Chapter 9

Immunity Mediated by B cells and

Antibodies

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Antibody production by B
lymphocytes

First priority- speed of production

low affinity IgM antibodies (useful but not optimal)

Second priority- improve quality of antibody

Somatic hypermutation and change isotype

Cross linking of B cell
receptors
• Surface IgM molecules of a
naïve mature B cell become
physically cross linked to each other
by antigen
• B cell receptors cluster and send signal
through Igα and Igβ
• Igα and Igβ have ITAMs which
get phosphorylated
• Receptor associated
tyrosine kinases Blk, Fyn,
Lyn phosphorylate tyrosine residues in
ITAMs
• Tyrosine kinase Syk binds Igβ
tails that are doubly
phosphorylated
• Interaction between Syk
molecules sends activation
signal to nucleus and changes
gene expression
 • Cross linking of B cell receptor necessary, but
not sufficient to activate naïve B cell
• B cell co-receptor required
• CR2 or (CD21)- complement receptor 2 which
recognizes iC3b and C3d derivatives of C3b
fragments on pathogen
• CD19- signaling chain
• CD81- binds CD19 and is essential for bringing it
B cell co- to B cell surface

receptor
Function of
B cell co-
receptor
• CR1 and CR2 present on
B cells
• CR1 can bind C3b
• C3b susceptible to
cleavage by factor I
resulting in iC3b and C3d
• C3d is B cell co-receptor
ligand
Signals generated from B cell
receptor and co-receptor
• Binding of B cell receptor to
antigen and co-receptor to C3d,
brings them close together
• Lyn phosphorylates cytoplasmic
tail of CD19
• Phosphorylated CD19 interacts
with signaling molecules
• Synergistic activation signal that
changes gene expression
• Increases overall signal 1000-
10,000 fold
• Individuals lacking B cell co-
receptor have low level of
antibody, very
little isotype switching (defective
CD81 or CD19)
• Poor B cell response to infections
and vaccines
B cell activation by a
thymus-independent
antigen
• Infants with DiGeorge Syndrome- who lack thymus have
normal number of B cells but can’t make effective antibody
response
• Die within first two years unless have a thymus transplant
• Can produce low affinity IgM antibodies
• CD5 expressing B-1 cells
• Don’t require T-cell help
• No isotype switching
• No affinity maturation
• Thymus independent antigens (TI antigens)- antibodies
against these antigens are present in athymic individuals
because population of B cells that produce these antibodies
are thymus independent
• B-2 cells nonfunctioning
Follicular dendritic
cells
• B cells depend on FDC- accessory cells dedicated to B cell
development and function
• Different from myeloid dendritic cells and
plasmacytoid dendritic cells
• FDCs organize B cell area of lymph node into primary follicle
• Serve as a depository of intact antigens available for
interaction with antigen receptors of circulating B cells
• Good at displaying intact antigens because:
• Extensive surface area of dendrites allows large
quantity of antigen to accumulate
• FDC have no phagocytic ability so antigens remain
intact on surface of cell from months to years
• C3b and C3d attached to pathogens
and antigens
• FDC’s CR2 and CR1 bind C3b and C3d
and hold them at surface of cell
• Subcapsular sinus macrophage- found
in subcapsular sinus of lymph node
and resemble FDCs
• Little phagocytic activity
• CR1 and CR2 bind C3b or C3d
and hold on cell surface
• CR2 made up of 15 CCP modules-form
long flexible stalk so can fish for C3d
tagged proteins and peptides in lymph
• Medullary sinus macrophage
• Highly phagocytic
• Filters lymph before it leaves
the node
• Removes remaining pathogens
or antigens
B cells home to
lymph node
• Circulating naïve B cells home to lymph node
• Enter through HEV
• Attracted into T cell area by CCL19 and
CCL21 and then into B cell follicle by
CXCL13
• Enter through lymph
• Enter node at subcapsular sinus. Naïve
B cells screen antigens at surface of
subcapsular sinus macrophage
• If specific antigen is found B cells enter
B cell area of follicle to interact with TFH
cells and complete its activation
B cells antigen
recognition
B cells recognize antigen
• If B cells recognize antigen on FDC B
cell is activated and induced to
express CD69
• CD69 prevents expression S1P
receptor which keeps the B cells in
the lymphoid tissue to continue
differentiation
B cells don’t recognize antigen
• Express S1P receptor and leave
through efferent lymph by S1P
gradient
Activated B cell-TFH
interaction
• Antigen activated B cells endocytose and process
complexes of B cell receptor with antigen
• B cells then present peptides from degraded antigen
on MHC class II molecules
• B cells induced to express CCR7, which binds CCL21
and CCL19 draws activated B cell to boundary between
B and T cell area
• T cells reduce secretion of CCR7, which facilitates
movement to the B cell/T cell boundary
• Antigen stimulated B cells interact with differentiated
TFH cells
• Conjugate pairs form if T cell recognize B cell MHC
class II:peptide
Conjugation between B cell and T cell

• Interaction induces T cell to

express CD40 ligand and bind B
cell’s CD40
• B cell activates NFκB and
increases expression ICAM-1
that binds T cell’s LFA-1
• Strengthens interaction
between the two cells
Conjugation
between B cell and
T cell
• T cell reorganizes
cytoskeleton and Golgi
apparatus that
facilitates delivery of
cytokines onto B cell
Primary focus of B cell expansion
• Conjugate pairs move out of T cell
area medullary cords
• Both cells begin to divide forming primary
focus of clonal expansion
• Lasts several days results in dividing B
lymphoblasts secreting IgM
• Antibody leaves node in efferent lymph
goes to blood and then site of infection
Differentiation to plasma
cells
• Some B lymphoblasts stay in medullary
cords and differentiate into plasma
cells due to IL-5 and IL-6 secreted by
TFH
• Transcription factor BLIMP-1 results in
terminal differentiation of lymphoblast
to plasma cell
• Stops transcription of genes
needed for proliferation
 • Some clones return to primary follicle in the cortex
• Proliferate and become second focus of B cell
expansion
• Germinal center is formed by expanding population
of specific B cells

Second • B cells undergo affinity maturation and isotype

switching

focus of B
cell
expansion
 IL-6, IL-15, 8D6 and BAFF made by FDC cells
force B cells to rapidly divide (1/6hr) and
become centroblasts

TFH cells also divide, make cytokines and

interact via CD40 ligand which induces B
cell to produce AID
Primary follicle
becomes secondary
follicle Centroblast is not expressing surface Ig-
purpose is expansion of large population of
B cells with switched isotype and V region
mutation

Primary follicle becomes secondary follicle

Germinal center in
secondary follicle
• Germinal center is dominant feature
• Rapidly dividing B and T cells
• Periphery of follicle- naïve B cells passing
through lymph node in search of specific
antigen and survival signals
• B cells form zone- mantle zone
• In primary immune response germinal
centers appear a week after infection starts
• Germinal center reaction- all activity that is
occurring in germinal center
Germinal Center
• Centroblasts
• Dark zone
• Closely packed
• Centroblasts become
centrocytes
• Centrocytes-
• divide more slowly
• begin to express surface
Ig that is mutated and
switched in isotype
• Centrocytes leave dark zone
move to light zone
• Light zone- lower density B
cells, high density FDCs and
TFH cells
• Centrocytes programed to die
by apoptosis unless their
surface Ig is bound by antigen
and CD40 bound by T cell
CD40 ligand
Fate
of centrocytes
Most B
cell lymphomas originate
with a germinal center
cell
 B cell
receptors
with
reduced
affinity
B cell receptors with higher affinity
 Isotype switching
Happening in Germinal Centers

Mouse B cells

• T cell cytokines induce switching by stimulating transcription from the

switch region 5’ to each heavy chain C gene
• Could be opening up chromatin making switch region accessible to
somatic recombination machinery that will bring a new C gene next
to V region sequence
Isotype switching
• Requires CD40 on B cell to bind CD40 ligand on
TFH cell
• Individuals lacking CD40 ligand – hyper-
IgM syndrome
• B cells can’t switch Ig isotypes
• High amount of IgM, almost no IgG and IgA
• No germinal centers
Plasma cell
or memory
cell
Comparison of naïve B cells and plasma cells
 • Initially pentameric IgM is made
• Affinity maturation and isotype
switching
• IgG and monomeric IgA
IgG • 2 high affinity binding sites as
effective as 10
• Smaller better able to access
infected tissue
• IgG –dominant blood borne Ab
• Monomeric IgA also contributes
 Transport of IgG
from blood to
tissues

FcRn- transport receptor

Similar to MHC class I
molecule
Dimeric IgA

• Protects mucosal surfaces that communicate with

external environment
• Lining of GI tract, respiratory tract, genital tract,
urinary tract
• Eyes, nose, throat
• mammary glands
• Made in patches of MALT in lamina propria, connective
tissue under mucosal epithelium
• IgA secreting plasma cells are on one side of the mucosal
epithelium and target pathogens on other side
• Dimeric IgA molecules transported individually across
epithelium by receptor on epithelial cells
Transcytosis of dimeric IgA
• Polymeric Immunoglobulin
receptor (PIgR)- specific for
IgA dimers and IgM
pentamers
• secretory component-
Small fragment of PIgR still
bound to dimeric IgA
• Mucins at surface hold IgA
• IgA bind microorganisms at
mucosal surface to prevent
attachment and
colonization
• Pathogens removed in
feces, sputum, tears and
other secretions
IgE and parasites
• Fc region of IgE bound by Fc receptor FcεRI
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• FcεRI binds only IgE Ab- cannot disassociate
• The small amounts if IgE secreted by plasma cells
quickly attaches to:
• surface mast cells in connective tissue
• activated eosinophils at mucosal surface
• circulating basophils
• Cross linking of 2 or more FcεRI in mast cell surface
activates cell to secrete active mediators that cause
violent reactions
• Sneezing, coughing, vomiting, diarrhea
• Ejects pathogens from respiratory and GI tracts
• Why this strategy of physical ejection?
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 Activation of mast
cells

• Variety of IgE
• Important in connective
tissue under mucosa of GI
and respiratory tract and
connective tissue along
blood vessels
• Granules- histamine
• Cross-linking of FcεRI
signals degranulation
• Cells synthesize and
package new set of
granules
Inflammatory mediators

• Smooth muscle contraction

• Increase permeability of local blood vessels
• Allow immune cells to move out of blood into
tissue
• Swelling, redness, pain, heat
• Inflammation recruits cells and proteins to site
of infection for defense
• Response is quick because cells are already
loaded with granules and display IgE
 Eosinophils act
directly
against multicellular
parasites

• Even small organisms

are too large to
be phagocytized
• Activated eosinophils bi
nd IgE
through FcεRI and
release granules

SL= schistosome larva

E= activated eosinophils
 • Allergy and asthma
• Response to innocuous
substances- allergens
• Second exposure
response
is inappropriate to
threat posed by antigen
• Extreme cases-
ingestion of allergen
results in life
threatening response-
anaphylaxis

Detrimental
response of IgE
Mothers provide Ab protection to baby before and
after birth

• IgG transported across placenta

by FcRn and delivered to
fetal bloodstream
• Dimeric IgA transferred to gut
of baby
• Passive transfer of immunity-
IgA in breast milk and IV
immunoglobulin to patients
with genetic defect in B cell
function
Immunoglobulin levels
IgG levels lowest in infants 3-12 months
 Neutralizing
antibodies against
viruses

• High affinity Abs bind

microbial ligand
toprevent
attachment
• Stop infection before it
starts
• Mucosal surface- start
of many infections so
dimeric IgA often
neutralizing Ab
Neutralizing
antibodies
against
bacteria
Adhesins- bacterial
surface protein that binds
epithelial cells and allows
bacteria to colonize
 Bacterial
toxins disrupt
normal
function of
cells
Neutralization of toxins by IgG Abs

• Passive immunization-
immunize large
domestic animals with
venom- use Abs to
treat patients exposed
to venom
• Venom from snakes,
scorpions etc. acts as
toxic polypeptide
• Exposure is rare so no
vaccines- use passive
immunization
Activate and fix
complement

• Only small number

of antibodies have
direct inhibitory effect
on pathogens

• More common is
that antibodies bind to Ag
and recruit other immune
cells
IgM activates complement classical pathway
 • End result: phagocytosis by neutrophil or macrophage
• Size of pentameric IgM restricts penetration into
infected tissues
• After affinity maturation and isotype switching
smaller IgG molecules important
• Adaptive immunity provides specificity and innate
Complement immunity provides strength
fixation by
classical
and alternative
pathway
 • Depends on amount and density of Ab bound
to pathogen surface
• Complex formation of antigen and IgG activate C1
classical pathway then follows same pathway as if
activated by IgM
• Soluble immune complexes can also activate
classical pathway and phagocytic cells can then take
up complexes from blood and lymph
Complement
activation
by IgG
Erythrocytes aid in removing
immune complexes from circulation

• Cells with CR1 receptor can bind C3b fragment on

pathogen
• Most numerous circulating cell with CR1 receptor is
erythrocyte
• Erythrocytes bind complex of complement, antigen and
antibody
• When erythrocytes pass through liver and spleen,
tissue macrophage remove and degrade
complexes, while leaving erythrocyte unharmed
• SLE (systemic lupus erythematosus)-immune
complexes in blood are deposited in large amounts
in kidneys which can result in kidney failure
 Antigen cross linking of bound IgG required before
Fc receptors on signal is produced
phagocytes trigger IgG3 and IgG1 have such high affinity for FcγRI that
uptake and breakdown individual IgG can bind receptor without being
of Ab coated pathogens bound to antigen
These interactions do not send signal
Fc receptor expressed by NK cells recognizes IgG coated target cell
and signals it to die
• Rituximab- anti CD20 antibody used to treat B cell lymphomas
• ADCC- antibody dependent cell-mediated cytotoxicity