- The concept of immunity
- The elements of innate immunity: Physical barriers, soluble mediators, cells and their receptors
- The hallmarks of adaptive immunity: Specificity, memory, tolerance
- Repertoire and receptor rearrangement
- Cells and molecules of adaptive immunity
- The adaptive immune response
- Immune tolerance
The concept of immunity
All multicellular organisms have evolved defense mechanisms against the onslaught of viruses, bacteria, and other parasites. The immune system of vertebrates combines evolutionarily innate (or natural) immunity with newer adaptive responses against specific antigens. It is important to keep in mind that the two systems are tightly integrated, and several molecules and cells, in addition to their fundamental role in one system, act as interfaces between the two. In the first part of this review we will briefly examine the main elements of innate and adaptive immune responses. An excellent reference for further reading is the textbook by Abbas and Lichtman 
The elements of innate immunity: Physical barriers, soluble mediators, cells and their receptors
The main elements of innate immunity are physical barriers, soluble mediators that directly inhibit foreign microorganisms and specialized cells endowed with recognition receptors and with the ability to phagocytose and kill microorganisms.
Physical barriers are sometimes underestimated as part of the immune system, even though it is common wisdom that an intact skin is a very effective protection from infection, while even superficial wounds can be easily colonized by foreign invaders. In addition to skin, physical barriers include the blood coagulation system that stops bleeding and forms a protective clot over wounds, and internal mucosae, protected by mucus and continuously cleansed by the action of ciliated cells that sweep away bacteria and other materials. The mildly acidic pH of skin inhibits bacterial proliferation, and the very acidic pH of stomach juices effectively sterilizes ingested material.
A variety of soluble molecules present on the skin, in secretions, in blood, and throughout the body directly kill bacteria and inhibit viral infections. Prominent examples are defenses (natural antibiotics), the lysozyme (an enzyme that lyses the wall of some bacteria), complement (a cascade of proteins activated by bacterial components that make holes in bacterial membranes), opsonins (complement fragments and other proteins that coat bacteria and facilitate phagocytosis) and some ancient cytokines such as type I (à and ß) interferons that are released by virus infected cells and induce an anti-viral state in neighboring cells.
Cellular migration toward invaders and phagocytosis are the basic cellular mechanisms of innate immunity, followed by intracellular destruction of the ingested microorganism. Extracellular killing is also possible if phagocytes release lytic substances in the environment. In mammals the two main cell types are granulocytes, which contain lytic substances packed in intracellular granules (also called polymorphonuclear cells because their nucles is of irregular shape) and macrophages. Neutrophil granulocytes have an important role in defense against bacterial infections, basophils have an anti-parasitic role. Macrophages derive from circulating monocytes that emigrate to practically all tissues and organs, sometimes under different names (e.g. Kupffer cells in the liver). In addition to immune functions, macrophages play important roles in the turnover of aging cell components, like red blood cells which are continuously phagocytosed by splenic macrophages. Natural killer (NK) cells are non-phagocytic elements that kill virus-infected cells; it is worth noting that the evolution of NK cells is relatively recent and goes in parallel with the evolution of lymphocytes, to which NK cells closely resemble from a morphological point of view.
Recognition in innate immunity is mediated by interactions with pathogen-associated molecular patterns expressed by microorganisms. Cellular receptors are globally referred to as "pattern-recognition receptors" and include the family of Toll-like receptors (TLR), mannose receptors (MR) and seven-transmembrane spanning receptors (TM7). TLR recognize bacterial and viral nucleic acids, flagellin, bacterial peptides, lipopolysaccharide (LPS) and other bacterial components. MR bind carbohydrate moieties on several pathogens, such as bacteria, fungi, parasites, and viruses. TM7 receptors are activated by bacterial peptides or by endogenous chemokines
The hallmarks of adaptive immunity: Specificity, memory, tolerance
The evolution of adaptive immunity accompanies the evolution of vertebrates, starting from sharks. It is therefore conceivable that part of the evolutionary success and long life spans of vertebrates could be attributed to improved defenses against exogenous pathogens. The functions of adaptive immunity are performed by a new type of cell (the lymphocyte) which recirculates between blood, tissues and a specialized circulatory system (the lymphatic vessels) and reside in specialized organs (lymphoid organs, which include the thymus, the spleen and lymph nodes).
The hallmarks of adaptive immune responses are specificity, immune memory and immune tolerance. The immunological meaning of specificity refers to the fact that a lymphocyte population is composed of millions of clones, each specific for a different antigen. Specificity is encoded in clonotypic antigen receptors generated by a process of DNA rearrangement that includes random events to produce billions of variant molecules from a relatively small pool of DNA sequences.
Specificity allows a considerable economy in adaptive immune responses to pathogens, because only the clones expressing receptors for a specific microorganism are activated upon infection, while all other clones remain inactive. In addition to the proliferation of specific lymphocyte clones, the first encounter with a given antigen (primary immune response) leaves behind a population of memory cells that will respond more promptly and more efficiently to subsequent encounters (secondary immune response). The presence of random events in the generation of specific antigen receptors implies the risk of producing autoreactive receptors. To prevent autoimmunity, the differentiation of lymphocytes is accompanied by selective mechanisms that ensure tolerance to autologous (self) components through the destruction or the inactivation of autoreactive clones.
Repertoire and receptor rearrangement
How many different antigens can be specifically recognized by the adaptive immune response?
A theoretical estimate is of the order of different genes. Given that mammalian genomes contain between and different genes, one might wonder how to encode such a large repertoire of specific antigen receptors. The answer resides in a unique DNA rearrangement that happens only in lymphocytes to produce antigen receptor genes. Antigen receptor genes comprise two or three sets of - alternative segments encoding the variable, antigen-binding part (V, D and J segments) separated by non-coding DNA segments and followed by one or a few segments encoding the constant (C) part of the receptor. During lymphocyte differentiation one segment from each variable set is chosen at random and brought in proximity to one another by enzymes that cut and discard the intervening DNA. At the end of the process the rearranged DNA of lymphocytes is different from that of all other cells of the organism, originally inherited by the individual (the so-called germline configuration), and each lymphocyte contains a slightly different sequence encoding the antigen receptor. Receptor diversity is further increased by enzymes that cause sequence alterations at the point of contact between the various segments (junctional diversity) and by the fact that functional membrane antigen receptors are dimers of molecules encoded by separate genes, each undergoing DNA rearrangement independently.
Cells and molecules of adaptive immunity
Lymphocytes comprise populations with different types of antigen receptors and diverse functions. The most fundamental distinction is between T and B cells. B cells use immunoglobulins (Ig) as membrane antigen receptors and, after receptor stimulation by the antigen, differentiate into plasma cells which actively secrete a soluble form of the receptor, called antibody.
Each molecule is a dimer of a heavy and a light chain, each with a variable and a constant part. The basic antibody molecule is Y shaped and has two independent antigen-binding sites (at the ends of the diagonal segments of the Y). The stem of the Y is constant and mediates the so-called effector functions of the antibody. Most mammals have multiple C segments in their genome, giving rise to different classes of antibodies. IgM are the first antibodies produced in primary immune response, IgM molecules are assembled from five basic molecules, thus have ten antigen-binding sites. IgG are the main class of antibodies produced in secondary immune responses and released in the bloodstream. In humans there are 4 different IgG types (IgG1, IgG2, IgG3 and IgG4) endowed with subtle differences in their functions. IgA are specialized for functioning in secretions like milk, tears, saliva and intestinal juices rather than in blood. IgE are best known in connection with allergies (see below). The function of antibodies is to bind to and inactivate their cognate antigens. This is accomplished through various mechanisms. Binding of multiple antibody molecules to multiple antigens can yield an insoluble antigen-antibody complex that is rapidly removed from the blood. Antibody binding to cell membranes activates complement components that can lyse the bound cell. Various leukocytes express surface receptors for the constant part of antibodies that mediate further functions such as phagocytosis (opsonization) and cell lysis (antibody-dependent cell-mediated cytotoxicity, ADCC). In this way cells of the innate immunity, such as the macrophages or NK cells acquire the antigen specificity of adaptive immunity. Other antibody receptors mediate the passage of maternal antibodies through the placenta and the intestinal uptake of antibodies present in maternal milk, thus conferring a passive protection to the fetus and to the lactating newborn.
T cells use a different type of surface antigen receptor (TCR). A major conceptual difference between T and B responses is that antibodies are at the same time the receptor and the effector molecule of B cell immunity, while the TCR is a membrane receptor that activates a signaling cascade culminating in effector actions mediated by other molecules. A second important difference is that antibodies bind practically any conceivable molecular species, from proteins to sugars and lipids, to small organic molecules, whereas the TCR recognizes small peptides derived from cellular metabolism of proteins. A third difference is that the antigen-binding site of antibodies recognizes the antigen in its free native form, whereas the TCR only recognizes peptides present on the cell membrane bound to a cellular protein called the major histocompatibility complex (MHC).
T cells comprise various subpopulations with different functions. Cytotoxic T cells (Tc, also referred to as CTL) directly kill cells expressing the antigen, helper T cells (Th) positively regulate the activity of B and Tc cells, while regulatory T cells (Treg) inhibit immune responses. The activity of Th and Treg is mediated both by direct cell-cell contact and by cytokine secretion. Th cells are further divided into type 1 and type 2 cells (Th1 and Th2) according to the spectrum of secreted cytokines and the type of immune response they help: Th1 cells release γ (interferon γ) and other cytokines to stimulate immune responses directed against viruses and intracellular bacteria, whereas Th2 cells release interleukin 4 (IL-4) to stimulate the immune response against parasites. There are various molecular markers that distinguish T cell subpopulations, among them CD8 for Tc and CD4 for Th. CD4 is also well known for being used by the human immunodeficiency virus (HIV) to infect helper T cells.
The adaptive immune response
The adaptive immune response involves a complex interplay of different cells. As an example we will follow from beginning to end an anti-viral response to explain the various cellular and molecular interactions. The immune response begins when viral proteins are captured in the periphery of the body (e.g. skin, mucosae, etc) by a specialized population of leukocytes called dendritic cells (DC). DC migrate to lymph nodes where they present to Th cells antigenic peptides generated from viral proteins and exposed on the DC membrane in association with MHC molecules. Intereleukin 12 (IL-12) secreted by DC is a potent activator of Th1 responses. The antigen-presenting function (APC) can be performed also by other cell types such as macrophages or B cells. Recognition of the peptide-MHC complex by a specific TCR activates the Th cell that proliferates and secretes cytokines, in turn activating antibody production by B cells and replication of Tc cells. Antibodies, Th and Tc cells reach the periphery where they encounter free viruses and virus-infected cells. Antibodies inactivate free viruses by direct binding and kill virus-infected cells through complement-mediated cytotoxicity and, with the help of NK cells, ADCC. Th cells secrete an array of cytokines that attract other leukocytes, inhibit viral replication, and stimulate hemopoiesis to increase the number of leukocytes in the body. Tc recognize and kill virus-infected cells expressing viral peptide-MHC complexes on their membrane.
Random generation of antigen receptors entails the risk of generating self-reactive immune responses (autoimmunity). Various differentiative and evolutionary biologically natural processes are intrinsically irreversible, and the natural way to cope with useless or potentially harmful cells is to produce the cells first and to destroy them afterwards.
T cells produced by hemopoiesis in the bone marrow migrate to the thymus where T cell rearrangement takes place followed by counterselection of clones expressing inappropriate receptors (clonal deletion). As soon as newly differentiated T cells express the rearranged TCR they interact with autologous MHC-peptide complexes expressed by thymic cells. Autoreactive T cells that recognize autologous MHC-peptide complexes die by apoptosis (negative selection), as do T cells with missing or defective TCR that completely fail to interact with autologous MHC. Only T cells whose TCR interacts with appropriate affinity with the MHC, thus potentially reactive with foreign antigens, survive and thrive (positive selection). It is said that T cells are educated in the thymus, but we must keep in mind that the thymus is a harsh mistress: more than 90% of T cells are killed during the selection process. It is interesting to note that thymic cells express a wide range of peptides, including those of proteins usually produced by specialized tissues and organs, like the pancreas and the thyroid, hence T cells in the thymus are exposed to a comprehensive sampling of the peptides they will subsequently encounter in the periphery.
Deletion of autoreactive clones in the thymus (central tolerance) is not 100% efficient, therefore all normal adults harbor autoreactive T cells that are rendered harmless by tolerogenic processes in the periphery of the body. Peripheral tolerance is the result of the dependence of T cell activation from antigen presentation by APC. Only APC express appropriate costimulatory molecules required (in addition to the MHC-peptide complex) to activate T cells. If a T cell encounters the antigen on a parenchymal cell lacking costimulatory molecules the ability of the T cell to be activated is inhibited (anergy).