The Specific Immune Response....continue...

Overview of the specific (adaptive) immune response - continue 1. Cell Mediated Immunity ( T cell mediated immunty) Key players : T lymphocytes. Two types: Cytotoxic T cells (CTL) or (CD8+) T helpers ( TH) cells (CD4+) - Cytotoxic T cell directly attack and destroy antigen-bearing cells especialy virally infected cells and tumours - Helper T cells act indirectly by secreting proteins called cytokines that activate other cells such as macrophages to destroy the antigen-bearing cells

Cellular immune response by TH cells (CD4+) - Particularly useful in eradicating pathogenic bacteria especially intracellular bacteria Can you give some examples for intracellular bacteria???? Act indirectly by secreting chemical mediators called cytokines that activate other cells such as macrophages to destroy the antigen-bearing cells Activated macrophages can then kill intracellular pathogens that would normally divide in a non-activated macrophages

Mechanism of cellular immune response by TH cells (CD4+) 1st step : foreign antigen will be captured and engulfed by the phagocytes (macrophages) and another cell type called dendritic cells at the site of infection (internalization) - What are dendritic cells?

Dendritic Cells Named so because they resemble dendrites of neurons THEY ARE NOT NEURONS!!! Their main function is to capture , concentrate and present antigens to lymphocytes (APC) Origin : stem cells in bone marrow Several Type Langerhans (LC) found in skin Circulating DCs Myeloid (MDC1 and MDC2) Plasmacytoid Interstitial DCs Heart, lungs, liver, intestines Interdigitating DCs, T-cell areas of lymph nodes and Thymic medulla

Mechanism of cellular immune response by TH cells (CD4+) Next , internalized antigen is processed inside the macrophages and dendritic cells where the antigen is degraded and fragment of it binds to MHC class II molecule (Major Histocompatibility Class II molecule(

Major Histocompatibility complex proteins are found on the surface of cells:: T cells cannot recognize foreign antigens unless they are associated with these MHC proteins Class II MHC proteins are only found on the surface of B lymphocytes, macrophages and other antigen presenting cells Class I MHC proteins are found on the surface of ALL nucleated cells ALL MHC proteins are imbedded in the cytoplasmic membrane of cells and project outward from the cell surface

Mechanism of cellular immune response by TH cells (CD4+) THEN , the processed antigens bind to Class II (Ag-MHC class II complex ) are transported to the cell surface where they expressed. - The macrophages and dendritic cells now move toward regional lymph nodes under the influence of certain chemical substances (chemotaxis)

Mechanism of cellular immune response by TH cells (CD4+) In the regional lymph nodes the phagocytes and dendritic cells present the antigen in association with MHC class II molecule to CD4+lymphocytes. That is why macrophages and dendritic cells are called antigen presenting cells (APC)

Class II MHC proteins and helper T cells (TH) The Class II proteins and antigen are expressed on B cells, APCs and macrophages 1. The APC takes up an external foreign protein via phagocytosis or endocytosis 2. Class II proteins are produced in the endoplasmic reticulum and assembled with a blocking protein (Ii) or invarient chain 3. The Class II proteins enter the phagolysosome where the Ii is degraded and the partially processed antigen binds to the class II molecule 4. The complex is translocated to the surface of the APC where it interacts with the TCR of a T helper cell

Mechanism of cellular immune response by TH cells (CD4+) The part of the CD4+ that comes in contact with the antigen - MHC class II complex is called TCR (T Cell Receptor). Cell- cell interaction mediated by TCR (from CD4+ T lymphocytes) and antigen - MHC class II complex (from macrophages or dendritic cells) will activate TH CD4 + to produce chemical mediators called cytokines (hormones of the immune system : Interferon - gamma (IFN- gamma( Tumour necrosis factor - alpha (TNF-alpha) Granulocyte monocyte- colony stimulating factor (GM-SF)

Mechanism of cellular immune response by TH cells (CD4+) These cytokines further stimulate macrophages to increase phagocytic activity and to in turn produce cytokines that promote inflammation

Class II MHC proteins and helper T cells (TH) Specialized TH cell involved in the inflammatory response Cell-cell interaction mediated by the TCR and the class II MHC-antigen complex activates The TH cell which produces cytokines TNF-alpha (tumor necrosis factor) IFN-gamma (interferon) GM-CSF (granulocyte-monocyte colony stimulating factor) These cytokines further stimulate macrophages to increase phagocytic activity and to in turn produce cytokines that promote inflammation

Types of Specific (adaptive) immunity Humoral immunity Cellular immunity

Specific immune response - humoral immunity B cell mediated immunity through the production of antibodies. Particularly effective against pathogens such as viruses and extracellular bacteria in the blood or lymph and also against soluble pathogen products such toxins Antibodies B - lymphocytes Humoral immunity = +

Humoral immunity: B- Lymphocytes Origin and Maturation: Bone marrow - B- lymphocytes from the bone marrow are released into circulation in a resting state and they do not secrete antibodies - Instead, resting B-lymphocytes display membrane bound antibodies (immunoglobulins) usually in the form of mIgD or mIgM - After activation by antigen, B- lymphocyte divides (clonal expansion( .Some differentiated into plasma cells which secrete antibodies, die within 1- 2 weeks. . Some change into memory cells- display same membrane bound antibodies as parent cell.

Phases of B-lymphocyte activation Antigen

Mechanism of humoral immune response by B - lymphocytes Resting B - lymphocyte is coated with membrane bound antibodies or immunoglobulin (mIg) on the surface of the lymphocytes The first step in the initiation of the humoral immune response is the binding of the antigen to the mIg mIg Resting B - lymphocyte B - lymphocyte + Antigens

Mechanism of humoral immune response by B - lymphocytes The mIg- antigen complex is then endocytosed and complexed with MHC class II molecule and then surface expressed Here , B - lymphocyte acts as APC where it presents the antigen-MHC class II complex to TH cells Now, TH cells start to secrete cytokines ( IL-4 and IL-5) that stimulate B-lymphocyte to divide (clonal expansion) and differentiate into plasma cells (1 B cell --> 4,000 Ab-secreting cells --> ~1012 antibody molecules/hour)

Mechanism of humoral immune response by B - lymphocytes Plasma cells start to secrete antibodies (short half life, die in 1-2 weeks). Some dividing B- lymphocytes change into memory cells where they display same mIg as parent B- cell and change rapidly into plasma cells when encountering same antigen for second time (secondary immune response. Primary immune response is usually mediated by IgM while the secondary immune response is stronger and mediated by IgG. Note : In secondary immune response , memory cells convert immediately to plasma cells and produce IgG in high amounts without the aid of helper T cells

B cells are coated with antibodies that react with specific antigens When the antigen binds to the antibody, the B cell first acts as an APC. The bound antigen is endo cytosed and complexed with MHC II and then surface expressed Class II MHC proteins, helper T cells that stimulate antibody producing cells—the B cells The surface expressed complex interacts with and activates TH cells that produce the cytokines interleukin 4 & 5 IL4 and 5 stimulates the B cells to produce identical memory B cells and antibody secreting plasma cells that secrete the same antibody

Secific immune response-Summary

Antibody (Immunoglobulin) Structure 5 classes: IgG, IgM, IgA, IgD and IgE Common structure , four polypeptide chains: - Two identical heavy (H) chains, each carrying covalently attached oligosaccharide groups (50-70 kDa) - Two identical, non-glycosylated light (L) chains (23kDa) Within the immunoglobulin, disulphide bonds join together: - Two heavy chains - Heavy chains to the light chains The disulphide bonds joining the antibody heavy chains are located in a flexible region of the heavy chain known as the hinge region. Heavy chain determines the Ig class: IgG : gamma HC IgA: alpha HC IgD: delta HC IgM: mu HC IgE:epsilon HC Light chain either kappa or lambda irrespetive of Ig class

Antibody (Immunoglobulin) Structure Based on variability of amino acid sequences, both H and L chains can be divided into: - VH and CH domains (variable and constant) VL and CL domains (variable and constant) The variable domains are attached to the constant domains. As the name implies, the variable domains vary in their amino acid sequence from one antibody molecule to another, providing the vast diversity the immune system needs to fight foreign invaders. The antigen binding site is formed where a heavy chain variable domain (VH) and a light chain variable domain (VL) come close together. These parts show the biggest difference among different antibodies. Antigen binding site

Proteolytic treatment of Ig with protease enzymes When the immunoglobulin is treated with proteolytic enzymes (proteases), such as pepsin or papain, it is broken at the hinge region into two fragments known as Fab (Fragment for antigen binding) and Fc (Fragment Crystalizable( The immunoglobulin specificty is determined by the Fab fragment, as well as its capability to react with the antigen. Fc) cannot bind with antigens, but is responsible for biological effector functions like complement fixation, binding to macrophages, natural killer cells and neutrophils.

Pentamer Dimer Monomer Secretions( tears, milk), blood, lymph IgG IgM IgA IgD IgE Structure Monomer Pentamer Dimer Serum % 80% 5-10% 10-15% 0.2% 0.002% Location Blood,lymph,intestine Blood,lymph,B cells as monomer Secretions( tears, milk), blood, lymph Blood, lymph, B cells Mast cells , basophils, blood Placenta transfer Yes No Complement fixation Function Neutralize viruses and toxins, enhance phagocytosis, protect fetus 1ry immune response Localized protection on mucous surfaces Serum function not known,initiation of immune response on B cells Allergic reaction and lysis of parasitic worms

Antibody (Immunoglobulin) functions 1. mIgs activate B- lymphocytes when comes in contact with antigen 2. Secreted Ig neutralizes the effect of viruses , extracellular bacteria and toxins

Antibody (Immunoglobulin) functions 3. Opsonization: bind pathogens for recognition by other immune cells (e.g. phagocytes) Opsonins- are the tagging proteins that make unrecognizable particles into “food” for phagocytes.

Antibody (Immunoglobulin) functions 4. Mast cell degranulation: Mast cells contain histamine in intracellular granules Binding of IgE to cell surface receptors on a mast cell primes the cell to respond to allergen Introduction of allergen and its subsequent binding to IgE stimulates the mast cell to degranulate and release of histamine Mast cell Mast cell Mast cell + IgE Antigens degranulation

Antibody (Immunoglobulin) functions 5. Antibody dependent-cellular cytotoxicity (ADCC) - Classically mediated by NK, but also by eosinophils and neutrophils - Part of the adaptive immune response (depend on antibodies)

Antibody (Immunoglobulin) functions 6. Complement activation Will be discussed in details in next lecture

Monoclonal Vs polyclonal antibodies B-lymphocyte Polyclonal antibody Multiple clones from multiple B - lymphocytes each of which recognizes different epitope on same antigen Antigen B-lymphocyte B-lymphocyte

Monoclonal Vs polyclonal antibodies Antigen Monoclonal antibody Single clone from single B - lymphocyte recognizes single specific epitope on antigen B-lymphocyte