Chapter 16 - Transplantation Immunology

MHC = closely linked ___ on chromosome # __

genes; 6

MHC Class 1 proteins

HLA-A, HLA-B, HLA-C

all nucleated cells

MHC class 2 proteins

HLA-DR, HLA-DQ, HLA-DP

pAPCs

Autograft

tissue transfer in same individual

Syngeneic / Isograft

transfer of cells/tissues to a genetically identical individual

Allograft

graft between genetically nonidentical individuals of the same species

Xenograft

transplant between members of different species

Direct vs Indirect Allorecognition

Direct

• Tc cells bind foreign HLA Ag and release cytotoxic factors (perforins, granzymes)

Indirect

• Host pAPC present foreign MHC Ag on graft to host Th cells

Hyperacute

minutes to hours

Preformed Ab to ABO, HLA, endothelial Ag

bind to vascular endothelium and activate C’ and clotting factors

Accelerated

several days

Preformed Ab to ABO, HLA, endothelial Ag

bind to vascular endothelium and activate C’ and clotting factors

Acute

days to months

Th cytokines produce DTH

CD8 T cells mediate cytotoxicity; Ab bind to C’and induce inflammation

Chronic

1 year or more

DTH (TH cytokines)

GVHD

lymphoid cells in graft mount immune response against host HLA Ag

cytokine release, inflammation, destruction of various tissues

100 days post-transplant

acute/chronic

Corticosteroids

anti-inflammatory

Antimetabolites

interfere with lymphocyte maturation and division

Calcineurin inhibitors

block cytokines synthesis in T cells

Monoclonal Ab

interfere with T/B cell function by binding to lymph surface molecules

Polyclonal Ab

animal-derived Ab against human cells that deplete thymocytes or lymphocytes

HLA Typing - Complement-Dependent Cytotoxicity (CDC)

HLA Ag identified by their reactivity with panels of antisera

Pt serum incubated with panels of lymphs

Anti-human Ig can increase test sensitivity

% panel reactive Ab (%PRA) reported

PCR with Sequence Specific Primers

PCR-SPP

PCR amplification of HLA genes with panels of primer pairs, each of which amplifies specific alleles or related allele groups

PCR-Sequence-Specific-Oligonucleotide-Probe Hybridization

single PCR reaction that will amplify all HLA gene variants at a specific locus (referred to as a generic amplification). The amplified gene is then subjected to hybridization with a panel of DNA probes, each specific for a unique HLA allele or allele group. Only those probes that specifically hybridize to the amplified DNA will be detected

Sequence-Base-Typing

SBT

sequencing of PCR-amplified HLA genes

SBT can be carried out using Sanger dideoxy chain terminator sequencing. A generic amplification of the HLA gene of interest is conducted, followed by a sequencing reaction using dideoxy nucleotides. The dideoxy terminators are fluorescently labeled. Incorporation into the synthesized DNA molecule is detected using automated DNA sequencers with fluorescent detectors. The sequence of the target gene is compared with an HLA sequence database to determine the specific HLA allele for the patient.

HLA genotyping advantages over CDC

Higher resolution

Reagent quality

High throughput

HLA Ab may develop in response to

multiple blood transfusions, prior organ transplants, paternally derived fetal Ag by pregnant women

HLA Ab can cause

Hyperacute graft rejection

Donors with HLA Ag that can react with the Ab in a transplant candidate should be eliminated from consideration

ELISA testing for HLA Ab

purified HLA Ag bound to microtiter wells

incubated with serum

Crossmatching

donor-recipient CDC test done to confirm the absence of donor-specific Ab

Flow Cytometry for HLA Ab

Pt serum incubated with mixture of beads coated with different HLA proteins

washed, add FITC-labeled anti-IgG

Wash; analyze fluorescence by flow cytometry

Post-transplant testing

monitoring

HLA Ab testing

• solid organ transplants

• bead array assay

Chimerism

• HSC transplants

• PCR amplification of STRs