Transcript HLA Antibody Screening and Identification and Their Role in

Tissue Typing & Cross
Matching - Recent
Advances
Transplantation Immunology
 MHC and Tissue Matching
 Graft Rejection
 Immuno-suppression
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Blood Transfusion
 First attempts were unsuccesful (MISMATCH)
 Discovery of blood groups (Red cell antigens)
A-B Landsteiner 1900
Rh Levine, Stetson 1939
 Succesful transfusion = Transplantation
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Others: Bone, Tissue-engineering, etc
Transplantation
 Organ Transplantation
Classification of Renal Transplantation
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Auto-RT
Cadaveric
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Related
Allograft RT
Living Donor
Unrelated
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Xenograft RT (In experimental)
Transplantation History
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experimental kidney transplantation -1912
 Alexis Carel-Nobel prize
1935 human kidney transplant in Russia - rejection
P.B. Medawar (1945) skin grafts
 Self skin accepted
 Relative not accepted ! 
difference ?
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What is the
  Immunologic mechanism
A. Mitchison (1950)
 Lymphocytes are responsible for rejection
Transplantation History
• Peter Gorer (~1935)
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 Identification of 4 group of genes for RBC
Gorer and Gorge Snell (~1950)
 Group II antigens are responsible for rejection
  Major HistoCompatibility genes (HLA)
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 Nobel prize 1980 George Snell
1954 Succesful kidney transplant between identical
twins in Boston – Peter Bent Brigham Hospital
 Joseph Murray 1991 Nobel prize
HISTORY OF RT
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1933 First clinical RT (Voronov);
1954 First long-term successful RT(Twin);
1958 Discovery of HLA(Human Lym Antigen);
1959 Radiation be used for immunosuppression;
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1961 Azathioprine (Aza);
1962 Prednisolone; Tissue Matching;
1966 Cross-Matching;
Late 1960’ Preservation the Kidney>24hr ;;
1978 Clinical use of Cyclosporine(CsA).
Key factors for succesful transplantation
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Knowledge of MHC haplotypes
Effective immunosuppression
Ability to identify and treat infections
Available donors
Pre-OP Selection
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ABO Blood Group: Compatible;
Cytotoxicity Test:
Donor Lymphocyte
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Cross matching
Donor Lymphocyte
Donor Serum
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Recipient Serum
Recipient Serum
Recipient’s Lymphocyte
Mixed Lymphocyte Culture
Tissue typing (HLA)
Transplantation Immunology
Histocompatibility Antigens
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Major histocompatibility antigens
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MHC class I molecules : almost all nucleated cells
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MHC class II molecules : APCs, endothelium of renal
arteries and glomeruli
Minor histocompatibility antigens : H-Y molecule
Major histocompatibility antigens
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Human leukocytic Antigen
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HLA I. (α1, α2, α3, β2-microglobulin)
Gene-Code alleles: A, B, C loci
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HLA II. (α1, α2, β1, β2)
Gene-Code alleles: DP, DQ, DR loci
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HLA III.
Gene-Code alleles: C4A, TNF, HSP70
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MHC complex: Gene
Major histocompatibility antigens
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MHC loci are highly polymorphic
Many alternative alleles at a locus
The loci are closely linked to each other
A set of alleles is called a HAPLOTYPE
One inherites a haplotype from mother and another
from father
The alleles are codominantly expressed
Inheritance of MHC alleles
A/C
A/D
B/C
Mother
Father
A/B
C/D
B/D
A/R1
R2/C
R2/R1
Possible children of parents with HLA haplotype A/B and C/D
R1=C-D recombination
R2=A-B recombination
Induction of Immune Responses Against
Transplants
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alloantigens and xenoantigens : antigens that serve
as the targets of rejection
the antibodies and T cells that react against these
antigens are said to be alloreactive and
xenoreactive, respectively.
allorecognition
 direct
 indirect
Rejection
From: Kuby: IMMUNOLOGY (fourth edition, 2000)
Tissue typing
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Microcytotoxicity assay
 Known antibody to WBCs of donor / recipient
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 Complement mediated lysis if Ab present on cell
surface
Mixed lymphocyte culture (MLC)
 Irradiated donor lymphocytes (stimulants)
 Incubated with recipient lymphocytes
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3H
Thymidin incorporatin measured
Flow cytometry cross typing
DNA analysis
 Genomic typing (very precise, many subtipes)
From which animals are we able to
transplant organs
1. The Chimpanzee:
Its DNA sequence
differs from ours by
only 2%
2. The Baboon:
Its organs are too
small for a large
adult human
3. The Pig:
Surprisingly similar
to our anatomy and
physiology
Organ breeding:
•A transgenic animal carries a foreign
gene inserted into its genome.
•The transgenic animal shows the
specific characteristics which are
coded on the inserted gene
 A gene which is responsible for the
construction of a human organ
makes the organism produce the
organ additionally.
The insert of a foreign gene into an animal
I. DNA microinjection
The DNA is inserted into the cell
with a small syringe
II. Retrovirus gene transfer
The DNA is carried into a cell by
a virus.
Suppression of immune system rejection
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The genes which are responsible for the own tissue
not being rejected can be injected into an animal
embryo
the organs of which are then similar to the ones of
the human.
It is possible to humanize the bred organs by making
certain genetic modifications.
Then the organs are accepted by the immune
system.
Transplantation-background
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Worse outcome for Rh-mismatched recipients— Rh(D)-positive
donor into a Rh(D)-negative recipient— 12 months
posttransplant
Clinical transplants 1988. Los Angeles, UCLA Tissue Typing Laboratory1988, p 409.
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Rh(D) mismatch had a negative impact on long-term graft
survival in cadaveric renal transplantation
Transplantation 1998; 65: 588.
Solid organ transplantation ABO blood group compatibility,
but the Rh(D) compatibility is an relevant obstacle .
Transplantation – Graft survival
Transplantation - Patient survival
Transplantation: Conclusion
• Rh incompatibility is not detrimental in live-donor
renal transplantation.
Transfusion and Transplantation
Conclusion
• Pre-transfusion compatibility testing only ABO
grouping, antibody screening and a major crossmatch, and D typing being discontinued
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Rh incompatibility is not detrimental in
transplantation.
CONTRAINDICATIONS TO RENAL
TRANSPLANTATION
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ABO incompatibility.
Cystoxic antibodies against HLA antigens of donor.
Recent or metastatic malignancy.
Active infection.
AIDS.
Severe extrarenal disease (cardiac, pulmonary, hepatic).
Active vasculitis or glomeulonephritis.
Uncorrectable lower urinary tract disease.
Noncompliance.
Psychiatric illness including alcoholism and drug addiction.
Morbid obesity.
Age > 70 years.
Primary oxalosis.
Persistent coagulation disorder.
Matching between Recipient And Donor
A- Tissue typing
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Determined by 6 antigens located on cell surface encoded for by the
HLA gene located on the short arm of chromosome 6.
Class I antigens (HLA-A and HLA-B) are expressed on the surface of
most nucleated cells.
Class II antigen (HLA-DR) are expressed on surface of APC and
activated lymphocytes.
These 6 antigens are referred to as major transplant antigens.
The match between donor and recipient can range from 0 to six.
Matching between Recepient And Donor
B- Cross matching
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A laboratory test that determines weather a potential transplant recipient
has preformed antibodies against the HLA antigens of the potential donor.
(Donor Lymphocytes + Recipient Serum)
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A Final CM is mandatory
C- Compatible ABO blood group.
Structure of the HLA class I and class II antigens.
Organization of the human HLA genes on chromosome 6.
Effect Of HLA Matching On The Graft Outcome
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Data from large registries indicate that, the better the HLA-match, the
better the long-term survival of the allograft.
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The benefits of matching are particularly noteworthy in recipients of
kidneys from donors with zero mismatch.
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The benefits of lesser degrees of matching have become less obvious
with the use of newer and more potent immunosuppressive drugs.
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Matching for DR antigens are more favorable than others.
The beneficial effect of HLA B and DR matching in
patients with and without the benefit of cyclosporine.
Factors Influencing The Longivity Of
Renal Allograft
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Age
HLA matching
Delayed graft function
Ischemia time.
Number of acute rejection episodes.
Native kidney disease.
Ethnicity.
Others
Relative incidence of causes of allograft dysfunction
during the year following transplantation.
Immune responses to renal allograft
Recent advances
• Over the past several years, significant advances in
HLA antibody detection have occurred
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Solid-phase, multiplex testing platforms have
replaced traditional cell-based assays, and have
provided better sensitivity and specificity in antibody
detection
As a direct result of improved antibody identification,
many programs are moving into the realm of the
'virtual cross-match'
The Virtual Cross-match
• The virtual cross-match has proven to be successful
in renal, cardiac and lung transplantation, and has
resulted in a greater percentage of sensitized
patients gaining access to transplantation
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Virtual cross-matching allows patients better access
to transplantation and enhances patient care
Conclusion
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Transplantation provides us the means of
restoring the function of a nonfunctional organ.
In the case of BMT it enables us to administer
such high doses of chemotherapy that would
destroy the BM as well as the residual tumor.
A lot immunologic knowledge had to be collected
to understand what is happening.
Conclusion
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HLA typing and matching is essential for allograft
transplantation.
Effective immunosuppressive therapy
(Cyclosporin) revolutionised organ transplantation.
The future is to transplant cells, that would restore
the function of the affected organ.
Gene therapy is growing, and will cause another
revolution like cyclosporin did in the 1980s.