Nonspecific host defensese. the immune response. types and forms of immunity. Nonspecific host defence factors of oral cavity

Antigens. The main attributes of antigenes. Antigens structure of bacterial cell and viruses. Methods of obtaine of bacterial antigenes. Immunoglobulins. Structure and classes. . Immunoglobulins of human oral cavity. Examination of immunoglobulins concentration in blood serum. Major histocompatibility complex

The structure of the immune system (Т– and B– system).  Cooperation of cells in immune response. Mechanism of antibacterial and antiviral immunity

 

 

 

Nonspecific host defenses. Types and  forms of immunity.

 

The term immunity (L. immunis freed from) usually means resistance of the body to pathogenic microbes, their toxins or to other kinds of foreign substances.

Immunity is a complex of physiological defence reactions which determine the relative constancy of the internal medium of the macroorganism, hinder the development of the infectious process or intoxication, and are capable of restoring the impaired functions of the organism.

In the process of evolution, organisms have developed the property of distinguishing ‘self and ‘non-self very accurately, which is just what protects them from being penetrated by foreign proteins, including pathogenic micro-organisms and heterogenic transplants. The ‘non-self is detected by the lymphocyte receptors.

Insusceptibility to infectious diseases depends on many factors grouped under the names of resistance and immunity. Resistance is the insusceptibility of the body to the effect of pathogenic factors. Resistance embraces a wider group of phenomena of insusceptibility than immunity. Non-specific resistance is the insusceptibility of the body to injury by pathogenic factors: mechanical (traumas, rocking), physical (barometric pressure, cooling, overheating, radiation energy, ionising radiation), chemical (oxygen deficiency, excess of carbon dioxide, action of poisonous substances, drugs, poisons of a chemical and bacterial origin), and biological (pathogenic protozoa, fungi, bacteria, rickettsiae and viruses).

There may be resistance of the entire body and of its separate systems, although mutual dependence of both exists. Resistance is associated with the anatomical-physiological characteristics of the body, development of the central nervous system, and endocrine glands. It depends on the phylogenetic development of the animal, the individual and functional state of the body, and in man it depends also on social factors. Mental traumas predispose to somatic and infectious diseases; chronic hunger and vitamin deficiencies lead to a decline in resistance; intoxication by alcohol, opium, cocaine and other narcotics has a negative effect on human resistance.

In the traditions and life of ancient people considerable allowance has been made for preventive measures including vaccines against various diseases. Thus, for example, the inhabitants of East Africa from time immemorial have successfully used vaccinations against the bites of poisonous snakes. For vaccines they used snake venom, contained in a paste from plants. The paste, applied to cross-like scarifications on the skin of the person being vaccinated, caused a prolonged inflammation, and after being absorbed gradually helped in producing immunity to the lethal bites of poisonous snakes. Repeated vaccinations were made over a period of several years. Africans produced an artificial immunity to tick-borne relapsing fever by natural immunization. They carried on their body ticks which had contained a virus for a long time.

 

Types and Forms of Immunity

Modem classification subdivides immunity into two types according to origin: (1) species inherited and (2) acquired.

Species immunity is insusceptibility of certain species of animals to diseases which attack other species. It is transmitted by heredity from one generation to the next. An example of species immunity is insusceptibility of man to cattle plague, chicken cholera and infectious horse anaemia. On the other hand, animals are not infected by many human infections such as enteric fever, scarlet fever, syphilis, measles, etc.

Species immunity is the result of a long evolution of interrelations between the macro-organism and pathogenic micro-organism. It depends on those biological peculiarities of a given species of organism, which were formed during historical development in the course of natural selection, variation and adaptation to the environmental conditions.

The underlying factors of the mechanisms of species immunity (hereditary resistance) to infectious diseases are the absence in the organism’s cells of receptors and substrates necessary for the adsorption (attachment) and reproduction of the causative agent, the presence of substances which block the reproduction of pathogenic agents, and the ability of the macro-organism to synthesize various inhibitors in response to the penetration of the pathogenic microbes.

Acquired immunity is subdivided into natural and artificial. Natural immunity in turn is divided into (1) active, that is acquired following an obvious (postinfection) or latent disease or repeated infection without clinical manifestations, (2) passive immunity of the newborn (maternal, placental), i e. immunity iewly born children, acquired from the mother in the period of intrauterine development, through the placenta in the process of ontogenesis. The duration of immunity of the newborn is short. After about six months this immune state disappears and children become susceptible to many infections (measles, diphtheria, scarlet fever, etc.). Artificial immunity is reproduced by active or passive immunization.

 

 

Immunity is manifested on the cell, molecular, and organism levels. The organism’s immune  system is a sum total of lymphoid organs consisting of central (the thymus, bone marrow) and peripheral (lymph nodes, spleen, lymphocytes of peripheral blood) organs.

The thymus is the central organ of cell immunity in which the differentiation of stem cells into immunologically competent T-lymphocytes occurs. The function of these lymphocytes is discussed in the corresponding sections.

The systems of T-lymphocytes determined cell immunity in tuberculosis, leprosy, brucellosis, tularaemia and other diseases. The Fabricius’ pouch in birds (its analogue in mammals are thePeyer’s patches) is the central organ of humoral immunity.

The system of B-lymphocytes is responsible for humoral immunity against most bacterial infections and intoxications.

 

 

Non-specific Resistance

This form of defence which includes defensive properties is associated with phagocytosis, barrier function of the skin, mucous membranes, lymph nodes and other tissues and organs.

Phagocytosis. The most ancient form of immunity is phagocytosis, a defence adaptation which entails the seizure and digestion of foreign particles, including bacteria and remains of disintegrated cells, by phagocytes. The phenomenon of phagocytosis is of great importance in defence reactions of heritable and acquired immunity. I. Metchnikoff established that amoeboid cells of the mesoderm in transparent marine animals are capable of swallowing and digesting various foreign particles.

Video – Phagocytosis

Video – Phagocytosis

During early embryonic development amoeboid (mesenchymal) cells are produced between the epithelial cells, which do not take part in building up organs, from which all types of motile erythrocyte cells and various species of migrating leukocytes originate. They are contained in the thymus, bone marrow, spleen, lymph nodes, tonsils, appendix, and interstitial tissues of parenchymatous organs.

For more than a quarter of a century, I. Metchnikoff and his pupils accumulated facts confirming the defence role of phagocytosis during infection of vertebrate animals with pathogenic microbes. This provided for the possibility of establishing in the evolution and phylogenesis of cells the relation between digestion and phagocytosis. I. Metchnikoff subdivided those cells able to carry out phagocytosis into microphages and macrophages.

Microphages include granular leucocytes, neutrophils, eosinophils and basophils, of which only neutrophils have quite a marked ability for phagocytosis. Eosinophils and basophils are characterized by a weak phagocytic activity, although this problem has not yet been studied sufficiently.

Phagocytosis takes place with the help of macrophages which may be motile (monocytes of the blood, cells of the lymph nodes and spleen, polyblasts, histiocytes, etc.) or non-motile (reticular cells of the spleen, cells of the lymphatic tissue, endothelium of the blood vessels, etc.).

Much importance in the mechanism of the phagocytic reaction is attached to lysosomes (oval three-layer structures) which possess bactericidal properties in relation to different bacterial species and are capable of destroying foreign substances.

The complex of lysosome enzymes and the permeability of the lysosome membranes are determined genetically. Phagocytosis differs in the accomplishment of its defence function depending on the completeness of the set of the lysosome enzymes and the function of the lysosome membranes.

 

The process of phagocytosis consists of four phases. The first phase involves the approach of the phagocyte to the microbe by means of a positive chemotaxis. Under the influence of the productsof the life activities of microbes excitation of the phagocytes occurs, which leads to a change in the surface tension of the cytoplasm, and gives the phagocytes amoeboid motility.

In the second phase adsorption of the micro-organism on the surface of the phagocyte takes place. This process is completed under the influence of an electrolyte which alters the electrical potential of the phagocytized object (microbe).

The third phase is characterized by submergence of the microbe into the cytoplasm of the phagocyte, which seizes minute objects quite rapidly and large ones (some protozoa, actinomycetes, etc.) are engulfed in pieces.

The phagocytosed bacteria perish under the bactericidal effect of the heightened hydrogen ion concentration due to an increase of lactic acid in the cytoplasm of the phagocytes.

In the fourth phase intracellular digestion of the engulfed microbes by the phagocytes takes place.

 

In the process of phagocytosis various changes in the microbes can be observed, e. g. the production of granules in cholera vibrios, swelling of enteric fever bacteria, fragmentation of diphtheria bacilli, destruction of anthrax bacilli and swelling of cocci. Eventually, the phagocytized microbes become completely disintegrated.

Factors which speed up phagocytosis include calcium and magnesium salts, the presence of electrolytes and antibodies (opsonins and bacteriotropins), histamine, pyrogenic substances capable of raising the temperature of the tissues and the entire organism. Phagocytosis proceeds more vigorously in the immune than in the non-immune organism.

 

 

 

 

 

 

 

 

 

 

Toxins of bacteria, leucocidin, capsular material of bacteria, cholesterol, quinine, alkaloids and also a block of the reticuloendothelial system inhibit phagocytosis.

Besides complete phagocytosis incomplete phagocytosis is observed in certain diseases (gonorrhoea, leishmaniasis, tuberculosis, leprosy) in which micro-organisms are absorbed by phagocytes, but do not perish, are not digested, and in some cases reproduce.

Viruses are also digested in the macrophages of immune animals under the effect of the acid content of the vacuoles and the enzymes of the phagocytes though, unlike bacteria, viruses are intracellular parasites and are capable, to a great degree, of resisting phagocytosis.

 

 

 

The skin, mucous membranes and lymph nodes. The data published by I. Metchnikoff on phagocytosis created wide interest and called for the necessity of carrying out numerous experimental investigations, as a result of which the defence mechanism of the skin, mucous membranes, lymph nodes and cells of many tissues and organs was established.

In a normal, uninjured state, the skiot only is a true mechanical protective barrier, but a bactericidal factor. It has been established that the clean skin of a healthy person has a lethal action on a number of microbes (haemolytic streptococcus, salmonellae of enteric fever and paratyphoid fever, colibacillus, etc.). Investigations confirmed that washing the hands not only aids in mechanically removing microbes from the surface of the skin, but also in increasing its bactencidal properties.

The mucous membranes of the eyes, nose, mouth, stomach and other organs have defence adaptations. Like the skin barrier, the mucous membranes perform antimicrobial function as a result of their impermeability to different microbes and the bactericidal action of their secretions. In lacrimal fluid, sputum, saliva, blood, milk, tissues and organs lysozyme is found. It is found in some bacterial cells.

Due to the establishment of this defence mechanism the biological role of lacrimal fluid, saliva, nasal mucus and sputum becomes apparent. A lack of lysozyme in the tears affects the cornea. When animals lick their wounds they transfer lysozyme into them. Microbes which have penetrated into the mucous membranes are continuously destroyed by the action of lysozyme. Nasal mucus is bactericidal for many microbes and viruses of influenza, herpes, poliomyelitis, etc.

Bactericidal properties are not limited to the action of lysozyme. There are other antibiotics produced by the organs and tissues, which are capable of inhibiting microbes. A special substance inhibin has been found in the saliva, and the antibiotic erythrin — in the erythrocytes Both preparations have a bacteriostatic action on diphtheria bacilli Interferon is one of the powerful inhibitors of viruses.

Of a certain significance in physiological immunity is hyaluronic acid which inhibits the penetration of microbes into tissues and organs. Gastric juice has quite marked bactericidal properties in relation to many causative agents, especially those of the Salmonella group and organisms responsible for food poisonings.

Besides the defence adaptations of the skin and mucous membranes, a large role is played iatural immunity by the lymph nodes in which the pathogenic microbes penetrating through the injured skin and mucous membranes are localized and rendered harmless. Inflammation develops in the lymph nodes.

The inflammatory reaction is characterized by the liberation from the tissues of a number of substances (leucotoxin, leucopenic factor, histamine, serotonin, etc.) under the influence of which changes in the leucocytes occur. As a result they become sticky and adhere to the capillary wall, where they enter into the tissues. They enhance (induce) proliferation of adjacent cells. Leucocytes accumulated in the inflammatory zone produce a protective barrier which hinders the spreading of microbes into the tissues, blood and organs. Phagocytosis plays a great role in the blocking and destruction of micro-organisms in the inflammation focus.

In the inflammatory focus the temperature rises and acidosis and hypoxia develop, which cause a fatal effect on the bacteria and viruses. The reproduction of bacteria and viruses diminishes in the acid medium and virus adsorption by the susceptible cells reduces.

An increase in the body temperature of the macro-organism also suppresses the activity of bacteria and viruses. Fever is considered an important factor in the recovery from a virus infection.

 

The excretory functions of the kidneys and saliva, the secretions of the respiratory passages, intestine, and mammary and sweat glands are a powerful factor of non-specific immunity.

 

 

Insusceptibility associated with the bactericidal properties of the blood is a later form of defence inherent in vertebrates.

A substance which has bactericidal properties with regard to a number of micro-organisms (causative agents of anthrax, tetanus, botulism, gas gangrene, and diphtheria, and staphylococci, pneumococci, bovine brucellae, etc ) is beta-lysin which is a substance of a complex nature, a thermostable fraction of normal serum, decomposing at temperatures of 63-70°C or under the action of ultraviolet rays. From human serum a fraction was isolated which is characterized by a bactericidal action in relation to diphtheria bacilli, and is not identical to beta-lysin.

From the blood of people with an elevated temperature a component X-lysin was isolated which dissolved mainly Gram-negative micro-organisms (meningococci, paratyphoid bacteria) and to a lesser degree –  Gram-positive organisms X-lysin acts without the participation of complement, and is thermostable (withstands a temperature of 68-100 °C).

Leukines,  thermostable substances freed of leucocytes, pertain to bactericidal substances They disintegrate at a temperature of 75-80 °C. Leukines render harmless Gram-positive as well as Gram-negative bacteria.

C-reactive protein (the name is associated with C-polysaccharide of the type II St pneumoniae) having immunological properties was discovered m 1930 in the serum of patients with pneumococcal diseases. C-reactive protein is considered to be conjugated with reactive, defensive, non-specific natural processes It has also been found in the serum of patients with typhus fever, tuberculosis and other infections The component parts of urine, prostatic fluid, extracts from the liver, brain, spleen and other tissues and organs are characterized by bactericidal properties.

 Interferons are glycoproteins that block virus replication and exert many immunomodulating functions. Alpha interferon (from leukocytes) and beta interferon (from fibroblasts) are induced by viruses (or double-stranded RNA) and have antiviral activity. Gamma interferon is a lymphokine produced primarily by the Th-1 subset of helper T cells. It is one of the most potent activators of the phagocytic activity of macrophages, NK cells, and neutrophils, thereby enhancing their ability to kill microorganisms and tumor cells. For example, it greatly increases the killing of intracellular bacteria; such as M tuberculosis, by macrophages. It also increases the synthesis of class I and II MHC proteins in a variety of cell types. This enhances antigen presentation by these cells.    

Under the influence of the virus the cells of affected tissues excrete interferon which does not have a specific action, but renders viruses harmless. Interferon is present m small amounts iormal human serum.

 

 

 

 

A tuberculostatic factor has been discovered in human blood which is characterized by the ability to kill tubercle bacilli In 1954 L Pillemer established that after treating serum with zymozan (obtained from yeasts) it loses its bactericidal activity A precipitate is formed m the serum After treating the precipitate a substance was isolated, which, on addition to serum, restored the lost bactericidal activity This substance was named properdin (L. perdin destroy). Properdin is a serum protein, an euglobulm, which plays an important part in immunity The greatest amount is found in the blood of rats then in  a decreasing order in the blood of mice, cows, pigs, humans, rabbits,  sheep and guinea pigs. It is possible that properdin is composed of a group of antigens related to thermolabile class M immunoglobulins.

The synthesis of complement, properdin, lysozyme, interferon, and other natural inhibitors is determined genetically, inherited, and belongs to the factors of species immunity.

A great role in humoral activity is played by antibodies (immunoglobulins), the origin and accumulation of which occur under the influence of antigens.

Investigations have confirmed that microbes which have penetrated into the blood are rendered harmless by substances in the plasma. J. Fodor, G. Nuttall and others established the bactericidal action of  blood, exudates and other fluids of animals and humans. G Buchner showed that serum has a lethal effect on microbes, but on heating it defensive forces considerably weaken. The bactericidal matter of fresh normal serum at first was named alexin (Gk alexin to ward off) then complement (L. complementum anything which completes) Since the complement dissolves some species of bacteria and cells, it is sometimes called lysin (alpha-lysin).

It was observed too that antiserum could exert two entirely different effects on gram-negative bacteria or red blood cells (RBCs). In one case, when fresh antiserum was used, such cells were lysed. On the other hand, if the antiserum was heated to 56°C for 30 minutes or aged about 1 week, it could no longer cause lysis but instead would agglutinate the bacteria or RBCs. However, when fresh normal serum, such as guinea pig serum, was added to the heated or aged antiserum, the ability to cause cell lysis was regained. The lytic effect, therefore, requires two factors: (1) specific antibody, and (2) a labile component present iormal serum. This latter substance has been given the name complement. Subsequent research has revealed that complement is a multicomponent system composed of many different proteins.

The activation of the complement proteins proceeds by two triggered enzyme systems, wherein a series of inactive proteolytic enzymes (zymogens) are converted into biologically active proteases, each possessing an extremely fine specificity for its substrate. There are two pathways for the activation of the system, each initiated by a different sequence of events. The classic pathway of complement activation is set in motion by antigen-antibody complexes, whereas the alternate pathway, which is phylogenetically much older, is entirely independent of antigen-antibody reactions. Instead, certain components are activated by the presence of a series of foreign substances, not the least of which are infecting bacteria and viruses. The two pathways, however, have much in common, particularly the fact that their final membrane-attack components are identical.

Biologic Function of the Complement System. Before the details of the classic pathway of complement activation are outlined, the biologic function of this system should be considered. This function can best be appreciated by noticing that after antibody has reacted with its antigen, it can do little more. In other words, antibody might precipitate an antigen or, if the antigen is cellular, might cause agglutination but, with the exception of the neutralization of toxins or of virus infectivity, antibody alone is an ineffective means of protection against infection. Thus, for practical purposes, the major function of an antibody is to recognize a foreign antigen and bind to it. By doing so, it provides a site for phagocyte interaction and for the initiation of the reactions of the complement system. It is the activation of this system that (1) leads to the lysis of foreign cells, (2) further enhances phagocytosis of invading microorganisms, and (3) causes local inflammation, stimulating the chemotactic activity of the host’s leukocytes.

In addition, after activation of the system, interaction of one or more complement components with specific receptors on cell surfaces can result in (1) enhancement of antibody-dependent cellular cytotoxicity (ADCC); (2) increased oxidative metabolism; (3) secretion of vasoactive amines and leukotrienes;  (4) secretion of monokines; (5) stimulation of prostaglandin and thromboxane pathways; (6) modulation of lymphocyte activation and antibody responses; and (7) mobilization of leukocytes from the bone marrow. The following sections are concerned with a step-by-step dissection of the component parts and reactions of this system and the role that it plays in the destruction of foreign cells.

 

 

Classic Pathway of Complement Activation. The operation of the complement system consists of anumber of reactions, each of which activates the next reaction in the series. A primary event must occur, however, to initiate the reactions that eventually involve the many components of the complement system.

In the case of the classic pathway, the initiating event occurs when the first component of complement reacts with antigen-antibody complexes in which the antibody is either IgM or IgG. IgA, IgD, and IgE are not effective in activating complement.

Once initiated, the activation of the complement system may have various effects, depending on the type of foreign cell involved in the antigen-antibody reaction. In the case of a gram-negative bacterium, the integrity of the cell membrane is destroyed, permitting the lysozyme-mediated lysis and death of the cell. Gram-positive organisms are not lysed, but the activation of complement by a gram-positive cell and antibody results in the release of fragments of complement components that aid in phagocytosis by binding to the antigen, providing a receptor for the host leukocyte. In addition, many eukaryotic cells, such as RBCs or virus-infected cells, are lysed by complement. The complex reactions that produce all these effects can be divided into three series of reactions involving the complement system: (1) the activation of the recognition unit, (2) the assembly of the activation unit, and (3) the assembly of the attack unit.

 

THE RECOGNITION UNIT. Of the nine known components of the classic pathway, only the first   component, C1, is involved in the recognition unit. This componentis composed of three different proteins—C1q, C1r, and C1s—which interact with each other on an antigen-antibody complex. It appears that antigen-antibody reactions bring numerous antibody molecules into aggregates that can be cross-linked by C1q. This clustering of antibody molecules can be mimicked by mild heating or chemically cross-linking antibodies to form an aggregate in the absence of antigen. Such artificial aggregates easily react with C1q, leading to complement activation.

The reaction begins when C1q, the recognition subunit of C1, binds to the constant region of the antibody complexes. The binding site for C1q is located in the fourth domain (C_H4) of IgM and in the second domain (C_H2) of IgG. Some variability exists in the capacity of the IgG subclasses to activate complement. The reason for this is unknown.

C1r and C1s are both present in the C1 complex as inactive proteases. Limited amino acid sequence studies have shown considerable homology between these two components, as well as some homology with other serine proteases. In the presence of Ca²⁺ they are bound to the collagen portion of C1q as a tetrameric complex consisting of two molecules each of C1r and C1s [(C1r₂C1s₂C1q]. In solution, C1q, C1r, and C1s exist in an easily dissociable complex, but after C1q cross-links antibody, this association becomes much more stable, presumably because of a conformational change in C1q. The actual binding to C1q appears to occur through C1r, because isolated C1r will bind to antibody-C1q, whereas C1s will not.

After binding to antibody-C1q, a conformational change occurs in C1r, which exposes an enzymatic site that catalyses its own hydrolysis, converting it into an active serine protease (C1r) whose only known substrate is C1s. (Complement components that have been modified to become enzymatically active are written with a bar over the complement designation, as in C1r). Once activated, C1r splits off a peptide from C1s, converting C1s also into a serine protease, C1s, which initiates the assembly of the activation unit.

ASSEMBLY OF THE ACTIVATION UNIT. The first step in the assembly of the activation unit occurs when C1s splits off a 77-amino acid vasoactive polypeptide (C4a) from the N-terminal chain of C4 to generate C4b. This exposes an intrachain thioester bond within C4b, which is the reactive site responsible for attachment to the cell membrane or to the cross-linked antibody molecules bound to C1q or to those immediately adjacent. Unbound molecules of C4b are rapidly inactivated. C1s also cleaves C2 into two components: C2a (a 70,000-dalton fragment) and C2b (a 30,000-dalton polypeptide).

At this point, C2a remains bound to the antibody-bound or membrane – bound C4b to form a new  active protease, C4b2a, which is called C3 convertase. The catalytic site of C3 convertase exists in the C2a fragment, and the role of C4b appears to be one of binding C2a, thus stabilizing the C3 convertase. The newly formed C3 convertase, C4b2a, cleaves C3 into two fragments, C3a and C3b, again exposing a highly reactive thioester bond in C3b. The C3b molecule reacts with residues on the cell surface, on the antigen-antibody complex, or on the C3 convertase itself, forming a new enzymatically active complex, C4b2a3b, called C5 convertase. Those C3b molecules that do not immediately react with sites nearby are inactivated by hydrolysis of this thioester bond. This C4b2a3b is the activation unit of the classic pathway, and its function is to split C5 into C5a and C5b, which initiates the formation of the membrane-attack complex. Interestingly, both C3 convertase and C5 convertase use the same catalytic site in the bound C2a. In the C5 convertase molecule, C3b acts as a binding site for the C5 substrate. It seems probable, however, that the C5 convertase continues to split C3 into C3a and C3b.

ASSEMBLY OF THE MEMBRANE-ATTACK COMPLEX. The splitting of C5 by C5 convertase is the last enzymatic reaction involved in the classic pathway of complement activation; the subsequent assembly of the remaining components of the attack complex are nonenzymatic, occurring spontaneously as follows:

1. C5b reacts with one molecule of C6, forming a relatively stable complex of C5b6.C5b alone is very unstable in serum, with a half-life measured in milliseconds; however, C5b is stabilized by reacting with C6 while still bound to the C3b of the C5 convertase.

2. In the fluid phase, C7 is added to C5b6 to create C5b67, the first component of the attack complex that has membrane-binding properties. C5b67 has a half-life of about 100 milliseconds in the fluid phase, but it is stabilized by binding directly to the membrane, independent of C3b.

3. C5b67 now expresses a C8-binding site, and when C8 binds to the complex, it is inserted into the lipid bilayer. At this point, the complex is capable of slow lysis of susceptible cells, which becomes more rapid with the addition of C9 in the next step. However, animals deficient in the C9 component are able to deal with most infections normally.

4. Aggregation of C5b-8 within the membrane then occurs, resulting in formation of a C9-binding site. Interaction of C9 with this site causes a conformational change in C9, exposing hydrophobic regions and facilitating its insertion into the membrane and its polymerization into a poly-C9 complex.

 

 

 

Structure of the C1 Molecule. The C1 protein is composed of three proteins: C1q, which binds to the Fc portion of the Ab molecule; C1s, which can enzymatically cleave the next complement component, C4; and C1r, which acts as a bridge connecting C1q to C1s.

 

 

 

 

 

 

 

 

 

 

OTHER MECHANISMS OF ACTIVATING THE CLASSIC PATHWAY. The classic pathway of complement activation is normally thought of as one initiated by antigen-antibody complexes but, actually, it involves C1, C2, and C4 to form the C3 convertase, because the alternate pathway of activation uses a different C3 convertase. With this definition, several other elements can be added that activate the classic pathway, namely, viral membranes, the lipid A portion of endotoxin, mitochondrial membranes, and miscellaneous polycations and polyanions such as heparin, protamine, and nucleic acids. The mechanism whereby these compounds initiate C1 activation is unknown, but it can be assumed that any substance possessing binding sites for C1q might initiate this pathway.

Interestingly, nonprimate retroviruses activate primate C1 directly. This activation is dependent on the C1s portion of C1. This may be the primary mode of defense against these viruses, because it is known that primates do not produce antibody to these nonprimate retroviruses. It may be that the viruses are cleared from the body by activating the classic pathway before antibody synthesis is induced.

 

Alternate Pathway of Complement Activation. The alternate pathway of complement activation (the properdin pathway) does not require the presence of antibodies for initiation and, as a result, provides a mechanism of nonspecific resistance to infection. Moreover, this pathway does not use C1, C4, or C2, which are the early reactants in the classic pathway of complement activation. Remember, however, that the overall result of this pathway is the same as that of the classic pathway: C3 is split into C3a and C3b, and C5 is cleaved to form C5a and C5b, thus permitting the spontaneous formation of the C5b-9 membrane-attack complex. The enzymes catalyzing these conversions are different from the C3 and C5 convertases described for the classic pathway of complement activation.

RECOGNITION AND ASSEMBLY. This pathway bypasses both the recognition unit and the  assembly of the activation unit as described for the classic pathway. Instead, there arc at least three  normal serum proteins that, when activated together with C3, form a functional C3 convertase and a C5 convertase. These are factor B, factor D, and properdin (P). To fully comprehend this pathway, the reader should keep the following facts in mind:

1. These are normal serum proteins, and the alternate pathway routinely undergoes activation in the absence of any stimulus.

2. In the absence of initiators (which is discussed later), the initial complexes of the alternate pathway are rapidly destroyed.

3. In the presence of these initiators, such complexes are stabilized, and complement is activated to form the identical membrane-attack complexes described for the classic pathway.

 

 

 

Reaction Sequence. The pathway appears to be initiated in the following steps:

1. The C3 molecule has an internal intrachain thioester bond formed by the reaction of the side chains of two amino acids: cysteine and glutamic acid. In the blood, this thioester bond reacts with a molecule of H₂O to form the unstable C3-H₂O complex. This unstable, yet active, complex then binds to serum factor B to form another unstable complex, C3B. The factor B portion of the C3B complex is split by factor D (a normal serine protease) into two fragments, Ba and Bb. Ba is a 33,000-dalton peptide that is released during the reaction, and Bb is a 60,000-dalton peptide that remains bound to C3 to form a C3Bb complex. The C3Bb acts, probably in the fluid phase, as an initial C3 convertase to split C3 into C3a and C3b.

2. C3b binds factor B, forming the transient intermediate C3bB, which is then subject to cleavage by factor D, to form C3bBb. This is the active C3 convertase of the alternate pathway.

Notice that there have been two C3 convertascs formed to this point (ie, C3Bb and C3bBb) that differ in the form of the C3 portion but that have the same enzymatic specificity. The C3bBb enzyme, however, is capable of reacting with surfaces (eg, the cell membrane) and thus can be stabilized.

3. As more C3b is generated by C3bBb, it continues to attach to the membrane. When an additional molecule of C3b becomes bound to the C3bBb, the specificity of the convertase is shifted to a C5 convertase (designated C3bBb3b).

4. Properdin enters the reaction sequence and binds to both the C3 and C5 convertase to protect the complex from the action of factor I (a normal component of serum that is capable of inactivating C3b). Properdin interaction, therefore, is the terminal event in the assembly of the activation unit for this pathway.

5. Once the C5 convertase is formed, C5 is cleaved to form C5a and C5b, and the spontaneous formation of the attack complex (C5b-9) quickly follows. The formation of this attack complex proceeds in the same manner as it does in the classic pathway discussed above.

 

 

 

The activation of either the classical complement pathway or the alternative pathway leads to the formation of C3 convertase (C4b2b or C3bBb). Cleavage of C3 and binding of C3b to C3 convertases results in the formation of C5-converting enzymes (C4b2bC3b or C3bBb3b). At the end, both pathways form MAC, which mediates lysis of target cells.

The Lytic Event. It is now well established that membrane-bound C5b-8 can cause slow lysis of a susceptible cell and that the addition of C9 greatly accelerates the cell lysis. However, no real consensus exists concerning exactly how lysis occurs. Aggregation of the C5b-8 complex in the membrane clearly forms a binding site for C9. Interaction of C9 with this site induces large conformational changes in C9, exposing hydrophobic regions responsible for insertion of C9 in the membrane and the formation ofpoly-C9. Poly-C9 is composed of 12 to 18 molecules ofC9 and is the ring-like structure seen in electron micrographs of cells treated with antibody and complement. The poly-C9 structure forms at high concentrations and is not required for efficient lysis. Lysis of the cell to which the membrane-attack complex is bound occurs primarily by extensive disruption of the lipid bilayer, an increase in membrane permeability, and marked changes in membrane potential, pH, and cytosoliccation concentrations, leading to complete loss of electrochemical gradients and rupture of the plasmamembrane.

Biological Consequences of Complement Activation

Complement activatioot only causes cell lysis, but other effects as well. Some of these include: contraction of smooth muscle, release of histamine from mast cells and platelets, enhanced phagocytosis, chemotaxis of phagocytes, and activation of lymphocytes and macrophages.

The main activity of C3a and C5a is anaphylaxis. These cause histamine release from mast cells and basophils, which can affect the activity of smooth muscle. Spasmogenicity, accounts for the ability of these molecules to induce an anaphylactic response in animals. In addition to spasmogenicity, histamine release induced by the anaphylatoxins as effects on inflammation. The cellular responses of neutrophils and monocytes to C5a include (1) degranulation and lysosomal enzyme release, (2) cell adherence, and (3) chemotactic migration.

CHEMOTAXIS. Any substance that attracts leukocytes to an area of inflammation is a chemotactic agent. Factors Ba (the split product from the alternate pathway) and C5a are both chemotactic for PMNs and macrophages, thus contributing to local inflammation. C5b67, the partially formed attack complex, also has been implicated as a chemotactic agent. Interestingly, although the removal of the terminal arginine from C5a by carboxypeptidase B completely eliminates all anaphylatoxic activity, its removal does not affect the chemotactic activity of C5a.

IMMUNE ADHERENCE. C3b is an effective opsonin, stimulating the phagocytosis of antigen-antibody aggregates, cells, and viruses. Its opsonic effectiveness stems from the presence of specific C3b receptors on PMNs, monocytes, macrophages, and mast cells. C3b also binds to antigen-antibody aggregates and to antibody- sensitized cells and viruses. Thus, C3b can act as a bridge to bring antibody-coated material into intimate contact with phagocytic cells, inducing their phagocytosis and destruction.

 

The acute inflammatory response is characterized by symptoms of redness, pain, swelling, and heat due to the action of C4a, C3a, C5a, and histamine. Inflammation’s primary goal is to set into motion a series of events that result in the elimination of foreign and damaged cells. This response can be mediated by the anaphylatoxins and their byproducts.

 

 

Interactions Between Complement and Other Protein Systems. As might be expected, a system as seemingly complex as the complement system does not operate in a void. Proteins of the complement system do interact with proteins of other complex systems, that is, the coagulation, fibrinolytic, and kinin systems. Examples of the interaction between components of the various systems are many. Plasmin has been shown to cleave C3 to give C3b- and C3a-like fragments, which possess chemotactic and anaphylatoxic activities. Protein S of the coagulation system circulates as a complex with C4-binding protein, although the biologic significance of this interaction is unknown. Kallikrein has been shown to cleave C5 with the release of a fragment that is chemotactic for PMNs.

Human platelets react directly with antigen-antibody complexes through cell-surface fg receptors. Experimental results suggest that activation of platelets to release vasoactive, granule-associated amines occurs by mechanisms requiring complement. This may occur because of platelet lysis via the classic pathway of complement activation or by nonlytic mechanisms that also involve complement.

 

 

NATURAL KILLER CELLS

Natural killer (NK) cells play an important role in the innate host defenses.

 

Important features of natural killer (NK) cells

 

They spe­cialise in killing virus-infected cells and tumor cells by secreting cytotoxins (performs and granzymes) similar to those of cytotoxic T lymphocytes and by participating in Fas-Fas ligand-mediated apoptosis. They are called “natural” killer cells because they are active without prior exposure to the virus, are not enhanced by exposure, and are not specific for any virus. They can kill without antibody, but antibody enhances their effectiveness, a process called antibody-dependent cellular cytotoxicity (ADCC). IL-12 and gamma interferon are potent activators of NK cells. From 5 to 10% of peripheral lymphocytes are NK cells.

 

NK cells are lymphocytes with some T cell markers, but they do not have to pass through the thymus in order to mature. They have no immunologic memory and, unlike cytotoxic T cells, have no T cell receptor; also, killing does not require-recognition of MHC proteins. In fact, NK cells have re­ceptors that detect the presence of class I MHC proteins on the cell surface. If a cell displays sufficient class I MHC proteins, that cell is not killed by the NK cell.

Many virus-infected cells and tu­mor cells display a significantly reduced amount of class I MHC proteins, and it is those cells that are recognized and killed by the NK cells.   

Summary of the Non-Specific Immune Response:

 

 

Antigens. The main attributes of antigenes. Antigens structure of bacterial cell and viruses. Immunoglobulins. Structure and classes. Methods of obtaine bacterial antigenes. examination of immunoglobulins concentration in blood serum. Major histocompatibility complex.

THE STRUCTURE OF THE IMMUNE SYSTEM (Т – AND B – SYSTEM).  COOPERATION OF THE CELLS  IN IMMUNE RESPONSES. MECHANISM  OF ANTIBACTERIAL AND ANTIVIRAL DEFENCE

 

The name antigens (Gk. anti against, genos genus) is given to organic substances of a colloid structure (proteins and different protein complexes in combination with lipids or polysaccharides) which upon injection into the body (subcutaneously, intracutaneously, cutaneously, into the mucous membranes, intramuscularly, intraperitoneally, intravenously, and orally) are capable of causing the production of antibodies and reacting specifically with them.

 

Antigens, consequently, are characterized by the following main properties: (1) the ability to cause the production of antibodies (antigenicity), and (2) the ability to enter into an interaction with the corresponding antibodies (antigenic specificity).

 

Antigenic substances are highly molecular compounds. They have certain properties: a specific action, heterogenicity for the body, a colloid structure, and solubility in the body fluids. The breakdown of proteins to peptones, amino acids and also a deep denaturation by physicochemical effects bring about a loss of antigenic ability, while the introduction of various radicals into the protein molecule causes the loss of species specificity. Substances composed of levorotatory amino acid isomers induce antibody production, while complexes of dextrorotatory amino acid isomers are devoid of antigenic functions.

 

 

Antigenic properties are pertinent to toxins of a plant origin (ricin, robin, abrin, cortin, etc.), toxins of an animal origin (toxins of snakes, spiders, scorpions, phalangia, karakurts, bees), enzymes, native foreign proteins, various cellular elements of tissues and organs, bacteria and their toxins, rickettsiae and viruses.

Not all substances (proteins and protein complexes in combination with lipids and polysaccharides) are characterized by having antigens with similar properties. There are complete and partial antigens.

Complete antigens are substances which cause the production of antibodies in the body, and react with them in vivo as well as in vitro (foreign proteins, sera, bacteria, toxins, rickettsiae, viruses and cellular elements).

 

 

                       

 

 

 

 

Partial antigens are known as haptens which do not cause the production of antibodies, but can react with them. Haptens include lipids, complex carbohydrates and other substances. The addition of proteins to haptens even in a small amount gives them the properties of complete antigens. In this case the protein carries out the function of a conductor.

It is well known that the properties of chemical, structural and functional speciflcity are inherent in all natural proteins. Proteins of different species of animals, plants, bacteria, rickettsiae and viruses can be differentiated by immunological reactions. The antigenic function of bacteria, rickettsiae and viruses is characterized not only by species, but also by type speciflcity.

The immunological speciflcity of antigens is linked with a determinant group found on the surface of the antigen as one or more active areas. The determinant group may be isolated in a relatively pure form, which makes it possible to improve the efficacy of vaccinal preparations significantly. Besides, inside each species of microbe there is a different amount of types which also have specific antigenic properties. Type specificity is associated with the presence of special polysaccharide com- plexes in the microbial cell. Besides species and type specificity group (generic) antigens have been revealed in closely related species. The presence of group antigens reflects the historical process of their development and genetic links.

When the antigenic structures of the host are similar to those of the causative agent, the macro-organism is incapable of producing immunity, as the result of which the disease follows a graver course. It is possible that in individual cases the carrier state and inefficacy of vaccination are due to the common character of the microbial antigens and the antigens of the person’s cells.

It has been established that human erythrocytes have antigens in common with staphylococci, streptococci, the organisms of plague, E. coli. Salmonella paratyphi, Shigella organisms, smallpox and influenza viruses, and other causative agents of infectious diseases. Such a condition is called antigenic mimicry.

In 1911 D. Forssman established that there are heterogenic or heterologic antigens (haptens) found in different species of animals (guinea pigs, dogs, cats, horses, chickens, fish and turtles). If a rabbit is immunised with an emulsion from the organs of guinea pigs, then antibodies appear in the serum of the rabbit which react not only with the emulsion of these organs but also with sheep erythrocytes. Thus, in the organs of the guinea pig and sheep erythrocytes there is a heterogenic antigen.

It has been proven that the non-specific properties of Forssman’s heterogenic antigen are associated with the presence of lipid or polysaccharide fractions closely related in composition, which bear  common properties in different species of animals, plants and microbes.

 

Isoantigens. Isoantigens are those substances which have antigenic properties and are contained in some individuals of a given species. They have been found in the erythrocytes of animals and man. At first it was established that in human erythrocytes there are two antigens (A and B), and in the sera — beta- and alpha-antibodies. Only heterogenic antigens and antibodies (agglutinins) can be found in human blood.

These combinations may be represented as follows:

 

 

Blood Group	Antigens on RBC	Antibodies (serum)	Can donate blood to	Can receive blood from
A	A	Anti-B	A and AB	A and O
B	B	Anti-A	B and AB	B and O
AB	A and B	None	AB	All groups
O	None	Anti-A and Anti-B	All groups	O

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

On the basis of antigenic structure the erythrocytes of all people can be subdivided into 4 groups. Subsequently, variants of antigens of erythrocytes in the second (A) and fourth (AB) group were isolated. The A group consists of two subgroups — A₁ and A₂ The AB group contains antigens A₁B and A₂B, and the antigens M and N, M₂, and N₂ etc., have been revealed. More than 15 systems of isoantigens including about 100 antigens are now known. Besides, the erythrocytes contain the rhesus factor (see ‘Blocking Antibodies’). Isoantigens of leukocytes, blood platelets, lymphocytes, granulocytes, blood sera, liver, and kidneys and inter-organ (cell nuclei, mitochondria, ribosomes, etc.) and pathological (cancerous, bum, radiation) isoantigens have been revealed. These data are taken into account during blood transfusion.

Autoantigens are substances capable of immunizing the body from which they are obtained. Thus, they become modified and are capable of bearing an antigenic function. These substances include the eye lens, spermatozoids, homogenates of the seminal gland, skin, emulsions of kidneys, liver, lungs and other tissues. Under ordinary conditions they do not come in contact with the immunizing systems of the body, therefore antibodies are not produced against such cells and tissues. However, if these tissues are injured, then autoantigens may be absorbed, and may cause the production of antibodies which have a toxic effect on the corresponding cells. The origination of autoantigens is possible under the influence of cooling, radiation, drugs (amidopyrine, sulphonamides, preparations of gold, etc.), virus infections “(virus pneumonias and mononucleosis), bacterial proteins and toxins of streptococci, staphylococci, tubercle bacilli, paraproteins, aseptic autolysis of brain tissue, and other factors. Probably, only haptens appear, which combine with proteins and produce complete antigens capable of causing the production of antibodies.

The production of autoantigens is the result of the disturbance of species specificity which provides for the antigenicity of a number of substances found in the given body. In relation to the reasons for the appearance of autoantigens, there are the following hypotheses: (1) endogenic substances become antigens only after preliminary changes; (2) endogenic substances can really be antigens, and (3) only certain endogenic substances which come from some tissues can be antigens.

Antigenic properties of bacteria, toxins, rickettsiae and viruses, used in the practice of reproducing artificial immunity against infectious diseases, are of most practical importance.

 

 

Major Histocompatibility Complex 

 

The success of tissue and organ transplants depends on the donor’s and recipient’s human leuko­cyte antigens (HLA) encoded by the HLA genes. These proteins are alloantigens; ic, they differ among members of the same species, If the HLA proteins on the donor’s cells differ from those on the recipient’s cells, an immune response occurs in the recipient. The genes for the HLA proteins are clustered in the major histocompatibility complex (MHC), located on the short arm of chromosome 6. Three of these genes (HLA-A, HLA-B, and HLA-C) code for the class I MHC proteins. Several HLA-D loci determine the class II MHC proteins, ie, DP, DQ, and DR (Figure 1).

Each person has two haplotypes, ie, two sets of these genes, one on the paternal and the other on the maternal chromosome 6. These genes are very diverse (polymorphic) (ie, there are many alleles of the class I and class II genes). For example, there are at least 47 HLA-A genes, 88 HLA-B genes, 29 HLA-C genes, and more than 300 HLA-D genes, but any individual inherits only a single allele  at each locus from each parent and thus can make no more than two class I and tl proteins at each gene locus. Expression of these genes is codominant; ie, the proteins encoded by both the paternal and maternal genes are produced. Each person can make as many as 12 HLA proteins: 3 at class I loci and 3 at class II loci, from both chromosomes.

 

In addition to the major antigens encoded by the HLA genes, there are an unknowumber of mi­nor antigens encoded by genes at sites other than the HLA locus. These minor antigens can induce a weak immune response that can result in slow rejection of a graft. The cumulative effect of several minor antigens can lead to a more rapid rejection response. There are no laboratory tests for minor antigens.

Between the class I and class II gene loci is a third locus (Figure 1), sometimes called class III. This locus contains several immunologically important genes, encoding two cytokines (tumor necrosis factor and lymphotoxin) and-two complement components (C2 and C4), but it does not have any genes that encode histocompatibility antigens.

 

MHC PROTEINS

Class I MHC Proteins.    These are glycoproteins found on the surface of virtually all nude ated cells. There are approximately 20 different proteins encoded by the allelic genes at the A locus, 40 at the B locus, and 8 at the C locus.

 

The complete class I protein is composed of a 45,000-moleculjar-weight heavy chaioncovalently bound to a β₂-microglobulin. The heavy chain is highly  polymorphic and is similar to an immunoglobulin molecule; it has hypervariable regions in its N-terminal region. The polymorphism of these molecules is important in the recognition of self and nonself. Stated another way, if these molecules were more similar, our ability to accept foreign grafts would be correspondingly improved. The heavy chain also-has a constant region where the CDS protein of the cytotoxicT cell binds.

 

 

 

Class II MHC Proteins. These are glycoproteins found on the surface of certain cells, includ­ing macrophages, B cells, dendritic cells of the spleen, and Langerhans cells of the skin. They are highly polymorphic glycoproteins composed of two polypeptides (MW 33,000 and 28,000) that are noncovalently bound. Like class I proteins, they have hypervariable regions that provide much of the polymorphism. Unlike class I proteins, which have only one chain encoded by the MHC locus (β₂-microglobulin is encoded on chromosome 15), both chains pf the class II proteins are encoded by the MHC locus. The two peptides also have a constant region where the CD4 proteins of the helper T cells bind.

 

BIOLOGIC   IMPORTANCE OF MHC

The ability of T cells to recognize antigen is dependent on association of the antigen with either class 1 or class II proteins. For example, cytotoxic T cells respond to antigen in association with class 1 MHC proteins. Thus, a cytotoxic Tcell that kills a virus-infected cell will not kill a cell in­fected with the same virus if the cell does not also express the appropriate class 1 proteins. This find­ing was determined, using inbred animals, by mixing virus-infected cells and cytotoxic T cells bearing.different class I proteins and observing that no killing of the virus-infected cells occurred. Helper T cells recognize class II proteins. Helper-cell activity depends in general on both the recognition of the antigen on antigen-presenting cells and the presence on these cells of self class II MHC pro­teins. This requirement to recognize antigen in association with a self MHC protein is called MHC restriction. Note that T cells recognize antigens only when the antigens are presented on the surface of cells (in association with either class I or II MHC proteins), whereas B cells do not have that requirement and can recognize soluble antigens in plasma with their surface monomer IgM act­ing as the antigen receptor.

MHC genes and proteins are also important in two other medical contexts. One is that many autoimmune diseases occur in people who carry certain MHC genes, and the other is that the success of organ transplants is, in large part, determined by the compatibility of the MHC genes of the donor and recipient (see below).

 

Expression of MHC molecules

MHC class I molecules are widely expressed, though the level varies between different cell types. MHC class II molecules are constitutively expressed only by certain cells involved in immune responses, though they can be induced on a wider variety of cells.

 

 

Antibodies (Immunoglobulins)

Antibodies are specific substances produced in the bodies of vertebrates upon the introduction of antigens; they are capable of a specific association with them. As a result of an obvious or latent infectious disease antibodies appear in the sera of animals and man. Antibodies appear not only as a result of an infection, but also due to immunization by live (attenuated) or dead bacteria, rickettsiae, viruses, toxins and other antigenic substances.

 

Antibodies which occur under the influence of active immunization are named immune antibodies in contrast to normal antibodies which are often found in the sera of man and animals who have not had infectious diseases and who have not been exposed to immunization. It is believed that they form under the effect of normal microflora during the development of the organism.

Any normal serum contains a small amount of cryogglutinins (cold haemagglutinins) which react with the erythrocytes of the same person. When cryogglutinins appear in a large number, paroxysmal haemoglobinuria develops. Normal as well as immune antibodies are capable of rendering harmless the causative agents of infectious diseases in various ways.

Young poisonous snakes of some species are devoid of antibodies, as a consequence of which they are sensitive to their own poison; adult snakes have antibodies which neutralize their own poison.

Immunoglobulins are synthesized by immunocytes which form from B-lymphocytes. The sources of immunocytes are the stem cells of the haematopoietic tissue, which acquire immunologic activity due to the effect of the thymus. Antigens come into contact with the cells continuously migrating from the thymus (T-cells) and bone marrow (B-cells) into the lymph nodes and spleen where with the participation of the macrophages clonal immunocytes are formed. The immune process is therefore realized as the result of cooperation of three groups of cells: T- and B-lymphocytes and macrophages which phagocytose and digest  the antigens and transmit the information concerning the antigen to the corresponding immunocom petent cells producing immunoglobulins.

 

IMMUNOGLOBUL1N STRUCTURE

Immunoglobulins are glycoproteins made up of light (L) and heavy (H) polypeptide chains. The terms “light” and heavy” refer to molecular weight; light chains have a molecular weight of about 25,000, whereas heavy chains have a molecular weight of 50,000-70,000. The simplest antibody molecule has a Y shape (Figure 2) and consists of four polypeptide chains: two H chains and two L chains. The four chains are linked by disulfide bonds. An individual antibody molecule always consists of identical H chains and identical L chains. This is primarily the result of two phenomena: allelic exclusion  and regulation within the B cell, which ensure the synthesis of either kappa (κ) or lambda (λ) L chains but not both.

 

L and H chains are subdivided into variable and constant regions. The regions are composed of three-dimensionally folded, repeating segments called domains. An L chain consists of one variable (VL) and one constant (CL) domain. Most H chains consist of one variable (VH) and three constant (CH) domains. (IgG and IgA have three CH domains, whereas IgM and IgE have four.) Each do­main is approximately 110 amino acids long. The variable regions are responsible for antigen-binding, whereas the constant regions are responsible for various biologic functions, eg, comple­ment     activation and binding to cell surface receptors. Carboxy terminal  end.

                                                                     

 

 

 

Figure 2. Structure of IgG The Y-shaped IgG molecule consists of two tight chains-and-two heavy chains. Each light chain consists of a variable region and a constant region. Each heavy chain consists of a variable region and a constant region that is divided into three domains: CH1, CH2, and CH3. The CH2 domain contains the complement-binding site, and the CH3 domain is the site of attachment of IgG to receptors oeutrophils and macrophages. The antigen-binding site is formed by the variable regions of both the light and heavy chains. The specificity of the antigen-binding site is a function of the amino acid sequence of the hypervariable regions.

 

The variable regions of both L and H chains have three extremely variable (“hypervariable”) amino acid sequences at the amino-terminal end that form the antigen-binding site. Only 5-10 amino acids in each hypervariable region form the antigen-binding site. Antigen-antibody binding involves electrostatic and van der Waals’ forces and hydrogen and hydrophobic bonds rather than covalent bonds. The remarkable specificity of antibodies is due to these hypervariable regions.

L chains belong to one of two types, k (kappa) or λ  (lambda), on the basis of amino acid differ­ences in their constant regions. Both types occur in all classes of immunoglobulins (IgG, IgM, etc), but any one immunoglobulin molecule contains only one type of L chain. The ammo-tion of each L chain participates in the antigen-binding site. H chains are distinct for each of the five immunoglobulin classes and are designated y, a, ji, e, and 8 (Table 1). The amino-terminal por­tion of each H chain participates in the antigen-binding site; die carboxy terminal forms the Fc frag­ment, which has the biologic activities described above and in Table 1.

If an antibody molecule is treated with a proteolytic enzyme such as papain, peptide bonds in the “hinge” region are broken, producing two identical Fab fragments, which carry the antigen-binding sites, and one Fc fragment, which is involved in placenta! transfer, complement fixation, attachment site for various cells, and other biologic activities (Figure 2).

 

Properties of immunoglobulins

 

IMMUNOGLOBULIN CLASSES

 

 

Each IgG molecule consists of two L chains and two H chains linked by disulfide bonds (molecular formula H2L2). Because it has two identical antigen-binding sites, it is said to be diva­lent. There are four subclasses, IgGl-IgG4, based on antigenic differences in the H chains and on the number and location of disulfide bonds. IgGl makes up most (65%) of the total IgG. IgG2 anti­body is directed against polysaccharide antigens and is an important host defense against encapsu­lated bacteria.

 

IgG is the predominant antibody in the secondary-response and constitutes an important defense against bacteria and viruses (Table 2). IgG is the only antibody to cross the placenta only its Fc portion binds to receptors on the surface of placenta! cells. It is therefore the most abundant un-munoglobulin iewborn. IgG is one of the two immunoglobulins that can activate complement; IgM is the other.

IgG is the immunoglobulin that opsonizes. It can opsonize, ie, enhance phagocytosis, because there are receptors for the y H chain on the surface of phagocytes. IgM does not opsonize directly, because 4here are no receptors on the phagocyte surface for the ji H chain. However, IgM activates complement, and the resulting C3b can opsonize because there are binding sites for C3b on the sur­face of phagocytes.

IgA. IgA is the main immunoglobulin in secretions such as colostrum, saliva, tears, and respira­tory, intestinal, and genital tract secretions. It prevents attachment of microorganiimi, eg, bacteria and viruses, to mucous membranes. Each secretory IgA molecule consists of two H2L2 units plus one molecule each of J (joining) chain and secretory component (Figure 3). The secretory com­ponent is a polypeptide synthesized by epithelial cells that provides for IgA passage to the mucosal surface. It also prelects IgA from being degraded in the intestinal tract. In serum, some IgA exists as monomeric H2L2.

 

 

 

 

 

 

 

 

 

 

 

IgM IgM is the main immunoglobulin produced early in the primary response. It is present as a monomer on the surface of virtually all B cells, where it functions as an antigen-binding receptor In serum, it is a pentamer composed of 5 H2L2 units plus one molecule of J (joining) chain (Figure 3). Because the pentamer has 10 antigen-binding sites, it is the most efficient immunoglobulin in agglutination, complement fixation (activation), and other antibody reactions and is important in de­fense against bacteria and viruses. It can be produced by the fetus in certain infections. It has the highest avidity of the immunoglobulins; its interaction with antigen can involve all 10 of its binding sites.

IgD This immunoglpbulin has no known antibody function but may function as an antigen recep­tor; it is present on the surface of many B lymphocytes. It is present in small amounts in serum.

IgE. IgE is medically important for two reasons: (1) it mediates immediate (anaphylactic) hypersensitivity, and (2) it participates in host defenses against certain parasites, eg, helminths (worms). The Fc region of IgE binds to the surface of mast cells and basophils. Bound IgE serves as a receptor for antigen (allergen), and this antigen-antibody complex triggers allergic responses of the immediate (anaphylactic) type through the release of mediators. Although IgE is present in trace amounts iormal serum (approximately 0.004%), persons with allergic reactivity have greatly increased amounts, and IgE may appear in ex­ternal secretions. IgE does not fix complement and does not cross the placenta.

IgE is the main host defense against certain important helminth (worm)infections, such as Strongyloides, Trichinella, Ascaris, and the hookworms. The serum IgE level is usually increased in these infections. Because worms are too large to be ingested by phagocytes, they are killed by eosinophils that release worm-destroying enzymes. IgE specific for worm proteins binds to receptors on eosinophils, triggering the antibody-dependent cellular cytotoxicity (ADCC) response.

 

 

 

 

 

ORIGIN OF IMMUNE CELLS

The capability of responding to immunologic stimuli rests mainly with lymphoid cells. During em­bryonic development, blood cell precursors originate mainly in the fetal liver and yolk sac; in post­natal life, the stem cells reside in the bone marrow. Stem cells differentiate into cells of the erythroid, myeloid, or lymphoid series. The latter evolve into two main lymphocyte populations: T cells and B cells (Figure 1 and Table 1). The ratio of T cells to B cells is approximately 3:1.

T cell precursors differentiate into immunocompetent T cells within the thymus. Stem cells lack antigen receptors and CD3, CD4, and CD8 molecules on their surface, but during passage through the thymus they differentiate into T cells that can express these glycoproteins. The stem cells, which initially express neither CD4 nor CD8 (double-negatives), first differentiate to express both CD4 and CD8 (double-positives) and then proceed to express either CD4 or CD8. A double-positive cell will differentiate into a CD4-positive cell if it contacts a cell bearing class II  MHC proteins but will differentiate into a CD8-positive cell if it contacts a cell bearing class I MHC proteins. (Mutant mice that do not make class II MHC proteins will not make CD4-positive cells, indicating that this inter­action is required for differentiation into single-positive cells to occur.) The double-negative cells and the double-positive cells are located in the cortex of the thymus,. whereas the single-posith cells are located in the medulla, from which they migrate outside of the thymus. Within the (hymns, two very important processes called thymic education occur.

 (1) CD-4-positive, CD8-positive cells, bearing antigen receptors for “self proteins, are killed (clonal deletion) by a process of “programmed cell death” called apoptosis (Figure 2). The removal of these self-reactive cells, a process called negative selection, results in tolerance to own proteins, ie, self-tolerance, and prevents autoimmune reactions.

(2) CD4-positiva, CDS-positive cells, bearing antigen receptors, that do not react with self MHC proteins are also killed. This results in a positive selection for T cells that react well with self MHC proteins.

These two processes produce T cells that are selected for their ability to react both with foreign antigens via their antigen receptors and with self MHC proteins. Both of these features are requir for an effective immune response by T cells.

Note that MHC proteins perform two essential functions in the immune response; one is the tive selection of T cells in the thymus, as just mentioned, and the other, which is described below, is the presentation of antigens to T cells, the initial step required to activate those cells. MHC teins are also the most important antigens recognized in the graft rejection process.

 

 

 

Table 1. Comparison of T cells and B cells

 

 

During their passage through the thymus, each double-positive T cell synthesizes a different, j highly specific antigen receptor called the T cell receptor (TCR). The rearrangement of the vari-i able, diversity, and joining genes that encode the receptor occurs early in T cell differentiation and accounts for the remarkable ability of T cells to recognize millions of different antigens.

Some T lymphocytes, perhaps as much as 40% of the total, do not develop in the thymus butJ rather in the “gut-associated lymphoid tissue” (GALT). These intraepithelial lymphocytes (lELs) are^! thought to provide protection against intestinal pathogens. Their antigen receptors and surface pro­teins are different from those of thymes-derived lymphocytes. lELs cannot substitute for thymus-derivcd lymphocytes because patients with DiGeorgc’s syndrome who lack a thymus (arc profoundly immunodeficient and have multiple infections.

The thymus involutes in adults, yet T cells continue to be made. Two explanations have been of­fered for this apparent paradox. One is that a remnant of the thymus remains functional throughout life and the other is that an extrathymic site takes over for the involuted thymus. Individuals who have had their thymus removed still make T cells, which supports the latter explanation.

 

Bone marrow stem сell

B cell

Bursa equivalent (bone marrow in humans)

B cell precursors differentiate into immunocompetent B cells in the bone marrow; they do not pass through the thymus. B cells also undergo clonal deletion of those cells bearing antigen recep­tors for self proteins, a process that induces tolerance and reduces the occurrence of autoimmune diseases.The site of clonal deletion of B cells is uncertain, but it is not the thymus.

Natural killer (NK) cells are large granular lymphocytes that do not pass through the thymus, do  not have an antigen receptor, and do not bear CD4 or CD8 proteins. They recognize and kill target cells, such as virus-infected cells and tumor cells, without the requirement that the antigens be pre­sented in association with class I or class II MHC proteins. Rather, NK cells target those cells to be killed by detecting that they do not display class I MHC proteins on the cell surface. For example, many cells after they have been infected by a virus lose their ability to synthesize class I MHC pro­teins.

In contrast to T cells, B cells, and NK cells, which differentiate from lymphoid stem cells, macrophages arise from myeloid precursors. Macrophages have two important functions, namely, phagocytosis and antigen presentation. They do not pass through the thymus and do not have an antigen receptor. On their surface, they display class II MHC proteins, which play an essential role in antigen presentation to helper T cells. The cell surface proteins that play an important role in the immune response are listed in Table 2.

T Cells. T cells perform several important functions, which can be divided into two main categories, namely, regulatory and effector. The regulatory functions are mediated primarily by helper (CD4-positive) T cells, which produce interleukins (Table 3). For example, helper T cells make (1) interleukin-4 (IL-4) and IL-5, which help B cells produce antibodies; (2) IL-2, which activates CD4 and CD8 cells; and (3) gamma interferon, which activates macrophages,  the main mediators of delayed hyper-sensitivity against intracellular organisms such as Mycobacterium tuberculosis. (Suppressor T cells are postulated to down-regulate the immune response, but evidence to support the existence of these cells is lacking.) The effector functions are carried out primarily by cytotoxic (CD8-positive) T cells, which kill virus-infected cells, tumor cells, and allografts.

CD4 and CD8 Types of T Cells.

 

 

 

Within the thymus, perhaps within the outer cortical epithelial cells (nurse cells), T cell progenitors differentiate under the influence of thymic hormones (thymosins and thymopoietins) into T cell subpopulations. These cells are characterized by certain sur­face glycoproteins, eg, CD3, CD4, and CD8. All T cells have CD3 proteins on their surface in asso­ciation with antigen receptors (T cell receptor [see below]). The CD3 complex of five transmembrane proteins is involved with transmitting, from the outside of the cell to the inside, the information that the antigen receptor is occupied. One of the CD3 transmembrane proteins, the  zeta chain, is linked to a tyrosine kinase called fyn, which is involved with signal transduction. The signal is transmitted via several second messengers, which are described in the section on activation (see below). CD4 is a single transmembrane polypeptide whereas CD8 consists of two transmem­brane polypeptides. They may signal via tyrosine kinase (the Ick kinase) also.

Table 2. Cell surface proteins that play an important role in the immune response.¹

 

There are many other cell surface proteins that play a role in the im­mune response, but the proteins listed in this table are the most im­portant for understanding the fundamental aspects of this response; ²TCR, T cell antigen receptor;  ³Macrophages and other “antigen-presenting cells.”

 

T cells are subdivided into two major categories on the basis of whether they have CD4 or CD8 proteins on their surface. Mature T cells have either CD4 or CD8 proteins but not both.

CD4 lymphocytes perform the following helper functions: (1) they help B cells develop into an­tibody-producing plasma cells; (2) they help CD8 T cells to become activated cytotoxic T cells; and (3) they help macrophages effect delayed hypersensitivity (eg, limit infection by M tuberculosis). These functions are performed by 2 subpopulations of CD4 cells: Th-1 cells help activate cytotoxic T cells by producing IL-2 and help initiate the delayed hypersensitivity response by producing pri­marily IL-2 and gamma interferon, whereas Th-2 cells perform the B cell helper function by produc­ing primarily IL-4 and IL-5 (Figure 3). One important regulator of the balance between Th-l cells and Th-2 cells is interleukin-12 (IL-12), which is produced by macrophages. IL-12 increases the number of Th-1 cells, thereby enhancing host defenses against organisms that are controlled by a delayed hypersensitivity response (Table 4). Another important regulator is gamma interferon which inhibits the production of Th-2 cells. CD4 cells make up about 65% of peripheral T cells and predominate in the thymic medulla, tonsils, and blood.

 To mount a protective immune response against a specific microbe requires that the appropriate subpopulation, ie, either Th-1 or Th-2 cells, play a dominant role in the response. For example, if an individual is infected with M tuberculosis and Th-2 cells are the major responders, then humoral im­munity will be stimulated rather than cell-mediated immunity.

 

Main functions of helper T cells

 Table 3

 

Humoral immunity is not protective against M tuberculosis and the patient will suffer severe tuberculosis. Similarly, if an individual is infected with Streptococcus pneumoniae and Th-1 cells are the major responders, then humoral im­munity will be not be stimulated and the patient will have severe pneumococcal disease. Precisely what component of a microbe activates either Th-1 or Th-2 cells is unknown.

 

   Comparison of Th-1 cells and Th-2 cells    

                 Table 4.

 

CD8 lymphocytes perform cytotoxic functions; that is, they kill virus-infected, tumor, and allograft cells. They kill by either of two mechanisms, namely, the release of performs, which destroy cell membranes, or the induction of programmed cell death (apoptosis). CD8 cells predominate in human bone marrow and gut lymphoid tissue and constitute about 35% of peripheral T cells,

 

Activation of T Cells. The activation of helper T cells requires that they recognize a complex on the surface of antigen-presenting cells (APCs), eg, macrophages consisting of both the antigen and a class II MHC protein               (Macrophages are the most important antigen-presenting cells, but B cells, dendritic cells in the spleen, and Langer-hans cells on the skin also present antigen, ic, have class II proteins on their surface: An essential first step for certain antigoH-prosnilinj; cells, eg. Umgcrhans cells ui the skin, is migration from the site of the skin infection to ihe local lymphoid tissue, where helper T cells are encountered). Within the cytoplasm of the macrophage, the foreign protein is cleaved into small peptides that associate with the class II MHC proteins. The complex is trans­ported to the surface of the macrophage, where the antigen, in association with a class II MHC pro­tein, is presented to the receptor on the CD4-positive helper cell. Similar events occur within a virus-infected cell, except that the cleaved viral peptide associates with a class I rather than a class II MHC protein. The complex is transported to the surface, where the viral antigen is presented to the receptor on a CD8-positive cytotoxic cell. Remember the rule, of eight: CD4 cells interact with class II (4 x 2 = 8), and CD8 cells interact with class I (8 x 1 = 8).

There are many different alleles within the class I and class II MHC genes; hence, there are many different MHC proteins. These various MHC proteins bind to different peptide fragnients. The poly­morphism of the MHC genes and the proteins they encode are a means of presenting many different antigens to the T cell receptor. Note that class I and class IIMHC proteins can only present peptides; other types of molecules do not bind and therefore cannot be presented. MHC proteins can present peptides derived from self proteins as well as from foreign proteins; therefore whether an immune response occurs is determined by whether a T cell bearing a receptor specific for that peptide has survived the positive, and negative selection processes in the thymus.

The first step in the activation process is the interaction of the antigen with the T cell receptor specific for that antigen (Figure 4). IL-1 produced by the macrophage is also necessary for efficient helper T cell activation. Note that when the T cell receptor interacts with the antigen-MHC protein complex, the CD4 protein on the surface of the helper T cell also interacts with the class II MHC protein. In addition to the binding of the CD4 protein with the MHC class II protein, other proteins interact to help stabilize the contact between the T cell and the APC; eg, LFA-1 protein on T cells (both CD4-positive and CDS-positive) binds to ICAM-1 protein on APCs (LFA proteins belong to a family of cell surface proteins called integrins, which mediate adhesion to other cells. Inte-grin proteins are embedded in the surface membrane and have both extracellular and intracellular domains. Hence they interact with other cells externally and with the cytoplasm internallyAbbreviations: LFA, lymphocyte function-asso­ciated antigen; 1CAM, intercellular adhesion molecule).

For full activation of helper T cells, an additional “costimulatory” signal is required; that is, B7 protein on the APC must interact with CD28 protein on the helper T cell (Figure 4). If the co­stimulatory signal occurs, IL-2 is made by the helper T cell, and it is this step that is crucial to pro­ducing a helper T cell capable of performing its regulatory, effector, and memory functions. If, on the other hand, the T cell receptor interacts with its antigen (epitope) and the costimulatory signal does not occur, a state of unresponsiveness called anergy ensues. The anergic state is specific for that epitope, since other helper T cells specific for other epitopes are not affected. After the T cell has been activated, a different protein called CTLA-4 appears on the T cell surface and binds to B7 by displacing CD-28.

The interaction of B7 with CTLA-4 inhibits T cell ac­tivation by blocking IL-2 synthesis. This restores the activated T cell to a quiescent state and thereby plays an important role in T cell homeostasis. Mutant T cells that lack CTLA-4 and there­fore cannot be deactivated participate with increased frequency in autoimmune reactions. Further­more, administration of CTLA-4 reduced the rejection of organ transplants in experimental animals.

T cells recognize only polypeptide antigens. Furthermore, they recognize those polypeptides only when they are presented in association with MHC proteins. Helper T cells recognize antigen in asso­ciation with class II MHC proteins, whereas cytotoxic T cells recognize antigen in association with class 1 MHC proteins. This is called MHC restriction; ie, the two types of T cells (CD4 helper and CD8 cytotoxic) are “restricted” because they are able to recognize antigen only when the antigen is presented with the proper class of MHC protein. This restriction is mediated by specific binding sites primarily on the T cell receptor, but also on the CD4 and CD8 proteins that bind to specific regions on the class II and class I MHC proteins, respectively.

Generally speaking, class I MHC proteins present endogenously synthesized antigens, eg, viral proteins, whereas class II MHC proteins present the antigens of extracellular microorganisms that have been phagocyti/cd, eg, bacterial proteins. One important consequence of these observations is that killed viral vaccines do not activate the cytotoxic (CD8-positive) T cells, because the virus does not replicate within cells and therefore viral epitopes are not presented in association with class I MHC proteins.

This distinction between endogenously synthesized and.extracellularly acquired proteins is achieved by processing the proteins in different compartments within the cytoplasm. The endoge­nously synthesized proteins, eg, viral proteins, are cleaved by a proteasome, and the peptide frag­ments associate with a ‘TAP transporter^” that transports the fragment into the rough endoplasmic reticulum, where it associates with the class I MHC protein. The complex of peptide fragment and class I MHC protein then migrates via the Golgi apparatus to the cell surface. In contrast, the extracellularly acquired proteins are cleaved to peptide fragments within an endosome, where the fragment associates with class II MHC proteins. This complex then migrates to the cell surface.

 

An additional protection that prevents endogenously synthesized proteins from associating with class II MHC proteins is the presence of an “invariant chain” that is attached to the class II MHC proteins when these proteins are outside of the endosome. The invariant chain is degraded by proteases within the endosome, allowing the peptide fragment to attach to the class II MHC proteins only within that compartment.

B cells, on the other hand, can interact directly with antigens via their surface immunoglobulins (IgM and IgD). Antigens do not have to be presented to B cells in association with class II MHC proteins. unlike T cells. Note that B cells can then present the antigen, after internalization and pro­cessing, to helper T cells in association with class II MHC proteins located on the surface of the B cells (see the section on B cells, below). Unlike the antigen receptor on T cells, which recognizes only peptides, the antigen receptors on B cells (IgM and IgD) recognize many different types of molecules, such as peptides, polysaccharides, nucleic acids, and small chemicals, eg, penicillin.

These differences between T cells and B cells explain the hapten-carrier relationship described. To stimulate hapten-specific antibody, the hapten must be covalently bound to the car­rier protein. A B cell specific for the hapten internalizes the hapten-carrier conjugate, processes the carrier protein, and presents a peptide to a helper T cell bearing a receptor for that peptide. The helper T cell then secretes lymphokines that activate the B cell to produce antibodies to the hapten.

When the antigen-MHC protein complex on the APC interacts with the T cell receptor, a signal is transmitted by the CD3 protein complex through several pathways that eventually lead to a large in­flux of calcium into the cell. (The details of the signal transduction pathway are beyond the scope of this book, but it is known that stimulation of the T cell receptor activates a series of phosphokinases which then activate phospholipase C, which cleaves phosphoinositide to produce inositol triphosphate, which opens the calcium channels.) Calcium activates calcineurin, a serine phosphatase. Calcineurin moves to the nucleus and is involved in the activation of the genes for IL-2 and the IL-2 re­ceptor. (Calcineurin function is blocked by cyclosporine, one of the most effective drugs used to prevent rejection of organ transplants.)

The end result of this series of events is the activation of the helper T cell to produce various lym­phokines, eg, IL-2, as well as the IL-2 receptor. IL-2, also known as T cell growth factor, stimu­lates the helper T cell to multiply into a clone of antigen-specific helper T cells. Most cells of this clone perform effector and regulatory functions, but some become “memory” cells (see below), which are capable of  being rapidly activated upon exposure to antigen at a later time. (Cytotoxic T cells and B cells also form memory cells.) Note that IL-2 stimulates CD8 cytotoxic T cells as well as CD4 helper T cells. Activated CD4-positive T cells also produce another lymphokine called gamma interferon, which increases the expression of class II MHC proteins on APCs. This enhances the ability of APCs to present antigen to T cells and upregulates the immune response.

The process of activating T cells does not function as a simply “on-off’” switch.The binding

 of an epitope to the T cell receptor can result in full activation, partial activation in” which only certain lymphokines are made, or no activation, depending on which of the signal transduction pathways is   stimulated by that particular epitope. This important observation may have profound implications .for our understanding of how helper T cells shape our response to infectious agents.

There are three genes at the class I locus (A, B, and C) and three genes at the class II locus (DP, DQ, and DR). We inherit one set of class I and one set of class II genes from each parent. Therefore, our cells can express as many as six different class I and six different class II proteins. Furthermore, there are multiple alleles at each gene locus. Each of these MHC proteins can pre­sent peptides with a different amino acid sequence. This explains, in part, our ability to respond to many different antigens.

Memory T Cells. Memory T (and B) cells, as the name implies, endow our host defenses with the ability to respond rapidly and vigorously for many years after the initial exposure to a microbe or other foreign material. This memory response to a specific antigen is due to several features: (1) many memory cells are produced, so that the secondary response is greater than the primary re­sponse, in which very few cells respond; (2) memory cells live for many years or have the capacity to reproduce themselves; (3) memory cells are activated by smaller amounts of antigen and require less costimulation than do naive, unactivated T cells; and (4) activated memory cells produce greater amounts of interleukins than do naive T cells when they are first activated.

T Cell Receptor. The T cell receptor (TCR) for antigen consists of two polypeptides, alpha and beta  ( Some TCRs have a different set of polypeptides called gamma and delta. Some of the T cells bearing these TCRs are involved in cell-mediated immunity against M tuberculosis),which are associated with CD3 proteins. TCR proteins are similar to immunoglobulin heavy chains in that (1) the genes that code for them are formed by rearrangement 6f multiple regions of  DNA; (2) there are V (variable), D (diversity), J (joining), and C (constant) seg­ments that rearrange to provide diversity, giving rise to an estimated number of more than 10⁷ differ­ent receptor proteins; and (3) the two genes (RAG-1 and RAG-2) that encode die recombinase en­zymes that catalyze these gene rearrangements are similar in T cells and 8 cells.

Note that each T cell has a unique T cell receptor on its surface, which means that hundreds of millions of different T cells exist in each person. Activated T cells, like activated B cells, clonally expand to yield large numbers of cells specific for that antigen.

Although TCRs and immunoglobulins are analogous in that they both interact with antigen in a highly specific manner, the T cell receptor is different in two important ways: (1) it has two chains rather than four, and (2) it recognizes antigen only, in conjunction with MMC proteins, whereas im­munoglobulins recognize free antigen.

 

 

 

 

Effect Of Superantigens on T Cells. Certain proteins, particularly staphylococcal enterotoxins and toxic shock syndrome toxin, act as “superantigens’¹ (Figure 5). In contrast to the usual antigen, which activates one (or a few) helper T cell, superantigens activate a large number of helper T cells. For example, toxic shock syndrome toxin binds directly to class II MHC proteins without in­ternal processing of the toxin. This complex interacts with the variable portion of the beta chain (Vβ) of the T cell receptor of many T cells . This activates the T cells, causing the release of IL-2 from the T cells and IL-1 from macrophages. These interleukins account for many of the findings seen in toxin-mediated staphylococcal diseases. Certain viral  proteins, eg, those of mouse mammary tumor virus (a retrovirus), also possess superantigen activity.

 

 

Figure 5.   Activation of helper T cells by superantigen. The helper T cell is activated by the presentation of processed antigen in assotiation with class II MHC protein to the antigen-soecific portion of the T cell receptor. Note that superantigen is not involved and that only one or a smoll number of helper T cells specific for the antigen are activated. 

 

Features of T Cells.    T cells constitute 65-80% of the recirculating poof of small lymphocytes.  Within lymph nodes, they are located in the inner, subcortical region, not in the germinal centers. (B cells iruike up most of the remainder of the pool of small lymphocytes and are found primarily in the germinal centers of lymph nodes.) The life span of T cells is long: months or years. They can be stimulated to divide when exposed to certain mitogens, eg/phytohemagglutinin or concanavalin A (endotoxin, a lipopolysaccharide found on the surface of gram-negative bacteria, is a mitogen for B cells but not T cells). Most human T cells have receptors for sheep erythrocytes on their surface«and can form “rosettes” with them; this finding serves as a means of identifying T cells in a mixed popu­lation of cells.

Effector Functions of T Cells. There are two important components of host defenses medi­ated by T cells: delayed hypersensitivity and cytotoxicity.

A.                      Delayed Hypersensitivity: Delayed hypersensitivity reactions are produced particularly against antigens of intracellular microorganisms including certain fungi, eg, Histoplasma and Coccidioides, and certain intracellular bacteria, eg, mycobacteria. Delayed hypersensitivity is medi­ated by macrophages and CD4 cells, in particular by the Th-1 subset of CD4 cells. Important lynv phokines for Ihese reactions fnclude gamma interferon, macrophage activation factor, and macrophage” migration inhibition factor. CD4 cells, produce the interleukins, and macrophages are the ultimate effectors of delayed hypersensitivity. A deficiency of cell-mediated immunity manifests itself as a marked susceptibility to infection by such microorganisms.

B.                      B. Cytotoxicity: The cytotoxic response is concerned primarily with destroying virus-infected cells and tumor cells but also plays an important role in graft rejection. In response to virus-infected cells, the CD8 lymphocytes must recognize both viral antigens and class I molecules on the surface of infected cells. To kill the virus-infected cell, the cytotoxic T cell must also receive a cy-tokine stimulus from a helper T cell. To become activated to produce these cytokincs, helper T cells rccogni/e viial anligcns bound to class II molecules on an APC, eg, a macrophage. The activated helper T cells secrete cytokines such as IL-2, which stimulates the virus-specific cytotoxic T cell to form a clone of activated cytotoxic T cells. These cytotoxic. T cells kill the virus-infected cells by in­serting “perforins” and degradative enzymes called granzymes into the infected cell. Performs form a channel through the membrane, the cell contents are lost, and the cell dies. Granzymes are proteases that degrade proteins in the cell membrane, which also leads to the loss of cell contents. They also trigger apoptosis, leading to cell death. After killing the virus-infected cell, the cytotoxic T cell itself is not damaged and can continue to kill other cells infected with the same virus. Cytotoxic T cells have no effect on free virus, only on virus-infected cells.

A third mechanism by which cytotoxic T cells kill target cells is the Fas-Fas ligand (FasL) inter­action. Fas is a protein displayed on the surface of many cells. When a cytotoxic T cell receptor rec­ognizes an epitope on the surface of a target cell, FasL is induced in the cytotoxic T cell. When Fas and FasL interact, apoptosis (death) of the target cell occurs. NK cells can also kill target cells by Fas-FasL-induced apoptosis.

        

 

 

 

 In addition to direct killing by cytotoxic T cells, virus-infected cells can be destroyed by a combi­nation of IgG and phagocytic cells. In this process, called antibody-dependent cellular cytotoxic­ity (ADCC), antibody bound to the surface of the infected cell is recognized by IgG receptors on the surface of phagocytic cells, eg, macrophages or NK cells, and the infected cell is killed. The ADCC process can also kill helminths (wormsjjk In this case, IgE is the antibody involved and eosinophils are the effector cells. IgE binds to surface proteins on the worm, and the surface of eosinophils dis­plays receptors for the epsilon heavy chain. The major basic protein located in the granules of the eosinophils is released and damages the surface of the worm.

 

 

Many tumor cells develop new antigens on their surface. These antigens bound to class I proteins are recognized by cytotoxic T cells, which are stimulated to proliferate by IL-2. The resultant clone of cytotoxic T cells can kill the tumor cells, a phenomenon called immune surveillance.

In response to allografts, cytotoxic (CD8) cells recognize the class I MHC molecules on the sur­face of the foreign cells. Helper (CD4) cells rccognize the foreign class II molecules on certain cells in the graft, eg, macrophages and lymphocytes. The activated helper cells secrete IL-2, which stimu­lates the cytotoxic cell to form a clone of cells. These cytotoxic cells kill the cells in the allograft.

Regulatory Functions of T Cells.    T cells play a central role in regulating both the humoral (antibody) and cell-mediated arms of the immune system.

A. Antibody Production: Antibody production by B cells usually requires the participation of helper T cells (T cell-dependent response), but antibodies to some antigens, eg, polymerized (mul­tivalent) macromolecules such as bacterial capsular polysaccharide, are T cell-independent. These polysaccharides are long chains consisting of repeated subunits of several sugars. The repeated subunits act as a multivalent antigen that cross-links the IgM antigen receptors on the B cell and acti­vates it in the absence of help from CD4 cells. Other macromolecules, such as DNA, RNA, and many lipids, also elicit a T cell-independent response.

In the following example illustrating the T cell-dependent response, B cells are used as the APC, although macrophages commonly perform this function. In this instance, antigen binds to surface IgM or IgD, is internalized within the B cell, and is fragmented. Some of the fragments return to-the surface in association with class II MHC molecules (Figure 6) (Note that one important difference between B cells and T cells is that B cells recognise antigen itself, whereas T cell’s recognize antigen onty in association with MHC proteins.)

  These interact with the recep­tor on the helper T cell, and, if the costimulatory signal is given by the B7 protein on the B cell inter­acting with CD28 protein on the helper T cell, the helper T cell is then stimulated to produce lymphokines, eg, IL-2, B cell growth factor (IL-4), and B cell differentiation factor (IL-8). IL-4 and IL-5 induce “class switching” from IgM, which is the first class of immunoglobulins produced, to other classes, namely, IgG, IgA, and IgE. These factors stimulate the B cell to divide and differentiate into many antibody-producing plasma cells.

Note that interleukins alone are not sufficient to activate B cells. A membrane protein on activated helper T cells, called CD40 ligand (CD40L), must interact with a protein called CD40 on the surface of the resting B cells to stimulate the differentiation of B cells into antibody-producing plasma-cells (Figure 6B). Furthermore, other proteins on the surface of these cells serve to strengthen the in­teraction between the helper T cell and the antigen-presenting B cell; eg, CD28 on the T cell inter­acts with B7 on the B cell and LFA-1 on the T cell interacts with ICAM-1 on the B cell. (There are also ICAM proteins on the T cell that interact with LFA proteins on the B cell.)

In the T cell-dependent response, all classes of antibody are made (IgG, IgM, IgA, etc), whereas in the T cell-independent response, primarily IgM is made. This indicates that lym-phokine’s produced by the helper T cell are needed for class switching. The T cell-dependent re­sponse generates memory B cells, whereas the T cell-dependent response does not; therefore, a secondary antibody response does not occur in the latter. The T cell-independent response is the main response to bacterial capsular polysaccharides, because these molecules are not effectively processed and presented by APCs and hence do not activate helper T cells. The most likely reason for this is that polysaccharides do not bind to class II MHC proteins whereas peptide antigens do.

 

Figure 6. A. B cell activation by helper T cells. B° is a resting B cell to which a multivalent antigen  is attach­ing to monomer IgM receptors (Y) The antigen is internalized, and a fragment ( a ) is returhed to the surface in con­junction with a class II molecule (n) A receptor on an activated T cell recognizes the complex on the B tell surface and the T cell produces B cell growth factor (BCGF, 1L-4,) and B cell differentiation factor (BCDF, IL-5; ) These fac­tors cause the progression of the B¹ cell to form B² and B³ cells, which differentiate into antibody-producing (eg, pen-tamer IgM) plasma cells CPC) Memory B cells are also produced.  B. Inducible protein B7 (w) on the B cell must interact with CD28 protein on the helper T cell in order for the helper T cell to be fully activated, and CD40L (CD40 ligand) on the helper T celJ must interact with CD40 on the B cell for the B cell to be activated and synthesize the full range of antibodies.

 

C. Suppression of Certain Immune Responses: Certain T cells can suppress antibody production. Failure of such regulation may result in unrestrained antibody production to self antigens, which can cause autoimmune diseases. There may not be a specific population of T cells that medi­ates suppression. There is evidence that in some situations CDS cells can suppress, but inhibitory lymphokines produced by CD4 cells also can play this role.

When there is an imbalance iumbers or activity between CD4 and CD8 cells, cellular immune mechanisms are greatly impaired. For example, in lepromatous leprosy there is unrestrained multi­plication of Mycobacterium leprae, a lack of delayed hypersensitivity to M leprae antigens, a lack of cellular immunity to that organism, and an excess of CD8 cells in lesions. Removal of some CD8 cells can restore cellular immunity in such patients and limit M leprae multiplication. In acquired immunodeficiency syndrome (AIDS), the normal ratio of CD4:CD8 cells (> 1.5) is greatly reduced. Many CD4 cells are destroyed by the human immunodeficiency virus (HIV), and the number of CD8 cells increases. This imbalance, ie, a loss of helper activity and an increase in suppressor activ­ity, results in a susceptibility to opportunistic infections and certain tumors.

One important part of the host response to infection is the increased expression of class I and class II MHC proteins induced by various cytokines, especially interferons such as gamma interferon. The increased amount of MHC proteins leads to increased antigen presentation and a more vigorous im­mune response. However, certain viruses can suppress the increase in MHC protein expression, thereby enhancing their survival. For example, hepatitis B virus, adenovirus, and cytomegalovirus  can prevent an increase in class I MHC protein expression, thereby reducing the cytotoxic T cell re­sponse against cells infected by these viruses.

 

B CELLS

B cells perform two important functions; (1) They differentiate into plasma cells and produce anti­bodies, and (2) they are antigen-presenting cells (APCs).

Origin. During embryogcncsis, B cell precursors are recognized first in the fetal liver. From there they migrate to the bone marrow, which is their main location during adult life. Unlike T cells, they do not require the thymus for maturation. Pre-B cells lack surface immunoglobulins and light chains but do have μ heavy chains in the cytoplasm. The maturation of B cells has two phases: the antigen-independent phase consists of stem cells, pre-B cells, and B.cells, whereas the antigen-dependent phase consists of the cells that arise subsequent to the interaction of antigen with the B cells, eg, activated B cells and plasma cells. B cells display surface IgM, which serves as a receptor for antigens. This surface IgM is a monomer, in contrast to circulating IgM, which is a pentamer. Surface IgD on some B cells may also be an antigen receptor. Pre-B cells are found in the bone marrow, whereas B cells circulate in the blood stream.

B cells constitute about 30% of the recirculating pool of small lymphocytes, and their life span is short, ie. days or weeks. Approximately 10⁹ B cells are produced each day. Within lymph nodes, they are located in germinal centers; within the spleen, they are found in the white pulp. They are also found in the gut-associated lymphoid tissue, eg, Peyer’s patches.

Clonal Selection. How do antibodies arise? Does the antigen “instruct” the B cell to make an antibody, or does the antigen “select” a B cell endowed with the preexisting capacity to make the an­tibody?

It appears that the latter alternative, ie, clonal selection, accounts for antibody formation. Each in­dividual has a large pool of B lymphocytes (about 10⁷). Each immunologically responsive B cell bears a surface receptor (either IgM or IgD) that can react with one antigen (or closely related group of antigens); ie, there are about l0⁷ different specificities. An antigen interacts with the B lympho­cyte that shows the best “lit” with its immunoglobulin surface receptor. After the antigen binds, the B cell is stimulated to proliferate and form a clone of cells. These selected B cells soon become plasma cells and secrete antibody specific for the antigen. Plasma cells synthesize the immunoglobulins with the same antigenic specificity (ie, they have the same heavy chain and the same light chain) as those carried by the selected B cell. Antigenic specificity does not change when heavy-chain class switching occurs.

Note that clonal selection also occurs with T cells. The antigen interacts with a specific receptor located on the surface of either a CD4-positive or a CD8-positive T cell. This “selects” this cell and activates it to expand into a clone of cells with the same specificity.

Activation of B Cells. In the following example, .the B cell is the APC. Multivalent antigen binds to surface IgM (or IgD) and cross-links adjacent immunoglobulin molecules. The immunoglobulins aggregate to form “patches” and eventually migrate to one pole of the cell to form a cap. Endocytosis of the capped material follows, the antigen is processed, and epitopes appear on the surface in conjunction with class II MHC proteins. This complex is recognized by a helper T cell with a receptor for the antigen on its surface.

(Macrophages bearing antigen bound to class II MHC proteins can also present antigen to the T cell, resulting in anti­body formation. In general, B cells are poor activators of “virgin” T cells, in the primary response because B cells do not make IL-1. B cells are, however, very good activators of memory T cells because little, if any, IL-I is needed). The T cell now produces various lymphokines (IL-2, IL-3, and IL-5) that stimulate the growth and differentiation of the B cell.

The activation of B cells to produce the full range of antibodies requires two other interactions in addition to recognition of the epitope by the T cell antigen receptor and the production of IL-4 and IL-5 by the helper T cell. These costimulatory interactions which occur between surface proteins on the T and B cells, are as follows: (1).CD28 on the T cell must interact with B7 on the B cell, and (2) CD40L on the T cell must interact with CD40 on the B cell. The CD28-B7 interaction is required for activation of the T cell to produce IL-2, and the CD40L-CD40 interaction is required for class switching from IgM to IgG and other immunoglobulin classes to occur.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Many plasma cells that produce large amounts of immunoglobulins specific for the epitope are the end result. Plasma cells secrete thousands of antibody molecules per second for a few days and then die. Some activated B cells form memory cells, which can remain quiescent for long pounds but are capable of being activated rapidly upon reexposure to antigen. Most memory B cells have surface IgG that serves as the antigen receptor, but some have IgM. Memory T cells secrete interleukins that enhance antibody production by the memory B cells. The presence of these cells ex­plains the rapid appearance of antibody in the secondary response.

 

Lymphocytes:

• Lymphocytes are differentiated into B lymphocytes (B cells), T lymphocytes (T cells), natural killer cells (NK cells), and killer cells (K cells); all have different roles in the immune defenses of the body.

• Lymphocytes are differentiated based on the presence of specific cell surface proteins or antigens bound to their plasma membranes.

• A surface marker that identifies a specific line of cells or a stage of cell differentiation because it interacts with a group or cluster of individual antibodies is called a CD (cluster of differentiation) antigen.

• B lymphocytes (B cells), which are differentiated in bone marrow, give rise to plasma cells when stimulated by antigen.

• Plasma cells produce and release antibodies. Each clone of a plasma cell line secretes a single specific antibody.

• T lymphocytes are differentiated in the thymus.

• Cytotoxic T cells (CD8⁺ cells) lyse abnormal cells.

• Helper T cells (CD4⁺ cells) activate the immune response.

• Suppressor T cells (CD8⁺ cells) regulate the immune response.

• Natural killer or null cells are a special subset of lymphocytes that are neither T nor B cells.

• Antigen-presenting cells (APCs) include macrophages and other specialized cells (dendritic cells) in skin and lymphoid organs. APCs help activate the immune response. APCs bind antigen on their surfaces in association with a special protein called the major histocompatibility complex (MHC).

• B cells that have processed antigens become antigenpresenting cells that activate the immune response.

• T cells respond to antigens associated with MHC on antigen-presenting cells.

 

 

Antibody-mediated Immunity:

• Antibody-mediated immunity is important in preventing and eliminating microbial infections.

• In the antibody-mediated immune response, specific glycoproteins called antibodies or immunoglobulins are made when foreign antigens activate B cells.

• The key to antibody immunity is the ability of antibodies to react specifically with antigens.

• The activation of a particular B cell leads to clonal selection and expansion of B cells specific to an individual antigen.

 

Cell-mediated Immunity:

• The cell-mediated immune response depends oatural killer cells and various T lymphocytes.      

• T cells recognize antigens by specific surface molecules called T cell receptors (TCRs).

• T cells produce cytokines when they are stimulated.

• Cytotoxic and aggressor T cells detect abnormal cells by antigens on tissue cell surface. These antigens are called major histocompatibility complex antigens. These MHC antigens are important.

 

 

 

 

 

 

 

 

 

Internet adresses:

http://www.online-medical-dictionary.org/Microbial+Antagonism.asp?q=Microbial+Antagonism

http://www.mansfield.ohio-state.edu/~sabedon/biol2035.htm

http://en.wikipedia.org/wiki/Antibiotic

http://www.nlm.nih.gov/medlineplus/antibiotics.html 

http://www.intmed.mcw.edu/AntibioticGuide.html

http://whyfiles.org/038badbugs/

http://www.niaid.nih.gov/factsheets/antimicro.htm

http://textbookofbacteriology.net/BSRP.html

http://www.bacteriamuseum.org/niches/pbacteria/pathogenicity.shtml

http://www.slic2.wsu.edu:82/hurlbert/micro101/pages/Chap20.html#Symbioses

http://www.slic2.wsu.edu:82/hurlbert/micro101/pages/Chap12.html#NSD_mechanisms

http://www2.hawaii.edu/~johnb/micro/m130/m130lect12.html

http://www.merck.com/mmpe/sec14/ch167/ch167b.html

http://en.wikipedia.org/wiki/Phagocytosis

http://en.wikipedia.org/wiki/Complementary

 http://www-micro.msb.le.ac.uk/MBChB/Merralls/Merralls.html               

http://en.wikipedia.org/wiki/Immune_system

http://www-micro.msb.le.ac.uk/MBChB/1b.html

http://www.answers.com/topic/lipopolysaccharide-1

http://www.answers.com/topic/flagellar-antigen

http://www.google.com/search?hl=en&q=K+antigen&btnG=Search

http://www.callutheran.edu/Academic_Programs/Departments/BioDev/omm/viral_antigens/molmast.htm

http://en.wikipedia.org/wiki/Major_histocompatibility_complex

http://www.cryst.bbk.ac.uk/pps97/assignments/projects/coadwell/MHCSTFU1.HTM

http://en.wikipedia.org/wiki/Antibody

http://www.fleshandbones.com/readingroom/pdf/291.pdf