Week 1 III

What are the 3 different types of granulocytes and what are their important features? What are their common features and how are they named?

* All possess cytoplasmic granules and a lobed nucleus * The granulocytes are named according to the ability of the granules to react with acid or basic dyes 1. Neutrophils - neutral granules - most numerous - important in anti-bacterial immunity 2. Eosinophils - granules stain red with the acid dye eosin - important in anti-parasite immunity 3. Basophils - stain with basic dyes - contain vasoactive amines - function ? important in allergic reactions

What are antibodies? What are they also known as? Where are they made and found? What do they bind to?

* Functional proteins - also known as Immunoglobulin - made by specialised B lymphocytes - found in body fluids - exposure to Ag induces the production of antibodies - antibodies bind to antigens • Y-shaped molecules - the fingertips are where the antigen binds and the trunk is where other things bind (e.g. complement proteins)

Eosinophil: Half-life Amount of time spent in tissues Function Defence mechanism Draw a picture

* Half life on the circulation is 30 min * They spend about 12 days in the tissues * Phagocytic function similar to neutrophils (only weakly phagocytic) * More suited to extracellular destruction of large parasites • Often look orange/pink on a stain * Important defense against parasitic and fungal infections

Basophil: What the granules contain Function

* Less common than the other granulocytes * The basophilic granules contain vasoactive amines such as histamine and serotonin * Function less well known May be involved in hypersensitivity and allergy

What are Natural Killer (NK) cells and what type of immunity are they involved in? What do they look like?

* Non T/B cytotoxic lymphocytes • Innate immunity • Not hugely abundant in blood • Not associated in adaptive immunity - associated with innate immunity • Can lyse and kill cancer cells and virus-infected body cells before adaptive immune system is activated • Part of large granular lymphocytes • Less picky than lymphocytes of the adaptive system - can eliminate a variety of cells by detecting lack of 'self' cell surface receptors and by recognizing certain surface sugars on the target cells • Not phagocytic - directly contact and induce cell to undergo apoptosis - uses same killing mechanisms used by cytotoxic T cells • Secrete potent chemicals that enhance the inflammatory response

What are antigens and what are some examples of them?

* Surface markers on cells called antigens, protein, sugars, lipids, CHO, nucleic acids * body can recognise these as self or non-self (foreign) * any molecule that can bind specifically to an Ab or generate peptide fragments that are recognised by a T cell receptor (TCR) - a molecule that can cause an immune response Examples: • Microbial antigens - toxins, cell wall proteins, flagella, enzymes • Blood group antigens (red cells), histocompatibility antigens - important for transplants ϖ Can have them on our own cells - can cause autoimmune disease • Can mobilise the immune system and provoke an immune response • A single antigen may stimulate the production of many antibodies and the activation of different types of lymphocytes

Neutrophils: Amount of time spent in bloodstream Life span Major function Defence mechanism Draw a picture of one

* They spend approximately 12 hours in the blood stream * Total life span is a few days - - They perform their function and then die - have granules that are quite toxic - Become phagocytic on encountering infectious material in the tissues * Major function - capture & destruction of foreign material (phagocytosis) * The respiratory burst is an important antibacterial defense

What are mast cells and what is their role? What do they look like?

* Tissue cells - rarely seen in blood • Interface between innate and adaptive immunity • Role in infection • Role in diseases e.g. allergy • Granules can contain histamine and serotonin • Involved in sensing invaders

What are some examples of different isotopes of antibodies?

* isotypes have different characteristics and different functions e.g. IgM, IgG (1-4), IgA, IgE (associated with allergy and parasite/worm infections - will have high levels of this in these circumstances), IgD (very rare)

What are the functions of IL-1, IL-6 and TNF-α both locally and systemically?

Il-1β o Local: Activates vascular endothelium, activates lymphocytes, local tissue destruction, increased access of effector cells o Systemic: Fever, production IL-6 TNF-α o Local: activates vascular endothelium increased vascular permeability = increased entry IgG, complement and cells to tissues and increased fluid drainage to lymph nodes o Systemic: fever, shock, mobilization metabolites IL-6 o Local: lymphocyte activation, increased antibody production o Systemic: fever, acute phase protein production

How do cytokines interact?

In an autocrine, paracrine or endocrine manner

Where are B cells developed and what percentage of pre-B cells are destroyed? Where do the B cells end up and which is the first Ab to be produced in a humeral response?

In humans the bone marrow serves as the primary lymphoid organ for B cells - Develop from lymphoid progenitor cell - Originate in the red bone marrow from hematopoietic stem cells • Up to 75% of pre-B cells generated are destroyed, possibly by macrophages - B cells that successfully make antigen receptors are positively selected and those whose antigen receptors are self-reactive are eliminated • B cells leave the BM, circulate in blood and end up in secondary lymphoid organs (LN, spleen, MALT e.g. Peyer's patches)) only allowed to leave the BM once they are developed • Naïve B cells are activated in these organs by antigen and interactions with Helper T cells • First Ab to be produced in a humoral response is IgM • activated B cells undergo subsequent isotype or class switching and secrete Ab of different isotypes • Some will become Ab producing plasma cells, some will become memory B cells • Lymph nodes have areas where the B cells and the T cells live • Internally are the macrophages in lymph node • Spleen also has T cell and B cell areas - only when they are activated do they come together and interact • T cells and B cells become immunocompetent in primary lymphoid organs - When they do this, they display a unique type of receptor on their surface which enable the lymphocyte to recognize and bind a specific antigen - Receptors on B cells are membrane-bound antibodies • When an antigen binds to a lymphocyte that has a receptor for it, the antigen selects that lymphocyte for further development • Once activated, the lymphocyte rapidly proliferates to form an army of cells all exactly like itself and bearing the same antigen-specific receptors (a clone) - Some members of the clone become effector cells and some become memory cells • Immunocompetent but naïve B lymphocyte is activated when matching antigens bind to its surface receptors and cross-link adjacent receptors together - Called clonal selection - Followed by proliferation and differentiation into effector cells

Define Inflammation. What does it involve?

Inflammation = complex biological response of body tissues to harmful stimuli, pathogens, damaged cells, or irritants • leads to increase blood flow and increased vascular permeability • protective response involving immune cells, blood vessels, and molecular mediators. • Immune cells also important for repairing the tissue (e.g. macrophages stimulating fibroblasts that produce molecules to repair the tissue) • Cytokines can affect the whole body - e.g. the brain, causing sickness behavior, they can also trigger BM to start releasing more immune cells required to fight the infection - Can cause the liver to create acute phase proteins (e.g. CRP) - its action is induced by the action of cytokines • DC's and macrophages can present the antigen and activate the adaptive immune response

What are some words used to describe innate immune responses?

Innate immune responses = Natural / Native / Non-specific

What is Interferon (IFN) Gamma secreted by and what are its functions?

Interferon (IFN) Gamma • Secreted by lymphocytes • Activates macrophages, stimulates synthesis and expression of more class I and class II MHC proteins and promotes differentiation of TH cells into TH1 • Cells infected by viruses can secrete interferons to protect cells that haven't been infected - stimulate synthesis of proteins which interfere with viral replication in health cells by blocking protein synthesis and degrading viral RNA

Describe how lymph flows through a lymph node...

Lymph Flow • Lymph enters convex side of lymph node (through afferent lymphatic vessels), then moves through a sinus (subscapular sinus), into smaller sinus (cut through cortex and enter medulla) - Carries phagocytic DC's • Lymph exits at hilum (on concave side) via efferent lymphatic vessels • Less efferent than afferent vessels - lymph stagnates - allows time for lymphocytes and macrophages to carry out their role • Naive lymphocytes enter the node from the bloodstream through specialized postcapillary venules (not shown) and leave with the lymph through the efferent lymphatic.

What does MHC stand for and what is it also known as? How many classes of molecules are there of MHC?

MHC = Major Histocompatibility Complex * In human the MHC is also known as the HLA (human leucocyte antigens) • There are two classes of MHC molecules with distinct subunit compositions but similar three-dimensional structures • MHC class I (CD8 T cell recognises) - found virtually on all body cells and class II (CD4 T cell recognizes) - found only on certain cells that act in the immune response- recognized by different T cells • Present peptide • Self-antigens, but are strongly antigenic to other people - Millions of combinations of these genes - Group of glycoproteins

Define the following properties of Antigens: a) Immunogenic b) Allergenicity c) Tolerogenicity

NOTE: All molecules that are immunogenic are also antigenic (these words are often used synonymously) a) Immunogenic = any molecule that on its own is able to elicit an adaptive immune response on injection in a person or animal - only certain parts of an antigen are immunogenic - free antibodies or lymphocytes bind to this site b) Allergenicity - capacity to induce allergy c) Tolerogenicity - capable of inducing immunological tolerance (immune system does not attack Ag)

What are some Neisseria Gonorrhoeae Infection outcomes in those suffering from complement deficiencies?

Neisseria Gonorrhoeae Infection Outcomes in those suffering from complement deficiencies • Properdin is a complement protein that is X-linked - synthesized by monocytes, granulocytic cells and T-cells. Mutant forms can decrease AP function • Deficiency increases susceptibility to bacterial infections of Neisseria family of organisms • As Neisseria gonorrhoea are extracellular encapsulated bacteria, people with complement deficiencies have higher risk of repeated infection. Dysregulation of complement system destabilises pathways and creates problems in forming MACs.

What are primary lymphoid organs and what are the examples?

Organs in which lymphocytes develop and mature (rearrange the receptor genes) - Bone marrow - where this occurs changes with age (more in pelvis in adults) - where B cells are developed - Thymus (just above the heart) - where T cells are developed - Bursa of Fabricius (birds) where the 'B' came from (not in humans)

What are secondary lymphoid organs and what are some examples?

Organs in which naive lymphocytes encounter antigen and are stimulated to become effector and memory cell populations - Spleen - Lymph nodes - Mucosal associated lymphoid tissue (MALT) Where they sit and wait to be activated by antigens

What are PAMPs and what do they work through to assist in the production of cytokines as part of early responses to infection and inflammation? Give an example of a type of PAMP.

Pathogen associated molecular patterns (PAMPs) on invading microorganisms are recognised by the body - Collective terms for things on a cell that are foreign (e.g. a lipopolysaccharide or a nucleic acid) - all things on a cell (e.g. microorganisms) that are foreign - can be recognized by cells of innate immune system - can be recognized by cells of the innate immune system • PAMPs are recognised by Toll-like receptors (TLRs) found on/in macrophages, DC, mast cells and mucosal epithelial cells (sensors) • Signals from TLR activate genes that are involved in the production of cytokines - triggering of inflammation Type: LPS - Histamines may also be produced which then stimulate inflammation - drives inflammatory cells to the area of infection (e.g. movement of neutrophils), which can then help clear the infection

How are PAMPs and DAMPs recognised? and what does recognition of these PAMPs and DAMPs cause?

Pattern-recognition receptors (PRRs) recognise PAMPs and DAMPs • Activation of PRRs by PAMPs or DAMPs triggers downstream signaling cascades and leads to production of proinflammatory cytokines.

Why are opsonins important?

Phagocytes must adhere to the pathogen and therefore must be able to recognize the pathogens 'carbohydrate signature' however many pathogens have an external capsule that conceals their 'carbohydrate signature' allowing them to avoid capture

What is hematopoiesis?

Process in which the cells of the blood and immune system are made. • Have lymphoid and myeloid cells

Which types of cells express pattern recognition receptors that provide an initial discrimination between self and non-self? How is NOD involved?

Sensor cells (e.g. macrophages, DCs) Macrophages, dendritic cells (and mast cells) switch on genes associated with inflammation - Have these on and in them - TLR-4 is a PRR - is able to bind a PAMP (which in this case is LPS, which is present on gram-negative bacteria) - sends a signal into macrophage to activate genes to secrete pro-inflammatory molecules (e.g. cytokines) - IL-1 beta particularly causes fever] - NOD is not on the surface, but sits inside the cell (like a TLR, but part of the MLR family) - really important for triggering the inflammatory cytokines Inflammosome - group of proteins that enhance the action of cytokines - cleave off a pro-peptide to release active cytokines such as IL-1 beta (NOD proteins are part of this inflammasome - can sense things inside the cell) • PRR = pattern recognition receptors - proteins expressed by cells of innate immune system to identify PAMPs (e.g. on macrophages and DC's) - When these join, the cells switch on genes associated with inflammation (e.g. macrophages may produce more cytokines)

Where are TLRs found on/in?

Sensor cells (macrophages, dendritic, mast cells) and mucosal epithelial cells

What are the inflammatory inducers, sensor cells, mediators and target tissues involved in inflammation?

Sensor cells (macrophages, mast cells, DCs) induce an inflammatory response by producing mediators such as chemokines and cytokines

What occurs if the neutrophils and macrophages are unable to kill the pathogen within the phagolysosome

Some pathogens (e.g. tuberculosis bacillus and certain parasites) are resistant to lysozyme enzymes and can even multiply within the phagolysosome - this calls for the assistance of helper T cells Helper T cells release chemicals that stimulate the macrophage and activating additional enzymes that produce a lethal respiratory burst The respiratory burst promotes killing of pathogens by: - Liberating a deluge of highly destructive free radicals (including superoxide) - Producing oxidizing chemicals (H2O2 and a substance identical to household bleach) - Increasing the phagolysosome's pH and osmolarity, which activates other protein-digesting enzymes that digest the invader

What are the features of Class I MHC proteins?

• All antigens on class I MHC proteins are endogenous - fragments of proteins synthesized inside the cell (usually self-antigens), but when cell is infected, the antigens expressed may be foreign • Crucial for activating naïve CD8 cells and informing cytotoxic T cells that infectious microorganisms are hiding in body cells • When displaying self-antigens, the cytotoxic T cells passing by get a signal to leave the cell alone • When they display foreign antigens, they sound a molecular alarm that signals invasion - They act as antigen holders AND form the self part of the self-nonself complexes that cytotoxic T cells have to recognize to kill

What are chemokines and how are they classified?

• Any group of low molecular weight cytokines identified on the basis of their ability to induce chemotaxis or chemokinesis in leucocytes in inflammation • Act as chemoattractants • Mainly classified into 2 groups: 1. The CC chemokines, which have two adjacent cysteine residues near the amino terminus 2. The CXC chemokines, in which the equivalent cysteine residues are separated by a single amino acid

Describe the structure of a lymph node...

• Bean shaped • Less than 2.5cm in length • Each surrounded by a dense fibrous capsule which has CT strands (trabeculae) who divide the node into compartments • Has internal framework (stroma) of reticular fibers • Has cortex (superficial part densely packed with follicles) - many with germinal centers heavy with dividing cells • DC's encapsulate follicles and abut deeper part of cortex - houses T cells in transit • T cells continuously circulate between blood, lymph nodes and lymph • Medullary cords = inward extensions from cortex - contain lymphocyctes and plasma cells • Lymph sinuses are found throughout the node (large lymph capillaries) • Macrophages lie on the reticular fibers • Lymph-borne antigens can leak into lymphoid tissue, activating lymphocytes • Paracortical area: T cells + DCs - combination of antigen presenting cells (dendrites) and naive t cells in the paracortical area is ideal for T cell activation. Active T cells then travel to germinal centre (to help activate B cells) • Medullary cords: strings of macrophages + plasma cells • Cortical sinus • Medullary sinus

Describe how blood flows through the spleen...

• Blood flows into spleen via splenic artery which branches into trabecular arteries - As these emerge from the CT, it is known as the artery of white pulp (central artery) which then continues into a lymphatic nodule as nodular artery which then becomes smaller and terminates in the red pulp - Blood vessels from white pulp (pulp arterioles) continue in red pulp as sheathed capillaries Blood carrying lymphocytes + Ag flows from trabecular artery central arteriole follicular artery white pulp trabecular vein splenic vein

What are the 3 main steps in inflammation and what do they involve?

• Cells can move between tight junctions of endothelial cells

What are cytokines and what are the 3 key ones produced by macrophages and their functions?

• Chemical messengers involved in cellular immunity - mediators that influence cell development, differentiation and responses in the immune system - E.g. IL-1 released by macrophages stimulates T cells to release IL-2 and synthesise more IL-2 receptors - positive feedback loop - T cells can secrete cytokines - some are cell toxins and others are inflammatory molecules - Bind to receptors on cell surface 3 Key cytokines produced by macrophages 1. TNF (tumor necrosis factor) - promotes inflammation, enhances phagocyte chemotaxis and nonspecific killing, slow tumour growth by selectively damaging tumour blood vessels and promotes cell death by apoptosis 2. IL-1 - promotes inflammation and T cell activation, causes fever (acts as a pyrogen that resets thermostat of hypothalamus) 3. IL-6 - active in inflammation and B-cell maturation and pyrogen, interacts with IL-6 receptor-alpha to induce transcription of inflammatory gene products

What are Toll-like receptors and what is the function of TLR-4?

• Class of proteins that play a key role in innate immune system - single and span membrane • Usually expressed in macrophages and DC's that recognize structurally conserved molecules derived from microbes • TLRs recognise molecular patterns characteristic of Gram-negative and Gram-positive bacteria, fungi and viruses. In particular, they recognise the lipoteichoic acids of Gram-positive bacterial cell walls and the lipopolysaccharide (LPS) of the outer membrane of Gram-negative bacteria. • TLRs activate the transcription factors to induce the expression of inflammatory cytokines and type I interferons. • TLR-4 - transmembrane protein that belongs to PRR family - Activation leads to an intracellular signaling pathway NF-KB and inflammatory cytokine production which is responsible for activating the innate immune system - Most well-known for recognizing and binding lipopolysaccharide (LPS), a cell-wall component present in many gram-negative bacteria (such as salmonella)

Why are innate immune responses needed?

These mechanisms are essential for the rapid response against microbes and react essentially in the same way to repeat infection

What are adaptive immune responses and what are its 2 types? What do these types depend on? What are the major components?

These responses are highly evolved and stimulated by exposure to the infectious agent and increase in magnitude • Depending the microbes the type of adaptive immune response could be either humoral (body fluids e.g. Ab, complement) or cell-mediated • The major components are the lymphocytes and their products

Why is inflammation observed in many different disease processes and what are the goals of inflammation?

Why inflammation is observed in many different disease processes: • Inflammation is part of the innate defences - non-specific and occurs for every disease the same - Prevents spread of damaging agents to nearby tissues - Disposes of cell debris and pathogens and sets stage for repair • Goals of inflammation: 1. Eliminate initial cause of tissue injury 2. Eliminate cell debris/ necrotic tissue 3. Initiating repair and an adaptive immune response 4. Stop the spread of pathogens to nearby tissue and stop further damage to the wound.

What are interferons and how do they act?

a family of immune modulating proteins; Diffuse to nearby cells that they stimulate to synthesize proteins that "interfere" with viral replication in still-healthy cells by blocking protein synthesis and degrading viral RNA. provide non virus specific protection

What is chemokine?

a type of cytokines involved in attracting cells (a chemotactic)

What are the 3 pathways of the complement system activated by?

• Classical Pathway - Mediated by 11 proteins (C1-C9) - Requires antigen-antibody interactions - Antibodies bind to invading organisms and subsequent binding of C1 binding to Fc region of an antibody - C1q specifically recognizes and binds to Fc region of antibody - C1r and C1s are inactive proteases • Alternative Pathway - Activated by presence of pathogen alone - initiated by spontaneous hydrolysis and activation of C3 - Triggered when factors B, D and P interact with polysaccharide molecules present on surface of certain microorganisms - Activation of each protein catalyses the activation of the next All pathways converge on C3, cleaving it into C3a and C3b - initiates a common terminal pathway that causes cell lysis, promoted phagocytosis and enhances inflammation • C3b bound to pathogen surface helps phagocytes recognise it • C3a helps recruite phagocytic cells to site of infection (increased inflammation) • Lectin Pathway - Similar to classical pathway - Mannose binding lectin (MBL), the Ficolines and Collectin can initiate the LP - Associated with these are enzymes called MASPs (MBL-Associated Serine Proteases) - they trigger the cleavage of complement proteins and activate the pathway - C2 and C4 also participate in the LP - First to react before adaptive immune response occurs Lectin-type proteins bind to CHO on pathogen surfaces

What are the 6 different types of effector T cell, their main functions in adaptive immune response and the pathogens that they target?

• Cytotoxic - kill cells - CD8 important in killing tumours - can be used in treatment of melanomas • CD4 have different types - helper cells - each has different effector function

Describe in detail the characteristics of adaptive immunity.

• Elite fighting force - attacks particular foreign substances - takes more time to mount than the innate response • Specificity - lymphocyte receptors are able to recognize subtle structural differences between distinct antigens • Diversity - the total number of antigen specificities of lymphocytes in an individual ie lymphocyte repertoire is large (109 to 1011) • Memory - enhances the ability to respond to reinfection - after an initial exposure, it recognizes and mounts even stronger attacks on the previously encountered pathogens • Self Limitation - the immune response wane of with time following the elimination of antigens and homeostasis is maintained at the basal rate. They proliferate to high numbers when stimulated (in a process called clonal expansion) but when they are no longer needed they die • Tolerance - immunological non-responsiveness to self antigens

What are APCs and which are the most potent antigen presenting cells during primary immune response in secondary lymphoid tissues? What are Naive CD4 T cells and Naive CD8 T cells activated by in antigen presentation?

• Engulf antigens and present fragments of them on their membrane where T cells can recognize them • APCs present antigens to the cells will deal with the antigens Professional antigen presenting cells (APCs) = dendritic cells (DC), macrophages and B cells - Unlike DC's and macrophages, B cells don't activate naïve T cells - they present antigens to a helper T cell to get help for their own activation instead • DC are the most potent antigen presenting cells during primary immune response in secondary lymphoid tissues • Naïve CD4 T cells activated by peptide Ag on MHCII • Naïve CD8 T cells activated by peptide Ag on MHCI • APC presents peptide to the T cell Receptor (TCR) through MHC - CD8 and CD4 T cells do completely different things

Outline the time course of a typical infection...

• Entry of organism and replication, then after a period of time, the amount of microorganism decreases (due to immune response) and adaptive immune responses kick in • Innate and adaptive responses fight the infection together • Later on, get memory T cells and B cells forming which can fight the infection more rapidly the next time around.

What is opsonisation and what is it mediated by?

• Facilitation of phagocytosis • Ingestion of complement-tagged pathogens by phagocytes is mediated by receptors for the bound complement proteins • The most important function of complement is to facilitate the uptake and destruction of pathogens (opsonisation) • CR1 (CD35) is the best characterised of complement receptors that bind C3b

What are the different cytokines secreted by macrophages and what are their local effects and/or systemic effects? How can they induce an acute phase response?

• IL-6 can trigger the acute phase response - can leak into circulation and then get into the liver - signals hepatocytes to pump out a number of different acute phase proteins • Cytokines secreted by macrophages and DCs induce a systemic reaction known as the Acute Phase Response

What are the normal blood cell counts for the different types of polymorphonuclear leucocytes and mononuclear cells? (Normal range and percentage)

• Important to recognize the main cells in peripheral blood • Some cells only have raised counts in certain sorts of pathologies - E.g. high neutrophils correspond with bacterial infections - High eosinophils suggest a parasitic/worm/allergic reaction - Lymphocytes - key to fighting viruses - may be high in viral infections - Low lymphocytes may suggest immunodeficiency (e.g. in AIDS - may have a low CD4 count)

What is an inflammasome and what is its role?

• Inflammasome - collection of proteins that activate cytokines by activating caspases o Probe cytokine is attached to the cytokine and causes the cytokine to be inactive → inflammasome cleaves off this probe cytokine and activates the cytokine

What does inflammation result in?

• Inflammation results in: - The recruitment of proteins and cells from the blood into infected tissues that help to directly destroy the pathogen - It increases the flow of lymph carrying microbes and antigen-presenting cells from the infected tissue to nearby lymphoid tissues, where they activate lymphocytes and initiate the adaptive immune response - Once adaptive immunity has been triggered, inflammation also recruits the effector components of the adaptive immune system-antibody molecules and effector T cells-to the site of infection.

What are the two types of adaptive immunity? describe.

o Humoral Response: Provided by antibodies present in the body's "humors," or fluids (blood, lymph, etc.). Antibodies circulate freely in the blood and lymph, where they bind primarily to extracellular targets - bacteria, bacterial toxins, and free viruses - inactivating them temporarily and marking them for destruction by phagocytes or complement. o Cell mediated Response: when lymphocytes themselves rather than antibodies defend the body. Cellular immunity also has cellular targets—virus-infected or parasite-infected tissue cells, cancer cells, and cells of foreign grafts. The lymphocytes act against such targets either directly, by killing the infected cells, or indirectly, by releasing chemicals that enhance the inflammatory response or activate other lymphocytes or macrophages.

What things induce macrophage cytokine production?

o Microbial invasion o Heat o Trauma o Chemicals

What are 5 factors that can influence immunogenicity?

1. Foreignness - a molecule must be recognised as non-self in order to elicit an immune response (tolerance of self antigens) 2. Molecular size - big 100kDa >small <5kDa 3. Chemical composition, charge, etc. 4. Susceptibility of antigen to processing and presentation 5. Individual, route, dose - these can influence the strength of the immune reaction

What are the 4 main components of an innate immune response?

1. Genetic susceptibility 2. Physical & chemical barriers - physical, biochemical & microbial 3. Cells - Phagocytic (neutrophils & macrophages) & NK cells 4. Protein molecules - Cytokines (tell cells what to do - involved in development and many other things - chemical messengers between cells),, complement, protein mediators of inflammation

2 main types of immunity in the body and their components

1. Immediate - innate immunity * Natural, present before infection, increase following infection - ready to go and non-specific (the immune response to one bacteria may be similar to that of another) - the molecules are already there * These are defences the body uses no matter what the invader is * Physical and chemical barriers - e.g. skin, mucous * Cellular responses - e.g. phagocytosis (neutrophils and macrophages), inflammation • - used when the first line of defence (physical barriers) have been breached - Inflammatory response enlists macrophages, mast cells, WBC's and chemicals - kill pathogens - Recognize surface carbohydrates unique to infectious organisms * Soluble mediators - cytokines, kinins, complement proteins Consists of 1st and 2nd lines of defence: 1st line of defence: - Physical - skin and mucous membranes - Chemical - acid, lysozyme, mucin (dissolved in water produced thick sticky mucous), defensins (antimicrobial peptides) - produced by the physical barriers) 2nd line of defence: - Cells involved: phagocytes, natural killer cells, antimicrobial proteins - Cellular responses (e.g. phagocytosis, inflammation) - Soluble mediators (e.g. cytokines, kinins, complements) - The hallmark of the 2nd line of defence is inflammation 2nd line is called in to action when the 1st line has been penetrated 2. Delayed - adaptive immunity (sometimes called specific or acquired immunity) - 3rd line of defence * Antibody (Ab) and cell-mediated responses (i.e. Lymphocyte-mediated responses) - Highly tuned - focused on a particular organism and its antigen • Specific, Systemic, Has memory Involves 3 crucial types of cells: o B lymphocytes o T lymphocytes o APC (antigen presenting cells)

What are the 4 main steps in the migration of leucocytes to sites of infection?

1. Interaction between activated vascular endothelium (selectins) and glycoproteins on the leucocyte 2. Tight binding triggered by chemokines (IL-8) - enhanced ability of leucocyte integrins to adhere to their receptor 3. Crossing of endothelial cell wall 4. Direction of migration follows gradient of chemokine • Macrophage sensor cell sends out signals which are cytokines • Adhesion molecules cause passing monocytes to slow down and facilitate the movement between the endothelial cells • Chemokine = a type of cytokine involved in cell movement - involved in recruiting cells to an area

Outline the process of inflammation in regard to the cardinal signs...

1. Process begins with an chemical 'alarm' - flood of inflammatory chemicals released into ECF - Macrophages have cell surface receptors (TLRs - Toll-like receptors) - play a central role in triggering immune responses - each of the 10 types recognize a specific class of attacking microbe - TLR triggers release of chemicals (cytokines) that promote inflammation and attract WBCs to the scene - Mast cells release histamine - Inflammatory mediators all cause small blood vessels in injured area to dilate - Antimicrobial proteins (interferons and complement proteins) attack microorganisms directly or hinder their ability to reproduce 2. As more blood flows into the area, hyperemia (congestion with blood) occurs - accounts for redness and heat of an inflamed region 3. Chemicals increase permeability of local capillaries - fluid containing clotting factors and antibodies seeps from the blood into the tissue spaces - causes local edema (swelling) - presses on nerve endings, causing pain ϖ Edema has a surge of protein-rich fluids into the tissue spaces - helps dilute harmful substances and delivers important proteins to the IF 4. Pain occurs because of release of bacterial toxins and effects of prostaglandins and kinins - Beta-defensins enter inflammatory sites where an epithelial barrier has been breached - antimicrobials - help maintain sterile environment of body passageways - help control bacterial and fungal colonization

What are the 4 cardinal signs of acute inflammation and what are they caused by?

1. Redness 2. Heat 3. Pain 4. Swelling • Redness - due to vasodilation - macrophages produce cytokines that promote inflammation - they cause small blood vessels in injured areas to dilate - as more blood flows into the area, hyperemia (congestion with blood) occurs - leads to redness and heat • Heat - due to blood flowing to inflamed area at core temperature • Pain - kinins and other things (such as bacterial toxins and effects of prostaglandins) can stimulate nerve endings • Swelling - fluid (plasma) leaves the circulation into the tissue - Chemicals increase permeability of local capillaries - fluid containing clotting factors and antibodies seeps from blood into tissue spaces - causes local edema - presses on nerve endings, causing pain

What are the 5 characteristics of adaptive immunity?

1. Specificity Lymphocyte receptors are able to recognise subtle structural differences between distinct antigens - can generate many different antibodies 2. Diversity The total number of antigen specificities of lymphocytes in an individual ie lymphocyte repertoire is large (109 to 1011) 3. Memory Enhances the ability to respond to reinfection (phagocytes don't have memory - they will act the same way each time) 4. Selflimitation The immune responses wane off with time following the elimination of antigens and homeostasis is maintained at the basal rate - proliferate to high numbers when they are activated, then die when they are not needed (this is helpful in the body) 5. Tolerance Immunological non-responsiveness to self antigens

What are some of the functions of antibodies (4)?

1. opsonisation - enhance phagocytosis, enhance complement mediated killing 2. ADCC 3. neutralise toxins 4. prevent pathogen attachment

What are the 3 pathways of activation of the complement system and which 'C' is central to all 3 pathways?

3 pathways of activation: • 1. Alternative 2. Classical 3. Lectin • C3 central to all 3 pathways • Needs activation, highly regulated

What is Interferon gamma (IFN- γ)?

A cytokine that activates macrophages and induces class II major histocompatibility complex (MHC) molecules

Name the important things within a lysosome

A lysosome contains: free radicals (toxic molecules), nitric oxide, superoxide, hydrogen peroxide, lysozyme

What is a fever? why/how does it occur? what are its effects?

A systemic response to invading microorganisms. When leukocytes and macrophages are exposed to foreign substances in the body, they release chemicals called pyrogens (pyro = fire). These pyrogens act on the body's thermostat (a cluster of neurons in the hypothalamus) raising the body's temperature above normal • A systemic response to invading microorganisms. • When leukocytes and macrophages are exposed to foreign substances in the body, they release chemicals called pyrogens (pyro = fire). These pyrogens act on the body's thermostat (a cluster of neurons in the hypothalamus) raising the body's temperature above normal Effects: - Liver and spleen to sequester iron and zinc, making them less available to support bacterial growth. - Increases the metabolic rate of tissue cells in general speeding up repair processes.

What is an epitope?

sites within antigens recognised by an Ab or other antigen receptor (e.g. TCR) - there may be antibodies directed against different epitopes of an antigen - there are different epitopes that interact with T cells ϖ T cells interact with antigens when it is broken up into small peptides ϖ MHC molecule binds to one of the peptides and presents it to a T cell

What are some acute phase proteins and how can an acute phase response occur?

Acute Phase Proteins: • C reactive proteins • Serum amyloid A • Prothrombin • Fibrinogen • Plasminogen • Mannose-binding lectin (MBL) • Ferritin • Ceruloplasmin • Alpha 2 macroglobulin • Alpha 1 antitrypsin Often CRP levels are indicative of the level of inflammation in the body

What are some words used to describe adaptive immune responses?

Adaptive immune responses = Acquired / Specific

What are adaptive immune responses initiated by? Which cells are involved in innate immunity? Which cells are involved in adaptive immunity?

Adaptive immune responses are initiated by antigen and antigen-presenting cells in secondary lymphoid tissues

What types of barriers are the first defence against pathogens? What are the steps in this process?

Anatomic and chemical barriers are the first defense against pathogens • First barriers are physical and chemical (e.g. mucous and acidic pH in stomach) • Preformed mediators (e.g. phagocytes, complement) can help contain the infectious agent - Can then get more phagocytes and neutrophils recruited • Antigen is transported to the lymphoid tissue and the lymphocytes are activated and another type of response occurs (e.g. T cells get activated)

Describe the different anatomical compartments of the spleen...

Anatomical Compartments • Served by splenic artery and vein - enter and exit hilum on concave surface • Provides site for lymphocyte proliferation and immune surveillance and response • Macrophages remove foreign matter and debris from the blood • Surrounded by a fibrous capsule, has trabeculae that extend inward - contain lymphocytes and macrophages • White pulp - Areas composed mostly of lymphocytes suspended on reticular fibers - forms cuffs around central arteries (branches of splenic artery) - form islands in the sea of red pulp - immune functions - Contains: - B cells - Ab production - APCs (Dendritic cells and macrophages) in the marginal zones can sample antigens from the blood, transport, process and display antigens to activate local naïve T cells in the PALS. T cells- cytotoxic or helper Surrounding the central arteriole is the periarteriolar lymphoid sheath (PALS), made up of T cells. Lymphocytes and antigen-loaded dendritic cells come together here. • B cell containing follicles are situated at intervals along the sheath • The follicles consist mainly of B cells; in secondary follicles, a germinal center is surrounded by a B-cell corona. - Follicles are surrounded by a so-called marginal zone of lymphocytes. - Marginal zone surrounding the follicle is rich in macrophages, T cells and marginal zone B cells • Red pulp - All remaining splenic tissue - venous sinuses and splenic cords - regions of reticular CT rich in macrophages - Disposes worn-out RBC's and bloodborne pathogens • Spleen also has T cell and B cell areas - only when they are activated do they come together and interact

How do complement proteins lyse a pathogen?

Cell lysis begins when C3b binds to the target cell's surface and triggers the insertion of a group of complement proteins called MAC (membrane attack complex) into the cell's membrane. MAC forms and stabilizes a hole in the membrane that allows a massive influx of water, lysing the target cell. The C3b molecules that coat the microorganism provide "handles" that receptors on macrophages and neutrophils can adhere to, allowing them to engulf the particle more rapidly.

Describe the cells in each compartment of the lymph node...

Cells in the compartments of each lymph node • Cortex (superficial) - follicles containing germinal centers with B cells - lymphocyte that matures in bone marrow and induced to replicate by antigen binding, usually followed by helper T cell interactions in lymphoid tissues. Clone members form memory cells and plasma cells - Germinal centers have the differentiation of B cells - plasma cells (produce specific antibodies) • outer cortex of B cells organized into lymphoid follicles and of adjacent, or paracortical, areas made up mainly of T cells and dendritic cells. - Primary lymphoid follicles have B cells which produce antibodies (memory or effector) • When an immune response is under way, some of the follicles—known as secondary lymphoid follicles— contain central areas of intense B-cell proliferation called germinal centers - eventually die out as germinal centers become senescent. • Medulla consists of strings of macrophages and antibody-secreting plasma cells known as the medullary cords. • Dendritic Cells - encapsulate follicles - antigen presenting cells - engulf and digest antigens, presenting parts of them on their plasma membrane (bound to the MHC protein) for recognition by T cells bearing receptors for the same antigen - essential for normal, cell-mediated responses • T cells are in transit • B cells live in cortical area • T cells live in paracortical area - helper (present Ag), effector (cytotoxic) and DC (present Ag and activate T cells) • Plasma cells - in medullary cords - antibody producing machines - produce large numbers of antibodies (immunoglobulins) with same antigen specificity. Specialised B cell clone descendent • Macrophages - on reticular fibers - type of phagocyte - detects, engulfs and destroys pathogens and apoptotic cells - Once ingesting a microbe, it presents an antigen on its surface which signals its presence to a corresponding helper T cell - Lymph nodes have areas where the B cells and the T cells live - Internally are the macrophages in lymph node

What are antimicrobial proteins? what are the 2 main types?

Cells that attack microorganisms directly or hinder their ability to reproduce. Complement proteins and interferons

What is the complement system and what is its purpose? How are they named and what type of immunity is it part of?

Complement = system of ~30 proteins, many enzyme precursors - All interact with each other in a cascade - C = complement protein - There are a number of ways in which these pathways can be activated - Even though the triggers are different, the pathways end up doing a similar thing - They can drive inflammation, allow inflammatory cells to come into tissues (chemotaxis) - Can participate in opsonisation = enhanced phagocytosis (allows the phagocytes to digest the molecule more easily) - Can lyse cells (e.g. bacteria and host cells also) - a number of the C proteins arrange themselves in a structure on the membrane of the cell, and cause a pore to form which causes lysis • Named with prefix C (e.g. C1, C2, C3 etc) • Present normally among plasma proteins or in tissues • Part of innate and adaptive immunity

What are opsonins?

Complement proteins or antibodies that provide 'handles' to which phagocyte receptors can bind

What are dendritic cells activated by and where do they move to to have their action?

Dendritic cells in tissues are activated via PAMPs/PRRs and move to lymphoid tissue to initiate adaptive immunity • Sensor cells are located just beneath the skin • DC switches on genes to get out of the infected tissue and go to the draining lymphoid tissue - in this case, they will move to the lymph nodes where they will present the antigen to the T cells which expand and proliferate and go back to the site of infection

What does the type of adaptive immune response utilised depend on?

Depending on the type of microbes invading the body

Macrophages: 5 main functions Where they are found Life span in relation to neutrophils Factors they produce

FUNCTIONS 1. Antigen presentation (APC) - Although macrophages (like DC's) can activate naïve T cells, they often present antigens to T cells to be activated themselves - Certain effector T cells release chemicals that prod macrophages to become activated macrophages, true "killers" that are insatiable phagocytes and secrete bactericidal chemicals. 2. Phagocytosis & destruction of bacteria 3. Secretion 4. Tumour cell destruction 5. Tissue reorganisation & wound healing • Key sensor cell • Secretion (e.g. of cytokines) * Macrophages are large phagocytic cells found throughout the body in tissues and they live longer than neutrophils * Antigens are partially degraded and bound to molecules called MHC for presentation to T helper cells * Macrophages are secretory cells producing factors which influence inflammation, healing and the body's response to infection

Describe the process of phagocytosis

Flowing cytoplasmic extensions bind to the particle and pull it inside a membrane lined vesicle becoming a phagosome A phagosome then fuses with a lysosome to form a phagolysosome Neutrophils and macrophages generally kill ingested prey by acidifying the phagolysosome and digesting its contents with lysosomal enzymes

What are the differences between Humoral and Cell-mediated/cellular immunity?

Humoral: • Ab-mediated immunity - provided by antibodies present in the body's humours (fluids) - produced by lymphocytes - circulate freely in blood and lymph - inactivate foreign complexes and mark them for destruction by phagocytes or complement • overseen by B lymphocytes Cellular/Cell-mediated: • lymphocytes themselves defend the body • has cellular targets - virus-infected or parasite-infected tissue cells, cancer cells and cells of foreign grafts • can act directly by killing foreign cells or indirectly by releasing chemical mediators to enhance the inflammatory response or activate other lymphocytes or macrophages • Constituted by T cells - they are non-antibody producing lymphocytes

What are the major pro inflammatory cytokines?

IL-1, IL-6 & TNF-α are major proinflammatory cytokines

What are the main steps in T cel development and activation?

• Lymphoid progenitor cells originally in BM, which then go to the thymus, where they switch on genes to become T cells • Any T cells that are selected leave the thymus and go into circulation and wait to be activated by an APC • Undergo a selection process: - Positive selection - ensure only T cells that are able to recognize self-MHC proteins survive (if not, they undergo apoptosis) - Negative selection - ensure that T cells do not recognize self-antigens displayed on self-MHC (basis for immunologic self-tolerance) • T cells can only recognize and respond to processed fragments of protein antigens displayed on surfaces of body cells (APCs and others) - unlike B cells and antibodies - Antigen presentation required for activation of naïve T cells and functioning of effector T cells - T cell antigen receptors (TCRs) bind to an antigen-MHC complex on the surface of an APC. - TCRs must perform double recognition: must recognize both MHC (self-antigen) and the foreign antigen it displays. - The binding of the TCR to the nonself-self complex triggers multiple intracellular signaling pathways that lead to T cell activation - CD4 and CD8 proteins that distinguish the two major T cell groups are adhesion molecules that help bind cells together during antigen recognition - they help with T cell activation - For a T cell to proliferate to form a clone, it must also bind one or more co-stimulatory signals - T cells are disposed of after a certain period of time - good because they can be dangerous because they produce large amounts of inflammatory cytokines

What are the functions of the complement system that protect the host?

• Lytic events begin when C3b binds to target cell's surface, triggering insertion of group of complement proteins called MAC (membrane attack complex) into cell's membrane - C3b helps activation C5 into C5b and C5a. C5b, C6-9 to produce a MAC • MAC creates a hole in the membrane and ensures lysis by inducing a massive influx of water • C3b molecules that coat the microorganism provide 'handles' that receptors on macrophages and neutrophils can adhere to, allowing them to engulf the particle more quickly (opsonisation) • C3a and other cleavage products formed during complement fixation amplify inflammatory response by stimulating mast cells and basophils to release histamine and by attracting other inflammatory cells to the area • CRP (produced by the liver) binds to C1 of classical complement pathway, activating it and results in deposit of C3 on the surface of bacteria (opsonisation)

What happens to the body if the complement system is not regulated?

• MAC can also form on host cells - autoimmune reactions • Need balance between complement activation and inhibition - can cause severe inflammation and cell death if it is not regulated properly

What is a basic definition of the complement system and what does activation of the complement system result in?

• Most important of the proteins of the innate response that destroy invading bacteria • The complement system is a set of proteins that act together to cause the destruction of invading microorganisms • Activation of the complement system results in: - inflammation due to chemotaxis - opsonisation (clearance) - cell lysis (by MAC complex - C5-C9) - regulation of the immune system

What are PAMPs recognised by?

• PAMPs (Pathogen associated molecular patterns) on invading microorganisms are recognized by Toll-like receptors (TLR's) found on/in macrophages - signals from these activate genes that are involved in the production of cytokines, triggering inflammation • bind to their specific receptor to induce an effect that is typically related to amplifying the effector mechanism of the target cell. • Things that may induce cytokine production y macrophages include viruses, fungi, parasites, tissue damage

Diagram of how innate and adaptive immune responses work together...

• Preformed mediators are there below the external barrier • Helper T cells can help the B cells make the antibodies - the whole thing is linked together

When does the primary immune response occur?

• Primary immune response = cellular proliferation and differentiation - has a lag period of 3-6 days after antigen encounter (because the few B cells specific for the antigen have to proliferate), plasma antibodies rise and reach peak in about 10 days • If a person is exposed to the same antigen, a secondary immune response occurs - faster, more prolonged and more effective - Within hours after recognizing the 'old enemy' antigen, a new army of plasma cells is being generated and antibodies reach higher levels more quickly - Secondary response antibodies bind with higher affinity and blood levels remain high for a long period of time

Why are Leukocyte Adhesion Deficiency Patients prone to recurrent infection and impaired wound healing?

• Prone to recurrent infection and impaired wound healing - Adhesion molecules strengthen the interaction of leucocytes with the endothelium ϖ aid their entry in large numbers into the infected tissue to form an inflammatory focus so that the inflammatory response is maintained and reinforced. - Leukocytes and other molecules which rely on adhesion molecules are unable to enter the tissue efficiently which results in uncontrolled infection and poor wound healing. • LAD type 1 = failure to express CD18 which composes the beta-2 subunit of LFA1 family (beta-2 integrins) - LFA1 plays an important role in lymphocyte adhesion to vascular endothelium and interactions to antigen presenting cells. Also plays a role in cytotoxic killing by T cells • Type 2 = rare - defect in expression of ligans for selectins due to lack of enzymes

What type of cells are dendritic cells, what are they derived from and what is their function? Where do immature ones and mature ones go in the body?

• Start the adaptive immune response * Derived from myeloid progenitors - Have lots of MHC on themselves * • Found at the body's frontiers (in the subepithelial layers of the skin - Langarhans cells) - ready to pick up antigens - In this position, they are immature - becomes mature when it is stimulated by a microbe Specialised antigen presenting cells (APC) * They take up antigen, process it and display the antigens for recognition by T cells * Immature DC migrate from the blood into tissue and reside in tissue and are both phagocytic and micropinocytic - Triggered by the PAMP interaction • Once they have internalized antigens by phagocytosis, they enter nearby lymphatics to get to a lymph node where they will present the antigens to T cells. * Mature DC migrate to secondary lymphoid tissue - where the T cells are waiting to be activated - key to innate and adaptive immune responses

Outline the steps in which inflammation initiates the immune responses...

• Stimulus could be a microbe (e.g. a LPS stimulating a sensor cell) • Pattern Recognition Receptors are ON the sensor cells (not like PAMPs which are on the microbes)

What is the difference between a PAMP and a DAMP and what do they cause?

• Surface and intracellular PRRs are activated by many different pathogen-associated molecular patterns (PAMPs) and danger associated molecular patterns (DAMPs) - DAMPs are things created by the host-cell itself (e.g. ATP) - host-cell derived metabolites • Innate immune function depends upon recognition of pathogen- associated molecular patterns (PAMPs), derived from invading pathogens and danger-associated molecular patterns (DAMPs), induced as a result of endogenous stress • DAMP-triggered inflammation (important in inflammatory diseases) is termed sterile inflammation when it occurs in the absence of any foreign pathogens - E.g. T2DM - inflammation not caused by the pathogen • Once they bind to their ligand, they initiate gene expression which then triggers phagocytosis • Lysosome in the cell contain lots of radical molecules that are detrimental to microbes • Fusion vesicle between the phagosome and lysosome = phagolysosome - where the microbe is killed - this is the part of the cell that generates the peptides to be attached to the MHC (particularly MHC II)

What occurs in the 3 main phases of the adaptive immune response?

• These phases take place at different times after some type of trigger • Recognition phase - Foreign material recognized - There are many types of cells involved - recognition between sensor cell and microbe • Activation phase - Moving into adaptive immunity - T and B cell lymphocytes recognize an antigen and are activated - expand in a process called clonal expansion - Only one or 2 specific T cells in the body will recognize a particular antigen - they will create clones of themselves (why it is called clonal expansion) - Lymphocytes start to develop effector functions (B cells produce antibodies) • Effector phase - The cells carrying out the effector function - may produce cytokines that recruit other cells into the immune response - Antibodies have multiple functions - can participate in complement activation, facilitate phagocytosis - Can eliminate antigen, return the body to homeostasis (there has been lots of proliferation of cells - these cells have to be removed from the body)

What are the features of Class II MHC Proteins?

• Typically found only on the surfaces of cells that present antigens to CD4 cells - dendritic cells, macrophages and B cells • Display exogenous antigens - antigens from outside the cell that have been engulfed by the cell • CD4 will help mount a defence against the antigen

How does the migration and homing of leucocytes occur? When are they recruited?

• WBC emigrate from blood to perform their effector function - Some may make their way to the thymus where they become T cells • Lymphocytes recirculate through the secondary lymphoid tissue - detained if they encounter Ag • Monocytes migrate into tissue and mature into macrophages - Don't have macrophages circulating in blood normally as they are tissue cells (blood counts are always monocyte counts, not macrophage counts) • Lymphocytes and granulocytes are recruited to extravascular sites in response to infection or injury • Need to get out into the periphery (not in the primary lymphoid tissue) - Their movement is controlled by cytokines which can drive cells in and out of tissues - lymphocyte movement is regulated by homing signals (CAMs) displayed on vascular endothelial cells • endothelium changes - increases number of adhesion molecules on its surface • this process used by cytotoxic T cells, monocytes and neutrophils - alerted to presence of danger due to change in endothelium

What are some of the different types of chemicals phagocytes can use to kill microbes?

• When the lysosome joins with the phagosome, it drops the pH which can help to kill the microbe

What are some examples of complement deficiencies and what are they caused by?

• increased susceptibility to bacterial infections (especially Neisseria infections) • syndromes resembling systemic lupus erythematosus (deregulation of complement cascade causes widespread inflammation) Factor H deficiency: membrano-proliferative glomerulonephritis and hemolytic uremic syndrome, age related macular degeneration