Fine needle aspiration (FNA) is a commonly used approach to determine the nature of a liver mass.¹, ², ³ The average sensitivity of FNA is about 85%, and the specificity has been reported to be as high as 100%.¹,⁴, ⁵, ⁶, ⁷, ⁸, ⁹, ¹⁰ It is important to understand the clinical history of the patient in order to make the best use of the FNA material. In situations where immediate on-site evaluation is being provided, the cytopathologist can play an important role in making sure that the biopsy material is optimally prepared. In many cases, cell block material may be required to perform immunostains or molecular studies. In addition, in cases where lymphoma may be suspected, needle rinses in RPMI can be performed for flow cytometry analysis. Moreover, in some cases, cytology and cell block material may be insufficient to determine if a hepatocytic tumor is benign or malignant. In such cases, it is prudent to suggest a core biopsy. In some cases, the clinician may opt to perform a core biopsy upfront, in which case touch imprints or smears can be prepared from the core.

Liver FNA biopsy can be performed by individual puncture, coaxial biopsy, or tandem needle biopsy.²,¹¹,¹² Each method has its own unique advantages, and the preferred method may vary according to the operators comfort level and the clinical circumstances of the patient. For instance, in the tandem method, an additional reference needle is utilized in parallel to guide the biopsy needle. This allows multiple passes to be performed, limiting the need for repeated imaging between passes.

FNA of normal or reactive liver parenchyma is comprised mostly of bland appearing hepatocytes. Hepatocytes appear as cells showing abundant polygonal cytoplasm with one to two uniform appearing nuclei (Fig. 22.1). The cytoplasm of hepatocytes shows a granular appearance and may contain lipofuscin pigment.¹³ Bile pigment may also be identified. The nuclei frequently possess an easily identifiable central nucleolus. The hepatocytes are frequently arranged as flat sheets, but scattered single cells may also be seen. Intrahepatic bile ducts can also be seen in benign FNA specimens, represented as small clusters or sheets of cells with indistinct cell borders and small round
evenly spaced nuclei. Endothelial cells are usually inconspicuous; however, the elongated nuclei of endothelial cells can occasionally be seen along the edges of hepatocyte clusters. Prominent endothelial wrapping around hepatocyte clusters is usually not seen in benign liver samples. Kupffer cells can be seen in liver aspirates as single cells attached to hepatocytes. Kupffer cells contain round to oval nuclei with some of the nuclei showing reniform contours.

Infections involving the liver can present as a mass lesion. An abscess is usually straight forward to identify because the smears are dominated by neutrophils and other inflammatory cells (Fig. 22.2). Clusters of epithelioid histiocytes, which may include multinucleated giant cells, indicate granulomatous inflammation. Although infection is a primary consideration when granulomas are seen, malignancies such as lymphoma, germ cell malignancy, and squamous cell carcinoma can also be associated with granulomatous inflammation. If a core biopsy or cell block is available, then special stains should be performed for microorganisms such as fungi, parasites, or mycobacterium. Parasites are difficult to detect on FNA but certain structures, such as the hooklets of Echinococcus, would be pathognomonic for parasitic infestation¹⁴,¹⁵ (Fig. 22.3).

Focal nodular hyperplasia and hepatic adenoma are common hepatocytic tumors that are often biopsied. Both tumors show bland appearing hepatocytes that cytologically overlap greatly with the appearance of reactive changes, such as those seen in cirrhotic nodules. In focal nodular hyperplasia, fragments of stromal tissue and bile duct epithelium can also be seen.¹⁶ Adenomas are typically comprised exclusively of hepatocytes; necrosis and hemorrhage can sometimes be seen in the background (Fig. 22.4).⁴

Bile duct adenomas and hamartomas are benign tumors of bile duct origin. Bile duct hamartoma and adenoma produce smears predominated by clusters of bland ductal cells with evenly spaced uniform nuclei (Fig. 22.5). The appearance of the clusters has been likened to that of a “honeycomb.”

Hepatocellular carcinoma usually occurs in the background of cirrhosis (˜80%). Patients with hepatocellular carcinoma are typically older male individuals with a history of chronic liver disease. Although the cytopathologist should avoid biasing himself or herself with the clinical history alone, a diagnosis of hepatocellular carcinoma should be rendered with care when the clinical findings are not typical.

The gross appearance of smears from a hepatocellular carcinoma usually has a dispersed granular appearance in contrast to benign hepatocytic lesions which tend to produce smears with larger tissue fragments.¹⁷

Well-differentiated hepatocellular carcinoma can be difficult to diagnose on cytomorphology alone because the nuclei frequently do not look overtly malignant.¹⁸,¹⁹ Cytologically, the low-power appearance of well-differentiated hepatocellular carcinoma usually consists of clusters of hepatocytes arranged as rounded nests and thickened trabeculae (Fig. 22.6). The trabaculae are usually three cells or more in thickness.

The clusters of tumor cells can be wrapped in endothelial cells (Fig. 22.7). Endothelial cell wrapping is relatively specific for hepatocellular carcinoma and is usually absent in nodules of cirrhosis.¹⁹ However, benign hepatic lesions can also occasionally show endothelial cell wrapping. Therefore, endothelial cell wrapping of hepatocyte clusters should be interpreted in the context of the overall cytomorphology.²⁰ Thin-walled capillaries coursing through the tumor clusters, called “transgressing vessels,” may also be present (Fig. 22.8). At times, the transgressing vessels show an arborizing pattern.²¹

Tumor clusters may be comprised of small hepatocytes with a high N/C ratio, as a consequence of a diminished amount of cytoplasm compared to normal hepatocytes. Large macronucleoli usually indicate malignancy. Moderately and poorly differentiated hepatocellular carcinoma tends to have granular appearing smears that result from the fragmentation of delicate cell membranes or from necrosis. As a result, numerous naked nuclei can be seen (Fig. 22.9). It has
been suggested that the presence of many naked nuclei will help distinguish hepatocellular carcinoma from other malignant tumors, but this not a completely specific feature.²² In the case of moderately differentiated hepatocellular carcinoma, the cytology shows small intact clusters of cells having a similar degree of nuclear atypia, but having abundant polygonal cytoplasm. In addition, some cases of moderately differentiated hepatocellular carcinoma retain trabecular aggregates with endothelial wrapping, strongly supporting the diagnosis of hepatocellular carcinoma. Poorly differentiated hepatocellular carcinoma can be difficult to recognize because many of the typical features of hepatocellular carcinoma are absent (Fig. 22.10). Poorly differentiated hepatocellular carcinoma can resemble other carcinomas, including intrahepatic cholangiocarcinoma. It should be noted that combined tumors containing both adenocarcinoma and hepatocellular carcinoma also exist. In cases of poorly differentiated hepatocellular carcinoma, acquisition of cell block material or a core biopsy is needed to perform immunohistochemical stains. Immunostains for markers of hepatic differentiation, such as HePar-1 and Arginase, as well as a battery of other stains to exclude various types of metastatic tumors, are usually needed.²³

Several variants of hepatocellular carcinoma can be mistaken for other tumor types. Hepatocellular carcinomas with psuedoglands can be mistaken for adenocarcinoma. When numerous gland-like or acinar structures are present, the diagnosis of adenocarcinoma should be excluded. Large three-dimensional rounded aggregates may be a clue to the diagnosis of adenocarcinoma. Wrapping of endothelial cells around acinar or rounded clusters of cells usually indicates hepatocytic differentiation. Clear cell hepatocellular carcinoma, as the name indicates, is comprised primarily of cells with clear cytoplasm because of the accumulation of glycogen. The tumor cells may appear vacuolated. As a result, clear cell hepatocellular carcinoma can mimic metastatic adrenal cortical carcinoma or renal cell carcinoma (Fig. 22.11). It should also be noted that large tumors directly involving the liver and either the adrenal gland or kidney can have a complicated clinical-pathologic picture without a clear site of origin. Clues that assist in the diagnosis of hepatocellular carcinoma include the presence of bile, trabecular arrangements of tumor cells wrapped by endothelial cells, and Mallory bodies.²⁴ Immunostains for hepatocytic differentiation are of great use in this scenario, as are immunostains for renal cortical differentiation such as PAX-8.

Fibrolamellar carcinoma (FLC) has unique clinical and morphologic features. Smears prepared from FLC are comprised mostly of single cells containing large
nuclei and prominent red nucleoli (Fig. 22.12). The tumor cells of FLC also contain abundant granular cytoplasm. Cytoplasmic pale bodies are also frequently seen. The abundance of cytoplasm may lead to the erroneous conclusion that the nuclei are not enlarged, because the N/C ratio may appear relatively normal. Trabecular aggregates and endothelial wrapping are not usually seen in cytology preparations of FLC.²⁵ Fragments of hyalinized stroma can also be identified, either in isolation or intimately admixed with tumor cells.