BMS150_PHL3-08_ToleranceV2_Win2023 (1).pdf

Immunology T-cell Development and Tolerance BMS 150 Week 4 e-learning Lymphocyte development – bone marrow https://upload.wikimedia.org/wikipedia/commons/1/1f/Hematopoiesis_%28human%29_diagram_en.svg T-cell Development - Overview After stem cells...

Immunology T-cell Development and Tolerance BMS 150 Week 4 e-learning Lymphocyte development – bone marrow https://upload.wikimedia.org/wikipedia/commons/1/1f/Hematopoiesis_%28human%29_diagram_en.svg T-cell Development - Overview After stem cells “choose” to become lymphoid progenitors, those progenitors divide under the influence of IL-7 ▪ FYI – stem cells = hematopoietic stem cells (HSC) and multi- potential progenitor cells (MPP) Unlike B-cells, T-cells undergo very little maturation in the bone marrow ▪ Need to travel to the thymus to develop functional T-cell receptors (TCRs) T-cell Development - Overview After stem cells “choose” to become lymphoid progenitors, those progenitors divide under the influence of IL-7 ▪ FYI – stem cells = hematopoietic stem cells (HSC) and multi- potential progenitor cells (MPP) Unlike B-cells, T-cells undergo very little maturation in the bone marrow ▪ Need to travel to the thymus to develop functional T-cell receptors (TCRs)  The Thymus Located superior to the heart, anterior to the great vessels ▪ Largest prior to puberty, shrinks afterwards ▪ Divided into a cortex and medulla, surrounded by a thin capsule Medullary and cortical thymic epithelial cells are key in guiding T-cell development ▪ mTEC and cTEC, respectively Thymus – key organ in T-lymphocyte development Within the thymus, precursor T-lymphocytes: ▪ Become terminally differentiated to the T-lymphocyte lineage ▪ Re-arrange the TCR ▪ “Learn” to only recognize antigen presented via HLA (HLA restriction) T-cells with TCRs that can bind to antigen presented via HLA WILL survive → positive selection ▪ “Learn” to ignore self-antigens presented via HLA T-cells with TCRs that bind self-antigen with high affinity WON’T survive → negative selection ▪ Are selected to express either CD4 or CD8  T-cell Development – An Overview 1. T-cell precursors from the bone marrow enter the thymus from vessels at the junction of the cortex and medulla 2. ▪ They don’t express CD8 or CD4 → double negative (DN) 2. DN cells commit to a T-cell lineage (known as DN3 cells) ▪ These cells are mostly found in the cortex of the thymus ▪ Proliferation begins, and 1. cells begin to genetically shuffle their TCRs  T-cell Development – An Overview 3. DN cells first rearrange the beta chain of the TCR ▪ If this is done correctly, then proliferation continues (DN4) ▪ If the beta chain is useless, 3. the cell dies 4. The alpha chain of the 4. TCR is then shuffled ▪ After both TCR chains have been recombined, the developing T-cell expresses both CD4 AND CD8 → double positive (DP)  T-cell Development – An Overview 5. DP cells with recombined TCRs sample thymic epithelial cells HLA (with self-antigens expressed) ▪ If very low affinity interactions → death of the DP T-cell ▪ If medium-low affinity → 5. survival of the DP T-cell ▪ If high affinity interactions → death of the DP T-cell T-cell Development - Selection Positive selection: ▪ When a DP T-cell that has arranged its TCR has a “low- medium” affinity for a thymic epithelial cell’s (TEC) HLA bound to self antigen, it is allowed to survive Rationale – the TCR is capable of interacting with an HLA molecule, but is unlikely to recognize self If no interaction occurs, the TCR has not been arranged in a productive fashion Negative selection: ▪ When a DP T-cell that has arranged its TCR has a “high” affinity for a TEC HLA bound to self antigen, it is killed Rationale – this is T cell with a TCR that recognizes self – it should not be allowed to survive Exception – some T cells that recognize self are allowed to exist as regulatory T-cells TCR affinity, survival, and selection  T-cell Development – An Overview 6. DP cells that have been positively selected and have survived negative selection then “make a choice” ▪ The DP cell with its useful TCR will “test” to see if the CD8 or CD4 co-receptor interacts better with the medullary TECs ▪ After this interaction, T cells 6. express only CD4 or CD8 T-cell Development – Interesting Facts 95% of developing T-cells do not survive positive and negative selection Events in the cortex of the thymus: ▪ T-cell commitment ▪ TCR recombination and testing ▪ Positive selection & first round of negative selection Events in the medulla of the thymus: ▪ Second round of negative selection (likely) ▪ “Choice” of CD8+ or CD4+ ▪ Exit into the bloodstream Peripheral vs. Central Tolerance Central tolerance? This happens in primary lymphoid organs with negative selection Can also consider it including thymus- generated Treg cells ▪ Most of these are CD4+ Peripheral tolerance? Treg polarization in SLOs or peripheral tissue Central tolerance Central role of the AIRE gene ▪ AIRE is a transcription factor-like protein that binds with a wide range of other transcription factors and activates them ▪ This causes thymic epithelial cells to express a very wide range of proteins on their HLA ▪ Therefore, TECs tend to express a lot of self-antigens Important if you want to generate thymic Treg cells or delete T- cells that recognize self (negative selection) Defects in AIRE are rare – they result in an autoimmune polyendocrinopathy syndrome (APS) ▪ Immune system attacks a wide range of endocrine glands, including the parathyroid glands, the adrenal glands, the pancreas, and the ovaries Peripheral Tolerance CD4+ Treg cells: Polarized by interaction with an APC and presence of high circulating levels of TGF- Remember – TGF- tends to be expressed in higher concentrations by APCs and other cells in “non- inflammatory” environments ▪ If it’s non-inflammatory, does the APC express CD80/86? Does a Treg need costimulation? Good question – this is not entirely clear See next slide – the answer may be low levels of CD80/86 → Treg costimulation ▪ Other stimuli likely play a role, i.e. retinoic acid (Vitamin A) Peripheral tolerance – T-lymphocytes CTLA-4 – expressed by helper T-cells, binds to B7 with higher affinity than CD28 ▪ Binding of CTLA-4 to B7 (CD80/CD86) molecules on APCs usually renders the CD4+ cell anergic however in some experimental models it can cause the CD4+ cell to express FoxP3 → Treg polarization Maybe this is what happens with more TGF-β, and this explains Treg polarization ▪ If a lot of B7 is present on the APC, then CD28 gets activated ▪ Therefore, if an effector CD4+ cell recognizes an antigen but there is a low-level of co-stimulatory molecules, then that effector CD4+ cell is shut down As well, T-reg cells often express CTLA-4 and can shut down CD80/86 expression on the APC ▪ Net result – if you knock out CTLA-4 in a mouse, it always gets very severe autoimmune disease Peripheral Tolerance CD4+ Treg cells: express high levels of the IL-2 receptor as well as FoxP3 Also express high levels of CTLA-4 after polarization CTLA-4 binds to CD80/86 → ▪ Downregulation of APC presenting antigen Reduction in pro-inflammatory cytokines Production of kynurenic acid (anti-inflammatory) FIGURE 16-2 CTLA-4 mediated inhibition of APCs by T REG cells. T REG can inhibit APCs by signaling through the CD80/CD86 (B7) receptor. In the APC, this engagement results in decreased expression of CD80/86 activation of indoleamine-2,3- dioxygenase (IDO; an enzyme that converts tryptophan to kynurenin) creating an immunoinhibitory microenvironment) changes in transcription leading to decreased expression of IL-6 and TNF- alpha Treg cells also express high levels of IL-2 receptor → “soaking up” IL-2 and preventing it from activating an effector cell IgA and peripheral tolerance The mucosal linings are exposed to antigens all the time → commensal bacteria that actually protect us ▪ How do we prevent an unnecessary attack on these commensals? This is a rich area of research, however a few generalities are known: ▪ Early in life we tend to develop tolerance to our flora at the mucosal barrier ▪ Secreted IgA tends to be recognize these bugs, but doesn’t kill them – it reduces their invasion ▪ The Th cells responsible for stimulating this IgA production do not tend to stimulate inflammation A general model of barrier tolerance Barrier tolerance When the barrier is healthy, with no tissue damage, then TGF-beta predominates in the mucosa ▪ TGF-beta and retinoic acid promotes class-switching to IgA by B-cells ▪ IgA in these circumstances will reduce invasion of microbes across the barrier ▪ In the presence of TGF-beta, there is limited or no inflammatory response to these relatively normal microbes, even if bound to IgA If a pathogenic microbe invades → damage and activation of different PRRs: ▪ Reduction of TGF-beta → development of other Th types that are specialized for fighting the invader (Th1, Th2, etc.) ▪ Increased inflammation, and a shift towards Th17 responses with increased innate cell and anti-microbial protein activity ▪ Likely production of antibodies that have undergone somatic hypermutation and are more specific for the invader “Tolerogenic” Situations Certain situations seem to increase the likelihood of developing Tregs and overall tolerance: ▪ Exposure to constant antigen levels for long periods of time ▪ Exposure to antigen early in life It is likely that young children are more likely to develop a more “tolerogenic” response (Tregs vs other Th types) to antigens as long as few damage signals are present ▪ Exposure to antigen in tissue that “looks” healthy More likely to develop Tregs Less likely for APCs to express costimulators Key Genes Implicated in Decreased Tolerance and Autoimmunity IL-2R-alpha: ▪ The alpha-receptor of the IL-2 receptor ▪ This is key for development of Tregs as well as “soaking up” IL-2 to prevent it from activating other lymphocytes ▪ Diseases associated: Multiple Sclerosis, Type 1 DM CTLA4: ▪ The “Treg” membrane receptor that binds to CD80/86 ▪ Diseases associated: Type I DM, Rheumatoid Arthritis  Central Tolerance in B-cells - review Pre-B-cell to immature B-cell The light chain is rearranged – at the end of successful light chain rearrangement (2nd checkpoint), the B-cell has a mature BCR, expressed at the cell surface as IgM These cells are “checked” for self-reactivity – if they are self-reactive, they rearrange their light chain (but only their light chain) to see if the receptor loses self-reactivity ▪ If self-reactivity is still present, usually clonal deletion occurs via apoptosis (central tolerance)  Central Tolerance in B-cells - review Transitional, immature B-cell to mature B-cell ▪ the immature B-cell leaves the bone marrow and (usually) heads to the spleen, where it starts expressing more and more IgD in relation to IgM ▪ B-cells that travel to the spleen to mature further are known as T1 – transitional-1 – Bcells The Spleen Big ovoid secondary lymphoid organ ▪ Immunological roles immune responses to antigens in the bloodstream ▪ No lymphatics ▪ Major effector – antibodies Maturation of T1 B cells (not quite mature) to T2 B- cells ▪ T2 B-cells go through another round of negative selection ▪ After this, they’re ready to travel through the body and become plasma cells Lymphocytes traffic frequently through the spleen – more than through all lymph nodes combined The Spleen Structures: ▪ White pulp – surrounds the arterioles “inner layer” – peri-arteriolar sheath, site containing many T-cells “middle layer” – B-cell-rich lymphoid follicles “outer layer” – marginal zone – specialized B-cells and macrophages here ▪ Red pulp – peripheral to the white pulp Main role of the red pulp is elimination of dysfunctional RBCs Mostly RBCs and macrophages, some lymphocytes ▪ Capsule Spleen –negative selection for B-cells Positive and negative selection for B-cells happens first in the bone marrow, and then is repeated in the spleen ▪ In the bone marrow, B-cells that encounter self-antigen have their receptor shuffled ▪ This doesn’t happen in the spleen – if they encounter self- antigen they undergo apoptosis Unsure how dendritic cells in the spleen know the difference between self and non-self T2 B-cells are cells that have survived negative selection ▪ T2 → mature B-cell progression is a key role of the spleen Mature B-cells are able to develop into plasma cells, can migrate, enter follicles, and produce specific antibodies in other SLOs (or they can stay in the spleen) Immunology of the spleen Circulating B-cells encounter antigens in follicles and PALS, T-cells encounter it in the PALS ▪ Lots of dendritic cells in PALS, access to B-cells and T- cells ▪ CD4+ T-cells will help B-cells, B-cells migrate to follicles → similar to germinal zone of lymph node ▪ CD4+ T-cells can also help activate CD8+ T-cells in PALS Immunology of the spleen There is evidence that the marginal zone is a bit different ▪ many B-cells here that produce somewhat non-specific, somewhat low-affinity IgM that recognize a number of different microbes ▪ These B-cells are called T cell-independent B-cells (TI) Don’t seem to require help from Th Although their antibodies are less diverse, these B-cells can produce large amounts of their antibody when exposed to antigen ▪ Likely most important in childhood ▪ Better characterized in mice, still poorly-understood in humans Immunology of the spleen How do you activate a B-cell without T-cell help? ▪ Antibody binds antigen AND either: C3d co-receptor activation PRR activation T-independent B-cells are important in mice, and their role is still being clarified in primates A brief segue – fluid flow through lymphatics https://commons.wikimedia.org/wiki/File:2202_Lymphatic_Capillaries.jpg  Lymphatic Conducting System What provides the driving force for lymph to enter lymph capillaries? ▪ Interstitial fluid build-up on the outside of the capillaries puts pressure on the capillary cells and forces them apart Once lymph enters, the external pressure dissipates ▪ Cells close again and the lymph is pushed forward Lymphatic Flow – Larger Channels What provides the driving force for lymph to circulate in the larger lymphatic vessels? ▪ Contraction of skeletal muscle surrounding the lymphatic vessels Skeletal muscle “squeezes” the vessels → movement of fluid forwards since there are one-way valves One-way valves prevent backflow ▪ Pressure created through breathing With each inhalation, the thoracic cavity expands ▪ This lowers the thoracic pressure, facilitating flow of lymph (and blood) from abdomen towards chest Lymphatic Anatomy - Basics Lymph vessels ▪ After exiting lymph nodes, vessels combine to form trunks ▪ Trunks combine to form one of two lymphatic ducts that drain into the blood system Right lymphatic duct ▪ Drains the right arm, thoracic region, and right head/neck ▪ Drains into the juncture of the right subclavian vein and right internal jugular Thoracic duct ▪ Drains the rest of the body ▪ Drains into the juncture of the left subclavian vein and left internal jugular Basic Lymphatic Anatomy https://upload.wikimedia.org/wikipedia/commons/2/2b/Anatomy_of_the_lymphatic_system.jpg