Priniciples of transplantation
February 19 2008
Goals and objectives
- Principal components
- Role of HLA in immunogenic response
- Understand the 3 signal pathway of T cell
activation and its clinical significance
Classification of grafts
* Autologous grafts
Grafts transplanted from one part of the body to another in the same
individual
* Syngeneic grafts (Isografts)
Grafts transplanted between two genetically identical individuals of
the same species
Allogeneic grafts (Allografts)
Grafts transplanted between two genetically different individuals of
the same species
Xenogeneic grafts (Xenografts)
Grafts transplanted between individuals of different species
2006-7year
Immunology
4
IMMUNE RESPONSES TO TRANSPLANTED TISSUES
* Transplant rejection caused by genetic differences between donor and recipient
* HLA and blood group antigens
*
Alloantigens
* Antigens which vary between members of same species
*
Alloreaction
* Immune response to an alloantigen
*
Alloreactions in transplantation
* Host-versus-graft (transplant rejection)
* Graft-versus-host
Figure 12-11
Effectors of rejection
*
*
*
*
Major players:
T Cells
B cells
Antigen presenting cells
MHC (Most important)
T cells
* Arise in thymus from bone marrow derived precursors
* Each T-Cell has unique T Cell receptor (Clone)
* Selection
Positive
Negative
* Subtypes
CD 4 T cells – Antigen specific immune response
CD8 T cells - Precursors of CTL – Class I MHC
B cells
*
*
*
*
Arise and mature in bone marrow
Negative selection
Express BCRs on their surface
When BCR is stimulated the B cell secrete
antibodies of same specificity as their BCRs
Antigen presenting cells
* Most important
* Activate T cells
* Endocytose antigen and display it on MHC
molecules
* T cells recognize and interact with antigen MHC
to become activated
MHC complex
* Encode molecules crucial to the initiation and
propagation of immune response
* The HLA complex on chromosome 6 contains
over 200 genes, more than 40 of which encode
leukocyte antigens
* The HLA genes that are involved in the immune
response fall into two classes, I and II, which are
structurally and functionally different
Location and Organization of the HLA Complex on Chromosome 6
Klein J and Sato A. N Engl J Med 2000;343:702-709
Types of MHC
* There are three classes of MHC molecules.
* Class I- encodes glycoproteins expressed on the surface of nearly all nucleated cell; the major
function of the class I gene is presentation of peptide antigens to cytotoxic T-cells
* Class II- encodes glycoproteins expressed primarily on antigen-presenting cells, examples:
macrophages, dendritic cells and B-cells, where they present processed antigenic peptides to
T helper cells.
* Class III- encodes various secreted proteins that have immune function including components
of the complement system; C2,C4, Factor B, &TNF, and molecules involved in
inflammation.
Nucleated cells
Class I MHC
RBCs
Class II MHC
APCs
Function of MHC
* The function of both class I and class II molecules is
the presentation of short, pathogen-derived peptides to
T cells, a process that initiates the adaptive immune
response
* Class I - Sample cytosolic proteins and detect foreign
proteins that would indicate an intracellular pathogen
such as virus or intracellular bacteria
* Recognised by CD 8 T cells and provide a
surviellance mechanism to target infected cells for
destruction
Antigen Processing and Presentation
Biological functions of Class I and Class II molecules
*
http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/H/HLA.html#cd8
Class I
* Present peptides derived from
endogenously synthesized proteins
* Responding T cells express CD8+
* Class II system is designated to sample
extracellular proteins by extracellular proteins by
specialized APC’s
* Class II are recognized by CD 4 helper T cells and
allow for the generation of immune response to
invading pathogens
Antigen Processing and Presentation
Biological functions of Class I and Class II molecules
*
Class II
* Present peptides derived from
exogenously synthesized proteins
* Responding T cells express CD4+
http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/H/HLA.html#class_II
Class I
* The class I genes code for the polypeptide chain of the
class I molecule; the ß chain of the class I molecule is
encoded by a gene on chromosome 15, the beta2microglobulin gene.
* There are some 20 class I genes in the HLA region; three
of these, HLA-A, B, and C, the so-called classic, or class Ia
genes, are the main actors in the immunologic theater
Structure of Class I MHC
*
Two polypeptide chains, a long α chain and
a short β (β2 microglobulin)
*
Four regions
* Cytoplasmic region containing sites for
phosporylation and binding to
cytoskeletal elements
* Transmembrane region containing
hydrophobic amino acids
Structure of Class I MHC
*
*
Four regions
* A highly conserved α3 domain to
which CD8 binds
* A highly polymorphic peptide binding
region formed from the α1 and α2
domains
Β2-microglobulin helps stabilize the
conformation
Structure of HLA Class I and Class II Molecules
Klein J and Sato A. N Engl J Med 2000;343:702-709
Class II
* The class II genes code for the alpha and ß polypeptide
chains of the class II molecules .
* The designation of their loci on chromosome 6 consists of
three letters: the first (D) indicates the class, the second (M,
O, P, Q, or R) the family, and the third (A or B) the chain (
or ß, respectively).
* HLA-DRB, for example, stands for class II genes of the R
family coding for the ß chains.
Structure of Class II MHC
*
*
Two polypeptide chains,α and β, of
roughly equal length
Four regions
* Cytoplasmic region containing sites
for phosporylation and binding to
cytoskeletal elements
Structure of Class II MHC
*
Four regions
* Transmembrane region containing
hydrophobic amino acids
* A highly conserved α2 and a highly
conserved β2 domains to which CD4
binds
* A highly polymorphic peptide binding
region formed from the α1 and β1
domains
Structure of HLA Class I and Class II Molecules
Klein J and Sato A. N Engl J Med 2000;343:702-709
Important aspects of MHC
*
*
*
Normally, the proteins that undergo recycling are the organism's own, but in infected
cells, proteins originating from the pathogen are also routed into the processing
pathways.
With the exception of jawed vertebrates, no organisms appear to make a distinction
between peptides derived from their own (self) proteins and those derived from foreign
(nonself) proteins.
Jawed vertebrates, by contrast, use the peptides derived from foreign (usually microbial)
proteins to mark infected cells for destruction
Important aspects of MHC
*
*
Protein processing and loading of peptides onto class I molecules are taking place all the
time in most cells. There is always plenty of material to feed the processing machinery,
because worn-out, damaged, and misfolded proteins are continuously being degraded
and replaced by new ones.
By contrast, the processing of exogenous proteins and the loading of peptides onto class
II molecules are normally restricted to B cells, macrophages, and dendritic cells, which
are very efficient in taking up material by endocytosis or phagocytosis.
Important aspects of MHC
*
*
*
*
The consequence of protein processing is that the surfaces of cells become adorned with
peptide-laden HLA molecules, amounting on a per cell basis to roughly 100,000 to
300,000 class I or class II products of each of the highly expressed HLA loci.
Since each HLA molecule has one peptide bound to it, each uninfected cell displays
hundreds of thousands of self peptides on its surface.
Each cell thus displays a heterogeneous collection of peptides, and the surface of a cell
resembles rows of well-stocked stalls at a bazaar, with bargain hunters scrutinizing the
wares.
But if, in this metaphor, the vendors are the HLA molecules and the peptides the goods,
who are the potential buyers? They are a group of lymphocytes reared in the thymus and
then turned loose to roam the body — the T cells.
Functions and Characteristics of HLA
*
HLA’s are cell-surface proteins involved in the recognition of self and non-self by the
immune system
*
HLA’s present foreign antigens to the immune system – resistance to viral and bacterial
pathogens
*
HLA’s are codominantly expressed
*
Highly polymorphic and polygenic
HLA genes are co dominant: A protein from
each parental gene is expresed on cellsurfaces
Polymorphism and polygeny
*
MHC genes are polymorphic: that is, there
are large numbers of alleles for each gene
*
MHC genes are polygenic: that is, there are
a number of different MHC genes.
Crossing over
results in new haplotypes
Structure of Class I MHC
Variability map of Class 1 MHC α Chain
Class I polymorphism
Locus
HLA - A
Number of alleles
(allotypes)
451
HLA - B
782
HLA - C
238
There are also HLA - E,
HLA - F and HLA - G
Relatively few alleles
Structure of Class II MHC
Variability map of Class2 MHC β Chain
Class II polymorphism
Locus
Number of alleles
(allotypes)
HLA - DPA
HLA - DPB
147
HLA - DQA
HLA - DQB
105
HLA - DRA
HLA - DRB1
HLA – DRB3
HLA – DRB4
HLA – DRB5
525
There are also HLA - DM and HLA - DO
Relatively few alleles
Why polymorphic?
* Multiple alleles of HLA in a population increases the
likelihood that the population will survive a pathogen
threat
* Unfortunately, it also cause histoincompatibility in organ
and tissue transplants
Important Aspects of MHC
* Primary HLA products that contribute to rejection are the
most polymorphic including HLA- A, - B and DR
* Efforts are made to match HLA-A, - B and DR genes and
proteins in kidney transplantation
HLA profiles
* Tissue typing
* Cross matching test
* Panel reactive antibodies
Tissue typing
* Helps to identify two alleles at each of the three loci
* One allele from mother and one from father
* Mother/Father: 25% chance of full match
* One Sibling: 25 % chance of full match
* Two Siblings: 44 % chance of full match
* HLA matching
3 year graft surivival 93 and 85% for HLA matched and mismatched
live donors
In cadaveric grafts 82 and 76%
Most benefit with zero mismatches
Cross matching test
* Serum of potential recipient is incubated with cells from
possible donor
* If recipient has antidonor antibodies there is a strong
likelihood that recipient would destroy transplant by
antibody mediated rejection
Panel reactive antibodies
* Anti HLA antibodies in the serum of a person can be
assessed as PRAs
* Testing the serum of the patient against a panel of cells or
antigens prepared from many different donors using
cytotoxicity or flow cytometry
* Results are expressed as percentage of positive donors
Response to antigenic stimulus
T cell activation
* Activation of T cell is a crucial step in generation of immune response
to specific antigens
* Naive T cells restricted to SLOs
* They interact with DCs that have migrated from the periphery in
response to infection or injury
* Once naïve T cells encountered their cognate antigen presented on
mature DCs, they become activated.
* Following activation CD 4 T cells help B cells to convert to plasma
cells
* Plasma cells produce antibodies
Generation of T cell effector function
* Alloreactive T cells can be found in both naive and memory T cells
* Naive T cells may be triggered by donor or recipient APCs to
proliferate and develop effector functions in SLOs
* Memory cells can be activated in the same manner or by recognizing
cells in allograft directly
* Reactions mediated by naïve T cells take longer to develop than those
mediated by memory T cells
Structure of the T cell Receptor
* Heterodimer with one α
and one β chain of
roughly equal length
* A short cytoplamic tail
not capable of
transducing an activation
signal
* A transmembrane region
with hydrophobic amino
acids
Structure of the T cell Receptor
* Both α and β chains
have a variable (V) and
constant (C) region
* V regions of the α and β
chains contain
hypervariable regions
that determine the
specificity for antigen
Structure of the T cell Receptor
* Each T cell bears TCRs
of only one specificity
(allelic exclusion)
TCR and CD3 Complex
* TCR is closely
associated with a
group of proteins
collectively called the
CD3 complex
*
*
*
*
γ chain
δ chain
2 ε chains
2 ξ chains
* CD3 proteins are
invariant
Role of CD3 Complex
* CD3 complex
necessary for cell
surface expression of
TCR during T cell
development
* CD3 complex
transduces signals to
the interior of the cells
following interaction
of Ag with the TCR
The “Immunological Synapse”
* The interaction between
the TCR and MHC
molecules are not strong
* Accessory molecules
stabilize the interaction
* CD4/Class II MHC or
CD8/Class I MHC
* CD2/LFA-3
* LFA-1/ICAM-1
The “Immunological Synapse”
* Specificity for antigen
resides solely in the TCR
* The accessory molecules
are invariant
* Expression is increased
in response to cytokines
The “Immunological Synapse”
* Engagement of TCR and
Ag/MHC is one signal
needed for activation of T
cells
* Second signal comes from
costimulatory molecules
* CD28 on T cells interacting
with B7-1 (CD80) or B7-2
(CD86)
* Others
* Costimulatory molecules are
invariant
* “Immunological synapse”
Costimulation is Necessary for T Cell Activation
* Engagement of TCR and
Ag/MHC in the absence of
costimulation can lead to anergy
* Engagement of costimulatory
molecules in the absenece of
TCR engagement results in no
response
* Activation only occurs when
both TCR and costimulatory
molecules are engaged with
their respective ligands
* Downregulation occurs if
CTLA-4 interacts with B7
* CTLA-4 send inhibitory signal
Clonal expansion
* Signals 1 and 2 activate calcium calcineurin pathway,
MAP kinase pathway and NF-Kb pathway
* These pathways activate transcription factors that trigger
the expansion of many new molecules including IL-2,
CD 154 and CD 25.
* IL-2 and other cytokines activate the TOR pathway to
provide signal 3, the trigger for cell proliferation
* A subset of activated helper T cells migrate to the B region
of lymph nodes located in the cortex and help to
differentiate B cells while the remainder of the effector T
cells leave the lymph node and proceeds the inflamed site
* The activated T cells rapidly accumulate in the interstitium
of the allograft as the response mounts in the first few
days.
* CD4 T cells are cytokine secreting cells that express IL-2
and alter a variety of cytokines
* CD4 cells help B cells to enhance their antibody
production through CD40 ligand
* Alloantibody produced during rejection is mainly IG g and
primarily participates in the destruction of vascular
endothelium of the graft
* CD 8 T cells participate in rejection through DTH or
cytotoxicty
Key Steps in T cell Activation
* APC must process and present peptides to T cells
* T cells must receive a costimulatory signal
* Usually from CD28/B7
* Accessory adhesion molecules help to stabilize binding of
T cell and APC
* CD4/MHC-class II or CD8/MHC class I
* LFA-1/ICAM-1
* CD2/LFA-3
* Signal from cell surface is transmitted to nucleus
* Second messengers
* Cytokines produced to help drive cell division
* IL-2 and others
Rejection
1. Hyperacute rejection
* Occurrence time
* Occurs within minutes to hours after host blood vessels are
anastomosed to graft vessels
* Pathology
* Thrombotic occlusion of the graft vasculature
* Ischemia, denaturation, necrosis
ORIGINS OF ANTIBODIES TO HLA AND ABO ANTIGENS IN HYPERACUTE
REJECTION
* Pregnancy
* Fetus is allograft in mothers body
* During birth, fetal cells can stimulate maternal immune response
* Blood transfusion
* HLA typing not performed for routine transfusion
* Leukocytes and platelets in whole blood
* Transplantation
* Persons with more than one transplant
Figure 12-15
* Complement activation
* Endothelial cell damage
* Platelets activation
* Thrombosis, vascular occlusion, ischemic damage
2006-7year
Immunology
77