1. Transplantation Immunology Major Histocompatibility Complex (MHC) The MHC represent a set of genes that control the types of cellular antigens known as MHC antigens or Human Leucocytes Antigens (HLA). These antigens differ from one individual to the other. The immune system differentiate foreign from self antigen via the recognition of the HLA antigens. These antigens represent the crucial factor in acceptance or rejection of transplanted / grafted tissues or organs. In early seventies, it was discovered that sera from multiparous women could agglutinate leucocytes of different individuals. Later, it became clear that these sera were anti-HLA antibodies fromed due to the differences between the women HLA and their carried emberyos (had also paternal HLA antigens).

2. The MHC Genes: In humans, the MHC or HLA genes occupy a portion on the short arm of the 6 th chromosome which control the MHC (HLA) antigens. These genes are found in regions ABC and D (Fig.). The genes in each region are clustered and located in one or more locus ( locus = location of a gene). There are two groups of MHC genes controlling the proper MHC antigens. A third group is also present which mainly related to the complement system, and it does not belong to the MHC antigens: 1. Genes of class I MHC : A, B, C. 2. Genes of Class II MHC : DP, DQ, DR (D related) 3. Genes of Class III MHC: C2, C4A, C4B, BF, 21A, 21B TNF- alpha & beta (comple- ment or steroid related) Heat Shock Protein….. cont..

4. Cont/... MHC genes Each individual inherits one chromosome from each parent. So, there are MHC genes of maternal and paternal origin. Each half is called haplotype. The genes of both haplotypes are co-dominant. Each individual have a formula for the alleles in both haplotypes. In each locus there alleles of the genes; there are about 767 alleles for A, 1178 for B, 439 for C, and more than 900 for the D-related (DP, DQ & DR) genes. By combination of the alternative, there will be a possibility of 1/ 2,000,000 for a person to have another identical individual. The genes of haplotypes are inherited according to Mandelian Law of heredity; 25% of brothers/ sisters are identical, 50% are partially identical and 25% are non-identical.

5. Nomenclature of HLA antigens 1. Serologic: Give number, e.g. HLA- B27, HLA-DR3, HLA-DR4. 2. Genetic: Give 4-8 digit number, e.g. HLA- B*01:02, HLA-B* 01:04: 78: 23 ‘or HLA-B*01027823). Some times add letters as L, N, Q, S (e.g N= Null) First 2: group of alleles, Second 2: Synonymous alleles, Third 2: mutation in genes, Fourth 2: mutation outside genes.

6. The MHC (HLA) Antigens: Class I MHC antigens: These are membrane glycoproteins on the surface of all nucleated cells and platelets. They are not present on trophoblast cells, sperms and RBC. These antigens are composed of: 1. Heavy polypeptide chain (alpha): This has M.W. of 43 KD and three extracellular domains alpha 1, 2, & 3). This chain has also transmembranous portion and a small cytoplasmic tail. 2. Beta- 2 microglobulin: The heavy chain is non- covalently bound to Beta- 2 microglobulin which has M.W. of 12 KD and one extracellular domain only. This chain is encoded by chromosome 15. All domains of class I antigen has –S-S- bond except alpha-1 (Figure).

8. Class II MHC Antigens: They are glycoproteins found chiefly on the surface of APC, B cells activated T cells and some actively dividing cells as basal layer of epidermis and some testicular cells. They consist of two polypeptide chains: 1. Alpha- chain: has M.W. of 29 KD, two extra- cellular domains (alpha 1 and 2), tansmembranous portion and a small cytoplasmic tail. 2. Beta- chain: has M.W. Of 33 KD, two extracellular domains (beta 1 & 2), transmembranous portion and a small cytoplasmic tail, Figure. All domains have –S-S- bonds except alpha-1.

9. Non – classical HLA Loci: There are loci recently discovered which are non- classical loci: 1. For class I ; there are 2 loci which are G and E. The G is related to Beta-2 microglobulin and is resposible for the foetal- mother immunological control (no response to foetal antigens). The E is responsible for making complexes with antigens to prevent NK cells from attaking normal body cells or tissues. 2. For class II ; there are DO and DM loci. The DM is responsible for promoting the loading of peptides intracellularly. The DO modulates the function of DM. Also, there are areas called Tapacin Binding Proteins “TBP” (TAP1 and TAP2), and Low Molecular Protein Areas (LTLT). Both the TBP and LTLT are responsible for controlling the peptides binding inside the cells.

10. Specificity of HLA Antigens: The heavy chain of class I has hypervariable regions in its N- terminal (alpha-1 domain) which contains the antigen binding groove and constant domains. They can bind to 8 – 10 amino acid derived from endogenous antigens. The alpha chain of class II has variable (V), Joining (J) and constant (C) regions, while beta chain has VDJC regions. The specificity of these chains is in alpha-1 and beta-2 domains in particular the latter one. The class II antigen has also a single peptide binding groove which binds to 13 – 18 amino acids derived from exogenous antigens. Due to the similarity between the structures of Ig & HLA antigens, the latter are called Histoglobulins.

12. HLA Typing: Indications: 1. Transplantation; inparticular for kidney and bone marrow transplatations. 2. Disease association; there are certain diseases associated with specific HLA types as SLE with HLA-DR3, rheumatoid arthritis with HLA-DR4, myasthenia gravis with HLA- B8 or ankylosing spondylitis with HLA-B27. This association could be due to: (a) HLA antigens are similar to (molecular mimicry) or receptors for the causative agents. (b) HLA antigens are markers for immuno- suppression or immuno-stimulation. 3. Paternity testing; it is accepted in courts. 4. Anthropological studies.

14. Methods of HLA typing: 1. Microcytoxicity: For HLA class I and HLA- DQ. Principle; HLA antigen + HLA antibody + complement leads to lysis in microtiter plate. 2. Mixed Lymphocytes Reaction (MLR): For HLA- DR. Principle; different lymphocytes reacts against each other and become blast cells. 3. Primed Lymphocytes typing (PLT): For HLA- DP. Principle is similar to MLR. 4. Polymerase Chain Reaction (PCR): For any type, but inparticular the D- related antigens.

15. Preparations for transplatations: 1. ABO & Rh compatibility test. 2. Screening for reaction between leucocytes of recipient and serum of donor. 3. HLA typing; 2 loci compatibility as A & B preferably with DR. 4. Vaccination against common bacterial and viral diseases. 5. Antibiotics umbrella as penicillins or cephalosporins with aminoglycosides specially the new ones. 6. Splenectomy is preferable. 7. Use of immunosuppressive agents

16. Immunosuppressive Agents: Immunosuppressive therapy is used to prevent or treat graft rejection by non-specifically interfering with the induction or expression of the immune response. The following agents or measures are in use: 1. Immunosuppressive drugs: A. Cyclosporine A is an antibiotic produced by a fungus. It prevents T cells activation and blocks the accompanying cytokine production. B. FK – 506 (Tacrolimus), rapamycin (Sirolimus) or Everolimus are relatively new immunosuppressive drugs with action similar to cyclosporine. C. Corticosteroids are given in big doses; they act on blocking IL-1 & 2 release and cytokines receptor expression and suppressing macrophages. D. Anti-mitotic drugs as Azathioprine and Methotrexate which inhibit DNA synthesis and block the growth of T cells. All used in prevention, but Cortisones is also used in treatment. 2. Anti-lymphocyte or anti-thymocyte globulin or anti- CD3 MCA. These agents lyses T cells and can block their functions. Used in prevention and treatment. 3. Antibody to block co-stimulatory molecules. 4. Total lymphoid irradiation while shielding the marrow, lungs and other vital organs before engraftment. 5. Antigen specific immunosuppression by induction of tolerance to the graft antigen.

17. Types of grafts : 1. Autograft is the transfer of an individual own tissues from place to place, e.g., skin. Accepted. 2. Isograft (syngeneic)is the transfer of tissues between genetically identical individuals, e.g., identical twins. Accepted. 3. Allografts (homograft) is a graft between genetically different members of the same species as humans, e.g., Kidney. Rejection may occur. 4. Xenograft (heterograft) is the transfer of tissues between different species. Always rejected. 5. Split graft: e.g. part of liver. 6. Domino graft: Combined lung/Heart.

18. Types of graft rejection: 1. Hyperacute: This occurs minutes or hours after transplantation. Graft should be removed. 2. Acute rejection: This occurs 10 – 30 days after transplantation. It is mainly cell mediated. 3. Chronic or late rejection: This occurs slowly over a period of months or years. It may be cell mediated or antibody mediated or both.

19. Graft Versus Host Disease (GvHD): This condition occurs when there are: 1. Histoincompatibility between donor and recipient tissues. 2. Immuno-compromized recipient (host). 3. Immunocompetent donor tissues. The grafted cells survive and react against the host cells or tissues (i.e. Instead of reaction of host against the graft, the reverse occurs). There are three types of GvHD: 1. Hyperacute: maculopapular skin rash. Dangerous. 2. Acute: Extensive skin rash with hepato-splenomegaly, lymphadenopathy, cardiac irritability, and confusion. The patient is very ill. 3. Chromic GvHD: Hepato-splenomegaly and lymphadenopathy with some skin rash. The transplanted organ is functionally suboptimal. Treatment: The GvHD is very serious condition. It has mortality up to 90%. The best treatment is combination of Methotrexate + corticosteroids + Anti-sera. So, the prevention is better than treatment.

20. Prevention of GvHD: 1. Anti- CD3 MCA to remove T cells from grafted tissue as the bone marrow. 2. The blood or blood products should be irradiated prior giving to the recipient using about 3000 Rad. 3. Take all the measures necessary before transplantation (precautions) to prevent GvHD rather than treating it.