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MICROBIOLOGY AND IMMUNOLOGY MOBILE  -  IMMUNOLOGY CHAPTER FIFTEEN 

MHC: GENETICS AND ROLE IN TRANSPLANTATION 
 

DEFINITIONS

• Histocompatibility (transplantation) antigens
  Antigens on tissues and cells that determine their rejection when grafted between two genetically different individuals
• Major histocompatibility (MHC) antigens
  Histocompatibility antigens that cause a very strong immune response and are most important in rejection
• MHC complex
  Group of genes on a single chromosome encoding the MHC antigens
• HLA (human leukocyte antigens)
  MHC antigens of man (first detected on leukocytes)
• H-2 antigens
  MHC antigens of mouse

Types of graft (figure 1)

• Xenograft
  Grafts between members of different species (also known as heterologous, xenogeneic or heterografts)
• Allograft
  Grafts between two members of the same species (also known as allogeneic or homograft)
• Isograft
  Grafts between members of the same species with identical genetic makeup (identical twins or inbred animals)

Haplotype
A group of genes on a single chromosome
 

PRINCIPLES OF TRANSPLANTATION (figure 2)

An immunocompetent host recognizes the foreign antigens on grafted tissues (or cells) and mounts an immune response which results in rejection. On the other hand, if an immunocompromised host is grafted with foreign immunocompetent lymphoid cells, the immunoreactive T-cells in the graft recognize the foreign antigens on the host tissue, leading to damage of the host tissue.

Host-versus-graft-reaction

The duration of graft survival follows the order, xeno- < allo- < iso- = auto- graft. The time of rejection also depends on the antigenic disparity between the donors and recipient. MHC antigens are the major contributors in rejection, but the minor histocompatibility antigens also play a role. Rejection due to disparity in several minor histocompatibility antigens may be as quick or quicker than rejection mediated by an MHC antigen. As in other immune responses, there is immunological memory and secondary response in graft rejection. Thus, once a graft is rejected by a recipient, a second graft from the same donor, or a donor with the same histocompatibility antigens, will be rejected in a much shorter time.
 

Graft-versus-host (GVH) Reaction
Histocompatible lymphoid cells, when injected into an immunocompromised host, are readily accepted. However, the immunocompetent T lymphocytes among the grafted cells recognize the alloantigens and, in response, they proliferate and progressively cause damage to the host tissues and cells. This condition is known as graft-versus-host (GVH) disease (figure 3) and is often fatal.

Common manifestations (figure 4) of GVH reaction are diarrhea, erythema, weight loss, malaise, fever, joint pains, etc. and ultimately death.
 

THE MHC GENE COMPLEX

The MHC complex contains a number of genes that control several antigens, most of which influence allograft rejection. These antigens (and their genes) can be divided into three major classes: class I, class II and class III. The class I and class II antigens are expressed on cells and tissues whereas as class III antigens are represented on proteins in serum and other body fluids (e.g.C4, C2, factor B, TNF). Antigens of class III gene products have no role in graft rejection.

Human MHC
The human MHC is located on chromosome 6.

Class I MHC
The class I gene complex contains three major loci, B, C and A and other undefined minor loci (figure 5). Each major locus codes for a polypeptide; the alpha-chain that contains antigenic determinants, is polymorphic (has many alleles). It associates with beta-2 microglobulin (beta-chain), encoded by a gene outside the MHC complex, and expressed on the cell surface. Without the beta-2 microglobulin, the class I antigen will not be expressed on the cell surface. Individuals with a defective beta-2 microglobulin gene do not express any class I antigen and hence have a deficiency of cytotoxic T cells.

Class II MHC
The class II gene complex also contains at least three loci, DP, DQ and DR; each of these loci codes for one alpha- and one beta-chain polypeptide which associate together to form the class II antigens. Like the class I antigens, the class II antigens are also polymorphic. The DR locus may contain more than one, possibly four, functional beta-chain genes.
 

Mouse MHC
The mouse MHC is located on chromosome 17.

Class I MHC
This consists of two major loci, K and D. Unlike the human MHC, the mouse class I gene complexes loci are not together but they are separated by class II and class III genes (Figure 6A).

Class II MHC
The class II gene complex contains two loci, A and E, each of which code for one alpha and one beta chain polypeptide, which form one class II molecule. The mouse class II gene complex is also known as the I region and the genes in this complex are referred to as Ir (immune response) genes since they determinel the magnitude of immune responsiveness of different mouse strains to certain antigens. Products of the A and E loci are also termed IA and IE antigens, collectively known as Ia antigens.
 

MHC ANTIGENS

Nomenclature
HLA specificities are identified by a letter for locus and a number (A1, B5, etc.) and the haplotypes are identified by individual specificities (e.g., A1, B7, Cw4, DP5, DQ10, DR8). Specificities which are defined by genomic analysis (PCR), are names with a letter for the locus and a four digit number (e.g. A0101, B0701, C0401 etc). 

Specificities of mouse MHC (H-2) are identified by a number. Since laboratory mice are inbred, each strain is homozygous and has a unique haplotype. The MHC haplotype in these strains is designated by a 'small' letter (a, b, d, k, q, s, etc.); for example, the MHC haplotype of Balb/c mice is H2d.

Inheritance
MHC genes are inherited as a group (haplotype), one from each parent. Thus, a heterozygous human inherits one paternal and one maternal haplotype, each containing three class-I (B, C and A) and three class II (DP, DQ and DR) loci. A heterozygous individual will inherit a maximum of 6 class I specificities (Figure 6: top). Similarly, the individual will also inherit DP and DQ genes and express both parental antigens. Since the class II MHC molecule consists of two chains (alpha and beta), with some antigenic determinants (specificities) on each chain, and DR alpha- and beta-chains can associate in ether cis (both from the same parent) or trans (one from each parent) combinations, an individual can have additional DR specificities (Figure 6B). Also, there are more than one functional DR beta-chain genes (not shown in the figure). Hence, many DR specificities can be found in any one individual.

Crossover
Haplotypes, normally, are inherited intact and hence antigens encoded by different loci are inherited together (e.g., A2; B27; Cw2; DPw6; DQw9; DRw2). However, on occasions, there is crossing over between two parental chromosomes, thereby resulting in new recombinant haplotypes. Thus, any one specificity encoded by one locus may combine with specificities from other loci. This results in vast heterogeneity in the MHC make-up in a given population.

MHC antigen expression on cells
MHC antigens are expressed on the cell surface in a co-dominant manner: products of both parental genes are found on the same cells. However, not all cells express both class I and class II antigens. While class I antigens are expressed on all nucleated cells and platelets (and red blood cells in the mouse), the expression of class II antigens is more selective. They are expressed on B lymphocytes, a proportion of macrophages and monocytes, skin associated (Langerhans) cells, dendritic cells and occasionally on other cells.

MHC detection by serological test
The MHC class I antigens are detected by serological assays (Ab and C). Tissue typing sera for the HLA were obtained, in the past, from multiparous women who were exposed to the child's paternal antigens during parturition and subsequently developed antibodies to these antigens. More recently, they are produced by monoclonal antibody technology. With most laboratories switching to PCR for tissue typing, the use of serology is rapidly diminishing.

MHC detection by mixed leukocyte reaction (MLR)
It has been observed that lymphocytes from one donor, when cultured with lymphocytes from an unrelated donor, are stimulated to proliferate. It has been established that this proliferation is primarily due to a disparity in the class II MHC (DR) antigens and T cells of one individual interact with allogeneic class-II MHC antigen bearing cells (B cells, dendritic cells, langerhans cells, etc.). This reactivity was termed mixed leukocyte reaction (MLR) and has been used for studying the degree of histocompatibility. In this test, the test lymphocytes (responder cells)are mixed with irradiated or mitomycin C treated leukocytes from the recipient, containing B-lymphocytes and monocytes (stimulator cells). The cells are cultured for 4 6 days. The responder T cells will recognize the foreign class II antigens found on the donor and undergo transformation (DNA synthesis and enlargement: blastogenesis) and proliferation (mitogenesis). The T cells that respond to foreign class II antigens are typically CD4+ TH-1 type cells. These changes are recorded by the addition of radioactive (tritiated, 3H) thymidine into the culture and monitoring its incorporation into DNA.
 

Generation of cytotoxic T lymphocytes
Another consequence of the MHC antigen and T cell interaction is the induction of cytotoxic T-lymphocytes. When T-lymphocytes are cultured in the presence of allogeneic lymphocytes, in addition to undergoing mitosis (MLR), they also become cytotoxic to cells of the type that stimulated MLR (figure 7). Thus, T-lymphocytes of 'x' haplotype cultured over 5 - 7 days with B lymphocytes of 'y' haplotype will undergo mitosis and the surviving T-lymphocytes become cytotoxic to cells of the 'y' haplotype. The induction of mitosis in MLR requires disparity of only class II antigens whereas the induction of cytotoxic T-lymphocytes (CTL) requires disparity of both class I and class II antigens. However, once cytotoxic cells have been induced, the effector cytotoxic cells recognize only class I antigens to cause cytotoxicity.
 

ALLOGRAFT REJECTION
The clinical significance of the MHC is realized in organ transplantation. Cells and tissues are routinely transplanted as a treatment for a number of diseases. However, reaction of the host against allo-antigens of the graft (HVG) results in its rejection and is the major obstacle in organ transplantation. The rejection time of a graft may vary with the antigenic nature of the graft and the immune status of the host and is determined by the immune mechanisms involved (Figure 8 and Table 1).

Hyper-acute rejection
This occurs in instances when the recipient has preformed high titer antibodies. A graft may show signs of rejection within minutes to hours due to immediate reaction of antibodies and complement.

Accelerated (2nd set; secondary) rejection
Transplantation of a second graft, which shares a significant number of antigenic determinants with the first one, results in a rapid (2 - 5 days) rejection. It is due to presence of T-lymphocytes sensitized during the first graft rejection. Accelerated rejection is mediated by immediate production of lymphokines, activation of monocytes and macrophages, and induction of cytotoxic lymphocytes.
 

Acute (1st set; primary) rejection
The normal reaction that follows the first grafting of a foreign transplant takes 1 - 3 weeks. This is known as acute rejection and is mediated by T lymphocytes sensitized to class I and class II antigens of the allograft, elicitation of lymphokines and activation of monocytes and macrophages.
 

Chronic rejection
Some grafts may survive for months or even years, but suddenly exhibit symptoms of rejection. This is referred to as chronic rejection, the mechanism of which is not entirely clear. The hypotheses are that this may be due infection, causes which led to failure of the first organ, loss of tolerance induced by the graft, etc.

Fetus as an Allograft
The fetus in an out-bred mammalian species bears antigens derived from both the father and the mother. Thus, truly, the fetus is an allograft and the mother should normally recognize the fetus as foreign and reject the fetus. Nonetheless, such rejections seldom occur. Thus, mammals have adapted in a way that allows implantation of their embryos in the mother's womb and their subsequent survival. There are multiple mechanisms that play a role, of which the most important being the unique structure and function of placenta.

Immunologically privileged sites and tissues
There are certain locations in the body in which allografts are not readily rejected. These include the brain, anterior chamber of the eye, testis, renal tubule, uterus, etc. This stems from the fact that such sites may lack of good lymphatic drainage. Also, such tissues may express molecules such as Fas ligand that kills any immune cell that may come in contact with these tissues. Additionally, such tissues, may have other immune suppressor mechanisms. Similarly, there are some tissues that can be transplanted without matching and without being rejected. Such tissues are called immunologically privileged tissues. Corneal graft is an excellent example that enjoys the highest success rate of any form of organ transplantation. The low incidence of graft rejection is impressive despite the fact that HLA antigen matching of donor and recipient is not normally performed. There are many explanations as to why such grafts are accepted. The avascularity of the graft bed prevents corneal alloantigens from reaching the regional lymphoid tissues. Also, the corneal antigens may be masked. Together, such mechanisms fail to activate the immune system of the recipient.

PROCEDURES TO ENHANCE GRAFT SURVIVAL

In clinical practice, the most successful transplantation programs have been with kidneys and corneas. However, other organs are being transplanted with increasing frequency. The success in these programs has been due to a better understanding of immunological mechanisms, definition of MHC antigens and development of more effective immunosuppressive agents.

Donor selection
Based on extensive experiences with renal transplants, certain guidelines can be followed in donor selection and recipient preparation for most organ transplants. The most important in donor selection is the MHC identity with the recipient; an identical twin is the ideal donor. Grafts from an HLA-matched sibling have 95-100% chance of success. One haplotype-identical parent or sibling must match at the HLA D region. A two haplotype-distinct donor with a reasonable match for D-region antigen can also be used. Organs from a two or one DR matched cadaver have been used also with some success. In every case, an ABO compatibility is essential.

Recipient preparation
The recipient must be infection-free and must not be hypertensive. One to five transfusions of 100-200 ml whole blood from the donor at 1-2 week intervals improves the graft survival and is practiced when possible.

Immunosuppression
Immunosuppressive therapy is most essential part of allo-transplantation. The most recent and effective family of agents is cyclosporin A, FK-506 (tacrolimus) and rapamycin. Cyclosporin A and FK506 inhibit IL-2 synthesis following Ag-receptor binding whereas rapamycin interferes with signal transduction following IL2 - IL2 receptor interaction. Thus, all three agents block T cell proliferation in response to antigen. Other chemical agents used to prevent graft rejection and their modes of action have been listed in Table 2. Whole body irradiation is used in leukemia patients before bone marrow transplantation. Antisera against T cells (anti-thymocyte globulin: ATG) or their surface antigens (CD3, CD4, CD45 on activated T-cells, CD25:IL-2 receptors) are being used also to achieve immunosuppression (Table 2).

Strategies for bone marrow transplantation
In bone marrow transplantation, the most crucial factor in donor selection is class II MHC compatibility. Once again an identical twin is the ideal donor. From poorly matched grafts, T lymphocytes can be removed using monoclonal antibodies (figure 10). The recipient must be immunosuppressed. Malignant cells must be eliminated from the recipient blood (in case of blood-borne malignancies). Methotrexate, cyclosporin and prednisone are often used to control GVH disease.
 

Other grafts
Corneal grafts do not contain D region antigens and consequently survival is frequent. Small grafts are better and corticosteroids are helpful.
Skin allografts have a very poor success rate and immunosuppressive therapy is relatively ineffective. Nevertheless, they are often used to provide a temporary covering to promote healing in severe skin damage. Indeed, there will be no rejection if the host and donor are perfectly matched (identical twins) or the recipient is tolerant to the donor MHC antigens (bone marrow chimeras).
 

 

MHC association with diseases
A number of diseases have been found to occur at a higher frequency in individuals with certain MHC haplotypes. Most prominent among these are ankylosing spondylitis (B27), celiac disease (DR3) and Reiter's syndrome (B27). Other diseases associated with different specificities of the MHC are listed in Table 3. No definite reason is known for this association. However, several hypotheses have been proposed: antigenic similarity between pathogens and MHC, antigenic hypo- and hyper-responsiveness controlled by the class II genes are included among them.
 

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