Lecture 33 Transplantation

•Placenta - embryo-derived cells bathed in maternal blood supply. Tolerogenic factors include:

-Don't express HLA-ABC, but express and secrete HLA-G -Express FasL and inhibitory C' regulatory proteins -Secrete suppressive cytokines (IL-10, IDO, TGF-β) -Induce KIR expression on maternal NK cells and differentiation of maternal Th2 and M2 macs (alternatively activated) -Secrete human chorionic gonadotropin (HCG) which chemoattracts Treg cells

Corticosteroids

-Downregulate cytokines (e.g. IL-2, TNF-α, GM-CSF) and adhesion molecules; increase lymphocyte apoptosis; decrease PMN and mac phagocytosis and NOS activity

Privileged tissues evade/suppress immunity by:

-MHC class I expression - ↓ HLA-ABC and ↑ HLA-G (immunosuppressive) -↑ expression of inhibitory C' regulatory proteins -↑ FasL expression (kills infiltrating Fas+ T cells) -↑ production of immunosuppressive cytokines (e.g. IL-10, TGF-β) and neuropeptides

•Multiple ways in which direct allorecognition can occur:

-Panel A - Correct interaction between recipient MHC-peptide and TCR = the normal process for T cell activation -Panel B - donor MHC "fits" the recipient TCR = polymorphic residues of the donor MHC fit into the peptide "pockets" on the recipient TCR -Panel C - a combination of molecules from donor MHC and donor (self) peptide "fits" the recipient TCR

The 4 Immunology-Focused Steps in Screening Prior to Transplantation:

1.Blood typing - basic ABO profile of donor and recipient 2.Tissue-typing - matching 12 HLA alleles (6 MHC Class I and 6 MHC Class II) of donor and recipient 3.Cross-matching - screening of serum of recipient for antibodies that would react with donor's cells (generally MHC-specific antibodies) 4.Serology - screening for the presence of pathogens, particularly those that are blood-borne or that infect macrophages/DCs - cells that are easily transferred from donor to recipient inside the transplanted organ

Acute Rejection

A primary immune response against alloantigens of graft. •occurs in days (7+ days) to weeks or yrs. •Highest risk = first 3 months. •Mechanisms of injury are predominantly cell-mediated and include: -CD8 T cells directly attack donor cells. -CD4 T helper cells secrete cytokines - adaptive and innate. -B cells secrete antibodies that directly attack donor cells.

Two forms of allorecognition: direct vs indirect

Direct recognition - recipient T cells recognize MHC-peptides presented on engrafted donor APCs. The peptide can be the donor's self-peptide or a peptide that was foreign to the donor. •Indirect recognition - donor MHC or protein is processed by the recipient's APCs, then presented to the recipient T cell, and recognized as a foreign antigen.

When do you tissue type??

If the tissue is NOT immunopriviliged IF you have time...certain diseases like heart failure/liver their may not be time to FULLY type

Serum based assay used for HLA tissue typing

•Detects antibodies in the recipient's serum which would recognize donor MHC as foreign, bind, and activate the classical C' pathway •Lymphocyte cytotoxicity assay 1)Cells expressing known MHC proteins are "glued" to the bottom of a tissue culture plate, and serum from the recipient is added. Unbound proteins are washed off, then complement proteins are added. If anti-MHC Abs bound to the cells, then C' will be activated and the MAC complex will poke holes in the cell surface. A vital dye (for selective staining of live cells) is then added. If C' was not activated (no holes in the cells), the dye is excluded from the cell cytoplasm. If Abs bound and C' was activated, the dye will enter the cell through the holes made by MAC, staining the dead cells.

•Cell-based assays used for HLA tissue typing, she mentioned two

•Detects non-self MHC proteins on the surface of donor tissue/organ •Hemagglutination, flow cytometry, immunofluorescence - quick, can be done in less than 1 day (usually used for blood typing) •Mixed lymphocyte reaction or mixed lymphocyte culture - slower, takes several days to complete. Donor cells irradiated and mixed with live recipient cells. If reaction, cells proliferate and make cytokines.

Why does hyperacute rejection occur??

•Due to pre-existing antibodies in the recipient which recognize donor antigens (protein or carbohydrate). Potential predisposing causes - previous pregnancy, transfusion, transplant. •Since donor is already immune, response is rapid - minutes to hours

Largest factor in graft rejection

•HLA (or MHC) mismatch in HLA between donor and recipient •Largest factor - polymorphism of MHC genes •nucleated cells (expressing both MHC Class I and maybe Class II) and highly vascularized tissues (i.e. lots of vessels containing lots of MHC-bearing cells) can make good targets for the host immune response.

Stages and Major Effectors of allograft rejection

•Hyperacute rejection - antibody, complement, and clotting cascades •Acute rejection - CD4 and CD8 T cells, antibody, macrophages •Chronic rejection - CD4 T cells, fibroblasts, macrophages

orthotopic vs heterotopic transplant

•Orthotopic transplantation - grafting of tissue in a natural position; the transplanted tissue is placed in the same anatomical location •Heterotopic transplantation - occurring in an abnormal position (happens with kidney and heart transplants)

treatment of hyperacute rejection

•Treat by removal of donor tissue Major hindrance to xenotransplantation •Prevent by ABO, HLA, and cross-match testing prior to transplant

Treatment of acute rejection

•Treat with more immunosuppression - other drugs, better compliance. •Prevent by cross-matching and immunosuppression.

Tissue typing or cross-matching is the matching of

•donor and recipient tissues through a variety of laboratory tests. •The degree of similarity between the donor and recipient can predict the success of the transplant and minimize the risk of graft rejection. •The extent of HLA mismatch may also guide selection of immunosuppressive therapies.

Bone Marrow and Hematopoietic Stem Cell Transplantation

•donor cells will be recognized as foreign (host vs graft) and will recognize recipient tissue as foreign (graft vs host), leading to rejection. To minimize: -Recipient is immunosuppressed PRIOR to transplant (radiation or chemo) -Mature T cells are removed from donor tissue (i.e. cells) -Lower success rates because enriched immune cells are the graft tissue! •In stem cell transplant, cells have the potential to become any cell of the hematopoietic lineage - RBC, T, B, PMN, mac, etc.

Immunoprivileged Sites

•tissues/sites which are not well-recognized by the immune response •Often the tissues are in isolated sites (eye, brain) that are not well-vascularized. •So immunoprivileged tissues are protected because they are isolated AND because they actively work to induce immunologic tolerance.

•Graft versus host disease (GVHD)

−Mature and activated donor T cells see the recipient tissue as foreign and attack it. −Occurs most frequently in bone marrow transplants where the basis of the transplant is the engrafting of immune system cells.

•Host versus graft disease or allorecognition

−The host immune response sees the donor MHC and/or the donor self peptide as foreign and targets these proteins. −Tissue cross-matching minimizes this problem.

Partial or mixed chimerism

achieved when there are persistence low levels of donor-derived DCs and when donor progenitor cells (e.g. stem cells) proliferate and populate recipient bone marrow. Result - lymphocytes of BOTH recipient and donor origin are found in the circulation with donor-derived DCs able to effectively activate new naïve T cells that arose from donor stem cells.

alloantigens and xenoantigens

alloantigens: human antigens from allografts xenoantigens: antigens from xenograft both are detected by recipients immune response

Transfusions are considered

are a tissue transplantation with rejection targets being the AB and Rh Ags - which are easy to access in sample and easy to assay

autologous graft (auto), syngeneic graft (iso), allogenic graft (allo), xenogenic graft (xeno).

autologous or AUTO: one's self syngeneic or ISO: identical twin allogeneic or ALLO: non identical human xenogeneic or XENO: a different species

Calcineurin inhibitors

cyclosporine, -Calcineurin = important signaling molecule for T cell activation, cytokine production, and other T cell functions. So inhibiting calcineurin interferes with T cell signaling and clonal expansion

treatment of chronic rejection

directly - none. Immunosuppressive therapies have little impact because the damage is already done.

Antibodies (for immunosuppresion)

for example, anti-CD25 (basiliximab, daclizumab), polyclonal anti-T cell, anti-CD52, CTLA4-Ig

Fetus is a

•"semiallograft" since 50% of genes/proteins are foreign (from dad), so could be an easy target for rejection. •Amniotic sac - secretes suppressive cytokines/factors. Alfa-feto protein (AFP) - made in fetal yolk sac and liver, major immunosuppressive factor.

Chronic rejection

•A slow and progressive tissue destruction by inflammatory processes. •Both cell-mediated immunity (esp. CD4 T cells) and Ab contribute. •Failure occurs months to years post-transplant. •Mechanisms causing damage to graft: −T cell responses (low magnitudes) damage engrafted cells. Cytokines released which stimulates fibroblasts and vascular smooth muscle cells in graft and leads to fibrosis. −Ab-mediated responses (e.g. C' activation) also contribute to fibrotic process. −Repeated episodes of acute rejection

DNA-based assays used for HLA tissue typing

•Also called genotyping. •Compares DNA sequences of donor's and recipient's MHC genes •Quick, can generally be done in <1 day 1. PCR genotyping - analyze specific DNA sequences 2. DNA fingerprinting - based on SNPs 3. Microhybridization on a chip - a combination