TRANSPLANTATION

IMMUNOLOGY
1. Introduction

1.1 What is Transplantation?

 Transplantation or graft is the act of transferring cells,
tissues, or organs from one site to another.
 Studying and understanding the mechanisms of the
immune system is critical.
 Organ donors may be:
– living, brain dead or dead via circulatory death.

 Organs that can be transplanted include:

 Unlike organs, most tissues (with the exception of
corneas) can be "banked".
 A particular problem is organ trafficking
 Some organs, such as the brain, cannot be transplanted.
 1.2 History

• In 1944, Medawar showed that skin allograft rejection is a host

versus graft response.
• The first successful identical twin transplant of a human kidney was
performed by Joseph E. Murray in 1954 in Boston,
• The first successful liver transplant by Dr. Thomas E. Starzl in
1967,
• The first heart transplantation by Christian Barnard in 1967,
• The first successful bone marrow transplant by E. Donnall Thomas
in 1968.
• Schwartz and Dameshek, in 1959, showed that 6-mercaptopurine
was immunosuppressive in rats,
– ushering in the era of immunosuppressive drug treatment.
1.3 Types of Transplant or Graft

 To define the terms used for transplants between

individuals and species:

1. Autograft
2. Allograft or Allotransplantation (homograft)
3. Isograft
4. Xenograft or xenotransplantation (heterograft)
 Other types of transplants include:
Domino Transplants
 This term also refers to a series of living donor transplants
 These transplants are otherwise impossible due to blood
type or antibody barriers to transplantation.
 E.g. In people with cystic fibrosis
ABO-incompatible Transplants
 Is the transplantation of organs or tissues from donors
which are not ABO-compatible
 The most important factors are that the recipients not have
produced
 isohemagglutinins and
 that they have low levels of T cell-independent antigen.
TRANSPLANTATION
REJECTION
2.1 Genetic Background
• Histocompatibility antigens.
• In humans, the MHC is called the human leukocyte
antigen (HLA) system
• MHC genes are co-dominantly expressed
• The MHC molecules are divided into 2 classes
 The class I molecules are normally expressed on all nucleated
cells
 class II molecules are expressed only on the antigen presenting
cells (APCs)
• The physiological function of the MHC molecules is to
present antigenic peptides to T cells
2.2 Mechaisms of Rejection by Adaptive Immunity

• The immune response to a transplanted organ

consists
– Cellular (lymphocyte mediated)
– Humoral (antibody mediated) mechanisms.
• T cells are central in the rejection of grafts.
• The rejection reaction consists of the
Sensitization stage and
Effector stage.
 Sensitization Stage
• CD4 and CD8 T cells, via their T cell
receptors, recognize the alloantigens
expressed on the cells of the foreign graft.
• Two signals are needed for recognition of an
antigen;
 The interaction of the T cell receptor with the
antigen presented by MHC molecules,
A co-stimulatory receptor/ligand interaction on the
T cell/APC surface.
• Direct and indirect pathways.
Direct Pathway
Indirect pathway
 Effector Stage
• Alloantigen dependent and independent factors
contribute to the effector mechanisms.
• Initially, non immunologic "injury responses"
(ischemia) induce a nonspecific inflammatory response
• Various T cells and T cell derived cytokines are up
regulated early after transplantation.
• Later, ßchemokines like RANTES,IP10, and MCP1
are expressed
• Endothelial cells activated by T-cells
• CD8positive T cells mediate cell mediated
cytotoxicity reactions by delivering a "lethal hit"
 Apoptosis

• The final common pathway for the cytolytic

processes is triggering of apoptosis in the target
cell
• CD8positive CTLs can also use the Fas
dependent path way to induce cytolysis and
apoptosis.
• Cell mediated cytotoxicity has been shown to
play an important role in acute allograft rejection.
 Role of Natural Killer Cells
• The natural killer (NK) cells are important in
transplantation
• In addition, they also possess stimulatory receptors
• These effector responses include
– both cytokine release and
– direct toxicity
• NK cells also provide help to CD28positive host T
cells, thereby promoting allograft rejection.
• NK cells are now being recognized as active
participants in the acute and chronic rejection of
solid tissue grafts.
2.3 Rejection Detection
• Diagnosis of acute rejection relies on clinical data
—patient signs and symptoms
• But also can be done by laboratory data such as
tissue biopsy.
• The laboratory pathologist generally seeks three
main histological signs
(1) infiltrating T cells, perhaps accompanied by
infiltrating eosinophils, plasma cells, and neutrophils.
(2) structural compromise of tissue anatomy,
(3) injury to blood vessels.
2.4 Mechanism of Rejection by Innate Immunity

• The up-regulation of pro-inflammatory mediators

in the allograft occur before the T cell response
• These innate mechanisms alone do not appear
sufficient to lead to graft rejection itself.
• However, they are important for
– optimal adaptive immune responses to the graft
– may play a major role in resistance to tolerance
induction.
 
2.5 Clinical stages of Rejection

• Once graft rejection has begun, it can be

classified in one of three ways in humans
– Hyperacute rejection,
– Acute rejection,
– Chronic rejection.

• These categorizations are based on how quickly

rejection occurs.
 Hyperacute rejection
• In hyperacute rejection-minutes to hours,
because vascularization is rapidly destroyed.
• It is humorally mediated
• The kidney is most susceptible to hyperacute
rejection especially due to rapid clumping
• The liver is relatively resistant
 Acute Rejection
• Acute rejection occurs to some degree in all
transplants, unless immunosuppression is achieved
• Acute rejection begins as early as one week after
transplant
• Highly vascular tissues such as kidney or liver often
host the earliest signs
• A single episode of acute rejection can be
recognized and promptly treated, but recurrent
episodes lead to chronic rejection.
• Acute cellular rejection is mediated by lymphocytes
 Chronic rejection
• Chronic rejection develops months to years after
acute rejection episodes have subsided.
• Chronic rejections are both antibody and cell
mediated.
• The use of immunosuppressive drugs methods has
increased the survival of allografts in the first year
• But chronic rejection is not prevented in most
cases.  
• Chronic rejection appears
– as fibrosis and
– scarring in all transplanted organs
• The following factors increase the risk of
chronic rejection:
Previous episode of acute rejection
Inadequate immunosuppression
Initial delayed graft function
Donor related factors (e.g., old age, hypertension)
Reperfusion injury to organ
Long cold ischemia time
Recipient related factors (e.g., diabetes,
hypertension, hyperlipidemia)
Post transplant infection (e.g., cytomegalovirus
[CMV])
Transplantation Rejection Therapy
and Treatments
3.1 Immune Tolerance
• Allograft may be accepted without
immunosuppressive measures
• In tissues that lack alloantigens no immunologic
barriers to transplantation
• Allograft may be accepted in two cases:
1. When cells or tissue are grafted to a so-called
privileged site.
2. When state of tolerance has been induced
biologically.
 
3.2 Immunosuppressive Therapy
• Allogeneic transplantation requires some degree of
immunosuppression if the transplant is to survive.
• Immunosuppressants diminish the body's immune system
• There are adverse side-effects that include
– a high incidence of opportunistic infection and
– transplant-related diseases in patients.

• A major goal of immunosuppression is:

– To identify the optimal balance of therapy
– Drug-related adverse effects, infection, and
malignancies are minimized.
 Immunosuppressive therapies are classified into:
A. General immunosuppressive therapy
• Nonspecific
• Any rapidly dividing non-immune cells are also
affected
B. Specific Immunosuppressive Therapy
• Is an antigen-specific immunosuppressant
• This goal has not yet been achieved in human
transplants
 Stages of immunosuppressive treatment are:
1. Inductive therapy
2. Maintenance therapy
3. Episodic treatment
1. Inductive therapy
– Inductive therapy refers to the prophylactic application of
perioperative antibodies

– The goal of the employment of these drugs is to induce hypo-

responsiveness

– The drugs must be strong enough to prevent rejection

– Initial efforts at inductive therapy utilized polyclonal

antibodies,

– Antimicrobial agents are a necessary component of

inductive therapy.
2. Maintenance therapy
– Maintenance immunosuppression refers
combination of therapy
– The combination includes
• corticosteroid,
• calcineurine inhibitor, and
• anti-proliferative.
– Distinct combined effects on each individual
– Balance between under-immunosuppression and
over-immunosuppression
Corticosteroids
• Anti-inflammatory and immunosuppressive
effects.
• Are non-specific
• induce a state of immune hypo-responsiveness
• They inhibit
– cytokine production,
– circulation of lymphocytes,
– block T cell activation and proliferation
• Prednisone and Methyl-prednisolone are two of
the most commonly prescribed corticosteroids
• Too much corticosteroid can cause
hypertension, hyperglycemia, and
opportunistic infection
• Too little corticosteroid can result in graft
rejection.
Calcineurine inhibitor
• To combat activated T cells
• Calcineurine inhibitors
– block the clonal expansion of T cells
• Cyclosporine and Tacrolimus are the two most
prominent drugs
 Anti-Proliferatives
• The final part of triple therapy includes anti-
proliferatives
– Mycophenolate Mofetil,
– Azathioprine, and
– Sirolimus.
• These anti-mitotic drugs inhibit DNA synthesis
• The role is more general and supportive
• Patients can be gradually weaned off of anti-
proliferatives
3. Episodic treatment
• Most transplanted organs do eventually fail.
• High doses of corticosteroid is used to combat
rejection
• Polyclonal and monoclonal antibodies are often
employed as a rescue therapy.
• Muromonab-CD3 and Basiliximab, are more
specific than their polyclonal counterparts;
• Anti-proliferatives have also been found to be
effective in treating rejection episodes.
3.3 Antibody-Based Treatment
• Antibody specific to select immune components can be
added to immunosuppressive therapy
• Antibody drugs are:
– Monoclonal anti-IL-2Rα receptor antibodies
– Basiliximab
– Daclizumab
– Polyclonal anti-T-cell antibodies
– Anti-thymocyte globulin (ATG)
– Anti-lymphocyte globulin (ALG)
– Monoclonal anti-CD20 antibodies
– Rituximab
• Cases refractory to immunosuppressive or antibody
therapies are sometimes given blood transfusions
3.4 Marrow Transplant
• Replace the transplant recipient's immune system
with the donor's.
• The marrow's hematopoietic stem must be of the
individual who donated the organ
• Graft-versus-host disease (GVHD).
• GVHD can result in generalized erythroderma of
the skin, gastrointestinal hemorrhage, and liver
failure
• Various treatments are used to prevent GVHD in
bone marrow transplantation.
– The transplant recipient is usually placed on a
regimen of immunosuppressive drugs
– the donor bone marrow is treated with anti-T-cell
antisera or monoclonal antibodies specific for T cells
– Partial T-cell depletion.
 ADVANCES IN
TRANSPLANTATION
• So, how to overcome transplant rejection?
1) Screening patients for particular types of cells.
(e.g. HLA)
2) Screening for genetic mutations to predict
whether a patient is prone to rejection.
3) Early detection
4) Gene sequencers to measure the amount of
foreign DNA in blood or in transplanted organs.
5) Drugs that suppress the immune response.
6) Testing for gene mutations, to predict rate of
processing of a drug and configuring doses.
 FUTURE DRUGS
• Importance of improving current drugs and
developing new approaches for
pharmacotherapy.

Minimize side effects associated with current

medications.
Improvement in effectiveness of drugs, as
most transplants ultimately fail.
 Basiliximab

• It’s used for induction therapy in renal

transplantation.
• Reduces acute rejection
• May serve as an alternative to corticosteroids in
acute rejection due to its:
 Overall efficacy
 Tolerability
 Ease of administration
 Cost effectiveness
 Reduction of incidence of adverse events
What is it Basiliximab?
• A chimeric monoclonal antibody specific for the
alpha chain of high affinity IL-2 receptor.
• The IL-2 receptor (IL-2R) is the surface antigen
on T lymphocytes that detects IL-2 (a cytokine).
• Has significant effects only on T lymphocytes.
 Anti-CD20
• Antibody
corresponding to
CD20 (Rituximab),
depletes B cells
from the blood and
lymphatic tissues.
• Possible treatment
for antibody-
mediated acute
vascular rejection.
CP-690,550
• targets enzymes found ONLY in immune cells
• It inhibits the enzyme Jak3; this protein is only found
in immune cells
Antibodies targeting costimulatory proteins
Monoclonal or polyclonal antibodies blockage of signals
for activation of T and B lymphocytes leads to anergy.
• Mature B and T cells needed for normal immune
function (such as common diseases) are spared.
• Thus, only limited impairment of the immune system
is experienced.
 Finding Alternatives
What if you could modify, or create, a transplant organ that
doesn’t send the patient’s body into a state of immune hysteria?

What if…you could synthesize organs from scratch??!

Recent advances include:

• 3D Printing
• Tissue Engineering
 3D Printing
“grow your own organ”

• Layer by layer construction of a particular organ

structure to form a cell scaffold.
• This can be followed by the process of
 Cell seeding.
OR
 Integrating cells into the printable material itself.
• How does it work?
1. Biopsy of the organ to isolate and grow cells in a
lab.
2. Grow cells, mix them in an oxygen-rich liquid
with other nutrients .
3. Print the slurry into the appropriate shape along
with a biomaterial for structure.
This makes use of Modified inkjet printers
• Printer cartridges are filled with a suspension of
living cells and a smart gel.
• Materials for 3D printing consist of integration of
cells with alginate or fibrin polymers.
Printing material --"bioink" must be:
• Biocompatible
• Biodegradable
• Customizable and adaptable

• Organs successfully printed and

implemented in a clinical setting are either
flat, vascular, or hollow.
TISSUE ENGINEERING
 Stem Cells in a Petri Dish
• Instead of “printing” the cells into a supportive
lattice, assist the more complex organs (spinal
cords, brains, etc.) in growing on their own.
•Tissue engineering incorporates use of stem cells
and therapeutic cloning to circumvent
Immunosuppression
Drawbacks
– Manufacturing problems.
– Ethical considerations.
An attempt to circumvent use of embryonic stem
cells= using adult stem cells
• Detergent solution to strips a donor organ of cells
that might set off an immune response in the
recipient.
• Then adult skin cells turned into pluripotent stem
cells using messenger RNA
 Pigs as “biological incubators”
• Injection of human stem cells into pig embryos
to produce chimeras.
1. CRISPR gene editing to remove DNA from a
newly fertilized pig embryo.
2. Then, human induced pluripotent (iPS) stem
cells are injected into the embryo.
Engineering of immune cells to protect organs
from transplant rejection
• Gene therapy that programs T regulatory cells
(Tregs)
• Harmless virus to insert the CAR gene into
Tregs,
• Modified Tregs could recognize transplanted
tissues and protect them from the immune
system.