Tissue
Typing And HLA Matching
Tissue typing of transplant
recipients is required to assess their immunological profile in order to find
an optimally matched organ. This involves identifying their ABO blood group,
and determining which human leucocyte antigens (HLAs) their cells express.
These tests are performed as part of the transplant assessment process, well in
advance of the actual transplant.
ABO antigens
ABO antigens are carbohydrate molecules found on the surface of red blood
cells and endothelial cells. Group O individuals (who lack A and B antigens) develop antibodies to both A and B anti-
gens. This is thought to be driven by cross-reactivity with microbial antigens.
In group A individuals, B antibodies are present, while group B individuals
have A antibodies. AB individuals have no A or B antibodies. Transplantation of
an organ into an ABO-incompatible recipient, e.g. an organ from a group A donor
into a group O recipient, would lead to immediate binding of A antibodies to
graft endothelium, and to hyperacute rejection. Thus, the first step of tissue
typing is to ascertain the recipient’s blood group so that an ABO-compatible
donor can
be selected. There are two methods used to perform ABO typing.
1
Forward typing – the recipient’s
erythrocytes are mixed with anti-A or anti-B serum. If the erythrocytes express
A antigens, then agglutination of the cells will occur when incubated with
anti-A serum, etc.
2
Reverse typing – the recipient’s serum is
mixed with erythrocytes of known ABO type. This test is used to confirm the
results of forward typing. It can also be used to determine the quantity of ABO
antibodies present by performing serial dilutions of the recipient’s serum
prior to incubation with erythrocytes. The ABO titre gives a measure of the
concentration of ABO antibodies, and is quantified as the final dilution at
which agglutination takes place,
e.g. 1 in 32. The latter test is used in preparation for ABO-incompatible
transplantation to assess the requirement for antibody removal during
desensitisation (see Chapter 12).
HLA antigens
The human leucocyte antigens (HLA), also known as the major
histocompatibility complex (MHC) molecules, are a family of highly polymorphic
glycoproteins found on the surface of cells. They are divided into class I and
class II molecules. Class I molecules are found on the surface of all nucleated
cells and are composed of a polymorphic α chain combined with an invariant
subunit (β2 microglobulin). Intracellular protein antigens are processed and
presented on class I molecules to CD8 T cells (see Chapter 8). Class II
molecules are found only on the surface of antigen-presenting cells (APCs) and
are composed of two highly polymorphic subunits, an α-chain and a β-chain. APCs
internalise extracellular antigens, process them and load peptides onto class
II molecules for presentation to CD4 T cells (see Chapters 8 and 9).
HLAs are encoded by a cluster of genes on chromosome 6. In humans, there
are 3 HLA class I genes (A, B and C). These genes are extremely variable, and
encode highly polymorphic α-chains. More than 700 variants of the A gene, 1000
variants of the B gene and 400 variants of the C gene have been identified.
This variation makes it unlikely that an unrelated donor and recipient will
have exactly the same HLA antigen on the surface of their cells. Such extensive
genetic variability is unusual in the human genome and is thought to have
arisen as a strategy to prevent a single viral mutation (which might prevent
viral peptide being loaded onto class I molecules) from conferring virulence
against all humans, as there would likely be a class I variant in some
individuals in the population which could present the mutated viral peptide.
The HLA class II genes (DP, DQ and DR) are also found on chromosome 6,
and are more complex than the class I genes.
HLA-DP is encoded by a polymorphic α-chain gene (HLA- DPA1; >25
different alleles described) and a polymorphic β-chain (HLA-DPB1; >130
alleles described).
HLA-DQ is encoded by a polymorphic α-chain gene (HLA- DQA1; >30
alleles described) and a polymorphic β-chain (HLA- DQB1; >90 alleles
described).
HLA-DR is encoded by a polymorphic α-chain gene (HLA- DRA; three
alleles described) and four highly polymorphic β- chain genes (HLA-DRB1, B3, B4
and B5; >600 variants described). The DRB1 gene encodes the β-chain of the
classical DR class II molecule. The most commonly observed DR antigen in UK
donors (arising from variants of the DRB1-β and DRα genes) is the DR4 antigen
(present in 35% of donors). The DRB3, 4 and 5 genes also encode β-chains that
can complex with the DR α- chain, and give rise to the HLA-DR52, 53 and 51 antigens
respectively.
HLA nomenclature
Two parallel systems of nomenclature are applied to HLA antigens.
1
Serological – this was the initial
system used to name HLA antigens based on their reactivity to standardised
antisera. In transplantation, 55 HLA-A, B and DR antigens are defined based on
reactivity to a set of broad antisera. Some of these antigens can be subdivided
using more specific antisera (e.g. HLA-A10 can be split into HLA-A25(10) and
HLA-A26(10)).
2
DNA sequence – advances in molecular
biology have allowed the specific sequences of different HLA genes to be
determined. Allele names are prefixed with a ‘*’; for example, the alleles
encod- ing the HLA-A3 antigen are named A*03. Different A3 alleles are then given
different numbers, e.g. A*0301, A*0302, etc.
In clinical practice in solid organ transplantation, HLA type is now
determined by DNA sequencing.
HLA matching
Given that an individual has two copies of each HLA gene, the maximum
number of mismatches that can occur between a donor and recipient is 12, i.e.
two A mismatches, two B mismatches, etc. However, in renal transplantation only
A, B and DR mismatches are considered, so the maximum number of mismatches
possible is six. Such a mismatch would be described as a 2-2-2 mismatch (2 A, 2
B and 2 DR mismatches). The best mismatch would be a 0-0-0 mismatch. The more
mismatches that are present, the more likely that the allograft will be
recognised as foreign and rejected. This is reflected in allograft survival
data which suggest that 80% of those patients receiving a 0-0-0 mismatched
kidney will still have a functioning allograft at 5 years compared with 60% of
those receiving a 2-2-2 mismatched kidney. DR mismatches are more significant
than A or B mismatches, therefore every effort is made id DR mismatches.
