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
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
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, morethan 40 of which encode leukocyte antigens • The HLA genes that are involvedin the immune response fall into two classes, I and II, whichare 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 IImolecules is the presentation of short, pathogen-derived peptidesto 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
Biological functions of Class I and Class II molecules • Class I • Present peptides derived from endogenously synthesized proteins • Responding T cells express CD8+ http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/H/HLA.html#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
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 • Theclass I genes code for the polypeptide chain of the class Imolecule; the ß chain of the class I molecule is encodedby a gene on chromosome 15, the beta2-microglobulin gene. • There are some 20 classI genes in the HLA region; three of these, HLA-A, B, and C,the so-called classic, or class Ia genes, are the main actorsin 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 polypeptidechains of the class II molecules . • The designationof their loci on chromosome 6 consists of three letters: thefirst (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 genesof 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 undergorecycling are the organism's own, but in infected cells, proteinsoriginating from the pathogen are also routed into the processingpathways. • With the exception of jawed vertebrates, no organismsappear to make a distinction between peptides derived from theirown (self) proteins and those derived from foreign (nonself)proteins. • Jawed vertebrates, by contrast, use the peptides derivedfrom foreign (usually microbial) proteins to mark infected cellsfor destruction
Important aspects of MHC • Protein processing and loading of peptides onto class I moleculesare taking place all the time in most cells. There is alwaysplenty of material to feed the processing machinery, becauseworn-out, damaged, and misfolded proteins are continuously beingdegraded and replaced by new ones. • By contrast, the processing of exogenous proteins and the loadingof peptides onto class II molecules are normally restrictedto B cells, macrophages, and dendritic cells, which are veryefficient in taking up material by endocytosis or phagocytosis.
Important aspects of MHC • The consequence of protein processing is that the surfaces ofcells become adorned with peptide-laden HLA molecules, amountingon a per cell basis to roughly 100,000 to 300,000 class I orclass II products of each of the highly expressed HLA loci. • Since each HLA molecule has one peptide bound to it, each uninfectedcell displays hundreds of thousands of self peptides on itssurface. • Eachcell thus displays a heterogeneous collection of peptides, andthe surface of a cell resembles rows of well-stocked stallsat a bazaar, with bargain hunters scrutinizing the wares. • Butif, in this metaphor, the vendors are the HLA molecules andthe peptides the goods, who are the potential buyers? They area group of lymphocytes reared in the thymus and then turnedloose 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 cell-surfaces
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.
Structure of Class I MHC Variability map of Class 1 MHC α Chain
Structure of Class II MHC Variability map of Class2 MHC β Chain
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