L18- Immunology of Transplant

Question 1

What did Peter Medawar et al experiment look at?

Answer 1

It showed that failure of skin grafting was caused by an inflammatory reaction, which they called rejection

• Rejection shows characteristics of memory and specificity mediated by lymphocytes.
• For instance, rejection occurs 10 to 14 days after the first transplant from a donor to a nonidentical recipient (called first-set rejection) and more rapidly after the second transplant from the same donor to this recipient (called second-set rejection), implying that the recipient developed memory for the grafted tissue.

Question 2

Define autologous graft

Answer 2

Also known as autograft

A graft transplanted from one individual to the same individual – autologous transplant

Question 3

Define syngeneic graft

Answer 3

It is a graft transplanted between two genetically identical individuals – also referred to as isologous transplant (identical twins)

Question 4

Define allogeneic graft

Answer 4

Also known as allograft

It is a graft transplanted between two genetically different individuals of the same species – homologous transplant

Question 5

Define xenogeneic graft

Answer 5

Also known as a xenograft

It is a graft transplanted between individuals of different species

Question 6

Define alloantigens

Answer 6

Molecules that are recognized as foreign in allografts

Question 7

Define xenoantigens

Answer 7

Molecules recognized as foreign in xenografts, also known as heterologous transplant

Question 8

Lymphocytes and antibodies that react with alloantigens or xenoantigens are described as being…?

Answer 8

Alloreactive and xenoreactive

Question 9

Are cells or organs transplanted between genetically identical individuals (identical twins or members of the same inbred strain of animals rejected?

Answer 9

Not rejected

Question 10

Are cells or organs transplanted between genetically non-identical people or members of two different inbred strains of a species rejected?

Answer 10

Almost always rejected

Question 11

Will the offspring of a mating between two different inbred strains of animal reject grafts from either parent?

Why?

Answer 11

Will not be rejected.

In other words, an (A × B) F1 animal will not reject grafts from an A or B strain animal.

Thus, an (A × B) F1 animal does not reject either A or B strain grafts because the F1 will express all the genes donated by each parent and therefore will be tolerant to their encoded proteins.

Question 12

Will a graft derived from the offspring of a mating between two different inbred strains of animal be rejected by either parent?

Why?

Answer 12

Yes, it will be rejected.

In other words, a graft from an (A × B) F1 animal will be rejected by either an A or a B strain animal.

Both A and B strain recipients reject an (A × B) F1 graft because a graft from an F1 animal will express proteins not present in each parent, and therefore the parent will not be tolerant to those proteins.

Question 13

The molecules responsible for strong and rapid rejection reactions are…?

Answer 13

Major histocompatibility complex (MHC) molecules that bind and present peptides to T cells

Question 14

Allogeneic MHC molecules of a graft can be presented for recognition by the recipient’s T cells in which two different ways?

Answer 14

Direct and indirect

Question 15

Define direct recognition (direct presentation) of alloantigens.

Answer 15

T cells of a graft recipient recognize intact, unprocessed MHC molecules in the graft

Question 16

Define indirect recognition (indirect presentation) of alloantigens

Answer 16

Recipient T cells recognize graft (donor) MHC molecules only in the context of the recipient’s MHC molecules, implying that the recipient’s MHC molecules must be presenting peptides derived from allogeneic donor MHC proteins to recipient T cells – basically, the same as the recognition of any foreign (e.g., microbial) protein antigen.

Question 17

Where does the initial T cell response to MHC alloantigens occur? (Whether it results in direct or indirect recognition?)

Answer 17

It most likely occurs in lymph nodes draining the graft

Question 18

In direct recognition, do intact MHC molecules displayed by cells in the graft be recognise by recipient T cells without a need for processing by host APCs?

Answer 18

Yes they are recognised with a need for processing by host APC. (Why it’s called direct)

Direct allorecognition may be seen as an

Question 19

In direct allorecognition, why is MHC molecules displayed by cells in a graft able to be recognised by recipient T cells without a need or processing by host APC’s?

Answer 19

It is an example of an immunologic cross-reaction in which a T cell that was selected to be self MHC restricted is able to bind structurally similar allogeneic MHC molecules with high enough affinity to permit activation of the T cell.

T cell responses to directly presented allogeneic MHC molecules are very strong because there is a high frequency of T cells that can directly recognize any single allogeneic MHC protein.

( It is estimated that as many as 1% - 10% of all T cells in an individual will directly recognize and react against an allogeneic MHC molecule on a donor cell (cf 1 in 10 5 -10 6 against microbes))

Question 20

Why in indirect recognition, donor (allogeneic) MHC molecules are captured and processed by recipient APCs, and peptides derived from the allogeneic MHC molecules are presented in association with self MHC molecules?

Answer 20

Peptides from the allogeneic MHC molecules are displayed by host APCs and recognized by T cells like conventional foreign protein antigens.

Question 21

Is MHC class I or Class II used for indirect presentation?

Answer 21

Mostly MHC class II.

The alloantigens are acquired by host APC’s primarily through the endosomal vesicular pathway as a consequence of phagocytosis and are therefore presented by MHC class II molecules.

Question 22

When would some antigens of phagocytosed graft cells enter the MHC class I pathway?

Answer 22

Some antigens of phagocytosed graft cells do enter the class I MHC pathway of antigen presentation and are indirectly recognized by CD8 + T cells.

This phenomenon is an example of cross-presentation or cross-priming in which dendritic cells ingest proteins of another cell, e.g. from the graft, the proteins are delivered to the cytosol where they are processed into peptides by proteasomes, and the peptides are presented on class I MHC molecules to activate (prime) CD8+ CTL.

Question 23

Explain what happens in the lymph nodes that drain the graft in direct allorecognition

Answer 23

Donor APCs in transplanted organs can migrate to regional lymph nodes and present, on their surface, unprocessed allogeneic class I or class II MHC molecules to the recipient’s CD8 + and CD4 + T cells, respectively.

Question 24

Explain what happens in the lymph nodes that drain the graft in indirect allorecognition

Answer 24

Host dendritic cells from the recipient may also migrate into the graft, pick up graft alloantigens, and transport these back to the draining lymph nodes, where they are displayed (the indirect pathway)

Question 25

Why does the allorecognition (whether it’s direct or indirect) occur in the lymph nodes that drain the graft?

Answer 25

The connection between lymphatic vessels in allografts and the recipient’s lymph nodes is surgically disrupted during the process of transplantation, and it is likely reestablished by growth of new lymphatic channels in response to inflammatory stimuli produced during grafting. The Donor APC’s migrate through lymph and can be picked up by host molecules. As well as host APC’s migrating towards the graft.

Question 26

Alloreactive CD4 + and CD8 + T cells that are activated by graft alloantigens cause rejection by distinct mechanisms. Explain 3 ways.

Answer 26

* CD8+ CTL generated by direct allorecognition of donor MHC molecules on donor APCs can recognize the same MHC molecules on parenchymal cells in the graft and kill those cells, and also secrete cytokines that cause damaging inflammation * CD8 + CTLs generated in response to indirect recognition of allogeneic MHC are restricted to recognition of peptides from these allogeneic MHC molecules bound to recipient (self) MHC molecules, therefore the T cells will not be able to kill the foreign graft cells because the graft does not express recipient MHC molecules * CD4 + effector T cells generated by direct or indirect recognition of allogeneic MHC cause rejection via inflammation caused by the cytokines they produce. Effector cells activated by the indirect pathway infiltrate the graft and recognize peptides from graft MHC molecules being displayed by host APCs that have also entered the graft.

Question 27

Antibodies against graft antigens, called donor-specific antibodies, also contribute to rejection. Explain.

Answer 27

* High-affinity alloantibodies are mostly produced by helper T cell–dependent activation of alloreactive B cells, much like antibodies against other protein antigens * Antigens most frequently recognized by alloantibodies are donor MHC molecules, including both class I and class II MHC proteins * Naive B lymphocytes recognize the allogenic MHC molecules, internalize and process these proteins, and present peptides derived from them to helper T cells that were previously activated by the same peptides presented by dendritic cells. * Donor-specific antibodies against non-HLA alloantigens also contribute to rejection * Alloreactive antibodies produced in graft recipients engage the same effector mechanisms that antibodies use to combat infections, including complement activation, and Fc receptor-mediated binding and activation of neutrophils, macrophages, and NK cells * MHC antigens are expressed on endothelial cells, and much of the alloantibody-mediated damage is targeted at the graft vasculature

Question 28

Define hyperacute rejection

Answer 28

* Characterized by thrombotic occlusion of the graft vasculature that begins within minutes to hours after host blood vessels are anastomosed to graft vessels * Mediated by preexisting antibodies in the host circulation that bind to donor endothelial antigens This causes complement activation, endothelial damage, inflammation and thrombosis

Question 29

Why is hyperacute rejection rare now.

Answer 29

Hyperacute rejection by anti-ABO antibodies is extremely rare now because all donor and recipient pairs are selected so that they have compatible ABO types • In the early days of transplantation, hyperacute rejection was often mediated by preexisting immunoglobulin M (IgM) alloantibodies specific for the carbohydrate ABO blood group antigens that are expressed on red cells and endothelial cells. These natural antibodies are present in most individuals.

Question 30

Define acute cellular rejection

Answer 30

The principal mechanisms of acute cellular rejection are CTL-mediated killing of graft parenchymal cells and endothelial cells and inflammation caused by cytokines produced by helper T cells. This causes parenchyma cell damage, interstitial inflammation and endothelitis (thrombus)

Question 31

As therapy for acute rejection has improved, what is the major cause of the failure of vascularized organ allografts?

Answer 31

Chronic rejection.

Question 32

What is the survival rate of kidney allografts?

Answer 32

Since 1990, 1-year survival of kidney allografts has been better than 90%, but the 10-year survival has remained approximately 60% despite advances in immunosuppressive therapy

Question 33

Why is chronic rejection an issue for kidney allografts?

Answer 33

A dominant lesion of chronic rejection in vascularized grafts is arterial occlusion as a result of the proliferation of intimal smooth muscle cells, and the grafts eventually fail mainly because of the resulting ischemic damage * The arterial changes are called graft vasculopathy or accelerated graft arteriosclerosis * Likely caused by activation of alloreactive T cells and secretion of IFN-γ and other cytokines that stimulate proliferation of vascular smooth muscle cells. As the arterial lesions of graft arteriosclerosis progress, blood flow to the graft parenchyma is compromised, and the parenchyma is slowly replaced by nonfunctioning fibrous tissue

Question 34

What is a way the reduce graft immunogenicity?

Answer 34

In human transplantation, the major strategy to reduce graft immunogenicity has been to minimize alloantigenic differences between the donor and recipient

Question 35

What laboratory tests are routinely performed to reduce the risk for immunological rejection of allografts?

Answer 35

* ABO blood typing * determination of HLA alleles expressed on donor and recipient cells, called tissue typing * detection of preformed antibodies in the recipient that recognize HLA and other antigens representative of the donor population * detection of preformed antibodies in the recipient that bind to antigens of an identified donor’s cells, called cross-matching

Question 36

What is the laboratory technique of cross matching?

Answer 36

Detection of preformed antibodies in the recipient that bind to antigens of an identified donor’s cells

Question 37

What is the laboratory technique of tissue typing?

Answer 37

Determination of HLA alleles expressed on donor and recipient cells

Question 38

What are the HLA antigens for Class I HLA?

Answer 38

A, B and C

Question 39

What cells express class I HLA?

Answer 39

All nucleated cells The density varies from tissue to tissue

Question 40

What are the HLA antigens for Class II HLA?

Answer 40

DR, DQ and DP

Question 41

What cells express class II HLA?

Answer 41

APC’s such as dendritic cells, macrophages B lymphocytes Activated T lymphocytes (can be induced on many other cell types too)

Question 42

What is the HLA diversity (multiple genes) with allelic polymorphism for; HLA-A

Answer 42

>2000 alleles

Question 43

What is the HLA diversity (multiple genes) with allelic polymorphism for; HLA-B

Answer 43

>3000 alleles

Question 44

What is the HLA diversity (multiple genes) with allelic polymorphism for; HLA-C

Answer 44

>1500 alleles

Question 45

What is the HLA diversity (multiple genes) with allelic polymorphism for; Beta2m?

Answer 45

1 allele

Question 46

What is the HLA diversity (multiple genes) with allelic polymorphism for; HLA-DRA?

Answer 46

7 alleles

Question 47

What is the HLA diversity (multiple genes) with allelic polymorphism for; HLA-DRB1?

Answer 47

>1353 alleles

Question 48

What is the HLA diversity (multiple genes) with allelic polymorphism for; HLA-DRB3/4/5?

Answer 48

>75 alleles

Question 49

What is the HLA diversity (multiple genes) with allelic polymorphism for; HLA-DQA1+B1?

Answer 49

>400 alleles

Question 50

What is the HLA diversity (multiple genes) with allelic polymorphism for; HLA-DPA1+B1?

Answer 50

>200 alleles

Question 51

Which HLA has the most alleles?

Answer 51

HLA-B with >3000

Question 52

Which HLA has the least amount of alleles?

Answer 52

HLA-DRA with 7

Question 53

What factors are vital for graft survival?

Answer 53

The larger the number of MHC alleles that are matched between the donor and recipient, the better the graft survival. Greater survival of grafts when donor and recipient have fewer HLA allele mismatches • Of all class I and class II MHC loci, matching at HLA-A, HLA-B, and HLA-DR is most important for predicting survival of kidney allografts. (HLA-C is not as polymorphic as HLA-A or HLA-B, and HLA-DR and HLA-DQ are in linkage disequilibrium, so matching at the DR locus often also matches at the DQ locus.)

Question 54

How many HLA mismatches between donor and recipient are possible for the HLA genes?

Answer 54

* Two codominantly expressed alleles are inherited for each of these HLA genes – possible to have zero to six HLA mismatches of these three loci between the donor and the recipient * Survival of grafts with two to six mismatches is significantly worse than that of grafts with zero- and one-antigen mismatches

Question 55

How long can kidney is stored before transplantation?

Answer 55

72 hours HLA matching in renal transplantation is possible because donor kidneys can be stored for up to 72 hours before being transplanted, and patients needing a kidney allograft can be maintained on dialysis until a well-matched organ is available

Question 56

Which organs can be used for HLA matching without issues with graft survival?

Answer 56

Kidneys can. HLA matching in renal transplantation is possible because donor kidneys can be stored for up to 72 hours before being transplanted, and patients needing a kidney allograft can be maintained on dialysis until a well-matched organ is available.

Question 57

Can HLA typing be considered for pairing with heart and liver diners and recipients? ]

Answer 57

No. It’s is not considered. The best approach is ABO blood group matching and anatomical compatibility. In the case of heart and liver transplantation, organ preservation is more difficult, and potential recipients are often in critical condition

Question 58

Patients in need of allografts are also tested for the presence of preformed antibodies against donor MHC molecules or other cell surface antigens. What test can be performed to test this?

Answer 58

Panel reactive antibodies (PRA)

Question 59

Describe why you would use the Panel reactive antibodies (PRA) test?

Answer 59

* Patients waiting for organ transplants are screened for the presence of preformed antibodies reactive with allogeneic HLA molecules prevalent in the population. * Results are interpreted as percentage of MHC allele panel with which the patient’s serum reacts

Question 60

Wha increases the risk of having antibodies against donor MHC molecules?

Answer 60

The presence of these antibodies, which may be produced as a result of previous pregnancies, transfusions, or transplantation, increases risk for hyperacute or acute vascular rejection

Question 61

How does Panel reactive antibodies (PRA) work?

Answer 61

HLA antibody characterization using a bead-based multiplex assay Beads are multiplexed in a suspension array on the Luminex platform and incubated with recipient serum. If anti-HLA antibodies are present, they bind to the corresponding HLA molecules on the surface of the microbeads. The beads are then incubated with phycoerythrin-labelled anti-human IgG and analysed using flow cytometry. A dual-laser instrument interrogates the beads, with one laser identifying the specific bead and the other determining the presence of bound antibody. The strength of any antibodies detected is measured as the mean fluorescence intensity

Question 62

Explain how HLA testing for a cross match works for; Complement-dependent cytotoxicity crossmatch

Answer 62

Recipient serum and exogenous complement are added to donor lymphocytes. If donor- specific anti- HL A antibodies are present in the recipient serum, they bind to HL A molecules on the surface of the donor lymphocytes and activate complement, leading to cell lysis. The lysed cells take up vital dye and can be detected using microscopy. The strength of the crossmatch is graded according to the proportion of lysed cells. As T cells and B cells have differential expression of class I and class II HL A molecules, both T cell and B cell crossmatch assays are performed. The sensitivity of the crossmatch test can be increased by the addition of anti-human globulin (AHG). Multiple AHG molecules bind to each HL A-bound donor- specific antibody (DSA), leading to amplification of complement activation and subsequent cell lysis.

Question 63

Explain how HLA testing for a crossmatch works for; Flow Cytometry Crossmatch test

Answer 63

Recipient serum is added to donor lymphocytes. If donor-specific anti-HL A antibodies are present in the recipient serum, they bind to HL A molecules on the surface of donor T or B cells. Following incubation with fluorochrome- labelled anti-human immunoglobulin G (IgG), HL A-bound anti-HL A antibodies can be detected using flow cytometry. The strength of the flow cytometry crossmatch is measured as the number of ‘channel shifts’ above the control sample.

Question 64

What is the major immunologic barrier to xenogeneic transplantation

Answer 64

A major immunologic barrier to xenogeneic transplantation is the presence of natural antibodies in the human recipients that cause hyperacute rejection

Question 65

Natural antibodies are rarely produced against carbohydrate determinants of closely related species, such as humans and chimpanzees. Why?

Answer 65

Ethical reasons

Question 66

What antigens are detected by the immune system in species that are evolutionarily distant, such as pigs, which have anatomically compatible organs?

Answer 66

More than 95% of primates have natural IgM antibodies that are reactive with carbohydrate determinants expressed by cells of species such as pigs.

Question 67

What happens during when a xenogeneic transplantation has induced a hyperacute rejection?

Answer 67

Natural antibodies against xenografts induce hyperacute rejection by the same mechanisms as those seen in hyperacute allograft rejection. These mechanisms include the generation of endothelial cell procoagulants and platelet-aggregating substances, coupled with the loss of endothelial anticoagulant mechanisms • Even when hyperacute rejection is prevented, xenografts are often damaged by a form of acute vascular rejection that occurs within 2 to 3 days of transplantation. This form of rejection has been called delayed xenograft rejection, accelerated acute rejection, or acute vascular rejection and is characterized by intravascular thrombosis and necrosis of vessel walls.

Question 68

When would you use HSC transplantation? | Haematopoietic Stem Cell Transplant

Answer 68

HSC transplantation is a clinical procedure to treat lethal diseases caused by intrinsic defects in one or more hematopoietic lineages in a patient * A patient’s own hematopoietic cells are destroyed, and HSCs from a healthy donor are then given to restore normal blood cell production in the patient * Transplantation of pluripotent HSCs was done in the past using an inoculum of bone marrow cells collected by aspiration, and the procedure is often called bone marrow transplantation * In modern clinical practice, HSCs are more often obtained from the blood of donors, after treatment of the donor with colony-stimulating factors that mobilize stem cells from the bone marrow * The recipient is treated before transplantation with a combination of chemotherapy, immunotherapy, or irradiation to kill the defective HSCs, and to free up niches for the transferred stem cells * After transplantation, the injected stem cells repopulate the recipient’s bone marrow and differentiate into all of the hematopoietic lineages * HSC transplantation is most often used clinically in the treatment of leukemias and pre-leukemic conditions. HSC transplantation is the only curative treatment for some of these diseases, including chronic lymphocytic leukemia and chronic myeloid leukemia. The mechanisms by which HSC transplantation cures hematopoietic neoplasms is in part the graft-versus-tumor effect, in which mature T cells and NK cells present in the bone marrow or stem cell inoculum recognize alloantigens on residual tumor cells and destroys them

Question 69

Explain autologous stem cell transplant

Answer 69

In an autologous bone marrow transplant, hematopoietic stem cells are harvested from the blood or bone marrow of a patient before the patient undergoes treatment for cancer. In order to remove tumour cells that may have been collected with the stem cells, the sample is incubated with antibodies that bind only to stem cells. The stem cells are then isolated and stored for later use, when they are reinfused into the patient.

Question 70

Define graft vs host disease (GVHD)

Answer 70

GVHD is caused by the reaction of grafted mature T cells in the HSC inoculum with alloantigens of the host. GVHD is the principal cause of mortality among HSC transplant recipients