chapter 1 9/14/13 12:35 PM

Medicine and history – immunity builds over time

Immunization = exposure to infection without causing infection

First line of defense = innate immunity

Physical and chemical barriers

Next = adaptive immunity = specific target

Refined over course of infection

Provides long lasting immunity

Commensal species = microbial species in the gut

Flora = microbial species from specific region

Pathogens

Bacteria

Virus

Fungi

Parasites

Endemic disease = ubiquidous

Childhood exposure

Skin = first barrier

Forms epithelium (keratinized cells)

Also in lining of respiratory and gastrointestinal and urogenital

tracts – usually musosal lining

o Glycoproteins, proteoglycans, enzymes

All epithelia produce antimicrobial peptides called defensin

o These kill pathogens on their turf

Tears and saliva have lysozymes – degrade cell wall

Acid from stomach, vagina, skin

Innate immune response

First recognition

Then effectors

Innate immunity inherited

Recognition

o Pathogen is present

o Soluble proteins and cell surface receptors bind to pathogen

and its products or human cells and serum proteins that

become altered in the presence of the pathogens

Effector mechanism

o Kills and eliminates

Effector cells engulf bacteria, kill virus infected cells, or

attack protozoan parasites, and a battery of serum

proteins called complement that help the effector cells

by marking pathogens with molecular flags but also

attack pathogens in their own right.

Overall effect of the innate immune response is to induce a states of

inflammation

Cytokines sent to damaged tissue

Calor – heat, dolor – pain, rubor – redness, tenor – swelling

Cytokines induce dilation of capillaries which increase blood flow

gaps in between cells of endothelium edema (swelling) pain

Cytokines cause white blood cells to stick to infected tissue/area

Adaptive immunity

Pathogen specific response

Clonal selection = select a small subset of lymphocytes for

proliferation ≈ felt week after infection

Clonal expansion = select a small subset of lymphocytes for

differentiation ≈ felt week after infection

Memory cells – faster response second time around

First exposure = primary immune response

Subsequent exposures = secondary immune response

Immune cells mainly leukocytes (white blood cells)

Hematopoiesis = continual generation of leukocytes

Leukocytes come from pluripotent hematopoietic stem cells – also make

erythrocytes (red blood cells), megakaryocytes (which become platelets)

Place of production changes with age

Embryo – blood cells from yolk sac then fetal liver

3-7 months of fetal life – spleen

4-5 months of fetal life – shifts towards bone marrow

birth onwards – bone marrow

adults – bone marrow of skull, ribs, sternum, vertebral column,

pelvis, and femurs

mature stem cells commit to one of three lineages:

erythroid

o lead to oxygen carrying erythrocytes and platelet producing

megakaryocytes

platelets are small non-nucleated cell fragments of

plate-like shape that maintain the integrity of blood

vessels… important to clotting

megakaryocytes are giant cells that arise from the

fusion of multiple precursor cells and have nuclei

containing multiple sets of chromosomes… permanent

residence in bone marrow

myeloid

o granulocytes… cytoplasmic granules containing reactive

substances that kill microorganisms and enhance

inflammation

neutrophil (most abundant granulocyte)

capture, engulfment, killing of microorganisms

main ingredient in pus, pimples, boils

neither acidic nor basic stains

eosinophils (second most abundant granulocyte)

defend against parasites

basic subsatances bind to acidic stain eosin

basophils (least abundant granulocytes)

regulate immune response to parasites

little known

acidic substances that bind basic stains

o monocytes, macrophages, and dendritic cells

monocytes = leukocytes that circulate in the blood

bigger than granulocytes, distict indented nucleus

macrophages

travel in blood to tissues

large phagocyte… phagocytosis

general scavenger cells of body

live longer

secrete cytokines that recruit neutrophils

and other leukocytes

Dendritic cells in body tissue

act as cellular messengers that are sent to

call an adaptive immune response when

needed

mast cells – in connective tissues

involved in activation and degranulation of mast

cells at infection site

lymphoid

o Natural killer cell/T cell precursor

Natural killer cells – effector cells of innate immunity

with large granular lymphocytes

Important for viral infections

Prevent spread of infection by killing virus

infected cells

Secrete cytokines that impede viral replication

Small lymphocytes are responsible for adaptive immune

response

Recognize pathogen and drive lymphocyte

selection, growth and differentiation

o B cells and T cells

B-cell cell surface receptors for pathogens =

immunoglobulins

B-cells express single immunoglobulin

T-cell cell surface receptors for pathogens = T-cell

receptors

T-cells express single t-cell receptor

T cells

Cytotoxic T cells

Kill cells infected with either viruses or

bacteria that live and reproduce inside

humans

NK cells of adaptive immunity

Helper T cells

Secrete cytokines that help other cells of

immune system become fully activated

effector cells

B cells

Plasma cells – effector cells that secrete soluble

immunoglobulin called antibodies

Most lymphocytes found in lymphoid tissue or lymphoid organs

Lymphoid organs = bone marrow, thymus, spleen, adenoids, tonsils,

appendix, lymph nodes, and Peyer’s patches

Lymphoid tissue = lining of mucosal surfaces of respiratory,

gastrointestinal, and urogenital tracts

o Two types

Primary/Central lymphoid tissue

Lymphocytes mature and develop to the stage at

which they are able to respond to pathogens

Bone marrow and thymus

B and T lymphocytes originate from bone marrow

B cells mature in bone marrow and enter

circulation

T cells leave bone marrow immature and

migrate in blood to thymus to mature

Secondary/Peripheral lymphoid tissues

Mature lymphocytes become simulated to

respond to invading pathogens

Lymph nodes – lie at junctions of lymphatic

vessels

Collect plasma that leaks out of blood

vessels and forms extracellular fluid

o This lymph returned to blood

Lymph flow driven by continuous movement

Adaptive immunity is initiated in secondary lymphoid tissues

Connective tissues usually site of infection (pathogens through

wound)

o Pathogen carried by lymphatic to lymph node receives fluid

collected at infected site ( draining lymph node) blood

cells call upon proper lymphocytes

Spleen provides adaptive immunity to blood infections

Spleen filters blood

Removes damaged or senescent red cells

Defends body against blood-borne pathogens

Splenic macrophages and dendritic cells simulate B and T cells from

blood

Spleen made of two different tissues

o Red pulp (RBC monitored and removed)

o White pulp (WBC gather to provide adaptive immunity)

No spleen = asplenia

o Genetic

o Susceptive to encapsulated bacteria (whose cells surrounded

by thick polysaccharide capsule)

Damaged spleen often removed

o Children with spleenectomy more vulnerable to bacterial

infections than adults

Adults have protective immunity from previous

infections

Most secondary lymphoid tissue associated with gut

GALT (gut associated lymphoid tissues) = tonsils, adenoids,

appendix, and Peyer’s patches (line small intestine)

BALT (bronchial associated lymphoid tissues) = lining of respiratory

epithelium

MALT (mucosa-associated lymphoid tissues) = mucosal surfaces like

gastrointestinal tract

Adaptive immunity leads to long lived immunological memory cells

Immune system can be compromised by immunodeficiencies and certain

pathogens

Mutations in both copies

Immunodeficiencies once cause death in infants

AIDS and HIV

Chapter 3: Adaptive Immunity 9/14/13 12:35 PM

Adaptive immunity is the body’s third line of defense

1st: physical barriers

2nd: innate immunity

adaptive immunity made up of B cells and T cells (both lymphocytes)

innate immunity and adaptive immunity differ in how they recognize

pathogens

B and T cells recognize pathogens by using cell surface receptors of

just one molecular type (B cell receptors and T cell receptors)

o These proteins can be made in infinite different versions

Adaptive immunity only in vertebrates, innate immunity shared with

many animals… and some plants

Advantages:

Strong immune response against particular pathogen effective by

targeting small differences that distinguish that pathogen from

others

Pathogen-specific B and T cells stored as memory cells in case

pathogen returns

Evolving vertebrates have defense against evolving microorganisms

B cell receptors = immunoglobulins

Effector B cells = plasma cells, secrete soluble immunoglobulins =

antibodies

T cell receptors only expressed as cell surface recognition … never

secreted

antigen = molecule, macromolecule, virus particle or cell that has

structure recognized and bound by immunoglobulin or T cell receptor

these t cell receptors or immunoglobulins = antigen receptors

immunoglobulins can bind many different structures, TCR can only

bind few

both have specificity for antigens they bind

immunoglobulins and TCR are structurally related

immunoglobulins are Y shaped polypeptides with two identical

heavy and two identical light chains

Both have variable and constant regions

Antibodies are secreted immunoglobulins

TCR has alpha chain and beta chain

Both have variable and constant regions

The constant regions of antibodies contain binding sites for cell surface

receptors on phagocytes and other inflammatory cells

Immunoglobulins have several different types of constant regions (Not

TCR) which confer different effector functions on the secreted antibodies

and can also target then to different sites in the body

Gene rearrangement leads to diversity of immunoglobulins and TCR

Variable regions of immunoglobulins and TCR have separate gene

segments called V, D, J

Immunoglobulin heavy chain and Beta TCR have V D, J segments

Immunoglobulin light chain and Alpha TCR only have V, J

Immunoglobulin and TCR genes are only human genes requiring

rearrangement in order to be functional = rearranging genes

Gene rearrangement: functional exon encoding variable region

consists of one V segment join with one J segment ( cut and paste,

excess DNA removed as circle)

o Germline configuration before

o Somatic recombination occurs

o Rearranged gene: excised DNA circle, and rearranged gene

Proliferation of pathogen stimulated lymphocytes leads to many identical

clones of immunoglobulin or TCR == increases population

Clonal selection = pathogens select lymphocytes for expansion

Clonal expansion = proliferation of selected clones

Adaptive immune responses are initiated in secondary lymphoid tissues

by antigen-bearing dendritic cells and T cells

Lymph nodes, white pulp of spleen, Peyer’s patches of gut

First, dendritic cell carries pathogen from infected site to nearest

2ndary lymphatic tissue

o Dendritic cell = phagocytic leukocyte like macrophages, found

in tissues

Link between innate and adaptive immunity

Second, antigen loaded dendritic cell meets and activates TCR with

that antigen receptor

o Native T cells = those that have not met their antigen

o Effector T cells = T cells with receptors specific for bacterial

antigens will bind their antigens on dendritic surface where

they activate to divide and differentiate

Effector T cells needed to activate B cells

TCR recognize degraded fragments of pathogen proteins

TCR antigens recognize short peptides… this simplifies the process

Antigen processing = production of antigenic peptides from

pathogen proteins by degradation of dendritic cell

TCR recognize peptide antigens bound to human cell surface molecules

MHC molecules = pathogen derived peptides bound tby

glycoproteins

MHC = major histocompatibility complex = genes that encode MHC

molecules

o 1 MHC molecule binds 1 peptide = MHC

o ligand for TCR

o antigen presenting cells = MHC complexes on cell surface

that stretch out peptide on MHC to make accessible to peptide

binding site

o MHC can bind many different peptides, TCR only binds a few,

thus only small fraction of all peptides produced by pathogen

breakdown are presented by MHC to TCR as antigens

Dendritic cells express many different MHC molecules to increase

efficiency

MHC polymorphism (allele variability) is the basis for tissue type and

cause of transplanted organ rejection

Two classes of MHC molecule present peptide antigens to two types of T

cell

Two types of microorganisms that infect human tissues

o Extracellular pathogens

Bacteria… live and replicate in space between human

cells

o Intracellular pathogens

Viruses… live and replicated inside human cells

Two types of MHC to accommodate two types of microorganisms

o MHC class I – antigens from intracellular pathogens

Peptides produced by degraded in cytosol of

intracellular pathogens are delivered to ER

Cytotoxic T cells defend against intracellular infections

Cytotoxic T cell recognizes pathogen derived

peptide presented by an MHC class I molecule

Carry CD8 so only recognize MHC class I

o MHC class II – antigens from extracellular pathogens

Peptides produced by lysosomal degradation of

extracellular pathogens reside in endosomal vesicles

Helper T cells defend against extracellular infections

Enhance phagocytosis of extracellular pathogens

by macrophages and neutrophils

By direct contact with macrophage

Secretion of cytokines to drive macrophages

to higher state of activation

Helps B cells make antibodies that opsonize

Engulfment NOT interaction

Express CD4

All t cells have CD 4 or CD8… NOT BOTH

MHC I most cells in the body because cytotoxic T cells can attack

the vast majority of infected cells

Helper T cells attack extracellular infections so only interact with

dendritic cells, macrophages, and B cells

MHC class I molecules present antigens of intracellular origin to CD8 T

cells

Viruses are the only group of pathogens that Must be inside human

cells to replicate and survive

o Cytotoxic T cells important for viral infections

Viral proteins synthesized on ribosomes of dendritic cells and

degraded by cytosolic protease

o Peptides transported to ER so can bind MHC class I

o Bound to MHC I, leaves ER through Golgi to plasma

membrane

MHC class II molecules present antigens of extracellular origin to CD4 T

cells

1/3 T cells are CD8

CD4 T cells activated by antigen, differentiate into helper T cells =

effector CD4 T cells

o Can secrete many different cytokines

More versatile than cytotoxic CD8 T cells

o CD4 T cells respond to pathogens in fluid and spaces that

separate cells

Dendritic cells use cell surface receptors to endocytose pathogens

Helper t cells trave in lymph and blood to infected tissue where they

secrete cytokines that attract neutrophils and monocytes to tissue

Effector CD4 T cells help B cells become antibody producing plasma cells

B cell purpose = make antibodies

Plasma cell = activated B cell by pathogen that has proliferated,

differentiated into dedicated antibody factory

Antibodies target pathogen for destruction by phagocyte or effector

cell

Antibodies recognize all parts of antigen structure (entire)

Native B cells = those that have not yet encountered their antigen

Receptor mediated endocytosis = bacterium binds BCR which

causes engulfing… delivered to endocytic vesicles in B cell

cytoplasm

B cells can act as antigen presenting cells to CD4 T cells

Interacting T cell and B cell must both be specific for the same or

different parts of the same antigen

Extracellular pathogens and their toxins are eliminated by antibodies

Immunoglobulins produced by B cells and plasma cells -- 5 classes

o Differ in heavy chain constant region

o IgA

Made in lymphoid tissues… mucus surfaces

o IgD

Antigen receptors on circulating B cells that have yet to

encounter antigen

o IgE

Bind tightly to receptors on mast cell surface

Respond when parasites present

Allergic reactions

o IgG

Most important function: facilitate engulfment and

destruction of extracellular microorganism and toxins by

phagocytes

Neutrophils and macrophages have cell surface

receptors that bind IgG heavy chains

o IgM

Antigen receptors on circulating B cells that have yet to

encounter antigen

First antibody to be secrete in immune response

Humoral immunity = immunity due to antibodies and their actions

IgM, IgA, IgG = main antibodies present in blood, lymph and

intercellular fluid

Antibodies secreted into blood from lymph nodes, spleen, and bone

marrow

Antibodies reduce infection by binding tightly to site in pathogen to

inhibit pathogen growth, replication or interaction with human cells

= neutralization

Opsonization = coating of surface of pathogen with molecules that

make it more readily ingested by phagocytosis

Antibody quality improves during the course of an adaptive immune

response

Somatic hypermutation = nucleotide substitution throughout

variable regions of rearranged immunoglobulin heavy and light

chains

o Strengthens the antigen-binding properties of the antibodies,

isotype switching improves recruitment of effector functions

Isotype switching = changes the isotype of the immunoglobulin but

not the antigen binding site

o Germinal center B cells remove gene segments encoding the

mu constant region and bring the rearranged variable region

into juxtaposition with a gene segment encoding a different

constant region

IgM to IgG, IgA or IgE

Lymph nodes and spleen = IgM IgG

Peyer’s patches of gut and lymphoid tissues in mucosa

= IgM IgE

Enable eosinophils, basophils and mast cells to be

recruited to the adaptive immune response

Somatic hypermutation and isotype switching – adaptive immune

response selects for antibodies that bind more tightly to pathogen

and are more efficiently delivered to site of infection and are better

able to involve phagocytes and other inflammatory cells in

pathogen elimination

Immunological memory is a consequence of clonal selection

Once pathogen terminated

o Effector t cells apoptose

o Plasma cells persist

o Pathogen specific antibodies maintained in circulation for

years

Prevent reoccurrence

Memory cells come from clonal expansion

Secondary adaptive immune response leads to earlier detection

Memory B cells make better immunoglobulins than native B cells

because they benefit from somatic hypermutation and isotype

switching that occurred in the primary response

Clonal selection makes T cells and B cells tolerant of self and responsive

to pathogens

Fraction of developing B cells and T cells carry receptors that bind

to normal components of the healty human body

Tolerance to some important self antigens is set up during

lymphocyte development

Immature T cells undergo two rounds of clonal selection

o First positive selection

Identifices and selects T cell bearing antigen receptors

that work effectively with the particular subset of MHC

class I and II molecules the person has

T cells recognize peptide antigens in combination with

own self MHC

Useful T cells recognize receptors for self MHC

and peptide

Those that bind weakly to self MHC apoptose

o Second negative selection

Identifies T cells with receptors that bind too strongly to

self MHC

Apoptose

T cells in circulation are self tolerant

During B cell maturation in bine marrow, negative selection used to

prevent emergence of some cells that could make harmful

antibodies

Regulatory T cells = effector CD4 cells that suppress responses of T

cells reactive against healthy cells and tissues

Unwanted effects of adaptive immunity cause autoimmune disease,

transplant rejection and allergy

Chapter 4: Antibody structure and the Generation of B cell Diversity 9/14/13 12:35 PM

Antibodies – clear extracellular pathogens and their toxins

Antibodies = secreted B cell antigen receptors

Recognize all types of biological macromolecules

o Proteins and carbohydrates most common

Antibody + virus/bacterium particle = disabled virus/bacterium

Protective immunity

Secreted form of immunoglobulins

Antigen binds receptor proliferates and differentiates into plasma cell

Secrete large amts of antibodies with same specificity as membrane

bound immunoglobulin

Ex. Of clonal selection

Variable region contains site of antigen binding and specificity of antibody

Constant region interacts with their immune system components

Isotypes distinguished based on structural differences in constant region

and effector functions

Antibodies are composed of polypeptides with variable and constant

regions

Glycoproteins with four polypeptide chains

o Two heavy chains (H chains)

o Two light chains (L chains)

Light chain has only two isotypes kappa and lambda

Each light chain has kappa OR lambda, not both

2/3 human antibodies contain kappa

immunoglobulin chains are folded into compact and stale protein domains

antibodies function in extracellular environments

VL and VH together form antigen binding site

An antigen binding site is formed from the hypervariable regions of a

heave chain V domain and a light chain V domain

Antigen binding sites vary in shape and physical properties

Epitope = where antibody binds antigen

Antibodies that bind to the end of a polysaccharide/polypeptide

chain use a deep pocket formed between heavy chain and light

chain V domains

Multivalent antigen = antigen that has more than one epitope or

more than one copy of the same epitope

Conformational/discontinuous epitopes = parts of protein separated

in AA sequence that are brought together in a folded protein

The binding of antigens to antibodies is based solely on noncovalent

forces – electrostatic forces, hydrogen bonds, van der Waals forces,

and hydrophobic interactions

Affinities = different binding strengths

Catalytic antibodies = antibodies that catalyze chemical reactions

involving bound antigen

Monoclonal antibodies are produced from a clone of antibody-producing

cells

Anticera

o B cells isolated from immunized animal and immortalized by

fusion with a tumor cell to form hybridoma cell lines that

grown and produce antibodies indefinitely

o Hybridomas separated

Monoclonal antibodies are used as treatments for a variety of diseases

Chimeric monoclonal antibodies = combine mouse V regions with

human C regions

o Used to reduce rejection rate

Humanize

Generation of immunoglobulin diversity in B cells before encounter with

antigen

Immunoglobulin genes are in a fragmented form that cannot be

expressed

o All but B cells

Immunoglobulin heavy and light chain loci have families of gene

segments

o Germline inheritance

Before expression, genes must rearrange to become functional

o only occurs in B cell development

o In bone marrow

DNA sequence encoding a V region is assembled from two or three gene

segments

Heavy chain on chromosome 14

Lambda light chain on chromosome 22

Kappa light chain on chromosome 2

Different segments encode the leader peptide (L), V region (V) and

the constant (C) of heavy and light chain

o L and C have exons and introns

o V regions encoded by 2 (VL) or 3 (VH)

Light chain V region = variable (V) and joining (J) gene segments

Heavy chain VJ to diversity (D) gene segment

Difference in V gene segments are in the sequences that encode

the first and second hypervariable region

o Third hypervariable region determined by the junction

between V and J

B cell

o Only one light chain loci (kappa or lambda) functional light

chain genes

o Heavy chain = one V one J one D

First two = differences in sequences of heavy chain V

gene

Third = differences in D gene and junction with V and J

respectively

C of heavy chain = 1 C

Random recombination of gene segments produces diversity in the

antigen binding site of immunoglobulins

Somatic recombination = VDJ segment recombination

Light chain

o All that’s needed is single recombination of V and J

(lambda/kappa)

Heavy chain

o Two recombinations

D to J

DJ to V

Somatic recombination done with enzymes that cut and rejoin DNA

o Recombination signal sequences (RSS)

o Flank 3’ side of V segment, both sides of D segment, and 5’

side of J

o Two types of RSS and recombination can only occur between

different types

Heptamer

Nonamer

o RSS makes sure gene segments joined in correct order

Recombination enzymes produce additional diversity in the antigen

binding site

V(D)J recombinase = enzymes required for VDJ segment combos

2 component proteins made only by lymphocytes

o RAG1 (Recombination activating genes) and RAG2

RAG1 and RAG2 interact with each other and other proteins to form

the RAG complex

One RAG binds one type RSS and other binds another

Two RAGs align two RSSs and cleaves DNA at end of

immunoglobulin gene segments so DNA hairpin at each end

Broken ends joined by DNA repair enzymes

o Brings coding regions together