IMMUNOPATHOLOGY

DR SUVARNA NALAPAT

ANA
Screening for autoimmune diseases is often

performed with the antinuclear antibody (ANA) test in which patient serum is incubated with a tissue substrate to which any autoantibodies to nuclear antigens will bind. Then, a fluorosceinated antibody is added and the tissue is observed under fluorescence microscopy to see if staining is present. Seen here is the typical "homogenous" pattern of nuclear staining of a positive ANA.

ANA

 Here is another positive ANA in which the

substrate cells are a human cell line known as Hep2. Any positive ANA is reported with a titer. The titer is simply the dilution of patient serum at which the test is still positive. The test has to be positive at greater than 1:16 or 1:20 (depending upon the lab) to be positive at all. In general, the higher the titer, the more likely a serious autoimmune disease is present. Thus, a 1:40 result might not mean much, but a 1:1012 certainly should not be ignored.

Rim pattern characteristic of SLE

Speckled patern -MCTD

This pattern is not

specific, seen in "mixed connective tissue disease" which is a mix between SLE, scleroderma, and polymyositis, but without serious renal or pulmonary disease. The autoimmune diseases are very hard to classify, even for the experts.

Nucleolar pattern
This is the so-called

"nucleolar pattern" of staining in which the bright fluorescence is seen within the nucleoli of the Hep2 cells. This pattern is more suggestive of progressive systemic sclerosis.

nucleolar pattern -reminiscent of a view from the Hubble space telescope

LE CELL

Malar(butterfly)rash

Vasculitis -SLE

 basal layer is

undergoing vacuolization and dissolution, and there is purpura with RBC's in the upper dermis (which are the reasons for the rash).

 immunofluorescence

stain with antibody to complement or immunoglobulin is performed, see the brightly fluorescing band along the dermal epidermal junction that indicates immune complex deposits are present

 immunofluorescence

staining pattern with antibody to IgG showing evidence for immune complexes at the dermal-epidermal junction. If such a pattern is seen only in skin involved by a rash, then the diagnosis is probably DLE, but if this pattern appears even in skin uninvolved by a rash, then the diagnosis is SLE.

 vascular thrombosis in

the dermis,with antiphospholipid syndrome (APS). This syndrome results from autoantibodies directed against anionic phospholipids, such as cardiolipin. This syndrome can occur in patients with SLE, antibody interferes with coagulation assays, giving it the name "lupus anticoagulant."

 These are Crithidia

organisms, whose sole purpose for existence is to serve as a substrate for the double stranded DNA test. Here the little critters have brightly fluorescing kinetoplasts indicative of a positive test. A positive double stranded DNA test strongly suggests a diagnosis of SLE.

 The periarteriolar

fibrosis ("onion skinning") seen in the spleen in patients with SLE at autopsy is quite striking, though of no major clinical consequence. This results from vasculitis.

 One of the feared

complications of the rheumatic diseases is renal failure. This is most likely to occur with SLE. Here is a glomerulus in which the capillary loops are markedly pink and thickened such that capillary lumens are hard to see. This is lupus nephritis.

 Here is a glomerulus

with thickened pink capillary loops, the so-called "wire loops", in a patient with lupus nephritis. The surrounding renal tubules are unremarkable

 If immunofluorescence

staining is performed, here with antibody to IgG, then a granular pattern of immunofluorescence is seen, indicative of deposition of immune complexes in the basement membranes of the glomerular capillary loops.

 Here is another

granular pattern of immunofluorescenc e in the glomerulus, this time with antibody to C1q complement, which is more specific for SLE.

 The thickened

basement membrane (arrow) that results from immune complex deposition in the glomerular capillary loop is prominent in this electron micrograph. The dark immune deposits are located mainly in a subendothelial position

 The electron dense

deposits by electron microscopy at high magnification are seen here.

 This is the linear pattern

of immunofluorescence with antibody to IgG in a patient with Goodpasture's syndrome. The even linear pattern is produced because the antibody is directed against the entire glomerular basement membrane.

 Here is a patient

demonstrating the taut and shiny skin typical of sclerodactyly. The skin becomes inelastic and it is hard to move the fingers. If sclerodactyly is seen along with calcinosis, Raynaud's phenomenon, esophageal dysmotility, and telangietastias, then the best diagnosis is CREST syndrome.

 Here is a higher

magnification of the taut, shiny, inelastic skin with sclerodactyly. Note also the cheilosis at the corners of the mouth from riboflavin deficiency as a result of the malabsorbtion that can occur with scleroderma.

 At low magnification,

the collagen of the dermis is increased. Chronic inflammatory cells are sparse with systemic sclerosis, unlike SLE.

 At high magnification,

the dermis is expanded by dense collagenous fibrosis in a patient with systemic sclerosis. Immunofluorescence staining of the skin is not helpful with scleroderma.

 A serious consequence

of the "R" in the CREST syndrome (limited scleroderma) is seen here. The fingertips are blackened and additional portions of the hand purplish with early gangrenous necrosis from vasospasm with Raynaud's phenomenon.

 This trichrome stain of

the stomach demonstrates intense blue staining in the submucosa from the collagen deposition. Such fibrosis can occur anywhere in the gastrointestinal tract, but is most common in the lower esophagus, leading to the esophageal dysmotility with systemic sclerosis.

 The mononuclear

inflammatory infiltrates, interstitial fibrosis, acinar atrophy of minor salivary gland in a biopsy of lip -typical for long-standing Sjogren's syndrome, (xerostomia) .lacrimal glands (xerophthalmia). Most patients are middleaged women. The autoantibodies SS-A (Ro) and SS-B (La) have specificity for Sjogren's.ANA in 50%pts+

 Renal disease

suggests diffuse scleroderma in this patient with hyperplastic arteriolosclerosis and malignant hypertension (blood pressure 300/150 mm Hg).

 This is contact

dermatitis, a form of type IV hypersensitivity in which pre-sensitized lymphocytes led to this inflammatory reaction a couple of days after contact with the offending plant material. Antigens such as those in poison oak and poison ivy are most often responsible for this appearance.

Mast cells,eosinophils,plasma cells

A form of localized

anaphylaxis with type I hypersensitivity occurs with "hay fever" when allergens in plant pollens contact IgE bound to mast cells, causing them to release their granules containing mediators such as histamine that promote vasodilation

 The acute laryngeal

edema seen here that killed the patient was due to an anaphylactic reaction to penicillin. Such an allergy is a form of type I hypersensitivity reaction in which there is preformed IgE antibody on mast cells that quickly reacts with an antigen. The mast cells release histamine and other mediators that lead to the edema.

 Besides the icterus

(yellow color, jaundice) in this skin there is a fine scaling rash in this patient following bone marrow transplantation with a 5 out of 6 antigen match. This is an example of graft versus host disease in which donor lymphocytes attack host tissues

 Microscopically, graft

versus host disease is one of the best examples of a process called "apoptosis" or single cell necrosis. There is vacuolization and dissolution of epidermal cells along the basal layer, along with lymphocytes. At the arrow is a rounded pink apoptotic body

 Graft versus host

disease also leads to marked cholestasis in the liver, seen here as large collections of yellow-green bile pigment in the bile canaliculi.

 The graft versus

host disease here is marked by yellowbrown collections of bile in the canaliculi, as well as chronic inflammatory cells within the liver parenchyma

 Immunologic disease

can also complicate solid organ transplantation. Here is a renal biopsy that demonstrates marked interstitial fibrosis in a patient with chronic vascular rejection.

 At high magnification,

the renal arteries with chronic vascular rejection are markedly thickened and fibrotic. There is interstitial fibrosis and chronic inflammation. Such chronic rejection usually occurs slowly over several months to years following transplantation. This disease, unlike acute rejection, is difficult to treat

 This is a form of acute

renal transplant rejection known as acute cellular tubulointerstitial rejection because most of the inflammation is in the interstitium. The glomerulus seen here is normal, but the tubules are infiltrated by many lymphocytes at the upper right.

 -H.Pthe lymphocytes

and plasma cells seen around renal tubule in a renal transplant patient with acute cellular rejection. This type of rejection can occur at any time following transplantation when immunosuppression is diminished. This is treated by administering cyclosporine and other immunosuppressive agents.

 The

immunofluorescence pattern with acute tubulointerstitial renal transplant rejection is shown here, in which immune deposits occur between glomeruli in the interstitium. Both type II and type IV immune hypersensitivity reactions contribute to this rejection reaction.

Acute vascular rejection in a heart transplant

The inflammatory reaction consists of lymphocytes , mainly around small arteries, a vasculitis. This can occur when the dose of immunosuppressive drugs is decreased in the months following transplantation. Increasing immunosuppresive therapy not effective as for acute cellular

 By

immunofluorescenc e, antibody to IgG is seen highlighting the vascular walls in this heart with acute vascular rejection.

 By immunoperoxidase

staining with antibody to CD-3, the Tlymphocytes in the myocardium involved in this acute cellular rejection phenomenon in a heart transplant recipient can be identified.

 This is a peculiar

disease known as amyloidosis. Amyloidosis is characterized by slow deposition over years of increasing amounts of an amorphous proteinaceous material in one or more tissues. Seen here in the heart between the darker red myofibers are pale pink amyloid deposits

 When stained with

Congo red and observed under polarized light, the amyloid has a characteristic "apple green" birefringence as seen here in a deposit around an artery in the heart.

 By electron microscopy,

amyloid is composed of a "beta-pleated sheet" of fibrils, seen here as irregular grey material. When the amyloid protein is made up of immunoglobulin light chains, then it is "AL amyloid" and when it is derived from serum amyloid-associated protein, then it is "AA amyloid." In terms of the effect upon the organs, "amyloid is amyloid".

 This is the

immunofluorescent appearance of the myocardium with antibody to lambda light chain. Thus, this is "AL amyloid".