VTMC 330 Immunology
Terms
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NUTRITION
PROLIFERATION -
Immune cell proliferation is highest in body
Requires:
- Protein
- Glucose
- Fatty acids
- Fat soluble vitamins
- trace minerals -
NUTRITIONAL ROLES
SEVEN -
Intestinal Lumen
Epithelial and Mucosal Barries
- Respiratory
- Intestinal
- Skin
Development of Lymphoid Tissue
Synthesis of Active Immunologic Sustances
Cellular Activation, Proliferation and Movement
Intracellular Killing
Modulation and Regulation of Immune process -
NUTRITION
PROTEIN -
Amino Acids are building blocks for:
Acute phase proteins
Cell
- proliferation
- maturation,
- secretions
Tissue repari
Restoration of normal immune function -
NUTRITION
GLUCOSE - Energy for cellular metabolism
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NUTRITION
FATTY ACIDS -
Essential long chained polyunsaturated fatty acids;
Omega-6
- linolenic acid and byproducts gamma-linolenic acid and arachidonic acid
- proinflammatory
Omega-3
- Alpha linolenic acid and by products eicosapentaenoic acid, docosahecaenoic acid
- anti inflammatory
Optimum Balance Omega-6:Omega-3
- Dogs 6:1
- Humans 4:1
C2 - C4 Shortchain Fatty Acids
- fermentation of soluble fibre
- prefered energy source of colon mucosa -
NUTRITION
VITAMIN C - Quench freed radicals in extra and intracellular fluid
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NUTRITION
ZINC -
Decreased barrier function
Decreased circulating thymic hormone
Essential for T lymphocytes and cell mediated immunity -
NUTRITION
IRON -
Balance between:
- sequester to prevent microbe growth
- release for host metabolism and oxygen transport -
NUTRITION
SELENIM / VITAMIN E -
Antioxidants quench free radicals
Prevents lipid peroxidation of cell membranes -
NUTRITION
COPPER -
Acute phase proteins
- ceruloplasmin
Quench free radicals
- super oxide dismutase -
NUTRITION
6 NUTRIENTS FOR
EPITHELIAL AND MUCOSAL BARRIERS -
Protein
Vitamin A
Vitamin D
Zinc
Omega 3 and 6 Fatty acids
Short Chain Fatty Acids -
NUTRITION
7 NUTRIENTS FOR
NEUTROPHIL AND MACROPHAGE FUNCTION -
Iodine
Copper
Iron
Vitamin D
Vitamin C
Vitamin E
Selenium -
NUTRITION
6 NUTRIENTS FOR
ANTIBODY RESPONSE -
Protein
Arginen
Glutamine
Chromium
Iron
Vitamin B6 -
NUTRITION
2 NUTRIENTS FOR
KILLER T CELLS -
Vitamain A
Arginine -
NUTRITION
3 NUTRIENTS FOR
CELL MEDIATED IMMUNITY -
Protein
Zinc
Chromium -
MEASUREMENT OF B CELLS AND B CELL RESPONSES
2 GENERAL METHODS -
Measurements of Immunoglobiulin
- Antgen Specific Ig
- Total Ig (Class/Subclass Ig)
Measurement of B Cells - IMMUNE COMPLEX
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Interation between antigen and immuoglobin
Forms compound upon which assay is based
Assay can be used to detect either antigen or immunoglobin
Not visible tf require detection system to make visible - ANTI-SERA
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Serum containing antibodies to immunoglobulin
Ig from one species create antibodies in the serum (anti-sera) of another
Anti-sera can be used in assays to detect presence of Ig of first species -
5 DETECTION METHODS FOR
VISUALIZING IMMUNE COMPLEXS -
Percipitation
Agglutination
Immunohistochemistry
- immunofluorescence
- immunoenzyme
ELISA
- enzyme linked immunosorbent assay
Western Blot - PERCIPITAION
-
Antibody and Antigen are placed in seperated wells of clear agar gell
Antigen and antibody diffuse towards each other meeting along straight line
If antigen and antibody are specific for each other immune complexes form
Immumne complexs form a lattice which percipitates forming an opaque line in gel
Not quantitative - COGGINS TEST
- Percipitin based assay for EIA (Equine Infectious Anemia) Virus antigen specific immunoglobulin
- AGGLUTINATION
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Antigen coated particles (ie latex beads or red blood cells) are mixed with antibodies
If antigen and antibodies are specific for each other immune complexes form on the surface of the beads
Bivalency (think Y shape) of antibodies allows for cross linking between immunce complexes which results in visible clumping of particles
Quasi quantitative via titer - TITER
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Highest (last) dilution of test serum that agglutinates antigen coated particles
Titer is proportional to the levels of antibodies in the serum - ASSAY FOR TOXOPLASMA GONDII
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Agglutination based assay for T.gondii antigen specific Ig
Patient serum is incrementally diluted and placed in wells containing red blood cells with surface T.gondii antigen
Negative
- redd cells settle into bottom of well forming small dot
Positive
- immunocomplexes form and cross link froming an agglutination that fans out over bottom of well -
IMMUNOHISTOCHEMISTRY
IMMUNOFLUORESCENCE -
Mainly used to detect antigen
Immune complexes are formed on tissues on a microscope slide
Immune complexes are made visble via:
IMMUNOFLUORESCENCE
- antibody is labled with ultra-violet visible fluorescein dye
- Direct: the anitbody forming the immune complex with the target antigen is labeled
- Indirect (most commonly used): antibody forming immune complex is unlabeled. Second labled antibody from anti-sera to first specific antibody binds to constant domain of first antibody
- tf only need one type of labled anti-sera antibody for a range of antigen specific antibodies from a given species
- also will get multible binding of labled anti-sera antibody tf more visible
IMMUNOENZYME
- Antibody is labled with an enzyme ie peroxidase
- if immune complexes form enzyme will be bound to tissue
- addition of substrate for the enzyme results in coloured reaction product which is visble under light microsope -
IMMUNOHISTOCHEMISTRY
IMMUNOENZYMES -
Antibody is labled with an enzyme ie peroxidase
If immune complexes form enzyme will be bound to tissue
Addition of substrate for the enzyme results in permenant coloured reaction product which is visble under light microsope
Can be direct or indirect -
IMMUNOHISTOCHEMISTRY
ABC -
Avidin-Biotin Complex Immunoenzyme Method
Specific antibody forms immune complex with tissue
Second Anti-srea bound with biotin is added
Avidin, a large molecule with multiple binding sites for biotin, is added and triggered with peroxidase
Resuling complexes are large providing upto 1000 x MAGNIFICATION
tf used for detection of sparse antigens or when antigen has been damaged by fixatives such as formalin
Can be used for detection of antigen or antibodies -
ASSAY FOR
LEPTOSIRA - Indirect immunoperoxidase test using peroxidase labled antisera to leptospira antigen specific rabbit IgG
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ASSAY FOR
NEOSPORA -
Indirect immunoperoxidase assay for ANTIBODIES specific to Neospora
Determines if primary IgG or ongoing IgM response via:
- add serial dilutions of patient serum to TWO SETS of wells containing neospora antigen
- add antisera to IgG to one set of wells and anti-sera to IgM to another set of wells -
ASSAY FOR
INSULIN ANTIBODIES -
ABC Avidin-Biotin Complex Immunoenzyme Method
- add patient serum to normal pancrease which will contain normal insulin
- add biotin labeled anti- sera to patient antibodies
- add avidin and peroxide to from visible complexes -
ASSAY FOR
RABIES -
ABC Avidin-Biotin Complex Immunoenzyme Method
- primary antibodies to rabies virus protein
- secontary anti-sera antibody, labeled with biotin, to specific species antibody
- avidin plus peroxidase from visible complexes aka Negri bodies - ELISA
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Enyme-Linked ImmunoSorbent Assay
Antibody or Antigen detection
Immune complexes form on a SOLID SURFACE
- antigen or antibody is coated on polyvinyl chloride plate or nitrocellulose membrane
- Patient serum/tissue containing antibody or antigen of interest is added
Immune complexes are labled with an enzyme coated secondary antibody that is species specific to antibody of interest (anti-sera)or is specific to antigen of interest
Immune complexes are made visible with an enzyme substrate which produces a soluble coloured reaction product
Quantitative
- color of reaction product is proportional to antibody or antigen in serum/tissue -
ASSAY FOR
BHV-1 -
Bovine Herpes Virus Type 1
Antibody detection ELISA -
ASSAY FOR
BORRELIA BURGDORFERI -
Lyme Disease
Antibody detection Rapid ELISA -
ASSAY FOR
CANINE HEARTWORM -
Antigen CAPTURE ELISA
- form of thin layer chromatography (TLC)
- labled antibody binds directly to antigen then imobilized with second antibody
- read location of colour reaction on strip - MONOCOLONAL ANTIBODIES
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Identicle Antibodies that derive from a single HYBRIDOMA and are specific to a single epitope
Hybridomas are formed by fusing immortal non secreting plasma cells (myelomas) with mortal antibody producing (ie from inmmunized animal spleen) B cells
Note polyclonal anti bodies dervive from a hetergenous population of B-cells ie the normal situation that exists in an animal - ANTIGENIC RELATIONSHIPS
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Antigneically-related organsims share some but not all antigens
Monoclonal antibodies can distinguish amoung antigenically related organisms by detection of the unique epitopes - COMPETITIVE ELISA
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Detection and measurement of antibodies binding a single epitope
- antigen of interest is bound to wall of well
- both monoclonal antibody to specific epitope of and patient serum are placed in well
- wash and add enzyme conjugated secondary antibody specific to MONOCLONAL antibody
- wash and add substrate
- if patient has ployclonal antibody specific to epitope then it will out COMPETE the monoclonal antibody and there will be no enzyme for substrate to react with
- tf NEGATIVE test means patient HAS polyclonal antibody for epitope
- POSITIVE test means that patient has no polyclonal antibody to specific epitope so the enyzme labled monolonal antibody can bind and there will be a colour reaction - WESTERN BLOT
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Quasi percipitation based assay
Separates antigens by size and charge via electrophoresis
Antigens are blotted and then washed with enzyme labeled antibody
Useful to identify which antigens or which species/strain of microrganism (based on size and charge characterstics) is generating immune response -
ASSAY FOR
STAPH BACTERIA - Western blot to determine strains based on characteistic proteins
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MEASUREMENT OF ANTIGEN SPECIFIC IMMUNOGLOBULINS
4 REQUIREMENTS -
Pateint serum and source of target antigen
Method to allow the formation of immune complexes
Detection system for making the interaction visible
- percipitation
- agglutination
- immunohistochemical
- ELISA
Means of quantifying the amounts of anitbody detected
- titer
- spectrometry -
MEASUREMENT OF TOTAL IMMUNOGLOBULIN LEVELS IN SERUM
4 REQUIREMENTS -
Antisera to immunoglobulin
- generate by injecting purified Ig into a heterologous species
Method to allow the formation of immune complexes
Detection system for making the interaction visible
- percipitation
- agglutination
- immunohistochemical
- ELISA
Means of quantifying the amounts of anitbody detected
- Immunoelectrophoresis
- Radial Immunodiffusion - IMMUNOELECTROPHRESIS
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Perciptiation based assay
Serum of patien applied to wells in an agar gel
Immunoglobulins diffuse into gel and separate based on size and charge via electrophoresis
various antisera applied at seperate locations along the gel and allowed to diffuse into the gel where it binds to the Ig forming an immune complex which percipitates forming arcs around the anti-sera spots
Location and size of arcs provides semi qualitative (size of arc) and qualitative estimate of differnet classes of immunoflobulin present in patients serum -
ASSAY FOR
PLASMA TUMORS -
aka monoclonal gammopathies
- B cell tumor
Immunoelectrophoresis
- shows extremely thick and skewed arc - RADIAL IMMUNODIFFUSION
-
Gold Standard
Antisera to immunoglobulin incorporated into agar gel
Serum samples from patient and standards with known Ig levels placed in wells
Ig in serum diffuses into gel forming immune complexes which percipitate froming rings around wells
Diameter of rings is proportional to serum Ig levels
- tf determine patient levels via standard curve
Requires 24 hrs for diffusion -
MEASUREMENT OF B/T CELL NUMBERS AND SUBSET NUMBERS
Fluorescent Activated Cell Sorter (FACS) or Flow Cytometry -
Fluorescent Activated Cell Sorter (FACS) or Flow Cytometry
Expose suspension of cells to primary monoclonal antibodies to specific CD (cluster of differentiation) cell surface molecule specific to the cell type/subtype of interest ie CD4 or CD8
Expose suspension of cells to differently fluoro labeled secondary antibodies
Count cell numbers via flow xytometry of fluorescent activated cell sorter (FACS)
Ratio of CD4:CD8 provides indication of humoral or cellular immunity -
MEASUREMENT OF B/T CELL NUMBERS AND SUBSET NUMBERS
IMMUNOHISTOCHEMICAL STAINING -
Stain for CD (cluster of differentiation) molecules or other cell surface markers
ie show distribution of B and T cells in tissues such as lymph nodes or skin -
T LYMPHOCYTE RESPONSE
3 MEASUREABLE CHARACTERISTICS -
Proliferation Responses
Cytokine Secretion Responses
Cytotoxicity to target cells
- CD8 - PROLIFERATION RESPONSES
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aka Clonal Expansion
Measurement of Blastogenesis which is a hall mark of T cell function
Mitogen Response
- mitogens are generally lectins that bind to CHO residues on cell surface glycoproteins stimulating division
- expose extracted patient lympocytes to mitogens and radioactive thymidine
- incorporation of H3thymidine in specific time period is proportional to proliferation - CYTOKINE PRODUCTION RESPONSES
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BIO-ASSAY
- expose extracted patient lympocytes to mitogens or antigens that will stimulate proliferation and cytokine release
- extract supernatent and add, along with H3tymidine to cells that are targeted by specific cytokine to proliferate
ELISA
- antigen capture method using surface impregneated with antibody specific for the cytokine and an enzyme conjugated antibody that is ALSO specific for the cytokine - MEASUREMENT OF T CELL CYTOXICITY RESPONSES
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CYTOTOXIC ASSAY
- radio labled target cells release radio activity when killed
- antigen specific
- MHC I restricted
IN VIVO
- expose antigen to skin
- measure (subjective) Delayed Type Hypersensitivity (DHT) response
- provides measure of T-Cell memory -
NEONATAL IMMUNITY
3 BASIC FACTOIDS -
Domestic animals become immunocompetent prior to birth
Partuition is associated with immunosuppressive levels of stress
- high levels of corticosteroids
Colostral immunoglobulin is critical for the survival of newborns
- except primates - DEVELOMENT OF IMMUNE RESPONSE IN FETUS
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Primary lymphoid organs develop in the first trimester
Secondary lymphoid organs follow shortly thereafter
Lymphoid organs and phagocytic capability functional by mid gestation
Immune responses including B and T Cells to foreign invadors are detected soon after the development of the lymphoid organs
Animals can respond to vaccine antigens in utero
ie chick embryos vaccinated for Mareks Disease (herpes virus) at 18 days - FETAL IMMUNE RESPONSES TO INTRAUTERINE INFECTIONS
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Consequence depends on the pathogen and stage of development of the fetus
ie BVD Virus
- early abortion
- early to mid tolerance and persistent infection because perceives virus as self
- mid to late malformations
- late normal
ie BHV-1 can result in death at any time because virus is highly lytic
Most pathogens do not cross placenta
Overall fetus is less capable than adult at mounting effective response
Fetal immune responses are PRIMARY (mainly IgM)
Elevated fetal serum Igs and pathogen specific IgM can be used to diagnose intrauterine infections -
EFFECT OF TIME OF GESTATION
BY THE TRIMESTER -
First
- more likely to result in death because no effective immune response
Second
- result in variable damage to fetus but fetus likely to remain viable
Third
- infections can provoke effective immune responses tf fetus often undamaged - NEONATAL IMMUNOSUPPRESION
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Parturition is triggered by fetal stress (high steroid levels)
Steroid levels are immunosuppresive resulting in:
- decreased phagocytosis
- decreased T Cell blastogensis and cytokine secretion
For the first 3 - 5 days of life:
- susceptible to infections which would not cause serious problems in older animals
- disease can be caused by organsisms and MODIFIED LIVE VIRUS VACCINES which are safe in older animals
Dam is also immunosuppressed -
TRANSFER OF MATERNAL IMMUNITY
IN UTERO -
Route determined by the nature of placenta
- hemochorial ie primates allows full transfer of IgG but not other Ig classes
- Endothelialchorial ie dogs and cats allows transfer of only 5 - 10% of IgG
- Syndesmochorial ie ruminants no transfer of maternal Ig prior to birth
- Epitheliochorial ie pigs and horses no transfer of maternal Ig prior to birth - COLOSTRUM
-
first milk/secretion produced by mammary gland after parturition
Vehicle for transfer of maternal Ig to neonate
- Mainly IgG
- also IgM, IgA, IgE
Receptors for Ig in mammary gland upregulated results in transfer from plasma to milk
- tf can vaccinate dam to protect neonate
- also high levels of lactoferring
- also high levels of gowth factors for development of GI and other organs
Thick yellow and viscous ie 50 - 100 g IgG/l vs 1 - 2 g/l normally
Normal milk Igs are locally produced in mammary gland not sourced from serum
- ruminants IgG is major milk Ig
- non ruminants IgA is major milk Ig - TRANSFER OF COLOSTRAL Ig TO NEONATE
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Igs reach small intestine intact and functional
- low proteolytic activity in new born
- high typsin injibitor levels in colostrum reduces degradation of Igs
Intestinal epithelium of neonate allows absorption of intact Igs DIRECTLY to blood stream
- binding of Fx receptors on epi cells results in uptake by pinocytosis
- Ig is passed into intestinal lacteals and capillaries then RESECRECTED back to mucosal surfaces
- transfer phenomina only lasts for a few hours - PROTECTION OF COLOSTRAL Ig
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Serum Ig protects from viremia and bacteremia
Re-excretion of Igs on mucosal surface blocks pathogen invasion MOST IMPORTANT
Half life of IgG is 3 weeks. Neonate antibody production begins at 5 weeks
- tf period of vunerabilitity is 5 - 12 weeks - FAILURE OF PASSIVE TRANSFER
-
FPT newborns are those with indadequete levels of colostral Ig to protect from disease
Adequate passive transfer results in 10g/l of IgG in serum at 24 hrs
Immunoglobulin production varies widely amongst dams -
FAILURE OF PASSIVE TRANSFER
3 MECHANISMS -
Too little or to late
Indaequate colostrum production because young or premature (leaking) lactation
Inadequate colostrum ingestion via poor mothering or suckling
Inadequate If absorbtion via low Ig [] or acidosis because of difficult birth
- requires 100 - 120 g IgG to be injested by calf by 6 hrs
- calf usually only injests 2 l tf concentration must be high enough -
FAILURE OF PASSIVE TRANSFER
DIAGNOSIS -
Colostral Ig peaks in serum of newborns at 24 - 36 hours after suckling
Measure via
- Radial Immunodiffusion
- antigen capture ELISA cows and horses -
FAILURE OF PASSIVE TRANSFER
CONSEQUENCES -
Less than half normal levels results in high morbidity or mortaliity due to bacterial septicemia
More than half normal levels results in variable morbidity/mortality most often due to diarrheas
Amount of disease and losses highly dependent on management
- reduce challenge level
- suppportive care
- serum tranfusion if over 12 hours - IMMUNE RESPONSES IN YOUNG
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Primary (IgM) responses can occur in utero
Neonates are unresponsive for 3 - 5days
Colostral Ig interfers with development of active immunity:
- sequestration of antigen
- negative feedback on specific B cells. Binding of antibody to FcReceptor blocks BCR activity
- tf time to effective vaccination of the young is inversely correleated to the titer of antibody found in the mother - ANTIGENS AND PATHOGENS TRANSFERED WITH COLOSTRUM
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Transfer of antibodies to blood group associated antigens
- neonatal iserthrolysis and thrombocytopenia
Transfer of cell associated infectious disease agents in macropages and neutrophils which directly enter the blood stream through "open gut"
- Johnes Disease - PASSIVE IMMUNITY IN BIRDS
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Naternal serum IgG is concentrated in the egg in the ovary and enter the bloodstream of the chick
IgM and IgA enter the abumin in the oviduct and are swallowed by the chick - INTENT OF NEONATAL IMMUNITY
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Fetus and newborn are immuo-compentent but immunologically naive
Colostral antibodies designed to provide initial high levels of passive protection from pathogens at time of birth
Gradual decline in the passive protection at the same time as exposure to low levels of pathogens
Young gradually replace the passive maternal protection with active responses
ie NATURAL CONDITIONS ARE GRADUAL EXPOSURE TO PATHOGENS otherwise system would not work this way - TOLERANCE
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Central Tolerance (Thymus)
- deletion of T cells that have nonfunctional TCRS ie no ability to bind self MHC or too strong binding
- All individuals have self reactive T cells which escape deletion in thymus
Peripheral Tolerance - other regulatory mechanisms that prevent self-reactive lymphocytes from damaging self
- anergy lack of co-stimulatory signals
- Suppression
- Immunological Ignorance ie self antigens are hidden (CNS) or cryptic - PHYSIOLOGICAL AUTOIMMUNITY
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Normal mechanism for removal of aged cells
- new immunogenic epitopes are formed as cells age ie band 3 on degenerating surface proteins of red blood cells and others - AUTOIMMUNE DISEASES
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Pathological conditions associated with ongoing tissue damage from self reactive antibodies (T Cell help) and or cytotoxic T Cells
Develop spontaneously
Idiopathic
Partial or complete failure of tolerance
Result of either and immune response directed to single self antigen or general defect such that may self antigens are recognized
Results from either:
- normal immune response to an abnormal self-antigen
- abnormal immune response to a normal self-antigen (worst situation)
Autoimmune conditions are failures of both tolerance and immunoregulation
- tolerance to self is broken
- normal regulatory mechanisms do not function to pervent or down regulate response to self antigens
Can be specific to one cell type or general to many cell types - 4 CAUSES OF LOSS OF TOLERANCE
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Exposure of HIDDEN antigens
New epitopes or antigens on self proteins
Cross Reactive antigens
Changes in antigen processing or presentation - EXPOSURE OF HIDDEN ANTIGEN
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antigens not normally exposed to cells of the immune system
- CNS, Testes, Cytoplasm
- cryptic epitopes normally hidden by conformation of antigen
These antigens are normally protected from exposure to lymphocytes tf no active tolerance
Inflammation can cause exposure of antigens leading to triggering of immune response and AID Auto immune disease
- antibodies t cardiac mitochondria after heart attack
- antibodies to sperm after inflammation
- antibodies to liver membrane proteins - NEW EPITOPES OR ANTIGENS
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Generated on self proteins by molecular changes
- rhumatoid factor: auto-antibodies to the Fc portion of IgG in immune complexes via conformation change when bound
- Immunoconglutinins: auto-antibodies to complement components created dureing complement activation esp C3 - CROSS REACTIVE ANTIGENS
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Molecular Mimicry
Immune response to an infectious agent which shares epitopes to self antigens
- T helper cells become activated to epitope - NAME 5 DISEASES WHICH CROSS REACT WITH SELF ANTIGEN
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Gp A Streptocacci (rheumatic fever
- AID of heart, kidney, joints
Mycoplasma cattle and pigs
- antibodies to normal lung antigens
Trypansomes
- mammalian cardia muscle and neureons
Epstein Barr Virus (MS)
- myelin basic protein
Leptospira interrogans Moon Blindness horses
- immune responses to cornea / uvea - CHANGES IN ANTIGEN PROCESSING OR PRESENTATION
-
Once immune response is triggered the antigen presenting cells in the area becaume activated and have enhanced interactions with T helpers
Enhanced antigenprocessing results in better ability to present self antigens
High levels of cytokines in the environment results in supplying the co-stimulatory signals needed to trigger self reactive cells
Both bacteria and viruses have been implicated in inducing auto immunity this way -
AID
2 BIG THINGS -
Auto Immune Disease
Failure of Tolerance and Immunoregulation
- Autoreactive lymphocyte loses tolerance and starts to react to self tissues
Regulatory Control Fails
- mechanisms not understood
- low level auto immune responses are normal
- usually response is regulated
- disease develops when ther is a failure of regulatory control of self antigens -
PREDISPOSITION TO LOSS OF TOLERNANCE AND IMMUNOREGULATION
6 FACTORS -
Genetics
- AIDs are familial
Age
- middle age or older
Gender
- females more affected
Infections
- bacteria, viruses
Vaccination
- adjuvanted vaccines
Altered Immune System
- lymphoid neoplasia
- immunosupressive therapy
- immunodeficiency states -
AUTO IMMUNITY
GENETICS -
MHC II genes are associated with development of AIDs
MHC II fundamentally controls immune response
- regulate the ability to respond to antigen
- onset of AIDs after reproductive age has resulted in lack of selection pressure against MHC antigen binding grooves that present self antigen
Most diseases are associated with particular combinations of MHC genes not a single MHC gene
AIDs tend to be familial in all species
Inbreeding can result in high levels of AIDs -
AUTOIMMUNITY
GENDER -
Hormones
- estrogen increases AID prevlaence if females 10x
Micro-Chimerism
- exchange of fetal and maternal cells during pregnancy
- cells survive for years
- fetus lymphocytes view dam as foreign
- dam lymphocytes view fetus cells as foreign -
AUTOIMMUNITY
INFECTIOUS AGENTS -
Cross Reactivity
Bystander Activation
- upregulation of immune response in area of infection activates local self reactive cells -
AUTOIMMUNITY
VACCINATION -
Administration of potent adjuvanted vaccine may trigger transient production of a variety of auto-antibodies
- ie via macrophage activation
Rare Events -
AUTOIMMUNITY
ALTERED IMMUNE SYSTEM AND LOSS OF IMMUNOREGULATION -
Neoplasia of lymphoid tissues associated with AIDs
- Thymoma: autoantibodies to Ach receptors block Ach binding
Immuno-deficent individuals have increased AIDs
- IgA defecient dogs have weakened mucosal barries which allows increased numbers of micro invadors to enter resulting in increased stimulation - DETERMINANT SPREADING
-
The increase with time in numbers of epitopes recognized in an autoimmune response
Response starts out confined to a one or oa fuw immunodominant peptide epitopes
With time immune response spreads to recognize othe epitopes on the same protein and then on other proteins
ie antibodies to heterlogous species insulin is induced by the foreign epitopes but rapidly recognizes all sites on host species insulin and foreign insulin -
VACCINATION
2 CRITEREA
2 METHODS -
Will immune response from vaccination actually protect against disease
Do benefits of vaccination outweigh risks and does risk of contacting disease outweigh risk of vaccination
PASSIVE
- temporary resitance achieved by transferring antibodies from a resistannt to a susceptable individual
ACTIVE
- adminsistering antigen to an animal so that it responds by producing a protective immune response - 5 ADVANTAGES OF LIVE VACCINES
-
Few inoculateing doses required
- risk that non pathogenic altered agent could revert or be contaminated with other virus
Adjuvants unnecessary
Less chance of hypersensitivity
- because of low amount of antigen used
Induction of interferon
- better cell mediated response because of accurate epitopes
Relatively cheap
- low amounts of antigen required because animal will amplify response - 3 ADVANTAGES OF INACTIVATED VACCINES
-
Stable during storage
Unlikely to cause disease through residual virulence
Unlikely to contain contaminating organisms - 3 WAYS OF GENETICALLY ENGINEERING ANTIGENS
-
Inactivated recombinant organisms or purified antigens derived from recombinant organisms
- ie isolate AG gene from virus and insert into bacterial plasmid aka Subunit vaccine
- antibody response only
- can be used for disease eradication by vaccinating for specific antigen subunit and then culling any animal with other subunits of pathogen
Live organisms that have virlulence genes deleted or contain heterologous marker genes
Vaccines that contain live expression vectors expressing heterlogous genes for immunizing antigens or other stimulantes
- canary pox, modified to carry rabies surface glycoportein gene, as vector. Has limited replication in mammals
- risk that when booster vaccination occurs body has developed response to canary pox vector
- advantage is cell mediated response - PARENTERALLY
-
Out of GI tract
ie
- Sub Q
- IM
- Intranasally
- orally - DEPOT ADJUVANTS
-
Aluminum Salts
- aluminum phospate
- aluminum hydroxide
Emulsify antigen
- sequester antigen (depot effect) slowing release which prolongs immune response
Inert and safe but not highly effective
- only adjuvant used in human vaccines in NA - IMMUNOSTIMULATORY ADJUVANTS
-
Anaerobic corynebacteria, BCG, LPS
- All MACROPHAGE STIMULATORS
- not neccesarily inert
- LPS stimulates specific response but also IL-1 proinflamatory cytokines
Saponin
- STIMULATES ANIGEN PROCESSING
- plant material used in veterinary vaccination
- fools system into processing it ENDOGENOUSLY
- tf cellular response but safe because not live agent - PARTICULATE ADJUVANTS
-
Liposomes
- STIMULATES ANTIGEN PROCESSING
- vesicles resemble cell membranes
- induce both antibody and cellular response (via endogenous processing)
Micro Particles
- DEPOT effect and ANTIGEN PROCESSING
- tiny solid particles degrade slowly
- antibody and cellular response - MIXED ADJUVANTS
-
Freund's Complete Adjuvant
- water in oil emulsion plus mycobacterium protein
- depot effect -
ANTIVIRAL VACCINES
5 TYPES -
Mixed parenteral vaccines
- modified live
- inactivated
Subunit Vaccines
Genetically Engeineered vaccines
Vector Vaccines
Intranasal Vaccines -
ANTI BACTERIAL VACCINES
3 TYPES -
TOXIODS
- inactivated (formaldehyde)
- given with adjuvant
- ie Tetanus, blackleg
BACTERINS
- killed bacteria (formaldehye)
- adjuvant
- Ananculture bacteria 9esp membranes + toxoid
- Autogenous Bacterins made from bacteria isolated from farm to get specific strain
- CORE antige J5 from LPS
LIVING BACTERIAL VACCINES
- Attenuated virus ie brucella 19, capsulless anthrax, raough salmonell dublin
- requires lower dose
- Th1 response - PROTOZOA VACCINES
-
Very Few Successful
Theileria Parva
- infection with low dose of sprozoites and treatement with tetracycline
- must get cellular response because part of life cycle is intracellular
Toxoplasma gondi
- live vaccine - HELMITH VACCINES
-
Very Few Successful
- large size
- diversity of life style
Dictyocaulus viviparus
- irradiated larva - EXPERIMENTAL CHALLENGE STUDIES
-
Naive vs Seronegative (no antibodies but has memory cells)
Randomization
Blinding
Relevance of model(ie IV) to natural disease (aereosolised)
Preventable Fraction
- PF = (%controls dying - %vaccinates dying)/%controls dying - FIELD TRIALS
-
Pen effect vs herd immunity effect or dilution with vaccinates
Randomization
Blinding
Outcomes
Murphy's law in reverse - IMMUNOLOGICAL STUDIES
-
Agents used in assays
difficuly in comparing serological results amoun laboratories
Tests of cell mediated immunity
Corelative challenge data
Human medicine can work inductively via serological studies because high $ and #
Veterinary medicine should work deductively via challenge studies and history - VACCINE FAILURE MODES
-
CORRECT ADMINISTRATION
Animal Responds
- vaccine too late already infected
- wrong strain or organism
- non portective antigens
Animal Fails to Respond
- prior passive immunixation
- immunosuppressed
- biological variation
- inadequate vaccine
INCORRECT ADMINISTRATION
- inappropriate route of administration
- death of live vaccine (dashboard)
- administration to passively protected animal - ADVERSE REACTIONS TO VACCINE
-
NORMAL TOXICITY
- Fever, Inflammation, Malaise, Pain
ERRORS
- manufacture/administrion
- Bacterial or Viral contamination
- abnormal toxicity
- residual virulence
INAPPROPRIATE RESPONSES
- Hypersensitivites
- Neuologic reactions
- Foreign Body reactions
- biological variance - IMMUNOTOXICOLOGY
-
Study of immunododulatory effects of chmicals on the immune system
Aimed at detection subclinical problems before they become clinical
Enviromentally realistic exposure ie normal paths of exposure are not IV
Requires a wide range of tests because immune system is compelex and highly redundant - IMMUNOTOXILOGICAL ENDPOINTS
-
Act as BIOMARKERS of
- exposure
- effects of exposure
Are used to evaluate risks to wildlife
Animal serv as sentineld for human risk from chemical exposure - XENOBIOTICS
-
Foreign compound
- effects on biological system
- if in body must be detoxified and excreted
- may cause immunosuppression or immunostimulation
- may cause disregulation or immunododulation - IMMUNOTOXICITY TESTS
-
TIER I
Post-Mortem
- immunopathology
- organ weights, spleen, thymus, bursa
TIER II
In Vivo
- PHA (mitogen) skin test
- Antibody response (DNP-KLH (keyhole lympet antigen), foreign RBCs)
- DTH (delayed type hypersensitivity) GOLD STANDARD
In Vitro
- wbc countand differential
- plasma proteins
- phagocytosis assay (macrophages, splenocytes)
- respiratory burst (macrophages, PMNs/heterophils)
- NK assay - TYPE I HYPERSENSITIVITY
-
Immediate Hypersensitivity
Acute inflammatory reactions
Mediated by IgE antibody responses
Effector cells are mast cells, basophils and eosinophils
ANAPHYLAXIS
- systemic and severe
- immediately life threatening
ALLERGY
- localized
- usually not immediately life threatening -
TYPE I HYPERSENSITIVITY
MECHANISM -
Antigen binds (crosslinks) IgE antibody bound to FcRI on mast cells, basophils triggers cell degranulation
Pathogenesi of tissue damage is due to the effects of released granules
- vasodilation
- puritis (itching)
- bronchoconstriction -
TYPE I HYPERSENSITIVITY
IgE -
200KdA Dimer ala IgG
Produced primarily at body surfaces (skin, gi, lung)
- Th2 Cytokines (IL-4 esp, 5, 10, 13) elicit B cell Class Switch to IgE
- Th1 Cytokines (gamma IFN, IL12)inhibit IL4 production
Half life of 11 - 12 days
Induced by parasites
Normally produced in low levels
Allergies result from IgE production to an antigen tf aka ALLERGEN - ATOPY
- Condition in which IgE produced at abnormally high levels to many antigens
- IgE RECEPTORS
-
FcRI
- receptor with very high affinity binding for IgE
- mainly found on mast cells and basophils and some on eosinophils
- also found ondendritic cells and monocytes of ATOPIC patients (ie differential)
FcRII
- receptor with much lower affinity fo rIgE
- found on NK cells, macrophages, dentritic cells, eosinophils, platelets and some B cells
ALLERY LOOP
- dendritic cells bind antigen via FcRI and IgE
- processing stimulates cytokine release which stimulates Th2 responses (IL4, 5, 10, 13)
- stimulation of B Cells produces more IgE
- more IgE binding and crosslinking on Mast cells causes them to release Th2 activating cytokines (IL-4, 5, 13) which further amplifies IgE production
- IgE bound and cross linked mast cells release allergic mediators - ALLERY LOOP
-
- dendritic cells bind antigen via FcRI and IgE
- processing stimulates cytokine release which stimulates Th2 responses (IL4, 5, 10, 13)
- stimulation of B Cells produces more IgE
- more IgE binding and crosslinking on Mast cells causes them to release Th2 activating cytokines (IL-4, 5, 13) which further amplifies IgE production
- IgE bound and cross linked mast cells release allergic mediators -
MAST CELL RESPONSES
ANTIGEN CROSSLINKED TO SPECIFIC IgE -
Cytokine Synthesis and Secretion
- positive feedback
Granule Exocytosis
- rapid within 60 seconds
Prostaglandin and Leukotriene synthesis
- pro inflammatory - SOLUBLE MEDIATIORS RELEASED BY MAST CELLS
-
SECONDS TO MINUTES Gronule Exocytosis
Histamine
- smooth muscle contraction
- broncho constriction
- exocrine secretions mucus, tears, saliva
Serotonin
- vasodilation, edema
Proteases
- trypsin, chymotrypsin
Kalikreins, Proteoglycans
MINUTES Eicosanoid sythesis and secretion
Arachidonic acid, protaglandins, thromboxanes, Leulotriene- dramitic effects on vasculature tone and permeability
HOURS Cytokine sythesis and secretion
IL-4, 5, 6, 13, TNF-alpha, MIP-1alpha
Animal dies of low Blood Pressure due to acute inflamatory response -
TYPE I HYPERSENSITIVITY
ROLE OF EOSINOPHILS -
Mobilizes and activated preferentially in Type I Hypersensitivity reactions
- IL5 form Th2 mobilize marrow esosinophils
- Mast cell degranulation, inflammation increases eosinophil chemotaxis
Peiecemeal Degranulation
- induced by binding of IgE/antigen to FcRII and by Th2 cytokines (IL5)
- small vesicles of the large secondary granules (crystaloid) move to plasma membrane and release granule contents
- very damaging ie peroxidase, neurotoxin etc
- keep inflammation going in chronic allergic reaction -
TYPE I HYPERSENSITIVITY
DEVELOPMENT -
Sensitization
- can't be allergic on first encounter with antigen
- phase of exposure such that IgE immune response is induced
- nature, site, dose (high) result in Th2 dominated immune response
IgE is produced and will bind to the high affinity FcI receptors on mast cells
Re-Exposure
- to antigen results in CROSS-LINKING of the IgE on mast cells causing release of granules initiating response
Sensitivity should tail off if no ongoing exposure for more that 11 days due to 1/2 life of IgE -
TYPE I HYPERSENSITIVITY
CLINICAL MANIFISTATIONS
ANAPHYLAXIS -
MAIN SPECIES VARIATION IS TARGET ORGAN
Symptoms depend on number and location of mast cells affected
Drugs (penecillin), Milk allergy, parasites arble pupae disruption, blackflies
Cattle, sheep, humans
- serotonin, kinins, leukotriens
- lung
- dyspnea via bronchoconstriciton and pulmonary edema
Horse
- seotonin, histamine
- lungs and intestine
- dispnea, diarrhea
Dog
- histamin, prostoglandin, leukotrienes
- liver
- weakness, collapse hypotension
Pigs
- histamine
- lungs
- syspnea
Cats
- histamine leukotrines
- lungs, skin
- dyspnea, puritis -
TYPE I HYPERSENSITIVITY
CLINICAL MANIFISTATIONS
ALLERGIES -
Site of symptoms do not always relate to site of exposure
- most allergens manifest in skin
- histological lesions edema and infiltration of mast cells, eosinophils results in acute inflammation
SKIN
- puritis via eosinophil granules impacting nerves
- 30% of K9 skin desease
- Urticaria (Hives), Angioedema sub Q
- Atopic Dermatitis genetic, percutaneous
- Flea Allery Dermatitis may also involve Type IV HS
- Eosinophilic granuloma complex cat high eosinophils, nasty lesions
FOOD ALLERY
- 2% of food absorbed as peptides large enough to be allergens
- most cases puritic dematitis, 15% GI, vomiting, diarrhea
- 4 -4 24 hrs of reexposure
- diagnose via exclusion diets
- May be associated with IgA which acts at mucosal surfaces
- tf if deficient get high peptide entry
RHINITIS/ASTHMA
- one of components in COPD horse
- dogs and cats but not common
- genetic -
TYPE I HYPERSENSITIVITY
TESTING - ELISA for specific IgE
-
TYPE I HYPERSENSITIVITY
TREATMENT -
Avoid allergens
Treat secondary infections
Antiinflamatory agents to prevent mast cell degranulation
- 2 types of catecholamine receptors block or stimulate
HYPOSENSITIZATION
- repeated injection of allergen sub Q (macrophages)at increasing doses
- promotes swtich to Th1 and IgG response
- maintenance every 2 - 4 weeks - TYPE II HYPERSENSITIVITY
-
Destruction of CELLS due to binding of antibodies (IgG, IgM) to cell surface
Mediated by:
- NK CELLS
- Complement
- Phagocytic Cells
PHAGOCYTOSIS
- occurs mainly in the spleen
- Extra vascular hemolysis
COMPLEMENT
- lyssis cccurs mainly in the bloodstream
- Intra-vascular hemolysis
Damage does not occur in isolation from other immune mediated damage II and IV
Damage also occurs during beneficial immune inflammatory conditions
- destruction of virus infected cells -
TYPE II HYPERSENSITIVITY
IMMUNE MEDIATED HEMOLYTIC ANEMIA -
Antibodies directed to
- self RBC antigen ie cross reactivity
- altered self RBC antigen ie drugs creating novel epitopes
- exposure of cryptic self antigens ie drugs, microbes, toxins
- adsorbed foreign antigens ie immune response destroys the RBC in the process of attacking the foreigner ie penicilling, ASA, EIA
- Alloantigens ie transfusions, neonatal isoerythrolysis
Direct Coombs Test
- agglutination test for IgG, IgM antibodies binding to rbcs and for fixation of C3
- -
TYPE II HYPERSENSITIVITY
NEONATAL ISOERYTHROLYSIS -
Alloantigens inherited from sire are expressed in the fetus
Late in gestation the placenta develops small tears which allow the feta tissues (aalo antigens) into dam
Dam mounts immune response ro foreign bllod group antigens
Antibodies concentrate in colostrum
Colostral antibodies caus RBC lysis in newborn
Diagnose via Indirect Coombs test
- stallion blood and mare serum plus anti sera to mare Ig -
TYPE II HYPERSENSITIVITY
DISEASES -
Cold Agglutinin Disease
- necrosis of the tips, tail ears, toes
- associated with cold reactive IgM (bigger) to RBCs
- formation of micro thrombi in small blood vessels
Immune Mediated Thrombocytopenia
- petechial hemmorrhages on skin and mucus membranes
- can be transferred by maternal antibodies, piglets or sheep via bovine colostrum (heterophile antibodies)
Skin Diseases
- Pemphigus and Pemphigoid target antigens in epithelium
- severity depends on depth of target antigen ie basement membrane is severe
Addisons Disease
- antibodies target adrenal cortex
- low levels of corticoids
Diabetes Mellitus
- target cells are insulin producing beta cells
Myasthenia Gravis
- antibodies to AchR block and initiate complement mediated injury to the post synaptic membrane
Immune mediated destruction of the thyroid
- antibodies to thyroid follicular cells and thyroglobulin
- thyroid high conserved tf antigens are the same across individuals and species - TYPE III HYPERSENSITIVITY
-
Immune Complex Disease
Immune complexes are formed with SOLUBLE ANTIGEN
Complement is fixed by the CLASSICAL pathway
Chemotaxic and activation substances (C3a, C5a) for NEUTROPHILS are generated from complement pathway
Neutrophil DEGRANUALATION is EXTRACELLULAR
- because the immune complexes formed from soluble antigen are on sufraces the neutrophil cant phagocytise
Effort to clear persisten antigen may trigger Type III
Clears if antigen can be cleared -
TYPE III HYPERSENSITIVITY
ARTHUS REACTION -
Antigen inject sub Q into animal with high levels of antibody
- 6 - 8 hrs peak of thrombosed blood vessels
- antigen diffuese into blood vessel walls
- antigen forms immune complexes with between and beneath vascular endothelial cells
- immune complexes fix complement in blood vessel walls
- neutrophil chemotaxis and degranulation
- problematic lesion is Vasculitis ie immune complexes and inflammation generated CLOT FROMATION and ISCHEMIA -
TYPE III HYPERSENSITIVITY
LOCAL -
Blue Eye
- anterior uvetis and crneal edema in dogs following vaccination with modified live adenovirus
Feline Infectious Peritonitis uveitis
- immune complexes
Hypersensitivity Pneumonitis
- lungs of immune animals after inhaling antigens
- mold spores encounter antibody and form immune complexes in alveolar walls
Tissue damage involved in immune responses to many pathogens may also be mediated through the formation of immune complexes
- Staphlococcal Hypersensitivity
- Neutrophilic dermal vasculitis -
TYPE III HYPERSENSITIVITY
GENERALIZED -
Large immune complexes or those bound to RBCs or platelets are removed by proffesional phagocytes
Small circulating immune complexes that cant be bound by phagocytes become deposited in blood vessels
- especially medium sized arteries of glomeruli, synovia, skin
- areas of high volume low pressure
Serum Sickness
- soluble immune complexes with horse serum
Systemic Lupus Erythematosus (SLE)
- development of autoantibodies to many self antigens
- mmune complexes deposited in basement membrane zone of skin, glomeruli of kidney, small vessels
- diagnose via anti nuclear antibodies
- implies failure of regulation to produce antibody to DNA (poor antigen)
Immune Mediated Arthritis
- rheumatoid arthritis
- synovial membrane of joint
- Aglutination test for rheumatoid factor (RF)
- RF is an IgM auto antibody against immune complexed IgG
Purpura Haemorrhagiica
- horses recovering from Srep. equi (strangles) - TYPE IV HYPERSENSITIVITY
-
Delayed Type Hypersensitivity (DTH)
CELL MEDIATED
- Activated Macrophages (main effect) and Cytotoxic T Lymphocytes
- Antigen via antigen presenting cell activates Th1 cells
- cytokine release activates macrophages and sometimes CD8 T Cells
Differs from I, II, III because cell mediated not antigen mediated
Type IV reactions can be transfered by Th1 lymphocytes
Normal mechanism for recovery from intracellular infections
- -
TYPE IV HYPERSENSITIVITY
DELAYED TYPE HYPERSENSITIVITY SKIN TEST
DTH -
Intradermal injection of antigen
- because immunitiy via Th1 cells no reaction if not previously exposed
- reaction if Th1 memory cells present
Antigen phagocytosised by APC
Presentation of antigen by MHC Class II to memory Th1 cells
- preferential recognition by Th1 cells because expanded specific population due to prior exposure
- Th1 binding along with IL12 prompts release of of Th1 cytokines (IFN-gamma, TNF, IL2,8)
Th1 cytokines
- activate macrophages
- increase expression of adherence molecules on local blood vessel endothelium
- chemotactic for neutrophils
- extravascularize neutrophils, lymphocytes, basophils
Tissue damage and inflammation peaks at 24 - 72 hours when lymphocytes show up -
TYPE IV HYPERSENSITIVITY
OUTCOMES -
Antigen is cleared
- nomral course of events
- repilcation and spread of virus is limited
- means of killing intracellular bacteria and protozoa
- classic test for cell mediated immunity
Antigen Persists
- granuloma formation
- accumulations of macrophages
- giant fused cells
- fibrosis and pathological consequences -
TYPE IV HYPERSENSITIVITY
PATHOLOGICAL CONSEQUENCES -
Feline Leprosy
- cutaneous mycobacteria causes grnuloma formation
Johnes Disease
- mycobacteria paratuberculosis
- macrophages in lamina propria of GI tract
Leishmaniasis
- protozoa in dermal macrophages resulting in grnulomatous dermatitis
Contact Hypersensitivity
- dermatits resuling from contact with wide range of chemical or biological substances
- contact irritant substance causes direct tissue inflammation
- Type IV hypersensitiviy from prolonged exposure to smarll reactive molecusle ie drucgs neomycin, dyes, plastic food bowles, metal etc
- occurs via haptens
Graft Rejection -
TYPE IV HYPERSENSITIVITY
ALLOGRAFT REJECTION
3 TARGETS -
Targets of Graft Rejection are the Histocompatibility Antigens
MHC I on all nucleated cells
- looks self MHC I + foreign peptide
- cytotoxic T cells triggered to kill all cells epxpressing foreign MHC I
- Helper CD4 T cells recognize a complex of MHC I and foreign antigen
- foreign MHC I antigens are interpreted as self + foreign
- Th cell is triggered to release helper cytokines
MHC II on APCs and B cells
- Helper CD4 T cells recognize a complex of MHC II and foreign antigen
- foreign MHC II antigens are interpreted as self + foreign
- Th cell is triggered to release helper cytokines
Blood goroup antigens
- occur on all nucleated cells as well as on RBCS
- associated with antibody responses
- trigger type II destruction of the graft -
TYPE IV HYPERSENSITIVITY
GRAFT VS HOST DISEASE -
Lymphocytes in graf develop and immune response to host tissues
- Recipient is usually immunosuppressed
- donor passenger lymphocytes with the graft mount a type IV response towards recipient cells -
TYPE IV HYPERSENSITIVITY
ALLOGRAFTS AND PREGNANCY -
Fetus is an allograft
Mechanism which prevent rejection:
- no expression of immunogenic MHC I or II molecules on the cell layer of the trophoblast in contact with maternal tissues
- fetal antigens entering the maternal cirulation initiate an antibody response which limits cell mediated immunity via destruction of antibodies in liver
- placenta nd fetus are source of many immunosuppressive factors ie estradiol, progesterone, alpha fetoprotein, indolamine
- Mild immunosuppression and kewing to anitbody response is feature of pregnancy -
PRIMARY IMMUNE DEFICIENCEIES
3 TYPES -
Inherited or Congemital
Defects in Phagocytosis
Defects in Antibody Production
Defects in T Cell Functions
Rare in outbred ie people, cats
More common in inbred ie dogs, dairy cattle , horses
Increased susceptibility of infections at young age
- especially opportunistic invaders
- pathogens
- increased IgE response
- Increeased autoimmune diseases
- increased malignancies
SCIDs Severe Combined Immuno Deficiency -
PRIMARY IMMUNE DEFICIENCEIES
DEFECTS IN PHAGOCYTOSIS -
Phagocytic cell function requires
- normal neutrophil cell numbers and morphology
- response to chemotctic factors
- Phagocytosis ie opsonization, engulfment, formation phgolysosome
- oxidative burst
- killing of micro organisms
Failure of Opsonization
Defect in formation of intracellular granules
- abnormally large granules associated with grey coat colour
BOVINE LEUKOCYTE ADESHION DEFICIENCY (BLAD)
- defect in the adhesion proteins that enable neutrophils to leave the bloodstream and enter the tissues in response to chemtaxic factors
- Integrin on surface of neutrophils defective
- neutophils cannot squeeze through vasculature
- results in uncontrolable bacterial infection in tissue
- pneumonia
- NO NEUTROPHILS IN TISSUE BUT BLOOD VESSELS PACKED FULL
- Holstein cattle (14% of bulls, 5% cows)
- Irish setters -
PRIMARY IMMUNE DEFICIENCEIES
LYMPHOCYTE DEFECTS -
B Cell and or T Cell Defects
Test via:
- numbers and morphology of cells
- Expression of CD markers
- cell function assays ie blastogenesis, cytokine release
- DTH or antibody response in vivo
SEVER COMBINED IMMUNODEFICIENCY (SCID)
- people cattle dogs, horses
- 8% of Arabs carriers in NA
- molecular test
- absence of a component of a DNA dependen torptein kinase which is required to rejjoin the ends of DNA when V, D, J genes are cut and rejoinded to produce BCRs and TCRs
- no production of functional T or B cells
- have NK cells
- undeveloped lymph nodes
- low circulating lymphocytes
- no IgM before suckling
- survive as long as colostral antibodies are present (t1/2 3 weeks)
- die from first opportunistic invader ie adenovirus or pheumocystis
- X linked SCID defect in gene that codes for the gamma chain of IL-2 Receptor and component of many cytokines tf lymphocytes csnt proliferate -
PRIMARY IMMUNE DEFICIENCEIES
B CELL DEFECTS -
Usually result in lowered levels of serum Ig
- measure via radial diffusion
Agammaglobinemia
- no Bcells, no serum Igs
Trasient Hypogammaglobinemia
- production of Igs delayed for 2 - 3 months
Selective IgA Deficiency
- infections at surfaces ie respiratory, skin, eyes
- most clinically relevant ie survivable
- increased autoimmunity, cancers
- German Shepards
Selctive IgM Deficiency
- IgA, IgG levels normal or elevated
- recurrent bacterial infections
- mortality after a few months from pneumonia
Selective IgG Deficiency
- IgA, IgM levels may be normal -
PRIMARY IMMUNE DEFICIENCEIES
T CELL DEFECTS -
T Cell defects tend to be incompatible with life
- tf fewer conditions perptuated
NUDE CONGENITAL HYPOTRICHOSIS AND THYMIC APLASIA
- cats, rats, pidgs, calves, guinea pigs, MICE
- NO THYMUS because of deficiency in epithelial cells
- tf animals have no hair
- used as living test tubes
- animals die once maternal antibody declines
Parvo virus
- breed susceptabilitiy suspected due to T cell defficency -
SECONDARY IMMUNE DEFICIENCEIES
VIRUS INDUCED -
Virus Induced Immunoduppression
- usually associated with infection of the lymphocytes
- damage/loss of lymphocytes results in decreased cell numbers and functions
- less often vuruses stimulate lymphoid cell activity ot patholgical extent ie unable to respond to other pathogens
- other viruses cause lymphoid neoplasia
- tf neoplastic cells replace normal lymphocytes
Infectious Bursal Disease Virus
- chickens
- targets bursa and secondayr lymphoid organs
Primate Retroviuses
- HIV I and HIV II
- targets CD4 cells damaging receptor
- tf failure of antibodiy and cell mediated response
FeLV (Feline Leukemia Virus)
- infectsts lymphocytes
- various effects including neoplasia
Feline Immunodeficiency Virus (FIV)
- ralated to HIV
- infects T cell, mega karyocytes, macrophages, dendritic cells, neurons
- inital reponse to virus is follicular hperplasia
- followed by gradual depletion of T Cells
Canine Distemper Virus (CDV)
- infects and destroys secondary lymphoid organs
- depresses activities of lymphocytes and macrophages via IL1,2
Bovine Viral Diarrhea Virus
- infects and can destroy lymphocytes
- may preferentially target Peyers Patches tf if infected early will have life long immunodeficiency
- acute infections result in long term decrease in immune responses -
SECONDARY IMMUNE DEFICIENCEIES
OTHER CAUSES OF IMMUNOSUPPRESSION -
Lymphoid Neoplasia
Nicrobial and Parasitic Infections
- chronic disease results in diminished immune reponses
- decreased IL2 production
- decreased proliferative response
Malnutrition
- first affects T Cell functions, complement, neutrophill funtion
- driven by protein loss
- zinc deficiency is especially immmono suppressive
- also vit A, Bqw folic acid, copper selenium, taurine
Exercise
- regular moderate excercise increases immune response
- strenuous excercise has opposite effect possibly via increased steroid levels
Trauma
- severe injuries rusultin immmunodeficiency via corticosteroids, prostoglandins, protein loss
Age
- B and T Cell response decrease with age
- Thymic involution lowers T cell production
- tf majority of cells in secondary lymphoid organs are memory cells
- loss of T cell division and response to IL2
- Bone marrow ok but B cell repitoire diminished because of lower Th cells
Stess
- many management situations are stressfull -
SECONDARY IMMUNE DEFICIENCEIES
DRUGS -
Nonspecific Immunosuppression
- drugs or radiation that have general effects on rapidly dividing celss
- GIT, skin, blood and immune system cells
CORTICOSTEROIDS
- steroids: wide variety of suppressive effects
- mechanisms: poorly understood
- Steroids dircetly absorbed and transported to nucleus
- block cytokine synthesis
- humans and lab animals more susceptable than domestics
- lymphopenia ie reduced emigration of neutrophils to tissues
- decreased chemotaxis and phagocytosis
- decreased cytokine production tf redueced T cell response
- inhibits acute inflammatory response via decrased macrophage production of protaglandins and IL1
CYTOTOXIC DRUGS
- designed to inhibit cell deivison
- inhibit immune response by reducing cell proliferation
- toxic effects on bone marrow tf thrombocytopenis, anemia, leukopenia
DRUGS THAT TARGET T CELLS
- developed from fungi
- blocks signal transduction oin Th1 cells tf lower production of IL2 and ganna INF
- VERY IMPORTANT IN GRAFT SURVIVAL
- ie renal graft cats can last more than a year -
TUMOR IMMUNITY AND IMMUNOTHERAPY
TUMORS -
Tumor = Cancer = Neoplasms
Benign
- remain in a single location
- not highly invasive
Malignant
- mestastic via blood or lymph
- highly invasive
Carcinoma
- arises from epithelial cells
Sarcoma
- arise from mesenchymal cells
Leukemia
- tumor cells of haemopoietic cells in blood -
TUMOR IMMUNITY AND IMMUNOTHERAPY
TUMOR SURVEILLANCE -
Although tumors arise after reproductive age immuno suppression results in increased rates of some cancers indicating some role for the immune system in the prevention of some cancers
But most tumors are not a consequence of falure of the immune responses
- nude mice are not at increased risk for most cancers -
TUMOR IMMUNITY AND IMMUNOTHERAPY
ANTIGENS -
Tumor cells are genetically unstable
- tf gain and lose surface antigens
- loss of MHC and or blood group antigens
- expression of developmental proteins
- new antigens from incomplete or altered synthesis of normal proteins
- virus related antigens in virus induced proteins
- expression of antigens normal to other tissues
- expression of antigens in unsusual quantities
Most structures that differ are CHO and lipids which are poor antigens -
TUMOR IMMUNITY AND IMMUNOTHERAPY
NATURAL KILLER NK CELLS -
Main Effector Cells in tumor immunity
- 15% of lymphocytes are NK
- found in secondary lymphocytes do not recirculate
- No T or B cell markers
- Not MHC restricted ie no specificity
- kill virus infected and tumor celss without pervious sensitization ie no memory
- serial killers not damaged in process of killing
- kill via perforins and ganule enzymes ala CD8 T cells
NK cells recognize Tumor cells via:
- loss of MHC I expression
- recognition of antibody via CD16 Fc receptor
- binding CD 16 triggers release of INF gamma and TNF
- recognition of MICA and MICB proteins expressed by stressed cells -
TUMOR IMMUNITY AND IMMUNOTHERAPY
OTHER EFFECTOR MECHANISMS -
Cytotoxic T cell
- most important in virus induced tumors that express viral antigens
Activated Macrophages
- via IFN gamma
- release TFN
Antibody
- may have effects through complement mediated lysis and NK mediated killing -
TUMOR IMMUNITY AND IMMUNOTHERAPY
FAILURE OF IMMUNITY -
NK cells are capable of killing oly certain types of tumors
- ineffective against carcinomas (70% of tumors)
- better against sarcomas
Tumors are poorly immunogenic
- tf other effector mechanisms are not activated
Tumor population is gentically unstable
- antigenic heterogenity tf only some tumor cells targeted
Failure to express Class I MHC antigens
- tf best killers, CD8 T cells not active
Immunosuppression
- relese of immunosuppressive factors ie prostaglandins and cytokines form tumor cells
- catabolic state tf protein drain
- Suppressor cells: regulatory poplulations of T cells result in AB mediated response which is not as effective
Blocking Antibodies
- antibodies bind tumor cell without harm but suppress cell mediated immunity
Tumor Cell Selection
- tumor cells appear normal
- do not trigger immune response until large size
- immune systme must kill every tumor cell
- if large high heterogenity tf hard to do -
TUMOR IMMUNITY AND IMMUNOTHERAPY
IMMUNOTHERAPY -
Nonspecific Immune Stimulation
- attenuated mycobacteruim bovis (BCG) activates macrophages tf increases cytokine release
Active Specific Immunization
Vaccination to prevent tumors
- difficult because of heterogenity
- used for viral induce tumores ie FeLV, Mareks Disease
Vaccination to TREAT tumors
- harvest tumor
- use to produce vaccine via adding adjuvant or treating tumor cells to increase immunogenicity
- papillomas, ocular squamous cell carcinomas in cattle
- equine lymphsorcoma via infecting tumor cells with vaccina virus which produces antigen for NK cells
- some results but NOT SUSTAINED
Monoclonal Antibodies
- antibody and complement dependent cytotoxicity
- coupled to a toxin
- coupled to a radioisotope
- coupled to an enzyme which specifically activates a cytotoxic drug
- heterogenity is problem
- low proportion of dose reaches tumor cells
- production of immune reponses to monoclonal Igs ie usually from mouse
- can build recombinant Ig
- anti tumor effects present but not clinical or sustained in most patients
Cytokine Tumor Immunotherapy
- Interferons successful in some tumors esp hhuman hiry cell leukemia and kaposi sarcomas
- cytokines are very toxic
- tf local application best ie injection inotp papillomas or carcinomas of cattle
Cytokine Activated Cell Therapy
- lymphokin Activated Killers (LAK) cells:
- blood lymphocytes harvested and incubated with IL2 then reinfused into patient
- mainly expands NK population
- Tumor Infliltrating Lymphocytes (TIL):
- lymphocytes harvested from tumor tissue
- cultured with IL2 and reinfused
- killers are both NK and T cells
- can produce remission -
IMMUNE MEMORY
CLONAL SELECTION -
Once a lymphocyte binds antigen it DIFFERENTIATES into either:
- Effector Cell
or
- Memory Cells
both then proliferate -
IMMUNE MEMORY
MECHANISMS OF STRONGER MEMORY RESPONSE
4 KEY WORDS -
Increased number of antigen specific RESPONDER cells
- ie the memory cells
Increased responsiveness to antigen presenting cells
- naive cells require more costimulatory bonds and more time to become activated
- memory cells have lower activation threshold and require less costimulatioN
- tf more effective RECOGNITION
Better expression of costimulatory molecules
Phenotypic commitment
More effective MIGRATION
- naive T cells recirculate searching for novel antigen
- memory cells located in tissues at invasion sites
More effective FUNCTION
- class switch has already occured for B cells
- broader range of cytokines -
IMMUNE MEMORY
KINETICS OF T AND B CELL RESPONSE -
IMMEDIATE 12 - 24 HRS
Primary - Induction
Secondary - Amplification, Differentiation
EARLY 24 - 96 HRS
Primary - Amplification, Differentiation
Secondary - Effector
LATE > 96 HRS
Primary - Amplification, Differentiation, Effector
Secondary - Effector -
IMMUNE MEMORY
CLINICAL DISEASE - Race between pathogen's ablility to replicate and/or cause disease and the speed with wchich to body can mount a protective immune response
-
IMMUNE MEMORY
IMMUNITY - Ability to mount MEMORY response
-
IMMUNE MEMORY
PROTECTION -
Clinical disease does not Develop
- ie difficult to have for rota virus because would need to have antibody in GIT
-ie easy to have for small pox because it has a long incubation period -
IMMUNE MEMORY
DURATION OF MEMORY
5 WAYS -
Memory cells PERSIST in lymph node follicle in presence of antigen
Dendritic cells in lymph nodes SEQUESTER antigen
- months to years
RE-EXPOSURE to low levels of endemic antigen
Exposure to related antigen via CROSS REACTIVITY
Physiological MAINTENANCE of memory cells via CYTOKINES
- non antigen dependent mechanism - HERD IMMUNITY
-
The disease sparing effect on susceptible individuals within a group of individuals that are immune
Immune individuals REDUCE the PROBABILITY of an ADEQUATE CONTACT with susceptable individuals -
HERD IMMUNITY
IMMUNITY IN POPULATIONS
3 FACTORS -
Persitence and Spread of disease within a population depends on:
Immunity of INDIVIDUALS
- ie innate, T-cells, B-cells, antibodies, cytokines
Nature of CAUSAL AGENT
- How contagious
- host specificity
- Duration of infection
- carrier status
Structure and Dynamics of Population
aka Disease Triangle or Ecology of disease -
HERD IMMUNITY
REED - FROST MODEL
3 CATAGORIES
3 ASSUMPTIONS -
Individules categorized as:
- Infected
- Susceptable
- Immune
Assumptions
- infection is only spread from infected individuals to others by adequate contact
- if an individual is susceptable and adequately contacted it will develop disease
- there is a fixed probability of adequate contact between individuals -
HERD IMMUNITY
5 EFFECTS
OF INCREASING # OF IMMUNE ANIMALS IN POPULATION -
Delays Epidemic
Increases duration of epidemic
Reduces Magnitude of Epidemic
Reduces total number of infected individuals
Can prevent epidemic -
HERD IMMUNITY
EFFECTS OF ADEQUATE CONTACT -
The rate of contact of cases (infecteds and contagious) with susceptable animals determines if an epidemic can start
- decrease contact disease dies out
- increase contact maintenace of disease in populationn or epidemic -
HERD IMMUNITY
BASIC REPRODUCTION RATE
HERD IMMUNITY THRESHOLD -
Basic reproduction rate (Ro)is the spreading potential of an infection in a population
- function of biological mechanism of transmission
- function of rate of contact between members of the host population
Herd Immunity Threshold (H) is the proportion of immune individuals which must be exceeded if disease incidence is to decrease
- ie a disease with lower transmissiblity requres a lower number of immune individuals to avoid disease
- many common contagious diseases have an immune threshold of 80 - 85% -
HERD IMMUNITY
2 WAYS OF ENHANCEMENT -
Increase the proportion of immune animals in a population
- increase exposure to disease when appropriate
- increase vaccination
Reduce the probability of adequate contact
- identify and isolate infected animals
- pervent clusters of increased contact -
HERD IMMUNITY
POPULATION DYNAMICS -
Clusters of Susceptable Animals
- age cohort
- sex cohort
- emigrant populations
- management groups
- house holds
Clusters of Adequeate Contact
- Pounds, kennels
- racetracks, horse shows
- auction markets, trucks
- vet clinics, treatment areas -
EVOLUTION OF IMMUNE RESPONSE
INVERTEBRATES -
PHYSICAL BARRIERS
PHAGOCYTOSIS
- remember Metchnikov and Starfish larvae
- universal defense mechanism
- hemocytes, coelomocytes
- chemotaxis, adherence, ingestion, digestion (proteases and oxidants)
- enhanced activity via cytokine like (TNF, IL1,6) molecules
PROPHENOL OXIDASE activating system
ANTIMICROBIAL PEPTIDES
COMPLEMENT
- ancient system
- alternate and lectin pathways (note lytic pathway (C6 and MAC) does not appear until lancets)
- C like proteins occur in echinoderms, sea urchins have C3, C2/factor B
Limulus Amoebocyte Lystate (LAL) Assay
- use of horseshoe crab clotting defense mechanism to detect endotoxin (LPS) -
EVOLUTION OF IMMUNE RESPONSE
IMMUNOLOGICAL BIG BANG -
Development of Specific Aquired Immunity via genes for B and T receptors
- started with jawed fish (sharks and above)
- inclusion of bacterial TRANSPOSON via integrase gene
- BCR and TCR require rearangement of V, D, J genes -
EVOLUTION OF IMMUNE RESPONSE
JAWED FISH -
INNATE
- phagocytosis similar to mammals
- granulocytes enter inflamatory sites first
- originate from head kidney (lympohid function)
- macrophages (blood) and lymphocytes
- melanin prevents oxidation
- lysozyme, defensisns, C, acute phase proteins
- classical (antibody dependent), alternate and lectin pathways
- MAC
ACQUIRED IMMUNITY
- full set of lymphoid organs except bone marrow
- pronephros (head kidney) has antibody forming cells and macrophages
- Immunoglobulins Ig cluster genes, IgG, NO EVIDENCE OF OPSONIZATION
Cell Mediated Immunity
- TCR, MHC I, II -
EVOLUTION OF IMMUNE RESPONSE
AMPHIBIANS -
Potent Antimicrobial Peptides in SKIN
C system similar to mammals
- works best at 16 deg C
Uordeles (newts and salamanders more primitive)
- lack bone marrow
- monomeric IgM
- do not react to soluble portein antigens ie toxins
Anurans (frogs and toads)
- IgM pentamers or hexameres
- IgY
- IgX
- secretory antibody (IgY, IgX)
- Functional TCR ie graft rejection
- Cytokines IL1,2, INFs
- MHC II larva
- MHC I adults
- immunosuppresed during metamorphis -
EVOLUTION OF IMMUNE RESPONSE
REPTILES -
Age and seasonal involution of Thymus
Splenic structure similar to mammals
Primative lymph nodes
- no germinal centres
C3 in venom of some snakes
Immunoglobulins
- IgM primary, IgY secondary
- Respond to BSA
- T cell dependent response
- no secondary response to LPS ie T independent via CHO ala mammals
Cell Mediated Immunity
- Graft rejection (temperature dependent)
- mixed luekocyte reactions
- Delayed Type Hypersensitivity (DTH) -
EVOLUTION OF IMMUNE RESPONSE
BIRDS -
Organs
- Thymus
- Bursa
- primative lymph nodes
- well developed germinal centres
Immunoglobulins
- IgM primary
- IgY secondary main and involved in opsonization
- truncated reduces lethal hypersensitivity
- IgA including secretion
- Diversity generated in ovo via one V and J gene tf less than mammals
Cell Mediated Immunity
- Graft Rejection
- DTH
- alpha,beta and gamma,delta T cells
- Th1,2 helper cells with appropriate cytokines -
IMMUNE GENETICS
MHC -
MHC I
- A, B , C genes
MHC II
- DP, DQ, DR genes
Homos 3 genes, Heteros 6 genes
MHC is DEGENERATE
- recognizes many epitopes
- ie has a peptide pocket on size fits most
- Specificity is deterimined by T cell recognizing specific bound epitope in context of MHC -
IMMUNE GENETICS
BCR -
GENES
- V = variable region
- J = joining
- D = diversity
- C = constant region
HEAVY CHAIN
- coded for by two gene clusters, one cluster for constant domain and one cluster for variable domain
VARIABLE REGION
- coded for by muliple genes
- gene rearrangement via looping out, deletion and splicing
CONSTANT REGION
- coded for by a single gene for each class
CLASS SWITCHING
- via combining any variable region with a constant region (class)
- recombination for class swtich occurs via excision (Rcombinase) and splicing (DNA PK repair enzymes - defect = Equine SCID)
- induced via Th signaling and cytokines
Light Chain Exclusion
- only one of kappa or lambda light chain is transcribed
HYPERVARIABLE REGION
- aka Complemntarity Determining Region (CDR)
- 6 - 10 ammino acids
- shape deterimines specificity of epitope binding -
IMMUNE GENETICS
TCR -
GENES
- V = variable region
- J = joining
- D = diversity
- C = constant region
- fewer VDJ genes than BCR but greater diversity via junctional diversity
TCR TYPES
- alpha/beta or gamma/kappa types are created by 3 gene clusters
HYPERVARIABLE REGION
- aka Complemntarity Determining Region (CDR)
- CDRs 1,2,3 in V region and V(D)J region
- gene rearrangement via looping out, deletion and splicing -
IMMUNE GENETICS
MECHANISMS TO GENERATE DIVERSITY
7 FOR BCR
4 FOR TCR
LOCATION AND IMPORTANT DIFFERENCE -
BCR
- occurs in Bone Marrow, Bural Equivalent ie Peyers Patches, Lymph node ie Germinal Centre
- VJ and VDJ gene recombination
- Base Deletion
- Base Insertion
- Combinatorial association ie combination of different K or L light chains wtih different heavy chains
- Gene Conversion ie segments of psuedo genes inserted into Variable region
- Receptor Editing ie rearrangement of VDJ genes after antigen exposure in Germinal Centres. Note if after 4 attempts B cell cant bind antigen targeted for apoptosis
- SOMATIC MUTATION ie after antigen exposure, mediated by cytosine deaminase converting cytosines to uracils which are then repaired as thymadines
Note if after 4 attempts via Receptor editing or somatic mutation B cell cant imporve antigen binding it is targeted for apoptosis ie cell selection
TCR
- occurs ONLY in the thymus
- VJ, VDJ and VDDJ gene recombination
- Base Deletion
- Base Insertion
- Combinatorial association ie combination of different K or L light chains wtih different heavy chains
Note the only thing worse than having somatic T cell Mutation in your body (would get mondo B cell response to self via Th not to mention killer T cells) would be to neglect to point out that SOMATIC MUTATION DOES NOT OCCUR IN T-CELLs on the exam -
IMMUNE GENETICS
HERIDITY AND DISEASE RESISTANCE -
Immune responsiveness is inherited via multiple genes
Immune system is one of most variable physiological systems
Primary Genes:
- BCR, TCR and MHC
Secondary genes:
- immune regulatory, innate
Parameters
- Ig []
- antibody response
- T cell proliferation
- DTH
- Macrophage function
Identify resitant individuals and characterized response
Select for appropriate parameters -
MUCOSAL SURFACES
3 FAST FACTS
3 FUNCTI0NS -
Large surface between body and evironment
More than 95% of pathogens enter body via mucosal surfaces
Mucosa-associated lymphoid tissue (MALT) represents LARGEST IMMUNE COMPARTMENT WITHIN THE BODY
Physical BARRIER
TOLERATES Antigens of commensal flora, nutritional and normal enviromental elements
Mounts rapid and psecific IMMUNE RESPONSE against invading pathogens -
3
DEFENSE MECHNAISMS
AT
BODY SURFACES -
Non Immunological
- Mucus, saliva tears
- Normal bacterial flora ie outcompete pathogens
- Enzymes ie lysozyme
- Fatty acids on skin
- mucocillary escalator
- vomiting
- diarrhea
Innate Immunity
- Antimicrobial Peptides
- Phagocytes ie eosinophilic granulocytes
- TLRs + PAMPs = activation of monocytes, macrophages and dendritic cells results in INDUCTION of innate and psecific immune repsonses
Specific Immunity
- Antibodies IgA monogastrics, IgG ruminants
- Cytotoxic T Cells - DENDRITIC CELLS
-
Link Innate and Specific Immunity
Immature dendritic cells take up antigen/pathogen at mucosal surface via TLRs
Binding TLRs activates NFkB intracellular system which upregulates cytokine production
- TNF alpha, IL12, INF alpha attracts/activates NK cells
- TNF, IL6 attracts B cells
- TNF, attracts T cells
ie Activates Innate immune system and ALERTS Specific immune system
Dentritic cell migrates to lymph nodes and becomes mature via changing surface receptors from antigen sensing to ANTIGEN PRESENTING
- up regulates costimulatory molecules stimulating T-cell proliferation
- ie ACTIVATES Specific immune system -
ANTIMICROBIAL PEPTIDES
FAST FACTS AND FUNCTIONS -
aka HOST DEFENSE PEPTIDES
- do much more than just make sieves out of microbes
Small peptides secreted by
- epithelial cells
- granulocytes
- macrophages
Two Families
- Defensins
- Cathelicidins
FUNCTIONS
Antimicrobial Activity
- several defensins insinuate into outer leaflet of cell membrane via cationic peptides then join to create pore
Chemmoattracts for Immune Cells
Activate antigen presenting cells
Immunomodulate
- specific peptides stimulate either Th1 or Th2
- tf can be used as adjuvants
Promote wound healing and angiognesis
Anti-endotoxic (LPS) effects
Increase inflammation
In short don't leave home without them safely stowed in your epithelium because they could just be the most important molecules in the whole immune system -
SPECIFIC IMMUNITY
MUCOSAL BASICS -
Cytotoxic TCD8 cells reside in the epithelium amongst the epithelial cells
- gamma-delta T cells prevelant in intestinal epithelium
Plasma Cells (B cells stimulated by Th cells) reside in Lamina Propria where the secrete antibody -
MALT
COMPARTEMENTS AND SIGNIFICANCE -
Mucosa Associated Lymphoid Tissues
Form the COMMON MUCOSAL IMMUNE SYSTEM
- composed of compartments
- GALT Gut associated lymphoid tissues
- BALT Bronchus associated lymphoid tissues
- NALT Nasal associated lymphoid tissues
- Genitlal associated lymphoid tissues
Compartements are connected by permenant but varied exchange of immune cells
- lymphocyte trafficking and homing
INDUCTIVE TISSUES
- afferent limb
- Peyer's patches, Tonsils, Solitary lymph nodules
- M-cells, Dendritic cells, Epthelial cells
EFFECTORS
- efferent limb
- lamina propria
- Intraepithelial Lymphocytes (IELs), Lamina proprial Lymphocytes (LPLs)
Allow infection in one mucosal location to generate memory response (immunity) in another -
MUCOSAL IMMUNITY
ANTIGEN UPTAKE
3 WAYS -
M CELLS
- reside in epithelium above Peyers patchs
- shorter micrvilli creates a pocket that traps particles under 5 micrometers
- antigen is passed through M cell to its basal surface
- basal surface is involuted forming large pocket which is full of lymphocytes and antigen presenting cells which go to town with the antigen
- actually the dendritic cells take the antigen to the nearest germinal center in the Peyers patch
DENDRITIC CELLS
- dendritic cells open up tight junctions between epithelial cells
- extend dendrites (imagine that!) into lumen and fish for antigen
- once bound with antigen they head to a germinal centre to make men and women out of the naive B cells and the T cells join in on the fun
EPITHELIAL CELLS
- not wanting to miss out on the fun epithelial cells can also take up antigen
- however being weekend warriors they are not very effective
- low or no secretion of costimulatory molecules so they are a bit like wall flowers at the dance
- more important in chronic disease situations with severe inflamations -
MUCOSAL IMMUNITY
EFFECTOR CELL POPULATIONS
WHO ARE THEY AND WHAT DO THEY DO -
LPL lAMINAL PROPRIA LYMPHOCYTES
- mainly B Cells and T helpers
- Secretors of Antigen
- IgA MOST IMPORTANT Ig on mucosal surfaces
- IgG also present on mucosal surfaces in ruminants
- IgE also secreted ie allergies and parasites
T HELPER CELLS
- provide micro environment (a satisfying little swirl of cytokines, interleukins and costimulatory embraces)
INTRAEPITHELIAL LYMPHOCYTES (IEL)
- shoulder to shoulder with epithelial cells
- make up to 40% of total peripheral T cells
- mainly of the gamma-delta receptor CD8 cytotoxic types
- express alpha/alpha CD8 homo dimers
- bind MHC Class Ib which is uniquely expressed by enterocytes
- uniquely use Vgamma and Vdelta genes to form TCR antigen binding site
- secrete IFN-g
- regulatory function ie oral tolerance to dietary antigens -
MUCOSAL IMMUNITY
THE IgA STORY
HOW DOES IT GET TO THE SURFACE -
SYNTHESIS
IgA secreted in lamina proprial by plasma cell
- forms dimer via J-chain
Taken into epithelial cell via POLY Ig RECEPTOR (pIgR)and transported to luminal surface where it is SECRECTED and forever known as SIgA
- being a good citizen, the epithelial cell recycle the pIgR rather than let it float around in the lumen
- however there is always someone willing to abuse the system as some pathogens use the recycled pIgR as route through the epithelial cell to the basal side
FUNCTIONS -
MUCOSAL IMMUNITY
THE IgA STORY
3 FUNCTIONS -
FUNCTIONS
Blocks Bacterial Adhesion
- bacteria use adhesins to bind to host cell surfaces
- IgA binds adhesins on pathogen so the bacteria slide on by
- tf E-coli vaccine uses bacterial fibril protein as antigen to stimulate IgA response
NEUTRALIZATION OF VIRUSES
- viruses like to use cell surface receptors (ie AIDS virus uses CCR5 on Th cell)to bind host cell
- IgA via its handy hypervariable region can bind the receptor binding sites on the virus and whisk it off the dance floor
- tf when making vaccines handy to know which proteins viruses use to bind host cell surface receptors
NEUTRALIZATION OF BACTERIAL TOXINS
- most toxins need to be internalized to be effective
- bind cell surface receptors to enter cell
- IgG in this case binds toxin keeping the receptors clear for friendly physiological molecules -
COMMON MUCOSAL IMMUNE SYSTEM
CELL TRAFFICKING -
Hallmark of immune response
NAIVE T CELLS
- remember they already have their specific antigen receptors
- migrated only to secondary lymphoid organs via L-selectin
ACTIVATED EFFECTOR/MEMORY T/B CELLS
Migrate preferentially to site of antigen encounter
- ie HOMING
Mediated via mucosal addressins
Effector cells leave lymph node through efferent lymph
Each tissue expresses different chemokines which are upregulated and expressed during inflammation
- ie CC secondary lymphoid tissue chemokine
- ie alpha4beta7 integrin on effector T cell binds MAdCam on Mucosa
- ie CCR9 is a receptor for TECK/CCL25
- TECK/CCL25 is expressed on endothelial cells of High Endothelial Venules (HEV), site of lymphocyte extravasation, of small small intestine but not the HEV of the colon
- CCR9 is highly expressed on Lamina Propria Lymphocytes (LPL), Intra Epithelial Lymphocytes (IEL) and IgA secreting B cells
- tf SPECIFIC HOMING of LPL, IEL and IgA B cells to small intestine but not colon via SPECIFIC receptor and chemokine EXPRESSION
DENDRITIC CELLS
- specific receptors for lymph nodes once bound with antigen -
COMMON MUCOSAL IMMUNE SYSTEM
HOMING EXTRAVASATION
I SAY MUCOSAL ADDRESSINS
YOU SAY -
Integrins and Selections expressed on surface of lymphocytes
Lymphocytes travel at speed in blood vessels
TETHERING via SELECTINS binding with chemokines expressed on endothelial tissue induces ROLLING which slows them down
ADHESION via INTEGRINS binding with chemokines expressed on endothelial tissue causes ADHESION which brings the lymphocytes to a stop
Once stopped, the lymphocytes can TRANSMIGRATE through the endothelium and follow the immflammatory chemokines to party central -
COMMON MUCOSAL IMMUNE SYSTEM
SUMMARY
FIVE KEY EXAM FRIENDLY POINTS -
Homing of primed lymphocytes is high specific
Mediated by mucosal adressins ie the homing molecules
Primed (Activated/Effector) lymphocytes home selectively to certain compartments of the immunce system
Chemokines are important in orchestratiing migration patterns
Important for development and application of vaccines -
COMMON MUCOSAL IMMUNE SYSTEM
ORAL VACINES
EFFECTIVE AND INEFFECTIVE RESPONSE SITES -
EFFECTIVE
Proximal Small Inestine
- site of e-Coli infections
Ascending colon
Mammary Glands
Salivary Glands
INEFFECTIVE
Distal large intestine
Genital mucosa
Tonsils -
COMMON MUCOSAL IMMUNE SYSTEM
RECTAL VACINES
EFFECTIVE AND INEFFECTIVE RESPONSE SITES -
EFFECTIVE
Rectum
INEFFECTIVE
Small intestine
Proximal Colon -
COMMON MUCOSAL IMMUNE SYSTEM
NASAL OR TONSILAR VACINES
EFFECTIVE AND INEFFECTIVE RESPONSE SITES -
EFFECTIVE
Upper airway,
Regional secretions
Genital Mucosa
INEFFECTIVE
Gut -
COMMON MUCOSAL IMMUNE SYSTEM
VAGINAL VACINES
EFFECTIVE AND INEFFECTIVE RESPONSE SITES -
EFFECTIVE
Genital mucosa
INEFFECTIVE
None tf highly focused -
MUCOSAL VACCINES
THE GOOD AND THE BAD -
ADAVANTAGES
Immune response where needed
Effective immune response
Needle-free injection
No injection site reactions
DISADVANTAGES
Inneffective Delivery Routes
- ie if oral have to contend with the effects of acid and enzymes on the delicate little antigen peptides
- difficult to identify as antigen not nutrition
- tf target M Cells via antibody mediated targeting
- tf use liposomes, microparticles, viral vectors
- tf us
Require large amounts of antigen to make up for inneffective delivery routes
Stimulation of mucosal immune response difficult due to complexity of inductive sites -
MUCOSAL IMMUNITY
REGULATION -
TOLERANCE
Tissue resident dendritic cells and macrophages take in antigen
Release TGF beta attracting other dendritic cells and neutrophils
- ie positive feed back
Regulatory T Cells in laminal propria down regulate immune response in effort to minimize energy expenditure
- ie negative feedback -
MUCOSAL IMMUNITY
SUMMARY
6 POINTS -
Cell Trafficking is essential for effective mucosal immune response
Naive lymphocytes permanently circulate through secondary lymphoid tissues in order to encounter antigen
Effector (activated) lymphocytes home to the mucosal effector tissues
Common mucosal immune system but site specific trafficking
Mucosal immune system is effectively divided into compartments by specificity of trafficking
Important for vaccine delivery - VASOACTIVE MOLECULES
-
HISTAMINE
- released from mast cells, basophils, platelets
- increase vascular
permeability
KININS
- released from plasma kninogens and tissues
- vasodilation, permeablilty
PROSTAGLANDINS
- arachidonic acid
- vasodilation, permeablilty
C3a AND C5a
- serum complement
- mast cell degranulation, sm contraction, Neutrophil chemotaxis (C5a) -
MAST CELLS
ACTIVATORS
SECRETIONS -
ACTIVATORS
- IgG or IgE and antigen
- LPS
- ethdothelins
- adenosin
- neuropeptides
SECRETIONS
- histamine
- IL 4,5,6,13,16
- TNF alpha
- CCL3 chemokine
Proinflammatory or promote Th2 responses - TNF alpha
-
SECRECTED BY
- macrophages
- mast cells
- lymphocytes
- endothelial cells
- fibroblasts
FUNCTIONS
Activates
- mast cells
- dendothelium
- macrphages
- lymphocytes
- neutrophils
INFLAMMATION
- adhesion
- procoagulants
- induces cute phase proteins
- promotes ganuloma formation
ENHANCES
- fibroblast growth
- collagen syntesis
- bone resoption
TOXIC
- kills tumor cells
- septic shock
- sickness behavior
- altered lipid metabolism - IL 1
-
SECRESTED BY
- MAINLY macrophages
- dentritic cells
- lymphocytes T, B, NK
- endothelium
FUNTIONS
- proinflammatory
- mucus production
- induces acute phase proteins
- tissue growth
- IFN gamma synthesis, integrin expression
- fever, drowsines loss of appetite
- Th2 cytokine production
- eosinophil and basophil degranulation
- cytotoxic to tumor cells -
MACROPHAGES
ACTIVATORS
SECRETIONS -
ACTIVATORS
- IL 2, 4, 13
- C 1, 3
- antibody
- PAMPS
- INF gamma
FUNCTIONS
- phagocytosis
- MHC II antigen presentation
- bactercidal activity
- tissue repair IL 4, 13
- IL 1 costimulates Th2, stimulates acute phase response
- IL 6 promotes B cell differentiation, stimulates acute phase response
- IL 12 costimulates Th1 cells
- IL 18 promotes INF gamma production by Th1 cells
- TNF alpha stimulates T cell growth, acute phase response, triggers inflamation -
Th1
FUNCTIONS
ACTIVATORS
SECRETIONS -
FUNCTIONS
- T cell cytotoxicity
- macrophage activation
- some IgG responses
ACTIVATORS
- CD80 costimulation
- IL 12 (dendritic cells or macrophages
INHIBITORS
- IL 10
SECRETIONS
- IL 2 activates T cells, B cells, NK cells and macrophages
- INF gamma inhibits Th2 cells, stimulates Th1 cells, activates NK cells and macrophages
- TNF alpha and beta -
Th2
FUNCTIONS
ACTIVATORS
SECRETIONS -
FUNCTIONS
- costimulation of B cells
- tf IgG response
- tf IgA and IgE production
ACTIVATORS
- CD86 costimulation
- IL 1, 4 (dendritic cells)
INHIBITORS
- INF gamma
SECRETIONS
- IL 4 simulates B cell growth and differentiation, activates mast cells
- IL 5 simulates B cell growth, activates eosinophils
- IL 9 T cell growth factor
- IL 10 inhibits Th1 and macrophages
- IL 13 stimulates B cell growth, inhibits macrophages -
IMMUNOGLOBULINS
NAME EM
SIZE EM
SOURCE EM -
IgM
- primary response
- pentamere
- spleen and lymph nodes
IgG
- major IgG, class switch to in secondary response
- monomere, smallest tf most mobile
- spleen and lymph nodes
IgA
- epithelial surfaces tf mucosal immunity
- dimer via a jazzy J chain
- intestinal and respiratory surfaces
IgE
- parasites, allergies
- monomere
- intestinal and respiratory tracts
IgD
- not in horses or rabbits
- monomere
- spleen and lymph nodes