Approach Considerations
The overwhelming factor in diagnosing amyloid A (AA) amyloidosis is considering the possibility that it is present. The development of proteinuria in any individual with chronic inflammatory disease or another associated condition should prompt a search for tissue AA deposition, most commonly in the kidney.
The diagnosis of all forms of amyloidosis is confirmed by Congo red staining in a biopsy specimen. AA amyloidosis is then identified through immunohistochemical analysis and genetic testing. With respect to site selection, rectal biopsy is more useful than subcutaneous fat aspiration in AA amyloidosis. Biopsy of a clinically affected organ is the most sensitive method and may also detect concomitant pathologies, but is invasive and carries the risk of complications. In the past, liver biopsy was a common procedure in the investigation of AA amyloidosis. Several reports of fatal liver rupture or bleeding, as well as the availability of sampling procedures with little or no morbidity and mortality, have resulted in its decreased use.
Laboratory Studies
No specific tests for AA amyloidosis exist. While the serum amyloid A (SAA) precursor is usually elevated, prolonged elevation does not necessarily indicate tissue deposition because many patients with inflammatory disease have very high levels of SAA without developing amyloidosis. Serum immunoglobulins should be evaluated because the presence of a monoclonal serum or urine protein suggests AL amyloidosis as a more likely diagnosis. Patients with AA amyloidosis tend to show polyclonal hypergammaglobulinemia, reflecting their underlying inflammatory condition.
Evaluate the parameters of kidney function to monitor the course of the nephrotic syndrome or kidney failure. Occasionally, patients show renal tubular acidosis as an early manifestation of renal involvement. Deterioration of a patient with the nephrotic syndrome may indicate progression of the amyloid renal disease, but consider the possibility of renal vein thrombosis because this complication can be observed in nephrotic syndrome due to any cause.
A serum creatinine level greater than 2 mg/dL and/or a serum albumin level less than 2.5 g/dL have been associated with diminished survival rates, including renal survival.
Biopsy
For the detection of amyloid, biopsy of a clinically affected organ is the most sensitive method and may also detect concomitant pathologies. However, such a biopsy is invasive and carries the risk of complications, in particular bleeding. Thus, if amyloidosis is clinically suspected, a less invasive procedure may be desirable.
In the early 1970s, Westermark and Stenkvist demonstrated that amyloid can be detected in subcutaneous fat. [43] During the subsequent decades, subcutaneous fat pad biopsy, obtained via fine-needle aspiration, was introduced as a screening test for the detection of amyloidosis, as it is safe, inexpensive, and rapid. [44] Nevertheless, the tissue with the highest yield, particularly in the patients with proteinuria or kidney failure, is the kidney. Technically adequate samples have a diagnostic yield close to 100%.
Rectal biopsy is more useful than subcutaneous fat aspiration in AA amyloidosis. It has been found to produce positive results (assuming that submucosa is included in the biopsy specimen) in 80-85% of patients ultimately found to have tissue amyloid at a clinically relevant site. Samples from either the subcutaneous fat aspirate or the rectal biopsy can be stained in the same manner as conventional tissue biopsies to determine the presence and nature of the amyloid precursor.
Occasional patients have positive results on subcutaneous fat aspirates in the presence of a negative result on rectal biopsy, while others may have deposits in the rectal tissue and not in the subcutaneous fat. Use of both procedures may increase the yield to 90%. Abdominal subcutaneous fat biopsy results are not very sensitive in AA amyloidosis caused by FMF and in dialysis-related amyloidosis. The results are usually negative, probably because beta2-microglobulin does not accumulate in this tissue.
Series from individual centers have shown that labial salivary gland or gastric mucosal biopsies can also be high-yield procedures. [45, 46] However, these have not been used widely for amyloidosis and their general utility remains to be definitively established.
Congo red staining
Congo red stain continues to be the criterion standard for detection of amyloid deposits. In AL and AA amyloidosis, Congo red staining of aspirated subcutaneous abdominal fat has a sensitivity of 70-90% for the diagnosis. Kidney, heart, or liver samples have also been used for Congo red staining, but biopsy of rectal mucosa, skin, or subcutaneous fat is often sufficient, except in the cases of FMF, when it is rarely, if ever, positive.
The tissue is stained with an alkaline solution of Congo red and examined it under polarized light, where positive (green) birefringence is detectable in the presence of amyloidosis of any type. The nature of the fibril precursor can be established by immunohistochemical staining with antibodies specific for the major amyloid precursors (AA, immunoglobulin L chains of κ or λ type, antitransthyretin). In AA amyloidosis, only the AA is positive. The amyloid nature of the deposit can by confirmed by staining with an antiserum specific for serum amyloid P-component (SAP).
Once histological diagnosis of amyloidosis has been established, the amyloid type should be defined based on immunohistochemical analysis and genetic testing. Immunoelectron microscopy characterizes the amyloid deposits by co-localizing the specific proteins with the fibrils and can be performed on abdominal fat samples.
Imaging Studies
Avoid intravenous pyelography in patients with suspected amyloidosis because exposure to contrast medium has been associated with more frequent acute kidney injury in individuals with substantial proteinuria.
Ultrasonography is useful in determining kidney size. However, kidneys may be large, small, or normal in size in patients with renal amyloidosis.
Single-photon emission computed tomography (SPECT) may be useful because technetium occasionally binds to soft-tissue amyloid deposits. This was originally reported as an incidental finding. However, SPECT does not yield great sensitivity, and reports concerning its specificity of CT scanning have varied considerably. If results are positive, SPECT can be used to monitor gross progression of the deposition in a given organ.
Magnetic resonance imaging may have a role in amyloidosis diagnosis in the future, but currently no formal studies have reported its use in a large series of patients.
Iodine-123–labeled serum amyloid P (SAP) component scintigraphy has been used in centers in London and France to demonstrate the total body burden of amyloid and its disappearance after successful treatment of the primary disease. This test has been most useful in AA amyloidosis because the major sites of deposition (ie, liver, kidneys, spleen, and adrenal glands) are readily accessible to the imaging agent. However, this method is not available in the United States because the SAP is of human origin and cannot tolerate stringent viral inactivation. [47]
Other Tests
In patients with AA amyloidosis who have cardiac involvement, conventional parameters of cardiac dysfunction—measured using electrocardiography, echocardiography, and cardiac catheterization with endomyocardial biopsy—provide the appropriate diagnostic information and tissue for the demonstration of AA (or other amyloid) deposition in the myocardium or coronary vessels. [36] Amyloid accumulation in the heart may be suggested by the following:
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Decreased voltage in the electrocardiogram limb leads
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Pseudoinfarction pattern in the form of Q waves in the anterior chest leads
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Echocardiogram evidence of thickening of the left ventricular wall disproportionate to the degree of current or prior hypertension
Histologic Findings
Infiltrated tissues show homogeneous eosinophilic staining with hematoxylin and eosin. The earliest deposits are usually vascular. In the kidney, early deposits may be mesangial, but later in the course, entire glomeruli may be obliterated. Distinguishing these from glomerulosclerosis and from other causes was difficult prior to Congo red staining. Congo red binding by itself may be observed in other states, particularly in collagen-rich tissues, but the green birefringence is characteristic on examination with polarized light and the amyloid nature of the deposit can be demonstrated by observing the characteristic beta pleated sheet on electron microscopy. The nature of the precursor can be established with certainty using antisera specific for various amyloid precursors. In this case, staining with anti-AA serum is positive, as described above.
