Lecture 7 - Autoimmunity, Autoimmune Diseases

• Disorders in which immune responses are targeted toward self-antigens and result in damage to organs and tissues in the body
• Could be T cell mediated or due to Abs to host Ags
• More than 100 autoimmune diseases have been discovered, and these can involve various organ system
• Autoimmune diseases are a leading cause of chronic illness and death, affecting about 5% of the world’s population, including more than 20 million people in the United States alone

1. Self-tolerance
- central (thymus or bone marrow)
- peripheral (lymph nodes, spleen)
2. Genetics
3. Hormonal influence
4. Exposure of cryptic antigens due to tissue damage following, e.g., infections, toxins, injury
5. Microbial infections: the main mechanism is molecular mimicry

- ability of the immune system to discriminate “self” from “non-self” or “foreign”, unresponsiveness to self Ags
- Lymphocytes must be “educated” at two levels (layers of protection) during development or immune response in order for self-tolerance to develop:

- Central: which occurs in thymus for T cells and in BM for B cells. CD4+ T cells or B cells having receptors reactive to self Ags will be removed, some self-reactive CD4+ T cells are not deleted but released into circulation to differentiate into Treg cells.
• Some self-reactive B cells also are released to modify their receptors, so they become non-reactive to Ags (receptor editing).
• If some B cells have BCRs that weakly react with self Ags, they will be induced to become anergic (downregulate their receptor)

Peripheral: occurs in spleen or lymph nodes
• T cells reactive to self Ags will be removed either through apoptosis, inhibition by Treg cells, by losing a co-stimulatory signal from APCs, or through binding of CTLA-4 (inhibits T cells)
• B cells reactive to self Ags will either be removed by apoptosis, become anergic after repetitive exposure to self Ags, or become inactivated through activation of CD22 receptor

In some individuals, self-tolerance can fail even after this second layer of protection; if this happens, autoimmunity can arise.

The development of autoimmune disease is thought to be caused by complex interactions between genetics, exposure to environmental factors, and defects in immune regulation.

• Important factor (autoimmune diseases more prevalent among family members, or more prevalent among monozygotic (genetically identical) twins than dizygotic (non-identical) twins or siblings
• Most studies in humans on MHC genes
• Polymorphism in Non-MHC genes can also be involved

- people inheriting HLA-B27 allele are at 100 times higher risk of developing ankylosing spondylitis

- IL2RA gene, which is involved in T-cell activation and maintenance of Tregs
- CTLA4 gene, which has an inhibitory effect on T-cell activation
- BLK gene, which is involved in B-cell activation and development

Estrogens promote type 2 helper cell (Th2) response, resulting in more B cell activation and antibody production, whereas androgens promote type 1 helper cell (Th1) response with activation of CD8+ T cells

- autoantibodies to the lens of the eye following an ocular injury
- autoantibodies to sperm after a vasectomy
- autoantibodies to DNA following damage to skin cells by overexposure to UV rays from the sun

• The main mechanism is molecular mimicry:
• Bacterial or viral agents contain antigens that closely resemble the structure or amino acid sequence of self-antigens. Exposure to such foreign antigens may trigger immune responses that cross-react with similar self-antigens

- association between Streptococcus pyogenes and rheumatic
fever
- an autoimmune disorder that primarily affects the joints and the heart. Patients develop antibodies to the M protein and N-acetyl glucosamine components of the bacteria, which cross-react with cardiac myosin, causing damage to the heart

Although the precise etiology of autoimmunity is unknown, there is much evidence that suggests that this heterogeneous disease entity is caused by complex interactions between genetic and environmental factors

• Chronic inflammatory disease
• Females are more likely to develop SLE than males, by a ratio of 9:1

• More than 100 autoantibodies identified
• Abs against: dsDNA, histones, as well as autoantibodies to lymphocytes, erythrocytes, platelets, phospholipids, ribosomal components, and endothelium. The typical patient has several circulating autoantibodies.
• Anti-dsDNA complexed with complement circulate (cannot be cleared from circulation) and deposit on glomerular basement membrane (GBM) of the kidneys
• Joint, skin, renal involvement
• Lupus anticoagulants

- CBC, platelet count, creatinine levels, and urinalysis to
assess renal function
- it may show leukopenia, possible thrombocytopenia and anemia, ESR may also be high, but CRP may be low or normal

- (anti-nuclear antibodies) ANA will be done which is the most important screening test
- After ANA screening is positive, further tests should be done to differentiate the type of ANA

Antibodies to dsDNA and antibodies to the Sm nuclear antigen are highly specific for SLE

Fluorescent anti-nuclear antibody (FANA) testing has been the most widely used and accepted test because it is highly sensitive, detects a wide range of antibodies, and is inexpensive and easy to perform

• A commercially prepared slide with fixed HEp2 cell line
• Incubation with patient’s serum (serial dilutions, starting with 1:80 or 1:160), ANA will bind to HEp2 nucleus
• Wash, to remove unbound Ab molecules
• Add anti-human Ig labeled with a fluorescent dye and incubate
• Wash again to remove unbound Ab
• Put a cover slip, mount the slide and view it under microscope (400× magnification)
• Titers above 160 are clinically significant

1. homogenous
2. speckled
3. nucleolar

- Homogenous!
- associated with antibodies to dsDNA, histones, and nucleosomes
- found in SLE patients, drug-induced lupus, chronic autoimmune hepatitis (AIH), and juvenile idiopathic arthiritis

- Speckled!
- associated with antibodies to ENAs
- found in SLE patients, Sjogren's syndrome, systemic sclerosis (SSc), and other SARDs

- Nucleolar!
- primarily caused by antibodies to RNA and RNP
- seen in patients with SSc, but can also be seen in other SARDs

• Caused by an inflammatory process that results in the destruction of bone and cartilage in the joints
• Infiltration of the inflamed synovium with T and B lymphocytes, plasma cells, dendritic cells, mast cells, and granulocytes is evidence of immunologic activity within the joint.

>100 genetic regions

- specific HLA-DRB1 alleles or PTPN22 gene polymorphisms
- these patients are positive for rheumatoid factor (RF) or antibodies to cyclic citrullinated peptides (CCP)

• Epigenetic factors, such as altered DNA methylation affecting the transcription of critical genes, are also believed to play a role in the pathogenesis of RA.
• The strongest environmental risk factor for RA is believed to be cigarette smoking, which doubles the risk of developing the disease.

• The balance between proinflammatory and anti-inflammatory cytokines in RA appears to be tipped toward continual inflammation: IL-1, IL-6, IL-17 and TNF-a (TNF-a plays the key role)
• These cytokines trigger the release of matrix metalloproteinases from fibroblasts and macrophages; these enzymes degrade important structural proteins in the cartilage
• Bone lesions due to hyperactivation of osteoclasts.
• TNF-a and RANKL ((receptor activator of nuclear factor kappa-B ligand), inhibit osteoblast differentiation and bone formation, they activate osteoclasts
• Two key Abs

1. Rheumatoid factor (RF)
2. anti-CCP or ACPA

an IgM against FC portion of IgG, enhances activity of macrophages and Ag presentation to APCs

- Anti-cyclic citrullinated peptide antibody [anti-CCP or ACPA])
- citrullinated proteins released from dead granulocytes or macrophages
- overexpression of these antigens may provoke an immune response in individuals with certain HLA-DRB1 alleles.

- initial test is RF
- negative RF does not exclude RA,
- positive RF is not specific to RA
- RF is also positive in about 5% of healthy individuals and in 10% to 25% of those over the age of 65. RF can be found in patients with other SARDs, such as SLE, Sjögren’s syndrome, scleroderma, as well as in people with some chronic infections

• RF is done using latex agglutination on a slide, detects only IgM isotype
• ELISA and nephelometry are more sensitive and precise, they detect IgG isotype (more specific)

• Anti-CCP is done using ELISA, great test for early diagnosis where RF may be negative
• A large number of RF-negative RA patients are anti-CCP positive