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Fine Needle Aspiration Biopsy Cytology of Liver: A Diagnostic Approach Based on Pattern Recognition
Acknowledgement
John M. Ferguson, MD, contributed the section on “Fine Needle Biopsy of Liver Lesions—A Radiologist’s Perspective.”
With the routine use of imaging to investigate patients with abdominal symptoms or to screen patients with cancer, increasing numbers of localized liver lesions are being detected. Fine needle aspiration biopsy (FNAB) of the liver is commonly performed to investigate these localized mass lesions or “tumors.” The aim of FNAB is to diagnose tumors either benign or malignant and where possible identify the specific tumor type. A cytologic diagnosis can be made quickly, accurately, and safely in a cost-effective manner. When necessary, a concurrent core biopsy can be performed. Most studies have shown that FNAB and core biopsy are complementary with respect to sensitivity and specificity.
Many lesions show a classical appearance enabling a cytodiagnosis on smears alone, but cytochemistry, immunocytochemistry (IHC), flow cytometry, molecular studies, and electron microscopy can be used to advance the diagnosis.
The FNAB is usually performed by a radiologist using ultrasound (U/S) or computed tomography (CT) scan guidance. But more recently, endoscopic ultrasound has been used effectively to target some liver lesions, in particular those involving the hilum and left lobe.
Most procedures cause the patient little discomfort, and serious complications are rare.
Patients often present clinically asymptomatic when a mass is identified incidentally when imaging is performed for an unrelated reason or when staging a previously diagnosed primary malignancy. Or they may be symptomatic due to abdominal pain and tenderness in the right upper quadrant (RUQ), hepatomegaly with a palpable mass or fullness, obstructive jaundice, or tumor rupture with peritonitis. They may show signs and symptoms of liver dysfunction in the setting of chronic liver disease or secondary to the mass effect. Systemic effects associated with malignancy, such as malaise, weakness, loss of appetite, weight loss, nausea, and vomiting, may be present.
Investigations that may provide useful clinical information include liver function tests (LFTs), viral hepatitis serology, autoimmune markers, metabolic screens, serum tumor markers such as alpha fetoprotein, and imaging.
Liver FNAB is generally considered not useful in the diagnosis of diffuse parenchymal disease, such as viral hepatitis. However, the cytologic features of various changes associated with diffuse chronic liver disease have been described.
Three common clinical scenarios are associated with liver FNAB:
- 1.
A mass lesion in a patient with documented chronic liver disease has a differential diagnosis (DD) of macroregenerative nodule, hepatocellular carcinoma, or, less likely, an unexpected other lesion.
- 2.
A solitary mass in a noncirrhotic liver often discovered incidentally on imaging for a nonrelated symptom, typically in a young or middle-aged female, with a DD of hepatocellular adenoma, focal nodular hyperplasia, or, less likely, primary or metastatic malignancy.
- 3.
Single mass or multiple masses in a patient with either no significant past history or a history of a primary malignancy elsewhere with a DD of metastatic malignancy or, less likely, primary malignancy such as hepatocellular or cholangiocarcinoma.
FNAB of Liver: Radiologist’s Perspective
Ultrasound is the preferred method of image guidance as real-time needle placement is possible without the need for radiation and selective sampling of the periphery of the lesion can be performed in case of central necrosis. But it must be possible to identify the lesion, which is not always possible on ultrasound, and CT may be required. Before FNAB it must be ascertained if the coagulation profile is normal and if the patient takes any drugs, which may increase bleeding risk.
The decision to opt for FNAB versus Tru-Cut biopsy in liver lesions is based on a number of factors and will vary from one institution to another but includes:
- 1.
Does the cytopathologist provide rapid on-site evaluation (ROSE) at the procedure and how experienced is the cytopathologist?
With ROSE providing immediate feedback, a single FNAB pass may provide enough material for diagnosis, thereby limiting needle passes. Conversely, there can be a progression to Tru-Cut biopsy if FNAB material is insufficient and the lesion is technically amenable. Valuable feedback at both the time of the FNAB and at multidisciplinary meetings leads to a better understanding of the benefits and limitations of FNAB for particular liver lesions.
- 2.
What is the preprocedure DD based on clinical history, laboratory tests, and imaging, and will the architecture be important for pathologic diagnosis?
Modern imaging can go a long way to establishing the diagnosis of a liver lesion or at least determining the benign versus malignant DD, especially with the more widespread use of liver magnetic resonance imaging (MRI) and positron emission tomography (PET) CT. Simple liver cysts, hemangiomas, and focal nodular hyperplasia do not require FNAB if they have typical imaging features. If ROSE demonstrates a primary hepatocellular lesion where the architecture of the lesion is required, such as the DD of hepatocellular adenoma versus well-differentiated hepatocellular carcinoma, core biopsy is recommended, generally using an 18-gauge (ga) needle. However, if the initial imaging suggests a nonresectable lesion or there is a history of prior primary malignancy elsewhere, then FNAB is preferred and is well tolerated by patients with low morbidity.
- 3.
Is a direct approach possible or will the biopsy needle traverse other structures?
Because of the FNAB small-size needle, generally 22 Gauge in the liver, access routes can traverse other structures, such as the inferior vena cava or small bowel, where necessary, and have a low complication rate.
- 4.
Can the patient cooperate with breath holding?
Deep inspiration may be required to make a lesion high up in the liver visible and accessible to the needle. To get a good accurate FNAB sample, it is important to watch the needle rapidly traverse the lesion repeatedly, and this may be difficult in a high superficial lesion if the patient cannot hold a breath for an adequate time.
- 5.
Is the tissue diagnosis procedure being combined with percutaneous therapy, such as radiofrequency ablation, at the same time?
Ablative therapies such as radiofrequency ablation are now frequently performed, either with curative intent or as a bridge to transplant in patients with hepatocellular carcinoma. Thus a lesion that has imaging characteristics of hepatocellular carcinoma on MRI in a cirrhotic patient may undergo FNAB biopsy with ROSE followed by core biopsy and ablation at the same time using a coaxial technique. The coaxial needle is large enough to accommodate either a FNAB or a core biopsy needle up to 16 Gauge, as well as the ablation probe, and may reduce track seeding rates. An optimum result for the patient comes from close collaboration in a multidisciplinary environment.
Approach to FNAB Diagnosis
Imaging guidance should ensure accurate needle placement, and optimally a cytopathologist or experienced cytotechnologist should attend the procedure for ROSE to confirm diagnostic material and to triage the material for cell block, histochemistry, IHC, flow cytometry, microbiology cultures, molecular studies, and electron microscopy. In some situations a core biopsy should be requested, particularly if architectural information is crucial to the diagnosis, for example, in distinguishing hepatic adenoma from well-differentiated hepatocellular carcinoma. If the material is to be smeared by the radiology staff, they should be trained, appropriate consumables should be supplied, and needle rinses for cell block or cytospin preparations must be collected.
Accurate interpretation requires past history, current clinical history, knowledge of the radiologic appearance, the site of the sample, and awareness of what is “normal” for that site.
Advantages of FNAB include that it is less invasive with fewer complications; it can be used to access lesions close to blood vessels, dilated bile ducts, or the gallbladder; multiple passes can be performed over a wide area and multiple sites sampled; ROSE can assess adequacy with cost-effective selection of appropriate ancillary tests; and it is a day procedure not requiring hospitalization.
Core biopsies are complementary as they yield more architectural information and do not require specific cytopathologist skills.
Contraindications for FNAB include bleeding disorders, difficult access or an uncooperative patient, and suspected hydatid disease, although steroid and albendazole cover can be considered.
FNAB complications include hemorrhage, biliary peritonitis, potential tumor seeding resulting in a worse morbidity and mortality, and potential anaphylaxis in hydatid disease.
General pitfalls include (1) the clinical history or radiologic appearance of a mass may be misleading, (2) cells from structures traversed on the way to the lesion such as bowel or mesothelium may predominate, and (3) the material may not be representative of the lesion.
Pattern Recognition: Specific Lesions and Their Cytopathological Criteria
Liver FNAB diagnosis relies on more than just assessment of the cellular morphology. Many diagnostic entities show a characteristic low-power pattern, which depends on the degree of cellular cohesion, architecture of tissue fragments, and smear background. Low-power assessment of patterns provides important diagnostic clues, which are supplemented by subsequent high-power examination of the cell morphology. One should get into the habit of making an initial low-power examination using a checklist approach to assess cellularity, cohesion, architecture, and background before moving on to higher power.
Moderate to marked cellularity generally implies a neoplastic process but may represent either a benign or malignant entity. Further diagnostic clues can be gleaned by assessing the arrangement of cells. Large tissue fragments, defined as large sheets with more than 100 cells, may have ragged edges suggesting they have been “torn” from the tissue and represent a “mini biopsy,” or they may have rounded edges implying a specific architectural pattern, such as trabecular or glandular. Small tissue fragments, defined as having 10 to 100 cells, may also be ragged or rounded and may show architectural features such as gland formation. Minute tissue fragments with fewer than 10 cells and dispersed cells imply poor cohesion and are a general feature of malignancy, with total dispersal associated typically with lymphoma or melanoma. Although cellularity may be a function of the FNAB technique, it can also be a characteristic of the lesion. Discussion with the attending radiologist regarding the ease of needle placement, texture of the lesion on FNAB, and length of time within the lesion may provide useful information to aid assessment.
Five distinctive smear patterns exist.
Pattern 1: Background Predominant ( Plates 6-1A and B ; Figs. 6-1 to 6-4 )
This is seen in many different types of lesions, both non-neoplastic and neoplastic, and may represent either the primary lesion such as an abscess or a nonspecific secondary change within or adjacent to a lesion, as in necrosis in a malignant tumor. The background pattern can be subclassified into inflammatory, bloody, cystic, mucinous, necrotic, and stromal (see Plate Table 6-1 ). Careful examination looking for diagnostic cells that may be scanty and obscured by the background is important. Correlation with the clinical history and imaging findings is crucial to reach a diagnosis in many cases. The background material may be perilesional and not representative of the lesion. Caution is advised in making a definitive diagnosis if there is any discrepancy when correlating with the clinical and radiologic findings, and ancillary tests may be useful.
Table 6-1
Smear Patterns in Liver FNAB
Pattern 6-1 : Background predominant ( Plate 6-1A )
Pattern 6-2 : Mildly cellular with small tissue fragments ( Plate 6-2A )
Pattern 6-3 : Moderately to markedly cellular with tissue fragments ranging from small to large without dispersed cells ( Plate 6-3A )
Pattern 6-4 : Moderately to markedly cellular with tissue fragments ranging from small to large with dispersed cells ( Pattern 6-4A )
Pattern 6-5 : Moderately to markedly cellular with dispersed cells ( Plate 6-5A ) |
Plate 6-1
Pattern 1 Background Predominant
Plate 6-1 Table
Pattern 1: Background Predominant
Entities in the Differential Diagnosis
|
Plate 6-2
Pattern 2 Mildly Cellular With Small Tissue Fragments
Plate 6-3
Pattern 3 Moderately to Markedly Cellular With Tissue Fragments Ranging From Small to Large Without Dispersal
Pattern 6-3 Table
Pattern 3: Moderately to Markedly Cellular With Small Ranging to Large Tissue Fragments Without Dispersal
Entities in the Differential Diagnosis
|
Plate 6-4
Pattern 4 Moderately to Markedly Cellular With Tissue Fragments Ranging From Small to Large and Dispersed Cells
Pattern 6-4 Table
Pattern 4: Moderately to Markedly Cellular With Tissue Fragments Ranging From Small to Large and Dispersed Cells
Entities in the Differential Diagnosis
|
Plate 6-5
Pattern 5 Moderately to Markedly Cellular With Dispersed Cells
Pattern 2: Mildly Cellular With Small Tissue Fragments ( Plates 6-2A and B ; Figs. 6-5 and 6-6 )
The presence of small tissue fragments implies either a cohesive, well-differentiated neoplasm such as hepatocellular adenoma or may signify a desmoplastic neoplasm of low cellularity such as cholangiocarcinoma.
Pattern 3: Moderately to Markedly Cellular With Tissue Fragments Ranging From Small to Large Without Dispersal ( Plates 6-3A and B ; Figs. 6-7 to 6-10 )
This pattern implies cohesion and is found in low-grade neoplasms, either benign or well-differentiated malignancies. If hepatocellular, the DD includes adenoma, focal nodular hyperplasia, and well-differentiated hepatocellular carcinoma; if not, the DD is well-differentiated adenocarcinoma, either cholangiocarcinoma or metastatic.
Pattern 4: Moderately to Markedly Cellular With Tissue Fragments Ranging From Small to Large and Dispersed Cells ( Plates 6-4A and B ; Figs. 6-11 to 6-14 )
This pattern usually implies a lesser degree of differentiation and malignancy. The presence or absence of vessels and their relationship to the cells, as well as the architecture of tissue fragments, are useful criteria. Hepatocellular carcinoma typically shows a trabecular architecture with smooth edges rimmed by endothelial cells, or it may have vessels running through or intersecting the fragments. Vessels are also seen in entities such as angiomyolipoma and metastatic renal cell carcinoma, where they usually only intersect ragged fragments. Metastatic adenocarcinoma from any site usually does not show a significant component of vessels.
Pattern 5: Moderately to Markedly Cellular With Dispersed Cells ( Plates 6-5A and B ; Figs. 6-15 and 6-16 )
This pattern can be seen in any poorly differentiated malignancy but usually implies a specific DD including lymphoma and melanoma. Carcinoid tumor is dispersed but usually shows some cohesion with small tissue fragments. In angiosarcoma the cells are typically spindled and the background bloody. Non-neoplastic inflammatory lesions can also present as a dispersed population but are generally diagnosed on cell morphology.
The lesions associated with each pattern are listed in Table 6-1 .
Normal Liver
Smear Pattern: Mildly Cellular With Small Tissue Fragments
Clinical Features
Normal liver may be sampled if needle placement is inaccurate and the lesion is missed. Problems arise when the imaging DD includes a benign hepatocellular lesion such as a regenerative nodule or focal nodular hyperplasia or hepatic adenoma. Confident distinction between these can be impossible, and close correlation with the clinical and radiologic findings is required.
Cytopathology
Cytologic diagnostic criteria ( Figs. 6-17 to 6-19 )
-
Cellularity: mild to moderate.
-
Pattern: mildly cellular with small tissue fragments.
-
Cohesive small sheets and trabecular or sinusoidal tissue fragments in a bloody background with scant dispersed cells and no stripped nuclei.
-
Polygonal or round hepatocytes with distinct cytoplasmic borders and low N:C ratio.
-
Single or occasionally binucleated, central, round nuclei with finely granular chromatin; single, small, prominent nucleolus; occasional intranuclear pseudoinclusions; and abundant, dense, and granular cytoplasm with some pigment (bile, lipofuscin) or lipid in some cells.
-
Mitoses rare.
-
Bile duct epithelium in tubular tissue fragments or sheets, evenly spaced nuclei with bland chromatin, smaller than hepatocytes with a higher N:C ratio.
-
Kupffer cells (sinusoidal macrophages) have elongated nuclei.
-
Sinusoidal endothelial cells.
-
Vessels and stroma.
-
Mesothelial sheets with a “honeycomb” pattern as contaminant.
Differential Diagnosis
There may be some variation in hepatocyte nuclear size and chromatin in disease states or representing secondary changes adjacent to a mass lesion.
Ancillary Diagnostic Tests
IHC on cell blocks is rarely required.
Hepatitis
Smear Pattern: Mildly Cellular With Small Tissue Fragments
Clinical Features
Broadly defined “hepatitis” indicates inflammation of the liver and is associated with a wide spectrum of primary and secondary liver diseases that are often difficult to distinguish without correlation with the clinical information. FNAB is not used in the diagnosis of diffuse inflammatory liver disease because the cytologic features are usually nonspecific, but inflammatory features do occur adjacent to a mass lesion and must be recognized, particularly when the DD is regenerative nodule versus hepatocellular carcinoma in a background of chronic liver disease.
Cytopathology
Cytologic diagnostic criteria ( Fig. 6-20 )
-
Cellularity: mild to sometimes moderate with inflammatory cells.
-
Pattern: mildly cellular with small tissue fragments.
-
Small sheets and tissue fragments of hepatocytes with lymphocytes and other inflammatory cells in the background, which may include debris.
-
Hepatocytes show ballooning degeneration and/or apoptosis in which hepatocytes are smaller and exhibit deeply stained cytoplasm and degenerate pyknotic nuclei.
-
Regenerating hepatocytes vary greatly in size and show uneven cytoplasmic staining, variable chromatin pattern, and large nucleoli.
-
Hepatocytes may show multinucleation, pleomorphism, and mitoses.
-
Prominent Kupffer cells with lipofuscin and iron.
-
Bile pigment in cholestasis.
In chronic liver disease, smears may also contain variable numbers of fibrotic tissue fragments and increased bile duct epithelium as seen in cirrhosis.
Ancillary Diagnostic Tests
IHC on cell blocks can confirm hepatitis B infection.
Cirrhosis
Smear Pattern: Mildly Cellular With Small Tissue Fragments
Clinical Features
Chronic liver disease of any etiology can progress to cirrhosis as the end stage. The underlying pathophysiology is inflammation, liver cell death, repair by fibrosis, and regeneration of surviving hepatocytes. The result is diffuse distortion of the liver architecture by fibrous septa surrounding nodules of regenerating hepatocytes. Bile ductules increase in number in response to disruption of the bile flow. Inflammation continues as long as there is hepatocyte damage. Liver cell dysplasia and hepatocellular carcinoma may complicate cirrhosis.
Cytopathology
If the FNAB enters a large regenerating nodule, the smears may show regenerating hepatocytes without evidence of cell necrosis, inflammation, fibrosis, or increased bile ductules, but when the FNAB samples a fibrous septum the smears may be poorly cellular and show mainly stromal tissue and inflammation.
Cytologic diagnostic criteria ( Fig. 6-21 )
-
-
Pattern: mildly cellular with small tissue fragments.
-
Irregular sheets of hepatocytes, some showing degenerate or reactive atypia.
-
Regenerating liver cells with occasional binucleated or multinucleated cells and mitoses.
-
Fibrotic tissue.
-
Increased bile duct epithelium.
-
Prominent Kupffer cells.
-
Variable inflammation, depending on the cause of the cirrhosis and stage of disease.
-
Cholestasis.
Liver Cell Dysplasia
Smear Pattern: Mildly Cellular With Small Tissue Fragments
Clinical Features
Liver cell dysplasia was first defined in 1973 as cellular enlargement, nuclear pleomorphism, and multinucleation of liver cells, but more recently, it has been divided into large cell dysplasia with enlarged hepatocytes with irregular nuclei, prominent nucleoli, and a normal N:C ratio and small cell dysplasia defined by small cells showing no atypia in a zone of “crowding” of nuclei with an increased N:C ratio.
The importance of liver cell dysplasia in cytology lies in its distinction from hepatocellular carcinoma because the criteria for dysplasia are also criteria for malignancy and dysplastic hepatocytes may be indistinguishable from malignant hepatocytes.
Cytopathology
FNAB smears from dysplastic nodules show the same features as cirrhosis with increased bile duct epithelium and inflammatory cells, as well as the dysplastic sheets.
Cytologic diagnostic criteria
-
Cellularity is mild.
-
Pattern: mildly cellular with small tissue fragments.
-
Trabecular architecture without a prominent vasculature.
-
Large cell dysplastic hepatocytes are mixed with “normal” hepatocytes.
-
Dysplastic hepatocytes are enlarged and polygonal with large, atypical nuclei but a normal N:C ratio, prominent nuclear membranes, abnormally clumped chromatin, and enlarged multiple nucleoli.
-
Occasional multinucleated forms.
Small cell dysplasia is difficult to identify with confidence as it requires sheets of hepatocytes showing increased N:C ratio contrasting with adjacent, more “normal” hepatocytes.
Differential Diagnosis
Multinucleation can also occur in some forms of active hepatitis or in the setting of hepatocellular carcinoma.
Hepatocyte atrophy, in which hepatocytes within a collapsed reticulin framework show reduced cytoplasm with small, often pyknotic nuclei, is often present around space-occupying lesions, often associated with cholestasis.
Ancillary Diagnostic Tests
There are no IHC studies to mark hepatocyte dysplasia.
Steatosis and Steatohepatitis
Smear Pattern: Mildly Cellular With Small Tissue Fragments
Clinical Features
Fat within hepatocytes may be seen focally or diffusely, when it represents steatosis (steatohepatitis) or a nonspecific finding in association with another disease, such as hepatitis C viral (HCV) infection.
Steatosis is divided into macrovesicular steatosis, in which the hepatocyte contains one or several large fat droplets displacing the nucleus to the periphery, and microvesicular steatosis, where the hepatocyte cytoplasm is filled with many small lipid droplets that indent but do not displace the nucleus. Macrovesicular steatosis is commonly associated with excess alcohol consumption or nonalcoholic fatty liver disease, which in turn can be associated with the “metabolic syndrome” including diabetes mellitus, obesity, or hyperlipidemia, or with drugs and HCV infection. Microvesicular steatosis is classically associated with Reye syndrome and fatty liver of pregnancy but can be seen in alcohol abuse or toxicity due to drugs such as tetracycline and valproic acid.
Cytopathology
Cytologic diagnostic criteria ( Fig. 6-22 )
-
-
Pattern: mildly cellular with small tissue fragments.
-
Small tissue fragments of hepatocytes showing some loss of cell cohesion.
-
Hepatocytes rounded and enlarged, containing variable-sized lipid vacuoles, eccentric nuclei, and nonspecific nuclear glycogenation.
-
Ballooning feathery degeneration, Mallory bodies, and neutrophils in steatohepatitis.
Mallory bodies are variously described as ropey eosinophilic strands or round or elongated eosinophilic perinuclear clumps or large, irregular, bluish-gray granules or bodies. If there is progression to fibrosis and cirrhosis, stromal elements and prominent bile ductules can be found.
Differential Diagnosis
Fatty change is rarely found in hepatocellular carcinomas.
Ancillary Diagnostic Tests
Positive staining with oil red-O and Sudan III can be seen on air-dried smears and a diastase periodic acid–Schiff (PAS) stain is negative. The stain for reticulin will show mild distortion of the reticulin pattern within tissue fragments in the cell block.
Cholestasis
Smear Pattern: Varies With the Etiology of the Cholestasis
Clinical Features
Cholestasis has many causes and is due to disruption of bile excretion at any point along the pathway from production within the hepatocyte, via the canaliculi and biliary tree, to excretion into the duodenum through the ampulla.
Morphologically, cholestasis manifests as visible aggregation of bile pigment within hepatocytes, bile canaliculi, and Kupffer cells. It is readily seen in cytologic preparations.
Cytopathology
Cytologic diagnostic criteria ( Fig. 6-23 )
-
-
Cellularity varies with cause.
-
Hepatocyte cytoplasmic feathery degeneration.
-
Smooth, intracytoplasmic bile droplets of varying size, yellow green, and glassy in the Papanicolaou (Pap) and khaki in the Giemsa stains.
-
Elongated bile plugs in canaliculi between liver cells, in Kupffer cells and free in the background.
-
Increased bile duct epithelium may be present.
Distinction between bile and hemosiderin in hepatocytes is usually not a problem as hemosiderin is yellow-brown and granular in the Pap and bluish black in the Giemsa. If in doubt, a histochemical stain for iron on tissue fragments and cells in the cell block is helpful.
Liver Granulomas
Smear Pattern: Mildly Cellular Arranged in Small Tissue Fragments
Clinical Features
Granulomas occur as an incidental finding or as the clinically suspected lesion and represent a primary hepatic disease or manifestation of a systemic disease. The etiology most likely to present as a mass, particularly in the developing world, is infection such as mycobacteria, which may mimic a pyogenic abscess or malignancy on imaging.
In hepatic tuberculosis the granulomas show central caseation. Direct immunofluorescence of the Pap-stained smear or a Ziehl-Neelsen stain on smears or cell block material will identify the acid-fast bacilli (AFB) in up to 70% of cases.
If granulomatous inflammation is identified by ROSE at the time of FNAB, fresh material should be sent for culture or, failing this, polymerase chain reaction (PCR) for mycobacteria can be performed on air-dried or fixed material.
Other causes of granulomas such as sarcoidosis with its noncaseating, rounded epithelioid granulomas; foreign material; drug sensitivities; primary biliary cirrhosis; idiopathic granulomatous hepatitis; and malignancy are more likely to be incidental findings but can be diagnostic.
Cytopathology
Cytologic diagnostic criteria ( Fig. 6-24 )
-
Cellularity: depends on the etiology of granulomas.
-
Pattern: mildly cellular with small tissue fragments.
-
Small tissue fragments of hepatocytes, along with granulomas, epithelioid histiocytes, lymphocytes, and in mycobacterial and some fungal infections, considerable caseous or other necrotic material in the background.
-
Epithelioid histiocytes are large with abundant, pale-staining cytoplasm and an eccentric, plump, elongated, “footprint” nucleus showing finely granular chromatin and small distinct nucleoli.
-
Langhans and/or foreign body type giant cells occasionally seen.
Differential Diagnosis
Once granulomas are recognized, the diagnosis hinges on identification of an organism or other etiologic agent. The presence of caseous necrosis suggests tuberculosis. Granulomas can be associated with metastatic carcinoma and seminoma, as well as with Hodgkin and other malignant lymphomas.
Ancillary Diagnostic Tests
The Ziehl-Neelsen stain for mycobacteria and Grocott methenamine silver stain for fungi are required on extra direct smears and cell block material, along with mandatory cultures for AFB and fungi, as well as PCR, if available.
Iron Overload
Smear Pattern: Depends on the Etiology of Iron Overload
Clinical Features
Iron overload can be primary as in genetic hemochromatosis or secondary associated with anemia, excessive ingestion of iron, frequent blood transfusions in leukemia or other malignancies, or cirrhosis, typically in the setting of steatohepatitis or HCV infection.
Iron accumulates in both hepatocytes and Kupffer cells. FNAB can indicate increased liver iron concentration but does not replace formal biopsy for histopathology and biochemical analysis of liver iron. Distinction between genetic hemochromatosis and hemosiderosis due to other causes is not possible.
Cytopathology
Cytologic diagnostic criteria ( Fig. 6-25 )
-
Pattern: varies with cause.
-
Fine and coarse, granular, yellow-brown pigment in hepatocytes and Kupffer cells.
-
Aggregates of extracellular pigment in heavy overload.
-
Blue granules on the Perls’ Prussian blue iron stain.
-
Evidence of fibrosis, regeneration, and increased bile duct epithelium if the liver is fibrotic or cirrhotic.
Lipofuscin pigment may be confused with hemosiderin. It is usually yellow-brown, perinuclear, rarely clumped, and not extracellular, and it is weakly PAS positive.
The absence of iron in malignant hepatocytes in contrast to heavy deposits in benign cells may be helpful in the diagnosis of hepatocellular carcinoma in patients with hemochromatosis.
Amyloidosis
Smear Pattern: Mildly Cellular With Small Tissue Fragments
Clinical Features
Amyloidosis occurs in a heterogenous group of diseases, all showing extracellular deposition of hyaline amorphous material. It can be classified in several ways, according to whether it is systemic versus localized, according to the specific disease association, or according to the chemical composition of the amyloid fibrils. There are up to 15 types of amyloid fibrils, the principal two being amyloid light chain, commonly associated with plasma cell dyscrasias, and amyloid A protein, which is associated with long-standing chronic inflammation and deposition of amyloid in blood vessel walls, portal tract fibrous tissue, or the parenchyma within the space of Disse, where progressive accumulation causes compression and atrophy of hepatocytes. The clinical presentation is usually one of progressive liver failure, but amyloid can be localized and therefore present as a mass lesion.
Cytopathology
Cytologic diagnostic criteria ( Fig. 6-26 )
-
Pattern: mildly cellular with small tissue fragments.
-
Extracellular amorphous material, which is vaguely fibrillary, glassy or waxy, and pink or green (Pap stain) or pink to purple (Giemsa stain).
-
Apple green birefringence on polarizing a Congo red–stained smear or cell block section.
-
Atrophic liver cells.
Differential Diagnosis
In FNAB smears the DD includes necrosis. The amyloid usually has tissue fragments of various thickness containing thin squashed fibroblasts, rather than a granular appearance of necrosis.
Liver Transplantation
Clinical Features
Although core biopsy is considered the “gold standard” in monitoring liver allografts, FNAB cytology is routinely used by some liver transplant centers for diagnosis of acute allograft rejection and monitoring the response to treatment. Distinction among acute rejection, HCV reactivation, and cytomegalovirus infection has been reported. Chronic rejection, the diagnosis of which relies on findings other than inflammation, cannot be reliably diagnosed.
Mass lesions in liver transplant patients can still be investigated by FNAB. The DD includes infectious masses and post-transplant lymphoproliferative disorders (PTLD), which are discussed under lymphomas (see page 274 ).
Cysts and Cystic Lesions
Smear Pattern: Background Predominant: Cystic
Clinical Features
Cystic lesions can be congenital or acquired, non-neoplastic, or neoplastic. They can be single or multiple and simple or complex. Imaging usually identifies the cystic nature of the lesion, and FNAB can provide diagnostic material, although there is a significant nondiagnostic rate, and FNAB can be therapeutic. The DD includes congenital cysts and infectious cysts discussed here and cystic primary and metastatic tumors discussed later in “Biliary Tumors: Benign” and “Biliary Tumors: Malignant.”
(i)
Congenital (Developmental) Cysts
Smear Pattern: Background Predominant: Cystic
Clinical Features
These comprise a complex group of disorders including solitary cysts, Caroli disease, and adult polycystic disease.
The majority of solitary cysts present in the fourth to sixth decades of life, often identified incidentally when the patient is being investigated for malignancy. The female-to-male ratio is 4:1. The etiology and pathogenesis are unknown. The cyst may be unilocular or multilocular, measure up to 10 cm in diameter, and contain usually clear fluid, but it may be milky, mucoid, purulent, bloody, or bile stained. The epithelial lining is usually simple cuboidal or columnar but can undergo squamous metaplasia or be ciliated and of foregut origin.
Caroli disease is defined by multiple dilatations of the intrahepatic and extrahepatic bile ducts up to 5 cm in diameter, with usually diffuse but occasionally focal involvement and continuity with the biliary tree, so the cysts contain bile and are prone to the formation of calculi. The cuboidal or columnar lining epithelium may be reactive or hyperplastic.
Adult polycystic disease is an autosomal dominant disease that presents in the fourth to fifth decade with a male-to-female ratio of about 4:1. The liver is grossly enlarged and diffusely cystic with cysts varying up to more than 10 cm in diameter. The cyst fluid is clear and light yellow, although occasionally bloody. The cysts are lined by simple cuboidal or columnar epithelium, which may become attenuated with increasing size. These cysts are rarely aspirated unless acutely symptomatic.
Cytopathology
Cytologic diagnostic criteria
-
Cellularity is low.
-
Pattern: background predominant: cystic.
-
Occasional epithelial cells and macrophages in a proteinaceous background.
-
Scant cuboidal or columnar epithelial cells, usually bland but can be hyperplastic, dysplastic, or rarely malignant.
-
Ciliated cells in hepatic foregut cyst have basal nuclei without prominent nucleoli, abundant apical cytoplasm, terminal plates, and fine, delicate cilia.
-
Squamous cells are rarely seen.
-
Hemosiderin and mucin may be present.
-
Abundant bile pigment in Caroli disease.
Differential Diagnosis
Rarely, malignant tumors arise in solitary cysts or Caroli disease. These are usually adenocarcinomas and rarely squamous cell carcinomas.
In multiloculated cysts malignant change may not be detected in smears unless the locule showing malignant change is aspirated.
(ii)
Hydatid Cyst
Smear Pattern: Background Prominent: Cystic
Clinical Features
The Echinococcus granulosus larva travels to the liver via the portal venous system after penetrating the wall of the colon. Here it encysts and multiplies. Usually the cysts are asymptomatic and found incidentally on imaging. Sometimes they compress structures such as the biliary tree or portal vein, causing jaundice or portal hypertension. Occasionally they may rupture and disseminate. The cyst wall consists of an external, chitinous, laminated membrane surrounded by fibrosis and compressed liver. The inner germinal layer forms brood capsules, which separate from the cyst wall into the lumen and produce numerous scolices, which in turn will become future tapeworms. Each scolex contains suckers and rows of hooklets. As the cyst enlarges, invaginations of the wall into the adjacent liver can give rise to daughter cysts. Partial cyst rupture can be associated with an inflammatory reaction.
Routine FNAB of a hydatid cyst is contraindicated because of the theoretical risk of anaphylaxis should the cyst contents spill into the abdomen.
Cytopathology
Cytologic diagnostic criteria ( Figs. 6-27A and B )
-
-
Pattern: background predominant: cystic.
-
Fragments of laminated membrane, pink in Pap and pale magenta in Giemsa stains.
-
Background is granular and proteinaceous with a variable number of inflammatory cells.
-
Scolices: bulging, rounded, blunt “heads” with attached suckers and rows of hooklets.
-
Detached hooklets, which are refractile and birefringent.
(iii)
Amoebic Abscess
Smear Pattern: Background Prominent: Suppurative and Necrotic
Clinical Features
The parasite Entamoeba histolytica reaches the liver from the colon via the portal vein. Patients present with an enlarged, tender liver; fever; and leucocytosis. The right lobe is more often affected, reflecting its greater portal venous drainage, and there are often multiple lesions. Serology is positive in most cases. FNAB is therapeutic and diagnostic. Amoebae secrete enzymes causing coagulative necrosis, which becomes confluent and forms a ragged cavity initially without a fibrous rim. The inflammatory cell response is limited. Amoebae can be identified in the necrotic lining or in the immediately surrounding liver parenchyma but not often in the center of the abscess. Granulation tissue and fibrosis occur with time.
Cytopathology
FNAB produces characteristic thick, reddish-brown, semifluid material classically described as “anchovy sauce.”
Cytologic diagnostic criteria
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Cellularity: low apart from variable numbers of inflammatory cells.
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Pattern: background predominant: suppurative and necrotic.
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Degenerate cells, necrotic debris, and blood, sometimes with admixed bile.
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Variable number of neutrophils, which degenerate.
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Oval amoebic trophozoites resemble histiocytes and in Pap smears are characterized by an eccentric spherical nucleus showing a central clear zone with a dotlike karyosome and margination of chromatin and vacuolated pale cytoplasm containing ingested red blood cells.
In the absence of the parasite the distinctive macroscopic and microscopic appearance of the aspirate allows a provisional cytological diagnosis of amoebic abscess.
Differential Diagnosis
Macrophages can contain ingested red cells, but the amoebic nuclei are distinctive. Both the amoebae and macrophages are Diastase-Periodic Acid Schiff positive.
(iv)
Pyogenic Abscess
Smear Pattern: Background Predominant: Purulent
Clinical Features
Most abscesses of liver are caused by pyogenic bacteria, usually gram-negative bacteria, particularly Escherichia coli or Klebsiella spp., but anaerobic bacteria and Streptococcus milleri are being reported with increasing frequency. Actinomyces israelii may rarely be a cause of liver abscess, arriving via the portal vein from infection in the female genital tract. Bacteria seed the liver via hematogenous, biliary, or direct routes. The patient usually presents with fever, chills and rigors, malaise, abdominal pain, anorexia, and weight loss and often jaundice, with a leukocytosis and abnormal LFT. CT scan or ultrasound usually identifies one or multiple cavities up to 20 cm in diameter with irregular, ill-defined walls. Macroscopically, multiple microabscesses form a honeycomb arrangement. Colonies of filamentous Actinomyces spp. matted together with fibrin may be seen as yellow “sulfur granules” surrounded by a wall of granulation tissue and fibrosis with prominent lipid-laden foamy macrophages.
The DD both clinically and radiologically of a pyogenic abscess includes primary and secondary tumors undergoing central cystic necrosis. FNAB produces pus, and a sample should be sent for microbiologic investigations for bacteria including mycobacteria and fungi.
The liver can be involved in disseminated fungal infections, usually starting in the lungs as an opportunistic infection in immunosuppressed patients. FNAB will demonstrate the fungal organism in a purulent or granulomatous background, and Aspergillus spp. and Phycomycetes may invade blood vessels causing thrombosis, infarction, and necrosis.
Cytopathology
Cytologic diagnostic criteria
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Cellularity is low.
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Pattern: background predominant: suppuration and necrosis.
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Numerous degenerate neutrophils, often caught up in fibrin, with necrotic liver cells, macrophages, debris, bacteria, and in actinomycosis, foamy histiocytes.
Routine gram, Grocott methenamine–silver, and Ziehl-Neelsen stains are required, and at ROSE extra smears should be made for these stains.
Diagnosis of fungal infections depends on identification of the fungus in the Giemsa or Pap smears as negative images or more easily in a Grocott methenamine–silver stain: hyphae of Aspergillus spp. are 8–10 μm in diameter, septate, and show acute branching, while phycomycoses have broad, ribbon-like, nonseptate hyphae, which branch at right angles, and Candida spp. have both pseudohyphae and yeasts.
Ancillary Diagnostic Tests
Cultures are mandatory for AFB, fungi, and bacteria.
Hepatocytic Benign Tumors
There are several liver lesions including focal fatty change, macroregenerative nodule, focal nodular hyperplasia, and hepatocellular adenoma, which can present with a classic clinical presentation and radiologic appearance while the FNAB shows predominantly benign hepatocytes and the DD includes normal liver and well-differentiated hepatocellular carcinoma. But there are indeterminate cases, and FNAB and core biopsy, which provide architectural information, can be useful.
(i)
Focal Fatty Change
Smear Pattern: Moderately to Markedly Cellular With Small to Large Tissue Fragments Without Dispersal
Clinical Features
Steatosis can be diffuse or focal, presenting as a localized lesion or lesions on imaging, while the usual risk factors may not be present. Macroscopically, lesions are rounded, well circumscribed, and yellow in a background of brown liver.
Cytopathology
Cytologic diagnostic criteria
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Cellularity is moderate.
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Pattern: moderately to markedly cellular with tissue fragments ranging from small to large without dispersal.
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Hepatocytes in tissue fragments varying from minute to large.
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Hepatocytes show uniform polygonal cytoplasm containing fat vacuoles of varying size with central round nuclei, or the intracytoplasmic vacuoles may push the nucleus to one side.
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Bile duct epithelium.
The cytopathologist needs to be confident that the material received is representative of the lesion to make a diagnosis of focal fatty change.
Differential Diagnosis
This includes benign hepatocytic tumors, well-differentiated hepatocellular carcinoma with fat, angiomyolipoma, and other mesenchymal tumors containing fat.
(ii)
Macroregenerative Nodule
Smear Pattern: Moderately to Markedly Cellular With Tissue Fragments Ranging From Small to Large Without Dispersal
Clinical Features
These occur in the setting of cirrhosis due to any cause and are arbitrarily defined as regenerating nodules of hepatocytes greater than 8 mm, with a DD of well-differentiated hepatocellular carcinoma, which also arises in this setting.
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