Positive and negative selection of T lymphocytes. Immunopoiesis: maturation of T - and B - cell receptors
The essence of the clonal selection theory of F. Bernet is that in the process of maturation of lymphocytes, strict culling of cells occurs according to the following characteristics:
Inability to recognize the MHC 1 and MHC 2 receptors of the body's own cells;
The ability to recognize self-antigens presented on MHC 1 and MHC 2.
Cells that have the above characteristics must be destroyed. The remaining lymphocytes continue differentiation and become the founders of clones - groups of lymphocytes that have an antigen recognition receptor of the same specificity.
T-lymphocyte selection
Immature T lymphocytes migrate from the bone marrow to the thymic cortex and begin to rapidly divide. In the thymic cortex, during contact with thymic epithelial cells that express both MHC I and MHC II molecules, positive selection occurs. Lymphocytes that are able to interact with MHC molecules receive a positive stimulus - a signal to reproduce, and cells that are unable to interact with MHC receive a negative signal to self-destruct (apoptosis).
Next, lymphocytes that have undergone positive selection migrate to the thymic medulla and negative selection of T lymphocytes occurs at the border of the cortex and medulla. Negative selection is carried out during their interaction with dendritic cells and macrophages, which present the body’s own antigens.
Autoaggressive T-lymphocytes receive a signal to self-destruct (negative selection), autotolerant ones continue to multiply and leave the thymic medulla, settling in the peripheral organs of the immune system. It has been shown that during the selection process, about 95% of T lymphocytes are not selected and die.
Lymphocytes located in the thymic cortex initially have both CD4 and CD8 receptors on their membrane. Next, cells that recognize MHC I lose CD4 and become CD8+, i.e. turn into CTLs, and cells that recognize MHC II, on the contrary, lose CD8 and turn into CD4+, i.e. in T helper cells.
T lymphocytes that have undergone differentiation and selection in the thymus are called “naive” T lymphocytes. After meeting the corresponding antigen, they turn into primed or effector T lymphocytes, ready to perceive cytokine signals for activation.
Selection of b-lymphocytes
In the bone marrow, immature B lymphocytes undergo negative selection. Lymphocytes that are able to bind their own antigens with their surface antigen-recognition receptor IgM receive a signal to self-destruct (apoptosis) and die. B-lymphocytes that have passed negative selection divide, and each of them forms a group of descendants, a clone, with the same specificity. Mature B lymphocytes leave the bone marrow into the bloodstream and populate the lymphoid organs.
Lecture 6. Immunity disorders
Immunity disorders include:
Hypersensitivity reactions;
Autoimmune reactions;
Immunodeficiency states.
Hypersensitivity reactions. Jell and Coombs classification – 4 types of hypersensitivity reactions.
Type 1 MIRV.
Asthma, hay fever, eczema, hives, food allergies.
Allergens: drugs, heterologous serum, plant pollen, feces of microdust mites, food products (eggs, milk, crabs, fish, etc.).
Factors facilitating the penetration of allergens into the mucous membrane are diesel exhaust particles (DEP) contained in the urban atmosphere.
Hereditary predisposition to type 1 allergic reactions is associated with the HLA-B8 and DR3 alleles.
Diagnosis: skin testing.
Treatment: hyposensitization - subcutaneous administration of increasing doses of the allergen, resulting in a switch to the predominant synthesis of IgG.
Prevention: avoiding contact with the allergen; if it is necessary to administer a heterologous therapeutic serum, fractional administration according to Bezredka. Prescribing antihistamines.
HSR type 2 – cytotoxic reactions involving IgG and complement. Observed when antibodies react with an antigen located on the cell membrane. In this case, complement is added to the resulting complex, the last fractions of which (C5-C9) are called perforins. The protein molecules of these fractions are embedded in the cell membrane, forming a large pore through which water enters the cell. As a result, cell lysis occurs. This type of hypersensitivity can develop with long-term use of drugs that can be adsorbed on red blood cells; An example is the antiarrhythmic drug quinidine. An example of type 2 HSR is hemolytic disease of newborns with Rh conflict (reticulocytosis). Another example is thrombocytopenic purpura.
Type 3 HSR are associated with the formation of a large number of immune complexes when a large amount of foreign protein enters the body without prior sensitization, for example, with the introduction of therapeutic or prophylactic heterologous antisera. As a result of temporary complement deficiency, small immune complexes are deposited in the walls of blood vessels, joints, and renal glomeruli. After completing the complement deficiency, it is fixed on small immune complexes (SICs) located in the tissues. Macrophages migrate to the formed large immune complexes (LIC), which absorb LIC and release cytokines that cause an inflammatory response. The result of type 3 HSR is the development of serum sickness, the manifestations of which are vasculitis, arthritis and glomerulonephritis.
Type 3 HSR can manifest itself in the form of the so-called Arthus phenomenon. Unlike serum sickness, the Arthus phenomenon is a violent local inflammatory reaction, which is accompanied by tissue necrosis at the site of antigen injection. A prerequisite for the development of the Arthus reaction is preliminary sensitization of the body with this antigen (foreign protein) and the presence in the blood serum of a high concentration of antibodies to this antigen.
Type 4 HSR occurs with the participation of cytotoxic lymphocytes.
There are 3 types of type 3 HSR: contact, tuberculin and granulomatous.
Contact hypersensitivity is characterized by an eczematous reaction at the site of antigen exposure. Sensitization of the body occurs, as a rule, with nickel and chromium compounds, substances included in detergents, i.e. haptens. The main APCs in contact hypersensitivity are dendritic cells of the skin - Langerhans cells. The contact hypersensitivity reaction occurs in 2 stages: sensitization and manifestation. The sensitization period lasts about 2 weeks. The hapten, having penetrated the skin, combines with protein. This complex is taken up by dendritic cells, which subsequently present the hapten-protein complex to T lymphocytes. In a sensitized organism, after repeated contact with the antigen within 48-72 hours, T-lymphocytes migrate to the site of contact with the antigen and a local inflammatory reaction develops.
Tuberculin-type hypersensitivity. Tuberculin is a filtrate of a killed tubercle bacillus culture containing bacterial antigens. It was first obtained by R. Koch.
A hypersensitivity reaction to tuberculin occurs only in individuals who have live tuberculosis pathogens in their bodies. After intradermal injection of tuberculin, monocytes and sensitized T-lymphocytes migrate to the injection site and secrete cytokines (TNF-alpha and beta). Cytokines increase the permeability of the vascular wall and an inflammatory infiltrate is formed at the site of tuberculin injection, which reaches its maximum size after 48 hours.
Granulomatous hypersensitivity. Granulomatous reactions develop in cases where the infectious agent remains viable in macrophages, for example, in tuberculosis and leprosy. An activated macrophage, inside of which there are live pathogens, is transformed into an epithelioid cell that actively produces cytokines - TNF. Epithelioid cells fuse with each other to form Langhans giant cells. In the center of the granuloma there are epithelioid cells, Langhans cells and macrophages. The center of the granuloma is surrounded by T lymphocytes. Outside the T-lymphocytes there is a zone of proliferating fibroblasts, which delimit the inflammatory zone from healthy tissue.
The main purpose of T lymphocytes is to recognize surface structures own body cells. If something on the surface of its cells “irritates” the T-lymphocyte (for example, an admixture of viral peptides), then it will try to organize the destruction of the damaged cell.
Unlike B-lymphocytes, T-lymphocytes do not produce soluble forms of Ag-recognition molecules and always “work” with their own “cell body”. Moreover, most T lymphocytes are not able to recognize and bind soluble Ag.
In order for a T-lymphocyte to “pay attention to Ag,” other cells must somehow pass Ag through themselves and display it on their membrane in complex with MHC-I/II. This is the phenomenon of Ag presentation to the T lymphocyte. Recognition of such a complex by T-lymphocytes is double recognition, or MHC restriction of T-lymphocytes.
receptor for t-lymphocyte antigen
Antigen-recognizing RCs of T-lymphocytes - TCRs - belong to the immunoglobulin superfamily (see Fig. 5.1). The Ag-recognition region of the TCR protruding above the cell surface is a heterodimer (i.e., consists of two different polypeptide chains) - an analogue of one Fab fragment of Ig. There are two known TCR variants, designated TCRαβ and TCRγδ; these variants differ in the composition of the polypeptide chains of the Ag-recognition region. Each T lymphocyte carries only one Rc variant. Tαβ became known earlier and was studied in more detail than Tγδ; Therefore, it is more convenient to describe the structure of RC T-lymphocytes for Ag using the example of TCRαβ. A completely transmembrane-located TCR consists of 8 or 10 (one or two α + β pairs plus the “2ε + δ + γ + 2ζ” complex) polypeptide chains (Fig. 6.1).
Rice. 6.1. TCR receptor ap T lymphocytes for antigen.
The Ag-binding region of the receptor is formed by α- and β-chains; chains γ, δ, ε (together called the CD3 complex) are necessary for the expression of α- and β-chains, their stabilization and, probably, signal transmission into the cell; The ζ chain, the most “intracellular” one, ensures signal transmission into the cell.
Transmembrane chainsα Andβ TCR. These are 2 approximately equal polypeptide chains - α (molecular weight 40-60 thousand, acidic glycoprotein) and β (molecular weight 40-50 thousand, neutral or basic glycoprotein). Each of these chains has two glycosylated domains in the extracellular part of the RC, a hydrophobic (positively charged due to lysine and arginine residues) transmembrane part and a short (5-12 AA residues) cytoplasmic region. The extracellular parts of both chains are connected by a single disulfide bond.
♦ V-region. The outer extracellular (distal) domains of both chains have a variable AK composition. They are homologous to the V region of Ig molecules, this is the V region of the TCR. It is the V regions of the α and β chains that interact with the MHC-I/II-peptide complex.
♦ C-area. The proximal domains of both chains are homologous to the Ig constant regions, these are the TCR C regions.
♦ Short cytoplasmic region(both α- and β-chains) cannot independently ensure signal transmission into the cell. For this purpose, 6 additional polypeptide chains are used: γ, δ, two ε and two ζ.
CD3 complex. The γ, δ, ε chains (together called the CD3 complex) are required for the expression of the α and β chains, their stabilization, and, possibly, signal transmission into the cell. The CD3 complex consists of an extracellular, transmembrane (negatively charged and
therefore electrostatically associated with the transmembrane regions of the α- and β-chains) and the cytoplasmic parts.
♦ ζ -Chains connected to each other by a disulfide bridge and, being mostly located in the cytoplasm, carry out the signal into the cell.
♦ ITAM sequences. The cytoplasmic regions of the polypeptide chains γ, δ, ε and ζ contain ITAM AK sequences (1 in the γ and δ chains, 2 in the ε chains, 3 in each ζ chain), which interact with cytosolic tyrosine kinases (activation of these enzymes and constitutes the beginning of biochemical reactions for signal transmission).
Ionic, hydrogen, van der Waals and hydrophobic forces participate in the binding of Ag, and the conformation of RC changes significantly. Each TCR is potentially capable of binding about 10 5 different Ags, not only structurally related (cross-reacting), but also not having homologies in structure.
TCR genes
The genes of the α-, β-, γ- and δ-chains (Fig. 6.2) are homologous to the Ig genes and undergo somatic DNA recombination during the differentiation of T-lymphocytes, which theoretically ensures the generation of about 10 16 -10 18 variants of antigen-binding centers (in reality, this diversity is limited the number of lymphocytes in the body is up to 10 9). Genesα -chains have 70-80 V-segments, 61 J-segments and one C-segment.
Rice. 6.2. Genesα- Andβ -chains of the T-lymphocyte receptor for antigen.
Genesβ -chains contain 52 V-segments, 2 D-segments, 13 J-segments, and 2 C-segments.
Genesδ -chains. Between the V and J segments of the α chain are the genes of the D, J and C segments of the TCRγδ δ chain. The V segments of the δ chain are interspersed among the V segments of the α chain.
Genesγ -chains TCRγδ have 2 C segments, 3 J segments before the first C segment and 2 J segments before the second C segment, 12 V segments.
Gene rearrangement
DNA recombination occurs when the V-, D- and J-segments combine and is catalyzed by the same recombinase complex as during the differentiation of B-lymphocytes.
After rearrangement of VJ in α-chain genes and VDJ in β-chain genes, as well as the addition of non-coding N- and P-nucleotides to DNA, RNA is transcribed. Fusion with the C-segment and removal of excess (unused) J-segments occurs during splicing of the primary transcript.
α-chain genes can rearrange multiple times while β-chain genes are already correctly rearranged and expressed, so there is some possibility that a single cell may carry more than one TCR variant.
TCR genes are not subject to somatic hypermutagenesis.
Coreceptor molecules CD4 and CD8
In addition to the TCR itself, each mature T lymphocyte expresses one of the so-called coreceptor molecules - CD4 or CD8, which also interact with MHC molecules on APCs or target cells. Each of them has a cytoplasmic region associated with the tyrosine kinase Lck, and probably contributes to the transmission of the signal into the cell upon recognition of Ag.
CD4 interacts with the invariant part (β2 domain) of the MHC-II molecule (belongs to the Ig superfamily, see Fig. 5.1B). CD4 has a molecular weight of 55 thousand and 4 domains in the extracellular part. When a T-lymphocyte is activated, one TCR molecule is “served” by two CD4 molecules (probably dimerization of CD4 molecules occurs).
CD8 binds to the invariant part (α3 domain) of the MHC-I molecule (belongs to the Ig superfamily, see Fig. 5.1A). CD8 is a heterodimer of α and β chains linked by a disulfide
communication In some cases, a homodimer of 2 α-chains is found, which can also interact with MHC-I. In the extracellular part, each of the chains has one immunoglobulin-like domain.
CONDUCTING SIGNALS FROM LYMPHOCYTE IMMUNORECEPTORS
Lymphocyte receptors for Ag (TCR and BCR) have a number of common patterns of registration and transmission of activation signals into the cell (see Fig. 5.8).
Receptor clustering. To activate a lymphocyte, clustering of RC and coreceptors is necessary, i.e. “linking” of several RCs with one Ag.
Tyrosine kinases. The processes of phosphorylation/dephosphorylation of proteins at tyrosine residues under the action of tyrosine kinases and tyrosine phosphatases play a significant role in signal transmission, leading to the activation or inactivation of these proteins. These processes are easily reversible and “convenient” for fast and flexible cell reactions to external signals.
Src kinases. Tyrosine-rich ITAM sequences of the cytoplasmic regions of immunoreceptors are phosphorylated by non-receptor (cytoplasmic) tyrosine kinases of the Src family (Fyn, Blk, Lyn in B lymphocytes, Lck and Fyn in T lymphocytes).
♦ The activity of Src kinases depends on the state of the C-terminal region of the molecule: its phosphorylation by the Csk kinase inactivates it, and dephosphorylation by the transmembrane tyrosine phosphatase CD45 activates the enzyme.
♦ Another mechanism for regulating the activity of Src kinases is their covalent binding to ubiquitin through the adapter protein Cb1. Binding to ubiquitin “directs” any protein to degradation in proteasomes.
Other kinases. The kinases Syk (in B lymphocytes) and ZAP-70 (in T lymphocytes), binding to phosphorylated ITAM sequences, are activated and begin to phosphorylate adapter proteins: LAT (Linker for Activation of T cells) and SLP-76 (Syk ), BLNK and SLP-65 (ZAP-70).
Phospholipase Cγ (see Fig. 4.3). Tec family kinases (Btk in B lymphocytes, Itk in T lymphocytes) bind adapter proteins
and activate phospholipase Cγ (PLCγ).
♦ PLCγ cleaves phosphatidylinositol diphosphate (PIP 2) of the cell membrane into phosphatidylinositol triphosphate (PIP 3) and diacylglycerol (DAG).
♦ DAG remains in the membrane and activates protein kinase C (PKC), a serine/threonine kinase that activates the evolutionarily “ancient” transcription factor NFkB.
♦ PIP 3 binds to its RC in the endoplasmic reticulum and releases calcium ions from the depot into the cytosol.
♦ Free calcium activates calcium-binding proteins - calmodulin, which regulates the activity of a number of other proteins, and calcineurin, which dephosphorylates and thereby activates the nuclear factor of activated T-lymphocytes NFAT (Nuclear Factor of Activated T-cells).
Small G proteins Ras in the inactive state are associated with GDP, but adapter proteins replace the latter with GTP, thereby transferring Ras to the active state.
♦ Ras has its own GTPase activity and quickly cleaves off the third phosphate, thereby returning itself to an inactive state (self-inactivation).
♦ In a state of short-term activation, Ras manages to activate another cascade of kinases called MAP (Mitogen Activated Protein kinase), which ultimately activate the transcription factor AP-1 (Activator Protein-1) in the cell nucleus.
DIFFERENTIATION OF T-LYMPHOCYTES
The differentiation processes occurring in the thymus have been studied in sufficient detail and represent the following sequence of events:
Thymocytes differentiate from a common precursor cell, which, outside the thymus, expresses membrane markers such as CD7, CD2, CD34 and the cytoplasmic form of CD3.
The precursor cell committed to differentiation into a T-lymphocyte migrates from the bone marrow to the subcapsular zone of the thymic cortex, where slow cell proliferation occurs for approximately 1 week. New membrane molecules CD44 and CD25 appear on thymocytes.
Then the cells move somewhat deeper into the thymic cortex, and the CD44 and CD25 molecules disappear from their membrane. At this stage
rearrangement of the genes of the β-, γ- and δ-chains of the TCR begins. If the genes of the γ- and δ-chains manage to rearrange themselves productively (i.e., without a reading frame shift) earlier than the genes of the β-chain, then the lymphocyte differentiates further as Tγδ. Otherwise, the β-chain is expressed on the membrane in complex with pTα (an invariant surrogate chain that replaces the real α-chain at this stage) and CD3. This serves as a signal to stop rearrangement of γ- and δ-chain genes. Cells begin to proliferate and express both CD4 and CD8 (double positive thymocytes). In this case, a mass of cells accumulates with a ready-made β-chain, but with not yet rearranged α-chain genes, which contributes to the diversity of α-β-heterodimers.
At the next stage, the cells stop dividing and begin to rearrange Vα genes, several times over the course of 3-4 days. Rearrangement of the α-chain genes results in irreversible deletion of the δ-locus located between the segments of the α-chain genes.
TCR expression occurs with each new variant of the α-chain, and selection (selection) of thymocytes occurs based on the strength of binding to the “peptide-MHC” complex on the membranes of thymic epithelial cells.
♦ Positive selection: thymocytes that have not bound any of the available peptide-MHC complexes die. As a result of positive selection, about 90% of thymocytes die in the thymus.
♦ Negative selection destroys clones of thymocytes that bind peptide-MHC complexes with too high affinity. Negative selection eliminates from 10 to 70% of cells that have undergone positive selection.
♦ Thymocytes that have bound any of the “peptide-MHC” complexes with correct(i.e., average in strength) affinity, we get signal for survival and continue differentiation.
For a short time, both coreceptor molecules disappear from the thymocyte membrane, and then one of them: thymocytes that recognize the peptide in complex with MHC-I express the CD8 coreceptor, and with MHC-II, the CD4 coreceptor. Accordingly, two types of T-lymphocytes reach the periphery (in a ratio of about 2:1): CD8 + (or T8) and CD4 + (or T4), functions which in upcoming immune responses are different.
♦ CD8+ T lymphocytes perform functions cytotoxic T lymphocytes(CTL), or “perforin-granzyme killers”. With their “cell body” they directly kill cells on the membrane of which they recognize Ag.
Rice. 6.3. The mechanism of action of a cytotoxic T-lymphocyte on a target cell. IN
killer granules with perforin, in response to an increase in Ca2+ concentration, merge with the cell membrane. The released perforin is incorporated into the membrane of the target cell with the subsequent formation of pores permeable to granzymes, water and ions. As a result, the target cell is lysed.
♦ CD4+ T lymphocytes. The functional specialization of immune CD4 + T lymphocytes is more diverse. From them, perforin-granzyme cytotoxic T-lymphocytes - CD4 + CTLs - can develop (in particular, such T-lymphocytes are found in significant quantities in the skin of patients with Lyell's syndrome). Apparently, a significant part of CD4 + T-lymphocytes in the process of developing an immune response become T-helpers - “professional” cytokine producers that “hire” other executor cells to destroy tissues damaged by the pathogen.
Immune deviation. A change in the terminal differentiation of immune CD4+ T lymphocytes towards the predominance of one or another subpopulation during the development of the immune response is called an immune deviation.
T helper subpopulations
Since the late 80s of the 20th century, it has been customary to distinguish two subpopulations of T helper cells (depending on what set of cytokines they produce) - Th1 and Th2. In a slightly transformed version, this concept (despite its significant conventionality) “has taken root” among immunologists and doctors, and it continues to be used, isolating the following types of T4 lymphocytes:
Th0- T4 lymphocytes in the early stages of immune development
response, they produce only IL-2 (a mitogen for all lymphocytes);
Th 1- a differentiated subpopulation of immune T4 lymphocytes, specializing in the production of IFNγ (manager of immune inflammation carried out by activated macrophages in the form of delayed-type hypersensitivity - DTH);
Th2- a differentiated subpopulation of immune T4 lymphocytes, specializing in the production of IL-4 and its “understudy” IL-13 (manager of the immune response with a predominance of IgE production and variants of immune inflammation dependent on it);
Th3- immune T4 lymphocytes at later stages of development of the immune response, switching to the production of transforming growth factor (TGFβ) - an inhibitor of lymphocyte proliferation;
T g - T4 regulators, producers of immunosuppressive cytokines - IL-10 (inhibitor of macrophage and Th1 activity) and TGFβ. It is also possible that inducers of apoptosis of activated and spent lymphocytes - FasL (Fas ligand), etc. - are expressed on the Tg membrane.
Subsequently, it became known that each mature immune T4 lymphocyte produces only one cytokine(only in rare cases, perhaps two), therefore, at present, most authors propose to talk not about different subpopulations of immune T4 lymphocytes, but about different types of immune response.
Types of immune response
Type I immune response
Properties. IFNγ and activated macrophages dominate. On the part of T lymphocytes, this response is promoted not only by CD4+ Th1, but also by other IFNγ producers - CD8 + lymphocytes and NK.
Biological effects of IFNγ , aimed at destroying cells infected from the inside: - direct antiviral effect at the level of nucleic acid enzymes (2"-5"-oligoadenylate synthetase, etc.); - strong stimulation of macrophages, correspondingly increased synthesis of toxic products of macrophages; - NK stimulation. - IFNγ supports the switching of immunoglobulin synthesis in B-lymphocytes to IgG, which activates phagocytes (neutrophils and macrophages), i.e. T-lymphocytes - producers of IFN - provide macrophage and
the cytotoxic nature of immune inflammation of tissues damaged by the pathogen.
Pathohistology. Immune inflammation type I consists of foci of hyperthyroidism, granulomas and similar changes in tissues.
Type II immune response
Characteristic. Type II immune response is a response driven by other cytokines (eg IL-4). IL-4 producers: CD4 + Th2, “null” (CD4/CD8) T-lymphocytes, mast cells.
♦ Th2 lymphocytes support the switching of the synthesis of immunoglobulin isotypes in B lymphocytes to IgE, IgG4 and IgA. Partner cells for these isotypes are mast cells, basophils and eosinophils. When they are activated, inflammatory processes develop with a pronounced vasoactive component and exudation or characteristic eosinophilic inflammation.
♦ Except in pathological cases of IgE-dependent allergic reactions, the type II immune response is generally considered to be anti-inflammatory.
Examples of immune inflammation. Pathological processes with a predominance of immune inflammation type I (Th1) or II (Th2) are listed below.
Th1 (I) (macrophage inflammation- HRT, granulomas: Hashimoto's thyroiditis; ophthalmopathy; diabetes mellitus type I; multiple sclerosis; rheumatoid arthritis; gastritis (Helicobacter pylori); Lyme borreliosis; chronic hepatitis C; acute allograft rejection; acute graft-versus-host disease; sarcoidosis; aplastic anemia; routine abortions.
Th2 (II) (Th2-dependent inflammation- exudative, eosinophilic, etc.): measles, Omenn's syndrome, atopic diseases; chronic graft-versus-host disease; allergic keratoconjunctivitis.
Lymphocytes Tγδ and thymus-independent antigens
99% of T lymphocytes undergoing lymphopoiesis in the thymus are Tαβ; less than 1% - Tγδ. The latter are mostly differentiated extrathymically, primarily in the mucous membranes of the gastrointestinal tract. Among all T-lymphocytes of the body, their share is estimated from 10 to 50%. In embryogenesis, Tγδ appears earlier than Tαβ.
Tγδ do not express CD4. The CD8 molecule is expressed on the Tγδ portion, but not as an αβ heterodimer, as on CD8 + Tαβ, but as a homodimer of two α chains.
T functionsγδ: cytokine producers and/or cytotoxic T lymphocytes.
Antigen recognition properties: TCRγδ are more Ig-like than TCRαβ, i.e. capable of binding native Ags regardless of classical MHC molecules - Tγδ does not require or does not require preliminary processing of Ags in APC.
TCR varietyγδ is greater than TCRαβ and Ig, i.e. in general, Tγδ are able to recognize a wide range of Ags (mainly phospholipid Ags of mycobacteria, carbohydrates, heat shock proteins).
Thymus-independent Ag. Substances of a similar chemical nature cannot be processed into complexes with MHC-I/II molecules due to their chemical properties and, therefore, cannot be presented for recognition and recognized by Tαβ lymphocytes. Such substances are called thymus-independent Ags and are divided into two classes.
♦ Thymus-independent Ag 1st class(TH-1) induce polyclonal activation of B lymphocytes and production polyclonal immunoglobulins. These substances are also called B-cell mitogens. The participation of T lymphocytes is not required at all.
The immune response of B-lymphocytes without the participation of T-lymphocytes is characterized by a number of properties: AT only class M (no class switching), no immunological memory, no “maturation” of affinity. But such a response also has an advantage: it develops already in the first 2 days after the penetration of Ag and begins to protect the body in the early stages of infection, while there is no thymus-dependent response yet.
♦ Thymus-independent Ag 2nd class(TN-2): bacterial wall polysaccharides containing many repeating structures. TH-2 (unlike TH-1) is capable of activating only mature B-lymphocytes. In immature B lymphocytes, repetitive antigenic epitopes induce anergy or apoptosis. It is TH-2 that predominantly B 1 lymphocytes “specialize”
Early precursors of T-lymphocytes are formed in the BM. Before other T-cell markers, CD7 is expressed on the surface of developing human T-series cells (already at the proT stage). These cells also carry the membrane marker CD38, which is characteristic of many hematopoietic cells at intermediate stages of development. Their reproduction is supported by stem cell factor and IL-7, the receptors of which are present on the surface of these cells. Cell proliferation can be caused by IL-3, 2, 9, 1 and 6. In the early stages, immature blast progenitors enter the thymus. All stages of differentiation will be associated with changes in surface markers for t-lymphocytes.
T lymphocytes – CD2+ CD3- CD4- CD8-
First, the β-chain is synthesized, then the α-chain. The chains are assembled and αβTCR CD3+ CD4+ CD8+ - cortical thymocytes - emerge. Feels. To apoptosis, cat. induc. corticosteroids and i.i.
From this moment the stages will begin. and deny. selection of T lymphocytes in the thymus. Positive selection is the process of selectively maintaining lymphocyte clones; negative selection is the process of eliminating lymphocyte clones. These processes lead to the correction of the primary antigen recognition complex (maintenance of clones that recognize peptides in the composition of “their” MHC molecules, and the elimination of completely autoreactive clones).
Earlier in time positive selection takes place in the deep layers of the thymus cortex. It is based on the interaction of thymocytes with epithelial cells carrying MHC class II molecules on the surface. At this stage, clones are maintained that are capable of recognizing both completely autologous combinations of MHC molecules and peptides, and autologous MHC molecules modified with foreign peptides. The basis of positive selection is the contact interaction of cells due to the complementarity of the thymocyte receptor and the MHC molecule of the epithelial cell. This interaction involves the already mentioned pairs of adhesive molecules that stabilize the interaction.
After completing the phase, it will be put. selection for more cells of surviving clones, increased expression of CD3-TCR and auxiliary molecules CD4 and 8. Cells that have undergone such changes become a substrate for deny. selection. It occurs in the medulla and cortico-medullary zone of the thymus in the process of interaction with dendritic cells rich in MHC class I and I products. If they recognize a high-affinity property. peptides – auto-AG – are then destroyed by apoptosis.
As a result of 2 phases of selection, those thymocyte clones that carry receptors specific to antigens that have no relation to autologous MHC, as well as to complexes of autologous antigen peptides with autologous MHC, are eliminated.
Subpopulations of T-lymphocytes, main functions. T-helpers, classification, differentiation mechanisms. Role in the development of the immune response Th1, Th2, Th17 and regulatory T lymphocytes.
Lymphocytes, entering the thymus from the BM, differentiate under the influence of thymic hormones into mature lymphocytes. At the same time, they go through different stages of development. There are 2 main subpopulations of T lymphocytes:
– T helper cells αβTCRCD4+- subpopulation =60%. Depending on what cytokines are produced by these lymphocytes during the development of imm. The answer is highlighted: T helper cells type 1– produce γ-interferon, interleukin-2, growth factor β. They activate macrophages and participate in cellular imm. response, take part in inflammation, in HRT reactions; T helper cells type 2 – produce interleukin-4,5,10,21,23. capable of activating B lymphocytes, i.e. are responsible for the development of special humoral imm. reactions, provide protection against helminths, parasites, and other implementation of all allergic areas in the organization; T helper cells 17– produce interleukin-17.36, interleukin-17A, F – vvl. in the development of autoimm. diseases, providing protection against bacteria, the cat has an extracellular reproduction cycle.
All lymphocytes are formed from naïve T-lymphocytes. Differentiation is determined by the local microenvironment and cytokines, cat. affect the diff.
For Th1 – interleukin-12, Th2 – interleukin-4, Th-17 – interleukin-6.23. Tregulate – transform. growth factor β.
– Cytotoxic lymphocytes– killer cells αβTCRCD8+ =30%.
They recognize and destroy foreign or own altered cells. The precursor cells of T killer cells recognize AG in multiple cells in association with class I MHC cells. They secrete perforins, granzymes, TNF, which cause membrane damage and cell death. Tk are capable of synthesizing interferon alpha, which has antiviral activity.
Immunopoiesis: maturation of T - and B cell receptors.The role of microenvironmental factors.
Mechanisms of positive and
negative selection.
Major subpopulations
lymphocytes.
Cycle 1 – immunology.
Lesson No. 3.
Central authorities
immunity - red
bone marrow and thymus.
In central authorities
immunity occurs
first,
antigen independent stage
differentiation
lymphocytes –
that is, "maturation"
unique
monospecific
receptors.
Occurs in the bone marrow
education and
differentiation of all
types of blood cells on
basis
self-sustaining
stem populations
cells,
Blymphocyte differentiation.
Thymus is a "school"
competence of Tlymphocytes", in
thymus gland
pre-T cells migrate
from bone marrow.
CENTRAL ORGANS OF IMMUNITY
Cells becomeimmunocompetent - then
are capable of distinguishing
various foreign molecules
structures.
This ability is inherent in
lymphocyte genome
presence of antigens on
this step is not required.
In central authorities
immunity is formed
cell ability
react in the future (to
periphery) to “alien” by
principle: one lymphocyte -
one antigen.
Central organs of immunity: thymus
thymus THYMUSLobulated structure with epithelial stromal cells and
connective tissue
The stroma provides a microenvironment for the development and selection of T
cells
Outside is the cortex, inside is the medulla, inside are thymocytes (Tlymphocytes that migrated from the bone marrow)
thymocyte
Epithelial
bark
cortex cell
Dendritic
cell
macrophage
Cerebral
layer
Epithelial
th cell
brain
layer
Cells of the cortex and medulla of the thymus
Thymus - biological clock: thymus mass
newborns1 – 5 years
6 – 10 years
11 – 15 years
16 – 20 years
21 – 25 years old
26 – 30 years old
31 – 35 years old
36 – 45 years old
46 – 55 years old
56 – 65 years old
66 – 90 years
15.15 g
25.6 g
29.4 g
29.4 g
26.2 g
21.0 g
19.5 g
20.1 g
19.0 g
17.3 g
14.3 g
14.06
Thymus - biological clock
For older people it is common to:large number of cells
memories (meetings with many
antigens)
reduced number of naive
T cells (thymic aging)
the decline is not only in numbers
naive T cells, but also
diversity of their repertoire
T cell receptors
reduced opportunity
formation of adequate
immune response to earlier
an unfamiliar infection.
Maturation of T-lymphocytes in the thymus: stage 1
Structuralpart of the thymus
Bark
Cells,
providing
T lymphocyte maturation
Functions
Selection
Nurse cages –
Synthesis of “hormones” + selection –
epithelial cells of the thymus - thymulin, are destroyed
thymus.
thymosins,
cells, not
thymopoietins,
In the cortex
capable
provide
thymus is located
bind
early
stages
most
own MNF
differentiated
thymocytes (85-95%)
-antigens, on
ki T lymphocytes output – either
CD4+ cells
(recognize MHC II
class) or CD8+
(MNS I)
Maturation of T-lymphocytes in the thymus: stage 2
Cells,Structures
providing
part
maturation of Tthymus
lymphocytes
Functions
Selection
comrade
Brain Dendritic
layer
cells,
macrophages
Meet CD4+ and
CD8+ cells on
border of cortical and
medulla,
present to them in
complex with MNS –
molecules
autoantigens
"-" selection:
those are destroyed
lymphocytes,
who answer
to autoantigen –
total dies
80-90% T cells
CENTRAL ORGANS OF IMMUNITY: positive and negative selection (selection) of cells
In central authoritiesimmunity occur
clone selection processes
lymphocytes (T-lymphocytes
- in the thymus, B-lymphocytes in the bone marrow).
Biological meaning
selection taking place in
central authorities
immunity - exit to
peripheral blood
functionally mature and
non-autoreactive
lymphocytes.
Selection is ensured
maintaining clones
recognizing peptides
as part of “our own”
main molecules
complex
histocompatibility
(positive
selection), and
eliminating
autoreactive clones
(negative
selection).
Thymocyte selection intensity
T cells mature intothymus,
but many more T cells die in
thymus (do not pass ±
selection).
98% of cells die in
thymus without development
inflammation and inflammation
resizing
thymus.
1 – Hassal’s body,
2 – thymocytes,
3 – apoptotic thymocytes. T cells mature in the thymus,
but many more T cells die
contains
1-2 x 108
cells
2 x 106 per day
98% of cells die in the thymus without developing inflammation and
changes in the size of the thymus.
Thymic macrophages phagocytose apoptotic thymocytes.
Structure of T receptors
The T cell receptor has and -chains (there arealternative receptors that have
and chains - provide immunity to mucous membranes
membranes, primary response to infection).
Each chain of the T receptor has:
- 1 external variable V domain
- 1 external constant C – domain;
- transmembrane segment;
- cytoplasmic tail (short). T cell receptor
Bondage place
AG
for comparison: BCR - Ig Fab fragment
VL
C.L.
V V
VH V
L
VH
CH
CH
CH CH
CH CH
C.L.
Fab
Fc
Domain structure: Ig genes
carbohydrates
monovalency
C C
+
+
Cytoplasmacy
chelic tail
+
Transmembrane
region
No alternative constants
regions
heterodimers, chains linked
disulfide bridges
Very short
cytoplasmic tail
Antigen binding site
formed by V and V regions
30,000 TcRs of one specificity
per cell
What cells emerge from the thymus to the periphery?
As a result of positive andnegative selection in
blood flow only
those T lymphocytes that:
have monospecific
T cell receptor
(TcR);
recognize MHC molecules
Class I (CD 8+Tcytotoxic) or MHC
Class II (CD 4+ T helper cells)
unable to recognize
autoantigens (that is, not
autoreactive T
lymphocytes).
Structure of the T receptor complex (TCR/CD3)
On the cellularsurface -T
cell receptor
(or) located
in the immediate
proximity to
complex,
called CD 3.
Through the CD 3 complex
is happening
signal transmission from
T cell
.
receptor into the cell
Structure of coreceptors (CD 4 or CD8)
Coreceptors (CD 4 orCD8) are located
on membrane T
lymphocyte next to
complex
TCR/ CD3.
Coreceptors “recognize”
MHC molecules
antigen presenter
Surface membrane of T lymphocyte
living cells, and
receptor
recognizes
fragments
antigen.
APC surface membrane Target cell
Antigen presenting cell
Thymocyte maturation process: stages of coreceptor formation
Coreceptors:CD4 – recognizes
MHC II molecules
CD8 – recognizes
MHC I molecules
The cortex contains immature
thymocytes:
double negative
(CD3/TcR CD4 - 8-)
double positive
(CD3/TcR CD4+ 8+)
During the transition to the brain
layer of cells lose either
CD4 or CD8 and
become
single-positive.
In the medulla - mature
single-positive
thymocytes, there are 2 types of them:
(CD3/TcR CD4+) –T –
helpers
(CD3/TcR CD8+) –T –
cytotoxic
This is how they come out
blood flow Transition from double positive T cells to single positive T cells
CD4+ THYMOCYTE
DOUBLE POSITIVE THYMOCYTES
TcR
TcR
√
X
CD8
3
MHC class I
CD4
TcR
TcR
2
CD8
3
CD4
2
MHC class II
MHC class I
MHC class II
Thymic epithelium
Signal from CD4 abolishes CD8 expression and vice versa
T cell receptor gene rearrangement
During the “maturation” of receptorsT lymphocytes in the thymus
α-β- or γ- and δ-chain genes
undergo recombination
DNA (gene rearrangement,
coding T cells
receptors).
In α-β-T lymphocytes, first
β-chain genes are rearranged, then α-chain genes T
cell receptor.
Theoretically
rearrangement
TCR genes
provides 10161018
T options
cellular
receptors;
is this real
diversity
limited in number
Lymphocyte TCR
body up to 109.
Rearrangement of genes encoding the α-chain of the T receptor
Rearrangement of genes encoding the T receptor α-chainThe initial configuration of the genes encoding the chain:
these genes are located on the chromosome in the form of repeating
segments belonging to three classes: V (variable), D
(diversity) and J (joining), as well as one or more
invariant constant regions C (constant).
V
D
J
C
Initial configuration
Tissue cell receptor (TCR) gene rearrangement
DNA recombination occurswhen combining V-, D- and J-segments, catalyzed
recombinase complex.
After rearrangement of VJ in α-chain genes and VDJ in β-chain genes, and
also after joining
non-coding N- and Pnucleotides, with DNA
transcribed by RNA.
Merger with C-segment and
removing unnecessary
(unused) J segments occurs when
splicing of primary
transcript.
Somatic hypermutagenesis
TCR genes are not affected..
Rearrangement of genes encoding T receptor by somatic recombination
Stage 1 – D-J gene fusionStage 2 - V-DJ gene fusion
Stage 3 – chain assembly
V
DJ
C
V
DJ
C
V-DJ fusion
D-J merger
Rearrangement of genes encoding the α-chain of the T receptor, assembly of the T receptor
Rearrangement of genes encoding the T chainreceptor, T receptor assembly
When rearranging genes
-T cell chain
receptor originate
the same steps as for
gene rearrangement
-chains.
Upon completion
gene rearrangements
chains
reading in progress
m RNA, protein construction,
joint assembly and
- chains, expression on
surface membrane
T-receptor
complex.
T cells can already
recognize antigen and
interact with
MHC I and II molecules
classes through
coreceptors - CD4 and
CD8.
After this they begin
processes
negative
selection (im
provide
autoantigens). How does autotolerance develop?
antigens
absent from the thymus?
T cells carrying TcR and entering
interaction with thymus antigens,
are destroyed (negative selection).
But! Some autoantigens are not
expressed in the thymus – i.e. with them thymocyte
will meet for the first time when he comes out on
periphery as a naïve T lymphocyte.
Conclusion: Cellular tolerance must
develop outside the thymus.
Costimulation (immune response); lack of costimulation (anergy, tolerance). immune response).
Presentation processantigens
accompanied by
or not
accompanied by
costimulation:
Agro-industrial complex is being experimented
or not
express
molecules
costimulation,
ligands for
which are
molecules on
surfaces
T cells.
Co-stimulatory molecular interactions on APC and T lymphocyte: CD40-CD 40L and B7-CD 28 complex
Damage and costimulation hypothesesFull expression of T lymphocyte functions depends on when
and where costimulatory molecules are expressed
Cells, contact
only
with autoAH
Cell death by
apoptosis.
Physiological
death.
No anxiety, no threat
agro-industrial complex
No anxiety, no threat
No APC activation, no immune response
agro-industrial complex Threat hypothesis
Cell death
by necrosis
eg damage
tissue, viral
infection
agro-industrial complex
ANXIETY
Pathogens,
recognized
receptors
agro-industrial complex
APCs that have detected threat signals express
costimulatory molecules
activate T cells and the immune response
Survivors as a result
positive and
negative
T cell selection
come out of the thymus
into the bloodstream is
naive T lymphocytes, still
never met
with antigen.
Naïve T cell circulating
by blood and periodically
enters the lymph nodes, where in the T-cell zone it contacts
antigen presenting
cells.
APC, presenting antigens,
"select" T lymphocytes, whose
the receptor is most suitable for
antigen, and give it to those
signals to cells
preferential
survival, activation,
proliferation and
differentiation – for
ensuring adequate
immune response to hypertension
Mature T lymphocytes: life course in the periphery
After encountering an antigen in a lymph nodeT cell using cytokines,
co-stimulatory molecules of APC
acquires the ability
be cloned (in all its descendants -
identical monospecific Treceptors recognizing AG).
Among the descendants of fissile T
lymphocytes appear:
central memory cells (TCM - stem
cell memory T cells),
short-lived effector cells,
carrying out an immune reaction
(SLEC or TEMRA cells),
effector cells precursor memory TEM,
All these cells are coming out
from the lymph node,
are moving
by blood.
Effector cells then
can go out
from the bloodstream for
implementation
immune response
in peripheral tissue
organ where it is located
pathogen (example:
viral infection).
By the time it ends
immune response
majority
effector cells
dies, 5-10%
remain as cells
memory.
.
Effector T cell emigration into tissue during viral infection
Emigration of effector T cells into tissue during viralinfections
Mature T lymphocytes: recirculating and resident
Lymphocytes recirculatethrough lymph and blood flow
in search of an antigen,
which you need
recognize and run
immune response.
Part of T-lymphocytes
is not in the blood
and not in the lymph nodes,
and in the organs
not related
to the immune system -
resident T
tissue lymphocytes,
which are
descendants of effector
T cells that have lost
ability
recycle.
Some peripheral for
immune tissue system,
(small intestinal mucosa,
abdominal cavity, etc.)
allow effector T lymphocytes to enter
free;
Other tissues (central nervous system, mucous membranes)
genital organs, lungs,
epidermis, eyes) practically not
allow T lymphocytes to pass through (not
express homing molecules
–addressins or express in
very small quantities);
a large flow of effector T cells into these tissues is observed
only during an inflammatory reaction. DISCOVERY OF B - CELL IMMUNITY
1954 - Bruce Glick, USA
Studying the function of Fabricius' bursa (bursa Fabricius), lymphoid
organ in the cloaca area of a chicken
Bursectomy in chickens did not result in
to visible effects
Burectomized chickens
used in
experiments to obtain
antibodies to antigens
Salmonella
None of them
bunionectomy
there were no chickens
antibodies detected
against Salmonella
It was found that the bursa is the organ in which
antibody-producing cells - that's why they are called B cells
bursa Fabricius is absent in mammals Origin of B cells and organ in which
B cells mature
It's in the blood
mature B cells
Transfer of labeled cells
fetal liver
Normal bone marrow
Mature
B cells
none
Defective bone marrow
B cells begin to develop in the fetal liver
After birth, their development continues in the bone marrow Bone marrow
S
M
M
E
Stages of B lymphocyte development
1). Stem cell2) General lymphoid
precursor for the B and T cell pathway
development - the most
early lymphoid
a cell for which
one of them was not determined
two directions
development;
3a) Early pro-B cell immediate descendant
previous cellular
type and predecessor
subsequent
advanced in
differentiation
cell types
(prefix "pro" from English.
progenitor);
3b) Late pro-B cell
4) pre-B cell - cell type,
finally released to B-cell
path of development (prefix "pre" from English.
precursor);
5) immature B cell - final
bone marrow development cellular
form that actively expresses B
receptor - surface immunoglobulin
and is in the selection stage for
ability to interact with
own antigens;
6) mature B cell - cell type
periphery, capable
interact only with aliens
antigens;
7) Plasma cell (plasmocyte)
effector, antibody-producing
cellular form that is formed from
mature B cell after its contact with
antigen and synthesizes antibodies
(immunoglobulins) Stages of B cell development
Early pro - B
Late pro - B
Stem cell
Big pre
-IN
peripheral
Small pre-V
Immature B
mature in cage
Gene rearrangement occurs at each stage of development
Ig heavy and light chains, surface Ig expression, expression
adhesion molecules and receptors for cytokines
Stages of B cell development
Early stages of developmentB lymphocytes
depend on direct
contact
interaction with
stroma.
As a result of these
contacts
is happening
proliferation of Blymphocytes and
their transition to
next stage
development – late
pro-B cells.
At later stages
development of B lymphocytes
requires humoral
bone stromal factors
brain – cytokines (IL-7)
Expressed on the surface of late pro-B cells
receptor for IL-7.
Under the influence of IL-7, pro-B lymphocytes proliferate and
differentiate into early
pre-B cells,
characterized by the presence in
their cytoplasm of the heavy μ (mu) polypeptide chain
immunoglobulin M.
Stages of B cell development
Subsequently, early pre-B cells are transformed intosmall pre-B lymphocytes,
some of whom have
cytoplasm in addition to μ
-heavy polypeptide
chains are revealed molecules
immunoglobulin
appear
light chains
immunoglobulin (or
kappa, or lambda), further
expression occurs on
surface membrane
monomeric
immunoglobulins M.
. Immunoglobulins M and
are antigen-recognizing
B cell receptors.
Antigen specificity
receptors genetically
determined.
Next, expression occurs on
immunoglobulin cells
class D (IgD).
With expression on lymphocytes
immunoglobulin D
The stage of antigen-independent maturation of B cells is completed.
Development of B cells in the bone marrow
More than 75% of those maturing in the bonebrain B cells do not enter
blood flow, but dies by
apoptosis and is absorbed
bone marrow macrophages.
+ selection occurs when
interaction between B cells and cells
stroma - B cells remain with
immunoglobulins (Ig).
- selection occurs when
interaction between B cells and
antigen-presenting
cells (APC) presenting
autoantigen fragments
Surviving cells
continue to mature
and reach
central
venous sinus.
At all stages
B cell maturation
plays an important role
B cell connection with
stromal cells
(microenvironment) and
presence of cytokines
- in particular,
interleukin-7. Development of B cells in the bone marrow: “maturation”
monospecific receptors, “+” and “-” selection
B
B
B
B
Regulation of B receptor maturation
Each B cell is monospecific
Destruction of autoreactive B cells
The release of full-fledged, but still immature
In cells to the periphery (first in
spleen, then to the lymphatic
nodes)
Bone marrow provides
MICROENVIRONMENT FOR MATURATION,
DIFFERENTIATION AND DEVELOPMENT OF CELLS Maturing B cells
Stroma cells B
B
Stroma cell Scheme of development of B cells in the bone marrow
predecessors
E
n
d
O
With
X
X
X
T
TO
ABOUT
WITH
T
N
ABOUT
Y
P
L
A
WITH
T
AND
N
TO
Immature and mature
Into the cells
Pre-B
stromal cells
macrophage
Central sinus B - cellular autotolerance - exit of mature
In cells from bone marrow
B
IgD
IgM
Small pre-B does not carry
receptors
Immature B lymphocyte
does not recognize auto Ag
YY
B
YY
Immature
IN
YY
B
YY
YY
Small
pre-B
IgD and IgM receptors
IgD
IgM
IgM
IgD
IgM
IgD
Mature B lymphocyte
comes out
to the periphery
Postulates of the theory of clonal selection
Each B lymphocyte hasunique receptor
specificity.
High affinity (durable)
receptor interaction
with antigen leads to
activation
B - lymphocyte.
Specificity
receptor is stored in
proliferation process
and differentiation
lymphocyte.
Lymphocytes with
receptors,
specific to
own
antigens (potentially
auto-aggressive),
are removed early
stages
differentiation.
Immunoglobulin (Ig) molecule genes
Each Ig molecule consistsof 2 heavy (H) and two
light (L) chains, in
each of these circuits
present
constant (C) and
variable (V)
areas.
Variable (V) and
constant (C) regions
immunoglobulin
molecules are encoded
separate genes.
For variable regions
there are many
genes (V1-Vn), and for
constant part
Ig molecules – one C-gene.
Light chains
immunoglobulins
encoded by genes
segments V and J.
Heavy chains are encoded
segments V and J, as well as
additional
segment (D). Stages of differentiation
determined by rearrangement of Ig genes
Stages
maturation
configuration
genes
IgH
Stem
cell
Early
pro-B
From
DH to
JH
Late
pro-B
From VH to DHJH
Big
pre-B
VHDHJH
Pre-B cage
express
smiles
receptor
Ig light chain genes have not yet been rearranged
Rearrangement of genes encoding the light chains of the Ig molecule
After the completion of perestroika(rearrangements) of genes encoding
heavy chains of the Ig molecule, begins
rearrangement of light chain genes.
There are 2 types of light chains - either
kappa or lambda.
After this, on the surface of immature B
lymphocyte appears B - cell
receptor consisting of two heavy
chains (H) and two light chains (L).
Positive and negative selection of B lymphocytes in the bone marrow
+ selection occurs through the interaction of B cells and stromal cells - B cells remain withproductive gene rearrangement
immunoglobulins (Ig), the rest -
are destroyed by apoptosis.
- selection – destruction of autoreactive Blymphocytes can also occur in the bone
brain, and in the spleen - the organ into which
the majority of newly formed B migrate
-cells during intrauterine development.
Genes of Ig molecules
Before meeting the antigen:Recombinations
limited number
gene segments
V, D and J
create infinity
number
monospecific
In receptors (there are many of them
more than antigens)
After encountering an antigen:
After antigenic stimulation -
during an immune response
for antigen
in the genes of light and heavy
chains of molecules
immunoglobulins in
proliferating B
lymphocytes occur
spot somatic
mutations
(fine “fit” of AT
to AG).
Further stages of B-lymphocyte development
Already selected B lymphocytes from the bone marrowenter the primary follicles with the bloodstream
spleen.
A number of functional stages occur in the spleen
"maturation" of B lymphocytes, including expression
MHC class II on their surface membrane.
Next B lymphocytes migrate to the lymph nodes
– to meet with a complementary receptor
antigen.
Before meeting the antigen, the B lymphocyte is called
"naive".
Release of mature B cells to the periphery
Only those B lymphocytes leave the bone marrowwho had a successful rearrangement
genes of the heavy and light chains of Ig molecules, and
these B lymphocytes are not activated in response to
autoantigens – that is, they are not
autoreactive.
All other cells die in the bone marrow
by apoptosis.
On the surface of selected B lymphocytes
IgM and IgD–Ig receptors are expressed,
which are synthesized from one DNA by
alternative splicing. Recirculating B cells meet “their own”
antigen in the lymph node
Vessels leave the cells
and enter the LU through VEV
Into the cells quickly
proliferate
Ag enters the lymph node through
afferent
YY
Y
Y
YYY
YY
Y
YYY
Y
Y
Germinal center
Intense proliferation
From the germinal center
come out into the cells,
which differences
were in plasmatic
which cells
YY
Y YY
Y
B
YY
B
YY
YY
Mature peripheral Recognized
B lymphocyte
non-auto-AG
on the periphery
B
Y
Y
Y
YY
YY
YY
YY
Y
YY
Differentiation of B cells at the periphery
Ig - secreting
plasma cell
Pattern of B-lymphocyte response to antigen
Subpopulations of B lymphocytes: B1 and B2
B 2 (CD 5-) lymphocytes bind protein antigens,they need the help of T-helpers, they synthesize
immunoglobulins of different classes in the process
adaptive humoral immune response.
B 1 (CD 5+) lymphocyte population responds to
bacterial capsule polysaccharides or their components
walls (such antigens are called T -
independent), when answering T- independent
B-lymphocyte antigens do not need helper help.
Since most antigens are protein in nature,
the population of B 2 lymphocytes is much larger
numerous compared to B1.
Subpopulation of B1 lymphocytes
After activation B1cells secrete
anti-polysaccharide
class M antibodies
(IgM), which
join
surfaces
bacterial cell.
Recognize epitopes of antigens with
repetitive
structures of phosphotidylcholine,
lipopolysaccharides and
etc.
Next comes activation
complement systems and
fast complement -
dependent lysis
bacterial cell.
No immunological memory; No
higher efficiency
response upon repeated administration
antigen.
In 1 cells produce
immunoglobulins only
class M. For this they do not
the help of T lymphocytes - helpers is required. Interaction of APC, T- and B-lymphocytes during the immune response to hypertension
Questions for lesson No. 3a
1.2.
3.
4.
5.
6.
7.
8.
9.
10.
What is the role of the thymus in the process of T lymphocyte differentiation?
What is the biological meaning of positive and negative
selection?
What are the mechanisms of formation of T-cell diversity?
receptors?
Describe the structure of the T-cell receptor.
List the main subpopulations of T lymphocytes.
Describe the structure of the B-cell receptor.
Name the subpopulations of B lymphocytes.
Describe the stages of antigen-independent differentiation of Blymphocytes.
Describe the process of antigen-dependent differentiation of Blymphocytes.
Which cells are the final stage of Blymphocyte development?
11. Antigen-independent differentiation of T lymphocytes
happens in:
thyroid gland
thymus
lymph nodes
spleen
hypothalamus
2. The specificity of the T-cell receptor is based on
stages:
double negative cell
double positive cell
single positive cell
after the T-lymphocyte leaves the thymus
when interacting with a macrophage
Test tasks for lesson No. 3
3. A common marker of T lymphocytes is the molecule:CD 3
CD 4
CD 8
CD 16
CD 34
4. Markers are characteristic of helper T-lymphocytes:
CD 3; CD 4
CD 3; CD 8
CD 4; CD 8
CD 16; CD 56
CD 4; CD 117
Test tasks for lesson No. 3
5. Markers characteristic of cytotoxic T-lymphocytes are:CD 3; CD 4
CD 3; CD 8
CD 4; CD 8
CD 16; CD 56
CD 4; CD 117
6. Antigen-independent differentiation of B lymphocytes
happens in:
thyroid gland
thymus
lymph nodes
spleen
red bone marrow
7. Germline genes for immunoglobulin heavy chains include
regions:
B
D.P.
V
D
J
8. The main markers of B lymphocytes are:
CD 3
CD 21
CD 19
CD 34
CD 4
Test tasks for lesson No. 3
9. On the surface of mature B-lymphocytes they are present as B-receptors:Ig E
IgM
IgG
IgD
IgA
10. The final stage of antigen-dependent differentiation of B lymphocytes is:
Natural killer cells
Macrophages
T lymphocytes
Plasma cells
B1 lymphocytes
1) Signaling proteins included in the BCR and TCR complexes. ITAM and ITIM motifs, SH2 domain.
2) BCR and TCR coreceptors.
3) The main receptors for costimulation signals on B and T cells.
| Receptor | BCR | TCR |
| Signaling proteins included in the complexes (they are needed to transmit a signal inside the cell, since the receptors themselves do not have intracellular parts) | Igα, Igβ (there are signaling areas on the intracellular parts - ITAM) | CD3 (have 2 subunits: gamma or delta and epsilon), there is also a separate zeta - without an extracellular part. There are signaling areas on the intracellular parts - ITAM |
| Coreceptors | CD21, CD19, CD18 - recognize the result of complement work (confirmation for B cells that there is a pathogen) | CD4, CD8 - nonspecifically recognize antigen (recognize MHC, not peptide; can distinguish between MHCI and MHCII) |
| Major receptors for costimulation signals | CD40 (binds to CD40L ligand, which is expressed by the T cell) | CD28 (interacts with the B7 complex on the surface of the antigen-presenting cell) |
FcR - receptors of constant parts of immunoglobulins
About the localization of ITIM we can say that NK cells have
SH2 domain (Src-homologous) is a structure that provides very affinity binding to phosphorylated tyrosine, serine or threonine in ITAM and ITIM. Without phosphate there is no interaction!
4) Immunological synapse. device and biological role
Immunological synapse - interaction of a T cell with an antigen presenting cell (APC). In this case, the T-lymphocyte tightly adheres to the target cell, and cytokines are injected into the cavity to act locally. The synapse also involves adhesion and costimulation molecules, receptors and coreceptors.
5) The role of lck kinase and CD45 phosphatase in signal transmission from lymphocyte antigen receptors.
Upon initiation of a signal from the TCR, CD45 phosphatase removes inhibitory phosphate from Lck kinase. Before this, the Lck kinase is maintained in an inactive state by the Csk kinase. In the inactive state, Lck kinase is curled up: its SH2 domain binds to its C-terminal phosphorylated tyrosine). After removal of the inhibitory phosphate, Lck kinase changes conformation, autoactivates, and phosphorylates ITAM. ITAM with two phosphorylated tyrosines is a substrate for Src kinases with two SH2 domains (for example, ZAP-70 kinase).
6) Major physiological consequences of TCR activation.
The main physiological consequences of TCR activation are that various signaling molecules (LAT and SLP-76) are phosphorylated, leading to:
● activation of transcription factors (via PLCγ)
● increasing cellular metabolic activity (via Act)
● actin polymerization and cytoskeleton reorganization (via Vav)
● enhancing the adhesive properties of the surface: enhancing “stickiness” and clustering of integrins (via ADAP)
7) Intracellular calcium and phospholipids in lymphocyte activation. NFAT, NFkB and AP1 families of transcription factors.
Activation of transcription factors occurs through PLCγ - phospholipase gamma. It breaks down PIP2 (phosphatidylinositol diphosphate) into IP3 (inositol triphosphate) and DAG (diacylglycerol).
IP3 opens calcium channels in the ER and outer membrane. DAG remains on the membrane and attracts PKC-θ and RasGRP (RAS guanyl-releasing protein), which trigger a MAP kinase cascade in which transcription factors of 3 families are activated (these are the most important):
● NFAT (via calcium and calcineurin); one of the molecules of this family is the target of cyclosporine A, a substance that can selectively suppress the T-cell response (important in organ transplantation)
● NFkB (via PKC-θ and CARMA)
● AP1 (via RasGRP, RAS and MAP kinase cascades)
8) Role of phosphatidylinositol 3-kinase (PI3K) in costimulation. Mechanism of action of CTLA-4.
The costimulation molecule (B7) appears in response to the innate immune response. It is recognized by the CD28 molecule. Through sites with tyrosine, phosphatidylinositol 3-kinase (PI3K) is recruited and activated. The result of its work is PIP3. PLCγ is activated only when it is recruited to the membrane and binds to membrane-anchored PIP3 there.
CTLA-4 is a CD28 antagonist. It competes with CD28 for binding to B7 and always wins because it binds more efficiently due to the fact that it does so in clusters. That is, in the presence of CTLA-4, CD28 does not bind to B7. CTLA-4 has inhibitory signals within ITIM.
9) The role of IL-2 and IL-2 receptor alpha chain in the activation of cytotoxic T cells.
To activate a killer T cell, simultaneous physical interaction with the same dendritic cell with which the helper T cell interacts is required. When a T-helper interacts with a DC, a co-stimulation molecule for the T-killer is released in the DC (you don’t need to remember it, but you can’t forget it - it’s called “four IBBL” 4-IBBL).
Strong positive feedback through IL-2 and CD25 (the alpha chain of the high-affinity IL-2 receptor on the surface of killer T cells) is very important. First, IL-2 is secreted by the helper T cell, and then IL-2 causes the production of IL-2 in the killer T cell itself. This is how the T-killer clone adjusts itself in growth during proliferation.
IL-2 is an example of the integration of signaling pathways in its gene promoter. Its promoter has binding sites for all transcription factors from question 3, that is, for its transcription to proceed well, both a signal from the TCR and a costimulation signal from CD28 are required.
10) Ligands for activating and inhibitory NK cell receptors.
Inhibitory receptors bind to a set of MHCI molecules. This is important when the NK cell learns to recognize its own - it will be activated by the absence of these ligands = “lack of self.”
Activating receptors bind to stress-induced ligands. The NK cell is activated in response to increased expression of stress proteins (“stress-induced self”).
11) The logic of lymphocyte development in primary lymphoid organs. Positive and negative selection.
Proliferation - expression of the first chain Pre-B or Pre-T - proliferation again - expression of the second chain (now there are complete antigen receptors) - selection (the future lymphocyte is eliminated if it binds very strongly to the antigen or does not bind at all).
Selection in the bone marrow: maturation of B cells expressing two immunoglobulins - IgM and IgD + selection of cells that respond only to foreign antigens.
B cells, whose immunoglobulin receptors are capable of interacting with self-antigens, either die as a result of apoptosis or become unresponsive (anergic).
Selection in the thymus: T cells express either CD4 (to be helpers) or CD8 (to be killers). The actual selection is based on the TCR signal: if it is absent or it is strong, the cell dies. If weak, there will be a naive T-cell, if moderate, there will be a T-regulatory cell.
12) Methods for removing potentially autoreactive clones from a population of mature lymphocytes. Editing of lymphocyte antigen receptors.
This was the example of B lymphocytes.
If a B lymphocyte contacts a dissolved autoantigen, then an anergic B cell is formed with a high level of IgD expression (induction of anergy without costimulation - a mechanism of immunological tolerance).
If the B lymphocyte has bound to a surface-bound autoantigen (on red blood cells, MHC), then it is possible to edit the receptor to change specificity (by continuing RAG expression, rearranging light chain genes).
If editing fails, apoptosis occurs.
13) Primary and secondary immune response. Switching from IgM to IgG, memory cells.
Primary immune response - lymphocyte receptors are represented by IgM, low specificity of binding to elements of the pathogenic architecture, IgM is assembled into pentamers (the first meeting with the pathogen, the response takes a long time to develop). Next - the production of highly specific IgG (“selection” - see vdj-recombination), lymphocytes throw out the gene for the constant part of IgM
Secondary immune response - a repeated encounter with the pathogen, there are already lymphocytes with the desired antigen, the response develops very quickly
Memory cells are lymphocytes that form the secondary immune response
14) Features of the physiology of antigen receptors B-1 and gamma delta T cells, allowing them to be classified as “innate lymphocytes”.
cell population B1- they develop in newborn liver. There they are produce IgM(when activated - soluble IgM). They have a limited repertoire of specificity, using only some V genes. Initially, they are tuned to frequently occurring glycans on the surface of bacteria, to some self-antigens (regulation, suppressor cytokines), to self-stress antigens (expressed in case of damage). B1 cells do not form immunological memory. Every time is like the first time. IgM is always in the same amount.
B1 cells add few nucleotides at the junctions of segments, and their repertoire of V genes is limited. Found in the peritoneal and pleural cavities. Unlike B2 cells, which live and die and are renewed in the BM, B1 cells, having settled in their locations, self-renew on the spot. They are able to produce IgM spontaneously at high levels and often have specificity for hydrocarbons. They do not form memories.
Gamma delta T cells can interact with MHCIb, but not with classical MHC molecules, and do not require antigen processing for recognition.
For most gamma delta T cells, the ligands are not precisely known.
There are quite a lot of gamma delta cells in the same place as B1 cells. They are necessary for the recognition of small phospholipids. There are in the peritoneal cavity, there are many of them in the epidermis of the skin, and some of them (which are in particular specific to proteins of the butyrophilin family) play the role of recognizing stress. Butyrophilins are expressed in the periphery as a result of stress. While there is no ligand, immunosuppressive cytokines are expressed. And when a sufficient amount of ligand appears, the immune response is triggered. They do not form immunological memory; they belong to the cells of the innate immune system.
15) Positive selection of CD4+ and CD8+ T cells in the thymus. MHC restriction.
In the cortical zone of the thymus, positive selection of T cells occurs - the removal of cells that have a very weak signal from the TCR.
Double positive thymocytes (CD4+CD8+) are subject to positive selection. Double positive cells are very sensitive to apoptosis (due to their low expression of antiapoptotic factors, such as Bcl-2 and Bcl-XL). Cells require microenvironmental support to survive. Thymocytes receive the signal necessary for survival through positive selection. Positive selection ensures that only those thymocytes that express TCRs with an affinity for MHC molecules are selected. If the TCR has an affinity for the MHC molecule, the thymocyte receives a supporting signal, the main results of which are an increase in the expression of the anti-apoptotic factor Bcl-2 and the advancement of the thymocyte through the cell cycle. An external sign of successful positive selection by thymocytes is the expression of the cell activation marker CD69, as well as the molecules CD5, CD27 and the costimulatory molecule CD28, accompanied by an increase in the density of expression of the TCR-CD3 receptor complex on the cell surface. T lymphocytes whose receptors lack affinity for MHC undergo apoptosis “by default.”
MHC restriction is the recognition of antigen fragments by T lymphocytes only in the context of MHC (with the exception of superantigens).
MNS are individual. T lymphocytes undergo positive selection on MHC molecules, which are different for each individual, and peptides that are able to bind to these alleles. Another individual will have a different set of MHC molecules, the resulting peptides will also be different, and as a result, the repertoire of specificities of mature lymphocytes will also be different.
Thymocyte development is dependent on MHC. If we implant receptors on MHC1 in a mouse, then at the output we will see only cytotoxic T cells. Developing thymocytes will in principle have no chance to express the receptor on MHC2. The same thing will happen if we add a receptor to MHC2 to a mouse.
16) Negative selection of thymocytes. Aire and Fas/FasL.
How do developing thymocytes manage to display all their own antigens, which are dangerous from the point of view of autoreactivity (this is negative selection)?
It was not clear until the gene was discovered AIRE is a transcription factor under the control of about 100 specific proteins, which it triggers from time to time in some cells of the thymic epithelium. Induces the expression of organ- or tissue-specific genes in the thymus:
Insulin (pancreas)
Interphotoreceptor retinoid-binding protein (IRBP) (eye)
Odorant binding protein 1a (tear glands)
Vomeromodulin (lungs)
If you run a small part for a while, nothing bad will happen, but in the end we will show everything that is needed. To put a gene under the control of AIRE, or remove it, you just need to change a couple of nucleotides in the promoter region of the gene (easy regulation).
This is one of the mechanisms of central tolerance. Very easily customizable - you just need to change a few nucleotides in the promoter for regulation.
Disturbances in AIRE function cause autoimmune polyglandular syndrome type 1, or chronic mucocutaneous candidiasis (autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy, APECED) - a rare hereditary disease characterized by an autoimmune response to the tissues of various organs, mainly the edocrine glands.
AIRE-dependent expression of tissue-specific antigens in thymic epithelial cells is one of the mechanisms central tolerance.
If the thymocyte is autoreactive, then sooner or later during maturation it will definitely encounter its peptide and will be destroyed by selection.
FasL and Fas ensure apoptosis (if the lymphocyte receptor has not been rearranged). This is a cytokine and its receptor that provide structural (instructional?) apoptosis. Knockout mice for these genes - hypertolerance of lymphocytes, autoimmune action.
17) The role of cytokines in the differentiation of T helper cells in the periphery.
18) Negative consequences of unbalanced differentiation of T helper cells.
