an evaluation of allergic rhinitis symptoms, the sneezing reaction, the number of nose scratching behaviors, the amount of nasal discharge, and the number of eosinophils in the nasal discharge were evaluated.

[0056] The sneezing reaction and the number of nose scratching behaviors were measured once a week from the first sensitization, that is, on the 0th, 7th, 14th, 21st, and 28th days from the first sensitization. On the measurement days of the sneezing reaction and the number of nose scratching behaviors on the 14th, 21st, and 28th days from the first sensitization, 6-hour hydrogen inhalation was performed before starting the measurement of the sneezing reaction and the number of nose scratching behaviors. The specific procedure for measuring the sneezing reaction and the number of nose scratching behaviors was the same as in Experimental Example 1.

[0057] On the other hand, the amount of nasal discharge was measured after intranasal administration of an OVA solution (200 μg / site) and after inhaling 2% hydrogen gas for 6 hours using the same procedure as described above. That is, 2 μL (total 4 μL) of the OVA solution (200 μg / site) was administered intranasally to both nasal cavities of the mouse with a micropipette, and 2% hydrogen gas was inhaled for 6 hours using the same procedure as described above. Then, the nasal discharge was collected by contacting the tip of Zone Quick (phenol red-based, manufactured by Ayumi Pharmaceutical Co., Ltd., Tokyo) with the left nasal cavity of the mouse for 15 seconds. Next, using calipers, the length of the portion stained red with the used Zone Quick was measured. The length of 1 mm of the stained portion of Zone Quick was calculated as 0.05 μL of nasal discharge volume.

[0058] The measurement of the number of eosinophils in nasal mucus was performed after intranasal administration of OVA solution (200 μg / site), following the same procedure as described above, after inhalation of 2% hydrogen gas for 6 hours. That is, 6 hours after intranasal administration of 2 μL (total 4 μL) of OVA solution (200 μg / site) to both nasal cavities of the mouse with a micropipette, the tip of Zone Quick (phenol red-based, manufactured by Ayumi Pharmaceutical Co., Ltd., Tokyo) was brought into contact with the left nasal cavity of the mouse for 15 seconds to collect nasal mucus. Next, using calipers, the length of the portion stained red by the used Zone Quick was measured, and 1 mm of the stained portion length of Zone Quick was calculated as 0.05 μL of nasal mucus volume. After allowing Zone Quick to air dry, the stained portion was cut out, placed in 20 μL of PBS, and left standing at 4°C for 10 minutes. Then, 20 μL of PBS containing cell sedimentation was mixed, 10 μL was collected and dropped onto a slide glass. After allowing the dropped liquid to air dry, staining was performed with eosin stain for 30 seconds, distilled water was added, and staining was further performed for 30 seconds. Then, washing was performed using distilled water and methanol, and the number of eosinophils stained red on the slide glass was calculated using a microscope.

[0059] <Results: Effect of hydrogen gas on sneezing reaction and nose scratching behavior> On days 14, 21, and 28 after the start of sensitization with OVA and ALUM, the measurement results of the number of sneezing reactions and nose scratching behaviors in mice that inhaled hydrogen gas at an inhalation concentration of 2% for a total of 30 minutes consisting of 10 minutes of environmental acclimation time and 20 minutes of allergic rhinitis symptom measurement time are shown in Figure 4. As shown in Figure 4, the number of sneezing reactions observed in the hydrogen gas inhalation group tended to be less compared to the control group, and on the 28th day of sensitization, a significant decrease was observed compared to the control group. This result indicates the inhibitory effect of a single inhalation of 2% hydrogen gas on the sneezing reaction. On the other hand, a single inhalation of 2% hydrogen gas did not show an inhibitory effect on the nose scratching behavior.

[0060] <Results: Effect of hydrogen gas on nasal mucus volume> On the 21st and 28th days from the start of sensitization with OVA and ALUM, after local sensitization, the measurement results of nasal discharge volume in mice that inhaled hydrogen gas at an inhalation concentration of 2% for 6 hours are shown in Fig. 5. As shown in Fig. 5, in both the 21st and 28th days from sensitization, inhalation of hydrogen gas did not show an inhibitory effect on nasal discharge volume.

[0061] <Results: Effect of hydrogen gas on eosinophil count> On the 21st and 28th days from the start of sensitization with OVA and ALUM, after local sensitization, the measurement results of eosinophil count in nasal discharge of mice that inhaled hydrogen gas at an inhalation concentration of 2% for 6 hours are shown in Fig. 6. As shown in Fig. 6, on the 28th day from sensitization, inhalation of hydrogen gas showed a significant inhibitory effect on the eosinophil count in nasal discharge compared with the control group.

[0062] Experimental Example 3: Hydrogen gas inhalation test using allergic conjunctivitis model mice A hydrogen gas inhalation test was conducted using allergic conjunctivitis model mice.

[0063] <Preparation of allergic conjunctivitis model mice> Female ICR strain mice (5 weeks old at the start of the experiment) (Japan SLC, Inc., Shizuoka) were intraperitoneally administered a solution in which 10 μg of OVA as an antigen and 100 μg of ALUM as an adjuvant were suspended in 200 μL of PBS on the 0th and 5th days of sensitization for systemic sensitization. Furthermore, as local sensitization, a solution of OVA (100 mg / mL) was instilled into both eyes at 3 μL each (total 6 μL) once a day with a micropipette for the period from the 14th to 28th days of sensitization.

[0064] <Hydrogen gas inhalation> On the 14th, 21st, and 28th days of sensitization, hydrogen gas was inhaled in the above-mentioned allergic conjunctivitis model mice in the same manner as in Experimental Example 1, except that hydrogen gas was inhaled at an inhalation concentration of 1%, 2%, or 3% for 6 hours. When inhaling hydrogen gas, it was confirmed by a hydrogen gas detector (XP - 3310, Shin Cosmos Electric Co., Ltd., Osaka) that the hydrogen concentration in the observation box was 1%, 2%, and 3% for 6 hours (Fig. 7).

[0065] <Measurement of the number of eosinophils in tears> As a specimen, tears were collected and the number of eosinophils in the tears was measured in the same manner as in Experimental Example 2, except that tears were used instead of nasal discharge. The measurement of the number of eosinophils in the tears was performed on the 28th day after sensitization.

[0066] <Results> Fig. 8 shows the effect of hydrogen gas on the number of eosinophils in tears in a mouse model of antigen-induced allergic conjunctivitis. Although a tendency of about 50% suppression was observed when 1% hydrogen gas was inhaled for 6 hours with respect to the number of eosinophils in the tears on the 28th day after sensitization, no significant inhibitory effect was recognized as compared with the control group not inhaling hydrogen gas. On the other hand, in the groups inhaling 2% and 3% hydrogen gas for 6 hours, a significant inhibitory effect on the number of eosinophils in the tears was confirmed. In particular, the inhibitory effect was more remarkable when 2% hydrogen gas was inhaled as compared with the case of inhaling 3% hydrogen gas. From the above results, it was shown that inhalation of hydrogen gas at an inhalation concentration of 2.0 ± 0.5 v / v% is particularly effective in the treatment of allergic diseases.

Industrial Applicability

[0067] According to the present invention, allergic diseases, preferably allergic rhinitis and / or allergic conjunctivitis, can be suppressed by an extremely simple and safe method of inhaling hydrogen gas at a predetermined inhalation concentration. Among the large number of patients suffering from allergic diseases, regardless of whether they are human or animal, and with an increasing trend year by year, the industrial applicability of the present invention that can easily and safely treat allergic diseases is great.

Claims

1. A composition for use in a method of treating an allergic disease in a subject, wherein the composition is a gaseous composition containing hydrogen gas as an active ingredient, and the method includes a step of causing the subject to inhale the composition containing hydrogen gas as an active ingredient so that the inhalation concentration of hydrogen gas falls within a predetermined range. Composition.

2. The composition according to claim 1, wherein the predetermined range is 2.0 ± 0.5 v / v%.

3. The composition according to claim 2, wherein the allergic disease is allergic rhinitis and / or allergic conjunctivitis.

4. The composition according to claim 3, wherein the subject is a subject in which the number of eosinophils is increased or suspected of being increased as compared with a healthy subject.

5. The composition according to claim 4, wherein the number of eosinophils is the number of eosinophils in nasal discharge and / or tear fluid.

6. The method includes a step of measuring the number of eosinophils in nasal discharge and / or tear fluid of the subject, and a step of determining whether the measured number of eosinophils is increased as compared with a healthy subject. The composition according to claim 4, wherein when it is determined that the measured number of eosinophils is increased as compared with a healthy subject, the subject is caused to inhale the composition containing hydrogen gas as an active ingredient.

7. The step of causing the subject to inhale the composition containing hydrogen gas as an active ingredient is performed by causing the subject to be present in a space in which the composition is supplied so that the concentration of hydrogen gas falls within the predetermined range. The composition according to any one of claims 1 to 6.

8. The composition according to any one of claims 1 to 6, wherein the composition containing hydrogen gas as an active ingredient is prepared in situ during use using a hydrogen gas generator.

9. An apparatus for use in the method of treating an allergic disease in a subject, comprising means for generating the composition according to any one of claims 1 to 6.

10. Furthermore, means for supplying the generated composition to a predetermined space, and means for maintaining the hydrogen gas concentration in the space within the predetermined range The apparatus according to claim 9, characterized by comprising.

11. The apparatus according to claim 10, wherein the subject can be accommodated in the predetermined space.

12. A method for treating allergic diseases in a subject (excluding humans), comprising: a step of causing the subject to inhale a composition containing hydrogen gas as an active ingredient such that the inhalation concentration of hydrogen gas falls within a predetermined range. **Claim 13** The method according to claim 12, wherein the predetermined range is 2.0 ± 0.5 v / v%. **Claim 14** The method according to claim 13, wherein the allergic disease is allergic rhinitis and / or allergic conjunctivitis. **Claim 15** The method according to claim 14, wherein the subject is a subject in which the number of eosinophils is increased or suspected of being increased as compared with a healthy subject. **Claim 16** The method according to claim 15, wherein the number of eosinophils is the number of eosinophils in nasal discharge and / or tear fluid. **Claim 17** a step of measuring the number of eosinophils in the nasal discharge and / or tear fluid of the subject; a step of determining whether or not the measured number of eosinophils is increased as compared with a healthy subject; The method according to claim 16, further comprising causing the subject to inhale the composition containing hydrogen gas as an active ingredient when the measured number of eosinophils is increased as compared with a healthy subject. **Claim 18** The step of causing the subject to inhale the composition containing hydrogen gas as an active ingredient is performed by causing the subject to be present in a space in which the composition is supplied such that the concentration of hydrogen gas falls within the predetermined range. The method according to any one of claims 12 to 17. **Claim 19** The method according to any one of claims 12 to 17, wherein the composition containing hydrogen gas as an active ingredient is prepared in situ during use using a hydrogen gas generator.

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