HLA-TYPING Dr Opoola A.O. Registrar,LUTH. Outline Definition of terms Classification of HLA types functions Characteristics Clinical significance.

HLA-TYPINGDr Opoola A.O.Registrar,LUTH

OutlineDefinition of termsClassification of HLA types functionsCharacteristicsClinical significanceReferences

Definition

The HLA system is a group of tissue antigens termed Human Leukocyte Antigens governed by a chromosomal region bearing a number of genetic Loci, each with multiple alleles that have relevance to transplantation rejection reactions and that mark the prevalence of

several diseases. This group of genes resides on the short

arm of Chromosome 6 and encodes Cell-Surface antigen-presenting proteins and many other genes.

BINDING OF ANTIGEN BY MHC AND T CELL RECEPTOR

Definition of terms CHROMOSOME: One of the threadlike

structures in a cell nucleus that carry the genetic information in the form of gene.

ALLELES: Variants of a single genetic locus.

MAJOR HISTOCOMPATIBILITY COMPLEX: A cluster of genes located in close proximity e.g. on human chromosome 6, that encode the histocompatibility antigens (MHC Molecules).

Definition of terms HISTOCOMPATIBLE: Sharing similar

antigens i.e in transplantation

ANTIBODY: A protein produced as a result of interaction with an antigen. The protein has the ability to combine with the antigen that stimulated its production.

ANTIGEN: A substance that can react with antibody. Not all antigens can induce antibody production; those that can are also called immunogens.

Definition of terms B LYMPHOCYTES: Strictly, a bursa-

derived cell in avian species and by analogy, a cell derived from the equivalent of the bursa in non-avian species. B Cells are the precursors of plasma cells that produce antibody.

T LYMPHOCYTE: A thymus – derived cell that participates in a variety of cell-mediated immune reactions.

ANTIGEN PRESENTING CELLS: Chiefly Macrophages or B Cells that process and present antigens to T Cells.

Definition of terms MACROPHAGES: A phagocytic

mononuclear cells derived from bone monocytes and found in tissues and at the site of inflammation.

Macrophages serve accessory roles

in immunity, particularly as APCs

COMPLEMENT: A set of plasma proteins that is the primary mediator of antigen – antibody reactions.

Functions The HLA System (MHC MOLECULES) bind

peptide antigens and present them to T-Cells.

Thus these transplantation antigens are responsible for antigen recognition by the T-Cell receptor.

While antibody molecules interact with antigens directly, the T-Cell receptor

only recognizes antigen presented by MHC molecules on an antigen presenting cells .

Function The T Cell receptor is also specific

for the MHC molecule.

If the antigen is presented by another allelic form of the MHC molecule there is no recognition by the T Cell receptor.

This phenomenon is known as MHC

RESTRICTION

ClassificationThe HLA system is classified into 3 based on their location on the chromosome and cell distribution. These are:-

1.Class 1: 3 major and 3minor

2.Class 2: 3major and 2 minor

3.Class 3: Complement proteins and several cytokines

Class 1MHC Proteins These major class I genes are encoded by

the HLA-A, - B, - C genes.

These proteins are made up of two chains:

(i) A transmembrane glycoprotein of MW 45,000, non-covalently associated with a non-MHC encoded polypeptide of MW 12,000 that is known as B2 – macroglobulin.

Class 1 molecules are to be found on virtually all nucleated cells in the body.

Minor genes are HLA – E, HLA – F and HLA-G.

HLA Class 1 molecule

Class 2 MHC proteinsMajor are HLA-DP,DQ and DR and

minor are HLA-DM and DO.

The genes of the Class II combine to form heterodimeric (αβ) protein receptors that are typically expressed on the surface of antigen presenting cells.

Class 2 MHC proteins1. HLA-DP α- chain encoded by HLA-DPA1 locus β-chain encoded by HLA-DPB1 locus

2. HLA-DQ α-chain encoded by HLA-DQA1 locus β-chain encoded by HLA-DQB1 locus

3. HLA-DR α-chain encoded by HLA-DRA locus 4 β-chains (only 3 possible per person) encoded by HLA-DRB1, DRB3, DRB4, DRB5 loci

MHC Class 2 Molecule

MHC Class 2 molecule

Class 2 MHC proteinsDM and DO are used in the internal

processing of antigens, loading the antigenic peptides generated from pathogens onto the HLA molecules of antigen-presenting cell.

The HLA – D Locus encoded proteins are made up of non-covalently associated transmembrane glycoprotein of about MW 33,000 and MW 29,000

Class 2 MHC proteins Unlike Class I proteins, they have a

restricted tissue distribution and are chiefly found on macrophages, B-Cells, and other APCs.

Their expression on the other Cells – e.g., endothelial cells can be induced by interferon-gamma.

CLASS 1 CLASS 11

GENETIC LOCI (PARTIAL LIST)

HLA – A, - B, - C HLA-DP, - DQ, - DR

POLYPEPTIDE COMPOSITION

MW 45,000 + B2 M (MW 12,000)

α CHAIN (MW 33,000)β CHAIN (MW 29,000)

CELL DISTRIBUTION ALL NUCLEATED CELLS (SOMATIC)

APCs (Macrophages, B-Cells etc) Activated human T-Cells

PRESENT ANTIGENS TO

CD8 T CELLS CD4 T CELLS

SIZE OF PEPTIDE BOUND

8 – 11 RESIDUES 10 – 30 OR MORE RESIDUES

TABLE SHOWING FEATURES OF CLASS I AND CLASS II PROTEINS

CHARACTERISTICS OF HLA THEY ARE POLYGENIC: There are

several genes for each class of molecule.

THEY ARE POLYMORPHIC: They have a large number of alleles in the population for each of the genes.

THEY ARE HAPLOTYPIC: Genes tend to be inherited as a block as there are relatively in-frequent cross-over events at the Locus.

CHARACTERISTICS OF HLA LINKAGE DISEQUILLIBRIUM: This is defined

as a deviation from Hardy – Weinberg equilibrium for alleles at linked loci. One consequence of this has been the resulting deficiency in assigning HLA- disease associations to a single allele at a single Locus.

Characteristics The proteins encoded by the HLAs are the

proteins on the outerpart of body cells that are unique to that person.

The immune system uses the HLAs to

differentiate self cells and non-self cells.

Any cell displaying that persons HLA type belongs to that person

Clinical Significance 1. In DISEASE DEFENSE

2. AGENTS OF HUMAN DISEASE IN AUTOIMMUNITY: Known to mediate many autoimmune diseases

3. AS ANTIGENS: Responsible for organ transplant rejection

4. IN REPRODUCTION – may be involved in male selection

5. IN CANCER – may be protective or fail to protect.

Clinical Significance IN INFECTIOUS DISEASE: When a foreign pathogen enters the body, specific cells called antigen-presenting cells (APCs) engulf the pathogen through a process called phagocytosis.

Proteins from the pathogen are digested into small peptides and loaded onto HLA antigens (Specifically MHC Class II).

They are then displayed by the APCs for the T Cells to produce a variety of effects to eliminate the pathogen.

Clinical Significance Through a similar process, proteins (both

native and foreign, such as the protein of viruses) produced inside most cells are displayed on HLA antigens (specifically MHC Class I) on the cell surface.

Infected cells can be recognised and destroyed by components of the immune system (Specifically CD8+ T Cells).

DR PROTEIN WITH BOUND STAPHYLOCOCAL ENTEROTOXIN LIGAND

Clinical Significance IN AUTOIMMUNITYHLA types are inherited, and some of them are connected with autoimmune disorders and other diseases.

People with certain HLA antigens are more likely to develop certain autoimmune diseases such as Type 1 Diabetes, Ankylosing Spondylitis, celiac disease, SLE, myasthenia Gravis, inclusion body myositis and Sjogren’s syndrome.

Clinical significance HLA typing in autoimmunity is being

increasingly used as a tool in diagnosis.

In Gluten Sensitive Enteropathy (GSE), it is the only effective means of discriminating between first degree relatives who are at risk from those who are not at risk, prior to the appearance of sometimes irreversible symptoms such as allergies and secondary autoimmune disease.

 Diseases Marker

Gene

Spondyloarthropathies Ankylosing spondylitis,Reiter’s syndrome, Acute anterior uveitis, Reactive arthritis(Yersinia, Salmonella, Shigella, Chlamydia), Psoriatic Spondylitis

B27 B*2702, –04, –05

Collagen-Vascular Diseases Juvenile arthritis, pauciarticular DR8,DR5  

Rheumatoid arthritis DR4 DRB1*0401, –04, –05

Sjögren's syndrome DR3  Systemic lupus erythematosus  White DR3    Japanese DR2  

Disease Marker Gene

Autoimmune Gut and Skin 

Gluten-sensitive enteropathy (celiac disease)

DQ2 DQA1*0501

    DQB1*0201

Chronic active hepatitis DR3  

Dermatitis herpetiformis DR3  

Psoriasis vulgaris Cw6  

Pemphigus vulgaris DR4 DRB1*0402

  DQ1 DQB1*0503

Bullous pemphigoid variant DQ7 DQB1*0301

Disease Marker Gene

Autoimmune Endocrine 

Type 1 diabetes mellitus DQ8 DQB1*0302   DR4 DRB1*0401, –

04   DR3    DR2 DQB1 *0602Hyperthyroidism (Graves') B8, DR3  

Hyperthyroidism (Japanese) B35  

Adrenal insufficiency DR3  Autoimmune Neurologic 

Myasthenia gravis B8, DR3  

Multiple sclerosis DR2 DRB1*1501

    DRB5*0101

Miscellaneous Diseases Markers

Gene

Behçet's disease B51  

Congenital adrenal hyperplasia B47 21·OH (Cyp21B)

Narcolepsy DR2 DQB1*0602

Goodpasture's syndrome (anti-GBM)

DR2  

Abacavir hypersensitivity B57 B*5701

HLA and autoimmunity diseases diseasesHLA allele Diseases with increased risk

HLA-B27Ankylosing spondylitis, post gonococcal arthritis, anterior uveitis

HLA-B47 21-hydroxylase deficiency

HLA-DR2 Systemic Lupus Erythematosus

HLA-DR3Autoimmune Hepatitis, Primary Sjogren syndrome, Diabetes mellitus type 1, SLE

HLA-DR4Rheumatoid Arthritis, Diabetes mellitus type 1

Clinical SignificanceIN GRAFT REJECTION Any cell displaying some other HLA type is “non-self” and is an invader resulting in the rejection of the tissue bearing those cells.

Because of the importance of HLA in transplantation, the HLA Loci are among the most frequent typed by serology or PCR relative to any other autosomal alleles.

HLA-A,B &DR-Important in Kidney transplant

HLA/TISSUE TYPING1.HLA- A1*,A2,B8*,B44,DR4*,DR152.HLA-A1*,A3,B7,B8*,DR4*,DR121A, 1B, 1DR mismatch 1.HLA- A2 - ; B27, B13, DR17, DR42.HLA- A2 A3, B8, B14, DR17 -

Once there is double DR mismatch-No transplant*

Clinical Significance IN CANCER:Some HLA mediated diseases are directly involved in the promotion of cancer.

Gluten sensitive enteropathy is associated with increased prevalence of enteritis associated T-Cell Lymphoma and DR3 – DQ2 homozygotes are within the highest risk group with close to 80% of gluten sensitive EATL cases.

Clinical Significance More often; however, HLA molecules play a

protective role, recognizing the increase in antigens that were not tolerated.

Abnormal cells may be targeted for apoptosis mediating many cancers before clinical diagnosis.

Prevention of cancer may be a portion of heterozygous selection acting on HLA

References The HLA sysyem and its value in clinical

medicine lecture by Dr Mankwe

The HLA system and its products. Harrison Principles of Internal Medicine 18th edition pages kasper et al pages 2691-2694