Application of HLA typing studies

Application of HLA typing studies

1. 1.CMH molecules

   • The major histocompatibility complex
     • HLA genes
     • Region I: class I genes
       • six loci code for CMH-I or class I molecules= ABCEFG
         • ■Classical class I molecules.
           • encoded by the A B C loci expressed on the surface of nucleated cells and mark them as self
         • ■Class I molecules
           • in the classical loci E F G immune cells
     • Region II: class II genes
     • Region III: class III genes
   • HLA inheritance
     • Haplotype is a group of genes that are inherited en bloc, with hardly any recombination.
       • Autosomal
       • Monofactorial
       • Dominant
         • Codominance
         • linkage disequilibrium
   • The expression of the molecules
     • CMH Class 1 molecules
     • These genes are not expressed indifferently in all cells
       • ■Class I genes
         • are expressed in the membrane of all nucleated cells, with the exception of trophoblasts and sperm.
       • ■Class II genes
         • are expressed only in antigen-presenting cells. These cells have their molecules CMH-I
     • Class II molecules
       • HLA-DP
       • HLA-DQ
       • HLA-DRE
   • CMH molecules
     • presentation of antigens to lymphocytes
     • CMH-II molecules
       • two chains, a and B. very similar to each other. Each of them is made up of two domains (a1, a2 and B1, B2).
     • CMH-I molecules
       • identification of one's own except for trophoblasts and sperm.
         • Chain a
         • Chain B
   • Genetic variability
     • polymorphic genes
   • Biological functions of CMH molecules
     • TCR antigen presentation
       • The peptide/CMH-II complex is recognized. by Th lymphocytes (CD4+).
       • The peptide/CMH-I group is recognized by Tc lymphocytes (CD8+).
     • Protect own cells from NK cytotoxicity
       • NK cells destroy all cells in which self-MHC is not expressed; That is why molecules are necessary, which act as ligands for NK inhibition receptors
     • Identify each individual
       • Polymorphism of CMH molecules
       • presentation of peptides to cytotoxic T lymphocytes
       • limited number of CMH molecules

2. 2.Histocompatibility studies

   • Serological techniques
     • Microlymphocytotoxicity technique
       • Dr. Paul Terasaki developed this technique in 1964. for HLA class I molecules separate B lymphocytes from T lymphocytes
       • Procedure: 1. Dispense each of the anti-HLA sera 2. Incubate for 30 min at room temperature so that the antibodies (anti-HLA) bind to the lymphocytes that have the specific antigens. 3. Add the complement formed immune complexes (HLA Ag-anti-HLA Ac 4. Add the trypan blue vital dye and let it act for 10 min.
       • Limitations
         • use a battery with a very high number of serums
         • There are no sera against rare alleles
       • Not suitable for HLA typing of donors corpses
         • complement-fixing antibodies are detected
         • Serum always face lymphocytes
       • Positive control: lymphocytes known specific antibodies + complement
         • Negative control: lymphocytes +
     • match test to determine
       • causes hyperacute rejection.
       • A lysis greater than 20% is considered a positive crossmatch. Transplantation is contraindicated
       • A lysis of less than 20% is considered a negative crossmatch
       • The transplant is viable
       • the presence of specific antibodies against HLA alleles in serum
       • False positives
         • presence of anti-HLA IgG, and The negative result .
         • ■antibodies (IgM). Negative results
         • These autoantibodies are IgM that react at low temperature
     • Detection of anti-HLA antibodies (PRA)
       • 20 individuals 40 alleles from each locus
       • This technique calculates the percentage of cells that are lysed in the assay
       • The higher the PRA, the lower the chances of being transplanted.
   • Molecular techniques
     • a single antigen defined with serological tests could consist of multiple variants
       • PCR-SSO
         • subsequent hybridization with specific probes (SSO) marked
         • reversa
         • process a high number of samples
       • PCR-SSP
         • verified by electrophoresis
         • produces amplification when the DNA contains the allele that corresponds to the set of primers for the particular reaction
       • PCR-SBT
         • gold standard
         • exon sequencing
         • they compare like this with databases

3. 3.Applications of histocompatibility studies

   • Histocompatibility for transplants
     • The graft
       • Autograft
         • The tissue comes from the same individual in which it was transplant.
       • Isograft
         • There is no rejection between univiteline twins
       • allograft
         • Same species
       • Xenograft
         • different species
     • Compatibility
       • better selection of compatible donor-recipient couples
       • better selection of compatible donor-recipient couples
     • Rejection
       • recognizes the TCR is the self/foreign peptide MHC complex
     • direct allorecognition
       • The recipient's LT recognizes the donor's CPA CMH molecule as non-self
     • indirect allorecognition
       • It is the recipient's CPA that recognizes foreign HSC in the graft
     • Sensitization of patients who are candidates for transplants.
       • identify if the patient has anti-HLA autoantibodies, see their specificity and monitor
     • bone marrow transplant
       • They recognize the receptor's MHC-peptide complex in different tissues of the recipient, which produces graft-versus-host disease of its own MHC, distinct from that of the recipient.
   • Paternity studies
     • The discovery of the ABO system of red blood cells in 1900
     • They are based on DNA repeated in tanden and the so-called minisatellites and microsatellites are used, which consist of repetitions of DNA fragments of variable numbers.
   • Anthropological studies
     • Know which are the most frequent alleles in certain population groups, Through HLA studies it is known that populations.
     • Found the most frequent alleles in certain population groups
   • Association of MHC alleles with diseases
     • HLA-B27 with ankylosing spondylitis
     • HLA complex of cla sell DR4 and DR1.rheumatoid arthritis
     • The 8*27:05 allele has been found mostly in all studied populations
     • Alleles B 27:01, 02, 08 and 09 have been described in Caucasoids
     • Alleles 8*27:04, 06 and 07 have only been observed in populations. oriental.
     • The B*27:03 allele has only been observed in populations from East Africa:

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