BIO 341 Unit 3 EU

Generation of T-cell immune response

Infection can occur anywhere in the body

Myeloid dendritic cells roam body, capture antigens, and bring them to secondary lymphoid tissue to present to naive T cells

• Infections in skin → draining lymph nodes

• Infections in blood → spleen

• Infections in mucosal tissue → mucosal secondary lymphoid tissues

Dendritic cell maturation

dendritic cells undergo a maturation process traveling from skin/periphery to lymph nodes

Immature dendritic cells in skin

• very good at capture, uptake, and processing of antigens

Mature dendritic cells in lymph node

• elaborate dendrites to aid in T cells activation

What happens to the rest of the T cells

pathogen specific T cells are 1 in 10^4-10^6 of all T cells

Naive T cells continue circulating

• condensed chromatin

• Little cytoplasm

• Little RNA and protein synthesis

Once activated, takes a few days for T cell to proliferate and differentiate into effector T cell = delay time between infection and immune response

Homing of naive T cells to lymph nodes

circulating T cell enters the high endothelial venule in the lymph node

Binding of L-selectin to GlyCAM-1 and CD34 allows rolling interaction

LFA-1 is activated by chemokines bound to extracellular matrix

Activated LFA-1 binds tightly to ICAM-1

Diapedesis - lymphocyte leaves blood and enters lymph node

Once inside lymph node… (T cell)

T cell initially binds dendritic cells roam body through low affinity LFA-1 and ICAM-1 interactions

Subsequent binding of T cell receptors send signal to LFA-1

Conformational change in LFA-1 increases affinity and prolongs cell-cell contact

Leaving the lymph node (T cell)

Most T cells won’t recognize antigen and will leave lymph by efferent vessel

S1P gradient directs cells toward efferent vessel

• Naive T cells have S1P receptor on cell surface to follow gradient

• Upon recognition of antigen, CD69 sequesters S1P receptor inside cell (T cells is stuck in lymph node)

• Mature/effector T cell lose CD69, express S1P receptor again, and can leave lymph

Co-stimulatory molecules

T cell must recognize peptide:MHC complex but that does not lead to full activation

Activation requires co-stimulatory signal

• CD28 receptor on T cells binds to B7 molecule on dendritic cell

• Only dendritic cells, macrophages, and B cells have B7 (only expressed during infection)

• Once T cells won’t recognize is activated, CD28 is replaced by CTLA4 (binds B7 20x stronger than CD28)

The SMAC

when naive T cell binds to dendritic cell, binding occurs in T ell synapse

Within the synapse are two areas

• c-SMAC (central)

• p-SMAC (peripheral)

ITAMs

Upon binding at T cell synapse, signaling beings by phosphorylation of ITAMs:

In the resting T cell the ITAMs of the CD3 complex are not phosprylated

Binding of MHC to the TCR leads to ITAM phosphorylation by Lck

Zap-70 binds phosphorylated zeta-chain ITAMs and is phosphorylated by Lck

effects of ZAP-70

T cell binds to dendritic cell → Lck phosphorylates ITAMs → ZAP-70 binds to phosphorylated ITAMs → PLC-gamma → PIP2→cleaved into DAG and IP3

→DAG

• →PKC-theta → NFkB

• Ras-MAPK → AP-1

→IP3

• increases Calcium → Calcineurin → NFAT

3 transcription factors (NFkB, AP-1, and NFAT) made that change the pattern of gene expression → cell division, proliferation, and differentiation to mature/effector T cells

IL-2

IL-2 is in activated T-cells

• cells constitutively express partial receptor

• IL-2 expression leads to full receptor function and autocrine signaling

T-cell anergy

some T ells will respond to self-antigen by accident

Lack co-stimulatory molecule → T cell anergy

Anergy = unresponsive and cannot ever be revived

differentiation of CD4 T cells

CD4 T cells are helper T cells

• recruit other cells to destroy pathogen

Several classes of CD4 T cells

• Cytokines environment determines which T cells develop

Polarized T cell responses

activation and differentiation of T cells by cytokines → positive feedback mechanism

• Polarized TH1 response = cell-mediated immunity

• Polarized TH2 response - humoral immunity

Activation of CD8 T cells

CD8 T cells are cytotoxic so they need a much stronger signal to be activated (because they kill)

Dendritic cell usually activates CD4 T cells to secrete IL-2 and CD8 T cell → IL2 from the CD4 T cell drives proliferation and differentiation of CD8 T cells

Dendritic cells can also send the signal themselves to activate CD8 T cells to effector status → CD8 T cell makes IL2 driving its own proliferation and differentiation

post activation of T cells

Once T cells are activated they leave secondary lymphoid tissue and travel to infected tissues to do their work

• leads to changes in many cell surface proteins

Relaxation of co-stimulators requirement

While activation of T cells required co-stimulators molecule, differentiated effector T cells do not have this requirement

Allows CD8 cells to bind to any cell with pathogen

Allows CD4 cells to bind more targets

cytokines and cytotoxins

effector T cell functions are mediated through a host of Tolkien’s and cytotoxins

• cytotoxins - kill cells

Cytokine regulation of gene expression

effector T ells can disrupt target cells by altering gene expression through the release of cytokines

JAK-STAT pathway often activated in this pathway

CD8 effector cell function

once inside of a cell, pathogens become inaccessible to antibodies

CD8 T cells secrete lyric granules (vesicles) to kill target cell as well as pathogen

Once they have enough lyric granules again, CD8 T ells can kill multiple cells in succession

killing by apoptosis

CD8 T cells kill their target cells by inducing apoptosis

• pokes holes in cell membrane

• Activate nucleases to chop up DNA

• Cell shrinks by shedding membrane-bound vesicles

Effector TH1 CD4 cells

Responsible for activating macrophages

• binding of CD40 and IFN-gamma lead to macrophage activation

TH1 cell and infected macrophage come together → T cell beings to and activates macrophage → killing intravesicular bacteria (remember macrophages aren’t activated which is why they aren’t killing pathogen in leprosy)

Effector TFH CD4 cells

TFH cells are responsible activating naive B cells

After differentiation they move to B cell area of lymph node to complete their work

TFH cell recognizes peptide derived from B cell’s antigen → Naive B cell and TFH cell exchange signals that being the process of B cell activation

B cell activation

B cell activation requires cross-linking of BCR by the antigen

• activates signaling pathway through ITAMs

Cross linking of B cell receptors by antigen → clustering of antigen receptors allows receptor-associated kinases to phosphorylate the ITAMs → Syk binds to doubly phosphorylated ITAMs and is activated on binding → changes in gene expression in nucleus

Co-receptors

Just like T ELLs, binding of B cells to antigen isn’t enough to fully activate

Must bind to co-receptor made up of 3 parts:

• CR2 and CD21 = recognizes C3b fragments on pathogen

• CD19 = signaling chain

• CD81 = brings CD19 to the surface

BCR/co-receptor function

BCR and co receptor cooperate in B cell activation by a pathogen

BCR and co-receptor cooperate in B cell activation by a soluble antigen

→ both lead to changes in gene expression in nucleus

TFH cells activating B cells and TI antigens

Most B cells are activated by TFH cells in the secondary lymphoid tissues

Thymus independent antigens

• Some B cells can be activated by certain antigens without T cell activation

• No need for T cell

Follicular dendritic cells (FDC)

FDCs and macrophages present antigens to B cells

Express CR2 to capture antigens and present to B cells

Activation of B and T cells in lymph nodes

Naive B cells search for specific antigen displayed by FDC in the B cells area; naive T cells search for specific antigen presented by dendritic cells in the T cell area

Antigen activated T cells proliferate and differentiate; antigen activated B cells move to the boundary region

Antigen activated B cells present antigens Thymus independent to effector TFH cells, forming cognate interactions and cognate pairs

TFH secrete cytokines to aid B cells

Helper TFH cells conjugates with the B ell and beings to synthesize cytokines and CD40 ligand

The helper TFH cell reorients its cytoskeleton and secretory apparatus toward the B cell

Cytokines are secreted into the narrow space between the TFH cell and the B cell

clonal expansion

after conjugate pair forms from B and T cells, they must being clonal expansion

Primary focus occurs in medullary cords

• lasts several days and produces B cells secreting IgM

• Some B cells stay there and differentiate into plasma cells

The primary focus for expansion of antigen activated B cells in in the medullary cords → the secondary focus for expansion of antigen activated B cells is in the primary follicle → expansion of antigen activated B cells in the primary follicle creates the germinal center

Secondary focus

many B/T cell conjugates will leave medullary cords and move to primary follicle for expansion → germinal center

Somatic hypermutation and isotype switching occur here

Centroblast = large, metabolically active B cells that divide rapidly

• cells have isotype switching occurring

Centrocytes = divide slowly, re-express cell-surface immunoglobulin that has been isotype switched

• derivatives of centroblasts

• Will die quickly if not bound to antigens

Selection of centrocytes

activated B cell→somatic hypermutation of immunoglobulin V regions in rapidly proliferating germinal center control blasts

→Germinal center centrocyte with low affinity surface immunoglobulin → BCR is not cross linked and centrcyte cannot present antigen to TFH cells → cell death

Or

→ germinal center centrocyte with high-affinity surface immunoglobulin → BCR is cross linked and antigen is presented to helper TFH cell → centrocyte receives help, survives and divides → centrocytes differentiate into plasma cells

cytokines from T helper cells determine B cell fate

Cytokines determine isotype switching (Ex: IL-4 induces IgG1)

If no binding to T cells through CD40, what will happen to B cells?

• B cells will only secrete IgM (can’t isotype switch if they can’t bind to T cell)

Differentiation to plasma vs memory cells

centrocytes with identical BCR

Antigen selected centrocytes mature under the influence of an IL-10 secreting helper TFH cell → centrocytes differentiate into plasma cells → make antibodies that fight infection

Or

Antigen selected centrocytes mature under the influence of an IL-4 secreting helper TFH cell death Or→ centrocytes differentiate into memory B cells → the investment that prevents future infections from causing disease

Antibody effects functions - IgM, IgG, monomeric IgA

while IgM is the first antibody produced, it undergoes isotype switching

IgM, IgG, and monomeric IgA protect internal tissues

Most common circulating molecule is IgG

• smaller than IgM and can reach more tissues

• Relies on Fc receptor (FcRN) to be carried into tissues

Dimeric IgA

responsible for protecting mucosal surfaces

IgE

found in very small amounts in body

Not secreted as a soluble antibody, remains on cell surface

Cross linking of IgE antibody leads to release of histamine from mast cells

• inflammatory mediator

Effects of inflammatory mediators

inflammatory mediators are released by mast ELLs, basophils, eosinophils

• smooth muscle contractions in airways and gut to expel parasites

• Increased permeability of blood vessels to help flush out parasites

• Physical removal of parasites that are too big to be phagocytosed

In developed countries, we aren’t exposed to many parasites so mast cells are considered a detriment

• some people make IgE that responds to innocuous substances like pollen

Antibodies from mothers

during pregnancy, from IgG from maternal circulation is transported across placenta and delivered to baby

At birth, baby has as high levels of IgG as mother with as wide a range of antigen specificities

Infants obtain IgA from breastmilk = passive transfer of immunity

• goes to gut and binds to microbes; excreted through feces

Baby’s first antibodies

after birth, antibody level gradullary decreases until 6 months when baby begins to make its own antibodies

• maternally derived IgG is catabolized and IgA is lost immediately in feces

• IgG levels are lowest in children age 3-12 months

  • Problematic for premature babies who didn’t get much of mother’s IgG and take longer for immune system to be fully functioning

Also age where genetic deficiencies in immune system are found

Neutralizing antibodies

specific to an antigen and can prevent microbe from attaching to human epithelium

neutralizing toxins

many microbes release toxins that can be neutralized by IgG or IgA

Antibody activation of complement

Most antibodies don’t directly kill pathogens but rather recruit other cells/molecules to destroy pathogen

• IgM, IgG3, and IgG1 can activate complement

Activation of complement by IgG

in order for IgG to activate the complement pathway, C1q must bind to at least 2 IgG molecules

• are we sure we’re gonna do this (cell death)

IgG molecules bind to antigens on bacterial surface → C1q binds to two or more IgG molecules and initiates complement activation

Or

IgG molecules bind to soluble multivalent antigen → C1q binds to soluble immune complex and initiates complement activation

The role of RBC’s

RBCs aid in removal of immune complexes from the circulation through CR1;C3b binding

Small antigen:antibody complex forms in circulation and activates complement → immune complex is coated with covalently bound C3b → CR1 on an erythrocyte surface binds C3b-tagged immune complex → erythrocyte carries immune complex to the liver or spleen, where it it detached and taken up by a macrophage

Fc(gamma) receptors

complement its powerful because C3b coats pathogens permanently, delivering them for uptake and elimination

• antibodies can bind just as well

Fc gamma receptors are specific for IgG and are expressed by macrophages and dendritic cells

• aid in uptake of pathogen coated in IgG

Antibody dependent cell mediated cytotoxicity (ADCC)

we can use our knowledge of Fc receptors in therapy:

B cell lymphoma cells express CD20

Treat patients with anti-CD20 antibody (Rituximab)

Fc receptor on NK cell binds to Rituximab and signals NK cell to kill B cell lymphoma cell