Lymphocytes

a basic for immunology

1. LYMPHOCYTES DR. SHILPA SONI MGMCH, JAIPUR

2. Basics of immune system

3. Lymphoid organs

4. Sites of lymphocytic cells

5. Cells of lymp.retic syst Lymphocytes- T- lymph B- “ Null cells/large granular lymphocytes -ADCC-antibody dependent cell mediated cytotoxic lymp . -NK cells Reticular sys cells- Phagocytic cells- macrophages & microphages Macrophg - monocytes (in Bd ) largest of lymp cells. - Histiocytes (in tissues)-microglia in CNS - Kupffer cells in liver -alveolar macrophages in lungs - osteoclasts in bone - sinushistiocytes in spleen. LN Microphg - polymorphnuc cells of Bd - N,E & B. Dendritic cells- dentritic , langerhans , follicular dendritic cells

6. MHC Located at short arm of Chr-6, codes for histocompatibility (transplantation )Ag. Binds peptide fragments of foreign Pr. for presentation to appr . Ag specific T cell. Classified as-

7. Lymphocytes Human body contains 10¹² lymphocytes out of which 10^9 are renewed daily. Mature B & T cells before encountering Ag are K/A Naïve cells. Types- T lymphocytes - B “

8. Normal Percentage Lymphocytes: 15-40% of  White Blood Cell s Total Lymphocytes: 800-2600/mm ³ Total T Lymphocytes: 800-2200/mm ³ T helper Cells: >400/mm ³ T suppressor Cells: 250-750/mm ³ Helper Cell to Suppressor Cell ratio: >0.9 CD2 Percentage of Lymphocytes: 65-85% CD4 Percentage of Lymphocytes: 45-75%

9. T cells Thymus derived lympc - 60-70% of periph lymphoc . Found in- paracortical areas of LN. - periarteriolar sheeths of spleen. Ag + TCR -> signal 1 All T cells contain CD 3 molecule->signal 1 Surface mol or receptors of T cells- CD 2 CD 4 – 60% of T cells CD 8 – 30% of T cells ( cytotoxic ) CD 11a CD 28 – binds to B 7-1(CD 80) & 7-2(CD 86) of APCs -> signal 2 CD 40

10. T Cell Subsets CD8/CTL CD4/T H cells 1) T H 1 - inflammatory response 2) T H 2 - anti-inflammatory, B cell response 3) T reg - inhibit immune responses

11. T-lymphocytes

12. T cells: Development in thymus Migration out of thymus into blood (Naïve T cells) Priming phase: Antigen-dependent differentiation in lymphoid tissues (to become m emory and effector T cells) Effector phase: Migration to sites of infection and effector function Human: Fetal development Birth School Job

13. Immunology’s Secret” Early experiments demonstrated that antigen-derived (i.e. TCR-mediated) signals alone are insufficient to initiate an immune response A second substance - an “adjuvant” - is required to prevent the induction of tolerance In vitro, triggering the TCR alone also leads to a tolerant state (known as “ anergy ”) “Two-Signal Hypothesis”: Activation of a naïve T cell requires signals from both the TCR (antigen-specific) and a second, co-stimulatory receptor (antigen-independent) Adjuvants work in part by inducing antigen presenting cells to express ligands for co-stimulatory receptors

14. Figure 6-1

15. Figure 6-2 Green: MHC class II Red: Lysosomal protein

16. Polarization of T Cells Part I: The Cytoskeleton T cell responses are directed at the APC/target cell, not in all directions This requires reorganization of the cell to have a “front” (toward the APC) and a “back” - induced by signals from the TCR and costimulatory molecules Result: Reorganization of the cytoskeleton causes reorientation of cytosolic organelles toward APC - Golgi, secretory vessicles , and microtubule organizing center (MTOC). The MTOC connects actin cytoskeletal changes with the tubulin cytoskeleton

17. Fig. 8.29 The polarization of T cells during specific antigen recognition

18. Polarization of T Cells Part II: Lipid Rafts Lipid rafts - also called GEMs ( glycolipid enriched microdomains ) and DIGs (detergent-insoluble glycolipid -rich domains) - are plasma membrane microdomains Enriched in cholesterol, glycolipids , and sphingolipids (i.e. lipids that are not glycerol-based), making them more rigid than the surrounding membrane Some membrane proteins are segregated - selectively enriched or depleted in rafts. Many key signaling molecules (esp. src -family kinases ) are constitutively localized to lipid rafts. Dynamic structures - small rafts can condense to form larger rafts During T cell activation, TCR, CD28, and many adhesion molecules are recruited into lipid rafts, where they can interact with signaling molecules

19. The T cell antigen receptor V a V b C a C b Carbohydrates Hinge Transmembrane region Cytoplasmic tail + + + Antigen combining site

20. The Immunologic Synapse - Putting it Together The combination of cytoskeletal rearrangement and lipid raft redistribution leads to the formation of a Supramolecular Activation Complex (SMAC) - a highly ordered arrangement of receptors, adhesion molecules, and signaling molecules

21. Th2 Additional Figures

22. t cells and their cytokines

23. T - lymphocyte in various dermatosis

24. Pppppppppp psoriasis The disorder is mediated largely by CLA( cutaneous lymphocyte associated antigen)-positive, CD8+ T cells with type 1 cytokines (interferon- g, interleukin- 2, and lymphotoxin ); these cells may be activated by an autoantigen in skin

25. Allergic contact dermatitis It is mediated by CLA-positive, CD8+ effector T cells that recognize contact-sensitizing antigens. The activated T cells have a variable cytokine profile (e.g., both type 1 and type 2 cytokines).

26. Atopic dermatitis initiated by CLA-positive, CD4+ T cells with type 2 cytokines (interleukin-4, 5, 10, and 13). T cells that produce type 1 cytokines may be involved in persistent lesions

27. Cut. T cell lymphoma The transformed T cells are found throughout the dermis and in the epidermis. Many reactive ( nontransformed ) CLA-positive T cells are also present in lesions.

28. Graft .vs. host rxn CLA-positive T cells producing type 1 cytokines (in acute disease) or type 2 cytokines (in chronic disease) are present in lesions.

29. Other T cell- skin disorders -LP -Eczemas - Pneumocystis jiroveci pneumonia and cryptosporidiosis.

30. Deffects of t cells T-cell defects -Wiskott–Aldrich syndrome ( WASP gene mutations) -Ataxia telangiectasia ( ATM mutations) -Chronic mucocutaneous candidiasis (AIRE, FoxP3 deficiency) -X-linked hyper- IgM (CD40 ligand deficiency); autosomal hyper- IgM syndrome (multiple defects) -Cartilage–hair hypoplasia -Idiopathic CD4+ T-cell lymphopenia

31. B lymphocytes 10%- 20% Of periph lymphoc Humoral immunity Presents in BM, peripheral lymphoid tissues-superficial cortex of LN, white pulp of Spleen, tonsils & extra Lymphatic organs e.g GIT Responds to free Ag Acts as APC Ag binding compon-IgM

32. Cont.. Other molecules are complement receptor, Fc receptors, CD 21 – recept for EBV, CD 40-essential for intraction of T & B cell -> B cell maturation (mutation in CD 40 ligand cause immunodeficiency called X-linked hyper IgM syndrome.

33. B cell development in the bone marrow B Regulates construction of an antigen receptor Bone Marrow provides a MATURATION & DIFFERENTIATION MICROENVIRONMENT for B cell development Ensures each cell has only one specificity B Checks and disposes of self-reactive B cells B Exports useful cells to the periphery B Provides a site for antibody production B

34. B-lymphocytes in bone marrow * The lymphoid stem cells differentiate into B cells * B-cells precursors mature, differentiate into immunocomptent B-cells with a single antigen specificity * Immature B-cells that express high affinity receptors for self antigens, die or fail to mature i.e negative selection or clonal deletion * This process induces central self tolerance and reduces autoimmune diseases

35. B B Stromal cell

36. Bone marrow stromal cell Maturing B cells

37. Stages of differentiation in the bone marrow are defined by Ig gene rearrangement B CELL STAGE IgH GENE CONFIGURATION Stem cell Early pro-B Late pro-B Large pre-B Germline D H to J H V H to D H J H V H D H J H Pre-B cell receptor expressed Ig light chain gene has not yet rearranged

38. B-lympocytes * Immature B cells - IgM receptors on the surface * Mature B cells - IgM , IgD molecules on surfaces IgM and IgD molecules serve as receptors for antigens * Memory B-cells - IgG or IgA or IgE on the surface * B-cells bear receptors for Fc portion of IgG and a receptor for C3 component of the complement * They express an array of molecules on their surfaces that are important in B-cells interactions with other cells such as MHC II, B7 and CD40

39. Activation of B cells to make antibody

40. Antibodies or immunoglobulins Structure & functions Definition: Glycoprotein molecules that are produced by plasma cells in response to an immunogen and which function as antibodies. * Produced by: B-lymphocytes in response to exposure to antigen * React specifically with antigen * Five classes of Antibodies: IgG IgM IgA IgD IgE

41. Antibody Structure Immunoglobulins are glycoproteins made up of - Four polypeptide chains ( IgG ): a- Two light (L) polypeptide chains b- Two heavy (H) polypeptide chains - The four chains are linked by disulfide bonds - Terminal portion of L-chain contains part of antigen binding site - H-chains are distinct for each of the five immunoglobulins - Terminal portion of H-chain participate in antigen binding site - The other (Carboxyl) terminal portion forms Fc fragment

42. Variable(V) and Constant (C) Regions - Each H -chain and each L -chain has V- region and C- region - V- region lies in terminal portion of molecule - V- region shows wide variation in amino a. sequences - Hypervariable region form region complementary to Ag determinant - It is responsible for antigen binding - C- region lies in carboxyl or terminal portion of molecule - C- region shows an unvarying amino acid sequence - It is responsible for biologic functions

43. Ab fragments & their functions Fab fragment: antigen binding site Fc ( crystallizable fragment): a- Complement fixation ( IgM and IgG ) b- Opsonization ( IgG ) C- Placental attachment ( IgG ) d- Mucosal attachment ( IgA ) e- Binding to mast cells ( IgE )

44. Struct of diff. imunog .

45. Properties of Immunoglobulins Property IgG IgA IgM IgE IgD Heavy chain symbol γ α µ ε δ Molecular weight 150 KDa 170-400 KDa 900 KDa 190 KDa 180 KDa Percentage in serum 75 % 15 % 10 % 0.004 % % 0.2 Complement fixation Yes No Yes No No Transplacental passage Yes No No No No Opsonization Yes No No No No

46. Primary and Secondary antibody response Primary antibody respone Secondary antibody response * first exposure to antigen * Subsequent exposure * lag period: days or weeks * Lag period: hours * Small amount immunogl . * large amount immunogl . * in Weeks then decline * Persist for long periods * Antibody is IgM * Antibody is IgG

47. B - cell deffects -X-linked agammaglobulinaemia - μ- chain deficiency; surrogate light-chain deficiency -SWAP-70 deficiency -Hyper- IgE syndrome (includes secondary neutrophil disorder) -CD79a ( Ig α chain) deficiency -BLNK deficiency -ICOS deficiency -TACI deficiency

48. B cell lymphoma

49. Differences between T & B lymphocytes. T cells B cells Ab binding site Ag rec (TCR with CD3) Surface Ig Fc receptor — + Complement receptor - + EAC rosette- C 3, CR 2, EBV rec - + E/SRBC rosette- Cd 2, measles rec + - Microvilli on surface - + Thymus specific Ag + - Blast transformation Occurs by anti CD 3, phytohemagglutinin , concanavalin Occurs by anti Ig endotoxin , staph aureus , EBV

50. Combined deffects -Severe combined immunodeficiency ( RAG-1/RAG-2 mutations, common γ-chain deficiency; CD3 ε deficiency) -Adenosine deaminase deficiency - Purine nucleoside phosphorylase deficiency DNA repair defects (DNA ligase IV); Bloom’s syndrome; xeroderma pigmentosum - Canale –Smith syndrome (autoimmune lymphoproliferative syndromes; fas , fas - ligand , caspase deficiencies) -X-linked lymphoproliferative syndrome (SAP deficiency; includes NK disorder)

51. Thank you

52. Increased ( Lymphocytosis ) Increased Absolute Lymphocyte Count (>4500/mm3) Non-activated Lymphocytes Influenza Pertussis Tuberculosis Mumps Varicella Herpes Simplex Virus Rubeola Brucellosis Chronic Lymphocytic Leukemia Acute  Lymphoblast ic  Leukemia

53. Activated Lymphocytes ( Atypical lymphocyte s) Cytomegalovirus  Infection Infectious  Mononucleosis Infectious Hepatitis Toxoplasmosis Post-transfusion Medication Mephenytoin Dilantin Para- aminosalicylic acid

54. Increased Relative Lymphocyte Count Normal finding in children under age 2 years Acute stage of viral infection Connective tissue disease Hyperthyroidism Addison's Disease Splenomegaly

55. Decreased AIDS Bone Marrow  suppression Aplastic Anemia Neoplasms Steroids Adrenocortical hyperfunction Neurologic Disorders Multiple Sclerosis Myasthenia Gravis Gullain Barre Syndrom