Adaptive Immunity and the Lymphoid System

Immune system Lymphatic system Computerized war machine Defense department of the body Cells of the immune system Myeloid lineage Lymphoid lineage Major Functions of the Immune System Block harmful agents Seek-out invasive pathogens Isolate and neutralize activity of that antigen Acquired immunity Type of resistance that is characterized by specificity for each individual pathogen and the ability to remember a prior exposure, which results in an increased response upon repeated exposure. Lymphocytes The key cell involved in adaptive immunity Represents approximately 20% of the circulating WBCs Has a large rounded nucleus which contains a dense nuclear chromatin 2 Major Stages of Lymphocyte Development 1. Antigen Independent Stage of Lymphopoiesis 2. Antigen Dependent Stage of Lymphopoiesis THE LYMPHOID SYSTEM Primary/Central Lymphoid Organs (developmental sites) Bone Marrow Thymus Secondary/Peripheral Lymphoid Organs (activation sites) Encapsulated organs spleen, lymph nodes Non-encapsulated organs MALT, BALT, GALT, CALT BONE MARROW Main source of hematopoietic stem cells Center for antigen-independent lymphopoiesis THYMUS Small, flat bilobed organ found in the thorax or chest cavity right below the thyroid gland and overlying the heart An endocrine gland Cortex where thymocytes can be found (85% of population of T cells) Medulla where mature T cells can be found Thymic Stromal cells include epithelial cells, macrophages and dendritic cells SPLEEN Largest secondary lymphoid organ that is found in the upper lower quadrant of the abdomen just below the diaphragm 2 Main types of Splenic Tissue Red Pulp involved in culling process White Pulp contains lymphoid tissue that is arranged around arterioles as PALS (Periarteriolar Lymphoid Sheath) o o T cells are found near the central arteriole Nave B cells are found on primary follicles

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Activated B cells are found on Secondary Follicles (germinal centers) Marginal zone contain macrophages

Sites of entry of Lymphocyte/Antigen into Spleen Specialized Capillaries on Marginal Sinus Via Trabecular Artery LYMPH NODES Junctional filter of the lymphoid system Sizes may range from 1mm 25mm Cortex B cell area; also contains primary follicles which small amount of T-cell and secondary follicles that contains germinal center Paracortex T-cell area Medulla contains differentiated cells and APCs Lymphocyte traffic/recirculation The movement of lymphocytes from the blood to lymphoid organs and back to the blood. Site of entry and exit of lymphocytes in the lymph node Afferent Arteriole Efferent Arteriole Post Capillary Traffic Endothelial Venule/High-walled Endothelial Venule MALT Mucosal Associated Lymphoid Tissue Small masses of lymphoid tissue found in: Intestinal Genitourinary tract Respiratory tract Through this system T and B cells are able to find its way to the lymph nodes Has addresins and other adhesion molecules CD32, CD 102 markers T and B-cells have homing receptors L-selectin or LAM

GALT BALT CALT Cutaneous Associated Lymphoid Tissue Intraepidermal lymphocytes Bronchus Associated Lymphoid Tissue Tonsils Adenoids Gut Associated Lymphoid Tissue Peyers patches (intestines) Appendix

Surface Markers Membrane proteins to differentiate T and B cells Detected through monoclonal Antibody test Distinguishes the developmental stage of the lymphocyte Cluster of Differentiation - reference in standardizing the membrane proteins found in the surface of T and B cells

Stages of B-cell differentiation 1. Pro-B cells markers include: CD19, CD45R

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TdT RAG-1, RAG-2 enzymes

Pre-B cells contains pre-B cell receptor that is made up of 2 heavy chains and a surrogate light chain whose function is to transmit signals to prevent rearrangement of any other heavy chain genes Immature B-cells recognized by the appearance of complete IgM molecules on their surface. Specificity of the surface immunoglobulin to be synthesized can already be predicted or noted. Surface markers that can be seen include receptors for complement components such as C3d, CD21 B-cells capable of producing antibody to selfantigens are deleted in the marrow through Apotosis Mature B-cells exhibit IgD and more IgM on their surface as well as MHC class II products Activated B-cells exhibit CD25 on surface. CD25 in turn acts as receptor to IL-2 Plasma cells are large spherical, ellipsoidal cells that contain abundant cytoplasmic immunoglobulins and little to no surface immunoglobulins

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Natural Killer cells Larger than T and B cells; contains a kidney shaped nucleus with condensed chromatin and prominent nucleoli Has high ctyoplasmic:nuclear ratio Cytopalsm has many azurophilic granules Constitutes 5-15% of the circulating lymphoid pool; found mainly in spleen and blood No specific surface markers Posess CD16, CD56, CD94 Lacks CD3, CD4 and CD8 Play a complementary role to CD8+ T cells Become LAK cells in response to IL-2 Mechanisms of NK cell Cytotoxicity It is brought about by the balance between activating and inhibitory signals that enables NK cells to distinguish healthy cells from infected or cancerous cells oKiller cell inhibitory receptors (KIRs) oNKG2D binds to MICA and MICB proteins on diseased or cancerous cells oIf an inhibitory signal is not produced, NK cells will release Perforins and Granzymes Antibody Mediated Cell Cytotoxicity Through binding of IgG-coated cell with CD16

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CD19 present in all population of B-cells Stages of T-cell Differentiation 1. 2. Pro-thymocyte possess CD44 and TdT Double-Negative Thymocyte lacks CD4 and CD8 antigens Possess CD2, CD5, CD7, Cd45R Double Positive Thymocyte Express both CD4 and CD8 antigens on their surface as well as CD3-a (TCR) Would undergo Positive and Negative Selection Mature T-cells represents those population of thymocytes that had survived positive and negative selection Express only 1 of either CD4 or CD8 on their surface 2/3 CD4+ 1/3 CD8+ Activated T-cells Express receptors for IL-2 and produce cytokines T memory cells

Laboratory Identification of Lymphocytes 1. Density Gradient Centrifugation with Ficoll-Hypaque 2. Fluorescence Microscopy Direct Immunofluorescence Use monoclonal antibodies with a fluorescent tag fluorscein and phytoerythrin (490nm) rhodamine (545 nm) Indirect Immunofluorescence Uses unlabeled antibody that first combines with the antigen by itself and a secongd antibody that is complexed with a dye

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3. Cell flow Cytometry/Fluorescence Activated Cell Sorter An automated system for identifying cells based on the scattering of light as cells flow in single file through a laser beam Fluorescent antibodies are used to screen of subpopulation of T and B cells

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Components: sample delivery system a laser for cell illumination photodetectors for signal detection computer based management system 4. Rosetting 5. ELISA