Introduction to Immunology

1. Introduction to Immunology

2. I. General Introduction

3. A. Definitions • Immunity: the state of protection from infectious disease involving specific and non-specific elements • Specific immunity (aka aquired immunity) employs components of the immune response that specifically recognize and selectively eliminate microorganisms and molecules perceived as foreign by the host

4. Nonspecific immunity (aka innate immunity) utilizes the basic resistance to disease that a species possesses and makes up the first line of defense • Antigen is anything that the adaptive immune response (IR) can recognize (antibody generating)

5. B. The Origin of Immunology • Edward Jenner (1796) used the cowpox virus (vaccinia) to confer induced protection fro human small pox • Vaccination – term now used to explain how healthy subjects are inoculated with attenuated (weakened) strains of pathogens to induce ACQUIRED protection (Active immunization) • Small pox successfully eradicated in 1979 – Announced by the WHO

6. Robert Koch (1843-1910) proved that bacteria were responsible for causing anthrax and tuberculosis. • He developed Koch’s postulates • A set of criteria to be used when establishing a causative link between a particular microorganism and a particular disease • Koch’s postulates are still followed today to show that pathology (disease) is caused by one of the four major groups of pathogens: • Viruses • Bacteria • Pathogenic fungi • Parasites

7. Louis Pasteur (1822-1895) – developed vaccines against cholera (Vibrio cholerae) and rabies (Rhabdovirus –Negribodies – RNA virus) • Emil von Behring (1845-1917) and Shibasaburo Kitasato (1852-1931) – using diphtheria toxin, identified that serum of vaccinated individuals contained antibodies that specifically interact with the immunogen (antisera, passive immunization when injected with immune serum)

8. Eli Metchnikoff (1845-1916) – reported the engulfment and degradation of microorganisms by a type of phagocytic cell he called macrophages • First line of defense • Innate (non-adaptive) immunity • Receptor-mediated endocytosis/phagocytosis

9. C. Major cells of Innate and Acquired Immune Responses • Involve action of white blood cells called leukocytes (lymphocytes, polymorphonuclear leukocytes and monocytes) and dendritic cells • Innate immunity • Involves mainly granulocytes • Granules in cytoplasm • Different cell types – many are phagocytic • Most are polymorphonuclear leukocytes = neutrophils (multilobed nuclei and cytoplasmic granules), eosinophils, basophils, mast cells (PMNLs) • Also involves monocytes (mononuclear)  Macrophages in tissues • Acquired (adaptive) immunity • Involves lymphocytes (B cells, T cells, Natural Killer Cells) • Lifelong immunity established through generation of memory cells (B and T cells have memory)

10. The Components of the Immune System

11. A. Hematopoietic Stem Cells Give Rise to White and Red Blood Cells • All circulating blood components originate in the bone marrow • The same precursor cell or progenitor gives rise to all of the various lineages = “Pluripotent Hematopoietic Stem Cell” • Differentiate or mature into two main progenitor populations • Lymphoid Lineage [B, T, and NK cells] • Myeloid Lineage [PMNL (Basophils, Mast cells, Eosinopils, Neutrophils), Monocytes/Macrophages • Erythroid Lineage (Megakaryocyte, Platelets and Erythrocytes) • Hierarachy of Cell Maturation: Pluripotent Stem Cells  Committed Progenitor Cells  Terminally Differentiated Cells

12. B. Maturation of Lymphocytes • Resting B and T cells • Small, inactive, heterochromatin,scantycytoplasm containing few organelles • No functional significance

13. Receptors to recognize specific antigen • B cell receptors • Membrane-bound antibody = surface immunoglobulin (Ig) • Following B cell activation  Differentiation into plasma cell  Cytoplasm enlarges, ER expands, active transcription, increase in organelles  Antibody secretion • T cell receptors • Related to Ig but distinctive • Following T cell activation (in peripheral lymphoid organ) • Cytotoxic T cells (Tcyt)  Kill infected target cells • Inflammatory (TH1) and Helper (TH2) T cells  Activation of other cells (macrophages and B cells, respectively)

14. Sites of Maturation  Central/Primary Lymphoid Organs • B cells mature in the bone marrow (or Bursa of Fabricius in birds) • T cells mature in the thymus • After maturation, lymphocytes are transported to the bloodstream and then traffic  to the Peripheral/Secondary Lymphoid Organs • This is where antigen is encountered • This is where lymphocytes divide (clonal expansion)

15. Adaptive IRs Occur in Peripheral Lymphoid Organs • Peripheral Lymphoid Organs • Lymph nodes • Spleen • Mucosal associated lymphoid tissue (MALT) • Bronchial associated lymphoid tissue (BALT) • Gut associated lymphoid tissue (GALT) • Tonsils • Adenoids • Appendix • Peyer’s patches • Function: to trap antigen from sites of infection and present it to circulating, resting lymphocytes to induce adaptive immune responses

16. Lymph nodes (LN) • The afferent lymphatic vessel delivers lymph draining the extracellular spaces of the body to the L.N. (e.g. in interstitial spaces of tissues) • Antigen (Ag) becomes trapped in the L.N. • The efferent lymphatic vessel takes lymph away from the L.N. medulla region

17. Anatomy of a lymph node • Post-capillary venules deliver naïve lymphocytes • Cortex • Outer cortex = B lymphocytes in 1o follicles and 2o follicles (germinal centers, where proliferation occurs if B cells are responsive to Ag) • Paracortex = T lymphocytes and dendritic cells • Medulla • Medullary cords = macrophages and plasma cells • Region where lymph leaves • Similar organization in spleen and Peyer’s patches. Important for T:B cooperation.

18. Spleen (Largest peripheral lymphoid organ) • Collects and traps Ag from the blood (via splenic artery) • Important for systemic infections • Not supplied by afferent lymphatics • Final stop for dying (senescent) RBC • Red pulp: major area and site of RBC disposal by splenic macrophages • White pulp forms around a central arteriole: • Periarteriolar lymphoid sheath (PALS) = Mainly T cells • B Cell corona and germina center • Arterioles  vascular sinusoids  splenic vein

19. D. Continuous Recirculation of Lymphocytes • Naïve lymphocytes are circulating continuously between blood and peripheral lymphoid regions • Homing to 2o lymphoid tissue • Involves binding to adhesion molecules on lymphocytes called L-Selectin, to its ligand called mucin-like vascular addressin (CD34 + GlyCAM-1 in L.N.; MAdCAM-1 in MALT) on high endothelial venules (HEV = capillaries delivering cells)

20. If Ag IS Encountered • 1. Ag enters L.N. through the afferent lymph (often via phagocytic cells) • 2. Ag is trapped by professional antigen presenting cells (APC) • 3. Ag is displayed to naïve lymphocytes • 4. Lymphocytes, which have a specific cell surface receptor that recognizes Ag, remain in peripheral lymphoid organ, proliferate, and then differentiate into effector cells • 5. Effector cells leave L.N. through efferent lymphatic vessel  return to blood via thoracic duct • 6. Emigration of WBC out of the bloodstream to sites of infection using adhesion molecules called integrins (extravasation) • T cells express increased levels of LFA-1, and begin to express VLA-4 once activated (binds ICAM-1 and VCAM-1, respectively) • Macrophages express MAC-1 (binds ICAM-1)

21. If Ag IS NOT Encountered • Lymphocyte leaves through efferent lymphatic vessel of 2o lymphoid • Returns to bloodstream via thoracic duct

22. Innate versus Adaptive Immunity

23. A. Distinction Between Innate and Adaptive Immune Responses • Innate immunity is non-adaptive and helps to initiate adaptive immune responses (= first line of defense – but LIMITED) • Immediate (0-4 hours) • Adaptive immunity provides a more universal line of defense and has long-lived memory to provide protection upon re-infection • Second line of defense • Generation of Ag-specific effector cells • Early (4-96 hours) • Late (>96 hours)

24. B. Innate Immune Responses • Innate defense is present in al individuals and can operate at various locations in the body

25. Seven types of defensive barriers • #1 – Anatomical Barriers • Skin: epidermis, dermis, keratin, sebum and other epithelial surfaces • Mucous membrane surfaces: saliva, tears, mucous secretions • #2 – Physiological barriers • Temperature, pH, O2 tension, soluble factors [lysozyme, interferons, acute phase proteins, complement, digestive enzymes, cytokines, chemokines, monokines (IL-1, IL-6, TNF alpha)]

26. Three Functions of Interferons • 1. Induce resistance to viral replication by activating cellular genes that: • Destroy viral mRNA • Inhibit translation of viral proteins • 2. Increase Major Histocompatibility Complex (MHC) Class I expression universally • Increases level of Ag presentation to Tcyt (CD8+) – cytotoxic T cells (aka killer T cells) • Increase resistance of uninfected cells to NK cell attack (more later) • 3. Activate NK cells to kill virus-infected cells

27. Acute Phase Proteins in Humans Act as Opsonins • Hepatocytes in liver produce APP in response to IL-1, IL-6 and TNF alpha • 1. Mannose binding protein (MBP) • Binds mannose residues on bacterial cells • Acts as an opsonin (enhances receptor mediated endocytosis by phagocytes) • Activates complement (lectin complement pathway) • Mimics activity of antibodies by acting as an opsonin and activating complement

28. 2. C-Reactive Protein (CRP) • Binds bacterial phosphorylcholine • Mimics activity of Ab by acting as opsonin and activating complement (classical pathway) • 3. Fibrinogen • Also an APP made by hepatocytes • Definition of opsonization: Alteration of the surface of a pathogen enabling its ingestion by phagocytic cells (neutrophils and macrophages) through RME. Examples: Ab, C’, CRP, MBP

29. #3 – Endocytic and Phagocytic Barriers • Endocytosis: Pinocytosis or receptor-mediated endocytosis or macromolecules (non-specific versus specific, respectively), followed by fusion with primary lysosomes where they are digested and processed (eliminated) • Phagocytosis: Ingestions of particulate material aided by microfilaments which fuse with lysosomes  phagolysosomes

30. #4 – Inflammatory Barriers • Major events: • Vasodilation • Increased capillary permeability • Influx of phagocytic cells • Vasodilation results in reduced blood flow velocity allowing leukocytes to move out of capillaries to vascular endothelium where they penetrate, resulting in accumulation of fluid (swelling, pain)

31. Macrophages produce MONOKINES which recruit more phagocytic cells and effector molecules to site of infection • IL-1, IL-6, IL-8, IL-12, TNF-alpha • Can also produce harmful, systemic effects • Systemic Shock + Disseminated Intravasclar Coagulation (DIC)  Organ Failure • Have a variety of effects at different locations

32. Inflammation – cont. • Other inflammatory mediators released/generated by macrophages and neutrophils include: • Plasminogen activator • Prostaglandins • Phospholipase • Platelet activating factor • Leukotrienes • Respiratory burst molecules: Nitric oxide, hydrogen peroxide, superoxide anion (toxic to bacteria, generated in phagolysosome)

33. Inflammatory mediators induce the expression of adhesion molecules that bind monocytes and PMNLs and aid in their recruitment to sites of infection in tissues  EXRAVASATION (4 steps) • 1. Rolling adhesion • 2. Tight binding • 3. Diapedesis (crossing the vascular endothelial wall) • 4. Migration • These 4 steps will now be reviewed.

34. Step 1: Rolling Adhesion • Endothelium is induced by inflammatory mediators to express SELECTINS • P-selectin induced by leukotriene B4, C5a or histamine • Appears immediately • Stored in endothelial granules called “Weibel-Palade Bodies” • E-selectin induced by TNF-alpha + Lipopolysaccharide (LPS)  appears after a few hours • Selectin ligand = glycoprotein sialyly-Lewisx on monocytes and neutrophils • Reversible binding  rolls along endothelium

35. Step 2: Tight Binding • ICAM-1 on endothelium induced by TNF-alpha • ICAM-1 binds integrins LFA-1 + MAC-1 (CR-3) • Tight binding induced by IL-8 (or other chemokines)  changes conformation of LFA-1 + MAC-1 • Binds better • Increases adhesion • Rolling stops

36. Step 3: Diapedesis • Crossing of endothelial wall = Extravasation (diapedesis) • Leukocytes squeeze through • Penetration of basement membrane (ECM)

37. Step 4: Migration • Migration through tissues via chemokines • Recruitment to appropriate location to enable phagocytosis/antigen processing

38. #5 – Normal Microbiological Flora (Microbiota) • Non-pathogenic organisms associated with epithelial surfaces compete with invading organisms for attachment sites • Compete for nutrients • Flora can produce anti-microbial substances (e.g. colicins made by Escherichia coli, oleic acid made by Propionibacterium acnes) • Often displaced by antibiotics enabling colonization by opportunistic bacteria

39. #6 – Alternative pathway of complement activation • Does not require antibody • Acts immediately • Activates the terminal complement components which destroy bacteria by creating holes (pores) in the bacterial membrane  Membrane Attack Complex • Opsonization also enhanced (C3b  binds to CR1)

40. #7 – Lymphoid lineages involved in non-adaptive responses • Natural killer (NK) cells (aka large granular lymphocytes – LGL) • Defend host against virus-infected cells • Kill sensitized targets • Activated by IL-12, alpha-interferon and beta-interferon • MHC class I involved • Present  Negative signal overrides activity of killing receptors

41. Intraepithelial gamma:delta T cells (gd) • A subset of T cells that are produced early during embryogenesis in waves • Homogeneous T cell receptors within any epithelium location • Do not recirculate • May reorganize alterations on the surfaces of epithelial cells as a result of infection • Exact function still unclear

42. CD5+ B cells (aka B-1 B cells) • Also arise early in embryogenesis • Limited rearrangement of V genes (ab genes), mainly IgM • Present as major lymphocyte in the peritoneum • Respond t polysaccharide antigens (TI-2 type – repeating subunit structure) • Exact function still debatable • Once IgM is bound, can activate complement

43. C. Adaptive Immune Responses • Clonal selection of lymphocytes • Lymphocytes express receptors with only one specificity • The specificity of each lymphocyte is unique • The body contains a pool of lymphocytes with a HUGE repertoire of different specificities • Lymphocytes with useful receptors are selected to survive • Most lymphocytes with self-reactive receptors are deleted (apoptosis) or rendered non-responsive (anergy)

44. Clonal expansion of lymphocytes • Because of huge variety of different receptors, the actual number of lymphocytes that can respond to a particular antigen is quite small • Lymphocytes will proliferate after activation prior to differentiating into effector cells

45. Stages of Clonal Expansion • Ag trapped in 2o lymphoid tissue is displayed to circulating naïve lymphocytes • Ag is recognized by a lymphocyte bearing a receptor with correct specificity for that Ag • Lymphocyte enlarges  Lymphoblast • Chromatin is less dense • Nucleoli appear • Cytoplasm increases • Transcription and translation begin

46. Cell division (2-4x every 24 hours for 3-5 days) • Can get up to 1000 daughter cells from one parent cell • Clones of daughter cells all have the same specificity for Ag as original activated cell • Differentiation into effector cells • B cells secrete Antibody (Ab)  Plasma cells • T cells destroy infected cells or “help” other cells to become activated

47. Some effector cells persist and develop into memory cells (more rapid 2o recall responses) • Lymphocytes with receptors that recognize host proteins (self) are deleted early in ontogeny and do not appear in the mature lymphocyte repertoire = TOLERANCE

48. Combinatorial diversity • Susumu Tonegawa (1976) demonstrated that Ig genes are a set of multiple gene segments that together encode the VARIABLE region of the antibody molecule (Nobel Price – 1987: Gene Rearrangement in Ab Synthesis) • Gene segments are joined together differently in each cell, generating a unique gene for the variable region (same process occurs in T cells)

49. Limited number of gene segments can give rise to large, diverse sets of products • Cells express unique Ag-receptors  huge repertoire of specificities • Genomic DNA recombined, changes are permanent  Somatic gene rearrangement (all daughter cells will have the same rearrangement) • ~108 different lymphocytes in our bodies each with unique specificity

50. IV. Antigen Presenting Cells