13- Effector responses: antibody and cell mediated immunity

Once B cells and T cells are activated, how do they kill pathogens and cancer cells?

They constantly patrol and target an immune response.

Learning Objectives

  1. Explain how humoral immune responses protect us against pathogens, toxins, or malignancy.

  2. For antibody responses, recognize how the multiple immunoglobulin classes and subclasses can have common, but also distinct, mechanisms for antigen inactivation, elimination, tissue distribution, and other biological activities.

  3. Describe the multistep process required for inducing the differentiation of TC precursors into effector cytotoxic T lymphocytes (CTLs).

  4. Compare and contrast the mechanisms by which CTLs and natural killer (NK) cells recognize and kill infected or tumor target cells.

  5. Explain how NK cells and NKT cells each have some properties of innate immune cells and adaptive immune cells.

Key Terms

  1. Humoral Immunity: antibodies target extracellular pathogens using class dependent effector functions

  2. Cell-mediated Immunity: cells target intracellular and extracellular pathogens as well as abnormal cells using apoptosis and degranulation

  3. Innate cells: NKT, NT, neutrophils, macrophages, eosinophiles

  4. FcR: Receptors on the surface of cells that recognize the constant fragments of antibodies

  5. neutralization: inactivates pathogens and prevents it binding to target cell

  6. agglutination: crosslinks multiple pathogens because antibodies are multivalent

  7. opsonization: marks pathogen for macrophage phagocytosis; macrophage contains Fc receptor that will bind to constant fragment of antibody

  8. complement cascade activation: lysis of bacteria if MAC becomes activated or opsonization

  9. Antibody cell-mediated cytotoxicity: NK cells bind to peptides and phagocytose

  10. degranulation and mediator release: granulocytes (esp. eosinophiles and mast cells) binds to antibody, releases granules and kills pathogen

  11. Natural antibodies: B1B T cell independent antigen nonconventional cells

  12. Transcytosis: transcellular transport of antibodies across interior of a cell captured by vesicles on one side and ejected on other side

  13. ITAM: immunoreceptor tyrosine activation motif; positive signaling

  14. ITIM: immunoreceptor tyrosine inhibition motif; negative signaling

  15. Myeloid lineage: granulocytes and mono, macro and baso/mast and dendritic cells (innate- BM maturation)

  16. Lymphoid lineage: lymphocytes (B and T) dendritic cells, and ILC (NK, NKT) (adaptive-lymphoid organ maturation)

  17. PolyIgR: polymeric immunoglobulin receptor is expressed by epithelial cells that initiate transcytosis of IgA and IgM from blood to lumen of tissues

  18. Neonatal FcR: transports IgG into fetus, from placenta, blood and into mucosal secretions

  19. Monoclonal Antibodies mAb: mouse derived antibodies

  20. Chimeric antibodies: mouse variable regions, human constant regions

  21. Humanized antibodies: CDRs are mouse derived

  22. Human antibodies: entirely human regions

  23. Rituximab: binds to CD20 (B-cell specific antigen); destroys B cells through complement MAC or CDCC activation

  24. Perforin: protein that creates pores in the cell membrane to allow granzyme inside

  25. Granzyme: serine protease induces apoptosis by activating Caspase signaling pathway

  26. Fas-FasL Pathway: This pathway involves the interaction between Fas, a death receptor on target cells, and Fas ligand (FasL) expressed on activated T cells, leading to apoptosis of the target cell.

  27. FADD: Fast activating Death domain that triggers the downstream signaling cascade necessary for apoptosis when Fas is engaged.

Notes

Effector responses target pathogens and abnormal cells through two immunity responses:

  1. Humoral (antibodies)

    1. EXTRACELLULAR PATHOGENS

    2. CLASS DEPENDENT ON Ig GAMED

    3. Function: isotype dependent

  2. Cell-mediated (Th and CD8+)

    1. INTRACELLULAR , EXTRACELLULAR AND ABNORMAL CELLS

    2. Helper T cells, CD8, NK, NKT, and Innate cells (Neutrophils, macrophages, dendritic cells, and eosinophils and monocytes)

    3. Function: apoptosis or degranulation

  3. FCR (Constant fragment (region) of antibodies- receptors) link both immunity responses by optimizing all cells that act as markers for phagocytosis

Effector functions of antibodies

  1. neutralization: inactivates pathogens and prevents it binding to target cell

  2. agglutination: crosslinks multiple pathogens because antibodies are multivalent

  3. opsonization: marks pathogen for macrophage phagocytosis; macrophage contains Fc receptor that will bind to constant fragment of antibody

  4. complement cascade activation: lysis of bacteria if MAC becomes activated or opsonization

  5. Antibody cell-mediated cytotoxicity: NK cells bind to peptides and phagocytose

  6. degranulation and mediator release: granulocytes (esp. eosinophiles and mast cells) binds to antibody, releases granules and kills pathogen

Each class of immunoglobulin will tailor specific pathogens.

Immunoglobulin

Function

Notes

IgM

complement activation and agglutination (opsonization and phagocytosis)

pentameric molecule; low affinity antibodies (naive cells), high avidity (compensates for low affinity)

IgG

neutralization and complement activation, opsonization and ADCC

most common; multiple subclasses 1-4

IgA

NOT pro-inflammatory (no complement activation); neutralization in secretion and opsonization in serum

dimer joined by J chain; two subclasses in serum and secretions, important in MALT (skin and mucosa surfaces) , help tolerate nonpathogenic bacteria good in gut

IgE

degranulation of eosinophils; releases histamine

allergy and asthma; parasitic multicellular pathogens; low in serum;

IdD

degranulation

terrible at everything; low in serum; in upper respiratory tract

FcR Receptors- Antibody sensors

  • recognize constant fragments of antibodies

  • mediate effector functions of antibodies

  • link humoral and cell-mediated immune responses

  • allows nonspecific (innate) immune cells to take advantage of antigen specific (adaptive) antibodies

  • immunoglobulin family domain

    • Majority: Positive ITAMS

    • Minority: Negative ITIMS (Fc gamma RIIB)

  • Differentiate by

    • Antibody class binding

    • Cell expression

    • Responses triggered

  • Multiple FcRs must be cross linked to initiate signal

  • Most diverse is Fc gamma because IgG IS MOST DIVERSE (1-4 SUBCLASSES)

    • Fcgamma : IgG (1-4)

    • Fcalpha : IgA

    • Fcmu: IgM

    • Fcepsilon: IgE (high and low)

    • Fcdelta: IgD

  • PolyIgR: polymeric immunoglobulin receptor is expressed by epithelial cells that initiate transcytosis of IgA and IgM from blood to lumen of tissues

    • carries Ab into tears and milk and gut mucosa with IgA and IgM

  • FcRn: neonatal Fc Receptor

    • transports IgG from placenta into fetus, milk in intestine into blood and into mucosal secretions

    • epithelial and endothelial cells

Making Connections

  • B cells encounter Ag, bind and become activated

  • Two types of B cells:

    • T independent : B1B and Marginal zone; TI-1 and TI-2

    • T dependent: B2B; Need Helper T cells to differentiate into different isotypes based on cytokines

      • Differentiate into:

        Phase 1:

        • GC- independent memory B cell: low affinity

        • Short Lived plasma cell (sLPC): low affinity

        • GC- dependent memory B cell

        Phase 2:

        • GC dependent memory B cells undergo SHM and class switching to become higher affinity

          • GC-dependent memory cells

          • Long lived plasma cells lLPC

            • AID: enzyme induces SHM

            • Dark to light for affinity selection testing on fDC

            • centroblasts → centrocytes

  • B cells become Ag secreting plasma cells

    • intracellular pathogens: class switch to isotypes that activate ADCC

    • extracellular pathogens: class switch to IgG subclasses

    • AB is carried to mult. sites for isotype specific effector functions that is sensed by FcRs on target cells

Therapeutic uses of Antibodies

We can manufacture antibodies with the specific protein/antigen and inject the antigen into a lab animal and wait for the organism to develop antibodies for that specific protein/antigen and harvest their B cells to culture in the lab.

However, we can’t inject ourselves with mouse/pig/horse/animal antibodies because our bodies will detect them as foreign OR detect ourselves as foreign and develop an immune response against it. SO, we can engineer modifications to make them look like human antibodies.

Monoclonal antibodies mAb: antibodies of mouse origin make up all therapeutic antibodies that end in -mab.

Three types:

  1. Chimeric: V (mouse) C (Human)

  2. Humanized: CDRs (mouse)

  3. Human: entirely human

Therapies:

  1. Antagonism- blocks ligand or receptor from binding; no activation

  2. Agonistic: antibody mimics ligand or receptor to activate pathway

  3. Complement activation: promotes lysis of target cell by activating the complement

  4. ADCC: antibody binds cancerous cells and T cells kill that cell

Examples:

Rituximab: binds to CD20 (B-cell specific antigen); destroys B cells through complement MAC or CDCC activation (marks them for destruction)

  • B cell cancer; Non-Hodgkin’s lymphoma

  • B cells replenished by bone marrow and hematopoietic stem cells

Cell Mediated Effector Responses

  • Cytotoxic effector cells: Adaptive, Branch and Innate Immunity arms

  • Apoptosis kills infected cells and abnormal cells

  • CTLs and NK involved in organ rejection

CD8+

  • CD8+ recognize infected and tumor cells by TCR activation on the cell surface of the T cell by binding MHC I peptide complex on target cell, local IL-2 cytokine signaling and costimulatory signals from CD28-CD80/86 interaction between pAPC and T cell

  • The CD8+ cells are activated by pAPC that have been licensed by CD4+ cells for crosspresentation

  • How do these cells kill target cells? They secrete cytotoxic granules containing perforin and granzyme inducing Caspase signaling pathway and Fas-FasL pathway

  • Fas-FasL pathway: Fas ligand on CD8+ cell membrane binds to Fas receptor on target cell, activates fast activating death domain (FADD) and cleaves Procaspase into caspase, and active Caspase 8 will trigger apoptosis of target cells.

NK- Natural Killer Cells

  • Innate lymphoid cell

  • Lack Ag specific receptors- are the first wave of immunity before CD8+ adaptive cells to respond to infected cells

  • Activated by IFN-alpha, beta and IL-2

  • Recognize inhibitory and activating receptors on target cells through two models: Missing MHCI self and Balance signal model

  • Viruses decrease expression of MHC I to try to evade CD8+ Tc cells, however, NK cells recognize the lack of self

  • Must be licensed and tested

  • NK cell memory exists despite it being innate cell

NKT- Natural Killer T cell

  • Bridge

  • Share characteristics of both

  • conserved TCR

  • Fas-FasL

  • no memory

Questions

  1. Match each antibody function to the correct definition:

    1. Neutralization: most effective mode of protection from toxins and viruses

    2. Agglutination: binds and crosslinks multiple antigens

    3. Opsonization: enhances the engulfment of antigens by phagocytes

    4. Complement activation: results in formation of the MAC cascade

    5. ADCC: activates the killing activity of cytotoxic cells including NK cells

  2. Match each description with correct antibody isotype:

    1. IgA: found in secretions; effective at neutralizing toxins and pathogens

    2. IgM: first antibody produced in primary response'; pentavalent

    3. IgG: includes several subclasses, some of which are particularly good at complement fixation

    4. IgD: promotes basophil and mast cell release of AMPs

    5. IgE: best known for roles in allergy, asthma, and parasitic worm infection

  3. What are the two types of immunity? Humoral and cell mediated

  4. Which immunity type targets extracellular pathogens only? humoral

  5. Which immunity type targets pathogens AND abnormal cells? cell-mediated

  6. What effector functions does cell-mediated immunity perform? degranulation and apoptosis

  7. What effector functions does humoral immunity perform? class-dependent because they’re antibodies Ig GAMED

  8. Cell-mediated immunity is mediated with the help of which adaptive cells? Helper T cells (CD4+) and CD8+ cytotoxic t cells

  9. Cell-mediated immunity is mediated by which innate cells? NKT, NK, and neutrophils, macrophages, and eosinophiles

  10. How does cell-mediated and humoral immunity interact with each other? FcRs: Constant fragment (region) of antibodies- receptors

  11. How do FcRs work? How does it link both immunity responses? Antibodies will opsonize a pathogen and the FcRs on the innate/adaptive cells will recognize the constant region of the antibodies and that will facilitate the phagocytosis/degranulation of the pathogen

  12. How many effector functions of antibody mediated immune responses are there? What are they? 6; neutralization, agglutination, opsonization, complement activation ADCC, degranulation

  13. Match the antibody with its effector function (from the 6):

    1. IgG: neutralization, MAC, opsonization, ADCC

    2. IgA: neutralization, opsonization, ADCC, degranulation

    3. IgM: agglutination, MAC

    4. IgE: degranulation

    5. IgD: degranulation

  14. What is the relationship between avidity and affinity? inverse; compensates for low affinity

  15. How does class switching in antibodies work to change isotypes? depends on the cytokines released from helper T cells

  16. What type of B cells bear IgM isotype? B1B T cell independent antigen cells (nonconventional)

  17. when are IgM antibodies released in B cell activation? Phase 1; germinal center independent

  18. Which antibodies are produced first? IgM

  19. The majority of IgM in circulation are called? natural antibodies; B1B cells

  20. Which functions are performed by IgM antibodies? complement (MAC) activation and agglutination

  21. Where are isotypes determined on the antibody structure? Heavy chain constant region

  22. Which isotype is “terrible”? IgD

  23. Which isotype is confined to the upper respiratory tract? IgD

  24. Which isotype is pentameric? IgM

  25. Which isotype is responsible for tolerating nonpathogenic bacteria in the gut? IgA

  26. Which isotype is a dimer joined by J chain? IgA

  27. Which isotypes have subclasses? IgA (1 and 2) & IgG (1-4)

  28. Which isotypes have only 1 effector function? Which function is it? IgE and IgD; degranulation

  29. Which isotype is found primarily on mucosal surfaces of barriers? IgA

  30. Which isotype is known for role in allergies/asthma? IgE

  31. Which isotype targets parasitic worms? IgE

  32. Which isotype is most common in serum? IgG

  33. Which isotype has the longest half-life? IgG

  34. Which isotype is the heaviest? IgM (pentameric) and IgA (dimer)

  35. Which isotype is the best at complement MAC activation? IgG

  36. Which isotype crosses placenta? IgG

  37. Which isotype is secreted? IgA

  38. Which cells secrete IgA? Plasma cells

  39. What are the Fc Receptors on epithelial cells for IgA called? poly-Ig receptor

  40. What is transport of an antibody from one domain into another called? Transcytosis

  41. To which superfamily do Fc receptors belong to? Immunoglobulin domain family

  42. The majority of FcR are part of which sub-family? R

  43. The majority of FcR are (inhibitory/activating)? activating

  44. Which FcR is the only inhibitory receptor? Why? Fc gamma RIIB- has ITIM

  45. Which FcR is the most diverse? Fc gamma R

  46. Which cells activate phagocytosis? macrophages

  47. Which cells induce degranulation? granulocytes (Neutrophils, Eosinophils, Basophils, Mast cells)

  48. Which cells activate ADCC (antibody dependent cell-mediated cytotoxicity)? NK cells

  49. Fc gamma RIIB is an inhibitory FcR. How does it inhibit? blocks B cell activation and proliferation

  50. When is Fc gamma RIIB expressed? when there are enough antibodies in circulation as a negative feedback regulatory mechanism

  51. Which Fc epsilon R is high affinity? FceRI (one)

  52. Which Fc epsilon R is low affinity? FceRII (two)

  53. Which Fc epsilon R has unknown function? FceRII

  54. Fc epsilon R triggers what effector function? Histamine, protease, inflammatory molecule release of IgE cells

  55. Fc mu R are expressed by which cells and perform which function? B/T and NK cells; regulate cell activation

  56. Fca/Fcm receptors are expressed on which cells? What function do they mediate? B, macrophages and fDC; binds both with high and low affinity to induce phagocytosis in IgM

  57. Which FcR is responsible for carrying antibodies from the blood into the lumen of tissues? Polygenic Immunoglobulin Receptor

  58. Why is poly-Ig receptor important? It carries antibodies from blood into lumen of tissues for tear, milk and gut mucosa secretion of IgA and IgM

  59. Which isotypes are transcytosed through mediation of poly-Ig receptor? IgM and IgA

  60. Which FcR is responsible for transporting IgG into the fetus? FcRn neonatal receptor

  61. What are the antibodies for therapeutical uses called? Monoclonal antibodies mAb

  62. What are the three types of mAB? chimeric, humanized and human

  63. What are the 4 types of therapies using antibodies? antagonism, agonism, complement MAC activation, and ADCC

  64. What does Rituximab target? B cell antigen CD20

  65. cell mediated effector responses include which 3 “arms”? adaptive, innate and bridge

  66. Which cells exhibit properties of both adaptive and innate responses in cell-mediated immunity? NKT

  67. Which adaptive cell carries out cell mediated effector responses more commonly? CD8+ Cytotoxic T cells

  68. Which cells are involved in organ/graft rejection? CTLs (CD8+) and NK

  69. How do CD8+ respond to organ rejection? tissue not matched to the host MHC is killed by CD8+ cytotoxic granules containing perforin and granzyme

  70. How do CD8+ cells recognize target cells? TCR activation (CD8 is a cytotoxic cell) by MHC1-peptide complex

  71. Which MHC does a CD8+ recognize? MHC I presenting self antigens

  72. How are dendrites licenced? Why do they need to be licensed? Licensed by engaging with CD4+ Th and to gain ability for cross presentation

  73. What are the two models of cross presentation? sequential and simultaneous

  74. What are the two pathways that CD8+ instruct apoptosis? Perforin/Granzyme AND Fas-FasL pathways

  75. Why do we have CD8+ and NK cells if they perform similarly? What are the smiliarites and differences? Both kill infected cells; CD8+ are part of the adaptive immunity and NK are part of innate immunity

  76. Which interferons are related to viral infections? IFN- alpha and Beta

  77. NKs proliferate in response to? IFNa, IFN-B, and IL-12

  78. How do NK cells recognize target cells? Missing self MHCI or balance signal models

  79. Which cells must be licensed to perform functions? DC and NK

  80. True/False: NK cell memory exists True, even though it’s innate cell, there is some memory