CN102657260B - Anti-allergic lactic acid bacteria - Google Patents
Anti-allergic lactic acid bacteria Download PDFInfo
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- CN102657260B CN102657260B CN2010105353158A CN201010535315A CN102657260B CN 102657260 B CN102657260 B CN 102657260B CN 2010105353158 A CN2010105353158 A CN 2010105353158A CN 201010535315 A CN201010535315 A CN 201010535315A CN 102657260 B CN102657260 B CN 102657260B
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Abstract
本发明涉及新的乳酸菌菌株，及含一种或多种所述菌株的组合物，其中所述菌株具有现有技术中未知的可用于增进抗过敏的能力，且该组合物为食品或药物组合物形式。本发明还公开了筛选测定乳酸菌菌株的方法，以及通过增进Th1型免疫反应来调控因过敏而反应过度的Th2型免疫反应的方法。The present invention relates to a novel lactic acid bacteria strain and a composition containing one or more of the strains, wherein the strain has an ability to enhance anti-allergy that is unknown in the prior art, and the composition is in the form of a food or pharmaceutical composition. The present invention also discloses a method for screening and measuring lactic acid bacteria strains, and a method for regulating Th2 type immune response that is excessively reactive due to allergies by enhancing Th1 type immune response.
Description
Technical field
The present invention relates to be applied in the antianaphylaxis lactic bacterium strains composition in food or the medicine, and the purposes in improving the antianaphylaxis ability.
Background technology
In recent years, anaphylactic disease had the trend that increases year by year, no matter be the situation that increase is arranged in the incidence of atopic dermatitis, allergic rhinitis and bronchial asthma.Think that at present the generation of anaphylactic disease may be relevant with civilization progress, comprise that the pollution of air and the minimizing of infection all are some possible causes that cause anaphylactic disease to increase.And the treatment of asthma mainly relies on pharmacological agent at present: as using steroid and suppressing medicine, trachea expanding agent and the immunotherapy etc. that mastocyte discharges the inflammation material.But, comprise the change allergic immune response that the pharmacological agent of anti-inflammatory drug and trachea expanding agent still can't be real, so the method that can only say so and take stopgap measures.And unique can saying so changes the therapeutic modality of irritated immune physique, is so-called hyposensitization, but subtracting quick treatment needs long period of time usually, and occurs some side effects sometimes, therefore can not be used for all patients.
The increase of relevant anaphylactic disease is thought at present and is followed the variation of overall situation that substantial connection is arranged, and more obvious relation is particularly arranged aspect environment.Probiotic bacterium meeting stimulating immune system effect in the human intestinal, if but the contained microbiotic of food or the content of steroid is too high in daily life, the capital causes the probiotic bacterium in the enteron aisle to reduce, thereby the generation of auxiliary type T cell 1 (Th1) in the immune stimulatory cell effectively, and the close ties that have of the generation of these auxiliary types T cell 1 and the relevant anaphylactic disease of auxiliary type T cell 2 (Th2).Therefore, if can utilize the protective foods stimulating immune system, the Th2 type immune response that excites the Th1 type immune response that can regulate in the allergic immune response to come balance allergy to cause can reach the effect of improving allergic constitution.
On the history of human using microbe, the application of milk-acid bacteria origin quite early.Find that from the processing of early stage milk-product edible food with lactobacillus-fermented helps to improve human intestines and stomach disease incidence rate and prolongs mean lifetime.Thus, cause many scholars and begin one's study the discussion milk-acid bacteria in the middle role that promotes health.Proposed in 1954 after the functional lactobacillus, constantly start the upsurge of research in the period of several, and formally propose the concept of Probiotic-probiotic bacterium in nineteen sixty-five by people such as Lilly DM, make it to become the general designation of functional lactobacillus.
The general edible product that contains milk-acid bacteria (LAB) only has the health effect of adjusting enteron aisle, though there is ten hundreds of lactic bacterium strains to be present in nature, only has the minority lactobacillus strain to have antianaphylactic speciality.The ability of the acidproof and bile tolerance ability that these bacterial strains of minority have, absorption mucodermis cell and the characteristics such as ability that still can survive after by intestines and stomach are the important evidence of screening when having the bacterial strain that promotes health effect.Even to this day, only have that several strains of lactic acid bacteria bacterial strains are verified to have an antianaphylactic health effect, and milk-acid bacteria is the specificity of bacterial strain (strain) but not bacterial classification (species) to healthy function, and this bacterial strain that human body health is had a special efficacy is called as functional probiotic bacterium (Guidelines for the evaluation of probiotics in food; Report of joint FAO/WHO working group on drafting guidelines for the evaluation of probiotics in food; " food probiotic bacterium assessment guide ", FAO/WHO joint working group is about drafting the report of food probiotic bacterium assessment guide, London Ontario, Canada, on April 30th, 2002 and May 1: 1-7).
Anaphylactic disease, for example allergic rhinitis and the allergic asthma of allergic eczema, urticaria, outbreak have repeatedly become serious social concern in Taiwan and other developed country.Cause the rising of anaphylactic disease popularity degree that reason is arranged, well known hygiology hypothesis is: the chance that the infant contacts the immunostimulation pathogenic agent is fewer, causes irritated relative disease (Strachan, 1989) more easily.When anaphylactic disease takes place, immune response meeting in the human body descends the Th1 cell quantity, produces the various kinds of cell hormone continuously and impels immune response towards the Th2 approach, forms humoral immune reaction, for example the generation of IgE and eosinophil are too much waited (Romagnani, 1994; Holt, 1995).
Find that in children's population of Estonia and these two countries of Sweden the lactobacillus in children's enteron aisle of trouble anaphylactic disease lacks (Bjo ¨ rksten etc., 1999) than the child of no anaphylactic disease.Lactobacillus is considered to promote the immune response of Th1 and then improves allergic symptom (Cross etc., 2001).Moreover, in the animal experimental model that brings out mouse allergy with Protalbinic acid, find, cheese milk-acid bacteria Shitota strain (Lactobacillus casei strain Shitota) after allowing mouse with the edible thermal treatment of oral way, the content (Matsuzaki etc., 1998) that can suppress IgE in the mice serum.In addition, prove in the animal experimental model that brings out mouse allergy with casein that the lactobacillus germ L-137 after the abdominal injection thermal treatment (Lactobacillus plantarum L-137) also can suppress the generation (Murosaki etc., 1998) of IgE.In the allergic symptom that is caused by the hogweed pollen hypersensitivity, oral enterococcus faecalis (Enterococcus faecalis) FK-23 extract, the eosinophil cell concentration that intraperitoneal is assembled reduces (Shimada etc., 2003).In the clinical trial of human body, treat to take between term sugar and lactobacillus rhamnosus bacterial strain GG (Lactobacillus rhamnosus strain GG) in the pregnant woman, discovery is 2 years (Kallioma ¨ ki etc. behind baby due, 2001) (Kallioma ¨ ki etc. and after infancy, 2003), can reduce risk and the incidence that allergic eczema takes place the infant.Lactobacillus rhamnosus 19070-2 and lactobacillus reuteri (Lactobacillus reuteri) DSM122460 also can moderately improve the children's that suffer from atopic dermatitis serious eczema allergic symptom (Rosenfeldt etc., 2003).
Summary of the invention
One aspect of the present invention broadly comprises the composition of being made up of following any biological pure growth and physiologically acceptable vehicle or thinner: Lactobacterium acidophilum (Lactobacillus acidophilus) PM-A0002 bacterial strain, be preserved in Chinese typical culture collection center, preserving number M 207,038 2007 on April 6; Jia Shi lactobacillus (Lactobacillus gasseri) PM-A0005 bacterial strain is preserved in Chinese typical culture collection center, preserving number M 207,039 2007 on April 6; Saliva lactobacillus (Lactobacillus salivarius) PM-A0006 bacterial strain is preserved in Chinese typical culture collection center, preserving number M 207,040 2007 on April 6; Yue Shi lactobacillus (Lactobacillus johnsonii) PM-A0009 bacterial strain is preserved in Chinese typical culture collection center, preserving number M 207,041 2007 on April 6; Lactobacterium acidophilum (Lactobacillus acidophilus) PM-A0013 bacterial strain is preserved in Chinese typical culture collection center, preserving number M 207,042 2007 on April 6.
In an embodiment of the present invention, said composition contains one or more described bacterial strains, preferred physiologically acceptable vehicle or thinner are a kind of food, and this food can be any in sour kefir milk, yogourt (yogurt), cheese, dairy drink, milk powder, tea drink or the coffee.Perhaps, said composition is pharmaceutical composition, and its vehicle or thinner should be pharmaceutically acceptable vehicle or thinner.
In another embodiment, adopted to have the biological pure growth that strengthens antianaphylaxis of the same race or mutant ability, physiologically acceptable following any bacterial strain: Lactobacterium acidophilum PM-A0002 bacterial strain, Jia Shi lactobacillus PM-A0005 bacterial strain, saliva lactobacillus PM-A0006 bacterial strain, Yue Shi lactobacillus PM-A0009 bacterial strain or Lactobacterium acidophilum PM-A0013 bacterial strain.
In addition, in another embodiment, but the present invention's broad sense comprises by promoting the immune response of il-1 2 (IL-12) or interferon-gamma regulation and control Th1 type to suppress Immunoglobulin IgE, improve the method for the excessive allergic phenomena of Th2 type immune response, described method comprises and gives any aforementioned biological pure growth that Mammals can reach stimulates Th 1 type immune effect dosage.
Moreover, in another embodiment, aforementioned more than one the biological pure growths in the bacterial strain that define have been adopted, preferably the form with composition gives this culture and physiologically acceptable vehicle or thinner, wherein said physiologically acceptable vehicle or thinner are preferably food, and this food is any in sour kefir milk, yogourt, cheese, dairy drink, milk powder, tea drink or the coffee preferably.Perhaps, said composition is pharmaceutical composition, and its vehicle or thinner should be pharmaceutically acceptable vehicle or thinner.
In another embodiment, provide above-mentioned arbitrary or multiple biological pure growth for the preparation of the purposes in the medicine of immune stimulatory emiocytosis antianaphylaxis relevant cell hormone.
But the present invention is comprising in the present specification separately or part, its constitutive requirements and the feature of sum total explanation of broad sense also, reach any comprise any combination any or multiple in these parts, constitutive requirements and the feature or its whole combination, and when known coordinator having occurred in the correlation technique relevant with the present invention as if the clear and definite complete things that reaches described herein, these known coordinators will be considered as independent item and be incorporated herein.
Description of drawings
Fig. 1 shows: respectively with 10
6To 10
8Individual cfu Lactobacterium acidophilum (Lactobacillus acidophilus) PM-A0002 bacterial strain, Jia Shi lactobacillus (Lactobacillus gasseri) PM-A0005 bacterial strain, saliva lactobacillus (Lactobacillus salivarius) PM-A0006 bacterial strain, Yue Shi lactobacillus (Lactobacillus johnsonii) PM-A0009 bacterial strain or Lactobacterium acidophilum (Lactobacillus acidophilus) PM-A0013 bacterial strain and 10
5To 10
7Individual human peripheral blood mononuclear cells (PBMC) co-cultivation was collected cell conditioned medium liquid after 48 hours, detected the content of interferon-gamma (IFN-gamma) in supernatant liquor with enzyme-linked immunosorbent assay.Wherein, with the negative control group of milk-acid bacteria (cheese milk-acid bacteria BCRC12249, Lactobacillus casei BCRC12249-food Industry in Taiwan Institute of Development Studies) of no antianaphylaxis function, the positive control group of PHA, the irriate concentration of detection interferon-gamma.The result shows test group stimulating human peripheral blood lymphocytes (PBMC) secretion significantly interferon-gamma, with negative control group significant difference is arranged.
Fig. 2 shows: respectively with heat-inactivated 10
6To 10
8Individual cfu Lactobacterium acidophilum (Lactobacillus acidophilus) PM-A0002 bacterial strain, Jia Shi lactobacillus (Lactobacillus gasseri) PM-A0005 bacterial strain, saliva lactobacillus (Lactobacillus salivarius) PM-A0006 bacterial strain, Yue Shi lactobacillus (Lactobacillus johnsonii) PM-A0009 bacterial strain or Lactobacterium acidophilum (Lactobacillus acidophilus) PM-A0013 bacterial strain and 10
5To 10
7Individual human dcs (dendritic cell) co-cultivation 48 hours is collected cell conditioned medium liquid, detects the content of interleukin I L-12 in supernatant liquor with enzyme-linked immunosorbent assay.Wherein, with the negative control group of milk-acid bacteria (cheese milk-acid bacteria BCRC12249-food Industry in Taiwan Institute of Development Studies) of no antianaphylaxis function, the positive control group of PHA, the irriate concentration of detection il-1 2 (IL-12).The result shows test group stimulating human dendritic cell secretion significantly il-1 2 (IL-12), with negative control group significant difference is arranged.
Fig. 3 shows: test-results shows the irritated milk-acid bacteria of commercial anti after the activation of substratum, and after handling via acidic buffer solution and cholate, the bacterium number obviously descends; And the present invention has the preservation lactobacillus strain of the irritated effect of adjusting after the substratum activation, handle with acidic buffer solution and cholate, PM-A0006, PM-A0009, PM-A0013 bacterium number are not changed by the hydrochloric acid in gastric juice choline to be influenced and sharply decline, PM-A0002 and PM-A0005 are not subjected to hydrochloric acid in gastric juice to influence (the bacterium number proves that it has tolerance to hydrochloric acid in gastric juice), and when bacterial strain handled via cholate, though the bacterium number has a little decline, but cholate is still had tolerance, prove that the antianaphylactic lactobacillus strain of this five strain can be by the test of the strict environment of digestion.
Fig. 4 shows: according to the analytical procedure of scholars such as Jacobsen (1999), when the average bacterium number in each visual field is less than 40, be judged to be " non-cohesive (nonadhesive) "; When the average bacterium number in each visual field during between 41 to 100, be judged to be " adhering to (adhesive) "; When the average bacterium number in each visual field during greater than 100, be judged to be " strong adhesion (strongly adhesive) ".The result shows that four strain antianaphylaxis bacterial strains such as Lactobacterium acidophilum (Lactobacillus acidophilus) PM-A0002 bacterial strain, Jia Shi lactobacillus (Lactobacillus gasseri) PM-A0005 bacterial strain, saliva lactobacillus (Lactobacillus salivarius) PM-A0006 bacterial strain, Yue Shi lactobacillus (Lactobacillus johnsonii) PM-A0009 bacterial strain all judge " strong adhesion ", and Lactobacterium acidophilum (Lactobacillus acidophilus) PM-A0013 bacterial strain then is judged to be " adhering to ".
Fig. 5 shows: the antianaphylaxis milk-acid bacteria is improved the design cycle of allergic experiment, and stage of tube feed antianaphylaxis milk-acid bacteria is divided into the fourth phase: the first phase is 0～22 day; The second phase is 23～36 days; The third phase is 37～50 days; The fourth phase is 51～53 days.With abdominal injection Protalbinic acid (OVA) sensitized mice, dividing four times sensitization therebetween, is the 2nd day of beginning tube feed for the first time; Be the 16th day of the beginning tube feed second time; Be beginning the 30th day of tube feed for the third time; The 4th time is the 44th day of beginning tube feed.To passing through abdominal injection Protalbinic acid (ovalbumin; OVA) mouse of sensitization is drawn blood every other week, checks the variation of IgE content in the mouse body.After continuous 2 days, detect the pulmonary respiration resistance response of mouse with nasal mucosa suction sensitized mice in putting to death mouse preceding 4 days (the 54th day), put to death the detection that mouse carries out other biochemical values every other day.
Fig. 6 shows: when replenishing saliva lactobacillus PM-A0006 lactobacillus strain for small white mouse, with Protalbinic acid (OVA) specific antibody titres part in the mouse serum of egg albumen sensitization, along with feeding dosage increases, Protalbinic acid (OVA) specific IgE antibody in Protalbinic acid (OVA) the sensitized mice serum has the trend of minimizing.
Fig. 7 shows: when replenishing saliva lactobacillus PM-A0006 lactobacillus strain for small white mouse, small white mouse pulmonary respiration resistance part with egg albumen sensitization, with respect to control group, all can under stimulating, high density methacholine (methacholine) significantly lower mouse Penh value.
Fig. 8 shows: when replenishing saliva lactobacillus PM-A0006 lactobacillus strain for small white mouse, with cell integral part in the small white mouse lung washing fluid of egg albumen sensitization, with respect to control group, significantly lower eosinophil (eosinophil) infiltration ratio is arranged.
Fig. 9 shows: with respect to control group, feeding PM-A0006 milk-acid bacteria can significantly reduce the secretory volume of chemotactic factor for eosinophils (eotaxin) in the lung flushing.
Figure 10 shows: with respect to control group, feeding PM-A0006 milk-acid bacteria can significantly reduce PGE in the lung flushing
2Secretory volume.
Figure 11 shows: spleen cell stimulates cultivation after 48 hours through ConA or Protalbinic acid (OVA), and under ConA stimulated, the interferon-gamma of the spleen cell secretion of feeding PM-A0006 milk-acid bacteria group was significantly higher than control group.
Embodiment
But the present invention's broad sense comprise following any biological pure growth: Lactobacterium acidophilum (Lactobacillus acidophilus) PM-A0002 bacterial strain is preserved in Chinese typical culture collection center, preserving number M 207,038 2007 on April 6; Jia Shi lactobacillus (Lactobacillus gasseri) PM-A0005 bacterial strain is preserved in Chinese typical culture collection center, preserving number M 207,039 2007 on April 6; Saliva lactobacillus (Lactobacillus salivarius) PM-A0006 bacterial strain is preserved in Chinese typical culture collection center, preserving number M 207,040 2007 on April 6; Yue Shi lactobacillus (Lactobacillus johnsonii) PM-A0009 bacterial strain is preserved in Chinese typical culture collection center, preserving number M 207,041 2007 on April 6; Lactobacterium acidophilum (Lactobacillus acidophilus) PM-A0013 bacterial strain is preserved in Chinese typical culture collection center, preserving number M 207,042 2007 on April 6.
Many documents point out that the cytohormone that increases auxiliary type T cell 1 comprises that the secretion of il-1 2 (IL-12), interferon-gamma (IFN-gamma) increases, and can effectively improve allergic symptom; Further Toll sample acceptor (Toil-like receptor) combination on specific lactobacillus strain and the dendritic cell is also pointed out in research, the albumen of translating in the active cells moves in the nuclear and discharges a large amount of cytohormones, a link that belongs to congenital immunity, therefore some specific lactic acid bacteria culturers is by its cell wall polysaccharides class material such as peptidoglycan (peptidoglycan), lipopolysaccharides (lipopolysaccharide), polysaccharide (polysaccharide) etc., through innate immune system, growth that really can activating T cell.
The lyophilize culture of this five strains bacterial isolates has been deposited in Chinese typical culture collection center, and the address is: Chinese Wuhan City Wuhan University.The detail file of preservation are as shown in table 1:
Table 1
| The bacterial strain name | Numbering | Date |
| Lactobacterium acidophilum PM-A0002 | M 207038 | On 04 6th, 2007 |
| Jia Shi lactobacillus PM-A0005 | M 207039 | On 04 6th, 2007 |
| Saliva lactobacillus PM-A0006 | M 207040 | On 04 6th, 2007 |
| Yue Shi lactobacillus PM-A0009 | M 207041 | On 04 6th, 2007 |
| Lactobacterium acidophilum PM-A0013 | M 207042 | On 04 6th, 2007 |
This five strains of lactic acid bacteria of having found preservation as listed above has antianaphylactic ability, comprises that the symptom for atopic dermatitis, urticaria, allergic rhinitis, food anaphylaxis and asthma has the function of slowing down and treating.
Embodiment
Morphology and the general aspects of 1: five strain antianaphylaxis of embodiment milk-acid bacteria
Confirm the feature of bacterial strain on taxonomy according to 16S rDNA sequential analysis and API Bacteria Identification systems analysis result.Find that eastern space strain number PM-A0002 is Lactobacterium acidophilum; East space strain number PM-A0005 is for adding the formula lactobacillus; East space strain number PM-A0006 is the saliva lactobacillus; East space strain number PM-A0009 is about formula lactobacillus; And eastern space strain number PM-A0013 is Lactobacterium acidophilum.The feature of this five strains bacterial strain on morphology and general aspects listed in table 2 in detail:
Table 2
Embodiment 2: the antianaphylaxis milk-acid bacteria is to the effect of regulating and control Th1 type immunological competence by the secretion that promotes interferon-gamma (be external effect verification platform with peripheral blood lymphocytes)
Detect five strain antianaphylaxis lactobacillus strains: Lactobacterium acidophilum PM-A0002 bacterial strain, Jia Shi lactobacillus PM-A0005 bacterial strain, saliva lactobacillus PM-A0006 bacterial strain, Yue Shi lactobacillus PM-A0009 bacterial strain or Lactobacterium acidophilum PM-A0013 bacterial strain are to the enhancement effect of Th1 type immunological competence, by measuring the secretory volume of interferon-gamma after human peripheral blood mononuclear cell and the antianaphylaxis milk-acid bacteria co-cultivation, screen the bacterial strain with antianaphylaxis ability.Use following experimental procedure:
1. extract an amount of human blood, at every turn about 200ml.
2. the blood cell parting liquid (Ficoll-paque) of getting equal proportion and blood under 18-20 ℃ with the centrifugal 30-40 of 400g minute.
3. (Peripheral blood mononuclear cell, PBMC) layer is behind buffered soln cleaning cell 2-3 time, with suitable substratum (for example RPMI-1640) suspension cell to get human peripheral blood mononuclear cells.
With cell with activated 3 days lactobacillus strain with 1: 10 ratio co-cultivation 48 hours.
5. collect clear liquid on the cell cultures.Utilize enzyme-linked immunosorbent assay (ELISA) to detect the content of interferon-gamma (IFN-gamma) in supernatant liquor.
Data statistic analysis such as table 3 and shown in Figure 1 are represented with mean value ± SD.Figure 1 shows that: respectively with 10
6To 10
8Individual cfu Lactobacterium acidophilum (Lactobacillus acidophilus) PM-A0002 bacterial strain, Jia Shi lactobacillus (Lactobacillus gasseri) PM-A0005 bacterial strain, saliva lactobacillus (Lactobacillus salivarius) PM-A0006 bacterial strain, Yue Shi lactobacillus (Lactobacillus johnsonii) PM-A0009 bacterial strain or Lactobacterium acidophilum (Lactobacillus acidophilus) PM-A0013 bacterial strain and 10
5To 10
7Individual human peripheral blood mononuclear cells (PBMC) co-cultivation 48 hours is collected cell conditioned medium liquid, detects the content of interferon-gamma (IFN-gamma) in supernatant liquor with enzyme-linked immunosorbent assay.Wherein, with the negative control group of milk-acid bacteria (cheese milk-acid bacteria BCRC12249) of no antianaphylaxis function, the positive control group of PHA (phyohenagglutimin) detects interferon-gamma irriate concentration.The result shows test group stimulating human peripheral blood lymphocytes (PBMC) secretion significantly interferon-gamma, with negative control group significant difference is arranged.Table 3 is for cultivating with human peripheral blood mononuclear cells with five strain antianaphylaxis bacterial strains respectively, stimulate interferon-gamma secretory volume (mean value ± SD):
Table 3
| Strain name | The secretory volume of interferon-gamma (pg/ml) |
| Lactobacterium acidophilum PM-A0002 | 19833±2767 |
| Add formula lactobacillus PM-A0005 | 46625±3624 |
| Saliva lactobacillus PM-A0006 | 25850±2347 |
| Yue Shi lactobacillus PM-A0009 | 25725±2008 |
| Lactobacterium acidophilum PM-A0013 | 17416±2803 |
| Negative control group (cheese milk-acid bacteria BCRC 12249) | 11±2.3 |
| Positive controls | 44666±2488 |
Embodiment 3: the antianaphylaxis milk-acid bacteria is to the effect of the Th1 type immunological competence of the secretion of interleukin (IL-12) regulation and control (be external effect verification platform with dendritic cell)
Detect five strain antianaphylaxis lactobacillus strains: Lactobacterium acidophilum PM-A0002 bacterial strain, Jia Shi lactobacillus PM-A0005 bacterial strain, saliva lactobacillus PM-A0006 bacterial strain, Yue Shi lactobacillus PM-A0009 bacterial strain or Lactobacterium acidophilum PM-A0013 bacterial strain are to the enhancement effect of Th1 type immunological competence, by the secretory volume of il-1 2 (IL-12) after mensuration human dcs and the antianaphylaxis milk-acid bacteria co-cultivation, screen the bacterial strain with antianaphylaxis ability.Use following experimental procedure:
1. extract an amount of human blood, at every turn about 200ml.
2. the blood cell parting liquid (Ficoll-paque) of getting equal proportion and blood under 18-20 ℃ with the centrifugal 30-40 of 400g minute.
3. get human peripheral blood mononuclear cells (PBMC) layer, behind buffered soln cleaning cell 2-3 time, with suitable substratum (for example RPMI-1640) suspension cell.
4. with CD14
+CD14 in microballoon (MiniMACS system) the purifying human peripheral blood lymphocytes (PBMC)
+Monocyte.
5. be divided into dendritic cell with cytohormone (IL-4) and tethelin GM-CSF irritation cell, behind 6-7 days the incubation time, collect the dendritic cell of having broken up.
6. with the activation in 3 days before co-cultivation of antianaphylaxis lactobacillus strain, afterwards with 100 ℃ of hot deactivation lactobacillus strains 30 minutes.
With dendritic cell and heat-inactivated lactobacillus strain with 1: 10 ratio co-cultivation 48 hours.
8. collecting cell culture supernatant.Utilize enzyme-linked immunosorbent assay (ELISA) to detect the content of interleukin (IL-12) in supernatant liquor.
Data statistic analysis such as table 4 and shown in Figure 2 are represented with mean value ± SD.Figure 2 shows that: respectively with hot deactivation 10
6To 10
8Individual cfu Lactobacterium acidophilum (Lactobacillus acidophilus) PM-A0002 bacterial strain, Jia Shi lactobacillus (Lactobacillus gasseri) PM-A0005 bacterial strain, saliva lactobacillus (Lactobacillus salivarius) PM-A0006 bacterial strain, Yue Shi lactobacillus (Lactobacillus johnsonii) PM-A0009 bacterial strain or Lactobacterium acidophilum (Lactobacillus acidophilus) PM-A0013 bacterial strain and 10
5To 10
7Individual human dcs (dentritic cell) co-cultivation 48 hours is collected cell conditioned medium liquid, detects the content of interleukin (IL-12) in supernatant liquor with enzyme-linked immunosorbent assay.Wherein, with the negative control group of no antianaphylaxis function milk-acid bacteria (cheese milk-acid bacteria BCRC 12249), the positive control group of PHA detects interleukin (IL-12) irriate concentration.The result shows test group stimulating human dendritic cell secretion significantly il-1 2 (IL-12), with negative control group significant difference is arranged.Table 4 is depicted as respectively and cultivates with dendritic cell with the hot deactivation antianaphylaxis of five strains bacterial strain, the secretory volume of stimulation interleukin (IL-12) (mean value ± SD):
Table 4
| Strain name | The secretory volume (pg/ml) of interleukin (IL-12) |
| Lactobacterium acidophilum PM-A0002 | 15019±569 |
| Add formula lactobacillus PM-A0005 | 19222±212 |
| Saliva lactobacillus PM-A0006 | 18625±365 |
| Yue Shi lactobacillus PM-A0009 | 18291±39 |
| Lactobacterium acidophilum PM-A0013 | 17836±168 |
| Negative control group (cheese milk-acid bacteria BCRC 12249) | 80±15 |
| Positive controls | 13786±341 |
Embodiment 4: the test of antianaphylaxis milk-acid bacteria stomach juice-resistant cholate
Detect five strain antianaphylaxis lactobacillus strains: Lactobacterium acidophilum PM-A0002 bacterial strain, Jia Shi lactobacillus PM-A0005 bacterial strain, saliva lactobacillus PM-A0006 bacterial strain, Yue Shi lactobacillus PM-A0009 bacterial strain or Lactobacterium acidophilum PM-A0013 bacterial strain, whether has the ability by the test of hydrochloric acid in gastric juice cholate, thereby be able to successfully bring into play its antianaphylactic function at enteron aisle, experiment process is as follows:
1. with five strain antianaphylaxis milk-acid bacterias activation 3 days.
2. get 1mL bacterium liquid and calculate original bacterium number, remaining milk-acid bacteria centrifugal 10 minutes with 500g adds the washed with de-ionized water milk-acid bacteria 2-3 time.
3. add and be deployed in the simulation hydrochloric acid in gastric juice solution of pH 2.5 with hydrochloric acid, milk-acid bacteria places 37 ℃ of incubators with after the substratum of pH 2.5 fully mixes.
4. per hour take out 1mL bacterium liquid with washed with de-ionized water 2-3 time after, the calculating survivaling cell number is till 3 hours incubation times.
5. (ox gall, Hui Rong in simulation choline solution Sigma) is after fully mixing, in 37 ℃ of cultivations to contain 1.5% (w/V) oxgall again for residue bacterium liquid centrifuged deposit thing.
6. the bacterium liquid that per hour takes out 1mL with washed with de-ionized water 2-3 time after, the calculating survivaling cell number is till 4 hours incubation times.
7. record lactobacter growth speed is calculated milk-acid bacteria to the tolerance of hydrochloric acid in gastric juice and cholate, and in the presence of hydrochloric acid in gastric juice and cholate, whether strain growth is suppressed with milk-acid bacteria in the comparative sample.
The capability result of stomach juice-resistant and analysis and arrangement Figure 3 shows that test-results shows the irritated milk-acid bacteria of commercial anti after the activation of substratum in table 5 and Fig. 3, via acidic buffer solution and cholate processing, the bacterium number obviously descends; Regulate irritated preservation lactobacillus strain of the present invention after the substratum activation, handle bacterial strain with acidic buffer solution and cholate, PM-A0006, PM-A0009, PM-A0013 bacterium number are not changed by the hydrochloric acid in gastric juice choline to be influenced and sharply decline, PM-A0002 and PM-A0005 are not subjected to hydrochloric acid in gastric juice to influence (the bacterium number proves that it has tolerance to hydrochloric acid in gastric juice), and when bacterial strain handled via cholate, though bacterial strain has a little decline, but cholate is still had tolerance, prove that antianaphylactic this five strains of lactic acid bacteria strain can be by the test of the strict environment of digestion.Table 5 is that the hydrochloric acid with pH value 2.5 mixes the result who tests its stomach juice-resistant ability with the antianaphylaxis milk-acid bacteria:
Table 5:
| Strain name | Original bacterium number | pH2.5，1hr | pH2.5，2hr | pH2.5，3hr |
| Lactobacterium acidophilum PM-A0002 | 8.20×10 8 | 6.50×10 8 | 2.87×10 8 | 2.51×10 8 |
| Add formula lactobacillus PM-A0005 | 2.65×10 9 | 2.06×10 9 | 1.19×10 9 | 7.10×10 8 |
| Saliva lactobacillus PM-A0006 | 2.55×10 9 | 9.70×10 8 | 1.42×10 9 | 1.43×10 9 |
| Yue Shi lactobacillus PM-A0009 | 6.87×10 8 | 4.05×10 8 | 4.15×10 8 | 4.35×10 8 |
| Lactobacterium acidophilum PM-A0013 | 1.78×10 9 | 1.83×10 9 | 1.86×10 9 | 1.73×10 9 |
The capability result of bile tolerance and analysis and arrangement are in table 6 and Fig. 3, and table 6 is to mix the result who tests its bile tolerance ability with the antianaphylaxis milk-acid bacteria with 1.5% bile.
Table 6
Embodiment 5: the antianaphylaxis milk-acid bacteria is to the adsorptive power test of human intestine's epidermic cell (CaCo-2)
Adopt the cell strain that has broken up to analyze the ability of five strain antianaphylaxis lactic bacterium strains absorption human intestine's epidermic cells (CaCo-2) in vitro.Earlier human intestine's epidermic cell (CaCo-2) monolayer cell is inoculated on the cover glass, grow to monolayer cell after, insert in the porous cell culture plate.Then with cell: milk-acid bacteria is with 1: 200 ratio, co-cultivation 1-3 hour, with 1 * PBS clean and with the methyl alcohol fixed cell and remaining adhere to the bacterium number after, carry out Gram dyeing and definite accompanying bacterium number.With reference to the analytical procedure of scholars such as Jacobsen (1999), 20 visuals field of counting when the average bacterium number in each visual field is less than 40, are judged to be " non-cohesive (nonadhesive) " under 1000 power microscopes; When the average bacterium number in each visual field during between 41 to 100, be judged to be " adhering to (adhesive) "; When the average bacterium number in each visual field during greater than 100, be judged to be " strong adhesion (strongly adhesive) ".
The statistical study of data is represented with mean value ± SD.On average, getting the calculated value in 45 visuals field. its result puts in order in table 7 and Fig. 4.Fig. 4 shows the analytical procedure according to scholars such as Jacobsen (1999), when the average bacterium number in each visual field is less than 40, is judged to be " non-cohesive (nonadhesive) "; When the average bacterium number in each visual field during between 41 to 100, be judged to be " adhering to (adhesive) "; When the average bacterium number in each visual field during greater than 100, be judged to be " strong adhesion (strongly adhesive) ".The result shows that four strain antianaphylaxis bacterial strains such as Lactobacterium acidophilum (Lactobacillus acidophilus) PM-A0002 bacterial strain, Jia Shi lactobacillus (Lactobacillus gasseri) PM-A0005 bacterial strain, saliva lactobacillus (Lactobacillus salivarius) PM-A0006 bacterial strain, Yue Shi lactobacillus (Lactobacillus johnsonii) PM-A0009 bacterial strain all are judged as " strong adhesion ", and Lactobacterium acidophilum (Lactobacillus acidophilus) PM-A0013 bacterial strain then is judged as " adhering to ".Table 7 is five strain antianaphylaxis milk-acid bacterias to the test-results of the adsorptive power of human intestine's epidermic cell (CaCo-2) cell strain:
Table 7
| The bacterial strain name | Mean value ± SD |
| Lactobacterium acidophilum PM-A0002 | 115.4±4.4 |
| Add formula lactobacillus PM-A0005 | 100.1±2.6 |
| Saliva lactobacillus PM-A0006 | 108.3±1.4 |
| Yue Shi lactobacillus PM-A0009 | 208.3±22.1 |
| Lactobacterium acidophilum PM-A0013 | 62.8±5.0 |
Embodiment 6: with the antianaphylaxis milk-acid bacteria as a supplement feed produced treatment or the effect of the allergic asthma of releiving
To give Protalbinic acid (ovalbumin; OVA) asthmogenic small white mouse feeding saliva lactobacillus PM-A0006 is example, carries out allergy by following experiment and improves assessment.
A. experimental design
1. laboratory animal
Use BALB/c mouse, 6-8 big female six weeks of mouse feeding in week, 14 every group of control group and experimental group.Buy the SPF level from platform large animal center, six all big female mouse of BALB/c, every mouse is raised respectively in Rotating Stainless Steel Cage, room temperature is controlled in 22 ± 2 ℃, light respectively is 12 hours (light morning six, and at 6 in afternoon is dark) dark cycling time, freely ingests, drinking-water and feed, with the body weight random packet, beginning gave tested milk-acid bacteria (2.6x10 in the tube feed mode when eight weeks of mouse were big
6To 2.6x10
7Cfu/ days) carry out sensitization simultaneously.
2. set up the respiratory inflammation zootype
The mouse sensitization step is summarized as follows: experiment beginning the 2nd day with 50 μ g Protalbinic acids (OVA) and adjuvant Alum 4mg abdominal injection to the mouse body, in experiment the 16th, 30,44 day again with 25 μ g Protalbinic acids (OVA) and 4mg Alum duplicate injection mouse.After the sensitization, experiment the 54th, 55 day, with mouse anesthesia, (intra-nasal, i.n.) mode that splashes into 100 μ g Protalbinic acids (OVA) was brought out the respiratory tract infection reaction with nasal cavity.
3. experiment flow improves the design cycle of allergic experiment for antianaphylaxis milk-acid bacteria as shown in Figure 5
The stage of tube feed antianaphylaxis milk-acid bacteria is divided into the fourth phase: the first phase is 0～22 day; The second phase is 23～36 days; The third phase is 37～50 days; The fourth phase is 51～53 days.With the mode sensitized mice of abdominal injection Protalbinic acid (OVA), dividing four times sensitization therebetween, is the 2nd day of beginning tube feed for the first time; Be the 16th day of the beginning tube feed second time; Be beginning the 30th day of tube feed for the third time; The 4th time is the 44th day of beginning tube feed.Whole mouse to abdominal injection Protalbinic acid (OVA) sensitization are drawn blood every other week, check the content of IgE in the mouse body.In putting to death mode sensitized mice that mouse preceding 4 days (the 54th day) sucks with nasal mucosa after continuous 2 days, detect the pulmonary respiration resistance response of mouse, put to death the detection that mouse carries out other biochemical values every other day.
B. experimental technique
1. the collection of the execution of animal and experiment material and analysis
A. the collection of serum sample
When experiment was carried out the 0th, 23,37,51 day and put to death, mouse is carried out contrary eye socket blood sampling (retro-orbital), collect blood sample.Behind mouse anesthesia, rapidly with micro-kapillary from eye socket venous sinus blood sampling, get 200-250 μ L blood approximately, 4 ℃ leave standstill 2-4 hour after, with the 12000rpm rotating speed centrifugal 20 minutes, collect serum, in-80 ℃ of preservations.
B. measure the enzyme immunoassay (ELISA) of Protalbinic acid (OVA) specific antibody
0.5mg antigen is dissolved in 100uL carbonate bag is cushioned liquid (carbonate coating buffer, 0.1M NaHCO
3) add microtest plate, in 4 ℃ of standing over night.The 2nd day to contain the 1X PBS buffer solution for cleaning microtest plate of 0.05%Tween 20.Carry out the sealing more than two hours with lock solution (the 1X PBS damping fluid that contains 1%BSA) then, after cleaning 3 times, then add 100 μ L samples to be tested, react on 4 ℃ of standing over night, after cleaning 4 times, add the anti-mouse IgE of 100 μ L antibody, reaction is one to two hour under room temperature, cleans 5 times again.Add avidin-HRP (the avidin-horseradish peroxidase conjugated of 100 μ L through suitably diluting, the avidin of horseradish peroxidase combination) reaction is one hour, clean 6 times, add at last 100 μ L ABTS (2.2 '-Azino-bis-3-Ethylbenzthiazoline-6-Sulfonic Acid, 2,2 '-azine-two-(3-ethyl benzo thiazole phenanthroline sulfonic acid) substrate solution, in color development at room temperature after about 30 minutes, with the 5%SDS termination reaction of 100 μ L, and read light absorption value with microplate reader (microplate reader).Measurement result is expressed as follows in the mode of enzyme immunoassay (ELISA) unit:
ELISA unit=(A
Sample-A
Blank)/(A
Positive control-A
Blank)
C. (airway hyperresponsiveness AHR) measures airway hyperreactivity
The anaphylaxis that whether can slow down sensitized mice for research oral anti-allergy milk-acid bacteria after anaphylactogen stimulates in twice tracheae, is accepted the test of resistance of respiratory tract every other day.Test macro be Buxco system (Biosystem XA, Buxco Electronics Inc.Sharon, CT, USA).Utilize sensor in the system (differential pressure pickup, differential pressure transducer, Buxco) and prime amplifier (preamplifier) (MAX II Buxco) collects the variable signal in mouse breathing road, thereby calculates the Penh value.The account form of Penh value is: intermittently * and PIF/PEF; Intermittently=(Te-Tr)/and Tr, (PIF: peak inspiratory flow (peak inspiratory flow); PEF: expiration amount peak value (peak expiratory flow); Te: expiratory duration (expiratory time); Tr: diastolic time (relaxation time)).Mouse is placed under the state of Consciousness in whole body volume scan instrument (the whole body plethysmograph) chamber, the physiological saline of ultrasound concussion gasification was imported chamber (chamber) three minutes, record the average pulmonary respiration Resistance Value of per minute Penh in the mouse three minutes then, again with the methacholine (Methacholine of cumulative concentration, 6.25,12.5,25,50mg/mL) atomization gives mouse stimulates, each concentration all after stimulating three minutes, records the physiological change in three minutes internal respiration roads.Record the mean P enh value of each minute in three minutes.
D. lung washing fluid and lung cells analysis
Test the pulmonary respiration resistance every other day, put to death mouse, cutting off neck skin and flesh exposes tracheae, put with vein and to stay pipe to insert tracheae, for the first time earlier with the aseptic HBSS damping fluid flushing of 1mL lung, second and third time be then with the HBSS that contains 2%BSA (Hanks ' balanced salt solution) damping fluid flushing lung, and the lung washing fluid that obtains first is through 1500rpm after centrifugal 5 minutes, separation of supernatant is stored in-80 ℃, treats subsequent analysis.Cell is partly incorporated in the twice lung washing fluid in back, with 1500rpm centrifugal 5 minutes, throw aside after the supernatant liquor, contain the HBSS damping fluid suspension cell of 2%BSA with 1mL, and with trypan blue (trypan blue) dyeing counting, get about 1 * 10
5Individual cell was made cell smear in centrifugal 3 minutes with cell centrifugation smearing machine (cytospin) 500rpm, after treating its natural air drying, successively with Liu A and Liu B dyeing, with a small amount of gum arabic mounting, use oily mirror (1000x) to read cell or the cell of sum more than 300 at least five visuals field, calculate its medium size lymphocyte and coenocytic ratio.
E. the cultivation of spleen cell
Under aseptic condition, open mouse peritoneal and take out spleen, place in the 20mm culture dish (petri dish) that adds 3mL HBSS damping fluid, with aseptic syringe tail end spleen is ground, its medium size lymphocyte is disengaged, cell suspending liquid is sucked in the 15mL centrifuge tube, after leaving standstill 5 minutes, get supernatant liquor with 1500rpm centrifugal 5 minutes, absorb supernatant liquor after, add 1mL RBC lysis buffer (lysis buffer), leave standstill and made globulolysis in 1 minute, add 5mL HBSS damping fluid, with 1500rpm centrifugal 5 minutes, absorb supernatant liquor; After repeating to wash twice with the HBSS damping fluid, add the RPMI-1640 perfect medium that 5mL contains 5%FBS again and make cell suspension.Calculate total cellular score with the trypan blue staining, adjust cell count to 1 * 10
7Individual cell/mL substratum.Get enchylema 0.5mL and add in 48 well culture plates (48-well plate), add the equal-volume nutrient solution again, contain the nutrient solution of fragmentation element or Protalbinic acid (OVA), making the cell ultimate density is 5 * 10
6Individual cell/mL substratum.Place 37 ℃ of constant incubators, 5%CO
2Cultivated 48 hours, and collected supernatant liquor and be stored in-80 ℃ and treat the wherein secretory volume of each cytohormone of later analysis.
F. the mensuration of cytohormone
Measure the content of the secreted cytohormone of spleen cell with sandwich ELISA method (ELISA).Be summarized as follows: earlier with the anti-cell hormone antibody with NaHCO
3Damping fluid (pH 9.6) dilution back adds 96 hole microtest plates, in 4 ℃ of standing over night.After culture plate being cleaned 3 times in second day, at room temperature react with the PBS that contains 1%BSA and to seal (blocking) at least in 1 hour.Can or place 4 ℃ of reactions to spend the night in 37 ℃ of reactions 2 hours after adding sample to be tested.Clean after 4 times anti-cell hormone antibody that the vitamin H that adds with the dilution of 1%BSA-PBS damping fluid engages (biotin-conjugated) in room temperature reaction 1 hour, clean 5 times, the peroxidase (streptavidin-conjugated peroxidase) that then adds the streptavidin joint reacted 1 hour again.After cleaning, add enzyme substrate solution tetramethyl benzidine (TMB) substrate (Tetramethylbenzidine substrate) of 100 μ L to every hole at last, reacted about 20 minutes in the room temperature lucifuge.Read the light absorption value of wavelength 450nm with microplate reader (microplate autoreader).
2. statistical method
Experimental result is represented (Means ± SEMs) with mean+/-standard error.Each group difference is analyzed with Student T check (t-test), p＜0.05 be considered as that there were significant differences (
*, p＜0.05;
*, p＜0.001).Be considered as p＜0.1 variant in addition.
Interpretation:
A. measure the result of the enzyme immunoassay (ELISA) of Protalbinic acid (OVA) specific antibody
Protalbinic acid in the serum (OVA) specific antibody titres part, the control group of feeding PM-A0006 milk-acid bacteria, Protalbinic acid (OVA) specific IgE antibody in Protalbinic acid (OVA) the sensitized mice serum has the trend of minimizing, and the result is illustrated in Fig. 6 in table 8.The measurement result of table 8 demonstration Protalbinic acid (OVA) specificity antibody IgE (mean value ± SEM, N=14, the P value=P)
Table 8
B. airway hyperreactivity (airway hyperresponsiveness, AHR) measurement result
Pulmonary respiration resistance part, with respect to control group, feeding PM-A0006 milk-acid bacteria is significantly lowered mouse Penh value under high density methacholine (Methacholine) stimulates.The results are shown in table 9, be illustrated among Fig. 7.Table 9 demonstration airway hyperreactivity measurement result (mean value ± SEM, N=14, the P value=P)
Table 9
C. lung washing fluid and lung cells analytical results
Cell integral part in the lung washing fluid, with respect to control group, the experimental group of feeding PM-A0006 milk-acid bacteria has significantly lower eosinophil (eosinophil) infiltration ratio.The results are summarized in table 10, be illustrated in Fig. 8.Table 10 demonstration lung washing fluid analytical results (mean value ± SEM, N=14, the P value=P)
Table 10
D. lung washing fluid cytohormone analytical results
With respect to control group, the experimental group of feeding PM-A0006 milk-acid bacteria can significantly reduce chemotactic factor for eosinophils's secretory volume and PGE in the lung flushing
2The trend that minimizing is also arranged.The results are summarized in table 11, be illustrated in Fig. 9 and Figure 10.Table 11 demonstration lung flushing cytohormone analytical results (mean value ± SEM, N=14, the P value=P)
Table 11
E. the supernatant liquor cytohormone analytical results cultivated of spleen cell
Spleen cell stimulates cultivation after 48 hours through ConA or Protalbinic acid (OVA), and under ConA stimulated, the IFN-γ of the spleen cell secretion of the experimental group of feeding PM-A0006 milk-acid bacteria was significantly higher than control group.The results are summarized in table 12, be illustrated in Figure 11.Table 12 shows the supernatant liquor that detects the spleen cell cultivation, and the influence that analysis different stimulated material is secreted the cytohormone of spleen cell (mean value ± SEM, N=14, the P value=P)
Table 12
Food compositions of the present invention, contain the various compositions that above-described any one or more than one bacterial strain combines, for example fermented-milk, yogourt, cheese, dairy drink milk powder, tea or coffee etc., and bacterial strain can viable bacteria or the form of dead bacterium be present in the composition.
Because milk-acid bacteria will be brought into play antianaphylactic effect, except will finding out the bacterial strain with specific function, to confirm that more can this bacterial strain also require this bacterial strain to have the favourable absorption ability to the mucous membrane of small intestine epidermic cell by the environment of human body hydrochloric acid in gastric juice cholate.Exactly because also this specific character makes antianaphylaxis milk-acid bacteria of the present invention that the medical use for the treatment of or the allergic symptom of releiving just can be provided.The anaphylaxis that antianaphylaxis milk-acid bacteria described in the invention can strengthen the Th1 approach simultaneously and regulation and control Th2 crosses Sheng.A target of the present invention continues popular to provide except steroid or antfhistamine new selection in the allergy treatment towards reaching these targets or being at least exactly, and the present invention finds out human body is had no side effect and wholesome antianaphylaxis milk-acid bacteria is used as the new selection of irritated treatment.
The above is by embodiment characteristics of the present invention to be described, its purpose is to make those skilled in the art can understand content of the present invention and implements according to this, and non-limiting claim of the present invention.Therefore, all other do not break away from disclosed spirit and the equivalence finished is modified or revise, and must be included in the claim scope of the following stated.
Culture presevation
Five strains of lactic acid bacteria bacterial strains of the present invention are preserved in Chinese typical culture collection center (CCTCC, China, Wuhan City) on April 6th, 2007, and preserving number and information see Table 13.The viability of these five kinds of cultures is detected on April 27th, 2007 by the preservation center and finishes, and the result is survival.
In addition, the lyophilize culture of five strain bacterial isolateses of the present invention once was deposited in food Industry in Taiwan Institute of Development Studies, and the address is: No. 331, food road, Hsinchu City, Taiwan Province.The detail file of Taiwan preservation are as shown in table 13.
Claims (10)
1. food compositions, it comprises:
Yue Shi lactobacillus (Lactobacillus johnsonii) PM-A0009:CCTCC NO:M207041, and any in physiologically acceptable vehicle and the thinner, it is characterized in that, but described food compositions immune stimulatory emiocytosis antianaphylaxis relevant cell hormone.
2. food compositions as claimed in claim 1 is characterized in that, described bacterial strain is viable bacteria or dead bacterium.
3. food compositions as claimed in claim 1 is characterized in that, described physiologically acceptable vehicle or thinner are a kind of food.
4. food compositions as claimed in claim 1 is characterized in that, described food is any in fermented-milk, milk powder, the tea or coffee.
5. food compositions as claimed in claim 1 is characterized in that, described food is dairy drink.
6. food compositions as claimed in claim 4 is characterized in that, described fermented-milk is cheese.
7. food compositions as claimed in claim 6 is characterized in that, described cheese is yogourt.
8. one kind is used for antianaphylactic pharmaceutical composition, it comprises Yue Shi lactobacillus (Lactobacillus johnsonii) PM-A0009:CCTCC NO:M207041, and pharmaceutically acceptable vehicle or thinner, it is characterized in that, but described pharmaceutical composition immune stimulatory emiocytosis antianaphylaxis relevant cell hormone.
9. pharmaceutical composition as claimed in claim 8 is characterized in that, described bacterial strain is viable bacteria or dead bacterium.
10. a biological pure growth is characterized in that in the purposes for the preparation of the medicine of immune stimulatory emiocytosis antianaphylaxis relevant cell hormone this biological pure growth is Yue Shi lactobacillus (Lactobacillus johnsonii) PM-A0009:CCTCC NO:M207041.
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001037865A1 (en) * | 1999-11-19 | 2001-05-31 | Probiall Pty Limited | Compositions and methods for treatment of allergic disorders |
| CN1760360A (en) * | 2004-10-15 | 2006-04-19 | 景岳生物科技股份有限公司 | Lactobacillus fermentum GM-090 and its use in stimulating INF-γ secretion and/or treating allergy |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001037865A1 (en) * | 1999-11-19 | 2001-05-31 | Probiall Pty Limited | Compositions and methods for treatment of allergic disorders |
| CN1760360A (en) * | 2004-10-15 | 2006-04-19 | 景岳生物科技股份有限公司 | Lactobacillus fermentum GM-090 and its use in stimulating INF-γ secretion and/or treating allergy |
Non-Patent Citations (2)
| Title |
|---|
| 《乳酸菌素治疗小儿消化道过敏症的探讨》;华稳珍;《温州医学院学报》;19971231;第27卷(第2期);第114页 * |
| 华稳珍.《乳酸菌素治疗小儿消化道过敏症的探讨》.《温州医学院学报》.1997,第27卷(第2期),第114页. |
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