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1 بسم هللا الرحمن الرحيم The last lecture we discussed the antigen processing and presentation and antigen recognition then the activation by T lymphocyte and today we will continue with B cell recognition of the antigen. As we know B cells have there receptor which is IgM or IgD expressed on there surface and they utilize it to recognize antigens that are soluble and don't involve other molecules. Immunoglobulin receptor is associated with invariant chain (Igα and Igβ) which carry ITAM motif that can transcript the signal in the manner cellular to take place in T cell with the help of CD3 and ايوتا and other chains. And antigen induce cross linking of BCR is important because clusters Igα and Igβ are set a single binding site is not single immunoglobulin molecule,cross linking mean two molecules are involved two immunoglobulin must be cross linked for signal 4:27 which cause clustering for Igα and Igβ and this will lead to phosphorylation of ITAM by Src family kinase like Lyn, Blk, or Fyn and this initiate a process similar to that take place in T lymphocyte where cross linking require identical epitope and phosphorylated ITAMs provide a docking site for the Syk tyrosine kinase, this the equvelant of ZAP-70 in T lymphocyte so this is 5:10 molecule in activation transduction of signal generated by antigen cross linking on B lymphocyte, Syk is an enzyme phosphorylates itself and a number of downstream signaling molecules, including PLC-Gamma and adaptor proteins so Plasma membrane PIP2 is cleaved by PLC-Gamma

2 into IP3 and DAG and the sequence of 6:04 then is similar to that take place in t lymphocyte where IP3 increases cytoplasmic Ca++and DAG induces enzymes such as PKC to activate transcription factors, including NFAT, NK-kB, Myc, and AP-1 (activating protein 1), In addition, PKC phosphorylates serine -threonine residues and also BCR cross-linking results in the Ras phosphorylation and leads to the activation of MAP kinase and this also exist in T-lymphocyte. Now the Different between B and T-lymphocyte is utilization of other receptor to activate B-lymphocyte and induce signal and this can be achieved by CR2/CD21 and CD19 because they carry ITAM so CR2/CD21 is a surface marker for B-lymphocyte. CD19 has a cytoplasmic tail with ITAM motifs and is associated with CR2 on the B cell surface,the engagement of a microbe for example by BCR and of the microbe bound C3b together by CR2 results in ITAM phosphorylation of Igα, Igβ, and CD19, and subsequent B cell activation and BCR-bound antigens can also internalized by endocytosis, enzymatically degraded, and displayed on the B cell surface as pmhc class II molecules recognize by CD4 molecule, Activated B cells enter the cell cycle and increase their Expression of CD80/86 and this bind to CD28,they express IL-2R, IL-4R this enable them to start production of anti body, and also express antiapoptotic proteins, B-lymphocyte which fail in receive a signal will die and this become activated express protein that protect them from apoptosis so antiapoptotic proteins are expressed by B-cell and they now await a T cell signal

3 remember that T-cell signal determine what antibody to be produce class switching somatical hypermutation all takes place after interaction with t-lymphocyte so activated B-cell express both pmhc class II and CD80/86 and CD4 T-lymphocyte will recognize pmhc class II by TCR and CD28 and these two signals will stimulate the T cell to express CD154 (CD40 ligand) and T-cell cytokines induce B-cell CD 40 expression and such interaction provide a signal for proliferation and differentiation for the plasma cell and T-cell cytokines stimulate B-cell proliferation and differentiation to produce antibody class switching and also associated in produce memory B-lymphocyte so activation of B-cell will genarate plasma cell and antibody and memory B-cell. So at one side mean the mechanism of the activatin of B-lymphocye involve cross linking of two immunoglobulin by an antigen and this will cause phosphorylation of ITAM present in Igα and Igβ by Syk family and this will phosphorylation of downstream signaling PCL-Gamma... The other way of activation involve antibody and CR2, CR2 is CD21 and antigen is opsonized to B-lymphocyte with the help of c3b which has CR2 receptor for c3b and now this will phosphorylate the ITAM with Syk until the production of nuclear protein that will make B-cell active now the B-cell is provided as a consequence presented an antigen by B-cell to T-lymphocyte and the second signal is receive from the T-cell in the form of cytokines So signal 1 is antigen binding and signal 2 is provided by T-cell by t-helper and that involve in prodction of CD80/86 with CD28 cd40 with cd40 ligand 13:02 // B cells are inherently prone to die by apoptosis Signal 1 & 2 unregulate

4 Bcl-XL in the B cell and Bcl-XL prevents apoptosis Signal 1 & 2 thus allow the B cell to survive so surival of B cell is t-dependent T cells regulate the survival of B cells and thus control the clonal selection of B cells, that's why we say that t-cell is key regulator of all type of immune response. The other issue is class switching and somatic hypermutation that take place as a consequnce of interaction between B AND T-lymphocyte where the set of signal by B-cell and t-cell* cause activate somatic hypermutation of CDR encoding part of immunoglobulin gene and serve to provide by the T-cell with helper regulatory T-cell. T-cells are clonally selected where low affinity Ig takes up and present antigen T-cell inefficiently, so inefficient to present Ag to T-cell doesn't induce CD40 and without the signal 2 delivered by CD40 low affinity B-cell will die, whereas only B-cell with high affinity Ig will survive,so T-cell determine the survival of B-cell and also responsible for class switching and somatic hypermutation that take place during antibody production. The issue of affinity is matter binding, we have 5 B-cell that can bind to antigens, the affinity of binding is variable and the second B-cell in the figure in slide 48 has best binding and the only cell with best binding can express CD40 and this cell can receive signal 2 and only this cell is rescued from apoptosis i.e clonally selection. The cells with lower affinity receptor will die by apoptosis by neglect. Effector Functions of the Adaptive Immune System

5 The effector cells that have effector function are : cytotoxic T lymphocyte, the late hypersensitivity cells and antibodies. Cells of adaptive immune system often use cells and molecules of innate immune system to focus and deliver fatal blow to an invader, this cells activated by production of cytokines that activated some cells like NK cells,macrophages that will get it of the invader,so invader is attacked by innate immune system but they are activated by the adaptive immune system which amplify and intensify the effector function of the innate system. As we mentioned earlier the term cellular and humoral immune response are actually misnomers,and all immune response have cellular basis. Cell-mediated immune response Delay type hypersensitivity: Delay type hypersensitivity is reaction localized to immediate area surrounding the site of injection that develop within hours, the proto type of this reaction is PPD skin test that's why we use here site of injection to inject the antigen and the local reaction will take place in the lung!!!! In the form of delay type hypersensitivity and assuming that we inject an antigen like PPD from micro bacteria and the action of induration and erthyma will develop and this will be followed by necrosis and ulceration and that's why region that develop in the lung will form granuloma. If we talk about skin test we don't measure erythema, erythema is a result of vaodilation but if we want to quantify the reaction

6 we measure the diameter of induration. We can label the test as +ve or -ve base on the diameter of induration and usually we have low,intermediate and high dose of PPD,we use intermediate dose and any reaction of 5mm or more consider +ve reaction of PPD. If we use a group of antigen to asses indicative immunity we should have +ve reaction for 2 antigen at least so if we use 5 antigen the individual should respond positively to two antigen where the sum of diameter 10mm or more (5 antigen). So the induration is developed in response to intracellular antigenic insult being mediated by CD4+Th1 cells which are responsible for such response. Now activated Th1 cells increase level of integrin namely LFA-1 and VLA-4 which during present in circulation,and phagocyte secrete TNF which signals nearby epithelial cells to increase the level of selectin and ligand for LFA-1 and VLA-4 on their luminal surfaces. Interaction between adhesion molecules and their ligands allow activated Th1 cells to localize at the epithelium overlying the site of inflammation followed by their migration to that site. Macrophage Activation Macrophage become activated upon arrival of Th1 at site of the inflammation and recognition of pmhc by Th1 TCR cause Th1 to express CD154 and secret INF-gamma which activate macrophage. The activated macrophage upregulates CD80/86 expression and they will secretes IL-1, IL-12, and TNF which increases

7 MHC class II expression and increase the production of degradative enzymes and reactive oxygen intermediates (ROI). * IL-1 and TNF attract polymorphonuclear cells to the Site of action. * Activated macrophages and neutrophils remove cellular debris, infected cells, and even some normal cells and also stimulate repair and mediate healing by secreting platelet-derived growth factor, TGFβ, and fibroblast growth factor. Cytotoxic T cell response Is the second arm of effector cells of delay type hypersensitivity, the target cell recognition subsequent to pmhc-tcr CD8 interaction leads to upregulation of Fas ligand. Contact between the CTL and target cell lasts for only a few minutes, during which a fatal hit is delivered to the target cell where Perforins and granzymes are released upon contact with the target cell. Perforin creates osmotic imbalance and granzymes enter through pores to activate caspases leading to apoptotic lysis. * regard -cell we have two arm CD4 and CD8, CD4 mediate the delay type hyper sensitivity whereas CD8 mediate cytotoxic T cell response. Humoral immune response This response is mediated by anti bodies, antibodies can be produce by T-dependent and T-independent antigen. T-dependent antigen: Antibody production requires assistance from T helper cells. Macrophages ingest ''process'' antigen and present it to TH

8 cell. TH cell stimulates B cells specific for antigen to become plasma cells. Antigens are mainly proteins on viruses, bacteria, foreign red blood cells, and hapten -carrier molecules. T-independent antigen : Antibody production does not require assistance from T cells. Antigens are mainly polysaccharides or LPS with repeating identical subunits (bacterial capsules); that's why they don't need processing and presentation. Weaker immune response than for T-dependent antigens. Activation mechanism of B-cell by T Independent Antigens (TI-1 ) : Bacterial Lipopolysaccharides, (TI-1 antigens), bind to host LPS binding protein in plasma. LPS/LPSBP is captured by CD14on the B cell surface. Toll - like receptor 4 (TLR4) interacts with the CD14/LPS/LPSBP complex. activation of B-cell. At high dose TI-1 antigens (like LPS) will POLYCLONALLY ACTIVATE all of the B cells irrespective of their specificity. So that's why TI-1 antigens are called MITOGENS. **Form slide 59 to 72 he just read the slide and there is some notes : Slide 67 point 2 :

9 Many of these cells will undergo apoptosis because there is no space and this process known activating killing cell. Slide 68 point 1 : They found that we can distinguish between memory cell and naive T-cell by CD40. Sorry for any mistake ركاد دمحم ابوركبة

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