Transplantation Xiang Li, Urology Department West China

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Transplantation

• Xiang Li, Urology Department
• West China Hospital, Sichuan University

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Acknowlegement

• To Dr. Lu Yiping and Dr. Wang jia
• To other Colleagues working on renal and liver
  transplantation

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Transplantation is a Dream?

• Dream of Paranoia
• Dream of excellent surgeon who wants to excel
  himself.
• Dream of excellent scientist who believe nothing
  is impossible.

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Can you imagine?
Can you imagine?
Can you imagine?
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Contents

• Basic concepts of transplantation
• Clinical Organ transplantation
• Renal Transplantation, RT
• Transplantation Immunology
• MHC and Tissue Matching
• Graft Rejection
• Immunosuppression

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Definition of Transplantation

• Implantation of non-self tissue into the body
• the process of taking cells, tissues, or organs
  called a graft (transplant), from one part or
  individual and placing them into another (usually
  different individual).
• donor the individual who provides the graft.
• recipient or host the individual who receives
  the graft.

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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
• Others Bone, Tissue-engineering, etc
• Transplantation
• Organ Transplantation

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Types

• Autologous graft (autograft) within an
  individual, autotransplantation
• Syngeneic graft (syngraft, isograft) identical
  twins, isotransplantation
• Allogeneic graft (allograft, homograft)
  non-identical, allotransplantation
• Xenogeneic graft (heterologous graft,
  heterograft) between species,
  xenotransplantation

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Classification of Renal Transplantation

• Auto-RT
• Cadaveric
• Allograft RT Living
  related
• Living Donor
• 
  Living unrelated
• Xenograft RT (In experimental)

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Transplantation History

• 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 ! ? What is the
  difference ?
• ? Immunologic mechanism
• A. Mitchison (1950)
• Lymphocytes are responsible for rejection

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Transplantation History

• Peter Gorer (1935)
• Identification of 4 group of genes for RBC
• Gorer and Gorge Snell (1950)
• Group II antigens are responsible for rejection
• ? Major HistoCompatibility genes (HLA)
• Nobel prize 1980 George Snell
• 1954 Succesful kidney transplant between
  identical twins in Boston Peter Bent Brigham
  Hospital
• Joseph Murray 1991 Nobel prize

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HISTORY OF THE RT

• 1933 First clinical RT (Voronov)
• 1954 First long-term successful RT(Twin)
• 1958 Discovery of HLA(Human Lym Antigen)
• 1959 Radiation be used for immunosupp-
• ression
• 1961 Azathioprine (Aza)
• 1962 Prednisolone Tissue Matching
• 1966 Cross-Matching
• Late 1960 Preservation the Kidneygt24hr
• 1972 First successful RT(LRD) in china
• 1978 Clinical use of Cyclosporine(CsA).

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Key factors for succesful transplantation

• Knowledge of MHC haplotypes
• Effective immunosuppression
• Ability to identify and treat infections
• Available donors

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Applications of allografting transplantation
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The importance of transplantation
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Clinical Organ Transplantation

• Liver Transplantation
• Renal Transplantation

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LIVER TRANSPLANTATION

• Indication End stage liver diseases (ESLD)
• Hepatic Disease to ESLD
• Congenital malfomation
• Congenital liver metabolic disorders
• Acute liver failure
• Chronic liver failure
• (1) Cirrhosis Hepatitis B, Alcoholic
• (2) Parasites Hydatid disease of liver, ect.
• liver malignance

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END STAGE RENAL DISEASES (ESRD)
RENAL TRANSPLANTATION

• Definition
• (1) Various causes
• (2) Irreversible injury
• (3) Functional failure.
• Morbidity
• Europe 50/million
• China 90-100/million

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TREATMENT OF ESRD

• DIALYSIS
• Chronic Ambulatory Peritoneal Dialysis
  (CAPD)
• Hemodialysis (HD).
• KIDNEY TRANSPLANTATION

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Renal Transplantation

• Renal transplantation is associated with as
  survival benefit for patients with ESRD when
  compared to dialysis
• Even marginal donor kidneys confer a significant
  survival advantage over maintenance dialysis.
• The preferred therapy for most of the Pts with
  ESRD
• More cost- effective Better survival Better
  life quality.

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CONTRAINDICATION

• Active invasive infection
• Active malignance
• High probability of operative mortality
• Unsuitable anatomic situation for technical
  success
• Severe psychological or financial problem.

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Pre-OP Selection

• ABO Blood Group Compatible
• Cytotoxicity Test
• Donor Lymphocyte Recipient Serum
• Cross matching
• Donor Lymphocyte Recipient Serum
• Donor Serum Recipients Lymphocyte
• Mixed Lymphocyte Culture
• Tissue typing (HLA)

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OPERATION

• DONOR
• (1) Living donor
• Nephrectomy via flank approach
• Nephrectomy via Laparoscope.

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• (2) Cadaveric Donor
• Total midline incision
• in situ flashing Euro-collins/UW solution
• Bilateral radical nephrectomy.
• Low temperature preservation.

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Potential Advantages of living versus cadaveric
kidney donor

• Better short-term result(about 95 versus 90
  1-yr function)
• Better long-term results(half-life of 12-20 yr
  versus 8-9 yr)
• More consistent early function and easy of
  management

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Potential Advantages of living versus cadaveric
kidney donor

• Avoidance of brain death stress
• Minimal incidence of delayed graft function
• Avoidance of long wait for cadaveric transplant

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Potential Advantages of living versus cadaveric
kidney donor

• Capacity of time transplantation for medical and
  personal convenience
• Immunosuppressive regime may be less aggressive
• Help relieve stress on national cadaver donor
  supply
• Emotional gain to donor.

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Potential disadvantages of live donation

• Psychological stress to donor and family
• Inconvenience and risk of evaluation
  process(i.e., intravenous contrast)
• Operative mortality(about 1 in 2000 Pts.)
• Major post operative complications (about 2 of
  Pts.)

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Potential disadvantages of live donation

• Minor postoperative complications(up to 50 of
  Pts.)
• Long-term morbidity(possible mild hyper-tention
  and proteinuria)
• Risk for traumatic injury to remaining kidney
• Risk for unrecognized covert chronic renal
  disease.

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Recipient Operation
Extraperitoneally in the contralateral iliac
fossa via Gibson incision.
Why contralateral ?
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RECIPIENT OPERATION

• Blood Vessel Anastomosis
• Donor renal V Recipientsexternal iliac V
• Donor renal A Recipients internal iliac
  A
• Ureter Anastomosis
• Donor ureter Recipients bladder
• Anti-reflux anastomosis

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Clinical phases of rejection

• Hyperacute rejection (minutes to hours)
• Preexisting antibodies to donor HLA antigens
• Complement activation, macrophages
• Accelerated rejection
• Acute rejection (around 10 days to 30 days)
• Cellular mechanism (CD4, CD8, NK, Macrophages)
• Chronic rejection (months to years !!)
• Mixed humoral and cellular mechanism
• CHRONIC REJECTION IS STILL HARD TO MANAGE !

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IMMUNOSUPPRESSION

• Immunosuppresents play a very impor-tant role in
  organ transplantation
• Immuosuppresents extremely increase the effect
  and the survival rate of organ transplantation

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IMMUNOSUPPRESSION

• Immunosuppresents are a double - edged sword
• the most important thing is to increase their
  positive effects, and in the same time decrease
  their side effects (i.e., organ toxicity,
  infection, tumors, ect.).

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Diagnosis of rejection

• Symptom/Sign
• fever
• urinary output ?
• graft tenderness
• graft size ?
• hypertension
• myalgia/arthragia.

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Laboratory Test

• Serum creatine, SCr
• Urinary creatine, Ucr
• Color doppler scan
• radiorenogram
• Ateriogram
• Biopsy
• (1) Fine needle aspiration biopsy
• (FNAB)
• (2) Core needle biopsy(CNB).

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Treatment of kidney rejection

• Hyperacute (Sometimes during the operation !)
• No therapy, usually results in graft failure
  kidney should be removed
• Acute (Most frequently in the first 4 weeks)
• BIOPSY !
• Increase immunosuppression
• Increase steroid dose
• Increase cyclosporin (monitor serum level !)
• ATG, ALG, OKT3
• Chronic
• ACE inhibitors, prostacyclin analog drugs
• Steroid, Imuran, Cellcept

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Transplantation Immunology
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Histocompatibility Antigens

• Major histocompatibility antigens
• MHC class I molecules almost all nucleated
  cells
• MHC class II molecules APCs, endothelium of
  renal arteries and glomeruli
• Minor histocompatibility antigens H-Y molecule

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Major histocompatibility antigens

• Human leukocytic Antigen
• HLA I. (a1, a2, a3, ß2-microglobulin)
• Gene-Code alleles A, B, C loci
• HLA II. (a1, a2, ß1, ß2)
• Gene-Code alleles DP, DQ, DR loci
• HLA III.
• Gene-Code alleles C4A, TNF, HSP70
• MHC complex Gene

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Major histocompatibility antigens

• 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

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Inheritance of MHC alleles
Mother
Father
A/B
C/D
A/C A/D B/C B/D
A/R1 R2/C R2/R1
Possible children of parents with HLA haplotype
A/B and C/D
R1C-D recombination R2A-B recombination
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Induction of Immune Responses Against Transplants

• 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

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Rejection

• Senzitization stage
• Not needed in hyperacute reactions !
• Effector stage
• Alloantibodies bind to endothelium, activate the
  complement system, and injure graft blood vessels

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Rejection

• Effector stage (Mostly cellular mechanism)
• Alloreactive T cells recruit and activate
  macrophages ---gt DTH response delayed type
  hypersensitivity
• Alloreactive CTLs CTL mediated cytotoxicity lyse
  graft endothelial and parenchymal cells directly
• ADCC
• CD4, CD8 lymphocytes, NK cells, macrophages

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Rejection
From Kuby IMMUNOLOGY (fourth edition, 2000)
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Tissue typing

• Microcytotoxicity assay
• Known antibody to WBCs of donor / recipient
• Complement mediated lysis if Ab present on cell
  surface
• Mixed lymphocyte culture (MLC)
• Irradiated donor lymphocytes (stimulants)
• Incubated with recipient lymphocytes
• 3H Thymidin incorporatin measured
• Flow cytometry cross typing
• DNA analysis
• Genomic typing (very precise, many subtipes)

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Hyperacute Rejection

• Occurs within minutes of transplantation
• Pre-existing IgM (natural) antibodies against
• ABO blood group antigens
• less well-characterized antigens in xenograft
• alloantigen, such as foreign MHC molecules, or
  alloantigen expressed on vasular endothelial
  cells

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Hyperacute Graft Rejection
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Acute Rejection

• Occurs within days or weeks after transplantation
• Acute vascular rejection
• Necrosis of cells of the graft blood vessels
  (vasculitis)
• Mediated by IgG antibodies against endothelial
  cell alloantigens and complement activation

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Acute Rejection

• Acute cellular rejection
• Necrosis of parenchymal cells with lymphocyte and
  macrophage infiltrates
• Effector mechanisms
• CTLs
• Activated macrophages
• Natural killer cells

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Acute Cellular Rejection
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Chronic Rejection

• Occurs over months or years
• Fibrosis with loss of normal organ structures
• Wound healing following the cellular necrosis
• A form of chronic DTH or a response to chronic
  ischemia caused by injury to blood vessels

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Prevention and Treatment of Allograft Rejection1

• Immunosuppression
• drugs that inhibit or kill T lymphocytes
• toxins that kill proliferating T cells
• antibodies that deplete or inhibit T cells
• anti-inflammatory agents

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Prevention and Treatment of Allograft Rejection2

• Reduce the immunogenicity of allografts
• ABO blood group typing
• HLA typing and matching
• induce donor-specific tolerance
• blood transfusion

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Graft-versus-host Disease (GVHD)

• Occurs in bone marrow recipients
• Initiated by T cell recognition of host
  alloantigens
• The effector cells are less well defined NK
  cells, CD8 CTLs, cytokines

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Acute GVHD

• Epithelial cell necrosis
• Skin
• Liver
• The gastrointestinal tract
• Characterized by skin rash, jaundice and diarrhea

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Acute GVH
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Immunosuppressive therapy 1.

• Allogenic transplantation always require
  immunosuppressive therapy
• Most of the drugs available are non-specific
• Common side effects of therapy
• Infection
• Cancer
• Bone-marrow depression

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Immunosuppression 2.

• Conventional drugs (1st phase)
• Steroids /Prednisone/ (0.1-10 mg / kg)
• Azathioprine /Imuran/ (0.5-3 mg/ kg)
• Cyclophosphamide (0.5-20 mg/ kg)
• Methotrexate (0.1-0.3 mg/ kg)

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Immunosuppression 3.

• New drugs (1)
• CYCLOSPORIN A (REVOLUTION !)
• 2-8 mg/ kg
• FK506 tacrolimus /PROGRAF/
• Sirolimus - rapamycin
• Gusperimus - dezoxyspergualin

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Effects of cyclosporins

• Receptor cytoplasmic immunophillin
• calcineurin blockage ? NF-Atc
• Rapamycin also binds to immunophillin, but
• the complex does not block calcineurin, it
  blocks proliferation in G1 phase.
• Highly lymphocyte specific. IL-2 action is
  impaired.
• T lymphocyte (Th) is blocked.

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Immunosuppression 4.

• Purin antagonists
• IMURAN
• CELLCEPT (micophenolate mofetil)
• Mizoribin bredinin
• Pirimidin antagonists
• Sodium-brequinar (highly lymphocyte specific)

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Monoclonal antibodies

• OKT3 (Anti CD3 mAb)
• CD3 T cells
• Anti-TAC (anti-IL2 receptor)
• Activated T lymphocytes
• Anti-CD4
• Anti-LFA1 anti-ICAM-1 experimental
• Anti-cytokine (IL-2, TNFa, IFN?)

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Problems of Transplantation

• There are not enough organs
• At least 150,000 patients in industrially
  developed countries badly need donor organs and
  tissues
• Every 14 minutes another name is added to the
  national transplant waiting list.
• About 16 people die because of the lack of
  available organs for transplant each day.
• Rejection
• When the immune system of the host detects
  foreign graft tissue, it launches an attack,
  resulting in tissue rejection

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Gene technology may as a solution

• Gene technology offers the possibility to breed
  the desired organs in animals.
• Lack of organs is no longer a problem

• Gene technology makes it possible to humanize the
  bred organs - the immune system identifies the
  organ as its own tissue.
• Immune system rejection is prevented

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From which animals are we able to transplant
organs
3. The Pig Surprisingly similar to our anatomy
and physiology
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
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• 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.

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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.
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Suppression of immune system rejection

• 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.

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Conclusion

• 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.

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Conclusion

• HLA typing and matching is essential for
  allografting 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.

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To seek what everybody has sought To think what
nobody has thought Try your best, everything
will be possible