WO2017164659A2 - Procédé d'étude de la capacité d'une substance naturelle à renforcer l'immunocompétence et procédé de fourniture d'un régime personnalisé l'utilisant - Google Patents

Procédé d'étude de la capacité d'une substance naturelle à renforcer l'immunocompétence et procédé de fourniture d'un régime personnalisé l'utilisant Download PDF

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WO2017164659A2
WO2017164659A2 PCT/KR2017/003107 KR2017003107W WO2017164659A2 WO 2017164659 A2 WO2017164659 A2 WO 2017164659A2 KR 2017003107 W KR2017003107 W KR 2017003107W WO 2017164659 A2 WO2017164659 A2 WO 2017164659A2
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Prior art keywords
cells
antigen
blood
natural material
natural
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English (en)
Korean (ko)
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WO2017164659A3 (fr
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백강민
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Ontle Union
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Ontle Union
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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/5005—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells
    • G01N33/5008—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6863—Cytokines, i.e. immune system proteins modifying a biological response such as cell growth proliferation or differentiation, e.g. TNF, CNF, GM-CSF, lymphotoxin, MIF or their receptors
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6863—Cytokines, i.e. immune system proteins modifying a biological response such as cell growth proliferation or differentiation, e.g. TNF, CNF, GM-CSF, lymphotoxin, MIF or their receptors
    • G01N33/6866—Interferon
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2500/00—Screening for compounds of potential therapeutic value
    • G01N2500/10—Screening for compounds of potential therapeutic value involving cells

Definitions

  • the present invention relates to a method for testing an individual's immune capacity by adding antigenic cells to blood of an individual and measuring the killing of antigenic cells and cytokines including IFN- ⁇ .
  • the present invention by adding antigen cells and natural material to the blood of the individual to measure the killing of antigen cells and cytokines including IFN- ⁇ to a method that can examine the extent to which the natural material affects the individual's immune capacity enhancement It is about.
  • the present invention relates to a method for providing a customized diet for enhancing an individual's immunity after identifying and selecting a natural material for enhancing an individual's immunity.
  • the immune system consists of innate and acquired immunity. Congenital immune cells have evolved to take early defense by responding to pathogens with limited receptors, and acquired immune cells are driven by T cells, and have a wider variety of memory and specific responses to specific antigens of infectious organisms. The attack system has evolved to enable immediate and effective response to reinfection of the same pathogens.
  • Innate immune responses are important for inducing early immune responses as well as for determining the direction and intensity of responses.
  • the acquired immune response itself acts to enhance the innate immune response, so that the two immune responses cooperatively form positive feedbacks to cope with infection effectively.
  • Innate immune responses and immunoregulatory cells regulate differentiation of acquired immune responses that is, they are controlled to differentiate into Th1 responses that primarily represent cytotoxic responses or Th2 responses that mainly represent humoral responses, including antibody responses, depending on the nature of the infectious agent. Induces an immune response that is optimal for the removal of infectious organisms.
  • NK cells Natural killer cells
  • cytotoxicity cytotoxicity
  • NK cells play a key role in regulating immune responses through direct interactions with dendritic cells, macrophages, T cells, and indirect interactions with cytokines.
  • This process is outlined as a mechanism by which NK cells are activated by interferons (IFNs) or macrophages-derived cytokines. That is, when exposed to IFN- ⁇ , IFN- ⁇ or IL-12, one of the cytokines produced at the beginning of infection, its activity is amplified 20 to 100 times to produce IFN- ⁇ . In other words, IFN- ⁇ and IFN- ⁇ synergize with IL-12, NK cells produce a large amount of IFN- ⁇ , and this IFN- ⁇ plays an essential role in controlling infection.
  • IFNs interferons
  • NK cells express various germ-line encoded immunoreceptors that induce or inhibit activity instead of lacking an antigen specific receptor. NK cells recognize target cells through these receptors and regulate their activity by a comprehensive signaling balance induced thereby.
  • NKT cells Natural killer T cells
  • Natual killer T cells have less than 1% of the total lymphocytes, but have been known to play an important role in various immune responses and pathologies.
  • NKT is a major cell population that performs multifaceted immunomodulatory functions in a wide range of immune diseases. Many studies of NKT cells in the past have reported that natural or artificial activation of NKT suppresses these diseases in a variety of diseases, including autoimmune diseases, transplant rejection, infectious diseases, and cancer.
  • NKT cells one of the immunoregulatory cells, are a heterogeneous group of T cells. They express NK cell markers, including NK cell receptors, on the cell surface. Unlike conventional T cell selection methods, CD 1d-glycolipid and Expresses the CD 1d-lipid complex. NKT cells are positively selected by CD4 + , CD8 + double positive (DP) thymic cells.
  • NKT cells It has different properties from other cell populations Within a short time With large amounts of IL-4 IFN secretes - ⁇ , Th1 and On Th2 reaction Central variety Cytokine Secrete. As a result, T-cell Th1 Of Th2 Plays an important role in controlling differentiation.
  • ILC innate lymphoid cells
  • ILC innate lymphoid cells
  • congenital lymphocytes do not possess antigen specificity, but they are anatomically similar to T cells and are congenital in that they play a role in T cells. 'S discovery caused a big wave in academia.
  • Congenital lymphocytes respond rapidly to innate cytokines (IL-1, IL-33, IL-25) and are responsible for maintaining tissue homeostasis, repairing damaged tissues, and protecting against pathogens. Although studies on the characteristics and functions of congenital lymphocytes are still underway, congenital lymphocytes can be classified into three groups.
  • Type 1 congenital lymphocytes have a phenotype that is quite similar to natural killer cells. Until recently, natural killer cells were thought to be primitives of congenital lymphocytes, and they were included in ILC1 in a broad sense, but in 2013, type 1 congenital lymphocytes (ILC1) were found in the colon of patients with Crohn's disease. The two are now being studied separately.
  • type 1 congenital lymphocytes Both natural killer cells and type 1 congenital lymphocytes require T-bet, a transcription factor, and secrete IFN- ⁇ in the differentiation process, but type 1 congenital lymphocytes are granzyme secreted by natural killer cells. The difference is that they do not secrete cytotoxic substances such as perforin. Instead, type 1 congenital lymphocytes can secrete large amounts of IFN- ⁇ in response to IL-7 and IL-12 .
  • NK cells natural killer cells
  • NKT cells natural killer T cells
  • ILC congenital lymphocyte cells
  • NK cells natural killer cells
  • NKT cells natural killer T cells
  • IFN- ⁇ congenital lymphocyte cells
  • Korean Patent Publication No. 10-2011-0010030 discloses an 'immunity test system for food selection'.
  • Korean Patent Publication No. 10-2004-0047899 discloses 'a novel method for measuring immunological activity'.
  • the prior art has a similar aspect to the present invention in that it measures immune cells and cytokines, the overall function of antigen cells is not measured by measuring each factor separately, and thus immunity based on innate and acquired immune mechanisms. The ability test can be said to be difficult.
  • the problem to be solved by the present invention is to provide a method for testing an individual's immune capacity by adding antigenic cells to individual blood and measuring the killing of antigenic cells and cytokines including IFN- ⁇ .
  • the problem of the present invention is to add the antigen cells and natural material to the blood of the individual to measure the killing of antigen cells and cytokines including IFN- ⁇ to examine the extent to which the natural material affects the individual's immune capacity enhancement. To provide a way.
  • the problem of the present invention is to provide a method for providing a customized diet for enhancing the individual's immunity after identifying and selecting a natural material for enhancing the individual's immunity.
  • the present invention comprises the steps of: a) collecting blood;
  • step a) culturing by adding antigen cells and natural material to the blood of step a);
  • step d) analyzing the cultured blood in step d) to determine the death of antigen cells and the concentration of cytokines including IFN- ⁇ ;
  • the present invention comprises the steps of performing the steps d) to f) on a plurality of natural products, respectively, analyzing the immune enhancing effect of each natural material; Providing a personalized diet providing method for immunological enhancement, comprising: providing a personalized diet based on the analysis result, to solve the technical problem.
  • the immunoassay method of the natural product material according to the present invention has a remarkable effect of providing an immunocompetence test method based on innate immunity and acquired immune mechanisms by examining the action on antigen cells.
  • NK cells natural killer cells
  • NKT cells natural killer T cells
  • innate lymphoid cells Innate lymphoid cells Holds.
  • the method of providing a customized diet according to the present invention has a remarkable effect of providing an optimal immune enhancing diet for each individual after examining the immune enhancement of each natural material.
  • 1 is a diagram schematically illustrating a method for testing an individual's immune capacity.
  • Figure 2 is a schematic diagram illustrating a method for testing the immune capacity enhancement of natural materials.
  • FIG. 1 is a view schematically showing a method for testing an individual's immune capacity
  • Figure 2 is a diagram schematically showing a method for testing the immune capacity enhancement of a natural material.
  • the sick includes a cancer patient or a person infected with a virus or the like.
  • Antigen cells are added and cultured to normal human blood, and the blood of the sick is cultured according to the characteristics of the antigen cells.
  • the antigen cells to be injected and the antigen cells of the sick are subject to floating cells. This is because adherent cells cannot be used because NK cells are floating cells.
  • the antigen cells to be added may be human cancer cells, preferably K562 cells.
  • K562 cells are a cell line derived from a patient with chronic myelogenous leukemia, and are originally Philaderphia chromosome positive (Ph + ), bcr-Abl positive cells. It shows pluripotent stem cell-like properties that differentiate into erythrocytes, granulocytes, and monocytes upon stimulation.In addition to its use in stem cell research, it has high susceptibility to cytotoxicity by NK cells. Can be.
  • DMEM and RPMI medium containing 10% FBS, 1% penicilin and streptomycin are used for culturing human cancer cells. Measure the number of cells at 1 ⁇ 10 8 cells / mL using a hemocytometer, and incubate in T25 cell culture flasks for 24 hours at 37 ° C., 5%, CO 2 incubator. Subcultures should be done once every two to three days. However, the culture conditions or the duration of the culture may be changed depending on the characteristics of the antigen cells.
  • the cultured blood is analyzed to determine the death of antigen cells and cytokines including IFN- ⁇ .
  • the killing assay of antigen cells can be made by measuring the cell proliferation rate using MTT assay.
  • MTT (3- [4,5-dimethyl thiazol-2-yl] -2,5-yl tetrazolium bromide) assay is a measure of the toxicity and viability of cells.
  • the yellow water-soluble substance MTT is responsible for the dehydrogenase activity in mitochondria. It uses the principle of producing purple formazan that is insoluble in water. To this end, each cell line is added 200 ⁇ L to 96well at the concentration of 1 ⁇ 10 4 cells / well and incubated in 37 ° C., 5% CO 2 incubator for 24 hours to stabilize. Prepared extracts are treated in medium at an appropriate concentration and incubated for 48 hours.
  • MTT (3- [4,5-dimethyl thiazol-2-yl] -2,5-yl tetrazolium bromide) assay reagent was added to the medium and reacted at 37 ° C., 5% CO 2 incubator for 4 hours, followed by MTT reaction solution. Remove it. In order to dissolve the generated formazan, 200 ⁇ L of DMSO was treated and the absorbance was measured at 540 nm with an ELISA spectrometer.
  • the concentration change of cytokines including IFN- ⁇ may be performed by ELISA technique.
  • binding buffer 0.1 M sodium phosphate buffer, pH 9.0
  • blocking buffer 1% BSA / PBS
  • the biotinylated anti-cytokine detection antibody was diluted in blocking buffer / Tween and treated with 100 ⁇ L / well and left at 37 ° C for 1 hour.
  • streptavidin-HRP diluted to 1/1000 was diluted in blocking buffer / Tween at 100 ⁇ g / well and left for 1 hour before washing.
  • the killing of antigen cells is analyzed and the difference between the concentration of cytokines analyzed in the blood of normal humans and the concentration of cytokines after injection of antigen cells is analyzed.
  • the values presented in the medical database may be utilized or a separate reference condition may be prepared to score immunity according to the death of antigen cells and changes in cytokine concentration.
  • step 1-2 Perform the same as in step 1-2) above, but additional natural material is added to the blood.
  • the ingredient of the natural product material may be a herbal preparation or a food preparation or a main ingredient of the food.
  • the natural product material may be an extract or concentrate extracted using hot water, ethanol, fermentation or fractions in the case of herbal preparations, and may be a sample obtained by pulverizing, powdering and dissolving foods in the case of food preparations or main ingredients of foods.
  • step 1-4 Perform the same as in step 1-4), but compare the results of step 1-4) to examine the immune enhancing ability of the natural material. In other words, if the killing of antigen cells and the secretion of cytokines including IFN- ⁇ were increased than the results analyzed in step 1-4), it can be said that the immune enhancing ability of the natural material to be tested was confirmed.
  • the optimal concentration may be analyzed by repeating steps 2-1) to 2-4) according to the concentration of the same natural material.
  • the optimal natural materials may be analyzed.
  • steps 1-1) to 1-4) are performed to measure the individual's immune capacity.
  • steps 2-1) to 2-4) are performed for various natural materials.
  • an optimal natural material for each individual can be selected.

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  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
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  • Biomedical Technology (AREA)
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  • Urology & Nephrology (AREA)
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  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
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Abstract

La présente invention concerne un procédé d'étude de l'immunocompétence de divers sujets, dans lequel des échantillons de sang prélevés chez divers sujets sont additionnés de cellules antigéniques et analysés pour déterminer le nombre de cellules antigéniques détruites et les taux de cytokines, dont ceux d'IFN-γ. La présente invention concerne un procédé d'étude de la mesure dans laquelle une substance naturelle renforce l'immunocompétence de divers sujets, les échantillons de sang prélevés chez les divers sujets étant additionnés de cellules antigéniques et de ladite substance naturelle et analysés pour déterminer le nombre de cellules antigéniques détruites et le taux de cytokines, dont celui d'IFN-γ. La présente invention concerne un procédé visant à fournir des régimes personnalisés pour renforcer l'immunocompétence de divers sujets, comprenant la détermination et la sélection de substances naturelles à des fins d'immunopotentialisation des divers sujets.
PCT/KR2017/003107 2016-03-23 2017-03-23 Procédé d'étude de la capacité d'une substance naturelle à renforcer l'immunocompétence et procédé de fourniture d'un régime personnalisé l'utilisant Ceased WO2017164659A2 (fr)

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KR1020160034495A KR20170110297A (ko) 2016-03-23 2016-03-23 천연물 소재의 면역증강 검사법 및 상기 검사법을 활용한 맞춤형 식단 제공방법
KR10-2016-0034495 2016-03-23

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20230135111A1 (en) * 2021-11-03 2023-05-04 Samuel B. Reichberg Test to Predict and Evaluate Innate Immune Responses to Infections and Methods of Treatment Thereof

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KR101981146B1 (ko) * 2018-03-26 2019-05-22 충남대학교산학협력단 수지상 세포를 이용한 면역 소재의 면역증진 효과 평가방법 및 면역 소재 스크리닝 방법
KR101981147B1 (ko) * 2018-09-27 2019-05-22 충남대학교산학협력단 수지상세포를 이용한 면역 소재의 면역증진 효과 평가방법 및 면역 소재 스크리닝 방법

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KR20040047899A (ko) * 2002-10-08 2004-06-05 오리엔트 캔서 세라피 컴퍼니 리미티드 신규한 면역활성 측정방법
AU2002952548A0 (en) * 2002-11-08 2002-11-21 Cellestis Limited Diagnostic assay
US7731937B2 (en) * 2003-09-17 2010-06-08 Incorporated Administrative Agency National Agriculture And Bio-Oriented Research Organization Diagnostic method for paratuberculosis
KR20110010030A (ko) * 2009-07-23 2011-01-31 주식회사 해림후코이단 식품 선택을 위한 면역검사시스템

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20230135111A1 (en) * 2021-11-03 2023-05-04 Samuel B. Reichberg Test to Predict and Evaluate Innate Immune Responses to Infections and Methods of Treatment Thereof
WO2023081291A1 (fr) * 2021-11-03 2023-05-11 Reichberg Samuel Test pour prédire et évaluer des réponses immunitaires innées à des infections et procédés de traitement de celles-ci

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WO2017164659A3 (fr) 2018-09-07

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