Anti-core or ANA and anti-nucleus-extractable antibodies, ENA

  1. Gastroepato

doctor's notes Claudio Italiano

Autoimmune diseases


Clinical Pathology, in recent years, has made considerable progress in the field of examinations that can be used for the diagnosis of rheumatic diseases. There are a whole series of laboratory investigations, in fact, very useful to approach the patient with problems of rheumatic diseases, from the Greek rheumatism, that is a pain that "migrates" or suffering from connective tissue diseases, such as Lupus, scleroderma. (see index of autoimmune diseases). Obviously, given the economic situations of our country, which is at the forefront of the world for assistance to citizens, given the precarious resources available today in health and the policy of non-waste, the wise doctor will not immediately prescribe expensive investigations, being in front of the rheumatic patient, but will start by prescribing the first level investigations, including the trivial inflammatory indexes, trying to aim the diagnosis at the organs involved in the pathology:

General or basic examinations

-emocromo -VES, CRP
-protidogramma
-ALT, alkaline phosphatase, γGT
-glycemia, cholesterolemia
-creatininemia, uricemia, urine test
-CK, LDH
-search for occult blood in the stool
In rheumatic diseases, in fact, already the single haemochromocytometric can direct the doctor to:
an inflammatory anemia,
a hypochromic anemia - hemolytic microcytic,
a leukopeniacon lymphocytopenia (LES),
a leukocytosis,
a thrombocytopenia (especially on an autoimmune basis)
a platelet.Eritema malare in pz con lupus

SPECIFIC EXAMINATIONS IN REUMATOLOGY

 -anti-citrulline antibodies (CCP) and rheumatoid factor
-automatic anti-nucleus antibodies (ANA)
-automatic anti-DNA antibodies
- ANCA autoantibodies
-aut anti-ENA antibodies
anti-phospholipid antibodies (aPL)
- complement components: C3 and C4
- Synovial fluid examination
- crioglobuline

The indices of inflammation, VES and PCR and the protidogramma, renal group and hepatic group:

 ESR (erythrocyte sedimentation rate) increases during disease activity, as well as C reactive protein (CRP) in diseases such as SLE or other rheumatic diseases.
The protidogramma may show an increase in a2-globulins, a hyperglobmagulinemia (in agreement with the abnormal production of autoantibodies), an hyperalbuminemia.
General blood tests and urinalysis can show an elevation of certain values (transaminases, bilirubin, azotemia, creatinine, etc.), expression of the pathology of the affected organ, liver or kidney).

SPECIFIC EXAMINATIONS IN REUMATOLOGY

Anti-CCP, Rheumatoid Factor and Rheumatoid Arthritis
Early detection of rheumatoid arthritis has important implications for therapeutic purposes; however the old research of the Rheumatoid Factor was today supported by other supporting investigations:
research of anti-peptide citrullinated antibodies (anti-citrulline or CCP), with high specificity, high diagnostic and prognostic predictive value in patients with rheumatoid arthritis.

FURTHER ASSESSMENTS OF THE CASE, THE ANAN ANTI-CARRIERS, ENA


We premise that the search for these anti-Core antibodies is the first step, that is the first level investigation, to get to the diagnosis of Autoimmune Rheumatic Disease, that is determined by a movement of antibodies which like "crazy missiles" turn against their own structures or Self of the human body itself. This is done with a special technique, that is with indirect immunofluorescence (IFI). In the presence, however, of a positive ANA-IFI result, we can not stop here, as we need to proceed with further investigation of the case and search for antibodies directed against other structures of the cell, that is against other antigens of the nucleus. In general, therefore, antibodies are sought for the so-called extractable nuclear antigens (ENA).
Their research is useful for the diagnosis of:
LES,
of neonatal lupus,
of Sjögren's syndrome (SS),
of mixed connective tissue disease (MCTD),
of polymyositis / dermatomyositis (PM / DM)
of systemic sclerosis (SSc),

Search for autoantibodies
-Anti-Sm: highly specific for LES; the clinical sensitivity varies from 15 to 30%. High titles have greater diagnostic significance.
-Anti-U1RNP: not specific. Present in 30-40% of patients with SLE; high-quality are an essential diagnostic criterion for the diagnosis of MCTD.
-Anti-SSA / Ro: not specific. In patients with SS there are up to 60% if tested with immunodiffusion and up to 90% if tested with ELISA; they are more frequently associated with extraglandular involvement. They can also be present in about 30% of patients with SLE. High ELISA titers, especially for anti-Ro 52, are a risk factor for neonatal lupus.
-Anti-SSV / La: not spefic. Both in patients with SS and those with SLE, they are present less frequently than SSA / Ro antibodies. They are associated with a higher incidence of cutaneous forms, vasculitis, leukopenia and lymphopenia, and neonatal lupus.
-Anti-centromere: associated with limited SSc. 32% sensitivity in all patients with SSc; 57% in limited SSc; high specificity.
-Anti-topoisomerase I (Scl70): associated with diffuse SSc. Clinical sensitivity variable, depending on the method, from 25 to 70%; analytical sensitivity 94-99%. High clinical specificity; analytical specificity ranging from 70 to 99%, depending on the method.
-Anti-Jo-1: specific for PM / DM. Sensitivity from 8 to 40%. Associated with pulmonary interstitiopathy and arthritis
Anti-ENA autoantibody specificities
-Anti-PM-Slc: is associated with SSc with myopathy.
-Anti-U3-RNP (fibrillarin): is present in 4-8% of patients with SSc.
-Anti-RNA polymerase I: is more frequent in progressive forms of SSc.
-Anti-polymerase III: common in SSc.
-Anti-ThTo: is associated with SSc with limited skin involvement.
-Anti-protein ribosomal P: is associated with LES and, in particular with greater frequency, to forms with psychotic manifestations.
-Anti-PCNA: it is associated with LES in active phase

The most specific use of laboratory tests in the definition of a rheumatic disease is based on the search for autoantibodies, as an expression of the autoimmune organism, which is the basis of many rheumatic diseases. In particular, autoantibodies are expressed against the autoantigens. , which are normally found constituents:
in the nucleus
in the cytoplasm of all cells,
in the cytoplasm of neutrophil granulocytes.

Correspondingly there are anti-nucleic autoantibodies (ANA), anti-DNA, extractable anti-nuclear antigens (ENA), anti-cytoplasm of neutrophil granulocytes (ANCA) and anti-phospholipids (aPL).
ANAs are directed towards nucleocytoplasmic antigens present in all human cells, unlike autoantibodies of organ-specific autoimmune diseases, such as pernicious anemia, autoimmune thyroiditis, primary biliary cirrhosis, autoimmune hepatitis etc. which autoantibodies to specific antigens of the single organ involved are found. The positivity of ANA research is considered one of the main characteristics of systemic autoimmune diseases, so much so that some of them fall within the diagnostic and classification criteria of the same diseases.
The research of ANA finds its clinical significance in the phase of:
a) diagnostic screening, because it has a function of diagnostic element or criterion;
b) deepening for the definition of subgroup or subset of systemic autoimmune disease (MAIS), eg the presence of autoantibodies towards CENP-B or Scl70 leads towards a clinical classification of localized or diffuse systemic sclerosis; SES-associated psychosis has a specific marker in anti p-ribosomal antibody;
c) monitoring, to define a follow-up of the course and / or of the disease, with significant variations in the degree of positivity.
ANA research must be motivated by a clinical suspicion or a risk factor for MAIS (autoimmune diseases):

MAIN SELF-ADVANTAGES IN SYSTEMATIC AUTOIMMUNE DISEASES

DsDNA Nucleic Acids
Proteins associated with DNA Histones
Ku
PCNA
Centomeric proteins
Topoisomerase I (Scl70)
Proteins associated with RNA Spliceosomal proteins
Nucleic proteins
Core-cytoplasmic prot
The research of ANA has meaning because, if carried out with the appropriate analytical technique, such as indirect immunofluorescence, it allows, on the basis of the title and the fluoroscopic picture identified, the direction towards a specific rheumatic pathology.
Fluoroscopic images concern HEp-2 human epithelial cells, used for ANA research: fluoroscopic appearance is defined based on the identified bright green immunofluorescence.
Positive ANA positives can be represented by:
- a nucleolar positivity,
- a speckled positivity, that is, dotted,
- homogeneous positivity,
- a positivity to diffuse granulitis.
The different positivity allows the identification of the involved autoantigen and the orientation of the clinician towards a defined clinical form, as schematized in the following tables.
ANA-IFA positivity
Homogeneous · nDNA Anticentromere · CENP-B
Speckled · Sm · RNP · SSA · SSB
Nucleolar · PMScl · RNA polymerase · Fibrillarin · Ku - To / Th
Diffuse grainy
Topoisomerases I PCNA · Cyclin


in particular we will have in the various pathologies:

Homogeneous ANA → LES, Rheumatoid Arthritis
ANA diffused granulosis → Systemic sclerosis
ANA speckled → Mixed Connectivity (Sharp's)
ANA end speckled → Sjögren's syndrome
ANA PCNA (proliferating cell nuclear antigen) → LES
ANA anticentromere → Systemic sclerosis (CREST or localized variant)
Nucleus ANA → Systemic Sclerosis

The predictability, ie the ability to identify a specific pathology, of ANAs is not absolute, depending on the influence of factors such as:
- clinical context (age, sex, diseases, drugs), so the positivity in a 80-year-old woman has a different meaning than the positivity in a 2-year-old girl or in a male compared to a female, finally there is the possibility of iatrogenic forms for which account should also be taken of the use of certain drugs in the clinical history sheet;
- analytical method used: immunofluorescence is a difficult and not very accessible method, but more predictive than others;
- fluoroscopic pattern: some are pathognomonic, markers of the disease;
- title of positivity: obviously a 1: 2480 positivity is much more significant than one
positivity 1:80; an ANA positivity (low titre) can easily be found in the following pathological and paraphysiological conditions:
neoplasms
leukemia
acute and chronic renal failure
kidney disease
viral infections (EBV, HIV)
healthy subjects (pregnancy, females> 40 years, elderly)
- persistence over time: certain viral infections (Cytomegalovirus, Hepstein-Barr virus), mixed polyclonal drugs or activations may result in a transient increase in autoantibodies.
The research of the ENAs should be understood as a II level survey for the identification of autoantibody specificities after the finding of ANA positivity, as the search for native DNA antibodies finds meaning in the patients in whom ANA-type positivity has already been found homogeneous and speckled with high titre, compatible with the presence of antibodies to DNA. The quantification of these antibodies is useful in the clinical and therapeutic monitoring of patients with SLE because there is a close relationship between title or better international units and activity of the disease itself.
In conclusion, the most useful laboratory tests for the diagnosis, prognosis and monitoring of rheumatic diseases are:
¨ diagnosis: identification of disease markers (CCP, FR, ANA, DNA, ENA)
¨ prognosis: quantitative DNA, clinical subset markers, bio-temporal examinations
¨ monitoring: quantitative DNA, CRP, bio-temporal examinations for organ pathology.

index topics on autoimmune disease