Immunology Exam 1

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Immunology

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-study of the body's natural defense mechansims against disease
-the study of the immune system, including the composition, function, and disorders
-a study developed when individuals who had a who had a particular infection, were later protected from the same infection
-immunis: latin for exempt
-immunity: state of protection from the same disease
-evolved to protect multicellular organisms from foreign material
-composed of a variety of molecules capable of specifically recognizing and eliminating foreign material

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Pathogens

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foreign materials that cause disease

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4 Major Categories of Pathogens

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1. Viruses
2. Bacteria
3. Fungi
4. Parasites

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Virus

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-unique because can't replicate themselves
-molecules that infect cells
-consist of genetic material (DNA/RNA) covered by a protein coat called a capsid
-your immune system must recogniize and eliminate a virus before it's life cycle is completed
-the later the response, the more viruses

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Fungi

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-major example is yeast (unicellular and eukaryotic)
-yeast is found on skin, intestinal tract, mucosal membranes, etc. (normal flora)
-multiply by budding
-yeast is a common cause of opportunisitc infections
-can develop/cause rashes/infection
-example: thrush is growth of yeast in mounth, often in infants

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Bacteria

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-unicellular and prokaryotic
-found on skin, intesinal tract, mucosal membranes, entire body → normal flora that can cause opportunisitc infections

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Parasites

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-multicellular and eukaryotic
-feeds off of other organsims → symbiosis
-NOT normal flora
-major example: worm
-most commonly found in undercooked food and the soil

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Antigen (Ag)

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any pathogen that binds to an antibody

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Two Major Activities of the Immune System

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1. Recognition
-recognize common molecular characteristics of pathogens
-detect subtle differences
-self-non-self discrimination
-able to recognize host/self cells that are altered and need to be removed
2. Response (elimination of pathogen)
-effector response: actual elimination of the pathogen
-memory response: effector response which is faster and heightened

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Effector Response

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actual elimination of the pathogen

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Memory Response

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effector response which is faster and heightened (because more cells, memory cells, are responding)

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Two Categories/Divisions of Immune System

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1. Innate Immunity
2. Adaptive Immunity

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Innate Immune System Overview

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-first line of defense
-utilizes molecules that you're born with
-main goal is to prevent the presence of the pathogen (creates barriers), and if the pathogen breaches the barrier, try to quickly eliminate it
-has self-nonself discrimination, but cannot distinguish between different pathogens

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4 Major Defense Barriers of the Innate Immunity System

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1. Anatomic Barriers
2. Physiologic Barriers
3. Endocytic and Phagocytic Barriers
4. Inflammatory Barriers

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Anatomic Barriers (physical structures) of Innate Immune System

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1. Skin
-mechanical and physical barrier with epidermis (thin outer layer) and dermis (thick inner layer)
-sebum: oil produced by the sebaceous glands in the SSE with a low pH so helps protect skin from entry of pathogens, especially bacteria whose optimal pH is 6-8 and its low pH also denatures proteins
2. Mucous Membranes
-found in: nose, mouth, lungs, eyelids, digestive tract, and genital tract
-cells secrete mucus, which is a thick substance with antibacterial enzymes and thus an acidic pH
-trap pathogens (because thick)
-acidic pH will prevent growth and denature pathogens
-contian cilia (usually respiratory tract) that help sweep away pathogens

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Physiologic Barriers of Innate Immune System

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1. Temperature (optimal temp for bacterial growth is 25-37)
-high temps denature pathogen
-high and low temps will prevent growth of pathogen
2. pH-Acidic (optimal pH for bacteria is 6-8)
-low/acidic pH will denature and prevent growth; found in tears, saliva, stomach, mucus, etc.
3. Oxygen-Tension
-low oxygen levels: aerobic bacteria don't grow
-high oxygen levels: anaerobic bacteria don't grow
4. Chemical/Soluble Factors
-lysozyme: antibacterial enzyme found in tears, saliva, and mucus that lyses and breaks down the cell wallof bacteria by affecting the peptidoglycan layer
-interferons: anti-viral molecules which are proteins produced by virus-infected cells; they bind to neighboring cells to ward off virus from entering by causing a physical hindrance so spokes of virus can't attach
-complement: protein "flags" pathogen for removal; induce lysis; can prevent virus entry into cell by physical hindrance

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Endocytic and Phagocytic Barriers of Innate Immune System

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1. Endocytosis: ingestion of small extracellular material
-pinocytosis: liquid; occurs in most cells
-receptor-mediated endocytosis: bind to receptor before engulfed; occurs in most cells
2. Phagocytosis: ingestion of particulate (large) matter
-occurs in phagocytes (special cells)

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Inflammatory Barriers of Innate Immune System

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-when pathogen breaches skin and mucous membranes, the pathogen can induce a cascade of events known as the inflammatory response
-generalized inflammation repsonse mechanism:
1. pathogen breaches skin and mucous membranes
2. causes tissue injury
3. endothelial cells and WBCs start secreting factors called cytokines which bind to blood vessel
4. binding causes vasodilation, which increases blood flow to area and thus the number of WBCs too
5. increase in vascular permeability
6. WBCs, fluid, enzymes leave vessel
7. WBCs move to pathogen
8. Destroy/get rid of pathogen

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Hallmark Signs of Inflammation

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1. Dolar (pain)
-the cytokine bradykinin secreted during inflammation functions to bind to pain receptors so that you get signal and feel pain
2. Rubor (redness)
-when blood vessels dilate that increases blood flow which increases redness
-also, increased vascular permeability allows fluid to seep out which causes redness under skin
3. Calor (heat)
-due to increased blood flow
4. Tumor (swelling)
-due to increased vascular permeablility

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Overview of Adaptive Immunity

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1. develops in response to a pathogen
2. adapts to recognize, eliminate, and develop a memory against the pathogen
3. relies on innate immunity for assistance (cooperative)
-adaptive immunity begins a few days after initial infection
-innate immunity is the first line of defense
4. provides a second and comprehensive line of defense that eliminates pathogens which evade the innate system or persist in spite of them
5. Antigen is the typical name for pathogen because they interact with and recognize the antibody

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2 Major Groups of Adaptive Immunity Cells

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1. Lymphocytes
2. Antigen-Presenting Cells

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Lymphocytes

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-type of WBC
-produced in bone marrow via process of hematopoiesis
-leave bone marrow → go into blood → move onto lymphoid organs
-two major types of lymphocytes: B and T

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B Lymphocytes

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-produced and mature in bone marrow
-each cell expresses a unique antigen-binding receptor on its membrane called a B cell receptor; this is a membrane bound antibody
-one B cell can have 10⁵ Ab receptors, but they will all bind the same Ag
-general mechanism of B cell development and action: naive B cell encounters an Ag that matches its Ab receptor and they bind, causing the cell to rapidly divide and differentiate into memory cells and effector B cells called plasma cells
-Memory B cells: long life span; membrane-bound receptors (Ab); recognizes the same Ag as the parent cell; produces a faster and heightened immune response
-Plasma Cells: short life span (few days); make Ab molecule in a secreted form (free Ab); still binds to same Ag, but lacks hydrophobic region to bind to membrane; can release 100,000s of Ab molecules/second
-Secreted/free Ab binds to Ag and induces Ag's elimination from body
-B cells also act as Antigen-Presenting Cells (APC) which can activate T cells (TH), which help eliminate pathogen

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Antibody

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-glycoprotein: has a carb/sugar attached to a string of AAs
-consists of two heavy and two light chains (proteins)
-four polypeptide chains linked by four disulfide bonds
-this molecule is found in B-cell membranes
-antigen-binding sites at open ends
-one receptor usually binds two antigens

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T Lymphocytes

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-produced in the bone marrow
-mature in the thymus
-maturing T cell expresses a unique antigen-binding receptor called a TCR on its membrane, which is a glycoprotein that varies depending on the T cell
-two major groups of T cells which are differentiated based on 2nd receptor: TH cells and TC cells
-recently characterized 3rd type is T regulatory cell
-TCRs can only recognize Ag bound to a molecule called MHC (whereas a BCR can recognize an Ag all by itself)
-general T cell development and mechanism of action: naive T cells encounter Ag-bound with MHC on cell which induces the T cell to proliferate and differentiate into memory T cells and effector T cells (which includes TH and Tc); interaction of TH with APC causes APC to secrete cytokines wich activate TH cells which secrete cytokines that activate themselves, B cells, macrophages, and other phagocytes; cytokines also induce Tc cells to turn into CTLs that can actually eliminate antigen bound to MHC-I (usually by getting rid of the whole altered self cell

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T Helper Cells (TH)

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-displays regular TCR and also has CD4
-CD4 helps TCR recognize and bind to the Ag
-subsets of TH cells:
1. TH₁: activate Tc cells and macrophages
2. TH₂: activate B cells

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T Cytotoxic Cells (Tc)

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-display a TCR and CD8

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T Regulatory Cells (Treg)

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-displays CD4
-differentiated from TH via presence of other cell surface markers

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Major Histocompatability Complex (MHC)

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-glycoproteins
-classes I and II

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MHC-I

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-nearly all cells express this
-Tc cells recognize Ag bound to MHC-I
-Ag: endogenous proteins (include: viral proteins, cancer proteins, and foreign tissue graft proteins)
-altered self cells (have MHC-I and this will interact with Tc cells)

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Endogenous Proteins (Ag)

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-Ag which is internally processed and alters the cell, usually by invading
-examples: viral, cancer, and foreign tissue graft proteins
-processed by the cytosolic pathway

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MHC-II

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-expressed by APCs, which include: B cells, dendritic cells, and macrophages
-TH are the one type of lymphocyte that can recognize MHC-II
-Ag bound to MHC-II molecules are typically exogenous Ag (bacteria, fungi, parasites)

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Exogenous Proteins (Ag)

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-stay outside the cell, don't alter it, but are phagocytized by cell
-examples: bacteria, fungi, and parasites
-processed by the endocytic pathway

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Cytosolic Pathway

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-mode of processing of endogenous Ag (which bind to MHC-I and are recognized by Tc cells)
-steps in example of virus:
1. virus invades cell
2. viral DNA exposed in cell
3. viral proteins are made
4. small % of these bind to MHC-I molecules inside and are packaged inside a vesicle
5. vesicle moves toward membrane, and MHC-I with bound Ag is displayed
6. free Tc cells come to recognize this Ag on the altered self cells

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Endocytic Pathway

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-mode of processing of exogenous Ag (which bind to MHC-II molecules and are recognized by TH cells)
-steps in example of bacteria:
1. phagocytose or endocytose bacteria
2. pinches off into a phagosome
3. fuses and binds with a lyzosome, which is an internal structure with hydrolytic enzymes
4. called a phagolyzosome and degrades bacteria
5. pieces of bacterial protein bind to MHC-II and in vesicle
6. moves to membrane
7. recognized by TH cells

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2 Categories of Adaptive Immunity

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1. humoral immunity
-involves Ab
-primarily protects host against exogenous antigens (bacteria, fungi, parasites, etc.)
2. cell-mediated immunity
-primarily involves T cells and sometimes other immune cells
-primarily protects host against endogenous Ags and some extracellular/exogenous Ags (esp. bacteria)

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Humoral Immunity

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1. mediated by Ab
2. B cell interacts with Ag → proliferates and differentiates into memory and effector (plasma) cells → secreted/free Ab free to bind to Ag → Ag eliminated by lysis or phagocytosis

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How does Ab induce the elimination of Ag?

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1. cross-linking: of several Ag together by Ab
-clusters them together which increases ease of recognition by phagocytes
-then, once recognized, phagocytosis
2. complement system activation by Ab: complement are molecules that can bind to Ag and cause it to lyse
-leads to lysis of Ag
-signals to phagocyte → increases phagocyte recognition → phagocytosis

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Cell-Mediated Immunity

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1. primarily mediated by T cells
2. TH cell mechanism:
-TH cells recognize and interact with Ag bound to MHC-II molecule of APC
-TH cells become active
-TH cells secrete cytokines which in turn activate B cells, macrophages, T cells, and macrophages
3. cytokines secreted by TH cells activate both humoral and cell-mediated response
4. B cells participate in:
-cell-mediated reponse: by acting as an APC which can further activate TH cells
-humoral response: because plasma cells can secrete free Ab

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4 Major Characteristics of Adaptive Immunity

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1. Specificity
2. Diversity
3. Self vs. Non-Self Recognition
4. Memory

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Specificity of Adaptive Immune System

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-can distinguish the differences between pathogens/antigens
-contributed by 3 classes of membrane-bound molecules
1. Ab of B cells
2. TCR of T cells
3. MHC-I and MHC-II

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Diversity of Adaptive Immune System

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-can recognize billions of unique structures on Ags, whereas innate immunity can recognize pathogens but cannot distinguish between different pathogens
-B cells:
1. Antigenic specificity of B cells is determined by its membrane-bound Ab
2. the specificity of the Ab receptor is created by random rearrangements of the genes that encode the Ab receptor
-at maturity, each B cell possesses one gene encoding the Ab heavy chain and one gene encoding the Ab light chain
-random gene arrangements during the B cell maturation in bone marrow generates a large number of different Ab receptor combos and thus, different antigenic specificities
3. Each B cell:
-expresses a unique Ab
-displays 10^5 receptors on one B cell
-each B cell displays Ab receptors which are specific for one Ag
-T cells:
1. T cell maturation also includes random rearrangements of genes which encodes the T cell receptor (TCR)
2. Each T cell:
-expresses unique TCR
-displays 10^5 TCR molecules on one T cell
-each T cell displays TCR molecules which are specific for one Ag
-MHC: MHC development includes random rearrangements of genes which encode MHC molecules

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Self vs. Non-Self Recognition of Adaptive Immune System

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-can distinguish between self and antigens
1. the antigenic specificity of each B and T cell is determined by its membrane receptors (Ab and TCR)
2. Once the Ag binds to the receptor, the B and T cells are antigenically-committed; the cells proliferate (expand) to produce cells that have the same antigenic specificity; any cell which recognizes self is destroyed or supressed later on if mature (CLONAL SELECTION)

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Memory of Adaptive Immune System

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-If the same or closely-related Ag infects a body again, memory cells help make a fast and heightened response:
1. Proliferated B and T cells either develop into memory cells or effector cells
2. Primary vs. Secondary Response
-primary response: initial encounter of Ag
-secondary response: later contact with Ag is faster, heightened, and lasts longer due to the presence of memory cells

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Consequences of a Dysfunctional Immune System

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-sometimes the system fails to protect the host adequately because of a deficiency
-common manifestations:
1. Allergies/Asthma: innapropriate response to a common antigen
2. Graft Rejection and Graft vs. Host Disease: immune system encounters foreign cells and strongly responds to eliminate them
3. Autoimmune Disease: lose self/non-self recognition
4. Immunodeficiency: defective immune system could be due to genetic disorder, chemicals, or environmental factors

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Cell Composition of Blood

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-most cells involved in the immune system circulate in the blood but can also be found in tissues, lymph, and lymphoid organs
1. Leukocytes: immune function
-Neutrophils
-Lymphocytes
-Monocytes
-Eosinophils
-Basophils
2. Erythrocytes: oxygen-carrying function
3. Thrombocytes: clotting function

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Blood Smear Technique

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1. place drop of blood on slide
2. using another slide, spread blood to make an even layer
3. allow to dry so cells are fixed
4. apply stain (usually Wright's' stain)
5. wash off excess
6. look under microscope

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Blood Smear Results

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1. RBC: red/pink donut shaped cell
2. Platelets: very small purple cells, look granular
3. WBC
-Neutrophils: multi-lobed nucleus with granular cytoplasm
-Lymphocytes: smallest WBC, large dark staining nucleus, non-granular cytoplasm (clear)
-Monocytes: large irregular nucleus, non-granular cytoplasm, generally large with irregular border
-Eosinophils: bilobed nucleus and granular cytoplasm wiith large pink/orange granules (because highly stained with acidic dye)
-Basophils: tri-lobed or bi-lobed nucleus and granular cytoplasm with large dark purple granules (because highly stained with blue/purple dye-basic)

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Lymphoid Lineage

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1. 20-40% of blood WBCs
2. 99% of WBCs in lymph and tissues
3. Subdivided into 3 major populations:
-B cells
-T cells
-Natural Killer Cells (NK)
4. Development of B and T Lymphocytes
-naive B and T lymphs: small and non-phagocytic, can barely see cytoplasm; arrested in G0 stage of cell cycle
-lymphoblasts: naive cell interacts with the Ag → progresses through cell cycle (G0, G1, S, etc.) → cells enlarge (because organelles double) and cytoplasm increased
-lymphocytes (effector and memory): lymphoblasts proliferate and differentiate into effector and memory cells; B lymphs → plasma cells; T lymphocytes → TH and Tc cells

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Natural Killer Cells (NK Cells)

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1. small, granular lymphocytes
2. 5-10% of blood lymphs
3. do NOT express CD4, CD8, TCR, or Ab membrane proteins
4. recognize tumor or virus infected cells (altered self cells)
5. eliminate cells via cytotoxic activity (cause cells to lyse)

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How do NK cells recognize a target cell?

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1. detect surface antigen on MHC-I displayed by some virus-infected cells
2. some tumor and virus-infected cells can induce immune response; anti-tumor and anti-virus can be recognized by NK cells
-How do NK cells bind to the Ab? CD16 on NK cells bind and recognize anti-tumor and anti-viral Ab

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Myeloid Lineage

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1. Mononuclear Cells
-Monocytes (blood)
-Macrophages (tissue)
2. Granulocytic Cells
-Neutrophils
-Eosinophils
-Basophils
3. Dendritic Cells
-Langerhan DC (epidermis)
-Interstitial DC (interstitial spaces of organs, except brain)
-Monocyte-derived DC
-Plasmacytoid-derived DC

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Mononuclear Cells

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1. consist of: monocytes (blood) and macrophages (tissue
2. development of mononuclear cells: myeloid progenitor cells in bone marrow → differentiate into promonocytes → leave bone marrow and enter blood → differentiate into mature monocytes → circulate in blood → migrate into tissue where they differentiate into tissue-specific macrophages
-this differentiation includes: enlargement; increase in # and complexity of organelles; increase in phagocytic activity/ability by increased synthesis of hydrolytic enzymes and increased secretion of soluble digestive factors in cytoplasm
3. macrophages can be dispersed throughout the body
-stay in a particular tissue: fixed macrophage
-remain motile: free macrophage
4. Macrophages are named according to their location:
-gut = intestinal macrophage
-lung = alveolar macrophage
-connective tissue = histiocytes
-liver = Kuppfer cells
-kidney = mesangial cells
-brain = microglial cells
-bone = osteoclasts
5. macrophages are phagocytes
6. opsonization
-if Ag is coated with Ab or other molecules, activation of macrophage is enhanced

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Opsonin

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any molecule bound to Ag that enhances phagocytosis

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Phagocytosis Mechanism of Macrophages

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1. Ag adheres to membrane (activates cell)
2. induces pseudopodia to extend around Ag
3. phagosome developed
4. phagosome fuses with lysosome and forms phagolysosome
5. Ag is degraded by enzymes and other molecules
--this is the stopping point for a normal phagosome, but if an APC:
6. small % of peptide will bind to MHC-II and move to the exterior of the cell
7. MHC-II + peptide activates TH cells

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How do we know that Ag moves to the exterior of the cell? (experiment)

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-use fluorescent microscopy
1. fluorescently labeled bacteria: membrane proteins of both the external membrane and of internal organelle membranes were labeled, and also cytoplasm has free proteins
2. added macrophages (APCs) (which have MHC-II)
3. allow time for macrophage to:
-phagocytose bacteria
-internally degrade bacteria into peptides
-allow peptide to combine with MHC-II
-allow MHC-II + peptide to move to exterior
4. washed macrophages to get rid of excess bacteria because don't have 100% phagocytosis because of time, ratio of macrophages to bacteria, and possibility of some defective macrophages
5. viewed with fluorescent microscope
-results were that they saw the signal on the outside membrane which helped prove that MHC-II + peptide does indeed move to exterior

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Granulocytic Cells (of Myeloid Lineage)

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-classified based on granules in cytoplasm
granules:; groups of proteins that stain with Wright stain and look like speckles
-subdivided into:
1. neutrophils
2. eosinophils
3. basophils

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Neutrophils

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1. 50-70% of blood WBCs
2. phagocytes (not APCs, so no MHC-II)
-lysosomes called primary granules (larger and denser) and secondary granules
3. circulate in blood (survive: 7-10 hrs.) or can move into tissue (survive few days)
4. Primary cause of increased number of neutrophils is infection because 1st WBC to the site
5. Leukocytosis: high WBC count; the primary cause is increased # of neutrophils
-normal: 4-10,000/microliter
-high: 15-20,000/microliter

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Eosinophils

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1. 1-3% of blood WBCs
2. phagocytes, but not main players
3. plays a major role in the elimination of pathogens, especially parasites
-secrete enzymes from granules
-these will damage the cell wall of parasite
-ultimate goal: lyse the parasite

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Basophils

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1. less than 1% of blood WBCs
2. NOT phagocytes
3. release granular proteins in response to allergies (ex. histamine)
4. when found in blood = basophils
when found in tissue = mast cells

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Dendritic Cells (DC)

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1. covered with long membrane extension that resemble the dendrites of a nerve cell
2. divided into 4 major categories:
-Langerhan DC: epidermis of skin
-Interstitial DC: interstitial spaces of organs (except brain)
-Monocyte-Derived DC: derived from monocytes which migrated from blood to tissue (different than macrophages)
-Plasmacytoid-derived DC: derived from plasmacytoid cells, which resemble plasma cells (no fingers), but function more like dendritic cells
3. APC - display Ag with MHC-II molecules
4. Special category not derived from bone marrow and do not act as typical APC: Follicular DC

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Follicular DC

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1. do NOT express MHC-II
2. exclusively found in lymph node structures → lymph follicles
3. do NOT express membrane-bound or secrete Ab
4. Assist in B cell maturation by presenting Ag to naive B cells

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Hematopoiesis

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-formation and development of blood cells
-general: start with pluripotent hematopoietic stem cell → under influence of growth factors and transcription factors, this cell differentiates into RBCs, WBCs, or platelets, and also self-renews

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Hematopoietic Stem Cells

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-pluripotent: can make different cell types, but committed to blood cell lineages
-self-renewing
-differentiate into one of two major pathways: lymphoid progenitor cells or myeloid progenitor cells

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Progenitor Cells

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1. lose ability for self-renewal
2. committed to a particular lineage
3. will eventually further differentiate into mature cells

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Lymphoid Progenitor

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differentiates into:
-B cell progenitor → B cells
-T cell progenitor → T cells
-Natural Killer Cells

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Myeloid Progenitor

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differentiates into:
-granulocyte → monocyte progenitor → monocyte OR neutrophil
-eosinophil progenitor → eosinophil
-basophil progenitor → basophil
-megakaryocyte → platelet
-erythroid progenitor → erythrocyte

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Where does hematopoiesis occur?

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1. hemaopoietic stem cells originated in yolk sac (during the first weeks of development)
2. migrate and move to fetal liver and spleen after 3 months of gestation
3. migrate to bone marrow after 7 months of gestation
4. maintained in bone marrow throughout the adult life

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How is hematopoiesis regulated?

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1. growth factors induce proliferation and differentiation of stem cells
2. Regulation of growth factor levels
-stromal cells: connective tissue cells found in bone marrow that are the primary source of growth factors; in bone marrow, form a scaffold on which hematopoietic stem cells grow and differentiate; secrete growth factors
-transcription factors (TF); regulate trascription of some genes encoding growth factors

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TF: GATA-1

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erythroid lineage

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TF: GATA-2

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erythroid, myeloid, and lymphoid lineages

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TF: PU.1

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erythroid, myeloid, and lymphoid

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TF: Bmi-1

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all hematopoeitic lineages

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TF: Ikaros

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lymphoid lineage

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TF: Oct-2

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B lymphoid

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Programmed Cell Death

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-each cell produced via hematopoiesis has a lifespan and dies via programmed cell death
-aka apoptosis
-does not initiate inflammation
-morphological changes:
1. decrease in volume
2. condensation of chromatin
3. degradation of the DNA
4. exterior modification which activated phagocytosis (flippase changes position of PS)
5. release apoptotic bodies which are phagocytosed

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Necrosis

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injured cell → swells → bursts → releases enzymes/proteins that were inside → tissue injury → inflammation

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Organs of the Immune System

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-Primary Lymphoid Organs: site of lymphatic maturation; include bone marrow and thymus
-Secondary Lymphoid Organs: sites where mature lymphs and Ag interact; include lymph nodes, spleen, and MALT

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Bone Marrow (Primary Lymphoid Organ)

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1. site of hematopoiesis
2. B lymphs are produced and mature in bone marrow (B cells interact with GFs/cytokines secreted by bone marrow stromal cells)
3. T lymphs are produced in bone marrow
4. cells move primarily into blood to be distributed elsewhere

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Thymus (Primary Lymphoid Organ)

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1. site for T lymph maturation
2. immature T lymphs are called thymocytes and they interact with GFs/cytokines secreted by stromal cells → become mature T lymphs
3. divided into 3 regions: capsule, cortex, medulla
-cortex and medulla are heavily populated with dendritic cells and macrophages
-stromal cells in the medulla release cytokines and GFs that cause T lymphs to mature

*anti-self programmed cell death*

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Lymph Nodes (Secondary Lymphoid Organs)

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1. organ which traps Ag from local tissue (lymph)
2. lymphatic system: blood circulating in body → liquid portion moves into tissue (ISF) and Ags are carried along → ISF + Ag moved into lymphatic vessel (called lymph now) → carried to lymphoid organs (1st are lymph nodes)
3. divided into 3 major regions: cortex, paracortex, and medulla
-cortex has macrophages and follicular dendritic cells which are a special class that lack MHC-II
-paracortex has T lymphs and dendritic cells
-medulla has plasma cells

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General Mechanism in Lymph Nodes

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1. Ag carried by lymph in afferent lymphatic vessel to lymph node
2. Ag moves across cortex to paracortex
3. dendritic cells bind and process Ag
4. dendritic cells with presented Ag activates TH cells
5. TH cells secrete cytokines to activate B cells
6. activated B cells group together in foci; B cells further develop into plasma cells, some of which move to the medulla
7. some activated TH and B cells move into groupings called primary follicles in cortex
8. B cells of primary follicles become activated and the primary follicle structure is then termed a secondary follicle with a middle region known as a germinal center which contains developing B cells

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Primary Follicle

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group of cells with naive/unactivated B cells and follicular dendritic cells located in the cortex of the lymph node

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Secondary Follicle

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group of cells now containing activated B cells because they have encountered Ag, and plasma cells generally congregate in middle called germinal center

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Spleen (Secondary Lymphoid Organ)

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1. traps Ag from blood
2. splenic artery carries Ag to spleen
3. microscopically divided into: red pulp and white pulp which is divided into PALS and marginal zone
4. red pulp: site where old and defective RBCs are destroyed and removed
5. white pulp: composed primarily of WBCs
-surrounds the splenic artery
-PALS: periarteriolar lymphoid sheath; populated with T lymphs; contains primary follicles (B cells)
-marginal zone: contains lymphocytes, macrophages, and dendritic cells; contains primary follicles (B cells)

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General Mechanism in Spleen

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1. Ag carried by blood to spleen via splenic artery
2. Ag is then carried onto marginal zone of the spleen
3. dendritic cells bind and process Ag
4. dendritic cells carry Ag peptide to the PALS region where they activate TH cells
5. activated TH cells then activate B cells
6. activated TH and B cells migrate to the primary follicles in the marginal zone
7. B cells of the primary follicles become activated and the follicle is now termed a secondary follicle with germinal center

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MALT: Mucosal Associated Lymphoid Tissue (Secondary Lymphoid Organ)

Back 87

1. Tissue found in:
-respiratory tract
-GI tract
-appendix
-tonsils
-Peyer's patches (intestinal)
2. contains largest population of plasma cells
3. epithelial cells of mucous membranes trap and deliver Ag to underlying MALT tissue
4. M cells: specialized epithelial cells that perform this task; these lack microvilli and contain pockets of cells containing B and T lymphs and DCs

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General Mechanism in MALT (in M cells)

Back 88

1. M cells endocytose Ag (vesicles + Ag)
2. vesicles move to pocket
3. Ag released from vesicle into pocket
4. Ag activates pocket cells
5. immune response elicited and Ag eliminated
OR
1. Ag endocytosed by M cells (vesicles + Ag)
2. vesicles move to underlying MALT tissue
3. Ag released from vesicle to MALT
4. Ag activates cells (lymphoid follicle and B cells)
5. plasma cells secrete Ab
6. immune response elicited

-side note: intraepithelial lymphs also act to trap Ag as it moves across eppithelium as a backup for M cells

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Innate Immunity

Back 89

-1st line of defense against pathogens
-prevent Ag from entering or remove quickly if enters

Front 90

Innate Recognition and Elimination

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1. innate molecules can discriminate between self and non-self
2. innate molecules recognize highly conserved molecules or structural motifs on pathogens called PAMPs (pathogen-associated molecular patterns) which are sugars, proteins, or lipids
3. innate molecules called pattern recognition receptors (PRRs) recognize and bind to PAMPs found on pathogen; PRRs may be soluble or membrane-bound

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Soluble Molecules (PRRs)

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-not hydrophobic, free-floating
-includes:
1. mannose-binding lectin
2. antimicrobial peptides
3. C-reactive protein
4. complement
5. lipopolysaccharide-binding protein

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Mannose-Binding Lectin (MBL)

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1. bind to mannose-containing PAMPs found on the pathogen
-indirect elimination, can't do it alone
2. activates the complement system
-composed of proteins that can bind to pathogen → embed itself in membrane → weaken/create holes → cell swells → bursts/lyses
-or these proteins can activate phagocytosis

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Antimicrobial Peptides

Back 93

1. examples: defensins (disrupt bacterial mebrane), cathelicidins (disrupt bacterial membrane and inhibit protein synthesis), interferon (blocks viral entry), etc.
2. attack bacteria, fungi, and viruses
3. How do they induce elimination of pathogens?
-in general, antimicrobial peptides may:
-disrupt membrane
-inhibit DNA/RNA/protein synthesis
-activate other antimicrobial peptides
-block entry

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Defensins

Back 94

-recognize and bind to bacteria
-very small proteins (29-35 AAs)
-highly positively charged (basic)
-some of the AAs are cysteines (S), which want to form disulfide bonds, which stabilize the folded structure
-secreted by a variety of cells (ex. granules of neutrophils make and secrete defensins)
-fast response: once bind to bacteria, can eliminate Ag within minutes
-mechanism: positively charged defensins bind closely to negatively charged membrane of bacteria → binding disrupts membrane lipid bilayer → embedded → weaken/create holes in membrane → lyse

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C-Reactive Protein (CRP)

Back 95

1. binds to C protein found on particular bacteria
2. activates the complement system
-disrupts the membrane → weaken → lyse
-acts as an opsonin → increase phagocytosis
3. acute phase protein (sudden inflammatory response)
-cytokines from inflammation travel to liver and tell it to increase synthesis of acute phase proteins; CRP is one, so CRP increased with inflammation
4. inflammation → cytokines → blood → liver → increased synthesis of acute phase proteins like CRP

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Complement

Back 96

1. lyses the pathogen
2. increases phagocytosis (opsonin)
3. acute phase protein

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Lipopolysaccharide-Binding Protein (LBP)

Back 97

1. binds LPS of gram negative bacterial cell walls
2. acts as an opsonin to increase phagocytosis

Front 98

Membrane-Bound Molecule (PRR)

Back 98

-must have hydrophobic AA region in order to stick in membrane; usually sticks out on either end
-major example: TLR

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Toll-Like Receptors (TLR)

Back 99

1. found primarily on dendritic cells and macrophages
2. receptor divided into:
-extracellular domain: LRR = Leucine Rich Region; sequence = XLXXLXLXX; this is the part that binds to the pathogen
-membrane domain: hydrophobic core of AAs that functions to anchor receptor to membrane
-intracellular domain: TIR because similar to Toll Interleukin-1 Receptor; binds to intracellular proteins to induce signal transduction pathways which is a domino effect until desired effect is produced
3. found on external and internal membranes
4. How does TLR induce elimination of pathogen? TLR signal-transduction mechanism

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TLR Signal-Transduction Mechanism

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1. Ag binds to extracellular domain of TLR
2. intracellular proteins (adaptor proteins) with TIR domains bind to the TIR domain of the TLR (ex. MyD88)
3. promotes association of IRAK1 and IRAK4 (protein kinases) to the TLR
4. IRAK4 becomes activated when bound to TLR, and then phosphorylates IRAK1
5. TRAF6 binds to phosphorylated IRAK1
6. TAK1 (protein kinase) binds to IRAK1:TRAF6 (complex)
7. TAK1 phosphorylates proteins of the MAP kinase and NFkB systems
8. Leads to:
-increased expression of inflammatory proteins like cytokines, complement, or other PRRs
-alteration of APCs, thus increasing their ability to present Ag (because the proteins help activate APCs)
-activation of phagocytes (because the proteins are opsonins)

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TLR 1 and TLR 2

Back 101

bacterial parasites; on cell membrane

Front 102

TLR 2 and TLR 6

Back 102

gram-positive bacteria and fungi; on cell membrane

Front 103

TLR 4

Back 103

gram-negative bacteria; on cell membrane

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TLR 5 and TLR 7

Back 104

flagellated bacteria; on cell membrane

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TLR 3

Back 105

viral dsRNA; on internal compartment

Front 106

TLR 7

Back 106

viral ssRNA; on internal compartment

Front 107

TLR 8

Back 107

viral ssRNA; on internal compartment

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TLR 9

Back 108

bacterial DNA elements; on internal compartment

Front 109

NFkB Pathway

Back 109

1. TAK1 phosphorylates IKK
2. IKK phosphorylates IKB while in complex with NFkB
3. complex dissociates
4. NFkB moves to nucleus
5. NFkB acts as a TF
6. genes transcribed to mRNA
7. mRNA leaves nucleus and translated to proteins
8. Proteins directly/inderectly lead to the elimination of the pathogen

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MAPK Pathway

Back 110

1. TAK1 phosphorylates and thus activates protein kinase Raf
2. Raf phosphorylates/activates protein kinase MEK
3. MEK phosphorylates/activates protein kinase ERK
4. active ERK moves into nucleus and phosphorylates TF
5. TF leads to transcription of particular gene → mRNA → translated into protein
6. protein leads to elimination of pathogen

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Extravasation Definition and Mechanism

Back 111

-process by which WBC moves across blood vessel into tissue
-mechanism:
1. cytokines and other factors secreted at the infection site bind to endothelial cells
2. induces an increased expression of CAM (cell adhesion molecules) proteins on blood vessel membranes
3. neutrophils (all WBCs really, but neutrophils are 1st) bind to endothelial cells because the neutrophil membrane protein mucin weakly binds to selectin protein on endothelial cells
4. neutrophils can move or "roll" along endothelial cells as they move from one selectin to the next
5. factors released at the infection site called chemokines bind to neutrophil
6. induces a conformational change that increases the affinity of the neutrophil membrane protein integrin for the blood vessel protein called ICAM (intracellular adhesion molecule)
7. neutrophil moves or "rolls" along blood vessel membrane until its integrin molecule binds to an ICAM on blood vessel membrane (strong bond)
8. neutrophil moves between endothelial cells into tissue

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Major Factors Secreted at Injury/Inflammation Site

Back 112

1. acute phase proteins
2. cytokines
-produced by WBCs and other immune cells
-hormone or growth factor-like proteins
-binds to target site and activates target site
3. chemoattractants
-attract WBC to infection site
-example of a chemokine
-example: complement

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How does a WBC know where to migrate?

Back 113

-it travels toward chemoattractants released at the injury site

Front 114

Major Innate Immunity Cells

Back 114

1. Neutrophils
2. Macrophages
3. Dendritic Cells
4. Natural Killer Cells

Front 115

Pathogen Elimination via Innate Immunity Cells

Back 115

-phagocytosis
1. primary defense
2. oxidative attack
-actually number 1 way to degrade pathogen
-during phagocytosis, increase of oxygen uptake by the cell, called respiratory burst
-this activates enzyme Phos
-production of reactive oxygen species (ROS), which damage DNA, oxidize lipids and AAs to make them nonfunctional; these will make holes in cell wall of pathogen and lead to lysis
-also, production of reactive nitrogen species (RNS): increased oxygen → activate Nos enzyme → NO + superoxide anion → reactive nitrogen species → damage cell walls
3. non-oxidative attack
-phagolysosome hydrolytic enzymes used to process pathogens
-typically secondary to oxidative attack

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ROS

Back 116

1. superoxide anion - ‧O₂⁻
2. hydrogen peroxide - H₂O₂
3. hypochloric acid - HOCl

Front 117

The recognition between antigen and Ab or TCR is determined by their:

Back 117

1. conformation
2. chemical properties

Front 118

Cross-Reaction

Back 118

antibodies can recognize distinct molecules on antigen(s) with similar configurations

Front 119

Immunogenicity

Back 119

1. ability to induce a humoral or cell-mediated response
2. Ag (immunogen) + B cell → memory B + effector B → plasma cell → secrete Ab (humoral)
3. Ag (immunogen) + T cell → memory T + effector T → TH (→ cytokines) + Tc (→ cytotoxic factors) (cell-mediated)

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Immunogen

Back 120

1. induce an immune response
2. all immunogens are antigens
3. all antigens are NOT imunogens

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Haptens

Back 121

-lack immunogenicity
-small molecule; body can see that they are foreign, so they are antigens
-but too small to produce an immune response, so not an immunogen

Front 122

Antigen vs. Immunogen

Back 122

-antigen = foreign; can be eliminated by binding to Ab or TCR, though they may not be able to induce formation of Ab and TCR
-immunogen = elicits an immune response

Front 123

Landsteiner's Experiment: How do we know that a hapten can induce an immune response when bound to a carrier?

Back 123

1. coupled hapten to a carrier:
-hapten = DNP (v. small)
-carrier = BSA (large)
-hapten + carrier = DNP + BSA
2. took each of these solutions and injected them into rabbits
3. allowed an immune response
4. obtained blood samples
5. analyzed Ab levels (looked for Ag+Ab agglutination)
6. results:
-DNP → none
-BSA → anti-BSA
-DNA + BSA → anti-DNP (most), anti-BSA (little), anti-DNP/BSA (little)
-demonstrated that DNP (haptens), with increased size are actually very immunogenic

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Antigenicity

Back 124

ability to bind with products of the humoral or cell-mediated responses (Ab, TCR)

Front 125

Immunogenicity vs. Antigenicity

Back 125

-a substance that is antigenic is not necessarily immunogenic; example is a hapten which can bind to antibodies but is too small to induce an immune response
-a substance that is immunogenic is also antigenic; example: bacteria, parasites, fungi

Front 126

Four Properties that Determine a Good Immunogen

Back 126

1. Foreigness
2. Molecular Size
3. Chemical Composition and Heterogeneity
4. Processing and Presentation of Antigen (Degradability)

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Foreigness (of Immunogens)

Back 127

1. to induce an immine response, the substance must be recognized as a non-self antigen
2. also needs tolerogenicity (unresponsiveness) to Ag (especially self Ag)
3. the greater the phylogenetic distance between two species, the greater the structural differences, and structural differences are a factor in determining immunogenicity
4. Example: injecting BSA into different animals and measuring amount of anti-BSA produced
-cow: none; low immunogenicity because BSA is from same species, so not viewed as foreign
-goat: some; cow and goat are closely related therefore only a small response
-chicken: abundant; cow and chicken are distantly related, therefore large response

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Molecular Size (of Immunogens)

Back 128

1. size and immunogenicity are directly proportional (increased size = increased immunogenicity)
2. generally 100,000 Da elicits an immune response

Front 129

Chemical Composition and Heterogeneity (of Immunogens)

Back 129

1. Immunogenicty is also dependent upon protein organization (primary, secondary, tertiary, quaternary structures and AA content)
2. example:
-homopolymer: all glycine; no Ab production; one AA is too simple to elicit an immune response
-heteropolymer: 2 AAs; glycine and lysine; some Ab production; different AAs increases immunogenicity
-heteropolymer with bulky AAs: glysine, lysine, and phenylalanine; larger response; different AA structures increase immunogenicity

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Processing and Presentation of Antigen (Degradability of Immunogens)

Back 130

1. robust humoral and cell-mediated response requires processing and presentation of Ag to the T cells
2. if a molecule (Ag) cannot be degraded and presented on MHC, it is a poor immunogen

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What factors determine how much each individual will respond to an immunogen?

Back 131

1. Genotype of Recipient
2. Immunogen Dosage
3. Route of Administration
4. Usage of Adjuvant (Freud's - First Developed)

Front 132

Genotype of Individual (as factor for individual response to an immunogen)

Back 132

1. determined by several different genes
2. example: MHC plays a major role in determining the degree of an immune response because MHC gene products (class I and II) present processed antigen to TH and Tc cells

Front 133

Immunogen Dosage (as factor for individual response to an immunogen)

Back 133

1. insufficient dose will not stimulate a response:
-no or little activation of lymphocytes (under threshold responsiveness)
-induce a state of tolerance (unresponsiveness)
2. high doses can also induce tolerance
3. single dose most often would not induce a strong response
4. increased # of doses will induce a strong response because more activatoin of lymphs; "boosters"

Front 134

Route of Administration (as factor for individual response to an immunogen)

Back 134

1. route influenced by where the antigen needs to go
2. routes:
-intravenous = vein → blood → spleen
-intradermal = skin → tissue → lymph → lymph node
-subcutaneous = beneath skin → tissue → lymph → lymph node
-intramuscular = muscle → tissue → lymph → lymph nodes
-intraperitoneal = peritoneal cavity → tissue → lymph → lymph node
-oral = Peyer's patches → MALT

Front 135

Usage of Adjuvant (as factor for individual response to an immunogen)

Back 135

1. adjuvants
-substances added to an Ag to enhance the Ag's immunogenicity (carrier proteins)
2. possible mechanisms:
-prolonged antigen persistence
-enhance co-stimulatory signals
-induce local inflammation (stimulate APC)
-non-specifically stimulates lymphocyte proliferation

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(Antigen) Epitopes

Back 136

region of an immunogen or antigen that can bind to an Ab or TCR

Front 137

B Cell Epitopes

Back 137

1. B cells recognize free/soluble Ag
2. epitope composed of protein, polysaccharide (carbohydrate), or lipid
3. typically highly accessible sites on exposed surface
4. composed typically of hydrophobic AAs
5. sequential epitopes (2, 3, 4, etc. AAs on protein)
6. non-sequential epitope (AAs from different parts of a chain)
7. flexible/mobile: epitopes within regions that are flexible allows the epitope to fit/match TCR or Ab
8. immunodominant: epitope which induces a more pronouned immune response than the other epitopes

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T Cell Epitopes

Back 138

1. T cells recognize Ag peptides bound to MHC
2. composed of proteins
3. T cells epitopes are typically:
-internal (hydrophobic); within the Ag itself
-sequential