Autoimmune disease is a group of disorders in which tissue injury is caused by humeral by autoantibodies or cells mediated immune response by auto reactive T cells to self antigens Normal the immune system does not attach the self However there is a large ID: 909052
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Slide1
Auto Immune Diseases
Slide2Autoimmune Diseases
Autoimmune disease is a group of disorders in which tissue injury is caused by humeral (by auto-antibodies) or cells mediated immune response (by auto reactive T cells) to self antigens.
Normal, the immune system does not attach the self. However, there is a large
g
roup of autoimmune diseases in which the immune system does attack self-cells.
The attack can be directed either against a very specific tissue or to a large no. of tissues.
Once started, autoimmune disease are hard to stop.
Slide3Causes of Autoimmune Diseases
Sequestered or Hidden antigens.
Ag in the secluded places – are not accessible to the immune system.
E.G. Lens Ag, Sperm Ag and Thyroglobulin.
Neo Antigens.
Altered or Modified Antigens –by physical (irradiation), chemical (drugs) or microbial agents (intracellular viruses)
Cessation of Tolerance
It may
result
when tolerance to the self Ag in abrogated
.
Slide4Cross reacting Antigens
A foreign Ag which resembles self a 2nd Ag.
Many Species share organ specific Ags.
E.g. Ag of Human brain and Ag of sheep brain, streptococcal M protein and heart muscles, Nephritogenic strains of streptococci Ags and Renal glomeruli shares similar epitopes.
Loss
of Immunoregulation.
Loss of Self tolerance – caused by over activity or
l
owered activity of T and B-cells.
Slide5Classification of Autoimmune Disease
Hemolytic autoimmune diseases
Localized autoimmune disease.
Systemic autoimmune diseases
Slide6Hemolytic autoimmune diseases
Clinical disorder due to destructions of blood components, auto Ab are formed against one’s own RBCs, Platelets or leucocytes.
E.g.:
Haemolytic
Anemia, Leucopenia,
Throbocytopenia
.
Slide7Localized autoimmune diseases.
Called also Organ specific autoimmune diseases.
A particular organ is affected due to auto Abs.
For Example:
Thyroiditis (attack the Thyroid).
Multiple sclerosis (attacks myelin coating of nerve axons).
Myasthenia gravis (attacks nerve-muscle junction).
Juvenile diabetes or Type I DM (attack insulin-producing cells).
Celiac disease.
Slide8Systemic autoimmune diseases.
Called also Non-organ specific autoimmune diseases.
Immune complexes accumulate in many tissue and cause inflammation and damage.
Affects many organs or the whole body.
For Examples:
Systemic Lupus Erythematous (anti-nuclear Ab); Harms kidneys, heart, brain, lungs, skin,……
Rheumatoid Arthritis (anti-
IgG
antibodies); Joints, hearts, lungs, nervous
Rheumatic fever: cross-reaction between antibodies to streptococcus and auto-antibodies.
Slide9Celiac disease
Celiac disease
-- also known
as
celiac
sprue
or
gluten-sensitive enteropathy
-- is a digestive and autoimmune disorder that results in damage to the lining of the small intestine when foods with gluten are eaten.
Gluten
is a form of protein found in some grains.
The
damage to the intestine makes it hard for the body to absorb nutrients, especially fat
, calcium
, iron, and folate
Slide10What Causes Celiac Disease?
When
people with celiac disease eat foods containing gluten, their immune system forms antibodies to gluten which then attack the intestinal lining.
This
causes inflammation in the intestines and damages the villi, the hair-like structures on the lining of the small intestine
.
Nutrients
from food are normally absorbed by the villi. If the villi are damaged, the person cannot absorb nutrients properly and ends up malnourished, no matter how much he or she eats.
Slide11Symptoms
of Celiac
Disease
Symptoms
of celiac disease vary among sufferers and include:
Digestive problems (abdominal bloating, pain, gas, diarrhea, pale stools, and weight loss)
A severe skin rash called dermatitis
herpetiformis
Iron deficiency anemia (low blood count)
Musculoskeletal problems (muscle cramps, joint and bone pain)
Growth problems and failure to thrive (in children)
Seizures
Tingling sensation in the legs (caused by nerve damage and low calcium)
Aphthous
ulcers (sores in the mouth)
Missed menstrual periods
Slide12Health Problems Accompany Celiac Disease
Celiac
disease can leave a person susceptible to other health problems, including:
Osteoporosis, a disease that weakens bones and leads to fractures. This occurs because the person has trouble absorbing enough calcium and vitamin D.
Miscarriage or infertility.
Birth defects, such as neural tube defects (improper formation of the spine) caused by poor absorption of such nutrients as folic acid.
Seizures.
Growth problems in children because they don't absorb enough nutrients.
Cancer of the intestine (very rare).
Slide13Diagnosis
The two major steps leading to diagnosis of celiac disease are:
Lab. tests
for gluten autoantibodies (These are IgA based tests accurate only while on a gluten-containing diet)
A
small bowel biopsy to assess gut damage. For those with suspected dermatitis
herpetiformis
, skin biopsies will be taken of the skin near the lesion.
Slide14Lab diagnosis
Anti-tissue transglutaminase antibody (anti-
tTG
), IgA:
Detects
antibodies to tissue transglutaminase, an enzyme that causes the crosslinking of certain proteins.
Anti-
tTG
, IgA is the most sensitive and specific blood test for celiac disease.
The
IgG class of anti-
tTG
may be ordered for people who have a deficiency of IgA
.
Deamidated
gliadin peptide (DGP) antibodies, IgA:
Detects anti-DGP
IgA antibodies; like anti-
tTG
, the IgG class may be performed for a person with an IgA deficiency
.
Slide15Other tests less commonly performed include:
Anti-
endomysial
antibodies (EMA), IgA class:
Detects antibodies to endomysium, the thin connective tissue layer that covers individual muscle fibers
Anti-
reticulin
antibodies (ARA), IgA class: not as specific or sensitive as the other autoantibodi
es
Lab diagnosis
Slide16Diagnosis of Auto-Immune Disease
Diagnosed by Clinical symptoms
Confirmed by detecting the auto Ab in the serum of the patients.
Autoantibodies are demonstrated by immunoflurescent Ab test , Haemagglutination, Complement fixation, immundodiffusion, Radio immuno assay, etc.
Slide17Immunofluorescence assay
Immunofluorescence is a technique allowing the visualization of a specific protein or antigen in tissue sections by binding a specific antibody chemically conjugated with a fluorescent dye such as fluorescein isothiocyanante (FITC)
The specific antibodies are labeled with a compound (FITC) that makes them glow an apple-green color when observed microscopically under ultraviolet light.
Slide18Fluorescence
is the property of certain molecules of fluorophores to absorb light at one wave length and emit light at longer wave length (emission wavelength) when it is illuminated by light of a different wave length (excitation wavelength).
The incident light excites the molecule to a higher level of vibration energy. As the molecules return to the ground state, the excited fluorophores emits a photon (=fluorescence emission).
Slide19Slide20Slide21Slide22Type of Immunofluorescence
Direct immunofluorescence : Staining in which the primary antibody is labeled with fluorescence dye.
Indirect immunofluorescence: staining in which a secondary antibody labeled with fluorochome is used to recognize a primary antibody.
Slide23Direct Immunofluorescence:
It is a simple procedure.
Ag is fixed on the slide
Fluorescein labeled
Ab’s
are layered over it.
Slide is washed to remove unattached
Ab’s
.
Examined under UV light using fluorescent microscope .
The site where the Ab attached to its specific Ag will show apple green fluorescence.
Use: direct detection of pathogens or their Ag’s tissues or in pathological samples.
Slide24Indirect Immunofluorescence
Indirect test is a double-layer technique.
The unlabeled antibody is applied directly to the tissue substrate.
Treated with a fluorochome-conjugated anti-immunoglobulin serum
Slide25Slide26Advangate
of indirect
Gives an amplification effect – more tag or label (signal) per molecule of target protein.
Requires only one labeled antibody to identify many protein– same labeled secondary antibody can be used to bind to (light up) many different proteins ( preparation of labeled antibody is difficult and expensive.
Slide27Slide28Slide29Systemic Lupus
Erythematosus
(SLE)
It is a skin disease due to the production of anti-nuclear factor (ANF) or anti-nuclear auto Ab.
ANF reacts with the breakdown products of nuclei in the normal wear and tear of cells and form immune complexes which cause the tissue damage.
In these patients, LE cell ( a mature neutrophil) appears in blood and bone marrow.
Slide30Characteristics of SLE
Appearance of blood red spots over the bridge of nose and cheeks. The lesion take the shape of a butterfly.
Connective tissue of the skin, kidney, heart, spleen and blood vessels are severely damaged resulting in joint pain, fever and anemia.
Glomerulonephritis due to deposition of immune complex in the glomerulus region.
It is a systemic disease affecting the whole body.
Slide31Malar Rash (SLE)
Slide32LE cells
the
LE cell reaction is positive in 50%-75% of individuals with acute disseminated lupus.
Positive reactions are also
seen in rheumatoid arthritis, chronic hepatitis, acquired hemolytic anemia, and Hodgkin disease.
It may also be positive in persons taking
phenylbutazone
and hydralazine.
Slide33LE cells
An LE cell is either a neutrophil or a macrophage that has engulfed (phagocytized) degraded nuclear material from another cell. The engulfed nuclear material takes up
Haematoxylin
stain strongly; this strongly-stained engulfed nuclear material is called LE body
Detection of LE cell is made through microscopic examination.
The
test was commonly used in the past to diagnose systemic lupus erythematosus. But currently, SLE is diagnosed by more sensitive and specific tests such as anti-nuclear antibody (ANA) blood test
Slide34Diagnosis of SLE
The
American
College of Rheumatology (ACR) criteria summarized features necessary to diagnose
SLE at 1982.
These
criteria were last updated in
1997.
The
presence of 4 of the 11 criteria yields a sensitivity of 85% and a specificity of 95% for SLE
Patients
with SLE may present with any combination of clinical features and serologic evidence of lupus
.
The Systemic Lupus International Collaborating Clinics (SLICC) group revised and validated the ACR SLE classification criteria in 2012
.
According to the revision, a patient is classified as having SLE if the patient has biopsy-proven lupus nephritis with ANA or anti-
dsDNA
antibodies or if the patient satisfies 4 of the diagnostic criteria, including at least 1 clinical and 1 immunologic criterion.
Slide35ACR
diagnostic
criteria in SLE
Serositis
- Pleurisy, pericarditis on examination or diagnostic electrocardiogram (ECG) or imaging
Oral ulcers - Oral or nasopharyngeal, usually painless; palate is most specific
Arthritis -
Nonerosive
, 2 or more peripheral joints with tenderness or swelling
Photosensitivity - Unusual skin reaction to light exposure
Blood disorders - Leukopenia (< 4 × 10
3
cells/µL on >1 occasion),
lymphopenia
(< 1500 cells/µL on >1 occasion), thrombocytopenia (< 100 × 10
3
cells/µL in the absence of offending medications), hemolytic
anemia
Slide36ACR
diagnostic
criteria in SLE
Renal
involvement – Based on presence of proteinuria (>0.5 g/day or 3+ positive on dipstick testing) or cellular casts (including red blood cells [RBCs], hemoglobin, granular, tubular, or mixed)
or based on the opinion of a rheumatologist or nephrologist
Neurologic disorder - Seizures or psychosis in the absence of other
causes
Malar rash - Fixed erythema over the cheeks and nasal bridge, flat or raised
Discoid rash - Erythematous raised-rimmed lesions with
keratotic
scaling and follicular plugging, often scarring
Slide37Antinuclear
antibodies (ANAs) - Higher titers generally more specific (>1:160); must be in the absence of medications associated with drug-induced lupus
Immunologic
phenomena -
dsDNA
; anti-Smith (Sm) antibodies;
antiphospholipid
antibodies (
anticardiolipin
immunoglobulin G [
IgG
] or immunoglobulin M [
IgM
] or lupus anticoagulant); biologic false-positive serologic test results for syphilis, lupus erythematosus (LE) cells (omitted in 1997 revised criteria
)
Slide38In
patients with high clinical suspicion and/or high ANA titers, additional testing is indicated.
This
commonly includes evaluation of antibodies to
dsDNA
, complement, and ANA subtypes such as Sm, SSA, SSB, and
ribonucleoprotein
(RNP) (often called the ENA panel), as well as screening
anticardiolipin
antibodies, lupus anticoagulant, and +/- beta-2 glycoprotein antibodies.
ENA :Extractable
Nuclear Antigen Antibodies
Slide39Serological tests for SLE
Test
Description
ANA
Screening test; sensitivity 95%; not diagnostic without clinical features
Anti-dsDNA
High specificity; sensitivity only 70%; level is variable based on disease activity
Anti-Sm
Most specific antibody for SLE; only 30-40% sensitivity
Anti-SSA (Ro) or Anti-SSB (La)
Present in 15% of patients with SLE and other connective-tissue diseases such as
Sjögren
syndrome
; associated with
neonatal lupus
Slide40Serological tests for SLE
Anti-ribosomal P
Uncommon antibodies that may correlate with risk for CNS disease, including increased hazards of psychosis in a large inception cohort, although the exact role in clinical diagnosis is
debated
Anti-RNP
Included with anti-Sm, SSA, and SSB in the ENA profile; may indicate mixed connective-tissue disease with overlap SLE,
scleroderma
, and myositis
Anticardiolipin
IgG
/
IgM
variants measured with ELISA are among the
antiphospholipid
antibodies used to screen for
antiphospholipid
antibody syndrome and pertinent in SLE diagnosis
Lupus anticoagulant
Multiple tests (
eg
, direct Russell viper venom test) to screen for inhibitors in the clotting cascade in
antiphospholipid
antibody syndrome
Slide41Serological tests for SLE
Direct Coombs test
Coombs test–positive anemia to denote antibodies on RBCs
Anti-histone
Drug-induced lupus ANA antibodies are often of this type (
eg
, with procainamide or hydralazine; p-ANCA–positive in minocycline-induced drug-induced lupus)
ANA
= antinuclear antibody; CNS = central nervous system; ds-DNA = double-stranded DNA; ELISA = enzyme-linked immunoassay; ENA = extractable nuclear antigen; Ig = immunoglobulin; p-ANCA =
perinuclear
antineutrophil
cytoplasmic antibody; RBCs = red blood cells; RNP = ribonucleic protein; SLE = systemic lupus erythematosus; Sm = Smith; SSA =
Sjögren
syndrome A; SSB =
Sjögren
syndrome B.
Slide42Other useful tests in suspected SLE
Standard laboratory studies that are diagnostically useful when systemic lupus erythematosus (SLE) is suspected should include the following:
Complete blood count (CBC) with differential
Serum creatinine
Urinalysis with microscopy
The CBC count may help screen for leukopenia,
lymphopenia
, anemia, and thrombocytopenia. Urinalysis and creatinine studies may be useful to screen for kidney disease.
Slide43Levels of inflammatory markers, including the ESR and CRP, may be elevated in any inflammatory condition, including SLE.
Measurement
of complement may be useful, because C3 and C4 levels are often depressed in patients with active SLE as a result of consumption by immune complex–induced inflammation. In addition, some patients have congenital complement deficiency that predisposes them to SLE
.
Other useful tests in suspected SLE
Slide44Indirect immunofluorescence assay:
A laboratory test used to detect antibodies in serum or other body fluid.
Example of autoantibodies
Anti dsDNA Abs.
ANA
APA
ASMA
AMA
Anti LKM
ANCA
Antithyroid Abs
Slide45Substrate
Autoantibodies are detected on specific substrates:
Autoantibodies
Substrate
Anti-dsDNA
on
Crithedia
Lucilae
substrate
ANA
on Hep-2
substrate
or on Mouse
stomach kidney substrate
APA
ASMA
AMA
on mouse stomach kidney
substrate
Anti LKM
on mouse Liver stomach kidney
substrate
ANCA
on
neutrophil substrate
Anti Thyroid Abs
on
thyroid tissue substrate
Slide46Advantage of Hep2 cells over rodent tissue
Higher sensitivity (greater Ag expression)
Human origin ensure better specificity.
Cell division rates are higher so cell cycle dependent Ab are easily identified.
Nucleus are much larger, visible and complete
nucleolar
detail can be seen.
Ags distribution are uniform not obscuring intercellular matrix.
Slide47Staining Patterns:
Diffuse ? Homogeneous: antibodies to histone
Rim: antibodies to nuclear envelope proteins and to dsDNA.
Speckled: antibodies to SM,RNP, Ro/SS-A, LA/SS-B, and other antigens.
Nucleolar: associated with diffuse scleroderma
Contromeric : highly specific to the CREST Syndrome.
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