Immunology PDF

IMMUNOLOGY Historical aspects of immunology The first written observations by man was recognized persons had contracted & recovered from epidemic plague in Athens, Greece, over 2500 years ago. They were not susceptible (i.e. were immune) to further attacks. Many attempts were...

IMMUNOLOGY Historical aspects of immunology The first written observations by man was recognized persons had contracted & recovered from epidemic plague in Athens, Greece, over 2500 years ago. They were not susceptible (i.e. were immune) to further attacks. Many attempts were made to induce this immune state (of no second attack or no fatal results). In ancient times the process of variolation (the inoculation of live organisms of small-pox obtained from diseased pustules from patients who were recovering from the disease) was practiced extensively in India and China. The success rate was very variable. The results were sometimes disastrous for the recipient. The father of immunology was Edward Jenner (1749-1823), English country doctor. He noticed that: 1- Similarity between pustules of small-pox and those of cowpox, a disease affects cow‟s udders. 2- Milkmaids contracted cowpox by handling diseased udders were immune to smallpox. 3- Inoculation of a young boy with cowpox & later challenge, after the body had recovered, with contents of a pustule from a person suffered from small-pox failed to induce the disease and subsequent rechallenges also failed. The process of vaccination (Latin, Vacca =Cow) was adopted as a preventative measure against small-pox. The mechanism by which this 1  immunity was induced was not understood. In 1801, Jenner, prophesied the eradication of small-pox by the practice of vaccination. In 1969: The disease infected 10 million person. WHO initiated a program of confinement and with the object of eradicating the disease by vaccination. In Somalia, 1977, last case of naturally acquired small-pox occurred, and in 1979, WHO announced the total eradication of small-pox. Louis Pasteur (1822-1895) He was a chemist interested in fermentation and putrefaction by different microorganisms. By 1860, Pasteur had moved from the views of the chemist to that of the biologist. In 1879, Pasteur made a chance discovery. In the spring of the year he had begun experiments on chicken cholera. After the summer vacation it was found that the cultures of chicken cholera bacillus that had been kept during the summer did not produce disease when inoculated into healthy chickens. Pasteur immediately recognized the analogy between his results and Jenner’s cowpox protection against smallpox. Pasteur established the general principle that an organism can be altered (attenuated) so that it does not cause disease but still retains the property of inducing immunity. Pasteur immunized sheep, goats, and cows with attenuated anthrax bacillus and then exposed them to a virulent anthrax. The animals 2  did not develop the anthrax (i.e. protected). Paul Ehrlich and the side-chain Theory (1854-1915) He presented the first theory of antibody formation in 1900. Ehrlich realized that the antibodies in the serum must come from cells and postulated the “side-chain Theory” as an explanation. According to the side-chain Theory, every cell capable of synthesizing antibody (exact cells were not known at the time) has on its surface an array of “side-chains,” each of which is capable to react with a different specific antigen (as in figure). He assumed that each cell has a side chain for each of the microbial antigens to which the individual can produce antibody. When the specific agent (antigen) infects the body, it reacts with its specific side chain on the cell. This interaction between side chain and infectious agent somehow causes the cell to cease producing all the other side chains and initiate production of only the specific side chain with which the agent has interacted. Eventually, however, the cell overproduces this side chain and the excess appears in the serum as antibody. 3 Science of immunology not only encompasses: 1-The body‟s immune responses to bacteria and viruses but involved in: 2-Tumor recognition and subsequent rejection. 3-Rejection of transplanted organs and tissues. 4-Elimination of parasites from the body. 5-Allergies. 6-Autoimmunity (when the body mounts reaction against its own tissues). 4 Body possesses efficient natural defenses which restricts pathogens (microorganisms) to area where they can be tolerated. A breach of this mechanism, allow microorganism to reach tissuesnormally inaccessible, results in infection. Invasion and multiplication of microorganism in host may result in a pathological condition, i.e. the clinical disease. Definitions Pathogen: is an organism has the ability to cause disease. Virulence: indicate the degree of pathogenicity of a microorganism. Attenuation: reduction in the normal virulence of a pathogen, can eventually result in m.o. losing its virulence completely & is termed avirulent. Conversely, any increase in virulence is termed exaltation. IMMUNE SYSTEM Has Two components: 1- Natural Immunity displays neither specificity (or only partial specificity) nor memory. Natural immunity is inducible, displays self-nonself discrimination, and plays important roles in resistance to disease. 2- Acquired Immunity (specific or adaptive immunity) displays specificity and memory. Acquired immunity is also inducible, displays self-nonself discrimination, and is present only in vertebrates. However, the innate and acquired immunity cooperate in important ways to produce more effective immunity, activate each other and work towards the final goal of destroying the invading microorganism. 5 NATURAL IMMUNITY (NON-SPECIFIC defence mechanisms or INNATE immune system) -Operative at all times against potentially pathogenic microorganisms. -Infectious agent has no intrinsic effect on these systems. -Innate immunity is conferred by all those elements with an individual is born which are always present and available at very short notice to protect the individual from challenges by “foreign” invaders. Natural immunity or Non-specific defences include the following: 1-Skin and mucous membranes. Intact skin is virtually inaccessible to microorganisms and only when damage occurs can invasion take place. -Many microorganisms fail to survive on skin surface for any length of time due to the inhibitory effects of fatty acids and lactic acids in sweat and sebaceous secretions. -Mucus, secreted by membranes lining inner surfaces of tracts, acts as a protective barrier by trapping microorganisms and other foreign particles and subsequently removed by ciliary action linked, in case of respiratory tract, with coughing and sneezing. 2-Body secretions. Many body secretions contain substances with bactericidal action: For example: a- Lysozyme in tears, saliva and nasal secretions. b- Hcl in stomach results in low pH (2-3). c- Basic polypeptides as spermine found in semen. 3-Phagocytosis. Phagocytosis attempted by phagocytes: which are responsible for engulfment and digestion of microorganisms. It is a major line of defence against microbes that breach initial barriers described above. 6 Two types of phagocytic cells are found in blood, (derived from bone marrow stem cells): 1-Monocytes: constitute about 5% of total blood leucocytes, they migrate into the tissues and mature into macrophages. 2-Neutrophils: (polymorphonuclear leucocytes, PMNs) which are the professional phagocytes of the body. They constitute >70% of total leucocyte population. They possess receptors for Fc and activated C3 which enhance their phagocytic ability. Macrophages: are another group of phagocytic cells. They are large, long-lived cells found in most tissues (and lining serous cavities & the lung and secondary lymphoid organs, such as spleen and lymph nodes where they are advantageously placed to filter out foreign material). Total body pool of macrophages constitutes the so-called reticuloendothelial system (RES). Macrophages are also involved with the presentation (processing) of antigen to the appropriate lymphocyte population. Mechanism of phagocytosis: a-Chemotaxis, Migration, and Attachment: Macrophages are attracted to the site of inflammation by chemotactic substances liberated from microbes and damaged tissues. These includes bacterial endotoxins (LPS), serum complement C5a, IL-8 (chemokine) & leukotrienes. The macrophages respond by producing soluble proteins (cytokines) which mediate the migration of leucocytes and monocytes from blood to tissues. The phagocytes have many 7  receptors (e.g. mannose receptors) on their surface through which they attach non-specifically to microbes. These receptors bind to mannose residues on glycoproteins of many bacteria and viruses. Attachment and ingestion are greatly enhanced if the organism is coated by its specific antibody, by activated complement C3b or by antibody & C3b (opsonins) in a process called opsonization. b-Ingestion: The phagocytes proceed to engulf the organism by extending pseudo- pods around it. These fuse and the organism is included into a vacuole called phagosome. Lysosomal granules are then fused with phagosome, forming phagolysosome, followed by digestion of microorganisms. c-Intracellular killing or digestion: A number of antimicrobial and cytotoxic substances produced by activated macrophages can destroy phagocytosed microorganisms. This can occur through: -The oxygen-independent killing which is the result of lysozomal granules content which include; lysozyme, lactoferrin, a group of cationic proteins, and a variety of hydrolytic and proteolytic enzymes. -The oxygen-dependent killing system in which oxygen is converted to superoxide anion, hydrogen peroxide, activated-oxygen and hydroxyl radicals. All are powerful microbicidal agents. 8 4-Complement system. Complement system comprises a group of heat –labile serum proteins Produced by liver which when activated, are associated with the destruction of (bacteria) microorganisms. It is present in low concentration in serum but, its action is linked intimately with a specific defense mechanisms. 5-Interferons. They are proteins produced by certain type of leucocytes and interfere with viral replication. There is type I IFN which consists of IFN-α produced by mononuclear phagocytes and IFN-β produced by fibroblasts, and type II IFN or IFN-γ produced by T cells. 9 6-Fever. (Elevated body temperature) The thermoregulatory centre in the hypothalamus regulates body temperature and can be affected by: 1-Endotoxins (heat-stable LPSs) of Gram-negative bacteria and by 2-Interleukin-1(IL-1) [Endogenous pyrogen] secreted by monocytes and macrophages. Notes: a-Antibody production and T cell proliferation have been shown to be enhanced at elevated body temperatures. b-Increased body temperature may restrict replication of some viruses. c-Increased body temperature may inactivate the toxin products of invading microorganisms. 7-Species-specific Host-parasite relationships: e.g. T.B., Typhoid, polio, and smallpox are specific human pathogens. Genetically defined susceptibility results in certain species-specific Host-Parasite relationship. Man is not susceptible to a whole range of microorganisms that infect animals. The absence of species-specific receptors for microorganisms on surface of cells probably play a role in this host-parasite relationship. 8-Acute phase proteins: These are substances that increase in response to inflammation and include: C-reactive protein (CRP), fibrinogen, and serum amyloidal A protein. They are synthesized in the liver in response to certain cytokines, namely, IL-1, IL-6, and TNF-α, which are produced by macrophages when stimulated by microbial products. It seems likely that the acute phase response achieves a beneficial effect through enhancing host resistance and minimizing tissue injury. 10 9-Inflammation: Redness and swelling increased in tissues at sites of infection. Tissue damage by a wound or by an invading pathogen, induces a complex sequence of events collectively called “inflammatory response”. This includes; vasodilatation of nearby capillaries leading to redness of tissues and increase of tissue temperature, increased capillary permeability and influx of fluids and cells (exudates), into the tissues, causing oedema, and influx of phagocytes from capillaries into site of tissue damage, these engulf bacteria and release lytic enzymes. A variety of chemical mediators are responsible for initiation of the inflammatory response. Some of these mediators are released from damaged tissues or invading microbes. Among these chemical mediators are the acute phase proteins, histamine, kinins, fibrin and cytikines. These participate in destroying and removing the invaders and in healing of tissues. 10-Normal bacterial flora: Present at the portal of entry suppress the growth of many pathogenic bacteria and fungi by competition for essential nutrients or by production of inhibitory substances such as colicins or acids. For example, in the adult vagina an acidic pH is maintained by normal flora namely lactobacilli, that interfere with the establishment of pathogenic organisms. Suppression of normal flora by antibiotics leads to super infection with potential pathogens. Cells involved in immunity In vertebrates (mammalian species) circulating blood cells originate as a small clusters of cells in bone marrow called Hematopoietic stem cells: are undifferentiated cells (characterized by an ability to 11  proliferate throughout life) from which all other specialized cells in blood develop. They are Pluripotent stem cellswhich are capable of developing into any of Unipotent (in bone marrow) as follow: A-One pathway of differentiation: Is myeloid differentiation givemyeloid stem cells whichdifferentiate (just before they go to blood) to 1-Erythroblast (in blood)  Erythrocytes. 2-Myeloblast (in blood) GranulocytesBasophil, Eosinophil, Neutrophil. 3-MonoblastAgranulocytes Monocytes (in blood)  Macrophages (in tissues). 4- Megakaryocytes (in blood) Platelets (in tissues). B-Other pathway of differentiation: Is lymphocytic differentiation which giveLymphoid stem cells (in bone marrow) which developed toLymphoblast (in bone marrow) which differentiate into: (all are non-granulated) 1- B-lymphocytes (In bone marrow) 2- T-lymphocytes (In thymus) 3- Natural killer cells (NK) Types and function of cells Cell type General function -T cells -Help, suppression, cytotoxicity, delayed hypersensitivity, memory -B cells -Antibody production, memory -NK(natural killer) -killing -Monocytes -Phagocytosis-killing, monokines -Antigen processing & Presentation -Neutrophils -Phagocytosis-killing -Eosinophils -Antiparasitic -Basophils -Immidiate hypersensitivity Antiparasitic -Platelets -Clotting -Erythrocytes -Oxygen & carbon dioxide transport 12  Lymphatic organs The Lymphatic organs are those organs in which lymphocyte- maturation, -differentiation and -proliferation take place. Lymphoid organs are generally divided into two categories: A-Primary lymphoid organs: Which include: Thymus gland, Bone marrow and Bursa of fabricius (in birds). In these organs the maturation of T-lymphocytes and B-lymphocytes into antigen recognizing lymphocytes occurs. Developing T and B cells acquire their antigen-specific receptors in primary lymphoid organs. 13 1-Thymus gland Lymphoid stem cells from the bone marrow migrate to the thymus gland, where they differentiate into T-lymphocytes. Maturation of T-lymphocyte involves the commitment of a given T-cells to recognize and respond to a given determinant or epitope of a foreign antigen. This recognition is achieved by a specific receptor on the T cell, which is acquired during differentiation in thymus. Mature T-lymphocytes migrate the thymus and enter peripheral blood circulation, through which they are transported to secondary lymphoid organs. In these organs T cells encounter and respond to foreign antigens. Mature T cells in thymus are capable of responding to foreign antigens in the same way that they would respond in secondary lymphoid organs 2-Bursa and Bone marrow In birds, B cells undergo maturation in bursa. No bursa in mammals, so after birth and for the live of the individual this function moves to bone marrow, a structure that is considered to be a primary lymphoid organ with functions equivalent to that of avian bursa. Each mature B cell bears antigen-specific receptors that have a structure and specificity identical to the antibody-later synthesized by that B cell. Mature B cells are transported by the circulating blood to the secondary lymphoid organs, where they encounter and respond to foreign antigens. 14 B-Secondary lymphoid organs: Which include: Spleen, Lymph nodes, Tonsils, Appendix and MALT (mucosa-associated lymphoid tissue) e.g. Peyer’s patches (clusters of lymphocytes distributed in lining of small intestine). In these organs, antigen- driven proliferation and differentiation take place, i.e., in secondary lymphoid organs, mature, antigen-committed lymphocytes are stimulated by antigen to undergo further division and differentiation. Secondary lymphoid organs have TWO major functions: 1-They are highly efficient in trapping and concentrating foreign substances. 2-They are the main sites of production of antibodies and the generation of antigen-specific T-lymphocytes. 15 1-Spleen. It is the largest secondary lymphoid organs. Spleen is the major organ in which antibodies are synthesized and from which they are released into the circulation. 50% of spleen cells are B cells, 30-40% are T cells. Following antigenic stimulation, B cells and plasma cells synthesize and release antibodies. 2-Lymph nodes. Lymph nodes contain B cells, T cells, and macrophages. Lymph nodes found in various regions throughout the body. Lymph and lymphocytes are transported through lymphatic channels to the thoracic duct then to the blood circulation. In lymph node, on antigenic stimulation, lymphocytes- mostly B cells, are propagated. Macrophages trap, process, and present antigen to the T cells that have specificity against that antigen, events that result in activation of the T cells. In the node, antigen interacts with macrophages, T cells, and B cells, and that interaction brings about an immune response, manifested by the generation of antibodies and antigen-specific T cells. Lymph, antibodies and cells leave the lymph node to the circulation. Antigen may enter the gastrointestinal or respiratory tract, where it lodges in the mucosa-associated lymphoid tissue (MALT). There it will interact with macrophages and lymphocytes. ACQUIRED IMMUNITY (Specific defence mechanism/adaptive immune system) Foreign agents including microorganisms which successfully overcome the non- specific defence mechanisms then have to contend with a second line of defence, the specific defence mechanisms (acquired immunity)which classified into: 16 1-Active Acquired Immunity. (With Own antibodies or other defenses) a- Natural : Exposure to infectious agent. b- Artificial: Immunization with a vaccine contains live, attenuated, or dead microorganisms or their toxin. 2-Passive Acquired Immunity. (With Ready-made antibodies) a-Natural: Maternal antibodies, e.g. antibodies cross the placenta (i.e. from mother to fetus) and Colostrum (first milk, i.e. from mother to new born baby). b-Artificial: Antibodies from other sources (Prepared in other hosts and given to the human). Acquired immune responses arise as a result of exposure to foreign stimuli. The compound that evokes the response is referred to either as „Antigen‟ or as„immunogen‟. Antigen: is any agent capable of binding (or reacting) specifically to components of the immune response, such as lymphocytes and antibodies. Immunogen: is any agent capable of inducing an immune response and binding (or reacting) specifically to components of the immune response, such as lymphocytes and antibodies. Many compounds are incapable of inducing an immune response, yet they are capable of binding with components of the immune system that have been induced specifically against them. Thus, all immunogens are antigens, but not all antigens need to be immunogens. Haptens: (Greek, means to grasp). Haptens are low molecular weight compounds, e.g. many antibiotics and drugs. By themselves, these compounds are incapable of inducing an immune response, but when they are coupled with much larger entities, such as proteins (carrier) resultant conjugate induces an immune response that is directed against various parts of conjugate, including the low-molecular-weight compound (Hapten). 17 Immunogenicity: Is the stimulation of specific humoral or cellular immune response by a foreign substance. OR the capacity to induce an immune respond by a foreign substance. Requirements for immunogenicity: 1-Foreigness: not self (if self it leads to a condition termed „autoimmunity‟). 2-High molecular weight: -compound less than 1000 daltons is not immunogenic (e.g. penicillin, aspirin) -compound between 1000-6000 may or may not be immunogenic (e.g. insulin, adrenocorticotropic) -compound greater than 6000 daltons are generally Immunogenic (albumin, tetanus toxin). 3-Chemical complexity: a certain degree of physico-Chemical complexity in a compound is required to be immunogenic. e.g. homopolymers e.g. poly--D-glutamic acid (capsular material of B.anthracis) of molecular weight 50,000 daltons is not immunogenic, because they are: 1- not sufficiently chemically complex. 2-D-amino acids polymers are resistant to enzymatic degradation & not immunogenic. Whereas their L-isomers are susceptible to enzymes & are immunogenic. Antigenicity: Is the ability of an antigen to bind or react specifically with antibodies or lymphocytes generates specifically in response to that antigen. OR “An immune response induced by an immunogen generates antibodies or lymphocytes that react specifically with the antigen”. Antigen-binding site of antibody or a receptor on lymphocyte has a unique structure that allow a complementary ”fit” to some structural aspect of the specific antigen. Antigenic determinant or epitope: Is the portion of the antigen that binds specifically with the binding site of an antibody or a receptor on a lymphocyte. 18  - B and T cells recognize different epitopes. - Some Molecules may contain a single epitope (hapten), some having numbers of the same epitope on the same molecule (polysaccharides). - The most common antigens (proteins) having varying numbers of different epitopes on the same molecules. Types of antigens A-Exogenous: Source of antigen is from outside the host e.g., microorganisms, Pollens, drugs, pollutants, …etc. These exogenous immunogens are responsible for a wide variety of human diseases ranging from infectious (as bacterial infection) to immunologically mediated diseases (as hypersensitivity). B-Endogenous: Antigen initiated inside the host e.g.: 1-Auto-antigens: is a self antigen as a result of exposure of a hidden organ to the immune system as in traumatic cornea, prostate, and sperms OR due to any immune disturbance as in autoimmune diseases. 2- Iso-antigens: Cause immune response in genetically identical individuals of the same species e.g., identical twins. 3- Allo-antigens: in individual of specified species & are capable of eliciting immune response in genetically different individuals of same species e.g., blood groups. 4-Heterophilic antigens: are derived from one species and is capable of stimulating an immune response in another species. These antigens share the antigenic determinants (epitopes) i.e. identical or similar, therefore antibody formed to any one of them could react with the other (cross-reaction).For examples: -Human heart valve tissue and certain antigen found in S.pyogenes. -Used in Weil Felix diagnostic test for Epidemic typhus (Ricketsial disease) by Proteus organism (bacteria). Major chemical classes of antigens 1-Carbohydrates (polysaccharides). Polysaccharides are potentially, but not always, immunogenic. An immune 19 response, consisting primarily of antibodies, can be induced against many kinds of molecules of polysaccharide, such as components of microorganisms, and of eukaryotic cells. In Human: An excellent example of antigenicity of polysaccharides is the immune response associated with the ABO blood groups, which are polysaccharides on the surface of the red blood cells. Microbial cell surface has different antigens. These antigens may be common to different species or types of microorganisms OR may be highly specific for one type only. 2-Lipids: Are rarely immunogenic (may be regarded as haptens). 3-Nucleic acids: Are poor immunogens by themselves, but they become immunogenic when they are conjugated to protein carriers. In clinical medicine, appearance of anti-DNA antibodies in patients with systemic lupus erythematosus. 4-Proteins: Most common immune responses are those to proteins. Four groups of antigens are found on surface of intact bacterial cell: A- H-antigens (protein antigens): H or Flagellar antigen associated with flagella and are therefore only found on motile bacteria. Antigenically, flagellar antigens are different so used in typing of different strains of salmonella. B- O-antigens: (Somatic Ag) O-antigen are associated with bacterial cell wall, and referred to as somatic antigen. In Gram-negative: specificity of reaction between antigen and antibody is due to nature & number of type-specific polysaccharide side chains attached to lipid A and core polysaccharide portion of lipopolysaccharide(LPS). Loss of O-specific side chain antigens, resulting in exposure of more deep- seated core polysaccharide. Changed from S (Smooth)-specific antigens to R (Rough)-non-specific antigens,(or genus-specific antigens) shared with other unrelated Gram-negative bacteria. In Gram-positive: teichoic acid moieties 20  associated with cell wall are the major type-specific antigens. C- Capsular antigens: Many bacteria possess a characteristic polysaccharides capsule external to the cell wall. Over than 80 serotypes of Pneumococcus group have been differentiated by capsular polysaccharide antigens. Salmonella may possess a polysaccharide microcapsule antigens and is thought to be responsible for the virulence of bacteria. It is termed Vi antigen and its presence is important in production of typhoid vaccines. In addition: D- Fimbrial antigens: Are surface antigens in fimbriated gram-negative bacilli. In addition to the bacterial cellular antigens, there are some bacterial soluble antigens which are products excreted into environment, e.g. exotoxins, enzymes, hemolysins, etc.. Immunologic cross-reactivity A situation in which two or more substances, which may have various degrees of dissimilarity, share epitopes and would therefore, react with the immune components induced against any one of these substances, e.g. Toxoid and Toxin. THE TWO ARMS OF ACQUIRED IMMUNITY: (SPECIFIC IMMUNITY). There are two “arms”(branches) that have different sets of participants and different sets of purposes but with one common aim : to eliminate a foreign substance in the body often triggers both kinds of responses. These two arms interact with each other and collaborate to achieve the final goal of eliminating the antigen. One of these arms is mediated mainly by B cells and circulating antibodies, hence it is termed humoral immunity (humor=fluid or liquid). The other is mediated by T cells that do not synthesize antibodies but instead synthesize and release various cytokines that affect other cells. Hence, this arm 21  is termed cellular or cell-mediated immunity. 1-Humoral Immunity. Synthesis and release of free antibodies into blood and other body fluid is termed the humoral immune response. All antibodies are immunoglobulins but not all immunoglobulins have antibody function. Humoral immunity depends first on the ability of B lymphocytes to recognize Specific antigens and second on ability to initiate responses that protect the body against foreign agents. Each B cell is only capable of recognizing one epitope (antigenic determinant) via specific receptor on its surface. This receptor has been shown to be antibody itself. In most instances the antigens are on the surfaces of infectious organisms. The most common response is the production of antibodies that will inactivate an antigen and lead to destruction of infectious organisms. Each kind of B cell carries its specific antibody on its membrane and can bind immediately to a specific antigen. The binding of an antigen sensitizes or activates, the B cell and causes it to divide many times: -some of the progeny are memory cells -but most are plasma cells. Plasma cells are large lymphocytes that synthesize and release many antibodies like those on their membranes (2000 Abs/second :by single active plasma cell). Responses of B cells can be influenced by certain T cells. When B cells encounter very large antigen molecules with many epitopes (multiple and variable), they appear to generate plasma cells and antibodies without the aid of T cells. However, most antigens found on infectious agents have relatively few epitopes. In such instances helper T cells assist B cells by “processing” antigens so that B cells can respond to them. Macrophages also present antigens to B cells. 22 Suppressor T cells can block activity of helper T cells and so may be important in terminating an antigen-antibody reaction. Helper and suppressor T cells secrete helper & suppressor factors, respectively. Their effect on B cells seems to depend on the relative quantities of the two factors present in the region of immune reaction. In a typical immune reaction, helper T cells facilitate growth and differentiation of plasma cells. After about a week this reaction reaches a peak and then subsides, largely because suppressor cells inhibit further antibody production. Properties of antibodies (-Immunoglobulins) Antibodies, or γ-immunoglobulins (Ig), are Y-shaped protein molecules composed of four polypeptide chains- two identical light (L) chains and two identical heavy (H) chains. The chains, which are held together by disulfide bonds, have constant regions and variable regions. Chemical structure of the constant regions determine the particular class that an immunoglobulin belongs to. The variable regions of each chain have a particular shape and charge that enable the molecule to bind a particular antigen. When an antibody is cleaved with papain enzyme at hinge region, two fab (antibody binding fragment) pieces and one Fc (crystallizable fragment) piece result. The Fab fragment binds to the epitope. The Fc region formed by parts of H chains in tail of the Y has a site that can: bind to and activate complement , participate in allergic reactions, and combine with phagocytes in opsonization. Biology of the B lymphocytes: The biology of B cells responsible for several major characteristics of Immune response including: 1-Specificity: The ability to discriminate among different antigenic epitopes, and to respond only to those that necessitate a response. 2-Memory: The ability to recall previous contact with a particular antigen, such that subsequent exposure leads to a more rapid and larger immune response. 23 3-Adaptiveness: The ability to respond to previously unseen antigens, which may never have existed before on earth. 4-Discrimination: between “self” and “nonself”: The ability to respond to those antigens that are not “self” but to avoid making responses to “self” antigens. The most widely accepted theory that best explains these features of the immune system is the clonal selection theory, which was originally developed for Ab- producing cells (B-lymphocytes), but now the basic postulates of the theory apply to both B and T cells. The essential features of clonal selection theory are: 1. B and T cells of all antigenic specificities exist prior to contact with antigen. 2. Each lymphocyte carries Ig or T-cell receptor molecules of only a single specificity on its surface. 3. Lymphocytes can be stimulated by antigen under appropriate conditions to give rise to progeny with identical antigenic specificity. In case of B-cells, 24  the antigen-specific receptor-Ig- is secreted as a consequence of stimulation. 4. Lymphocytes potentially reactive with “self” are deleted or in some way inactivated. This ensures that no immune response is mounted against self components. Lymphocytes reactive to “self” molecules that are not deleted or inactivated may give rise to autoimmunity. Primary and secondary responses: Primary response occurs when the antigen is first recognized by host B cells. B cells divide to form plasma cells, which begin to synthesize antibodies. In a few day antibodies begin to appear in blood plasma and increase in concentration over a period of time. First antibodies are IgM, which can attack foreign substances directly. As IgM production decreases, IgG production accelerates. Eventually, it too wanes. Concentrations of IgM and IgG can become so low as to be undetectable in plasma samples. However, memory cells persist in lymphoid tissues. They do not participate in initial response, but they retain their ability to recognize a particular antigen. They can survive without dividing for many months to many years. Secondary response occurs when an antigen, enters the blood, recognized by memory cells. Presence of memory cells makes the secondary response much faster than primary response. Some memory cells divide rapidly producing plasma cells, and others remain as memory cells. Plasma cells quickly synthesize and release large quantities of antibodies. In secondary response, as in primary response, IgM is produce before IgG. However, IgM is produced in smaller quantities over a shorter period, and IgG is produced sooner and in much larger quantities than in the primary response. Thus, secondary response is characterized by a rapid increase in antibodies, most of which are IgG. 25  Classes of -immunoglobulins: Five classes of -immunoglobulins can be distinguished in humans and have different molecular weights. Each class has a particular kind of constant region , which gives that class its distinguishing properties. The five classes are identified as IgG, IgM, IgA, IgE, and IgD. Each unit consists of:1-one pair of heavy chain and 2-one pair of light chain joined by disulphide bonds Heavy chains are given the name of corresponding Greek letter: =gamma=IgG, η=mu=IgM, α=alpha=IgA, δ=delta=IgD, ε=etta=IgE. Light chains are one of two types: kappa(K) or Lambda(λ) chains. Hyper-variable regions: In the variable regions of both L and H chains there are extremely variable (hyper-variable) amino acid sequences that form the antigen-binding site. IgM: -First antibody secreted into the blood (serum) during the early stage of primary & secondary immune response. Since it is short-lived, its presence indicates recent infection, after which is usually disappear. -Can be made by both B cells and plasma cells /with IgD expressed on surface of B cell where it acts as an antigen receptor. -IgM consists of five units connected by their tails to J chain and has 10 epitope-binding sites. 26 -The monomeric form serves as the Ag receptor on the B cell surface. -Activate and fix the complement. -It is the antibody of the inherited ABO blood types. -Because of its size, IgM is unable to cross the placenta. -Important in bacteraemia, effective agglutinating agents. 27 IgG: -The main class of antibodies found in serum, take over the defensive role of IgM somewhat later in course of infection. -Produced in largest quantities during a secondary response. 28  -IgG consists of one unit, attach to antigens on microoganisms through antigen-binding sites , and its tissue-binding sites (Fc) attach to receptors on phagocytic cells enhancing the process of phagocytosis (opsonization), thus as microorganism is surrounded by IgG, a phagocytic cell is brought into position to engulf the microorganism. -Activate complement and fix it, lead to lysis of microorganisms and attract and stimulate phagocytes. -Cross the placenta from mother to fetus & provide protection to newborn for about 6 months after birth/colostral IgG cross gut mucosa. -Its half life time is 23 days and is the longest of all immunoglobulins. -It diffuses into the extravascular spaces neutralizing bacterial toxins (antitoxin). IgA: -Secretory IgA consists of 2 units held together by J chain (Joining chain) and occurs in large amounts in body secretions such as tears, milk, saliva, and mucous. Secretory IgA has secretory component which protects IgA from proteolytic enzymes and facilitates its transport. -Surface IgA consists of 1 unit and occur as attached to the lining of the digestive, respiratory, and genitourinary tracts. -IgA secreted into blood, transported through epithelial cells that line these tracts and either: released in secretion OR attached to lining by tissue binding sites. -IgA are abundant in colostrum, but not cross the placenta. -IgA inhibit adherence of microorganism to surface of mucosa, thus protect external body surfaces from microbial attack. -IgA activates the complement, which helps to kill the microorganisms. 29 IgE (Reagin): -IgE found in very low concentration in serum. -It binds to basophils in blood or mast cells in tissues by tissue-binding sites via Fc region, Leaving antigen-binding sites (on IgE) free to bind antigens to which human can develop allergies, such as drugs, pollens, and certain foods. -Reaction of cell-bound IgE with antigen triggers the degranulation of these cells with release of histamine, serotonine, and other vasoactive mediators. -Asthma and hay fever are common allergic diseases (associated with immediate type hypersensitivity). -IgE plays a role in immunity to helminthic (worm parasites). IgD: -IgD found on mature B cells. Its function is unknown. -Surface IgD (monomeric) can no longer be detected after activation of B cells. Thus IgD may be involved with differentiation of B cells. -Very small amount in serum. Like IgM, it function as an Ag receptor. Monoclonal antibodies: Are antibodies produced in the laboratory by a clone of cultured cells that make one specific antibody. To make monoclonal antibodies, myeloma cells (malignant cells of immune system) are mixed with sensitized lymphocytes (to specific Ag). Malignant cells are used because they will keep dividing indefinitely. Lymphocytes are used to make a particular antibody. When the two cells are mixed in cultures, they fused together to make a cell called a hybridoma. Hybridomas, contain genetic information from each original cell, divide indefinitely, and produce large quantities of antibody. Antibody is determined by antigen to which the lymphocytes 30 were sensitized before their progeny were mixed with myeloma cells. HAT Medium (hypoxanthine-aminopterin-thymidine medium) -For preparation of monoclonal antibodies, in Hybridoma tech. -splenocytes (sensitize B cells) are isolated from mammal, then B cells are fused with immortalized myeloma cells using polyethylene glycol. -Fused cells are incubated in the HAT medium. Aminopterin blocks the de novo pathway. So, un-fused myeloma cells die, as they cannot produce nucleotides by de novo or salvage pathway -Un-fused B cells die as they have a short lifespan. -In this way, only B cell-myeloma hybrids survive, These cells produce antibodies (a property of B cells) and are immortal (a property of myeloma cells). Incubated medium is diluted into multi-well plates to such an extent that each well contains only 1 cell. A- Among the diagnostic uses of monoclonal antibodies are: 1-detect pregnancy only 10 days after conception. 2-Rapid diagnosis of Ags on microbes or infected cells as in cases of hepatitis, influenza, herpes virus, and chlamydial infections. 3-Determination of CD (cluster of differentiation) markers on T-cells. 4-Tissue typing. 5-Detection of tumor Ags (markers). 6-Hurmonal assay. 7-Radio-labeled Abs used in detection & location of breast cancer metastases. B- Among the Therapeutic uses of monoclonal antibodies are: 1-Anti-tumor therapy, they selectively damage malignant cells without damaging normal cells. 2-Anti-CD20 used in treatment of B cell lymphoma. 3-Anti-Rh D to prevent Rh-incompatibility. 4-Passive immunotherapy for protection against viral infections e.g., varicella-zoster & cytomegalovirus (CMV). 5-Anti-snake venom. 6-Anti-TNF (Tissue Necrotic Factor) used in treatment of rheumatoid arthritis. 7-monoclonal Abs bind to IgE & prevent them from binding to mast cells 31  (in treatment of allergy). The great advantage of therapeutic monoclonal antibodies is that they selectively damage infected or malignant cells without damaging normal cells. 32 Kinds of antigen-antibody interactions The interaction between antigen and antibody in vitro is widely used for diagnostic purposes, for the detection and identification of either 33  antigen or antibody. The utilization of the in vitro reaction between antigen and serum antibody is termed serology. The formation of antigen-antibody complexes is an important component of the inactivation of infectious agents because it is the first step in removing such agents from the body. However, means of inactivation varies according to the nature of antigen and kind of antibody with which it reacts. Inactivation can be accomplished by such processes as: agglutination, precipitation, opsonization, activation of complement, cell lysis, and neutralization. These reactions occur naturally in the body and can be made to occur in laboratory. The interaction of antigen with antibodies may result in precipitation (if the antigen is soluble), and agglutination (if the antigen is particulate, i.e., an insoluble particle). Agglutination, precipitation, and complement activation are caused by: interaction between multivalent antigens and antibodies that have at least two combining sites per molecule. Consequences of these interaction do not represent primary interaction between antibodies and a given epitope but, rather, depend on secondary phenomena (i.e. cross linking), which result from interactions between multivalent antigens and antibodies. These reactions would not occur if antibodies with two or more combining sites reacted with a hapten (i.e., unideterminant, univalent antigen), nor would they occur as a result of interaction between a univalent fragment of antibody, such as Fab, and an antigen, even if antigen is multivalent. Cross-linking is possible only if antigen is multivalent and antibody is divalent (either intact, or F(ab’)2). In contrast, no cross linking is possible if antigen or antibody is univalent. 34 Agglutinatioin reactions: Agglutination reaction is due to the reaction of antibody with a multivalent Ag that is particulate (insoluble) and results in cross-linking and eventually in clumping or agglutination of antigen particles by antibodies, e.g. Fig Agglutination reaction The following are examples of agglutination tests: 1-Brucella test (tubes) 2-Widal test (tubes agglutination test) It is test for determination of serum antibodies, in patient with typhoid fever. The procedure involves addition of a suspension of dead typhoid bacterial cells to a series of tubes containing the various dilutions of patient's serum. Tubes are then incubated for 30 minutes at 37°C. After centrifugation, the reciprocal of the highest dilution showing agglutination is designated as the antibody titer. For example, if the highest dilution at which agglutination occurs is 1:160, the titer is 160 antibody units per milliliter of serum. High or increasing titer indicates active infection 3-Coombs test (for hemolytic disease of the newborn, erythroblastosis fetalis): Direct test: The addition of anti-Rh IgG to a suspension of baby‟s erythrocytes would result in the binding of anti-Ig to the maternal IgG on surface of RBCs and would cause agglutination. Indirect test: The detection of free Ig, in the serum of an Rh-negative women, by addition of specific antigens on the particle, may fail to cause agglutination. The subsequent addition of anti-Ig will cause agglutination. Thus, direct Coombs test measures bound antibody while indirect test measures serum (free) antibody. 35 4-Erythrocyte typing in blood banks. Hemagglutination, or agglutination of red blood cells, is similar to the agglutination tests except the antigens are on the surface of red blood cells. Hemagglutination is used in blood typing. 5-Latex test for pregnancy (which involves the detection of human Chorionic gonadotropin (HCG) hormone (Ag) in the urine of pregnant women. Agglutination can also be used with soluble Ags, by attaching them to latex particles (insoluble). Precipitation reaction: Precipitation reaction takes place when antibodies and soluble antigen are mixed. As in agglutination, precipitation of Ag-Ab complexes occurs because the: divalent antibodies molecules cross-link multivalent antigen molecules to form a lattice. When it reaches a certain size, this Ag-Ab complex loses its solubility and precipitates out of solution. Precipitation reactions in gel (Immunodiffusion) [in semisolid phase, agar or gel]: Precipitation reactions between antigens and antibodies can take place 36  not only in solution but also in semisolid media such as agar gels. Single diffusion (Radial immunodiffusion): The wells contain Ag at different concentrations, while antibodies are distributed uniformly in agar gel. Thus, a precipitin ring around the well is formed. Precipitin ring is directly proportional to the concentration of Ag in the well. Relationship between concentration of Ag in a well and diameter of precipitin ring can be plotted. Thus, unknown concentration of same Ag can be determined. This test is used clinically to measure concentrations of serum proteins. Double diffusion: When soluble Ag and Abs are placed in wells cut in gel, the reactants diffuse in gel and form gradients of concentration, with the highest concentrations closest to the wells. Somewhere between the two wells, the reacting Ag and Abs will be present at proportions that are optimal for formation of precipitate. 37 Double diffusion method, developed by Ouchterlony, where Ag and Ab diffuse toward each other, is useful for establishing the antigenic relationship between various substance as follow: 3 pattern could be seen: Pattern of identity: Central well contains antibodies and peripheral wells contain identical antigens. Antibodies diffuse from central well toward antigens, since they are identical, form one continuous, coalescing precipitin line. This pattern, formed when the two antigens are identical. Pattern of nonidentity: Central well contains antibodies to Ag1 and Ag2 (unrelated Ag) and peripheral wells contain two unrelated antigens. Antigens antibodies diffuse towards each others. Each antigen forms an independent precipitin line with its corresponding antibody at an equivalent point. Precipitin lines cross each other since each antigen diffuses across the band formed by the antigen until it meets its specific antibody diffusing toward it, precipitin lines cross each other denotes nonidentity of 2 antigens. 38 Pattern of partial identity: Center well contains antibodies to various epitopes of Ag1. Reaction of antibodies with Ag1 results in a precipitin line. Ag2 contains some (not all) of epitopes of Ag1. Thus, some of antibodies to Ag1 will also combine with Ag2. Thus gives a line of identity (coalescence). However, antibodies that do not bind Ag2 will pass through this line of precipitate, combine with Ag1 on other side, and form a spur, which denotes partial identity, signifying that Ag1 and Ag2 share epitopes, with Ag1 having more epitopes. 2. Elek's test:- The double diffusion technique is used to detect toxin-antitoxin reactions as in Elek's test. An isolated strain of Diphtheria is not considered pathogenic unless it proves to be toxigenic. Principle: To determined the toxigenic strain of C. diphtheria. Toxin production by C. diphtheriae can be demonstrated by a precipitation reaction between exotoxin and diphtheria antitoxin. This test is done as follows: A strip of filter paper saturated with Diphtheria antitoxin is embedded in a serum agar plate. A heavy inoculum of the Diphtheria bacilli to be tested is inoculated at right angle to the strip of filter paper. Plates are examined after 2 days incubation at 37oC. If the organism is toxigenic, the toxin will diffuse sideways from the streak and the antitoxin diffuses from the filter paper, and where they meet at optimum concentrations a precipitate is formed which will appear as fine white lines commencing from the streak. Results: Positive test: formation of four radiating lines resulting from the precipitation reaction 39 between exotoxin and diphtheria antitoxin Immunoelectrophoresis: Immunoelectrophoresis involves separating a mixture of proteins in an electrical fields (electrophoresis) followed by their detection with antibodies diffusing into the gel. For example: in clinical characterization of human serum proteins, a small drop of human serum is placed in a well cut in the center of a slide that is coated with agar gel. Serum is then subjected to electrophoresis, which separates the various components according to their mobilities in electrical field. After electrophoresis, a trough is cut along the side of the slides, and antibodies to human serum proteins are placed in trough. Antibodies diffuse in agar, as do separated serum proteins. At an optimal antigen: antibody ratio for each antigen and its corresponding antibody, series of precipitate arcs are formed. Comparison of pattern and intensity of lines of normal human serum with patterns and intensity of lines obtained with sera of patients may reveal an absence, or over abundance, or other abnormality of one or more serum proteins (e.g. human myeloma protein). Normal human serum is separated into 5 major proteins i.e., albumin, α1-globulin, α2-globulin, β-globulin, and γ-globulin. Immunoelectrophoresis combines elecrophoretic separation diffusion and Immune precipitation of proteins. 40 COMPLEMENT FIXATION TEST: The body‟s natural defenses use complement to bind Ag-Ab complexes, helping to destroy pathogens. This ability is used in laboratory or clinic to detect very small quantities of antibodies. It is a multi-step procedure that begin with: 1-inactivation of complement from a patient‟s serum by heating, 2-serum is then diluted, and known quantities of non human complement and test antigen which is specific for antibody being thought, are added separately, 3-Mixture is incubated to allow antigen to react with antibody, 4-Next, an indicator system consisting of sheep RBCs-Anti-sheep‟s RBCs is added, 5-If antibody to test antigen was present in patient‟s serum, Ag-Ab reaction will have fixed (combined with) the C. Hence RBCs will not lyse, and the test will be positive, forming a red button of undamaged cells. But if antibody was not present, free C in the mixture will be fixed by indicator system, resulting in lysis of the cells, the test will be negative. This test is used to diagnose bacterial diseases as pertussis, gonorrhea; and fungal diseases, e.g. histoplasmosis. In addition the test is used in diagnosis of syphilis (Wassermann test). NEUTRALIZATION REACTIONS: Neutralization reaction can be used to detect bacterial toxins and antibodies to viruses. Immunity to diphtheria, which depends on presence of diphtheria antitoxins 41  (antibodies to diphtheria toxin) can be detected by Schick test. In this test a person is inoculated with a small quantity of diphtheria toxin. If the person is immune to the disease, diphtheria antitoxins (circulating in the blood) will neutralize the toxin, and no adverse reaction will occur. If the person is not immune and the antitoxin is not present, the toxin will cause tissue damage, detected as a swollen reddened area at injection site after 48 hours. Viral neutralization: occurs when antibodies bind to viruses and neutralize or prevent them from infecting cells. In laboratory, patient‟s serum & test virus are added to cell culture or chick embryo. If serum contains antibodies to virus, these antibodies will neutralize the virus and prevent cells of culture or embryo from becoming infected. TAGGED ANTIBODY TESTS. The most sensitive immunological tests used to detect antibodies or antigens use antibodies that have a ”molecular tag” that is easy to detect even at very low concentrations. In fact, the concentrations are so low that precipitation or agglutination does not occur. Western blots (Immunoblots): In Western blot technique, antigen (or a mixture of antigens) is first separated in gel by an electric current, as in immunoelectrophoresis. Separated material is then transferred “blotted” onto nitrocellulose sheet to which antigen binds strongly. Antibodies (serum patient), which is then applied to nitrocellulose sheet, binds with its specific antigen. Antibody may be labeled (e.g. with radioactivity or enzyme), or a labeled Anti-Ig may be used to localize the antibody and the antigen to which the first antibody is bound. Such Ag-Ab complexes can be visualized by the addition of an enzyme labeled anti-human antibody. When an enzyme substrate is added, colored bands appear on nitrocellulose sheet. This technique is used in diagnosis of AIDS, by application of patient‟s serum to nitrocellulose sheet on which HIV antigens are bound. Finding of specific antibody is strong evidence of infection by the virus. 42 Radioimmunoassay (RIA): Radioimmunoassay used to detect very small quantities (nanograms) of antigens or antibodies. To measure an antibody in specimen, a known antigen in saline solution is placed in a well in plastic plates. Some antigens molecules attach to plastic; those do not attach are washed away. Antibody need to be measured is added and allowed to bind with antigen. Then a radioactively tagged anti-antibody is applied. After an incubation period, the excess unbound antibody is removed by washing. Radioactive material remaining in well is measured with a radiation counter to determine the concentration of antibody in test specimen. Immunofluorescence: (Immunohistochemical technique, direct & indirect). 43 Immunofluorescence makes use of antibodies (usually IgG) to which fluorescent dye molecules are bound (tagged) at the tail (Fc) ends of antibodies. For example, IgG tagged with fluorescein isothiocyanate glow a bright yellow green when exposed to UV light. Fluorescent tagged antibodies can be used to detect antigens, other antibodies, or complement at their locations on cells or within tissues by the aid of fluorescent microscope, a fluorescent tagged antibody that detects another antibody is known as Anti-Ab; one that detect complement is an anti-complement Ab. Immunofluorescence is helpful in diagnosis of syphilis, gonorrhea, HIV infection, and Legionnaires‟ disease. Enzyme-linked immunosorbent assay (ELISA): [on solid phase]. ELISA is a modification of RIA in which the Anti-Ab, instead of being radioactive, has an enzyme tag attached to it. After the antibody need to be measured has reacted with the antigen, the Anti-Ab-Enzyme complex is added. Finally, a substrate that the enzyme converts to a colored product is applied. Amount of colored product is proportional to the concentration of antibody. RIA and ELISA are among the most widely used tests for antibodies or for antigens. ELISA is used in detection of HIV Abs. Test was developed to screen the nation‟s blood supply and protect recipients of blood products from infection. 44 2-Cell-mediated (Cellular) Immunity. -In contrast to humoral immunity, which involves B cells and Igs, cell- mediated immunity involves the direct action of T cells. -Host defenses against extracellular infectious agents are mediated Mainly by antibody, complement, and macrophages. -However, once the infectious agent invades the host cell, cell mediated immunity (CMI) is required for recovery from these intracellular infections. T cells interact directly with other cells that display foreign antigens. These interactions: 1-clear the body of viruses and other pathogens that have invaded host cells (intracellular parasite), and 2-account for rejection of tumor cells, 3-some allergic reactions, and 4-immunological responses to transplanted tissues. -Cell-mediated immune response involves the differentiation and actions of different types of T cells and the production of chemical mediators call lymphokines. T cells differ from B cells in that they do not make Abs. -However, they do have a particular cell membrane receptor protein that corresponds to Abs of B cells, & other receptors proteins as well. Identification and characterization of cells: 45  A uniform system of nomenclature has been adopted in which all cell- surface markers are designated CD, followed by a number indicating its order of discovery. The acronym CD (cluster of differentiation) describes the cluster antigens which are identified by using monoclonal Abs. The list extends at least to CD130, not all of these surface molecules have a currently understood function. Surface expression of a particular molecule may not be specific for just one cell or even for a cell lineage. T cell express glycoprotein molecules (receptors) e.g. CD3, CD4, CD8,.., on the surface. -CD3 receptors are expressed by all stages of T-cells, while, -CD4 or CD8 are expressed by mature T-cells. -CD4 on Helper cells (TH1 and TH2) and T-delayed hypersensitivity. -CD8 on cytotoxic cell (Tc) and T suppressor (Ts) cell. Cell-mediated immune reaction: Cell-mediated immune reaction typically begins with the processing of an antigen (associated with pathogenic organism) by macrophages. 1-Macrophages phagocytize pathogens, they ingest & degrade pathogen, then insert some of pathogen’s antigen molecules into their own cell membranes. THIS CONSTITUTES PROCESSING OF ANTIGEN. 2-When a macrophage presents antigen to T cells having proper antigen receptor, antigen and receptor bind. Macrophage also has on its surface genetically determined histocompatibility (MHC) proteins that bind to T cell receptors. 3-Binding with macrophages cause mature T cells to divide and differentiate into different types of T cell, including memory cells. Each type of T cells, sensitized to specific antigen that initiate the process, has a different function in cell-mediated immune reactions. Some cells act directly (e.g. cytotoxic cell) and others release lymphokines (e.g.T-helper cells), which are chemical substances that trigger certain immunologic reactions. e.g.: 46 4-Macrophages that have processed the antigen secrete the lymphokine, interleukin-1 (IL-1) which activates helper T (TH) cells which in turn, secrete lymphokines such as gamma interferon and Interloeukin-2 (IL-2). -IL-1 from macrophages and IL-2 from TH cells activate other T cells as suppressor T(Ts)cells, delayed hypersensitivity T (TD) cells, and cytotoxic (killer) T (Tc) cells. Also, IL-1, IL-2, and gamma interferon together cause null cells to become natural killer (NK) cells. -At the same time during differentiation of above cells, some T-memory cells are formed. As in humoral immunity, persistence of memory cells in cell-mediated immunity allows the body to recognize antigens to which T cells have previously reacted and to mount more rapid subsequent responses. -TH cells stimulate the growth and differentiation of B cells, and TS cells suppress TH cells. Such suppression apparently helps to prevent both humoral and cell-mediated immune processes from getting out of hands. TH cells have receptors that recognize the antigen (peptide fragment) on MHC class II on macrophage. Binding causes TH cells to become activated. Activated TH cells then differentiated into either TH1 cells activate infected macrophage to destroy internal bacterial infections, OR TH2 cells activate B cells (humoral immune responses) by binding to MHC class II : Ag (peptide fragment) presented by B cells. Presenting the same Ag (peptide fragment) on MHC class I processed by macrophages to TC cells activates these cells to attack infected cells, especially abnormal or virus-infected cells. 47 Activated TD (delayed hypersensitivity T cells) cells release various lymphokines, these includes: 1-Macrophage chemotactic factor, which helps macrophages to find microbes. 2-Macrophage activating factor, which stimulate phagocytic activity. 3-Migration inhibitory factor, which prevents macrophages from leaving sites of infection. 48 4-Macrophage aggregation factor, which causes macrophages to aggregate at such sites. TD cells also participate in delayed hypersensitivity (kind of allergic reaction). TC cells and NK cells kill infected host cells. When pathogens have evaded humoral immunity and established themselves inside cells, they can cause long-term infections unless the infected cells are destroyed by cell-mediated immunity. An agent that infects T cells (e.g. HIV) is especially devastating because it destroys the cells that might have combated the infection. HIV virus invades TH cells, prevents them from carrying out their normal immunological functions, and eventually kills them. Lack of TH cells impairs both humoral and cellular immune responses, including destruction of malignant cells. Thus, because of extensive destruction of TH cells, AIDS patients are susceptible to opportunistic infections and to various malignancies. How killer cells kill TC cells and NK cells kill other cells by making a lethal protein and firing it at target cells. Eosinophils have a similar protein, which they may use to kill certain helminthes and other parasites. Cytotoxic T cells act mainly on virally infected cells, whereas NK cells act mainly on tumor cells, cells of transplanted tissues, and possibly on cells infected with intracellular agents such as Rickettsias & Chlamydias. Tc cells bind to Ags presented by macrophages and then attack virus- infected cells. In contrast, NK cells bind directly to malignant or other target cells without the help of macrophages. Both kinds of killer cells contain granules of a lethal protein, perforin, which is released when they bind to a target cell. Perforin bores (makes) holes in the target cell membranes so that essential molecules leak out and the cells die. This process is similar to the action of complement. 49 By killing infected cells while they are few in number and before new virus particles are released from them, TC cell prevent the spread of infection-but the expense of destroying host cells. Similarly, NK cells destroy malignant cells before they have a chance to multiply. Both kinds of killer cells can withdraw from cells they have damaged and move onto other target cells. Extracellular killing: -The phagocytic process represents intracellular killing- that is, the microbe is degraded within a defensive cell. However, other microbes, such as viruses and parasitic worms, are destroyed without being ingested by a defensive cell; they are destroyed extracellularly. - Neutrophils and macrophages are too small to engulf a large parasite such as worms (helminthes). Therefore, another leukocyte, the eosinophil, takes the leading role in defending the body. Eosinophils are best suited for excreting toxic enzymes such as major basic protein (MBP) that can damage or perforate a worm’s body. Once such parasites are destroyed, macrophages can engulf the parasite fragments. -Viruses must get inside cells to multiply. TC and Natural killer (NK) cells are responsible for killing intracellular viruses. NK cells are a type of lymphocyte whose activity is greatly increased by exposure to interferons and cytokines. TC and NK cells probably recognize specific glycoproteins on cell surface of virus-infected cells. Such recognition does not lead to phagocytosis; rather, the cells secrete cytotoxic proteins that trigger the death of infected cell. Antigen-presenting cells (APC): Such as macrophages. Although these cells do not have Ag-specific receptors as do the lymphocytes, their important function is to “process” the Ag and “present” the Ag to the specific receptors on T cells. Ag-presenting cells have on their surface “two types” of special molecules that function in Ag presentation. These molecules, called MHC class I and MHC class II molecules, are encoded by a complex set of genes that are also responsible for rejection or acceptance of transplanted tissue. This set of genes is referred to as “Major Histocompatibility Complex (MHC)” and the class I and class II molecules are commonly referred to as MHC class I and MHC class II molecules. 50 Processed Ag is noncovalently bound to MHC class I or class II molecules or both and is thus presented to specific receptors on T cell. Interestingly, MHC I molecules present Ags to TC cells, while MHC II molecules present Ags to T-helper cells. Role of activated macrophages: Some bacteria, such as those cause tuberculosis, leprosy, listeriosis, and brucellosis, can continue to grow even after they have been engulfed by macrophages. TD cells combat such infections by releasing lymphokine macrophage activating factor. This factor causes macrophages to increase production of toxic hydrogen peroxide, along with enzymes that attack the phagocytized organisms and accelerate the inflammatory response. Organisms that survive these defenses are walled off in granulomas. As in case of intracellular infection by bacteria. COMPLEMENT Complement or complement system, refer to a set of more than 20 large regulatory proteins that play a key role in host defense. They are produced by the liver and circulate in plasma in an inactive form. Although complement can be activated by immune reactions, its effects are nonspecific- it exerts the same defensive effects regardless of which microorganism has invaded the body. The general functions of the complement system are: 1-enhance phagocytosis by phagocytes, 2-lyse microorganism, bacteria, and enveloped virus directly, and 3-generate peptide fragments that regulate inflammation and immune responses. Complement goes to work as soon as an invading microbe is detected; the system makes up an effective nonspecific host defense long before specific host immune defenses are mobilized. The complement system works as a cascade. A cascade is a set of reactions that amplify some effect- that is, more product is formed in the second reaction than in the first, still more in the third, and so on. Of the 20 different serum proteins, identified in complement system, 13 participate in cascade itself, and 7 activate or inhibit reactions in the 51  cascade. Complement function: Two pathways have been identified in the sequence of reactions carried out by complement system. They are called the classical pathway and the alternative pathway (or properdin pathway). Classical pathway begins when antibodies bind to (cross-link) antigens such as microbes and involves complement proteins C1, C4, and C2 (C stands for complement). Alternative pathway is activated by contact between complement proteins (called Factor B, Factor D, and Factor P (properdin) and polysaccharides at the pathogen surface. However, the complements of both pathways activate reactions involving C3 through C9. Consequently, the effects of complement systems are the same regardless of the pathway by which C3 is produced. However, alternative pathway is activated even earlier (i.e. non-specific) in an infection than is classical pathway (specific). The contributions of complement system to nonspecific defenses depend on C3, a key protein in the system. Once C3 is formed, it immediately splits into C3a and C3b, which then participate in three kinds of molecular defenses: Opsonization, inflammation, and membrane attack complexes. 52 Opsonization: It is known that some bacteria with capsule or surface proteins (M proteins) can prevent phagocytes from adhering to them. The complement system can counteract these defenses (by microbes), making possible a more efficient elimination of such bacteria. First, special antibodies called opsonins bind to and coat the surface of infectious agent. C1 binds to these antibodies, initiating the cascade. C1 causes the cleavage of C4 into C4a and C4b. C4b and C1 then cause C2 to split into C2a and C2b. The C4bC2a complex in turn leads to splitting of C3 into C3a and C3b. then C3b binds to surface of microbe. C3b molecules bind to complement receptors on plasma membrane of phagocytes. This recognition stimulates phagocytosis. This process, initiated by opsonins, is called “opsonization”, or “immune adherence”. Inflammation: Complement system is also potent in initiating and enhancing inflammation. C3a, C4a, and C5a enhance the acute inflammatory 53 reaction by stimulating chemotaxis and thus phagocytosis (by neutrophils). These 3 complement proteins also adhere to membranes of basophils and mast cells causing them to release histamine and other substances that increase the permeability of blood vessels. Membrane attack complexes: Another defense triggered by C3b is cell lysis. By a process called immune cytolysis, complement proteins produce lesions in cell membranes of microorganisms and other types of cells. The lesions cause cellular content to leak out. To cause immune cytolysis, C3b initiates the splitting of C5 into C5a and C5b. C5b then binds C6 and C7, forming a C5bC6C7 complex. This protein complex is hydrophobic and inserts into the microbial cell membrane. C8 then binds to C5b in the membrane. Each C5bC6C7C8 complex causes the assembly in cell membrane of up to 15 of C9 molecules. By extending all the way through cell membrane, these proteins form a pore and constitute the membrane attack complex (MAC). MAC is responsible for the direct lysis of invading microorganiswms. Importantly, host plasma membranes contain proteins that protect against MAC lysis. These proteins prevent damage by preventing the binding of activated complement proteins to host cells. The MAC forms the basis of complement fixation test, used to detect antibodies against any microbial antigens. 54  IMMUNIZATION Acquired immunity is induced by immunization, which can be achieved through: 1.Active immunization: refers to immunization of an individual by administration of an antigen. 2.Passive immunization: refers to immunization through the transfer of specific antibody from an immunized individual to a nonimmunized individual. Each year nearly 3.5 million children, most under 5 years, die of 3 infectious disease (measles, tetanus, and whooping cough) for which immunization is available. Another 4 million die of various kinds of diarrhea, against some of which immunization is possible. Most of these deaths occur in developing countries. These deaths provoke three important facts about immunization: 1-Immunization can prevent significant numbers of deaths. 2-Methods of immunization are not yet available for some infectious diarrheal diseases. Most organisms that cause diarrhea exert their effects 55  in digestive tract, where antibodies and other immune defenses can not reach them. 3-Much greater effort is needed to make immunizations available in developing countries. Active immunization: To develop active immunity, the immune system must be induced to recognize and destroy infectious agents whenever they are encountered. Active immunization is the process of inducing active immunity. It can be conferred by administering vaccines or toxoids. A vaccine: is a substance that contains an immunogen (i.e. antigen) to which the immune system responds. Antigens can be derived from live attenuated (weakened) organisms, dead organisms, or parts of organisms. A toxoid: is an inactivated toxin that is no longer harmful, but retains its antigenic properties. Mechanism of active immunization: When the vaccine or toxoid is administered, the immune system recognizes it as foreign and produces antibodies, or sometimes cytotoxic T cells, and memory cells. Immune response is the same as the one that occurs during the course of a disease, but the disease symptoms do not occur. In other words, vaccine or toxoid keeps antigenicity but lacks pathogenicity (capability to induce disease) or toxicity, respectively. Vaccines made with live organisms confer longer-lasting immunity than those made with dead organisms, parts of organisms, or toxoids. e.g.: 1.measles (both Rubella & Rubeola) vaccines and oral poliomyelitis 56  vaccine, which contain live viruses, usually confer life long immunity. 2.IM polio vaccine (killed virus) and typhoid vaccine (dead bacteria) confer immunity lasts 3-5 years. 3.Tetanus and diphtheria toxoids confer immunity of about 10 years. Because active immunity is not always lifelong, “booster shots” are often needed to maintain immunity. First dose of vaccine or toxoid stimulates a primary immune response. Subsequent doses stimulate a secondary immune response. Vaccines and toxoids, must be properly stored to retain their effectiveness, e.g. refrigeration. In general, active immunization can not be used to prevent a disease after a person has been exposed. This is because the time required for immunity to develop is greater than the incubation period of the disease. Rabies immunization is an exception of this rule. Because rabies typically has a long incubation period, active immunization can be used with some hope that immunity will develop before the virus reaches the brain. The farther the virus must travel to reach the brain, the greater the chance of effective immunization. So, a bite on the ankle received while kicking off a rabid animal may be less hazardous than one on the trunk or neck. Hazards of vaccines: Fever, malaise and soreness at the site of injection, joint pain (e.g. rubella vaccine), convulsions (e.g. pertussis), and allergic reactions are among the hazard of vaccines. Thus, patients already suffering from fever and malaise should not receive immunization because of the worsening of their conditions. An exceedingly small number of vaccine recipients die or suffer 57  permanent damage from vaccines (e.g. oral poliovaccine). Live vaccines pose particular hazards to pregnant women, cross placenta and infect fetus, with immature immune system, may cause birth defects. Also pose hazards to immunodeficient patients, patients receiving immunosuppressants as radiation or corticosteroid drugs. Passive immunization: Passive immunization is induced by introduction of ready-made antibodies into an unprotected individual. Since antibodies are found in serum protein of blood, these products are often called antisera. Induce passive immunity which is quick but it is only temporary. It lasts only as long as there is a sufficiently high titer of circulating antibodies. Passive immunization is established by administering a preparation such as: 1-Gamma globulin (Immune serum globulin) Consists of pooled gamma globulin fractions (portion of serum containing antibodies) from many individuals. Typically, it contains sufficient antibodies to provide passive immunity to a number of common diseases, such as mumps, measles, and hepatitis A. 2-Hyperimmune serum (convalescent sera) Consists of gamma globulins that have high titers of specific kinds of antibodies. For example, gamma globulin from persons recovering from mumps or from recent recipients of mumps vaccine consists especially high titers of anti-mumps antibodies. Also can be manufactured by introducing particular antigen, e.g. tetanus toxin, into another animal, such as a horse, and subsequently collecting the antibodies from the animal‟s serum. 58  3-Antitoxins Are antibodies against specific toxins, such as those that cause botulism, diphtheria, or tetanus. Passive immunization gives immediate immunity to a nonimmune person who is exposed to a disease. Before the advent of antibiotics, passive immunization was frequently used to prevent or lessen the severity of several kinds of pneumonia and a variety of other infectious diseases and counteract the effects of snake and other insect venoms. Anti-Rh-antibodies are given to Rh-negative mother within 72 hours after the birth of first Rh-positive child and after subsequent births, miscarriages, or abortions. Anti-Rh-antibodies bind to Rh-positive RBCs, so the cells are destroyed before the mother immune system can make antibodies to them. Hazards of passive immunity Are allergic reactions, e.g. antitoxin prepared in animals as horses, may contain antigenic protein. For example, allergic reactions to large IgG, if gamma globulins or hyperimmune sera are accidently given intravenously instead of intramuscular route. Immunity to various kinds of pathogens: Bacteria: antibodies produced by plasma cells are the chief immunological defense against bacterial antigens. Most immune responses to bacteria serve to promote phagocytosis of the invading cells. Viruses: viral infected cell is combated by nonspecific defenses, interferon, and antibodies. In addition, TC cells of cell-mediated responses and NK cells are important in destroying virus-infected cells. 59 Fungi: immune responses to fungi involves IgA antibodies and are primarily cell-mediated. Protozoa and helminthes: are largely cell-mediated. T cells release cytokines that activate macrophages and attract other leucocytes (eosinophils). Allergic reactions to helminthes can be damaging to host than to parasite. ,BCG BCG = Bacillus calmette-Guerin for Tuberculosis (TB). IMMUNOLOGICAL DISORDERS Previously, we emphasized how specific immune responses defend the body against harmful substances. However, such responses are not always beneficial. Sometimes the humoral or cell-mediated responses react in ways that are 60  physiologically unpleasant or even life-threatening, e.g. runny nose and watery eyes of hay fever season, and anaphylaxis. An immunological disorder is a condition that results from an inappropriate or inadequate immune response. Most inappropriate responses involve some type of hypersensitivity, whereas inadequate responses are due to immunodeficiency. HYPERSENSITIVITY (ALLEREGY) Occurs when the immune system reacts in an exaggerated or inappropriate way to a foreign substance. There are four types of hypersensitivity: 1-Immediate hypersensitivity (Type I), 2-Cytotoxic hypersensitivity (Type II), 3-Immune complex hypersensitivity (Type III), and 4-Cell-mediated or delayed hypersensitivity (Type IV). The type of allergy depends on: which components of immune response are involved and how quickly the reaction develops. 1-Immediate (Type I) hypersensitivity: Immediate hypersensitivity or anaphylaxis, typically produces an immediate response upon exposure to an allergy-inducing antigen (also known as allergen). Nonallergic persons do not respond to such antigens. The term anaphylaxis is derived from the Greek ana, which means ”away from” and phylaxis, which means “protection”. Anaphylactic reactions are mediated by IgE antibodies, which bind through the Fc portion to receptors on mast cells (in tissue)and basophils (in blood). When cross-linked by antigens, IgE antibodies trigger the mast cells and basophils to release pharmacologically active agents that are responsible for symptoms of anaphylaxis. Reactions are rapid, occurring 61  within minutes after challenge, that is, responsible to antigen. Anaphylaxis provided first example of ability of immune system to cause harm. The sequence of events involved in the development of anaphylactic sensitivity can be divided into several phases: 1. Sensitization phase, 2. Activation phase, and 3. Effector phase. 1.Sensitization phase: During this phase, IgE antibody is produced in response to an antigenic stimulus and binds to specific receptors on mast cells and basophils. Thus IgE is the antibody responsible for type I hypersensitivity, formerly 62 called reagin or reaginic antibody. All normal individuals can make IgE antibody specific for a variety of antigens when Ag is introduced parenterally in the appropriate manner. Approximately 50% of population generates an IgE response to airborne antigens (referred to as allergens) that are encountered only on mucosal surfaces, such as the lining of the nose and lungs and the conjunctiva of the eyes. However, 10% of general population develops clinical symptoms (e.g. hay fever) after repeated exposure to these airborne allergens such as pollen grains, mold spores, and animal dander. This syndrome is called atopy (which means uncommon), refers to localized allergic reactions and the term atopic to describe affected patients. Only a small proportion of the population exposed to the same allergen is developed such atopy response. This could be due to the differences in mucosal permeability which may restrict the effects of antigens lodging on the surface. IgE antibody production is T-cell dependent since interleukin-4 (IL-4) produced by CD4+ TH 2 cells is an important requirement for IgE production (since neutralizing antibodies against IL-4 can suppress IgE responses). Also, low level of IgE antibody in nonallergic individuals is maintained by suppressor T cells and by gamma interferon, which decreases IgE production. Thus a balance is maintained between those factors that up-regulate and down-regulate IgE responses. Viral infection may disturb this balance and stimulate IgE-producing B cells. Therefore, allergic sensitization may result from failure of a control mechanism. In any events, once adequate exposure to allergen has been achieved by 63  repeated mucosal contact, ingestion, or parenteral injection, and IgE antibody has been produced, an individual is considered to sensitized. IgE antibody is made in small amounts and very rapidly becomes attached to mast cells and basophils as it circulate them. Mast cells (main effector cells responsible for Type I responses) are found around blood vessels in connective tissue, in the lining of the gut, and in the lungs. Mast cells are large, mononuclear, and heavily granulated cells. Basophils are circulating, polynuclear granulated cells also take part in Type I responses and functions in the same way as the tissue-based mast cells. Once bound, IgE molecules persist at cell surface for weeks, and that cell will remain “sensitized” as long as enough antibody remains attached, and will trigger the activation of cell when it comes into contact with Ag. Sensitization may also be achieved passively by transfer of serum that contains IgE antibody to a specific antigen. 2.Activation phase: Anaphylactic reaction may be triggered by injection of specific antigen into the skin of a sensitized individual. This is commonly referred to as the challenge. When such challenge is performed by intradermal injection into the skin, may result in local cutaneous (localized) anaphylaxis; when challenging allergen is distributed throughout the body (following IV injection), the challenge may trigger systemic (generalized) anaphylaxis. The response to intradermal challenge, called “wheal and flare”, is characterized by erythema (redness due to dilation of blood vessels) and edema (swelling produced by release of serum into tissue). Anaphylactic reaction is the most rapid of all hypersensitivity reactions 64 and reaches its peak within 10-15 minutes, then it fades without leaving any residual damage. Size of local skin reaction is roughly indicative of the degree of sensitivity to that particular substance. Activation phase begins with triggering of mast cell to release its granules, and their pharmacologically active contents. It requires that at least two of the receptors for Fc portion of IgE molecules be bridged together in a stable configuration. This linkage is accomplished: by 1)a multivalent Ag that can bind a different molecule of IgE to each of several epitopes on its surface, thus cross-linking them and effectively triggering the cell to respond, or by 2)addition of antibody that is specific for IgE molecules, or by 3)addition of antibody that is specific for IgE receptor molecules on surface of mast cells. -Also, mast cell may be activated by anaphylatoxins C3a and C5a products of complement activation, as well as various drugs such as codeine, morphine; or by physical factors such as heat, cold, or pressure, e.g. in cold-induced urticaria (an anaphylactic rash induced in certain individuals by chilling an area of skin). Triggering of mast cell by bridging of its receptors initiates a rapid and complex series of events culminating in the degranulation of mast cell and the release of pharmacologically potent molecules (granules moved to cell surface, fuse with cell membrane and the contents are released to the exterior). Mast cell can release some or all of its granules, depending on extent of cross-linking on cell surface. Mast cell not lysed or died after degranulated, 65 but regenerate its content. 3.Effector phase: Symptoms of anaphylaxis are entirely attributable to the pharmacologically active materials released by the activated mast cells, which include: Preformed mediators (stored in granules): A)Histamine: binds rapidly to a variety of cells via two major types of receptors, H1, causing constriction of smooth muscle and increase vascular permeability of endothelial cells; and H2, which involved in increase vascular permeability, increase mucous secretion, and release of acid from stomach mucosa. Major signs in systemic anaphylaxis: asthma (difficulty in breathing), asphyxiation (constriction of bronchi smooth muscle), drop in blood pressure due to increase vascular permeability. H1 receptors, but not H2, are blocked by antihistamines. B)Serotonin: in some species, not in human. It causes contraction of smooth muscle and increase vascular permeability. C)Eosinophilic chemotactic factor: attract eosinophils to site of release, which has a role in parasitic worm infection. Then eosinophils activated and degranulated giving histsaminase and helminthotoxin. D)Neutrophil chemotactic factors. E)Heparin : inhibit coagulation. Newly synthesized mediators: Synthesized & released from mast cells after degranulation has occurred. F)Leukotrienes: slow-reacting substance of anaphylaxis (SRS-A) is another mediator that cause a slow, long-lasting airway constriction. SRS-A consists of 3 leukotriene mediators which cause prolonged constriction of smooth muscle. 66 G)Prostaglandins: vasoactive agents which cause broncho-constriction and chemotactic for neutrophils, eosinophils, basophils, and monocytes. H)Platlet-activating factor. Clinical aspects of Type I: Release of histamine locally as a result of intradermal injection of antigen (such as mosquito saliva), would induce a typical “wheal and flare” reaction consisting of blood vessel dilation (flare) and increase in permeability (wheal). Release of histamine systemically, would induce much more severe consequences include: difficulty in breathing (asthma) because of constriction of bronchiolar muscles, uterine cramps, or involuntary urination and defecation. In addition, widespread vascular permeability could produce a massive loss of fluid into tissue spaces (hives and edema) and a drastic fall in blood pressure (shock). The symptoms of anaphylaxis in human are confronted with three different possibilities of lethal outcome: (1)asphyxiation from laryngeal of edema, (2)suffocation from bronchiolar contraction, or (3)loss of adequate blood pressure from overwhelming peripheral edema. Many allergic reactions are due to food allergens. Various foods or their metabolic derivative sensitize GIT, leading to production of IgE antibodies. Subsequent exposure to same allergen result in mast cell degranulation and release of mediators that cause changes in gut permeability, with possible dissemination of Ag-Ab complexes into distal areas of the body such as skin or lungs, initiating urticaria or asthmatic disorders. Localized anaphylaxis. Atopy refers to localized allergic reactions. Atopic immune reactions 67 occur first at site where allergen enters the body. If allergen enters the skin, it causes a wheal and flare reaction, characterized by redness, swelling, and itching. If allergen is inhaled, mucous membranes of respiratory tract become inflamed, and the patient has a runny nose and watery eyes. If allergen is ingested, mucous membrane of digestive tract become inflamed, and the patient may have abdominal pain and diarrhea. Some ingested allergens, such as foods & drugs, also cause skin rashes. Hay fever, or seasonal allergic rhinitis, is a common kind of atopy. Hay fever can be distinguished from common cold by the increased numbers of eosinophils in nasal secretions. Finding elevated numbers of eosinophils in blood also suggests allergy (or infection with helminthes). Generalized anaphylaxis. Some are severe, and immediately life-threatening. In sensitized person, a generalized reaction begins with sudden reddening of skin, skin itching, and hives, especially over the face, chest, and palms of hands. Disorder can then progress to respiratory anaphylaxis or anaphylactic shock. 1-In respiratory anaphylaxis airway become severely constricted and filled with mucous secretions, and allergic individual may die of suffocation. Asthma is caused by inhaled or ingested allergens, emotional stress, aspirin, or cold, dry air, or to endogenous m.os. (e.g. Moraxella catarrhalis). 2-In anaphylactic shock: blood vessels suddenly dilate and become more permeable, causing an abrupt and life-threatening drop in blood pressure. Insect bites and stings are a common cause in people sensitized to insect venoms. Anaphylactic shock must be treated immediately. Unless epinephrine (adrenaline) is administered immediately, death can occur. Epinephrine acts by relaxing smooth muscle of respiratory passage ways 68 and constricting blood vessels. 2-Cytotoxic (Type II) hypersensitivity: (Mediated by IgM and IgG) In cytotoxic hypersensitivity, specific antibodies react with cell surface antigens interpreted as foreign by immune system, leading to phagoctosis, killer cell activity, or complement mediated lysis. The cells to which antibodies are attached, as well as surrounding tissues, are damaged because of the resulting inflammatory response. Antigens that initiate cytotoxic reaction typically enter the body in mismatched blood transfusions or during delivery of an Rh-positive infant to an Rh- negative mother. Mechanism of cytotoxic reactions: When an antigen on cell surface is first recognized as foreign, B cells become sensitized and ready for antibody production upon a subsequent antigen exposure. During subsequent exposures with surface antigen, antibodies bind to antigen activate complement. Phagocytic cells, such as macrophages and neutrophils, are attracted to the site. This type appear to be responsible for the tissue damage in cases of: 1.rheumatic fever following a streptococcal infection, 2.in certain viral diseases, 3.in transfusion reaction, and 4.in hemolytic disease of the newborn (mother-infant Rh incompatibility). Examples of cytotoxic reactions: Transfusion reactions: Normal human RBCs have genetically determined surface antigen (blood group systems) that form the basis for different blood types. Transfusion reaction can occur when matching antigens and antibodies are present in patient‟s blood at the same time. Antigens A and antigens 69 B are responsible for determination of ABO blood group system. Four blood types- A, B, AB, and O- are named according to whether RBCs have antigen A, antigen B, both A and B antigens, or neither antigen. Normally, human serum has no IgM antibodies against antigens present on own RBCs. In sensitized patient receives RBCs with different RBCs antigen during blood transfusion, IgM antibodies cause a Type II hypersensitivity reaction against foreign antigen. Foreign RBCs are agglutinated (clumped), complement is activated, and hemolysis (rupture of RBCs) occurs within blood vessels. Symptoms include fever, low blood pressure, back and chest pain, nausea and vomiting. 70 Hemolytic disease of newborn: (Erythroblastosis fetalis) In addition to ABO blood group, RBCs can have Rh antigens. Blood with Rh antigens on RBCs is designated Rh-positive, cells lacking Rh- antigen are designated Rh-negative. Anti-Rh antibodies normally are not present in serum of either Rh- positive or Rh-negative blood. Consequently, sensitization is necessary for an Rh antigen-antibody reaction. Sensitization occurs when an Rh-negative mother carries Rh-positive fetus (inherited from father). Fetal Rh antigen rarely enters mother‟s circulation during pregnancy but can leak across the placenta during 71  delivery, miscarriage, or abortion. Rh-negative mother becomes sensitized to Rh antigen and can produced anti-Rh antibodies upon re- exposure to the same Rh antigen. Because sensitization usually occurs at delivery, first Rh-positive child rarely suffers from hemolytic disease. But on subsequent pregnancies, sensitized Rh-negative mother produces Anti-Rh antibodies (IgG) that cross the placenta and cause a Type II reaction in Rh-positive fetus. If this occurs, fetal RBCs agglutinate, complement is activated, and RBCs are destroyed. The result is hemolytic disease of newborn. Baby is born with an enlarged liver and spleen caused by efforts of these organs to eliminate damaged RBCs. Such babies exhibit yellow skin color of jaundice due to excessive bilirubin- a product of breakdown of RBCs- in their blood. Treatment: by giving Rh-negative mother IM injections of Anti-Rh IgG antibodies within 72 hours after delivery. Antibodies presumably bind to Rh antigens on fetal RBCs that have leaked into mother‟s blood. These Anti-Rh antibodies destroy the fetal RBCs before they can act to sensitize her immune system. 3-Immune complex (Type III) hypersensitivity: Immune complex hypersensit